Modulators of atp-binding cassette transporters

TECHNICAL FIELD OF THE INVENTION

0001The present invention relates to modulators of ATP-Binding Cassette ("ABC") transporters or fragments thereof, including Cystic Fibrosis Transmembrane Conductance Regulator ("CFTR"), compositions thereof and methods therewith. Also described herein are methods of treating ABC transporter mediated diseases using such modulators.

BACKGROUND OF THE INVENTION

0002ABC transporters are a family of membrane transporter proteins that regulate the transport of a wide variety of pharmacological agents, potentially toxic drugs, and xenobiotics, as well as anions. ABC transporters are homologous membrane proteins that bind and use cellular adenosine triphosphate (ATP) for their specific activities. Some of these transporters were discovered as multidrug resistance proteins (like the MDR1-P glycoprotein, or the multidrug resistance protein, MRP1), defending malignant cancer cells against chemotherapeutic agents. To date, 48 ABC Transporters have been identified and grouped into 7 families based on their sequence identity and function.

0003ABC transporters regulate a variety of important physiological roles within the body and provide defense against harmful environmental compounds. Because of this, they represent important potential drug targets for the treatment of diseases associated with defects in the transporter, prevention of drug transport out of the target cell, and intervention in other diseases in which modulation of ABC transporter activity may be beneficial.

0004One member of the ABC transporter family commonly associated with disease is the cAMP/ATP-mediated anion channel, CFTR. CFTR is expressed in a variety of cells types, including absorptive and secretory epithelia cells, where it regulates anion flux across the membrane, as well as the activity of other ion channels and proteins. In epithelia cells, normal functioning of CFTR is critical for the maintenance of electrolyte transport throughout the body, including respiratory and digestive tissue. CFTR is composed of approximately 1480 amino acids that encode a protein made up of a tandem repeat of transmembrane domains, each containing six transmembrane helices and a nucleotide binding domain. The two transmembrane domains are linked by a large, polar, regulatory (R)-domain with multiple phosphorylation sites that regulate channel activity and cellular trafficking.

0005The gene encoding CFTR has been identified and sequenced (See <nplcit id="ncit0001" npl-type="s"><text>Gregory, R. J. et al. (1990) Nature 347:382-386</text></nplcit>; <nplcit id="ncit0002" npl-type="s"><text>Rich, D. P. et al. (1990) Nature 347:358-362</text></nplcit>), (<nplcit id="ncit0003" npl-type="s"><text>Riordan, J. R. et al. (1989) Science 245:1066-1073</text></nplcit>). A defect in this gene causes mutations in CFTR resulting in Cystic Fibrosis ("CF"), the most common fatal genetic disease in humans. Cystic Fibrosis affects approximately one in every 2,500 infants in the United States. Within the general United States population, up to 10 million people carry a single copy of the defective gene without apparent ill effects. In contrast, individuals with two copies of the CF associated gene suffer from the debilitating and fatal effects of CF, including chronic lung disease.

0006In patients with cystic fibrosis, mutations in CFTR endogenously expressed in respiratory epithelia leads to reduced apical anion secretion causing an imbalance in ion and fluid transport. The resulting decrease in anion transport contributes to enhanced mucus accumulation in the lung and the accompanying microbial infections that ultimately cause death in CF patients. In addition to respiratory disease, CF patients typically suffer from gastrointestinal problems and pancreatic insufficiency that, if left untreated, results in death. In addition, the majority of males with cystic fibrosis are infertile and fertility is decreased among females with cystic fibrosis. In contrast to the severe effects of two copies of the CF associated gene, individuals with a single copy of the CF associated gene exhibit increased resistance to cholera and to dehydration resulting from diarrhea - perhaps explaining the relatively high frequency of the CF gene within the population.

0007Sequence analysis of the CFTR gene of CF chromosomes has revealed a variety of disease causing mutations (<nplcit id="ncit0004" npl-type="s"><text>Cutting, G. R. et al. (1990) Nature 346:366-369</text></nplcit>; <nplcit id="ncit0005" npl-type="s"><text>Dean, M. et al. (1990) Cell 61:863:870</text></nplcit>; and <nplcit id="ncit0006" npl-type="s"><text>Kerem, B-S. et al. (1989) Science 245:1073-1080</text></nplcit>; <nplcit id="ncit0007" npl-type="s"><text>Kerem, B-S et al. (1990) Proc. Natl. Acad. Sci. USA 87:8447-8451</text></nplcit>). To date, > 1000 disease causing mutations in the CF gene have been identified (http://www.genet.sickkids.on.ca/cftr/). The most prevalent mutation is a deletion of phenylalanine at position 508 of the CFTR amino acid sequence, and is commonly referred to as ΔF508-CFTR. This mutation occurs in approximately 70% of the cases of cystic fibrosis and is associated with a severe disease.

0008The deletion of residue 508 in ΔF508-CFTR prevents the nascent protein from folding correctly. This results in the inability of the mutant protein to exit the ER, and traffic to the plasma membrane. As a result, the number of channels present in the membrane is far less than observed in cells expressing wild-type CFTR. In addition to impaired trafficking, the mutation results in defective channel gating. Together, the reduced number of channels in the membrane and the defective gating lead to reduced anion transport across epithelia leading to defective ion and fluid transport. (<nplcit id="ncit0008" npl-type="s"><text>Quinton, P. M. (1990), FASEB J. 4: 2709-2727</text></nplcit>). Studies have shown, however, that the reduced numbers of ΔF508-CFTR in the membrane are functional, albeit less than wild-type CFTR. (<nplcit id="ncit0009" npl-type="s"><text>Dalemans et al. (1991), Nature Lond. 354: 526-528</text></nplcit>; Denning et al., supra; <nplcit id="ncit0010" npl-type="s"><text>Pasyk and Foskett (1995), J. Cell. Biochem. 270: 12347-50</text></nplcit>). In addition to ΔF508-CFTR, other disease causing mutations in CFTR that result in defective trafficking, synthesis, and/or channel gating could be up- or down-regulated to alter anion secretion and modify disease progression and/or severity.

0009Although CFTR transports a variety of molecules in addition to anions, it is clear that this role (the transport of anions) represents one element in an important mechanism of transporting ions and water across the epithelium. The other elements include the epithelial Na⁺ channel, ENaC, Na⁺/2Cl^-/K⁺ co-transporter, Na⁺-K⁺-ATPase pump and the basolateral membrane K⁺ channels, that are responsible for the uptake of chloride into the cell.

0010These elements work together to achieve directional transport across the epithelium via their selective expression and localization within the cell. Chloride absorption takes place by the coordinated activity of ENaC and CFTR present on the apical membrane and the Na⁺-K⁺-ATPase pump and Cl- channels expressed on the basolateral surface of the cell. Secondary active transport of chloride from the luminal side leads to the accumulation of intracellular chloride, which can then passively leave the cell via Cl^- channels, resulting in a vectorial transport. Arrangement of Na⁺/2Cl^-/K⁺ co-transporter, Na⁺-K⁺-ATPase pump and the basolateral membrane K⁺ channels on the basolateral surface and CFTR on the luminal side coordinate the secretion of chloride via CFTR on the luminal side. Because water is probably never actively transported itself, its flow across epithelia depends on tiny transepithelial osmotic gradients generated by the bulk flow of sodium and chloride.

0011In addition to Cystic Fibrosis, modulation of CFTR activity may be beneficial for other diseases not directly caused by mutations in CFTR, such as secretory diseases and other protein folding diseases mediated by CFTR. These include, but are not limited to, chronic obstructive pulmonary disease (COPD), dry eye disease, and Sjögren's Syndrome.

0012COPD is characterized by airflow limitation that is progressive and not fully reversible. The airflow limitation is due to mucus hypersecretion, emphysema, and bronchiolitis. Activators of mutant or wild-type CFTR offer a potential treatment of mucus hypersecretion and impaired mucociliary clearance that is common in COPD. Specifically, increasing anion secretion across CFTR may facilitate fluid transport into the airway surface liquid to hydrate the mucus and optimized periciliary fluid viscosity. This would lead to enhanced mucociliary clearance and a reduction in the symptoms associated with COPD. Dry eye disease is characterized by a decrease in tear aqueous production and abnormal tear film lipid, protein and mucin profiles. There are many causes of dry eye, some of which include age, Lasik eye surgery, arthritis, medications, chemical/thermal bums, allergies, and diseases, such as Cystic Fibrosis and Sjögrens's syndrome. Increasing anion secretion via CFTR would enhance fluid transport from the corneal endothelial cells and secretory glands surrounding the eye to increase corneal hydration. This would help to alleviate the symptoms associated with dry eye disease. Sjögrens's syndrome is an autoimmune disease in which the immune system attacks moisture-producing glands throughout the body, including the eye, mouth, skin, respiratory tissue, liver, vagina, and gut. Symptoms, include, dry eye, mouth, and vagina, as well as lung disease. The disease is also associated with rheumatoid arthritis, systemic lupus, systemic sclerosis, and polymypositis/dermatomyositis. Defective protein trafficking is believed to cause the disease, for which treatment options are limited. Modulators of CFTR activity may hydrate the various organs afflicted by the disease and help to elevate the associated symptoms.

0013As discussed above, it is believed that the deletion of residue 508 in ΔF508-CFTR prevents the nascent protein from folding correctly, resulting in the inability of this mutant protein to exit the ER, and traffic to the plasma membrane. As a result, insufficient amounts of the mature protein are present at the plasma membrane and chloride transport within epithelial tissues is significantly reduced. In fact, this cellular phenomenon of defective ER processing of ABC transporters by the ER machinery has been shown to be the underlying basis not only for CF disease, but for a wide range of other isolated and inherited diseases. The two ways that the ER machinery can malfunction is either by loss of coupling to ER export of the proteins leading to degradation, or by the ER accumulation of these defective/misfolded proteins [<nplcit id="ncit0011" npl-type="s"><text>Aridor M, et al., Nature Med., 5(7), pp 745- 751 (1999</text></nplcit>); <nplcit id="ncit0012" npl-type="s"><text>Shastry, B.S., et al., Neurochem. International, 43, pp 1-7 (2003</text></nplcit>); <nplcit id="ncit0013" npl-type="s"><text>Rutishauser, J., et al., Swiss Med Wkly, 132, pp 211-222 (2002</text></nplcit>); <nplcit id="ncit0014" npl-type="s"><text>Morello, JP et al., TIPS, 21, pp. 466- 469 (2000</text></nplcit>);<nplcit id="ncit0015" npl-type="s"><text> Bross P., et al., Human Mut., 14, pp. 186-198 (1999</text></nplcit>)]. The diseases associated with the first class of ER malfunction are Cystic fibrosis (due to misfolded ΔF508-CFTR as discussed above), Hereditary emphysema (due to al-antitrypsin; non Piz variants), Hereditary hemochromatosis, Coagulation-Fibrinolysis deficiencies, such as Protein C deficiency, Type 1 hereditary angioedema, Lipid processing deficiencies, such as Familial hypercholesterolemia, Type 1 chylomicronemia, Abetalipoproteinemia, Lysosomal storage diseases, such as I-cell disease/Pseudo-Hurler, Mucopolysaccharidoses (due to Lysosomal processing enzymes), Sandhof/Tay-Sachs (due to β-Hexosaminidase), Crigler-Najjar type II (due to UDP-glucuronyl-sialyc-transferase), Polyendocrinopathy/Hyperinsulinemia, Diabetes mellitus (due to Insulin receptor), Laron dwarfism (due to Growth hormone receptor), Myeloperoxidase deficiency, Primary hypoparathyroidism (due to Preproparathyroid hormone), Melanoma (due to Tyrosinase). The diseases associated with the latter class of ER malfunction are Glycanosis CDG type 1, Hereditary emphysema (due to α1-Antitrypsin (PiZ variant), Congenital hyperthyroidism, Osteogenesis imperfecta (due to Type I, II, IV procollagen), Hereditary hypofibrinogenemia (due to Fibrinogen), ACT deficiency (due to α1-Antichymotrypsin), Diabetes insipidus (DI), Neurohypophyseal DI (due to Vasopvessin hormone/V2-receptor), Nephrogenic DI (due to Aquaporin II), Charcot-Marie Tooth syndrome (due to Peripheral myelin protein 22), Pelizaeus-Merzbacher disease, neurodegenerative diseases such as Alzheimer's disease (due to βAPP and presenilins), Parkinson's disease, Amyotrophic lateral sclerosis, Progressive supranuclear palsy, Pick's disease, several polyglutamine neurological disorders such as Huntington, Spinocerebellar ataxia type I, Spinal and bulbar muscular atrophy, Dentatorubral pallidoluysian atrophy, and Myotonic dystrophy, as well as Spongiform encephalopathies, such as Hereditary Creutzfeldt-Jakob disease (due to Prion protein processing defect), Fabry disease (due to lysosomal α-galactosidase A) and Gerstmann-Sträussler-Scheinker syndrome (due to Prp processing defect).

0014In addition to up-regulation of CFTR activity, reducing anion secretion by CFTR modulators may be beneficial for the treatment of secretory diarrheas, in which epithelial water transport is dramatically increased as a result of secretagogue activated chloride transport. The mechanism involves elevation of cAMP and stimulation of CFTR.

0015Although there are numerous causes of diarrhea, the major consequences of diarrheal diseases, resulting from excessive chloride transport are common to all, and include dehydration, acidosis, impaired growth and death.

0016Acute and chronic diarrheas represent a major medical problem in many areas of the world. Diarrhea is both a significant factor in malnutrition and the leading cause of death (5,000,000 deaths/year) in children less than five years old.

0017Secretory diarrheas are also a dangerous condition in patients of acquired immunodeficiency syndrome (AIDS) and chronic inflammatory bowel disease (IBD). 16 million travelers to developing countries from industrialized nations every year develop diarrhea, with the severity and number of cases of diarrhea varying depending on the country and area of travel.

0018Diarrhea in barn animals and pets such as cows, pigs and horses, sheep, goats, cats and dogs, also known as scours, is a major cause of death in these animals. Diarrhea can result from any major transition, such as weaning or physical movement, as well as in response to a variety of bacterial or viral infections and generally occurs within the first few hours of the animal's life.

0019The most common diarrhea causing bacteria is enterotoxogenic E-coli (ETEC) having the K99 pilus antigen. Common viral causes of diarrhea include rotavirus and coronavirus. Other infectious agents include cryptosporidium, giardia lamblia, and salmonella, among others.

0020Symptoms of rotaviral infection include excretion of watery feces, dehydration and weakness. Coronavirus causes a more severe illness in the newborn animals, and has a higher mortality rate than rotaviral infection. Often, however, a young animal may be infected with more than one virus or with a combination of viral and bacterial microorganisms at one time. This dramatically increases the severity of the disease.

0021Accordingly, there is a need for modulators of an ABC transporter activity, and compositions thereof, that can be used to modulate the activity of the ABC transporter in the cell membrane of a mammal.

0022<patcit id="pcit0001" dnum="WO2005075435A1"><text>WO 2005/075435 A1</text></patcit> describes modulators for ABC transporters or fragments thereof. <patcit id="pcit0002" dnum="WO2005016884A1"><text>WO 2005/016884 A1</text></patcit> relates to cyclopropyl derivatives as NK3 receptor agonists being useful in the treatment of e.g. psychotic disorders. <patcit id="pcit0003" dnum="WO2004041277A1"><text>WO 2004/041277 A1</text></patcit> describes carbonylamino-benzimidazole derivatives as androgen receptor modulators. These compounds are useful in the enhancement of weakened muscle tone and the treatment od conditions caused by androgen deficiency or which can be ameliorated by androgen administration.

0023<patcit id="pcit0004" dnum="EP1380576A1"><text>EP 1 380 576 A1</text></patcit> describes a novel 1-H-indazole compound having excellent JNK inhibitory action.

0024<patcit id="pcit0005" dnum="WO2004056745A2"><text>WO 2004/056745 A2</text></patcit> relates to adamantyl acetamides as 11-beta hydroxysteroid dehydrogenase inhibitors. <patcit id="pcit0006" dnum="WO03002533A1"><text>WO 03/002533 A1</text></patcit> describes triamide-substituted indoles, benzofuranes and benzothiophenes as inhibitors of microsomal triglyceride transfer protein (MTP) and/or apolipoprotein B (APO B) secretion. These compounds are useful for the treatment of obesity and lipid disorders. <patcit id="pcit0007" dnum="WO2004035571A1"><text>WO 2004/035571 A1</text></patcit> describes substituted indoles and their use as HCV inhibitors. The compounds disclosed therein are useful to inhibit HCV infection of a cell and in particular for treating HCV infection in a mammal.

0025There is a need for methods of treating ABC transporter mediated diseases using such modulators of ABC transporter activity.

0026There is a need for methods of modulating an ABC transporter activity in an <i>ex vivo</i> cell membrane of a mammal.

0027There is a need for modulators of CFTR activity that can be used to modulate the activity of CFTR in the cell membrane of a mammal.

0028There is a need for methods of treating CFTR-mediated diseases using such modulators of CFTR activity.

0029There is a need for methods of modulating CFTR activity in an <i>ex vivo</i> cell membrane of a mammal.

SUMMARY OF THE INVENTION

0030It has now been found that compounds of this invention, and pharmaceutically acceptable compositions thereof, are useful as modulators of ABC transporter activity, particularly CTFR activity. These compounds have the general formula Ic: <chemistry id="chem0001" num="0001"><img file="EP2007756B1_D0001.tif" /></chemistry> or a pharmaceutically acceptable salt thereof, wherein R₁, R₂, ring A, ring B, and n are defined below.

0031These compounds and pharmaceutically acceptable compositions are useful for treating or lessening the severity of a variety of diseases, disorders, or conditions, including, but not limited to, cystic fibrosis, hereditary emphysema, hereditary hemochromatosis, coagulation-fibrinolysis deficiencies, such as protein C deficiency, Type 1 hereditary angioedema, lipid processing deficiencies, such as familial hypercholesterolemia, Type 1 chylomicronemia, abetalipoproteinemia, lysosomal storage diseases, such as I-cell disease/pseudo-Hurler, mucopolysaccharidoses, Sandhof/Tay-Sachs, Crigler-Najjar type II, polyendocrinopathy/hyperinsulinemia, diabetes mellitus, laron dwarfism, myeloperoxidase deficiency, primary hypoparathyroidism, melanoma, glycanosis CDG type 1, hereditary emphysema, congenital hyperthyroidism, osteogenesis imperfecta, hereditary hypofibrinogenemia, ACT deficiency, diabetes insipidus, neurohypophyseal DI, nephrogenic DI, Charcot-Marie Tooth syndrome, Pelizaeus-Merzbacher disease, neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, progressive supranuclear palsy, Pick's disease, several polyglutamine neurological disorders such as Huntington, spinocerebellar ataxia type I, spinal and bulbar muscular atrophy, dentatorubral pallidoluysian atrophy, and myotonic dystrophy, as well as spongiform encephalopathies, such as hereditary Creutzfeldt-Jakob disease, Fabry disease, Gerstmann-Sträussler-Scheinker syndrome, COPD, dry-eye disease, and Sjögren's disease.

DETAILED DESCRIPTION OF THE INVENTION

I. DEFINITIONS

0032As used herein, the following definitions shall apply unless otherwise indicated.

0033The term "ABC-transporter" as used herein means an ABC-transporter protein or a fragment thereof comprising at least one binding domain, wherein said protein or fragment thereof is present in vivo or in vitro. The term "binding domain" as used herein means a domain on the ABC-transporter that can bind to a modulator. See, e.g., <nplcit id="ncit0016" npl-type="s"><text>Hwang, T. C. et al., J. Gen. Physiol. (1998): 111(3), 477-90</text></nplcit>.

0034The term "CFTR" as used herein means cystic fibrosis transmembrane conductance regulator or a mutation thereof capable of regulator activity, including, but not limited to, ΔF508 CFTR and G551D CFTR (see, e.g., http://www.genet.sickkids.on.ca/cftr/, for CFTR mutations).

0035The term "modulating" as used herein means increasing or decreasing, e.g. activity, by a measurable amount. Compounds that modulate ABC Transporter activity, such as CFTR activity, by increasing the activity of the ABC Transporter, e.g., a CFTR anion channel, are called agonists. Compounds that modulate ABC Transporter activity, such as CFTR activity, by decreasing the activity of the ABC Transporter, e.g., CFTR anion channel, are called antagonists. An agonist interacts with an ABC Transporter, such as CFTR anion channel, to increase the ability of the receptor to transduce an intracellular signal in response to endogenous ligand binding. An antagonist interacts with an ABC Transporter, such as CFTR, and competes with the endogenous ligand(s) or substrate(s) for binding site(s) on the receptor to decrease the ability of the receptor to transduce an intracellular signal in response to endogenous ligand binding.

0036The phrase "treating or reducing the severity of an ABC Transporter mediated disease" refers both to treatments for diseases that are directly caused by ABC Transporter and/or CFTR activities and alleviation of symptoms of diseases not directly caused by ABC Transporter and/or CFTR anion channel activities. Examples of diseases whose symptoms may be affected by ABC Transporter and/or CFTR activity include, but are not limited to, Cystic fibrosis, Hereditary emphysema, Hereditary hemochromatosis, Coagulation-Fibrinolysis deficiencies, such as Protein C deficiency, Type 1 hereditary angioedema, Lipid processing deficiencies, such as Familial hypercholesterolemia, Type 1 chylomicronemia, Abetalipoproteinemia, Lysosomal storage diseases, such as I-cell disease/Pseudo-Hurler, Mucopolysaccharidoses, Sandhof/Tay-Sachs, Crigler-Najjar type II, Polyendocrinopathy/Hyperinsulinemia, Diabetes mellitus, Laron dwarfism, Myeloperoxidase deficiency, Primary hypoparathyroidism, Melanoma, Glycanosis CDG type 1, Hereditary emphysema, Congenital hyperthyroidism, Osteogenesis imperfecta, Hereditary hypofibrinogenemia, ACT deficiency, Diabetes insipidus (DI), Neurohypophyseal DI, Nephrogenic DI, Charcot-Marie Tooth syndrome, Pelizaeus-Merzbacher disease, neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease, Amyotrophic lateral sclerosis, Progressive supranuclear palsy, Pick's disease, several polyglutamine neurological disorders such as Huntington, Spinocerebellar ataxia type I, Spinal and bulbar muscular atrophy, Dentatorubral pallidoluysian atrophy, and Myotonic dystrophy, as well as Spongiform encephalopathies, such as Hereditary Creutzfeldt-Jakob disease, Fabry disease, Gerstmann-Sträussler-Scheinker syndrome, COPD, dry-eye disease, and Sjogren's disease.

0037For purposes of this invention, the chemical elements are identified in accordance with the <nplcit id="ncit0017" npl-type="b"><text>Periodic Table of the Elements, CAS version, Handbook of Chemistry and Physics, 75th Ed</text></nplcit>. Additionally, general principles of organic chemistry are described in "<nplcit id="ncit0018" npl-type="b"><text>Organic Chemistry", Thomas Sorrell, University Science Books, Sausolito: 1999</text></nplcit>, and "<nplcit id="ncit0019" npl-type="b"><text>March's Advanced Organic Chemistry", 5th Ed., Ed.: Smith, M.B. and March, J., John Wiley &amp; Sons, New York: 2001</text></nplcit>.

0038As described herein, compounds of the invention may optionally be substituted with one or more substituents, such as are illustrated generally above, or as exemplified by particular classes, subclasses, and species of the invention.

0039As used herein the term "aliphatic" encompasses the terms alkyl, alkenyl, alkynyl, each of which being optionally substituted as set forth below.

0040As used herein, an "alkyl" group refers to a saturated aliphatic hydrocarbon group containing 1-12 (e.g., 1-8, 1-6, or 1-4) carbon atoms. An alkyl group can be straight or branched. Examples of alkyl groups include, but are not limited to, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, n-heptyl, or 2-ethylhexyl. An alkyl group can be substituted (i.e., optionally substituted) with one or more substituents such as halo, phospho, cycloaliphatic [e.g., cycloalkyl or cycloalkenyl], heterocycloaliphatic [e.g., heterocycloalkyl or heterocycloalkenyl], aryl, heteroaryl, alkoxy, aroyl, heteroaroyl, acyl [e.g., (aliphatic)carbonyl, (cycloaliphatic)carbonyl, or (heterocycloaliphatic)carbonyl], nitro, cyano, amido [e.g., (cycloalkylalkyl)carbonylamino, arylcarbonylamino, aralkylcarbonylamino, (heterocycloalkyl)carbonylamino, (heterocycloalkylalkyl)carbonylamino, heteroarylcarbonylamino, heteroaralkylcarbonylamino alkylaminocarbonyl, cycloalkylaminocarbonyl, heterocycloalkylaminocarbonyl, arylaminocarbonyl, or heteroarylaminocarbonyl], amino [e.g., aliphaticamino, cycloaliphaticamino, or heterocycloaliphaticamino], sulfonyl [e.g., aliphatic-SO₂-] sulfinyl, sulfanyl, sulfoxy, urea, thiourea, sulfamoyl, sulfamide, oxo, carboxy, carbamoyl, cycloaliphaticoxy, heterocycloaliphaticoxy, aryloxy, heteroaryloxy, aralkyloxy, heteroarylalkoxy, alkoxycarbonyl, alkylcarbonyloxy, or hydroxy. Without limitation, some examples of substituted alkyls include carboxyalkyl (such as HOOC-alkyl, alkoxycarbonylalkyl, and alkylcarbonyloxyalkyl), cyanoalkyl, hydroxyalkyl, alkoxyalkyl, acylalkyl, aralkyl, (alkoxyaryl)alkyl, (sulfonylamino)alkyl (such as (alkyl-SO₂-amino)alkyl), aminoalkyl, amidoalkyl, (cycloaliphatic)alkyl, or haloalkyl.

0041As used herein, an "alkenyl" group refers to an aliphatic carbon group that contains 2-8 (e.g., 2-12, 2-6, or 2-4) carbon atoms and at least one double bond. Like an alkyl group, an alkenyl group can be straight or branched. Examples of an alkenyl group include, but are not limited to allyl, isoprenyl, 2-butenyl, and 2-hexenyl. An alkenyl group can be optionally substituted with one or more substituents such as halo, phospho, cycloaliphatic [e.g., cycloalkyl or cycloalkenyl], heterocycloaliphatic [e.g., heterocycloalkyl or heterocycloalkenyl], aryl, heteroaryl, alkoxy, aroyl, heteroaroyl, acyl [e.g., (aliphatic)carbonyl, (cycloaliphatic)carbonyl, or (heterocycloaliphatic)carbonyl], nitro, cyano, amido [e.g., (cycloalkylalkyl)carbonylamino, arylcarbonylamino, aralkylcarbonylamino, (heterocycloalkyl)carbonylamino, (heterocycloalkylalkyl)carbonylamino, heteroarylcarbonylamino, heteroaralkylcarbonylamino alkylaminocarbonyl, cycloalkylaminocarbonyl, heterocycloalkylaminocarbonyl, arylaminocarbonyl, or heteroarylaminocarbonyl], amino [e.g., aliphaticamino, cycloaliphaticamino, heterocycloaliphaticamino, or aliphaticsulfonylamino], sulfonyl [e.g., alkyl-SO₂- cycloaliphatic-SO₂-, or aryl-SO₂-], sulfinyl, sulfanyl, sulfoxy, urea, thiourea, sulfamoyl, sulfamide, oxo, carboxy, carbamoyl, cycloaliphaticoxy, heterocycloaliphaticoxy, aryloxy, heteroaryloxy, aralkyloxy, heteroaralkoxy, alkoxycarbonyl, alkylcarbonyloxy, or hydroxy. Without limitation, some examples of substituted alkenyls include cyanoalkenyl, alkoxyalkenyl, acylalkenyl, hydroxyalkenyl, aralkenyl, (alkoxyaryl)alkenyl, (sulfonylamino)alkenyl (such as (alkyl-SO₂-amino)alkenyl), aminoalkenyl, amidoalkenyl, (cycloaliphatic)alkenyl, or haloalkenyl.

0042As used herein, an "alkynyl" group refers to an aliphatic carbon group that contains 2-8 (e.g., 2-12, 2-6, or 2-4) carbon atoms and has at least one triple bond. An alkynyl group can be straight or branched. Examples of an alkynyl group include, but are not limited to, propargyl and butynyl. An alkynyl group can be optionally substituted with one or more substituents such as aroyl, heteroaroyl, alkoxy, cycloalkyloxy, heterocycloalkyloxy, aryloxy, heteroaryloxy, aralkyloxy, nitro, carboxy, cyano, halo, hydroxy, sulfo, mercapto, sulfanyl [e.g., aliphaticsulfanyl or cycloaliphaticsulfanyl], sulfinyl [e.g., aliphaticsulfinyl or cycloaliphaticsulfinyl], sulfonyl [e.g., aliphatic-SO₂- aliphaticamino-SO₂-, or cycloaliphatic-SO₂-], amido [e.g., aminocarbonyl, alkylaminocarbonyl, alkylcarbonylamino, cycloalkylaminocarbonyl, heterocycloalkylaminocarbonyl, cycloalkylcarbonylamino, arylaminocarbonyl, arylcarbonylamino, aralkylcarbonylamino, (heterocycloalkyl)carbonylamino, (cycloalkylalkyl)carbonylamino, heteroaralkylcarbonylamino, heteroarylcarbonylamino or heteroarylaminocarbonyl], urea, thiourea, sulfamoyl, sulfamide, alkoxycarbonyl, alkylcarbonyloxy, cycloaliphatic, heterocycloaliphatic, aryl, heteroaryl, acyl [e.g., (cycloaliphatic)carbonyl or (heterocycloaliphatic)carbonyl], amino [e.g., aliphaticamino], sulfoxy, oxo, carboxy, carbamoyl, (cycloaliphatic)oxy, (heterocycloaliphatic)oxy, or (heteroaryl)alkoxy.

0043As used herein, an "amido" encompasses both "aminocarbonyl" and "carbonylamino". These terms when used alone or in connection with another group refer to an amido group such as -N(R^X)-C(O)-R^Y or -C(O)-N(R^X)₂, when used terminally, and -C(O)-N(R^X)- or -N(R^X)-C(O)- when used internally, wherein R^X and R^Y are defined below. Examples of amido groups include alkylamido (such as alkylcarbonylamino or alkylaminocarbonyl), (heterocycloaliphatic)amido, (heteroaralkyl)amido, (heteroaryl)amido, (heterocycloalkyl)alkylamido, arylamido, aralkylamido, (cycloalkyl)alkylamido, or cycloalkylamido.

0044As used herein, an "amino" group refers to -NR^XR^Y wherein each of R^X and R^Y is independently hydrogen, aliphatic, cycloaliphatic, (cycloaliphatic)aliphatic, aryl, araliphatic, heterocycloaliphatic, (heterocycloaliphatic)aliphatic, heteroaryl, carboxy, sulfanyl, sulfinyl, sulfonyl, (aliphatic)carbonyl, (cycloaliphatic)carbonyl, ((cycioaliphatic)aliphatic)carbonyl, arylcarbonyl, (araliphatic)carbonyl, (heterocycloaliphatic)carbonyl, ((heterocycloaliphatic)aliphatic)carbonyl, (heteroaryl)carbonyl, or (heteroaraliphatic)carbonyl, each of which being defined herein and being optionally substituted. Examples of amino groups include alkylamino, dialkylamino, or arylamino. When the term "amino" is not the terminal group (e.g., alkylcarbonylamino), it is represented by -NR^X-. R^X has the same meaning as defined above.

0045As used herein, an "aryl" group used alone or as part of a larger moiety as in "aralkyl", "aralkoxy", or "aryloxyalkyl" refers to monocyclic (e.g., phenyl); bicyclic (e.g., indenyl, naphthalenyl, tetrahydronaphthyl, tetrahydroindenyl); and tricyclic (e.g., fluorenyl tetrahydrofluorenyl, or tetrahydroanthracenyl, anthracenyl) ring systems in which the monocyclic ring system is aromatic or at least one of the rings in a bicyclic or tricyclic ring system is aromatic. The bicyclic and tricyclic groups include benzofused 2-3 membered carbocyclic rings. For example, a benzofused group includes phenyl fused with two or more C_4-8 carbocyclic moieties. An aryl is optionally substituted with one or more substituents including aliphatic [e.g., alkyl, alkenyl, or alkynyl]; cycloaliphatic; (cycloaliphatic)aliphatic; heterocycloaliphatic; (heterocycloaliphatic)aliphatic; aryl; heteroaryl; alkoxy; (cycloaliphatic)oxy; (heterocycloaliphatic)oxy; aryloxy; heteroaryloxy; (araliphatic)oxy; (heteroaraliphatic)oxy; aroyl; heteroaroyl; amino; oxo (on a non-aromatic carbocyclic ring of a benzofused bicyclic or tricyclic aryl); nitro; carboxy; amido; acyl [e.g., (aliphatic)carbonyl; (cycloaliphatic)carbonyl; ((cycloaliphatic)aliphatic)carbonyl; (araliphatic)carbonyl; (heterocycloaliphatic)carbonyl; ((heterocycloaliphatic)aliphatic)carbonyl; or (heteroaraliphatic)carbonyl]; sulfonyl [e.g., aliphatic-SO₂- or amino-SO₂-]; sulfinyl [e.g., aliphatic-S(O)- or cycloaliphatic-S(O)-]; sulfanyl [e.g., aliphatic-S-]; cyano; halo; hydroxy; mercapto; sulfoxy; urea; thiourea; sulfamoyl; sulfamide; or carbamoyl. Alternatively, an aryl can be unsubstituted.

0046Non-limiting examples of substituted aryls include haloaryl [e.g., mono-, di (such as <i>p,m</i>-dihaloaryl), and (trihalo)aryl]; (carboxy)aryl [e.g., (alkoxycarbonyl)aryl, ((aralkyl)carbonyloxy)aryl, and (alkoxycarbonyl)aryl]; (amido)aryl [e.g., (aminocarbonyl)aryl, (((alkylamino)alkyl)aminocarbonyl)aryl, (alkylcarbonyl)aminoaryl, (arylaminocarbonyl)aryl, and (((heteroaryl)amino)carbonyl)aryl]; aminoaryl [e.g., ((alkylsulfonyl)amino)aryl or ((dialkyl)amino)aryl]; (cyanoalkyl)aryl; (alkoxy)aryl; (sulfamoyl)aryl [e.g., (aminosulfonyl)aryl]; (alkylsulfonyl)aryl; (cyano)aryl; (hydroxyalkyl)aryl; ((alkoxy)alkyl)aryl; (hydroxy)aryl, ((carboxy)alkyl)aryl; (((dialkyl)amino)alkyl)aryl; (nitroalkyl)aryl; (((alkylsulfonyl)amino)alkyl)aryl; ((heterocycloaliphatic)carbonyl)aryl; ((alkylsulfonyl)alkyl)aryl; (cyanoalkyl)aryl; (hydroxyalkyl)aryl; (alkylcarbonyl)aryl; alkylaryl; (trihaloalkyl)aryl; <i>p</i>-amino-<i>m-</i>alkoxycarbonylaryl; <i>p</i>-amino-<i>m</i>-cyanoaryl; <i>p</i>-halo-<i>m</i>-aminoaryl; or (<i>m</i>-(heterocycloaliphatic)-o-(alkyl))aryl.

0047As used herein, an "araliphatic" such as an "aralkyl" group refers to an aliphatic group (e.g., a C_1-4 alkyl group) that is substituted with an aryl group. "Aliphatic," "alkyl," and "aryl" are defined herein. An example of an araliphatic such as an aralkyl group is benzyl.

0048As used herein, an "aralkyl" group refers to an alkyl group (e.g., a C_1-4 alkyl group) that is substituted with an aryl group. Both "alkyl" and "aryl" have been defined above. An example of an aralkyl group is benzyl. An aralkyl is optionally substituted with one or more substituents such as aliphatic [e.g., alkyl, alkenyl, or alkynyl, including carboxyalkyl, hydroxyalkyl, or haloalkyl such as trifluoromethyl], cycloaliphatic [e.g., cycloalkyl or cycloalkenyl], (cycloalkyl)alkyl, heterocycloalkyl, (heterocycloalkyl)alkyl, aryl, heteroaryl, alkoxy, cycloalkyloxy, heterocycloalkyloxy, aryloxy, heteroaryloxy, aralkyloxy, heteroaralkyloxy, aroyl, heteroaroyl, nitro, carboxy, alkoxycarbonyl, alkylcarbonyloxy, amido [e.g., aminocarbonyl, alkylcarbonylamino, cycloalkylcarbonylamino, (cycloalkylalkyl)carbonylamino, arylcarbonylamino, aralkylcarbonylamino, (heterocycloalkyl)carbonylamino, (heterocycloalkylalkyl)carbonylamino, heteroarylcarbonylamino, or heteroaralkylcarbonylamino], cyano, halo, hydroxy, acyl, mercapto, alkylsulfanyl, sulfoxy, urea, thiourea, sulfamoyl, sulfamide, oxo, or carbamoyl.

0049As used herein, a "bicyclic ring system" includes 8-12 (e.g., 9, 10, or 11) membered structures that form two rings, wherein the two rings have at least one atom in common (e.g., 2 atoms in common). Bicyclic ring systems include bicycloaliphatics (e.g., bicycloalkyl or bicycloalkenyl), bicycloheteroaliphatics, bicyclic aryls, and bicyclic heteroaryls.

0050As used herein, a "carbocycle" or "cycloaliphatic" group encompasses a "cycloalkyl" group and a "cycloalkenyl" group, each of which being optionally substituted as set forth below.

0051As used herein, a "cycloalkyl" group refers to a saturated carbocyclic mono- or bicyclic (fused or bridged) ring of 3-10 (e.g., 5-10) carbon atoms. Examples of cycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, adamantyl, norbornyl, cubyl, octahydro-indenyl, decahydro-naphthyl, bicyclo[3.2.1]octyl, bicyclo[2.2.2]octyl, bicyclo[3.3.1]nonyl, bicyclo[3.3.2.]decyl, bicyclo[2.2.2]octyl, adamantyl, or ((aminocarbonyl)cycloalkyl)cycloalkyl.

0052A "cycloalkenyl" group, as used herein, refers to a non-aromatic carbocyclic ring of 3-10 (e.g., 4-8) carbon atoms having one or more double bonds. Examples of cycloalkenyl groups include cyclopentenyl, 1,4-cyclohexa-di-enyl, cycloheptenyl, cyclooctenyl, hexahydro-indenyl, octahydro-naphthyl, cyclohexenyl, cyclopentenyl, bicyclo[2.2.2]octenyl, or bicyclo[3.3.1]nonenyl.

0053A cycloalkyl or cycloalkenyl group can be optionally substituted with one or more substituents such as phosphor, aliphatic [e.g., alkyl, alkenyl, or alkynyl], cycloaliphatic, (cycloaliphatic) aliphatic, heterocycloaliphatic, (heterocycloaliphatic) aliphatic, aryl, heteroaryl, alkoxy, (cycloaliphatic)oxy, (heterocycloaliphatic)oxy, aryloxy, heteroaryloxy, (araliphatic)oxy, (heteroaraliphatic)oxy, aroyl, heteroaroyl, amino, amido [e.g., (aliphatic)carbonylamino, (cycloaliphatic)carbonylamino, ((cycloaliphatic)aliphatic)carbonylamino, (aryl)carbonylamino, (araliphatic)carbonylamino, (heterocycloaliphatic)carbonylamino, ((heterocycloaliphatic)aliphatic)carbonylamino, (heteroaryl)carbonylamino, or (heteroaraliphatic)carbonylamino], nitro, carboxy [e.g., HOOC-, alkoxycarbonyl, or alkylcarbonyloxy], acyl [e.g., (cycloaliphatic)carbonyl, ((cycloaliphatic) aliphatic)carbonyl, (araliphatic)carbonyl, (heterocycloaliphatic)carbonyl, ((heterocycloaliphatic)aliphatic)carbonyl, or (heteroaraliphatic)carbonyl], cyano, halo, hydroxy, mercapto, sulfonyl [e.g., alkyl-SO₂- and aryl-SO₂-], sulfinyl [e.g., alkyl-S(O)-], sulfanyl [e.g., alkyl-S-], sulfoxy, urea, thiourea, sulfamoyl, sulfamide, oxo, or carbamoyl.

0054As used herein, the term "heterocycle" or "heterocycloaliphatic" encompasses a heterocycloalkyl group and a heterocycloalkenyl group, each of which being optionally substituted as set forth below.

0055As used herein, a "heterocycloalkyl" group refers to a 3-10 membered mono- or bicylic (fused or bridged) (e.g., 5- to 10-membered mono- or bicyclic) saturated ring structure, in which one or more of the ring atoms is a heteroatom (e.g., N, O, S, or combinations thereof). Examples of a heterocycloalkyl group include piperidyl, piperazyl, tetrahydropyranyl, tetrahydrofuryl, 1,4-dioxolanyl, 1,4-dithianyl, 1,3-dioxolanyl, oxazolidyl, isoxazolidyl, morpholinyl, thiomorpholyl, octahydrobenzofuryl, octahydrochromenyl, octahydrothiochromenyl, octahydroindolyl, octahydropyrindinyl, decahydroquinolinyl, octahydrobenzo[<i>b</i>]thiopheneyl, 2-oxa-bicyclo[2.2.2]octyl, 1-aza-bicyclo[2.2.2]octyl, 3-aza-bicyclo[3.2.1]octyl, and 2,6-dioxa-tricyclo[3.3.1.0^3,7]nonyl. A monocyclic heterocycloalkyl group can be fused with a phenyl moiety to form structures, such as tetrahydroisoquinoline, which would be categorized as heteroaryls.

0056A "heterocycloalkenyl" group, as used herein, refers to a mono- or bicylic (e.g., 5-to 10-membered mono- or bicyclic) non-aromatic ring structure having one or more double bonds, and wherein one or more of the ring atoms is a heteroatom (e.g., N, O, or S). Monocyclic and bicyclic heterocycloaliphatics are numbered according to standard chemical nomenclature.

0057A heterocycloalkyl or heterocycloalkenyl group can be optionally substituted with one or more substituents such as phosphor, aliphatic [e.g., alkyl, alkenyl, or alkynyl], cycloaliphatic, (cycloaliphatic)aliphatic, heterocycloaliphatic, (heterocycloaliphatic)aliphatic, aryl, heteroaryl, alkoxy, (cycloaliphatic)oxy, (heterocycloaliphatic)oxy, aryloxy, heteroaryloxy, (araliphatic)oxy, (heteroaraliphatic)oxy, aroyl, heteroaroyl, amino, amido [e.g., (aliphatic)carbonylamino, (cycloaliphatic)carbonylamino, ((cycloaliphatic) aliphatic)carbonylamino, (aryl)carbonylamino, (araliphatic)carbonylamino, (heterocycloaliphatic)carbonylamino, ((heterocycloaliphatic) aliphatic)carbonylamino, (heteroaryl)carbonylamino, or (heteroaraliphatic)carbonylamino], nitro, carboxy [e.g., HOOC-, alkoxycarbonyl, or alkylcarbonyloxy], acyl [e.g., (cycloaliphatic)carbonyl, ((cycloaliphatic) aliphatic)carbonyl, (araliphatic)carbonyl, (heterocycloaliphatic)carbonyl, ((heterocycloaliphatic)aliphatic)carbonyl, or (heteroaraliphatic)carbonyl], nitro, cyano, halo, hydroxy, mercapto, sulfonyl [e.g., alkylsulfonyl or arylsulfonyl], sulfinyl [e.g., alkylsulfinyl], sulfanyl [e.g., alkylsulfanyl], sulfoxy, urea, thiourea, sulfamoyl, sulfamide, oxo, or carbamoyl.

0058A "heteroaryl" group, as used herein, refers to a monocyclic, bicyclic, or tricyclic ring system having 4 to 15 ring atoms wherein one or more of the ring atoms is a heteroatom (e.g., N, O, S, or combinations thereof) and in which the monocyclic ring system is aromatic or at least one of the rings in the bicyclic or tricyclic ring systems is aromatic. A heteroaryl group includes a benzofused ring system having 2 to 3 rings. For example, a benzofused group includes benzo fused with one or two 4 to 8 membered heterocycloaliphatic moieties (e.g., indolizyl, indolyl, isoindolyl, 3H-indolyl, indolinyl, benzo[<i>b</i>]furyl, benzo[<i>b</i>]thiophenyl, quinolinyl, or isoquinolinyl). Some examples of heteroaryl are azetidinyl, pyridyl, 1H-indazolyl, furyl, pyrrolyl, thienyl, thiazolyl, oxazolyl, imidazolyl, tetrazolyl, benzofuryl, isoquinolinyl, benzthiazolyl, xanthene, thioxanthene, phenothiazine, dihydroindole, benzo[1,3]dioxole, benzo[b]furyl, benzo[b]thiophenyl, indazolyl, benzimidazolyl, benzthiazolyl, puryl, cinnolyl, quinolyl, quinazolyl,cinnolyl, phthalazyl, quinazolyl, quinoxalyl, isoquinolyl, 4H-quinolizyl, benzo-1,2,5-thiadiazolyl, or 1,8-naphthyridyl.

0059Without limitation, monocyclic heteroaryls include furyl, thiophenyl, 2H-pyrrolyl, pyrrolyl, oxazolyl, thazolyl, imidazolyl, pyrazolyl, isoxazolyl, isothiazolyl, 1,3,4-thiadiazolyl, 2H-pyranyl, 4-H-pranyl, pyridyl, pyridazyl, pyrimidyl, pyrazolyl, pyrazyl, or 1,3,5-triazyl. Monocyclic heteroaryls are numbered according to standard chemical nomenclature.

0060Without limitation, bicyclic heteroaryls include indolizyl, indolyl, isoindolyl, 3H-indolyl, indolinyl, benzo[<i>b</i>]furyl, benzo[<i>b</i>]thiophenyl, quinolinyl, isoquinolinyl, indolizinyl, isoindolyl, indolyl, benzo[<i>b</i>]furyl, bexo[<i>b</i>]thiophenyl, indazolyl, benzimidazyl, benzthiazolyl, purinyl, 4H-quinolizyl, quinolyl, isoquinolyl, cinnolyl, phthalazyl, quinazolyl, quinoxalyl, 1,8-naphthyridyl, or pteridyl. Bicyclic heteroaryls are numbered according to standard chemical nomenclature.

0061A heteroaryl is optionally substituted with one or more substituents such as aliphatic [e.g., alkyl, alkenyl, or alkynyl]; cycloaliphatic; (cycloaliphatic)aliphatic; heterocycloaliphatic; (heterocycloaliphatic)aliphatic; aryl; heteroaryl; alkoxy; (cycloaliphatic)oxy; (heterocycloaliphatic)oxy; aryloxy; heteroaryloxy; (araliphatic)oxy; (heteroaraliphatic)oxy; aroyl; heteroaroyl; amino; oxo (on a non-aromatic carbocyclic or heterocyclic ring of a bicyclic or tricyclic heteroaryl); carboxy; amido; acyl [e.g., aliphaticcarbonyl; (cycloaliphatic)carbonyl; ((cycloaliphatic)aliphatic)carbonyl; (araliphatic)carbonyl; (heterocycloaliphatic)carbonyl; ((heterocycloaliphatic)aliphatic)carbonyl; or (heteroaraliphatic)carbonyl]; sulfonyl [e.g., aliphaticsulfonyl or aminosulfonyl]; sulfinyl [e.g., aliphaticsulfinyl]; sulfanyl [e.g., aliphaticsulfanyl]; nitro; cyano; halo; hydroxy; mercapto; sulfoxy; urea; thiourea; sulfamoyl; sulfamide; or carbamoyl. Alternatively, a heteroaryl can be unsubstituted.

0062Non-limiting examples of substituted heteroaryls include (halo)heteroaryl [e.g., mono- and di-(halo)heteroaryl]; (carboxy)heteroaryl [e.g., (alkoxycarbonyl)heteroaryl]; cyanoheteroaryl; aminoheteroaryl [e.g., ((alkylsulfonyl)amino)heteroaryl and ((dialkyl)amino)heteroaryl]; (amido)heteroaryl [e.g., aminocarbonylheteroaryl, ((alkylcarbonyl)amino)heteroaryl, ((((alkyl)amino)alkyl)aminocarbonyl)heteroaryl, (((heteroaryl)amino)carbonyl)heteroaryl, ((heterocycloaliphatic)carbonyl)heteroaryl, and ((alkylcarbonyl)amino)heteroaryl]; (cyanoalkyl)heteroaryl; (alkoxy)heteroaryl; (sulfamoyl)heteroaryl [e.g., (aminosulfonyl)heteroaryl]; (sulfonyl)heteroaryl [e.g., (alkylsulfonyl)heteroaryl]; (hydroxyalkyl)heteroaryl; (alkoxyalkyl)heteroaryl; (hydroxy)heteroaryl; ((carboxy)alkyl)heteroaryl; (((dialkyl)amino)alkyl]heteroaryl; (heterocycloaliphatic)heteroaryl; (cycloaliphatic)heteroaryl; (nitroalkyl)heteroaryl; (((alkylsulfonyl)amino)alkyl)heteroaryl; ((alkylsulfonyl)alkyl)heteroaryl; (cyanoalkyl)heteroaryl; (acyl)heteroaryl [e.g., (alkylcarbonyl)heteroaryl]; (alkyl)heteroaryl, and (haloalkyl)heteroaryl [e.g., trihaloalkylheteroaryl].

0063A "heteroaraliphatic" (such as a heteroaralkyl group) as used herein, refers to an aliphatic group (e.g., a C_1-4 alkyl group) that is substituted with a heteroaryl group. "Aliphatic," "alkyl," and "heteroaryl" have been defined above.

0064A "heteroaralkyl" group, as used herein, refers to an alkyl group (e.g., a C_1-4 alkyl group) that is substituted with a heteroaryl group. Both "alkyl" and "heteroaryl" have been defined above. A heteroaralkyl is optionally substituted with one or more substituents such as alkyl (including carboxyalkyl, hydroxyalkyl, and haloalkyl such as trifluoromethyl), alkenyl, alkynyl, cycloalkyl, (cycloalkyl)alkyl, heterocycloalkyl, (heterocycloalkyl)alkyl, aryl, heteroaryl, alkoxy, cycloalkyloxy, heterocycloalkyloxy, aryloxy, heteroaryloxy, aralkyloxy, heteroaralkyloxy, aroyl, heteroaroyl, nitro, carboxy, alkoxycarbonyl, alkylcarbonyloxy, aminocarbonyl, alkylcarbonylamino, cycloalkylcarbonylamino, (cycloalkylalkyl)carbonylamino, arylcarbonylamino, aralkylcarbonylamino, (heterocycloalkyl)carbonylamino, (heterocycloalkylalkyl)carbonylamino, heteroarylcarbonylamino, heteroaralkylcarbonylamino, cyano, halo, hydroxy, acyl, mercapto, alkylsulfanyl, sulfoxy, urea, thiourea, sulfamoyl, sulfamide, oxo, or carbamoyl.

0065As used herein, "cyclic moiety" and "cyclic group" refer to mono-, bi-, and tricyclic ring systems including cycloaliphatic, heterocycloaliphatic, aryl, or heteroaryl, each of which has been previously defined.

0066As used herein, a "bridged bicyclic ring system" refers to a bicyclic heterocyclicaliphatic ring system or bicyclic cycloaliphatic ring system in which the rings are bridged. Examples of bridged bicyclic ring systems include, but are not limited to, adamantanyl, norbornanyl, bicyclo[3.2.1]octyl, bicyclo[2.2.2]octyl, bicyclo[3.3.1]nonyl, bicyclo[3.2.3]nonyl, 2-oxabicyclo[2.2.2]octyl, 1-azabicyclo[2.2.2]octyl, 3-azabicyclo[3.2.1]octyl, and 2,6-dioxa-tricyclo[3.3.1.0^3,7]nonyl. A bridged bicyclic ring system can be optionally substituted with one or more substituents such as alkyl (including carboxyalkyl, hydroxyalkyl, and haloalkyl such as trifluoromethyl), alkenyl, alkynyl, cycloalkyl, (cycloalkyl)alkyl, heterocycloalkyl, (heterocycloalkyl)alkyl, aryl, heteroaryl, alkoxy, cycloalkyloxy, heterocycloalkyloxy, aryloxy, heteroaryloxy, aralkyloxy, heteroaralkyloxy, aroyl, heteroaroyl, nitro, carboxy, alkoxycarbonyl, alkylcarbonyloxy, aminocarbonyl, alkylcarbonylamino, cycloalkylcarbonylamino, (cycloalkylalkyl)carbonylamino, arylcarbonylamino, aralkylcarbonylamino, (heterocycloalkyl)carbonylamino, (heterocycloalkylalkyl)carbonylamino, heteroarylcarbonylamino, heteroaralkylcarbonylamino, cyano, halo, hydroxy, acyl, mercapto, alkylsulfanyl, sulfoxy, urea, thiourea, sulfamoyl, sulfamide, oxo, or carbamoyl.

0067As used herein, an "acyl" group refers to a formyl group or R^X-C(O)- (such as alkyl-C(O)-, also referred to as "alkylcarbonyl") where R^X and "alkyl" have been defined previously. Acetyl and pivaloyl are examples of acyl groups.

0068As used herein, an "aroyl" or "heteroaroyl" refers to an aryl-C(O)- or a heteroaryl-C(O)-. The aryl and heteroaryl portion of the aroyl or heteroaroyl is optionally substituted as previously defined.

0069As used herein, an "alkoxy" group refers to an alkyl-O- group where "alkyl" has been defined previously.

0070As used herein, a "carbamoyl" group refers to a group having the structure -O-CO-NR^XR^Y or -NR^X-CO-O-R^Z, wherein R^X and R^Y have been defined above and R^Z can be aliphatic, aryl, araliphatic, heterocycloaliphatic, heteroaryl, or heteroaraliphatic.

0071As used herein, a "carboxy" group refers to -COOH, -COOR^X, -OC(O)H, -OC(O)R^X, when used as a terminal group; or -OC(O)- or -C(O)O- when used as an internal group.

0072As used herein, a "haloaliphatic" group refers to an aliphatic group substituted with 1-3 halogen. For instance, the term haloalkyl includes the group -CF₃.

0073As used herein, a "mercapto" group refers to -SH.

0074As used herein, a "sulfo" group refers to -SO₃H or -SO₃R^X when used terminally or -S(O)₃- when used internally.

0075As used herein, a "sulfamide" group refers to the structure -NR^X-S(O)₂-NR^YR^Z when used terminally and -NR^X-S(O)₂-NR^Y- when used internally, wherein R^X, R^Y, and R^Z have been defined above.

0076As used herein, a "sulfonamide" group refers to the structure -S(O)₂-NR^XR^Y or -NR^X-S(O)₂-R^Z when used terminally; or -S(O)₂-NR^X- or -NR^X -S(O)₂- when used internally, wherein R^X, R^Y, and R^Z are defined above.

0077As used herein a "sulfanyl" group refers to -S-R^X when used terminally and -S-when used internally, wherein R^X has been defined above. Examples of sulfanyls include aliphatic-S-, cycloaliphatic-S-, aryl-S-, or the like.

0078As used herein a "sulfinyl" group refers to -S(O)-R^X when used terminally and - S(O)- when used internally, wherein R^X has been defined above. Exemplary sulfinyl groups include aliphatic-S(O)-, aryl-S(O)-, (cycloaliphatic(aliphatic))-S(O)-, cycloalkyl-S(O)-, heterocycloaliphatic-S(O)-, heteroaryl-S(O)-, or the like.

0079As used herein, a "sulfonyl" group refers to -S(O)₂-R^X when used terminally and -S(O)₂- when used internally, wherein R^X has been defined above. Exemplary sulfonyl groups include aliphatic-S(O)₂-, aryl-S(O)₂-, (cycloaliphatic(aliphatic))-S(O)₂-, cycloaliphatic-S(O)₂-, heterocycloaliphatic-S(O)₂-, heteroaryl-S(O)₂-, (cycloaliphatic(amido(aliphatic)))-S(O)₂- or the like.

0080As used herein, a "sulfoxy" group refers to -O-SO-R^X or -SO-O-R^X, when used terminally and -O-S(O)- or -S(O)-O- when used internally, where R^X has been defined above.

0081As used herein, a "halogen" or "halo" group refers to fluorine, chlorine, bromine or iodine.

0082As used herein, an "alkoxycarbonyl," which is encompassed by the term carboxy, used alone or in connection with another group refers to a group such as alkyl-O-C(O)-.

0083As used herein, an "alkoxyalkyl" refers to an alkyl group such as alkyl-O-alkyl-, wherein alkyl has been defined above.

0084As used herein, a "carbonyl" refer to -C(O)-.

0085As used herein, an "oxo" refers to =O.

0086As used herein, the term "phospho" refers to phosphinates and phosphonates. Examples of phosphinates and phosphonates include -P(O)(R^P)₂, wherein R^P is aliphatic, alkoxy, aryloxy, heteroaryloxy, (cycloaliphatic)oxy, (heterocycloaliphatic)oxy aryl, heteroaryl, cycloaliphatic or amino.

0087As used herein, an "aminoalkyl" refers to the structure (R^X)₂N-alkyl-.

0088As used herein, a "cyanoalkyl" refers to the structure (NC)-alkyl-.

0089As used herein, a "urea" group refers to the structure -NR^X-CO-NR^YR^Z and a "thiourea" group refers to the structure -NR^X-CS-NR^YR^Z when used terminally and -NR^X-CO-NR^Y- or -NR^X-CS-NR^Y- when used internally, wherein R^X, R^Y, and R^Z have been defined above.

0090As used herein, a "guanidine" group refers to the structure - N=C(N(R^XR^Y))N(R^XR^Y) or -NR^X-C(=NR^X)NR^XR^Y wherein R^X and R^Y have been defined above.

0091As used herein, the term "amidino" group refers to the structure - C=(NR^X)N(R^XR^Y) wherein R^X and R^Y have been defined above.

0092In general, the term "vicinal" refers to the placement of substituents on a group that includes two or more carbon atoms, wherein the substituents are attached to adjacent carbon atoms.

0093In general, the term "geminal" refers to the placement of substituents on a group that includes two or more carbon atoms, wherein the substituents are attached to the same carbon atom.

0094The terms "terminally" and "internally" refer to the location of a group within a substituent. A group is terminal when the group is present at the end of the substituent not further bonded to the rest of the chemical structure. Carboxyalkyl, i.e., R^XO(O)C-alkyl is an example of a carboxy group used terminally. A group is internal when the group is present in the middle of a substituent of the chemical structure. Alkylcarboxy (e.g., alkyl-C(O)O- or alkyl-OC(O)-) and alkylcarboxyaryl (e.g., alkyl-C(O)O-aryl- or alkyl-O(CO)-aryl-) are examples of carboxy groups used internally.

0095As used herein, an "aliphatic chain" refers to a branched or straight aliphatic group (e.g., alkyl groups, alkenyl groups, or alkynyl groups). A straight aliphatic chain has the structure -[CH₂]_v-, where v is 1-12. A branched aliphatic chain is a straight aliphatic chain that is substituted with one or more aliphatic groups. A branched aliphatic chain has the structure -[CQQ]_v- where each Q is independently a hydrogen or an aliphatic group; however, Q shall be an aliphatic group in at least one instance. The term aliphatic chain includes alkyl chains, alkenyl chains, and alkynyl chains, where alkyl, alkenyl, and alkynyl are defined above.

0096The phrase "optionally substituted" is used interchangeably with the phrase "substituted or unsubstituted." As described herein, compounds of the invention can optionally be substituted with one or more substituents, such as are illustrated generally above, or as exemplified by particular classes, subclasses, and species of the invention. As described herein, the variables R₁, R₂, and R₃, and other variables contained in formulae described herein encompass specific groups, such as alkyl and aryl. Unless otherwise noted, each of the specific groups for the variables R₁, R₂, and R₃, and other variables contained therein can be optionally substituted with one or more substituents described herein. Each substituent of a specific group is further optionally substituted with one to three of halo, cyano, oxo, alkoxy, hydroxy, amino, nitro, aryl, cycloaliphatic, heterocycloaliphatic, heteroaryl, haloalkyl, and alkyl. For instance, an alkyl group can be substituted with alkylsulfanyl and the alkylsulfanyl can be optionally substituted with one to three of halo, cyano, oxo, alkoxy, hydroxy, amino, nitro, aryl, haloalkyl, and alkyl. As an additional example, the cycloalkyl portion of a (cycloalkyl)carbonylamino can be optionally substituted with one to three of halo, cyano, alkoxy, hydroxy, nitro, haloalkyl, and alkyl. When two alkoxy groups are bound to the same atom or adjacent atoms, the two alkxoy groups can form a ring together with the atom(s) to which they are bound.

0097In general, the term "substituted," whether preceded by the term "optionally" or not, refers to the replacement of hydrogen radicals in a given structure with the radical of a specified substituent. Specific substituents are described above in the definitions and below in the description of compounds and examples thereof. Unless otherwise indicated, an optionally substituted group can have a substituent at each substitutable position of the group, and when more than one position in any given structure can be substituted with more than one substituent selected from a specified group, the substituent can be either the same or different at every position. A ring substituent, such as a heterocycloalkyl, can be bound to another ring, such as a cycloalkyl, to form a spiro-bicyclic ring system, e.g., both rings share one common atom. As one of ordinary skill in the art will recognize, combinations of substituents envisioned by this invention are those combinations that result in the formation of stable or chemically feasible compounds.

0098The phrase "stable or chemically feasible," as used herein, refers to compounds that are not substantially altered when subjected to conditions to allow for their production, detection, and preferably their recovery, purification, and use for one or more of the purposes is one that is not substantially altered when kept at a temperature of 40 °C or less, in the absence of moisture or other chemically reactive conditions, for at least a week.

0099As used herein, an "effective amount" is defined as the amount required to confer a therapeutic effect on the treated patient, and is typically determined based on age, surface area, weight, and condition of the patient. The interrelationship of dosages for animals and humans (based on milligrams per meter squared of body surface) is described by<nplcit id="ncit0020" npl-type="s"><text> Freireich et al., Cancer Chemother. Rep., 50: 219 (1966</text></nplcit>). Body surface area may be approximately determined from height and weight of the patient. See, e.g., <nplcit id="ncit0021" npl-type="s"><text>Scientific Tables, Geigy Pharmaceuticals, Ardsley, New York, 537 (1970</text></nplcit>). As used herein, "patient" refers to a mammal, including a human.

0100Unless otherwise stated, structures depicted herein are also meant to include all isomeric (e.g., enantiomeric, diastereomeric, and geometric (or conformational)) forms of the structure; for example, the R and S configurations for each asymmetric center, (Z) and (E) double bond isomers, and (Z) and (E) conformational isomers. Therefore, single stereochemical isomers as well as enantiomeric, diastereomeric, and geometric (or conformational) mixtures of the present compounds are within the scope of the invention. Unless otherwise stated, all tautomeric forms of the compounds of the invention are within the scope of the invention. Additionally, unless otherwise stated, structures depicted herein are also meant to include compounds that differ only in the presence of one or more isotopically enriched atoms. For example, compounds having the present structures except for the replacement of hydrogen by deuterium or tritium, or the replacement of a carbon by a ¹³C- or ¹⁴C-enriched carbon are within the scope of this invention. Such compounds are useful, for example, as analytical tools or probes in biological assays, or as therapeutic agents.

0101Compounds of the present invention are useful modulators of ABC transporters and are useful in the treatment of ABC transporter mediated diseases.

II. COMPOUNDS

A. Generic Compounds

0102The present invention relates to compounds of formula Ic useful as modulators of ABC transporter activity: <chemistry id="chem0002" num="0002"><img file="EP2007756B1_D0002.tif" /></chemistry> or a pharmaceutically acceptable salt thereof, wherein <ul id="ul0001" list-style="none" compact="compact"><li>R₁ is -Z^AR₄, wherein each Z^A is independently a bond or a branched or straight C_1-6 aliphatic chain wherein up to two carbon units of Z^A are optionally and independently replaced by -CO-, -CS-, -CONR^A-, -CONR^ANR^A-, -CO₂-, -OCO-, -NR^ACO₂-, -O-, -NR^ACONR^A-, -OCONR^A-, -NR^ANR^A-, -NR^ACO-, -S-, -SO-, -SO₂-, -NR^A-, -SO₂NR^A-, -NR^ASO₂-, or -NR^ASO₂NR^A-,</li><li>Each R₄ is independently R^A, halo, -OH, -NH₂, -NO₂, -CN, or -OCF₃,</li><li>Each R^A is independently hydrogen, an aliphatic, a-cycloaliphatic, heterocycloaliphatic, an aryl, or a heteroaryl;</li><li>Each R₂ is independently -Z^BR₅, wherein each Z^B is independently a bond or a branched or straight C_1-6 aliphatic chain optionally substituted with halo, -OH, cyano, C_3-10 cycloaliphatic, 3-10 membered heterocycloaliphatic, monocyclic or 8-12 membered bicyclic aryl, 4-15 membered heteroaryl, or combinations thereof wherein up to two carbon units of Z^B are optionally and independently replaced by -CO-, -CS-, -CONR^B-, -CONR^BNR^B-, -CO₂-, -OCO-, -NR^BCO₂-, -O-, -NR^BCONR^B-, -OCONR^B-, -NR^BNR^B-, -NR^BCO-, -S-, -SO-, -SO₂-, -NR^B-, -SO₂NR^B-, -NR^BSO₂-, or -NR^BSO₂NR^B-,</li><li>Each R₅ is independently R^B, halo, -OH, -NH₂, -NO₂, -CN, -CF₃, or -OCF₃,</li><li>Each R^B is independently hydrogen, a C_3-10 aliphatic, a C_3-10 cycloaliphatic, a 3-10 membered heterocycloaliphatic, a monocyclic or 8-12 membered bicyclic aryl, or a 4-15 membered heteroaryl;</li><li>or, any two adjacent R₂ groups together with the atoms to which they are attached form a carbocycle or a heterocycle;</li><li>Ring A is a 3-7 membered monocyclic ring having 0-3 heteroatoms selected from N, O, and S;</li><li>Ring B is a group having formula Ia: <chemistry id="chem0003" num="0003"><img file="EP2007756B1_D0003.tif" /></chemistry> or a pharmaceutically acceptable salt thereof, wherein</li><li>p is 0-2,</li><li>Each R₃ and R'₃ is independently -Z^CR₆, where each Z^C is independently a bond or a branched or straight C_1-6 aliphatic chain optionally substituted with halo, hydroxy, or combinations thereof, wherein up to two carbon units of Z^C are optionally and independently replaced by -CO-, -CS-, -CONR^C-, -CONR^CNR^C-, -CO₂-, -OCO-, -NR^CCO₂-, -O-, -NR^CCONR^C-, -OCONR^C-, -NR^CNR^C-, -NR^CCO-, -S-, -SO-, -SO₂-, -NR^C-, -SO₂NR^C-, -NR^CSO₂-, or -NR^CSO₂NR^C-,</li><li>Each R₆ is independently R^C, halo, -OH, -NH₂, -NO₂, -CN, or -OCF₃,</li><li>Each R^C is independently hydrogen, an aliphatic optionally substituted with halo, hydroxy, or combinations thereof, a cycloaliphatic optionally substituted with aliphatic, halo, hydroxy, nitro, cyano, or combinations thereof, a heterocycloaliphatic_optionally substituted with aliphatic, halo, hydroxy, nitro, cyano, or combinations_thereof, an aryl, or a heteroaryl, any of which may be optionally substituted with halo, hydroxy, or combinations thereof,</li><li>or, any two adjacent R₃ groups together with the atoms to which they are attached form a heterocycle; and</li><li>n is 1-3, provided that</li><li>when ring A is unsubstituted cyclopentyl, n is 1, R₂ is 4-chloro, and R₁ is hydrogen, then ring B is not 2-(tertbutyl)indol-5-yl, or (2,6-dichlorophenyl(carbonyl))-3-methyl-1H-indol-5-yl; and when ring A is unsubstituted cyclopentyl, n is 0, and R₁ is hydrogen, then ring B is not <chemistry id="chem0004" num="0004"><img file="EP2007756B1_D0004.tif" /></chemistry> or <chemistry id="chem0005" num="0005"><img file="EP2007756B1_D0005.tif" /></chemistry></li></ul>

B. Specific Compounds

1.

R

<u>1</u>

Group

0103R₁ is -Z^AR₄, wherein each Z^A is independently a bond or an optionally substituted branched or straight C_1-6 aliphatic chain wherein up to two carbon units of Z^A are optionally and independently replaced by -CO-, -CS-, -CONR^A-, -CONR^ANR^A-, -CO₂-, -OCO-,-NR^ACO₂-, -O-, -NR^ACONR^A-, -OCONR^A-, -NR^ANR^A-, -NR^ACO-, -S-, -SO-, -SO₂-, -NR^A-, -SO₂NR^A-, -NR^ASO₂-, or -NR^ASO₂NR^A-. Each R₄ is independently R^A, halo, -OH, -NH₂, -NO₂, -CN, or -OCF₃. Each R^A is independently hydrogen, an optionally substituted aliphatic, an optionally substituted cycloaliphatic, an optionally substituted heterocycloaliphatic, an optionally substituted aryl, or an optionally substituted heteroaryl.

0104In several embodiments, R₁ is -Z^AR₄, wherein each Z^A is independently a bond or an optionally substituted branched or straight C_1-6 aliphatic chain and each R₄ is hydrogen.

0105In other embodiments, R₁ is -Z^AR₄, wherein each Z^A is a bond and each R₄ is hydrogen.

2.

R

<u>2</u>

Group

0106Each R₂ is independently -Z^BR₅, wherein each Z^B is independently a bond or an optionally substituted branched or straight C_1-6 aliphatic chain wherein up to two carbon units of Z^B are optionally and independently replaced by -CO-, -CS-, -CONR^B-, -CONR^BNR^B-, - CO₂-, -OCO-, -NR^BCO₂-, -O-, -NR^BCONR^B-, -OCONR^B-, -NR^BNR^B-, -NR^BCO-, -S-, -SO-, -SO₂-, -NR^B-, -SO₂NR^B-, -NR^BSO₂-, or -NR^BSO₂NR^B-. Each R₅ is independently R^B, halo, -OH,-NH₂, -NO₂, -CN, -CF₃, or -OCF₃. Each R^B is independently hydrogen, an optionally substituted aliphatic, an optionally substituted cycloaliphatic, an optionally substituted heterocycloaliphatic, an optionally substituted aryl, or an optionally substituted heteroaryl. Alternatively, any two adjacent R₂ groups together with the atoms to which they are attached form an optionally substituted carbocycle or an optionally substituted heterocycle or heteroaryl.

0107In several embodiments, R₂ is an optionally substituted aliphatic. For example, R₂ is an optionally substituted branched or straight C_1-6 aliphatic chain. In other examples, R₂ is an optionally substituted branched or straight C_1-6 alkyl chain, an optionally substituted branched or straight C_2-6 alkenyl chain, or an optionally substituted branched or straight C_2-6 alkynyl chain. In alternative embodiments, R₂ is a branched or straight C_1-6 aliphatic chain that is optionally substituted with 1-3 of halo, hydroxy, cyano, cycloaliphatic, heterocycloaliphatic, aryl, heteroaryl, or combinations thereof. For example, R₂ is a branched or straight C_1-6 alkyl that is optionally substituted with 1-3 of halo, hydroxy, cyano, cycloaliphatic, heterocycloaliphatic, aryl, heteroaryl, or combinations thereof. In still other examples, R₂ is a methyl, ethyl, propyl, butyl, isopropyl, or tert-butyl, each of which is optionally substituted with 1-3 of halo, hydroxy, cyano, aryl, heteroaryl, cycloaliphatic, or heterocycloaliphatic. In still other examples, R₂ is a methyl, ethyl, propyl, butyl, isopropyl, or tert-butyl, each of which is unsubstituted.

0108In several other embodiments, R₂ is an optionally substituted branched or straight C_1-5 alkoxy. For example, R₂ is a C_1-5 alkoxy that is optionally substituted with 1-3 of hydroxy, aryl, heteroaryl, cycloaliphatic, heterocycloaliphatic, or combinations thereof. In other examples, R₂ is a methoxy, ethoxy, propoxy, butoxy, or pentoxy, each of which is optionally substituted with 1-3 of hydroxy, aryl, heteroaryl, cycloaliphatic, heterocycloaliphatic, or combinations thereof.

0109In other embodiments, R₂ is hydroxy, halo, or cyano.

0110In several embodiments, R₂ is -Z^BR₅, and Z^B is independently a bond or an optionally substituted branched or straight C_1-4 aliphatic chain wherein up to two carbon units of Z^B are optionally and independently replaced by -C(O)-, -O-, -S-, -S(O)₂-, or -NH-, and R₅ is R^B, halo, -OH, -NH₂, -NO₂, -CN, -CF₃, or -OCF₃, and R^B is hydrogen or aryl.

0111In several embodiments, two adjacent R₂ groups form an optionally substituted carbocycle or an optionally substituted heterocycle. For example, two adjacent R₂ groups form an optionally substituted carbocycle or an optionally substituted heterocycle, either of which is fused to the phenyl of formula I, wherein the carbocycle or heterocycle has formula Ib: <chemistry id="chem0006" num="0006"><img file="EP2007756B1_D0006.tif" /></chemistry>

0112Each of Z₁, Z₂, Z₃, Z₄, and Z₅ is independently a bond, -CR₇R'₇-, -C(O)-, -NR₇-, or -O-; each R₇ is independently -Z^DR₈, wherein each Z^D is independently a bind or an optionally substituted branched or straight C_1-6 aliphatic chain wherein up to two carbon units of Z^D are optionally and independently replaced by -CO-, -CS-, -CONR^D-, -CO₂-, -OCO-, - NR^DCO₂-, -O-, -NR^DCONR^D-, -OCONR^D-, -NR^DNR^D-, -NR^DCO-, -S-, -SO-, -SO₂-, -NR^D-, -SO₂NR^D-, -NR^DSO₂-, or -NR^DSO₂NR^D-. Each R₈ is independently R^D, halo, -OH, -NH₂, -NO₂, -CN,-CF₃, or -OCF₃. Each R^D is independently hydrogen, an optionally substituted cycloaliphatic, an optionally substituted heterocycloaliphatic, an optionally substituted aryl, or an optionally substituted heteroaryl. Each R'₇ is independently hydrogen, optionally substituted C_1-6 aliphatic, hydroxy, halo, cyano, nitro, or combinations thereof. Alternatively, any two adjacent R₇ groups together with the atoms to which they are attached form an optionally substituted 3-7 membered carbocyclic ring, such as an optionally substituted cyclobutyl ring, or any two R₇ and R'₇ groups together with the atom or atoms to which they are attached form an optionally substituted 3-7 membered carbocyclic ring or a heterocarbocyclic ring.

0113In several other examples, two adjacent R₂ groups form an optionally substituted carbocycle. For example, two adjacent R₂ groups form an optionally substituted 5-7 membered carbocycle that is optionally substituted with 1-3 of halo, hydroxy, cyano, oxo, cyano, alkoxy, alkyl, or combinations thereof. In another example, two adjacent R₂ groups form a 5-6 membered carbocycle that is optionally substituted with 1-3 of halo, hydroxy, cyano, oxo, cyano, alkoxy, alkyl, or combinations thereof. In still another example, two adjacent R₂ groups form an unsubstituted 5-7 membered carbocycle.

0114In alternative examples, two adjacent R₂ groups form an optionally substituted heterocycle. For instance, two adjacent R₂ groups form an optionally substituted 5-7 membered heterocycle having 1-3 heteroatoms independently selected from N, O, and S. In several examples, two adjacent R₂ groups form an optionally substituted 5-6 membered heterocycle having 1-2 oxygen atoms. In other examples, two adjacent R₂ groups form an unsubstituted 5-7 membered heterocycle having 1-2 oxygen atoms. In other embodiments, two adjacent R₂ groups form a ring selected from: <chemistry id="chem0007" num="0007"><img file="EP2007756B1_D0007.tif" /></chemistry><chemistry id="chem0008" num="0008"><img file="EP2007756B1_D0008.tif" /></chemistry><chemistry id="chem0009" num="0009"><img file="EP2007756B1_D0009.tif" /></chemistry><chemistry id="chem0010" num="0010"><img file="EP2007756B1_D0010.tif" /></chemistry>

0115In alternative examples, two adjacent R₂ groups form an optionally substituted carbocycle or an optionally substituted heterocycle, and a third R₂ group is attached to any chemically feasible position on the phenyl of formula I. For instance, an optionally substituted carbocycle or an optionally substituted heterocycle, both of which is formed by two adjacent R₂ groups; a third R₂ group; and the phenyl of formula I form a group having formula Ic: <chemistry id="chem0011" num="0011"><img file="EP2007756B1_D0011.tif" /></chemistry>

0116Z₁, Z₂, Z₃, Z₄, and Z₅ has been defined above in formula Ib, and R₂ has been defined above in formula I.

0117In several embodiments, each R₂ group is independently selected from hydrogen, halo, -OCH₃, -OH, -CH₂OH, -CH₃, and -OCF₃, and/or two adjacent R₂ groups together with the atoms to which they are attached form <chemistry id="chem0012" num="0012"><img file="EP2007756B1_D0012.tif" /></chemistry><chemistry id="chem0013" num="0013"><img file="EP2007756B1_D0013.tif" /></chemistry><chemistry id="chem0014" num="0014"><img file="EP2007756B1_D0014.tif" /></chemistry><chemistry id="chem0015" num="0015"><img file="EP2007756B1_D0015.tif" /></chemistry>

0118In other embodiments, R₂ is at least one selected from hydrogen, halo, methoxy, phenylmethoxy, hydroxy, hydroxymethyl, trifluoromethoxy, and methyl.

0119In some embodiments, two adjacent R₂ groups, together with the atoms to which they are attached, form <chemistry id="chem0016" num="0016"><img file="EP2007756B1_D0016.tif" /></chemistry>

3.

Ring A

0120Ring A is an optionally substituted 3-7 membered monocyclic ring having 0-3 heteroatoms selected from N, O, and S.

0121In several embodiments, ring A is an optionally substituted 3-7 membered monocyclic cycloaliphatic. For example, ring A is a cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, or cycloheptyl, each of which is optionally substituted with 1-3 of halo, hydroxy, C_1-5 aliphatic, or combinations thereof.

0122In other embodiments, ring A is an optionally substituted 3-7 membered monocyclic heterocycloaliphatic. For example, ring A is an optionally substituted 3-7 membered monocyclic heterocycloaliphatic having 1-2 heteroatoms independently selected from N, O, and S. In other examples, ring A is tetrahydrofuran-yl, tetrahydro-2H-pyran-yl, pyrrolidone-yl, or piperidine-yl, each of which is optionally substituted.

0123In still other examples, ring A is selected from <chemistry id="chem0017" num="0017"><img file="EP2007756B1_D0017.tif" /></chemistry><chemistry id="chem0018" num="0018"><img file="EP2007756B1_D0018.tif" /></chemistry><chemistry id="chem0019" num="0019"><img file="EP2007756B1_D0019.tif" /></chemistry><chemistry id="chem0020" num="0020"><img file="EP2007756B1_D0020.tif" /></chemistry><chemistry id="chem0021" num="0021"><img file="EP2007756B1_D0021.tif" /></chemistry>

0124Each R₉ is independently -Z^ER₁₀, wherein each Z^E is independently a bond or an optionally substituted branched or straight C_1-5 aliphatic chain wherein up to two carbon units of Z^E are optionally and independently replaced by -CO-, -CS-, -CONR^E-, -CO₂-, -OCO-, -NR^ECO₂-, -O-, -NR^ECONR^E-, -OCONR^E-, -NR^ENR^E-, -NR^ECO-, -S-, -SO-, -SO₂-, -NR^E-, -SO₂NR^E-, -NR^ESO₂-, or -NR^ESO₂NR^E-, each R₁₀ is independently R^E, -OH, -NH₂, -NO₂,-CN, -CF₃, oxo, or -OCF₃. Each R^E is independently hydrogen, an optionally substituted cycloaliphatic, an optionally substituted heterocycloaliphatic, an optionally substituted aryl, or an optionally substituted heteroaryl.

0125q is 0-5.

0126In other embodiments, ring A is one selected from <chemistry id="chem0022" num="0022"><img file="EP2007756B1_D0022.tif" /></chemistry><chemistry id="chem0023" num="0023"><img file="EP2007756B1_D0023.tif" /></chemistry>

0127In several embodiments, ring A is <chemistry id="chem0024" num="0024"><img file="EP2007756B1_D0024.tif" /></chemistry>

4.

Ring B

0128Ring B is a group having formula Ia: <chemistry id="chem0025" num="0025"><img file="EP2007756B1_D0025.tif" /></chemistry> or a pharmaceutically acceptable salt thereof, wherein p is 0-3.

0129Each R₃ and R'₃ is independently -Z^CR₆, where each Z^C is independently a bond or an optionally substituted branched or straight C_1-6 aliphatic chain wherein up to two carbon units of Z^C are optionally and independently replaced by -CO-, -CS-, -CONR^C-, -CONR^CNR^C-, -CO₂-, -OCO-, -NR^CCO₂-, -O-, -NR^CCONR^C-, -OCONR^C-, -NR^CNR^C-, -NR^CCO-, -S-, -SO-, -SO₂-, -NR^C-, -SO₂NR^C-, -NR^CSO₂-, or -NR^CSO₂NR^C-. Each R₆ is independently R^C, halo, -OH, -NH₂, -NO₂, -CN, or -OCF₃. Each R^C is independently hydrogen, an optionally substituted aliphatic, an optionally substituted cycloaliphatic, an optionally substituted heterocycloaliphatic, an optionally substituted aryl, or an optionally substituted heteroaryl. Alternatively, any two adjacent R₃ groups together with the atoms to which they are attached form an optionally substituted carbocycle or an optionally substituted heterocycle, or R'₃ and an adjacent R₃, i.e., attached to the 2 position of the indole of formula Ia, together with the atoms to which they are attached form an optionally substituted heterocycle.

0130In several embodiments, ring B is <chemistry id="chem0026" num="0026"><img file="EP2007756B1_D0026.tif" /></chemistry>

0131wherein q is 0-3 and each R₂₀ is -Z^GR₂₁, where each Z^G is independently a bond or an optionally substituted branched or straight C_1-5 aliphatic chain wherein up to two carbon units of Z^G are optionally and independently replaced by -CO-, -CS-, -CONR^G-, -CO₂-, -OCO-, -NR^GCO₂-, -O-, -OCONR^G-, -NR^GNR^G-, -NR^GCO-, -S-, -SO-, -SO₂-, -NR^G-, -SO₂NR^G-, -NR^GSO₂-, or -NR^GSO₂NR^G-. Each R₂₁ is independently R^G, halo, -OH, -NH₂,-NO₂, -CN, or -OCF₃. Each R^G is independently hydrogen, an optionally substituted aliphatic, an optionally substituted cycloaliphatic, an optionally substituted heterocycloaliphatic, an optionally substituted aryl, or an optionally substituted heteroaryl.

0132For example, ring B is <chemistry id="chem0027" num="0027"><img file="EP2007756B1_D0027.tif" /></chemistry>

0133In several embodiments, R'₃ is hydrogen and R₃ is attached to the 2, 3, 4, 5, 6, or 7 position of the indole of formula Ia. In several other examples, R₃ is attached to the 2 or 3 position of the indole of formula Ia, and R₃ is independently an optionally substituted aliphatic. For instance, R₃ is an optionally substituted acyl group. In several instances, R₃ is an optionally substituted (alkoxy)carbonyl. In other instances, R₃ is (methoxy)carbonyl, (ethoxy)carbonyl, (propoxy)carbonyl, or (butoxy)carbonyl, each of which is optionally substituted with 1-3 of halo, hydroxy, or combinations thereof. In other instances, R₃ is an optionally substituted (aliphatic)carbonyl. For example, R₃ is an optionally substituted (alkyl)carbonyl that is optionally substituted with 1-3 of halo, hydroxy, or combinations thereof. In other examples, R₃ is (methyl)carbonyl, (ethyl)carbonyl, (propyl)carbonyl, or (butyl)carbonyl, each of which is optionally substituted with 1-3 of halo, hydroxy, or combinations thereof.

0134In several embodiments, R₃ is an optionally substituted (cycloaliphatic)carbonyl or an optionally substituted (heterocycloaliphatic)carbonyl. In several examples, R₃ is an optionally substituted (C_3-7 cycloaliphatic)carbonyl. For example, R₃ is a (cyclopropyl)carbonyl, (cyclobutyl)carbonyl, (cyclopentyl)carbonyl, (cyclohexyl)carbonyl, or (cycloheptyl)carbonyl, each of which is optionally substituted with aliphatic, halo, hydroxy, nitro, cyano, or combinations thereof. In several alternative examples, R₃ is an optionally substituted (heterocycloaliphatic)carbonyl. For example, R₃ is an optionally substituted (heterocycloaliphatic)carbonyl having 1-3 heteroatoms independently selected from N, O, and S. In other examples, R₃ is an optionally substituted (heterocycloaliphatic)carbonyl having 1-3 heteroatoms independently selected from N and O. In still other examples, R₃ is an optionally substituted 4-7 membered monocyclic (heterocycloaliphatic)carbonyl having 1-3 heteroatoms independently selected from N and O. Alternatively, R₃ is (piperidine-1-yl,)carbonyl, (pyrrolidine-1-yl)carbonyl, or (morpholine-4-yl)carbonyl, (piperazine-1-yl)carbonyl, each of which is optionally substituted with 1-3 of halo, hydroxy, cyano, nitro, or aliphatic.

0135In still other instances, R₃ is optionally substituted (aliphatic)amido such as (aliphatic(amino(carbonyl)) that is attached to the 2 or 3 position on the indole ring of formula Ia. In some embodiments, R₃ is an optionally substituted (alkyl(amino))carbonyl that is attached to the 2 or 3 position on the indole ring of formula Ia. In other embodiments, R₃ is an optionally substituted straight or branched (aliphatic(amino))carbonyl that is attached to the 2 or 3 position on the indole ring of formula Ia. In several examples, R₃ is (N,N-dimethyl(amino))carbonyl, (methyl(amino))carbonyl, (ethyl(amino))carbonyl, (propyl(amino))carbonyl, (prop-2-yl(amino))carbonyl, (dimethyl(but-2-yl(amino)))carbonyl, (tertbutyl(amino))carbonyl, (butyl(amino))carbonyl, each of which is optionally substituted with 1-3 of halo, hydroxy, cycloaliphatic, heterocycloaliphatic, aryl, heteroaryl, or combinations thereof.

0136In other embodiments, R₃ is an optionally substituted (alkoxy)carbonyl. For example, R₃ is (methoxy)carbonyl, (ethoxy)carbonyl, (propoxy)carbonyl, or (butoxy)carbonyl, each of which is optionally substituted with 1-3 of halo, hydroxy, or combinations thereof. In several instances, R₃ is an optionally substituted straight or branched C_1-6 aliphatic. For example, R₃ is an optionally substituted straight or branched C_1-6 alkyl. In other examples, R₃ is independently an optionally substituted methyl, ethyl, propyl, butyl, isopropyl, or tertbutyl, each of which is optionally substituted with 1-3 of halo, hydroxy, cyano, nitro, or combination thereof In other embodiments, R₃ is an optionally substituted C_3-6 cycloaliphatic. Exemplary embodiments include cyclopropyl, 1-methyl-cycloprop-1-yl, etc. In other examples, p is 2 and the two R₃ substituents are attached to the indole of formula Ia at the 2,4- or 2,6- or 2,7- positions. Exemplary embodiments include 6-F, 3-(optionally substituted C_1-6 aliphatic or C_3-6 cycloaliphatic); 7-F-2-(-(optionally substituted C_1-6 aliphatic or C_3-6 cycloaliphatic)), 4F-2-(optionally substituted C_1-6 aliphatic or C_3-6 cycloaliphatic); 7-CN-2-(optionally substituted C_1-6 aliphatic or C_3-6 cycloaliphatic); 7-Me-2-(optionally substituted C_1-6 aliphatic or C_3-6 cycloaliphatic) and 7-OMe-2-(optionally substituted C_1-6 aliphatic or C_3-6 cycloaliphatic).

0137In several embodiments, R₃ is hydrogen. In several instances, R₃ is an optionally substituted straight or branched C_1-6 aliphatic. In other embodiments, R₃ is an optionally substituted C_3-6 cycloaliphatic.

0138In several embodiments, R₃ is one selected from: -H, -CH₃, -CH₂OH, -CH₂CH₃, -CH₂CH₂OH, -CH₂CH₂CH₃, -NH₂, halo, -OCH₃, -CN, -CF₃, -C(O)OCH₂CH₃, -S(O)₂CH₃, -CH₂NH₂, -C(O)NH₂, <chemistry id="chem0028" num="0028"><img file="EP2007756B1_D0028.tif" /></chemistry><chemistry id="chem0029" num="0029"><img file="EP2007756B1_D0029.tif" /></chemistry><chemistry id="chem0030" num="0030"><img file="EP2007756B1_D0030.tif" /></chemistry><chemistry id="chem0031" num="0031"><img file="EP2007756B1_D0031.tif" /></chemistry><chemistry id="chem0032" num="0032"><img file="EP2007756B1_D0032.tif" /></chemistry><chemistry id="chem0033" num="0033"><img file="EP2007756B1_D0033.tif" /></chemistry><chemistry id="chem0034" num="0034"><img file="EP2007756B1_D0034.tif" /></chemistry><chemistry id="chem0035" num="0035"><img file="EP2007756B1_D0035.tif" /></chemistry><chemistry id="chem0036" num="0036"><img file="EP2007756B1_D0036.tif" /></chemistry>

0139In another embodiment, two adjacent R₃ groups form <chemistry id="chem0037" num="0037"><img file="EP2007756B1_D0037.tif" /></chemistry>

0140In several embodiments, R'₃ is independently -Z^CR₆, where each Z^C is independently a bond or an optionally substituted branched or straight C_1-6 aliphatic chain wherein up to two carbon units of Z^C are optionally and independently replaced by -CO-, -CS-, -CONR^C-, -CONR^CNR^C-, -CO₂-, -OCO-, -NR^CCO₂-, -O-, -NR^CCONR^C-, -OCONR^C-, -NR^CNR^C-, NR^CCO-, -S-, -SO-, -SO₂-, -NR^C-, -SO₂NR^C-, -NR^CSO₂-, or -NR^CSO₂NR^C-. Each R₆ is independently R^C, halo, -OH, -NH₂, -NO₂, -CN, or -OCF₃. Each R^C is independently hydrogen, an optionally substituted aliphatic, an optionally substituted cycloaliphatic, an optionally substituted heterocycloaliphatic, or an optionally substituted heteroaryl. In one embodiment, each R^C is hydrogen, C_1-6 aliphatic, or C_3-6 cycloaliphatic, wherein either of the aliphatic or cycloaliphatic is optionally substituted with up to 4 -OH substituents. In another embodiment, R^C is hydrogen, or C_1-6 alkyl optionally substituted with up to 4 -OH substituents.

0141For example, in many embodiments, R'₃ is is independently -Z^CR₆, where each Z^C is independently a bond or an optionally substituted branched or straight C_1-6 aliphatic chain wherein up to two carbon units of Z^C are optionally and independently replaced by -C(O)-, -C(O)NR^C-, -C(O)O-, -NR^CC(O)O-, -O-, -NR^CS(O)₂-, or -NR^C-. Each R₆ is independently R^C, -OH, or -NH₂. Each R^C is independently hydrogen, an optionally substituted cycloaliphatic, an optionally substituted heterocycloaliphatic, or an optionally substituted heteroaryl. In one embodiment, each R^C is hydrogen, C_1-6 aliphatic, or C_3-6 cycloaliphatic, wherein either of the aliphatic or cycloaliphatic is optionally substituted with up to 4 -OH substituents. In another embodiment, R^C is hydrogen, or C_1-6 alkyl optionally substituted with up to 4 -OH substituents.

0142In other embodiments, R'₃ is hydrogen or <chemistry id="chem0038" num="0038"><img file="EP2007756B1_D0038.tif" /></chemistry>

0143wherein R₃₁ is H or a C_1-2 aliphatic that is optionally substituted with 1-3 of halo,-OH, or combinations thereof. R₃₂ is -L-R₃₃, wherein L is a bond, -CH₂-, -CH₂O-,-CH₂NHS(O)₂-, -CH₂C(O)-, -CH₂NHC(O)-, or -CH₂NH-; and R₃₃ is hydrogen, or C_1-2 aliphatic, cycloaliphatic, heterocycloaliphatic, or heteroaryl, each of which is optionally subsitututed with 1 of -OH, -NH₂, or -CN. For example, in one embodiment, R₃₁ is hydrogen and R₃₂ is C_1-2 aliphatic optionally substituted with -OH, -NH₂, or -CN.

0144In several embodiments, R'₃ is independently selected from one of the following: -H, -CH₃, -CH₂CH₃, -C(O)CH₃, -CH₂CH₂OH, -C(O)OCH₃, <chemistry id="chem0039" num="0039"><img file="EP2007756B1_D0039.tif" /></chemistry><chemistry id="chem0040" num="0040"><img file="EP2007756B1_D0040.tif" /></chemistry><chemistry id="chem0041" num="0041"><img file="EP2007756B1_D0041.tif" /></chemistry><chemistry id="chem0042" num="0042"><img file="EP2007756B1_D0042.tif" /></chemistry><chemistry id="chem0043" num="0043"><img file="EP2007756B1_D0043.tif" /></chemistry><chemistry id="chem0044" num="0044"><img file="EP2007756B1_D0044.tif" /></chemistry><chemistry id="chem0045" num="0045"><img file="EP2007756B1_D0045.tif" /></chemistry><chemistry id="chem0046" num="0046"><img file="EP2007756B1_D0046.tif" /></chemistry><chemistry id="chem0047" num="0047"><img file="EP2007756B1_D0047.tif" /></chemistry><chemistry id="chem0048" num="0048"><img file="EP2007756B1_D0048.tif" /></chemistry><chemistry id="chem0049" num="0049"><img file="EP2007756B1_D0049.tif" /></chemistry>

5.

n term

0145n is 1-3.

0146In several embodiments, n is 1. In other embodiments, n is 2. In still other embodiments, n is 3.

0147C. Exemplary compounds of the present invention Exemplary compounds of the present invention include, but are not limited to, those illustrated in Table 1 below. <tables id="tabl0001" num="0001"><img file="EP2007756B1_D0050.tif" /></tables><tables id="tabl0002" num="0002"><img file="EP2007756B1_D0051.tif" /></tables><tables id="tabl0003" num="0003"><img file="EP2007756B1_D0052.tif" /></tables><tables id="tabl0004" num="0004"><img file="EP2007756B1_D0053.tif" /></tables><tables id="tabl0005" num="0005"><img file="EP2007756B1_D0054.tif" /></tables><tables id="tabl0006" num="0006"><img file="EP2007756B1_D0055.tif" /></tables><tables id="tabl0007" num="0007"><img file="EP2007756B1_D0056.tif" /></tables><tables id="tabl0008" num="0008"><img file="EP2007756B1_D0057.tif" /></tables><tables id="tabl0009" num="0009"><img file="EP2007756B1_D0058.tif" /></tables><tables id="tabl0010" num="0010"><img file="EP2007756B1_D0059.tif" /></tables><tables id="tabl0011" num="0011"><img file="EP2007756B1_D0060.tif" /></tables><tables id="tabl0012" num="0012"><img file="EP2007756B1_D0061.tif" /></tables><tables id="tabl0013" num="0013"><img file="EP2007756B1_D0062.tif" /></tables><tables id="tabl0014" num="0014"><img file="EP2007756B1_D0063.tif" /></tables><tables id="tabl0015" num="0015"><img file="EP2007756B1_D0064.tif" /></tables><tables id="tabl0016" num="0016"><img file="EP2007756B1_D0065.tif" /></tables><tables id="tabl0017" num="0017"><img file="EP2007756B1_D0066.tif" /></tables><tables id="tabl0018" num="0018"><img file="EP2007756B1_D0067.tif" /></tables><tables id="tabl0019" num="0019"><img file="EP2007756B1_D0068.tif" /></tables><tables id="tabl0020" num="0020"><img file="EP2007756B1_D0069.tif" /></tables><tables id="tabl0021" num="0021"><img file="EP2007756B1_D0070.tif" /></tables><tables id="tabl0022" num="0022"><img file="EP2007756B1_D0071.tif" /></tables><tables id="tabl0023" num="0023"><img file="EP2007756B1_D0072.tif" /></tables><tables id="tabl0024" num="0024"><img file="EP2007756B1_D0073.tif" /></tables><tables id="tabl0025" num="0025"><img file="EP2007756B1_D0074.tif" /></tables><tables id="tabl0026" num="0026"><img file="EP2007756B1_D0075.tif" /></tables>

III. SUBGENERIC COMPOUNDS OF THE PRESENT INVENTION

0148Another aspect of the present invention provides a compound that is useful for modulating ABC transporter activity. The compound has formula Id: <chemistry id="chem0050" num="0050"><img file="EP2007756B1_D0076.tif" /></chemistry> or a pharmaceutically acceptable salt thereof.

0149R₁, R₂, and ring A are defined above in formula I, and ring B, R₃ and p are defined in formula Ia.

0150However, when R₁ is H, n is 0, ring A is an unsubstituted cyclopentyl, and ring B is an indole-5-yl substituted with 1-2 of R₃, then each R₃ is independently -Z^GR₁₂, where each Z^G is independently a bond or an unsubstituted branched or straight C_1-6 aliphatic chain wherein up to two carbon units of Z^G are optionally and independently replaced by -CS-, - CONR^GNR^G-, -CO₂-, -OCO-, -NR^GCO₂-, -O-, -NR^GCONR^G-, -OCONR^G-, -NR^GNR^G-, -S-,-SO-, -SO₂-, -NR^G-, -SO₂NR^G-, -NR^GSO₂-, or -NR^GSO₂NR^C-, each R₁₂ is independently R^G, halo, -OH, -NH₂,-NO₂, -CN, or -OCF₃, and each R^G is independently hydrogen, an unsubstituted aliphatic, an optionally substituted cycloaliphatic, an optionally substituted heterocycloaliphatic, an unsubstituted aryl, or an optionally substituted heteroaryl; or any two adjacent R₃ groups together with the atoms to which they are attached form an optionally substituted heterocycle. Futhermore, when R₁ is H, n is 1, R₂ is 4-chloro, ring A is an unsubstituted cyclopentyl, and ring B is an indole-5-yl substituted with 1-2 of R₃, then each R₃ is independently -Z^HR₂₂, where each Z^H is independently a bond or an unsubstituted branched or straight C_1-3 aliphatic chain wherein up to two carbon units of Z^H are optionally and independently replaced by - CS-, -CONR^HNR^H, -CO₂- -OCO-, -NR^HCO₂-, -O-, -NR^HCONR^H-, -OCONR^H-, -NR^HNR^H-, -S-, -SO-, -SO₂-, -NR^H-, -SO₂NR^H-, -NR^HSO₂-, or -NR^HSO₂NR^H-, each R₂₂ is independently R^H, halo, -OH, -NH₂, -NO₂, -CN, or -OCF₃, and each R^H is independently hydrogen, a substituted C₄ alkyl, an optionally substitituted C_2-6 alkenyl, an optionally substituted C_2-6 alkynyl, an optionally substituted C₄ alkenyl, an optionally substituted C₄ alkynyl, an optionally substituted cycloaliphatic, an optionally substituted heterocycloaliphatic, an optionally substituted heteroaryl, an unsubstituted phenyl, or a mono-substituted phenyl, or any two adjacent R₃ groups together with the atoms to which they are attached form an optionally substituted heterocycle.

0151Another aspect of the present invention provides a compound that is useful for modulating ABC transporter activity. The compound has formula II: <chemistry id="chem0051" num="0051"><img file="EP2007756B1_D0077.tif" /></chemistry> or a pharmaceutically acceptable salt thereof.

0152R₁, R₂, and ring A are defined above in formula I; R₃, R'₃, and p are defined above in formula Ia; and Z₁, Z₂, Z₃, Z₄, and Z₅ are defined above in formula Ib.

0153Another aspect of the present invention provides a compound that is useful for modulating ABC transporter activity. The compound has formula IIa: <chemistry id="chem0052" num="0052"><img file="EP2007756B1_D0078.tif" /></chemistry> or a pharmaceutically acceptable salt thereof.

0154R₁, R₂, and ring A are defined above in formula I; R₃, R'₃, and p are defined above in formula Ia; and Z₁, Z₂, Z₃, Z₄, and Z₅ are defined above in formula Ib.

0155Another aspect of the present invention provides a compound that is useful for modulating ABC transporter activity. The compound has formula IIb: <chemistry id="chem0053" num="0053"><img file="EP2007756B1_D0079.tif" /></chemistry> or a pharmaceutically acceptable salt thereof.

0156R₁, R₂, and ring A, are defined above in formula I; R₃, R'₃, and p are defined above in formula Ia; and Z₁, Z₂, Z₃, Z₄, and Z₅ are defined above in formula Ib.

0157Another aspect of the present invention provides a compound that is useful for modulating ABC transporter activity. The compound has formula IIc: <chemistry id="chem0054" num="0054"><img file="EP2007756B1_D0080.tif" /></chemistry> or a pharmaceutically acceptable salt thereof.

0158R₁, R₂ and n are defined above in formula I; and R₃, R'₃, and p are defined in formula Ia.

0159Another aspect of the present invention provides a compound that is useful for modulating ABC transporter activity. The compound has formula IId: <chemistry id="chem0055" num="0055"><img file="EP2007756B1_D0081.tif" /></chemistry> or a pharmaceutically acceptable salt thereof.

0160Both R₂ groups, together with the atoms to which they are attached form a group selected from: <chemistry id="chem0056" num="0056"><img file="EP2007756B1_D0082.tif" /></chemistry><chemistry id="chem0057" num="0057"><img file="EP2007756B1_D0083.tif" /></chemistry><chemistry id="chem0058" num="0058"><img file="EP2007756B1_D0084.tif" /></chemistry><chemistry id="chem0059" num="0059"><img file="EP2007756B1_D0085.tif" /></chemistry>

0161R'₃ is independently selected from one of the following: -H, -CH₃, -CH₂CH₃, -C(O)CH₃, -CH₂CH₂OH, -C(O)OCH₃, <chemistry id="chem0060" num="0060"><img file="EP2007756B1_D0086.tif" /></chemistry><chemistry id="chem0061" num="0061"><img file="EP2007756B1_D0087.tif" /></chemistry><chemistry id="chem0062" num="0062"><img file="EP2007756B1_D0088.tif" /></chemistry><chemistry id="chem0063" num="0063"><img file="EP2007756B1_D0089.tif" /></chemistry><chemistry id="chem0064" num="0064"><img file="EP2007756B1_D0090.tif" /></chemistry><chemistry id="chem0065" num="0065"><img file="EP2007756B1_D0091.tif" /></chemistry><chemistry id="chem0066" num="0066"><img file="EP2007756B1_D0092.tif" /></chemistry><chemistry id="chem0067" num="0067"><img file="EP2007756B1_D0093.tif" /></chemistry><chemistry id="chem0068" num="0068"><img file="EP2007756B1_D0094.tif" /></chemistry><chemistry id="chem0069" num="0069"><img file="EP2007756B1_D0095.tif" /></chemistry><chemistry id="chem0070" num="0070"><img file="EP2007756B1_D0096.tif" /></chemistry> and each R₃ is independently selected from -H, -CH₃, -CH₂OH, -CH₂CH₃, -CH₂CH₂OH, -CH₂CH₂CH₃, -NH₂, halo,-OCH₃, -CN, -CF₃, -C(O)OCH₂CH₃, -S(O)₂CH₃, -CH₂NH₂, -C(O)NH₂, <chemistry id="chem0071" num="0071"><img file="EP2007756B1_D0097.tif" /></chemistry><chemistry id="chem0072" num="0072"><img file="EP2007756B1_D0098.tif" /></chemistry><chemistry id="chem0073" num="0073"><img file="EP2007756B1_D0099.tif" /></chemistry><chemistry id="chem0074" num="0074"><img file="EP2007756B1_D0100.tif" /></chemistry><chemistry id="chem0075" num="0075"><img file="EP2007756B1_D0101.tif" /></chemistry><chemistry id="chem0076" num="0076"><img file="EP2007756B1_D0102.tif" /></chemistry><chemistry id="chem0077" num="0077"><img file="EP2007756B1_D0103.tif" /></chemistry><chemistry id="chem0078" num="0078"><img file="EP2007756B1_D0104.tif" /></chemistry><chemistry id="chem0079" num="0079"><img file="EP2007756B1_D0105.tif" /></chemistry>

IV. GENERIC SYNTHETIC SCHEMES

0162The compounds of formulae (I, Ic, Id, II, IIa, IIb, IIc, and IId) may be readily synthesized from commercially available or known starting materials by known methods. Exemplary synthetic routes to produce compounds of formulae (I, Ic, Id, II, IIa, IIb, IIc, and IId) are provided below in Schemes 1-22 below.

0163Preparation of the compounds of the invention is achieved by the coupling of a ring B amine with a ring A carboxylic acid as illustrated in Scheme 1. <chemistry id="chem0080" num="0080"><img file="EP2007756B1_D0106.tif" /></chemistry>

0164Referring to Scheme 1, the acid 1a may be converted to the corresponding acid chloride 1b using thionyl chloride in the presence of a catalystic amount of dimethylformamide. Reaction of the acid chloride with the amine <chemistry id="chem0081" num="0081"><img file="EP2007756B1_D0107.tif" /></chemistry> provides compounds of the invention 1. Alternatively, the acid 1a may be directly coupled to the amine using known coupling reagents such as, for example, HATU in the presence of triethylamine.

0165Preparation of the acids 1a may be achieved as illustrated in Scheme 2. <chemistry id="chem0082" num="0082"><img file="EP2007756B1_D0108.tif" /></chemistry><chemistry id="chem0083" num="0083"><img file="EP2007756B1_D0109.tif" /></chemistry>

0166Referring to Scheme 2, the nitrile 2a reacts with a suitable bromochloroalkane in the presence of sodium hydroxide and a phase tranfer catalyst such as butyltriethylammonium chloride to provide the intermediate 2b. Hydrolysis of the nitrile of 2b provides the acid 1a. In some instances, isolation of the intermediate 2b is unnecessary.

0167The phenylacetonitriles 2a are commercially available or may be prepared as illustrated in Scheme 3. <chemistry id="chem0084" num="0084"><img file="EP2007756B1_D0110.tif" /></chemistry>

0168Referring to Scheme 3, reaction of an aryl bromide 3a with carbon monoxide in the presence of methanol and tetrakis(triphenylphosphine)palladium (0) provides the ester 3b. Reduction of 3b with lithium aluminum hydride provides the alcohol 3c which is converted to the halide 3d with thionyl chloride. Reaction of 3d with sodium cyanide provides the nitrile 2a.

0169Other methods of producing the nitrile 2a are illustrated in schemes 4 and 5 below. <chemistry id="chem0085" num="0085"><img file="EP2007756B1_D0111.tif" /></chemistry><chemistry id="chem0086" num="0086"><img file="EP2007756B1_D0112.tif" /></chemistry><chemistry id="chem0087" num="0087"><img file="EP2007756B1_D0113.tif" /></chemistry>

0170Preparation of <chemistry id="chem0088" num="0088"><img file="EP2007756B1_D0114.tif" /></chemistry> components is illustrated in the schemes that follow. A number of methods for preparing ring B compounds wherein ring B is an indole have been reported. See for example <nplcit id="ncit0022" npl-type="s"><text>Angew. Chem. 2005, 44, 606</text></nplcit>; <nplcit id="ncit0023" npl-type="s"><text>J. Am. Chem. Soc. 2005, 127, 5342</text></nplcit>,); <nplcit id="ncit0024" npl-type="s"><text>J. Comb. Chem. 2005, 7, 130</text></nplcit>; <nplcit id="ncit0025" npl-type="s"><text>Tetrahedron 2006, 62, 3439</text></nplcit>; <nplcit id="ncit0026" npl-type="s"><text>J. Chem. Soc. Perkin Trans. 1, 2000, 1045</text></nplcit>.

0171One method for preparing <chemistry id="chem0089" num="0089"><img file="EP2007756B1_D0115.tif" /></chemistry> is illustrated in Scheme 6. <chemistry id="chem0090" num="0090"><img file="EP2007756B1_D0116.tif" /></chemistry><chemistry id="chem0091" num="0091"><img file="EP2007756B1_D0117.tif" /></chemistry>

0172Referring to Scheme 6, a nitroaniline 6a is converted to the hydrazine 6b using nitrous acid in the presence of HCl and stannous chloride. Reaction of 6b with an aldehyde or ketone CH₃C(O)R₃ provides the hydrazone 6c which on treatment with phophoric acid in toluene leads to a mixture of nitro indoles 6d and 6e. Catalytic hydrogenation in the presence of palladium on carbon provides a mixture of the amino indoles 6f and 6g which may be separated using know methods such as, for example, chromatography.

0173An alternative method is illustrated in scheme 7. <chemistry id="chem0092" num="0092"><img file="EP2007756B1_D0118.tif" /></chemistry><chemistry id="chem0093" num="0093"><img file="EP2007756B1_D0119.tif" /></chemistry><chemistry id="chem0094" num="0094"><img file="EP2007756B1_D0120.tif" /></chemistry><chemistry id="chem0095" num="0095"><img file="EP2007756B1_D0121.tif" /></chemistry><chemistry id="chem0096" num="0096"><img file="EP2007756B1_D0122.tif" /></chemistry><chemistry id="chem0097" num="0097"><img file="EP2007756B1_D0123.tif" /></chemistry><chemistry id="chem0098" num="0098"><img file="EP2007756B1_D0124.tif" /></chemistry><chemistry id="chem0099" num="0099"><img file="EP2007756B1_D0125.tif" /></chemistry><chemistry id="chem0100" num="0100"><img file="EP2007756B1_D0126.tif" /></chemistry><chemistry id="chem0101" num="0101"><img file="EP2007756B1_D0127.tif" /></chemistry><chemistry id="chem0102" num="0102"><img file="EP2007756B1_D0128.tif" /></chemistry><chemistry id="chem0103" num="0103"><img file="EP2007756B1_D0129.tif" /></chemistry><chemistry id="chem0104" num="0104"><img file="EP2007756B1_D0130.tif" /></chemistry><chemistry id="chem0105" num="0105"><img file="EP2007756B1_D0131.tif" /></chemistry><chemistry id="chem0106" num="0106"><img file="EP2007756B1_D0132.tif" /></chemistry><chemistry id="chem0107" num="0107"><img file="EP2007756B1_D0133.tif" /></chemistry><chemistry id="chem0108" num="0108"><img file="EP2007756B1_D0134.tif" /></chemistry><chemistry id="chem0109" num="0109"><img file="EP2007756B1_D0135.tif" /></chemistry><chemistry id="chem0110" num="0110"><img file="EP2007756B1_D0136.tif" /></chemistry><chemistry id="chem0111" num="0111"><img file="EP2007756B1_D0137.tif" /></chemistry><chemistry id="chem0112" num="0112"><img file="EP2007756B1_D0138.tif" /></chemistry><chemistry id="chem0113" num="0113"><img file="EP2007756B1_D0139.tif" /></chemistry><chemistry id="chem0114" num="0114"><img file="EP2007756B1_D0140.tif" /></chemistry><chemistry id="chem0115" num="0115"><img file="EP2007756B1_D0141.tif" /></chemistry><chemistry id="chem0116" num="0116"><img file="EP2007756B1_D0142.tif" /></chemistry><chemistry id="chem0117" num="0117"><img file="EP2007756B1_D0143.tif" /></chemistry><chemistry id="chem0118" num="0118"><img file="EP2007756B1_D0144.tif" /></chemistry><chemistry id="chem0119" num="0119"><img file="EP2007756B1_D0145.tif" /></chemistry><chemistry id="chem0120" num="0120"><img file="EP2007756B1_D0146.tif" /></chemistry><chemistry id="chem0121" num="0121"><img file="EP2007756B1_D0147.tif" /></chemistry><chemistry id="chem0122" num="0122"><img file="EP2007756B1_D0148.tif" /></chemistry><chemistry id="chem0123" num="0123"><img file="EP2007756B1_D0149.tif" /></chemistry><chemistry id="chem0124" num="0124"><img file="EP2007756B1_D0150.tif" /></chemistry><chemistry id="chem0125" num="0125"><img file="EP2007756B1_D0151.tif" /></chemistry><chemistry id="chem0126" num="0126"><img file="EP2007756B1_D0152.tif" /></chemistry><chemistry id="chem0127" num="0127"><img file="EP2007756B1_D0153.tif" /></chemistry><chemistry id="chem0128" num="0128"><img file="EP2007756B1_D0154.tif" /></chemistry><chemistry id="chem0129" num="0129"><img file="EP2007756B1_D0155.tif" /></chemistry><chemistry id="chem0130" num="0130"><img file="EP2007756B1_D0156.tif" /></chemistry><chemistry id="chem0131" num="0131"><img file="EP2007756B1_D0157.tif" /></chemistry><chemistry id="chem0132" num="0132"><img file="EP2007756B1_D0158.tif" /></chemistry><chemistry id="chem0133" num="0133"><img file="EP2007756B1_D0159.tif" /></chemistry><chemistry id="chem0134" num="0134"><img file="EP2007756B1_D0160.tif" /></chemistry><chemistry id="chem0135" num="0135"><img file="EP2007756B1_D0161.tif" /></chemistry><chemistry id="chem0136" num="0136"><img file="EP2007756B1_D0162.tif" /></chemistry><chemistry id="chem0137" num="0137"><img file="EP2007756B1_D0163.tif" /></chemistry><chemistry id="chem0138" num="0138"><img file="EP2007756B1_D0164.tif" /></chemistry><chemistry id="chem0139" num="0139"><img file="EP2007756B1_D0165.tif" /></chemistry>

0174In the schemes above, the radical R employed therein is a substituent, e.g., RW as defined hereinabove. One of skill in the art will readily appreciate that synthetic routes suitable for various substituents of the present invention are such that the reaction conditions and steps employed do not modify the intended substituents.

V. FORMULATIONS, ADMINISTRATIONS, AND USES

0175Accordingly, in another aspect of the present invention, pharmaceutically acceptable compositions are provided, wherein these compositions comprise any of the compounds as described herein, and optionally comprise a pharmaceutically acceptable carrier, adjuvant or vehicle. In certain embodiments, these compositions optionally further comprise one or more additional therapeutic agents.

0176It will also be appreciated that certain of the compounds of present invention can exist in free form for treatment, or where appropriate, as a pharmaceutically acceptable derivative or a prodrug thereof. According to the present invention, a pharmaceutically acceptable derivative or a prodrug includes, but is not limited to, pharmaceutically acceptable salts, esters, salts of such esters, or any other adduct or derivative which upon administration to a patient in need is capable of providing, directly or indirectly, a compound as otherwise described herein, or a metabolite or residue thereof.

0177As used herein, the term "pharmaceutically acceptable salt" refers to those salts which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of humans and lower animals without undue toxicity, irritation, allergic response and the like, and are commensurate with a reasonable benefit/risk ratio. A "pharmaceutically acceptable salt" means any salt or salt of an ester of a compound of this invention that, upon administration to a recipient, is capable of providing, either directly or indirectly, a compound of this invention or an inhibitorily active metabolite or residue thereof.

0178Pharmaceutically acceptable salts are well known in the art. For example, <nplcit id="ncit0027" npl-type="s"><text>S. M. Berge, et al. describes pharmaceutically acceptable salts in detail in J. Pharmaceutical Sciences, 1977, 66, 1-19</text></nplcit>, incorporated herein by reference. Pharmaceutically acceptable salts of the compounds of this invention include those derived from suitable inorganic and organic acids and bases. Examples of pharmaceutically acceptable, nontoxic acid addition salts are salts of an amino group formed with inorganic acids such as hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid and perchloric acid or with organic acids such as acetic acid, oxalic acid, maleic acid, tartaric acid, citric acid, succinic acid or malonic acid or by using other methods used in the art such as ion exchange. Other pharmaceutically acceptable salts include adipate, alginate, ascorbate, aspartate, benzenesulfonate, benzoate, bisulfate, borate, butyrate, camphorate, camphorsulfonate, citrate, cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate, formate, fumarate, glucoheptonate, glycerophosphate, gluconate, hemisulfate, heptanoate, hexanoate, hydroiodide, 2-hydroxy-ethanesulfonate, lactobionate, lactate, laurate, lauryl sulfate, malate, maleate, malonate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palmitate, pamoate, pectinate, persulfate, 3-phenylpropionate, phosphate, picrate, pivalate, propionate, stearate, succinate, sulfate, tartrate, thiocyanate, p-toluenesulfonate, undecanoate, valerate salts, and the like. Salts derived from appropriate bases include alkali metal, alkaline earth metal, ammonium and N⁺(C_1-4alkyl)₄ salts. This invention also envisions the quaternization of any basic nitrogen-containing groups of the compounds disclosed herein. Water or oil-soluble or dispersible products may be obtained by such quaternization. Representative alkali or alkaline earth metal salts include sodium, lithium, potassium, calcium, magnesium, and the like. Further pharmaceutically acceptable salts include, when appropriate, nontoxic ammonium, quaternary ammonium, and amine cations formed using counterions such as halide, hydroxide, carboxylate, sulfate, phosphate, nitrate, loweralkyl sulfonate and aryl sulfonate.

0179As described above, the pharmaceutically acceptable compositions of the present invention additionally comprise a pharmaceutically acceptable carrier, adjuvant, or vehicle, which, as used herein, includes any and all solvents, diluents, or other liquid vehicle, dispersion or suspension aids, surface active agents, isotonic agents, thickening or emulsifying agents, preservatives, solid binders, lubricants and the like, as suited to the particular dosage form desired. <nplcit id="ncit0028" npl-type="b"><text>Remington's Pharmaceutical Sciences, Sixteenth Edition, E. W. Martin (Mack Publishing Co., Easton, Pa., 1980</text></nplcit>) discloses various carriers used in formulating pharmaceutically acceptable compositions and known techniques for the preparation thereof. Except insofar as any conventional carrier medium is incompatible with the compounds of the invention, such as by producing any undesirable biological effect or otherwise interacting in a deleterious manner with any other component(s) of the pharmaceutically acceptable composition, its use is contemplated to be within the scope of this invention. Some examples of materials which can serve as pharmaceutically acceptable carriers include, but are not limited to, ion exchangers, alumina, aluminum stearate, lecithin, serum proteins, such as human serum albumin, buffer substances such as phosphates, glycine, sorbic acid, or potassium sorbate, partial glyceride mixtures of saturated vegetable fatty acids, water, salts or electrolytes, such as protamine sulfate, disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, zinc salts, colloidal silica, magnesium trisilicate, polyvinyl pyrrolidone, polyacrylates, waxes, polyethylene-polyoxypropylene-block polymers, wool fat, sugars such as lactose, glucose and sucrose; starches such as corn starch and potato starch; cellulose and its derivatives such as sodium carboxymethyl cellulose, ethyl cellulose and cellulose acetate; powdered tragacanth; malt; gelatin; talc; excipients such as cocoa butter and suppository waxes; oils such as peanut oil, cottonseed oil; safflower oil; sesame oil; olive oil; corn oil and soybean oil; glycols; such a propylene glycol or polyethylene glycol; esters such as ethyl oleate and ethyl laurate; agar; buffering agents such as magnesium hydroxide and aluminum hydroxide; alginic acid; pyrogen-free water; isotonic saline; Ringer's solution; ethyl alcohol, and phosphate buffer solutions, as well as other non-toxic compatible lubricants such as sodium lauryl sulfate and magnesium stearate, as well as coloring agents, releasing agents, coating agents, sweetening, flavoring and perfuming agents, preservatives and antioxidants can also be present in the composition, according to the judgment of the formulator.

0180In yet another aspect, the present invention provides a method of treating a condition, disease, or disorder implicated by ABC transporter activity. In certain embodiments, the present invention provides a method of treating a condition, disease, or disorder implicated by a deficiency of ABC transporter activity, the method comprising administering a composition comprising a compound of formulae (I, Ic, Id, II, IIa, IIb, IIc, and IId) to a subject, preferably a mammal, in need thereof.

0181In certain preferred embodiments, the present invention provides a method of treating Cystic fibrosis, Hereditary emphysema, Hereditary hemochromatosis, Coagulation-Fibrinolysis deficiencies, such as Protein C deficiency, Type 1 hereditary angioedema, Lipid processing deficiencies, such as Familial hypercholesterolemia, Type 1 chylomicronemia, Abetalipoproteinemia, Lysosomal storage diseases, such as I-cell disease/Pseudo-Hurler, Mucopolysaccharidoses, Sandhof/Tay-Sachs, Crigler-Najjar type II, Polyendocrinopathy/Hyperinsulinemia, Diabetes mellitus, Laron dwarfism, Myeloperoxidase deficiency, Primary hypoparathyroidism, Melanoma, Glycanosis CDG type 1, Hereditary emphysema, Congenital hyperthyroidism, Osteogenesis imperfecta, Hereditary hypofibrinogenemia, ACT deficiency, Diabetes insipidus (DI), Neurohypophyseal DI, Nephrogenic DI, Charcot-Marie Tooth syndrome, Pelizaeus-Merzbacher disease, neurodegenerative diseases such as Alzheimer's disease, Parkinson' disease, Amyotrophic lateral sclerosis, Progressive supranuclear palsy, Pick's disease, several polyglutamine neurological disorders asuch as Huntington, Spinocerebellar ataxia type I, Spinal and bulbar muscular atrophy, Dentatorubral pallidoluysian atrophy, and Myotonic dystrophy, as well as Spongiform encephalopathies, such as Hereditary Creutzfeldt-Jakob disease (due to Prion protein processing defect), Fabry disease, Gerstmann-Sträussler-Scheinker disease, secretory diarrhea, polycystic kidney disease, chronic obstructive pulmonary disease (COPD), dry eye disease, and Sjögren's Syndrome, comprising the step of administering to said mammal an effective amount of a composition comprising a compound of formulae (I, Ic, Id, II, IIa, IIb, IIc, and IId), or a preferred embodiment thereof as set forth above.

0182According to an alternative preferred embodiment, the present invention provides a method of treating cystic fibrosis comprising the step of administering to said mammal a composition comprising the step of administering to said mammal an effective amount of a composition comprising a compound of formulae (I, Ic, Id, II, IIa, IIb,IIc, and IId), or a preferred embodiment thereof as set forth above.

0183According to the invention an "effective amount" of the compound or pharmaceutically acceptable composition is that amount effective for treating or lessening the severity of one or more of Cystic fibrosis, Hereditary emphysema, Hereditary hemochromatosis, Coagulation-Fibrinolysis deficiencies, such as Protein C deficiency, Type 1 hereditary angioedema, Lipid processing deficiencies, such as Familial hypercholesterolemia, Type 1 chylomicronemia, Abetalipoproteinemia, Lysosomal storage diseases, such as I-cell disease/Pseudo-Hurler, Mucopolysaccharidoses, Sandhof/Tay-Sachs, Crigler-Najjar type II, Polyendocrinopathy/Hyperinsulinemia, Diabetes mellitus, Laron dwarfism, Myeloperoxidase deficiency, Primary hypoparathyroidism, Melanoma, Glycanosis CDG type 1, Hereditary emphysema, Congenital hyperthyroidism, Osteogenesis imperfecta, Hereditary hypofibrinogenemia, ACT deficiency, Diabetes insipidus (DI), Neurohypophyseal DI, Nephrogenic DI, Charcot-Marie Tooth syndrome, Pelizaeus-Merzbacher disease, neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease, Amyotrophic lateral sclerosis, Progressive supranuclear palsy, Pick's disease, several polyglutamine neurological disorders asuch as Huntington, Spinocerebellar ataxia type I, Spinal and bulbar muscular atrophy, Dentatorubral pallidoluysian atrophy, and Myotonic dystrophy, as well as Spongiform encephalopathies, such as Hereditary Creutzfeldt-Jakob disease, Fabry disease, Gerstmann-Sträussler-Scheinker disease, secretory diarrhea, polycystic kidney disease, chronic obstructive pulmonary disease (COPD), dry eye disease, and Sjögren's Syndrome.

0184The compounds and compositions, according to the method of the present invention, may be administered using any amount and any route of administration effective for treating or lessening the severity of one or more of Cystic fibrosis, Hereditary emphysema, Hereditary hemochromatosis, Coagulation-Fibrinolysis deficiencies, such as Protein C deficiency, Type 1 hereditary angioedema, Lipid processing deficiencies, such as Familial hypercholesterolemia, Type 1 chylomicronemia, Abetalipoproteinemia, Lysosomal storage diseases, such as I-cell disease/Pseudo-Hurler, Mucopolysaccharidoses, Sandhof/Tay-Sachs, Crigler-Najjar type II, Polyendocrinopathy/Hyperinsulemia, Diabetes mellitus, Laron dwarfism, Myleoperoxidase deficiency, Primary hypoparathyroidism, Melanoma, Glycanosis CDG type 1, Hereditary emphysema, Congenital hyperthyroidism, Osteogenesis imperfecta, Hereditary hypofibrinogenemia, ACT deficiency, Diabetes insipidus (DI), Neurophyseal DI, Neprogenic DI, Charcot-Marie Tooth syndrome, Perlizaeus-Merzbacher disease, neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease, Amyotrophic lateral sclerosis, Progressive supranuclear plasy, Pick's disease, several polyglutamine neurological disorders asuch as Huntington, Spinocerebullar ataxia type I, Spinal and bulbar muscular atrophy, Dentatorubal pallidoluysian, and Myotonic dystrophy, as well as Spongiform encephalopathies, such as Hereditary Creutzfeldt-Jakob disease, Fabry disease, Straussler-Scheinker disease, secretory diarrhea, polycystic kidney disease, chronic obstructive pulmonary disease (COPD), dry eye disease, and Sjögren's Syndrome. The exact amount required will vary from subject to subject, depending on the species, age, and general condition of the subject, the severity of the infection, the particular agent, its mode of administration, and the like. The compounds of the invention are preferably formulated in dosage unit form for ease of administration and uniformity of dosage. The expression "dosage unit form" as used herein refers to a physically discrete unit of agent appropriate for the patient to be treated. It will be understood, however, that the total daily usage of the compounds and compositions of the present invention will be decided by the attending physician within the scope of sound medical judgment. The specific effective dose level for any particular patient or organism will depend upon a variety of factors including the disorder being treated and the severity of the disorder; the activity of the specific compound employed; the specific composition employed; the age, body weight, general health, sex and diet of the patient; the time of administration, route of administration, and rate of excretion of the specific compound employed; the duration of the treatment; drugs used in combination or coincidental with the specific compound employed, and like factors well known in the medical arts. The term "patient", as used herein, means an animal, preferably a mammal, and most preferably a human.

0185The pharmaceutically acceptable compositions of this invention can be administered to humans and other animals orally, rectally, parenterally, intracisternally, intravaginally, intraperitoneally, topically (as by powders, ointments, or drops), bucally, as an oral or nasal spray, or the like, depending on the severity of the infection being treated. In certain embodiments, the compounds of the invention may be administered orally or parenterally at dosage levels of about 0.01 mg/kg to about 50 mg/kg and preferably from about 1 mg/kg to about 25 mg/kg, of subject body weight per day, one or more times a day, to obtain the desired therapeutic effect.

0186Liquid dosage forms for oral administration include, but are not limited to, pharmaceutically acceptable emulsions, microemulsions, solutions, suspensions, syrups and elixirs. In addition to the active compounds, the liquid dosage forms may contain inert diluents commonly used in the art such as, for example, water or other solvents, solubilizing agents and emulsifiers such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol, dimethylformamide, oils (in particular, cottonseed, groundnut, corn, germ, olive, castor, and sesame oils), glycerol, tetrahydrofurfuryl alcohol, polyethylene glycols and fatty acid esters of sorbitan, and mixtures thereof. Besides inert diluents, the oral compositions can also include adjuvants such as wetting agents, emulsifying and suspending agents, sweetening, flavoring, and perfuming agents.

0187Injectable preparations, for example, sterile injectable aqueous or oleaginous suspensions may be formulated according to the known art using suitable dispersing or wetting agents and suspending agents. The sterile injectable preparation may also be a sterile injectable solution, suspension or emulsion in a nontoxic parenterally acceptable diluent or solvent, for example, as a solution in 1,3-butanediol. Among the acceptable vehicles and solvents that may be employed are water, Ringer's solution, U.S.P. and isotonic sodium chloride solution. In addition, sterile, fixed oils are conventionally employed as a solvent or suspending medium. For this purpose any bland fixed oil can be employed including synthetic mono- or diglycerides. In addition, fatty acids such as oleic acid are used in the preparation of injectables.

0188The injectable formulations can be sterilized, for example, by filtration through a bacterial-retaining filter, or by incorporating sterilizing agents in the form of sterile solid compositions which can be dissolved or dispersed in sterile water or other sterile injectable medium prior to use.

0189In order to prolong the effect of a compound of the present invention, it is often desirable to slow the absorption of the compound from subcutaneous or intramuscular injection. This may be accomplished by the use of a liquid suspension of crystalline or amorphous material with poor water solubility. The rate of absorption of the compound then depends upon its rate of dissolution that, in turn, may depend upon crystal size and crystalline form. Alternatively, delayed absorption of a parenterally administered compound form is accomplished by dissolving or suspending the compound in an oil vehicle. Injectable depot forms are made by forming microencapsule matrices of the compound in biodegradable polymers such as polylactide-polyglycolide. Depending upon the ratio of compound to polymer and the nature of the particular polymer employed, the rate of compound release can be controlled. Examples of other biodegradable polymers include poly(orthoesters) and poly(anhydrides). Depot injectable formulations are also prepared by entrapping the compound in liposomes or microemulsions that are compatible with body tissues.

0190Compositions for rectal or vaginal administration are preferably suppositories which can be prepared by mixing the compounds of this invention with suitable non-irritating excipients or carriers such as cocoa butter, polyethylene glycol or a suppository wax which are solid at ambient temperature but liquid at body temperature and therefore melt in the rectum or vaginal cavity and release the active compound.

0191Solid dosage forms for oral administration include capsules, tablets, pills, powders, and granules. In such solid dosage forms, the active compound is mixed with at least one inert, pharmaceutically acceptable excipient or carrier such as sodium citrate or dicalcium phosphate and/or a) fillers or extenders such as starches, lactose, sucrose, glucose, mannitol, and silicic acid, b) binders such as, for example, carboxymethylcellulose, alginates, gelatin, polyvinylpyrrolidinone, sucrose, and acacia, c) humectants such as glycerol, d) disintegrating agents such as agar--agar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates, and sodium carbonate, e) solution retarding agents such as paraffin, f) absorption accelerators such as quaternary ammonium compounds, g) wetting agents such as, for example, cetyl alcohol and glycerol monostearate, h) absorbents such as kaolin and bentonite clay, and i) lubricants such as talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate, and mixtures thereof. In the case of capsules, tablets and pills, the dosage form may also comprise buffering agents.

0192Solid compositions of a similar type may also be employed as fillers in soft and hard-filled gelatin capsules using such excipients as lactose or milk sugar as well as high molecular weight polyethylene glycols and the like. The solid dosage forms of tablets, dragees, capsules, pills, and granules can be prepared with coatings and shells such as enteric coatings and other coatings well known in the pharmaceutical formulating art. They may optionally contain opacifying agents and can also be of a composition that they release the active ingredient(s) only, or preferentially, in a certain part of the intestinal tract, optionally, in a delayed manner. Examples of embedding compositions that can be used include polymeric substances and waxes. Solid compositions of a similar type may also be employed as fillers in soft and hard-filled gelatin capsules using such excipients as lactose or milk sugar as well as high molecular weight polethylene glycols and the like.

0193The active compounds can also be in microencapsulated form with one or more excipients as noted above. The solid dosage forms of tablets, dragees, capsules, pills, and granules can be prepared with coatings and shells such as enteric coatings, release controlling coatings and other coatings well known in the pharmaceutical formulating art. In such solid dosage forms the active compound may be admixed with at least one inert diluent such as sucrose, lactose or starch. Such dosage forms may also comprise, as is normal practice, additional substances other than inert diluents, e.g., tableting lubricants and other tableting aids such a magnesium stearate and microcrystalline cellulose. In the case of capsules, tablets and pills, the dosage forms may also comprise buffering agents. They may optionally contain opacifying agents and can also be of a composition that they release the active ingredient(s) only, or preferentially, in a certain part of the intestinal tract, optionally, in a delayed manner. Examples of embedding compositions that can be used include polymeric substances and waxes.

0194Dosage forms for topical or transdermal administration of a compound of this invention include ointments, pastes, creams, lotions, gels, powders, solutions, sprays, inhalants or patches. The active component is admixed under sterile conditions with a pharmaceutically acceptable carrier and any needed preservatives or buffers as may be required. Ophthalmic formulation, eardrops, and eye drops are also contemplated as being within the scope of this invention. Additionally, the present invention contemplates the use of transdermal patches, which have the added advantage of providing controlled delivery of a compound to the body. Such dosage forms are prepared by dissolving or dispensing the compound in the proper medium. Absorption enhancers can also be used to increase the flux of the compound across the skin. The rate can be controlled by either providing a rate controlling membrane or by dispersing the compound in a polymer matrix or gel.

0195As described generally above, the compounds of the invention are useful as modulators of ABC transporters. Thus, without wishing to be bound by any particular theory, the compounds and compositions are particularly useful for treating or lessening the severity of a disease, condition, or disorder where hyperactivity or inactivity of ABC transporters is implicated in the disease, condition, or disorder. When hyperactivity or inactivity of an ABC transporter is implicated in a particular disease, condition, or disorder, the disease, condition, or disorder may also be referred to as a "ABC transporter-mediated disease, condition or disorder". Accordingly, in another aspect, the present invention provides a method for treating or lessening the severity of a disease, condition, or disorder where hyperactivity or inactivity of an ABC transporter is implicated in the disease state.

0196The activity of a compound utilized in this invention as a modulator of an ABC transporter may be assayed according to methods described generally in the art and in the Examples herein.

0197It will also be appreciated that the compounds and pharmaceutically acceptable compositions of the present invention can be employed in combination therapies, that is, the compounds and pharmaceutically acceptable compositions can be administered concurrently with, prior to, or subsequent to, one or more other desired therapeutics or medical procedures. The particular combination of therapies (therapeutics or procedures) to employ in a combination regimen will take into account compatibility of the desired therapeutics and/or procedures and the desired therapeutic effect to be achieved. It will also be appreciated that the therapies employed may achieve a desired effect for the same disorder (for example, an inventive compound may be administered concurrently with another agent used to treat the same disorder), or they may achieve different effects (e.g., control of any adverse effects). As used herein, additional therapeutic agents that are normally administered to treat or prevent a particular disease, or condition, are known as "appropriate for the disease, or condition, being treated".

0198The amount of additional therapeutic agent present in the compositions of this invention will be no more than the amount that would normally be administered in a composition comprising that therapeutic agent as the only active agent. Preferably the amount of additional therapeutic agent in the presently disclosed compositions will range from about 50% to 100% of the amount normally present in a composition comprising that agent as the only therapeutically active agent.

0199The compounds of this invention or pharmaceutically acceptable compositions thereof may also be incorporated into compositions for coating an implantable medical device, such as prostheses, artificial valves, vascular grafts, stents and catheters. Accordingly, the present invention, in another aspect, includes a composition for coating an implantable device comprising a compound of the present invention as described generally above, and in classes and subclasses herein, and a carrier suitable for coating said implantable device. In still another aspect, the present invention includes an implantable device coated with a composition comprising a compound of the present invention as described generally above, and in classes and subclasses herein, and a carrier suitable for coating said implantable device. Suitable coatings and the general preparation of coated implantable devices are described in <patcit id="pcit0008" dnum="US6099562A"><text>US Patents 6,099,562</text></patcit>; <patcit id="pcit0009" dnum="US5886026A"><text>5,886,026</text></patcit>; and <patcit id="pcit0010" dnum="US5304121A"><text>5,304,121</text></patcit>. The coatings are typically biocompatible polymeric materials such as a hydrogel polymer, polymethyldisiloxane, polycaprolactone, polyethylene glycol, polylactic acid, ethylene vinyl acetate, and mixtures thereof. The coatings may optionally be further covered by a suitable topcoat of fluorosilicone, polysaccarides, polyethylene glycol, phospholipids or combinations thereof to impart controlled release characteristics in the composition.

0200Another aspect of the invention relates to modulating ABC transporter activity in a biological sample or a patient (e.g., in vitro or in vivo), which method comprises administering to the patient, or contacting said biological sample with a compound of formula I or a composition comprising said compound. The term "biological sample", as used herein, includes, without limitation, cell cultures or extracts thereof; biopsied material obtained from a mammal or extracts thereof; and blood, saliva, urine, feces, semen, tears, or other body fluids or extracts thereof.

0201Modulation of ABC transporter activity in a biological sample is useful for a variety of purposes that are known to one of skill in the art. Examples of such purposes include, but are not limited to, the study of ABC transporters in biological and pathological phenomena; and the comparative evaluation of new modulators of ABC transporters.

0202In yet another embodiment, a method of modulating activity of an anion channel in vitro or in vivo, is provided comprising the step of contacting said channel with a compound of formulae (I, Ic, Id, II, IIa, IIb, IIc, and IId). In preferred embodiments, the anion channel is a chloride channel or a bicarbonate channel. In other preferred embodiments, the anion channel is a chloride channel.

0203According to an alternative embodiment, the present invention provides a method of increasing the number of functional ABC transporters in a membrane of a cell, comprising the step of contacting said cell with a compound of formulae (I, Ic, Id, II, IIa, IIb, IIc, and IId). The term "functional ABC transporter" as used herein means an ABC transporter that is capable of transport activity. In preferred embodiments, said functional ABC transporter is CFTR.

0204According to another preferred embodiment, the activity of the ABC transporter is measured by measuring the transmembrane voltage potential. Means for measuring the voltage potential across a membrane in the biological sample may employ any of the known methods in the art, such as optical membrane potential assay or other electrophysiological methods.

0205The optical membrane potential assay utilizes voltage-sensitive FRET sensors described by Gonzalez and Tsien (See, <nplcit id="ncit0029" npl-type="s"><text>Gonzalez, J. E. and R. Y. Tsien (1995) "Voltage sensing by fluorescence resonance energy transfer in single cells" Biophys J 69(4): 1272-80</text></nplcit>, and <nplcit id="ncit0030" npl-type="s"><text>Gonzalez, J. E. and R. Y. Tsien (1997) "Improved indicators of cell membrane potential that use fluorescence resonance energy transfer" Chem Biol 4(4): 269-77</text></nplcit>) in combination with instrumentation for measuring fluorescence changes such as the Voltage/Ion Probe Reader (VIPR) (See, <nplcit id="ncit0031" npl-type="s"><text>Gonzalez, J. E., K. Oades, et al. (1999) "Cell-based assays and instrumentation for screening ion-channel targets" Drug Discov Today 4(9): 431-439</text></nplcit>).

0206These voltage sensitive assays are based on the change in fluorescence resonant energy transfer (FRET) between the membrane-soluble, voltage-sensitive dye, DiSBAC₂(3), and a fluorescent phospholipid, CC2-DMPE, which is attached to the outer leaflet of the plasma membrane and acts as a FRET donor. Changes in membrane potential (V_m) cause the negatively charged DiSBAC₂(3) to redistribute across the plasma membrane and the amount of energy transfer from CC2-DMPE changes accordingly. The changes in fluorescence emission can be monitored using VIPR™ II, which is an integrated liquid handler and fluorescent detector designed to conduct cell-based screens in 96- or 384-well microtiter plates.

0207In another aspect the present invention provides a kit for use in measuring the activity of a ABC transporter or a fragment thereof in a biological sample in vitro or in vivo comprising (i) a composition comprising a compound of formulae (I, Ic, Id, II, IIa, IIb, IIc, and IId) or any of the above embodiments; and (ii) instructions for a.) contacting the composition with the biological sample and b.) measuring activity of said ABC transporter or a fragment thereof. In one embodiment, the kit further comprises instructions for a.) contacting an additional composition with the biological sample; b.) measuring the activity of said ABC transporter or a fragment thereof in the presence of said additional compound, and c.) comparing the activity of the ABC transporter in the presence of the additional compound with the density of the ABC transporter in the presence of a composition of formulae (I, Ic, Id, II, IIa, IIb,IIc, and IId). In preferred embodiments, the kit is used to measure the density of CFTR.

0208In order that the invention described herein may be more fully understood, the following examples are set forth. It should be understood that these examples are for illustrative purposes only and are not to be construed as limiting this invention in any manner.

VI. PREPARATIONS AND EXAMPLES

General Procedure I: Carboxylic Acid Building Block

0209<chemistry id="chem0140" num="0140"><img file="EP2007756B1_D0166.tif" /></chemistry>

0210Benzyltriethylammonium chloride (0.025 equivalents) and the appropriate dihalo compound (2.5 equivalents) were added to a substituted phenyl acetonitrile. The mixture was heated at 70 °C and then 50 % sodium hydroxide (10 equivalents) was slowly added to the mixture. The reaction was stirred at 70 °C for 12-24 hours to ensure complete formation of the cycloalkyl moiety and then heated at 130 °C for 24-48 hours to ensure complete conversion from the nitrile to the carboxylic acid. The dark brown / black reaction mixture was diluted with water and extracted with dichloromethane three times to remove side products. The basic aqueous solution was acidified with concentrated hydrochloric acid to pH less than one and the precipitate which began to form at pH 4 was filtered and washed with 1 M hydrochloric acid two times. The solid material was dissolved in dichloromethane and extracted two times with 1 M hydrochloric acid and one time with a saturated aqueous solution of sodium chloride. The organic solution was dried over sodium sulfate and evaporated to dryness to give the cycloalkylcarboxylic acid. Yields and purities were typically greater than 90%.

Example 1: 1-Benzo[1,3]dioxol-5-yl-cyclopropanecarboxylic acid

0211<chemistry id="chem0141" num="0141"><img file="EP2007756B1_D0167.tif" /></chemistry>

0212A mixture of 2-(benzo[d][1,3]dioxol-5-yl)acetonitrile (5.10 g 31.7 mmol), 1-bromo-2-chloro-ethane (9.00 mL 109 mmol), and benzyltriethylammonium chloride (0.181 g, 0.795 mmol) was heated at 70 °C and then 50% (wt./wt.) aqueous sodium hydroxide (26 mL) was slowly added to the mixture. The reaction was stirred at 70 °C for 24 hours and then heated at 130 °C for 48 hours. The dark brown reaction mixture was diluted with water (400 mL) and extracted once with an equal volume of ethyl acetate and once with an equal volume of dichloromethane. The basic aqueous solution was acidified with concentrated hydrochloric acid to pH less than one and the precipitate filtered and washed with 1 M hydrochloric acid. The solid material was dissolved in dichloromethane (400 mL) and extracted twice with equal volumes of 1 M hydrochloric acid and once with a saturated aqueous solution of sodium chloride. The organic solution was dried over sodium sulfate and evaporated to dryness to give a white to slightly off-white solid (5.23 g, 80%) ESI-MS <i>m</i>/<i>z</i> calc. 206.1, found 207.1 (M+1)⁺. Retention time 2.37 minutes. ¹H NMR (400 MHz, DMSO-d₆) δ 1.07-1.11 (m, 2H), 1.38-1.42 (m, 2H), 5.98 (s, 2H), 6.79 (m, 2H), 6.88 (m, 1H), 12.26 (s, 1H).

General Procedure II: Carboxylic Acid Building Block

0213<chemistry id="chem0142" num="0142"><img file="EP2007756B1_D0168.tif" /></chemistry>

0214Sodium hydroxide (50 % aqueous solution, 7.4 equivalents) was slowly added to a mixture of the appropriate phenyl acetonitrile, benzyltriethylammonium chloride (1.1 equivalents), and the appropriate dihalo compound (2.3 equivalents) at 70 °C. The mixture was stirred overnight at 70 °C and the reaction mixture was diluted with water (30 mL) and extracted with ethyl acetate. The combined organic layers were dried over sodium sulfate and evaporated to dryness to give the crude cyclopropanecarbonitrile, which was used directly in the next step.

0215The crude cyclopropanecarbonitrile was refluxed in 10% aqueous sodium hydroxide (7.4 equivalents) for 2.5 hours. The cooled reaction mixture was washed with ether (100 mL) and the aqueous phase was acidified to pH 2 with 2M hydrochloric acid. The precipitated solid was filtered to give the cyclopropanecarboxylic acid as a white solid.

General Procedure III: Carboxylic Acid Building Block

0216<chemistry id="chem0143" num="0143"><img file="EP2007756B1_D0169.tif" /></chemistry>

Example 2: 1-(2,2-Difluoro-benzo[1,3]dioxol-5-yl)-cyclopropanecarboxylic

acid

0217<chemistry id="chem0144" num="0144"><img file="EP2007756B1_D0170.tif" /></chemistry>

2,2-Difluoro-benzo[1,3]dioxole-5-carboxylic acid methyl ester

0218A solution of 5-bromo-2,2-difluoro-benzo[1,3]dioxole (11.8 g, 50.0 mmol) and tetrakis(triphenylphosphine)palladium (0) [Pd(PPh₃)₄, 5.78 g, 5.00 mmol] in methanol (20 mL) containing acetonitrile (30 mL) and triethylamine (10 mL) was stirred under a carbon monoxide atmosphere (55 PSI) at 75 °C (oil bath temperature) for 15 hours. The cooled reaction mixture was filtered and the filtrate was evaporated to dryness. The residue was purified by silica gel column chromatography to give crude 2,2-difluoro-benzo [1,3] dioxole-5-carboxylic acid methyl ester (11.5 g), which was used directly in the next step. <chemistry id="chem0145" num="0145"><img file="EP2007756B1_D0171.tif" /></chemistry>

(2,2-Difluoro-benzo[1,3]dioxol-5-yl)-methanol

0219Crude 2,2-difluoro-benzo[1,3]dioxole-5-carboxylic acid methyl ester (11.5 g) dissolved in 20 mL of anhydrous tetrahydrofuran (THF) was slowly added to a suspension of lithium aluminum hydride (4.10 g, 106 mmol) in anhydrous THF (100 mL) at 0 °C. The mixture was then warmed to room temperature. After being stirred at room temperature for 1 hour, the reaction mixture was cooled to 0 °C and treated with water (4.1 g), followed by sodium hydroxide (10% aqueous solution, 4.1 mL). The resulting slurry was filtered and washed with THF. The combined filtrate was evaporated to dryness and the residue was purified by silica gel column chromatography to give (2,2-difluoro-benzo[1,3]dioxol-5-yl)-methanol (7.2 g, 38 mmol, 76 % over two steps) as a colorless oil. <chemistry id="chem0146" num="0146"><img file="EP2007756B1_D0172.tif" /></chemistry>

5-Chloromethyl-2,2-difluoro-benzo[1,3]dioxole

0220Thionyl chloride (45 g, 38 mmol) was slowly added to a solution of (2,2-difluoro-benzo[1,3]dioxol-5-yl)-methanol (7.2 g, 38 mmol) in dichloromethane (200 mL) at 0 °C. The resulting mixture was stirred overnight at room temperature and then evaporated to dryness. The residue was partitioned between an aqueous solution of saturated sodium bicarbonate (100 mL) and dichloromethane (100 mL). The separated aqueous layer was extracted with dichloromethane (150 mL) and the organic layer was dried over sodium sulfate, filtrated, and evaporated to dryness to give crude 5-chloromethyl-2,2-difluoro-benzo[1,3]dioxole (4.4 g) which was used directly in the next step. <chemistry id="chem0147" num="0147"><img file="EP2007756B1_D0173.tif" /></chemistry>

(2,2-Difluoro-benzo[1,3]dioxol-5-yl)-acetonitrile

0221A mixture of crude 5-chloromethyl-2,2-difluoro-benzo[1,3]dioxole (4.4 g) and sodium cyanide (1.36 g, 27.8 mmol) in dimethylsulfoxide (50 mL) was stirred at room temperature overnight. The reaction mixture was poured into ice and extracted with ethyl acetate (300 mL). The organic layer was dried over sodium sulfate and evaporated to dryness to give crude (2,2-difluoro-benzo[1,3]dioxol-5-yl)-acetonitrile (3.3 g) which was used directly in the next step. <chemistry id="chem0148" num="0148"><img file="EP2007756B1_D0174.tif" /></chemistry>

1-(2,2-Difluoro-benzo[1,3]dioxol-5-yl)-cyclopropanecarbonitrile

0222Sodium hydroxide (50% aqueous solution, 10 mL) was slowly added to a mixture of crude (2,2-difluoro-benzo[1,3]dioxol-5-yl)-acetonitrile, benzyltriethylammonium chloride (3.00 g, 15.3 mmol), and 1-bromo-2-chloroethane (4.9 g, 38 mmol) at 70 °C.

0223The mixture was stirred overnight at 70 °C before the reaction mixture was diluted with water (30 mL) and extracted with ethyl acetate. The combined organic layers were dried over sodium sulfate and evaporated to dryness to give crude 1-(2,2-difluoro-benzo[1,3]dioxol-5-yl)-cyclopropanecarbonitrile, which was used directly in the next step. <chemistry id="chem0149" num="0149"><img file="EP2007756B1_D0175.tif" /></chemistry>

1-(2,2-Difluoro-benzo[1,3]dioxol-5-yl)-cyclopropanecarboxylic acid

02241-(2,2-Difluoro-benzo[1,3]dioxol-5-yl)-cyclopropanecarbonitrile (crude from the last step) was refluxed in 10% aqueous sodium hydroxide (50 mL) for 2.5 hours. The cooled reaction mixture was washed with ether (100 mL) and the aqueous phase was acidified to pH 2 with 2M hydrochloric acid. The precipitated solid was filtered to give 1-(2,2-difluoro-benzo[1,3]dioxol-5-yl)-cyclopropanecarboxylic acid as a white solid (0.15 g, 1.6% over four steps). ESI-MS <i>m</i>/<i>z</i> calc. 242.04, found 241.58 (M+1)⁺; ¹H NMR (CDCl₃) δ 7.14-7.04 (m, 2H), 6.98-6.96 (m, 1H), 1.74-1.64 (m, 2H), 1.26-1.08 (m, 2H).

Example 3: 2-(2,2-Dimethylbenzo[d][1,3]dioxol-5-yl)acetonitrile

0225<chemistry id="chem0150" num="0150"><img file="EP2007756B1_D0176.tif" /></chemistry>

(3,4-Dihydroxy-phenyl)-acetonitrile

0226To a solution of benzo[1,3]dioxol-5-yl-acetonitrile (0.50 g, 3.1 mmol) in CH₂Cl₂ (15 mL) was added dropwise BBr₃ (0.78 g, 3.1 mmol) at -78 °C under N₂. The mixture was slowly warmed to room temperature and stirred overnight. H₂O (10 mL) was added to quench the reaction and the CH₂Cl₂ layer was separated. The aqueous phase was extracted with CH₂Cl₂ (2 × 7 mL). The combined organics were washed with brine, dried over Na₂SO₄ and purified by column chromatography on silica gel (petroleum ether/ethyl acetate 5:1) to give (3,4-dihydroxy-phenyl)-acetonitrile (0.25 g, 54%) as a white solid. ¹H NMR (DMSO-<i>d</i>₆, 400 MHz) δ 9.07 (s, 1 H), 8.95 (s, 1 H), 6.68-6.70 (m, 2 H), 6.55 (dd, <i>J=</i> 8.0, 2.0 Hz, 1 H), 3.32 (s, 2 H). <chemistry id="chem0151" num="0151"><img file="EP2007756B1_D0177.tif" /></chemistry>

2-(2,2-Dimethylbenzo[d][1,3]dioxol-5-yl)acetonitrile

0227To a solution of (3,4-dihydroxy-phenyl)-acetonitrile (0.20 g, 1.3 mmol) in toluene (4 mL) was added 2,2-dimethoxy-propane (0.28 g, 2.6 mmol) and TsOH (0.010 g, 0.065 mmol). The mixture was heated at reflux overnight. The reaction mixture was evaporated to remove the solvent and the residue was dissolved in ethyl acetate. The organic layer was washed with NaHCO₃ solution, H₂O, brine, and dried over Na₂SO₄. The solvent was evaporated under reduced pressure to give a residue, which was purified by column chromatography on silica gel (petroleum ether/ethyl acetate 10:1) to give 2-(2,2-dimethylbenzo[d][1,3]dioxol-5-yl)acetonitrile (40 mg, 20%). ¹H NMR (CDCl₃, 400 MHz) δ 6.68-6.71 (m, 3 H), 3.64 (s, 2 H), 1.67 (s, 6 H).

Example 4: 1-(3,4-Dihydroxy-phenyl)-cyclopropanecarboxylic acid

0228<chemistry id="chem0152" num="0152"><img file="EP2007756B1_D0178.tif" /></chemistry>

1-(3,4-Bis-benzyloxy-phenyl)-cyclopropanecarbonitrile

0229To a mixture of (n-C₄H₉)₄NBr (0.50 g, 1.5 mmol), toluene (7 mL) and (3,4-bis-benzyloxy-phenyl)-acetonitrile (14 g, 42 mmol) in NaOH (50 g) and H₂O (50 mL) was added BrCH₂CH₂Cl (30 g, 0.21 mol). The reaction mixture was stirred at 50 °C for 5 h before being cooled to room temperature. Toluene (30 mL) was added and the organic layer was separated and washed with H₂O, brine, dried over anhydrous MgSO₄, and concentrated. The residue was purified by column on silica gel (petroleum ether/ethyl acetate 10:1) to give 1-(3,4-bis-benzyloxy-phenyl)-cydopropanecarbonitrile (10 g, 66%). ¹H NMR (DMSO 300 MHz) δ 7.46-7.30 (m, 10 H), 7.03 (d, <i>J=</i> 8.4 Hz, 1 H), 6.94 (d, <i>J=</i> 2.4 Hz, 1 H), 6.89 (dd, <i>J=</i> 2.4, 8.4 Hz, 1 H), 5.12 (d, <i>J =</i> 7.5 Hz, 4H), 1.66-1.62 (m, 2 H), 1.42-1.37 (m, 2 H). <chemistry id="chem0153" num="0153"><img file="EP2007756B1_D0179.tif" /></chemistry>

1-(3,4-Dihydroxy-phenyl)-cyclopropanecarbonitrile

0230To a solution of 1-(3,4-bis-benzyloxy-phenyl)-cyclopropanecarbonitrile (10 g, 28 mmol) in MeOH (50 mL) was added Pd/C (0.5 g) under nitrogen atmosphere. The mixture was stirred under hydrogen atmosphere (1 atm) at room temperature for 4 h. The catalyst was filtered off through a celite pad and the filtrate was evaporated under <i>vacuum</i> to give 1-(3,4-dihydroxy-phenyl)-cyclopropanecarbonitrile (4.5 g, 92%). ¹H NMR (DMSO 400 MHz) δ 9.06 (br s, 2 H), 6.67-6.71 (m, 2 H), 6.54 (dd, <i>J =</i> 2.4, 8.4 Hz, 1 H), 1.60-1.57 (m, 2 H), 1.30-1.27 (m, 2 H). <chemistry id="chem0154" num="0154"><img file="EP2007756B1_D0180.tif" /></chemistry>

1-(3,4-Dihydroxy-phenyl)-cyclopropanecarboxylic acid

0231To a solution of NaOH (20 g, 0.50 mol) in H₂O (20 mL) was added 1-(3,4-dihydroxy-phenyl)-cyclopropanecarbonitrile (4.4 g, 25 mmol). The mixture was heated at reflux for 3 h before being cooled to room temperature. The mixture was neutralized with HCl (0.5 N) to pH 3-4 and extracted with ethyl acetate (20 mL × 3). The combined organic layers were washed with water, brine, dried over anhydrous MgSO4, and concentrated under <i>vacuum</i> to obtain 1-(3,4-dihydroxy-phenyl)-cyclopropanecarboxylic acid (4.5 g crude). From 900 mg crude, 500 mg pure 1-(3,4-dihydroxy-phenyl)-cyclopropanecarboxylic acid was obtained by preparatory HPLC. ¹H NMR (DMSO 300 MHz) δ 12.09 (br s, 1 H), 8.75 (br s, 2 H), 6.50-6.67 (m, 3 H), 1.35-1.31 (m, 2 H), 1.01-0.97 (m, 2 H).

Example 5: 1-(2-Oxo-2,3-dihydrobenzo[d]oxazol-5-yl)cyclopropane-carboxylic acid.

0232<chemistry id="chem0155" num="0155"><img file="EP2007756B1_D0181.tif" /></chemistry>

1-(4-Methoxy-phenyl)-cyclopropanecarboxylic acid methyl ester

0233To a solution of 1-(4-methoxy-phenyl)-cyclopropanecarboxylic acid (50 g, 0.26 mol) in MeOH (500 mL) was added toluene-4-sulfonic acid monohydrate (2.5 g, 13 mmol) at room temperature. The reaction mixture was heated at reflux for 20 hours. MeOH was removed by evaporation under <i>vacuum</i> and EtOAc (200 mL) was added. The organic layer was washed with sat. aq. NaHCO₃ (100 mL) and brine, dried over anhydrous Na₂SO₄ and evaporated under <i>vacuum</i> to give 1-(4-methoxy-phenyl)-cyclopropanecarboxylic acid methyl ester (53 g, 99%). ¹H NMR (CDCl₃ 400 MHz) δ 7.25-7.27 (m, 2 H), 6.85 (d, J= 8.8 Hz, 2 H), 3.80 (s, 3 H), 3.62 (s, 3 H), 1.58 (q, <i>J</i> = 3.6 Hz, 2 H), 1.15 (q, <i>J</i> = 3.6 Hz, 2 H). <chemistry id="chem0156" num="0156"><img file="EP2007756B1_D0182.tif" /></chemistry>

1-(4-Methoxy-3-nitro-phenyl)-cyclopropanecarboxylic acid methyl ester

0234To a solution of 1-(4-methoxy-phenyl)-cyclopropanecarboxylic acid methyl ester (30.0 g, 146 mmol) in Ac₂O (300 mL) was added a solution of HNO₃ (14.1 g, 146 mmol, 65%) in AcOH (75 mL) at 0 °C. The reaction mixture was stirred at 0 ∼ 5 °C for 3 h before aq. HCl (20%) was added dropwise at 0 °C. The resulting mixture was extracted with EtOAc (200 mL × 3). The organic layer was washed with sat. aq. NaHCO₃ then brine, dried over anhydrous Na₂SO₄ and evaporated under <i>vacuum</i> to give 1-(4-methoxy-3-nitro-phenyl)-cyclopropanecarboxylic acid methyl ester (36.0 g, 98%), which was directly used in the next step. ¹H NMR (CDCl₃, 300 MHz) δ 7.84 (d, <i>J=</i> 2.1 Hz, 1 H), 7.54 (dd, <i>J=</i> 2.1, 8.7 Hz, 1 H), 7.05 (d, <i>J</i> = 8.7 Hz, 1 H), 3.97 (s, 3 H), 3.65 (s, 3 H), 1.68-1.64 (m, 2 H), 1.22-1.18 (m, 2 H). <chemistry id="chem0157" num="0157"><img file="EP2007756B1_D0183.tif" /></chemistry>

1-(4-Hydroxy-3-nitro-phenyl)-cyclopropanecarboxylic acid methyl ester

0235To a solution of 1-(4-methoxy-3-nitro-phenyl)-cyclopropane-carboxylic acid methyl ester (10.0 g, 39.8 mmol) in CH₂Cl₂ (100 mL) was added BBr₃ (12.0 g, 47.8 mmol) at -70 °C. The mixture was stirred at -70 °C for 1 hour, then allowed to warm to -30 °C and stirred at this temperature for 3 hours. Water (50 mL) was added dropwise at -20 °C, and the resulting mixture was allowed to warm room temperature before it was extracted with EtOAc (200 mL × 3). The combined organic layers were dried over anhydrous Na₂SO₄ and evaporated under <i>vacuum</i> to give the crude product, which was purified by column chromatography on silica gel (petroleum ether/ethyl acetate 15:1) to afford 1-(4-hydroxy-3-nitro-phenyl)-cyclopropanecarboxylic acid methyl ester (8.3 g, 78%). ¹H NMR (CDCl₃, 400 MHz) δ 10.5 (s, 1 H), 8.05 (d, <i>J</i> = 2.4 Hz, 1 H), 7.59 (dd, <i>J=</i> 2.0, 8.8 Hz, 1 H), 7.11 (d, <i>J</i> = 8.4 Hz, 1 H), 3.64 (s, 3 H), 1.68-1.64 (m, 2 H), 1.20-1.15 (m, 2 H). <chemistry id="chem0158" num="0158"><img file="EP2007756B1_D0184.tif" /></chemistry>

1-(3-Amino-4-hydroxy-phenyl)-cyclopropanecarboxylic acid methyl ester

0236To a solution of 1-(4-hydroxy-3-nitro-phenyl)-cyclopropanecarboxylic acid methyl ester (8.3 g, 35 mmol) in MeOH (100 mL) was added Raney Nickel (0.8 g) under nitrogen atmosphere. The mixture was stirred under hydrogen atmosphere (1 atm) at 35 °C for 8 hours. The catalyst was filtered off through a Celite pad and the filtrate was evaporated under <i>vacuum</i> to give crude product, which was purified by column chromatography on silica gel (petroleum ether/ethyl acetate 1:1) to give 1-(3-amino-4-hydroxy-phenyl)-cyclopropanecarboxylic acid methyl ester (5.3 g, 74%). ¹H NMR (CDCl₃, 400 MHz) δ 6.77 (s, 1 H), 6.64 (d, <i>J</i>= 2.0 Hz, 2 H), 3.64 (s, 3 H), 1.55-1.52 (m, 2 H), 1.15-1.12 (m, 2 H). <chemistry id="chem0159" num="0159"><img file="EP2007756B1_D0185.tif" /></chemistry>

1-(2-Oxo-2,3-dihydro-benzooxazol-5-yl)-cyclopropanecarboxylic acid methyl ester

0237To a solution of 1-(3-amino-4-hydroxy-phenyl)-cyclopropanecarboxylic acid methyl ester (2.0 g, 9.6 mmol) in THF (40 mL) was added triphosgene (4.2 g, 14 mmol) at room temperature. The mixture was stirred for 20 minutes at this temperature before water (20 mL) was added dropwise at 0 °C. The resulting mixture was extracted with EtOAc (100 mL × 3). The combined organic layers were dried over anhydrous Na₂SO₄ and evaporated under <i>vacuum</i> to give 1-(2-oxo-2,3-dihydro-benzooxazol-5-yl)-cyclopropanecarboxylic acid methyl ester (2.0 g, 91%), which was directly used in the next step. ¹H NMR (CDCl₃, 300 MHz) δ 8.66 (s, 1 H), 7.13-7.12 (m, 2 H), 7.07 (s, 1 H), 3.66 (s, 3 H), 1.68-1.65 (m, 2 H), 1.24-1.20 (m, 2 H). <chemistry id="chem0160" num="0160"><img file="EP2007756B1_D0186.tif" /></chemistry>

1-(2-Oxo-2,3-dihydrobenzo[d]oxazol-5-yl)cyclopropanecarboxylic acid

0238To a solution of 1-(2-oxo-2,3-dihydro-benzooxazol-5-yl)-cyclopropanecarboxylic acid methyl ester (1.9 g, 8.1 mmol) in MeOH (20 mL) and water (2 mL) was added LiOH.H₂O (1.7 g, 41 mmol) in portions at room temperature. The reaction mixture was stirred for 20 hours at 50 °C. MeOH was removed by evaporation under <i>vacuum</i> before water (100 mL) and EtOAc (50 mL) were added. The aqueous layer was separated, acidified with HCl (3 mol/L) and extracted with EtOAc (100 mL × 3). The combined organic layers were dried over anhydrous Na₂SO₄ and evaporated under <i>vacuum</i> to give 1-(2-oxo-2,3-dihydrobenzo[d]oxazol-5-yl)cyclopropanecarboxylic acid (1.5 g, 84%). ¹H NMR (DMSO, 400 MHz) δ 12.32 (brs, 1 H), 11.59 (brs, 1 H), 7.16 (d, <i>J</i>= 8.4 Hz, 1 H), 7.00 (d, <i>J</i>= 8.0 Hz, 1 H), 1.44-1.41 (m, 2 H), 1.13-1.10 (m, 2 H). MS (ESI) m/e (M+H⁺) 218.1.

Example 6: 1-(6-Fluoro-benzo[1,3]dioxol-5-yl)-cyclopropanecarboxylic acid

0239<chemistry id="chem0161" num="0161"><img file="EP2007756B1_D0187.tif" /></chemistry>

2-Fluoro-4,5-dihydroxy-benzaldehyde

0240To a stirred suspension of 2-fluoro-4,5-dimethoxy-benzaldehyde (3.00 g, 16.3 mmol) in dichloromethane (100 mL) was added BBr₃ (12.2 mL, 130 mmol) dropwise at -78 °C under nitrogen atmosphere. After addition, the mixture was warmed to -30 °C and stirred at this temperature for 5 h. The reaction mixture was poured into ice water and the precipitated solid was collected by filtration and washed with dichloromethane to afford 2-fluoro-4,5-dihydroxy-benzaldehyde (8.0 g), which was used directly in the next step. <chemistry id="chem0162" num="0162"><img file="EP2007756B1_D0188.tif" /></chemistry>

0241<b>6-Fluoro-benzo[1,3]dioxole-5-carbaldehyde</b> To a stirred solution of 2-fluoro-4,5-dihydroxy-benzaldehyde (8.0 g) and BrClCH₂ (24.8 g, 190 mmol) in dry DMF (50 mL) was added Cs₂CO₃ (62.0 g, 190 mmol) in portions. The resulting mixture was stirred at 60 °C overnight and then poured into water. The mixture was extracted with EtOAc (200 mL × 3). The combined organic layers were washed with brine (200 mL), dried over Na₂SO₄, and evaporated <i>in vacuo</i> to give crude product, which was purified by column chromatography on silica gel (5-20% ethyl acetate/petroleum ether) to afford 6-fluoro-benzo[1,3]dioxole-5-carbaldehyde (700 mg, two steps yield: 24%). ¹H-NMR (400 MHz, CDCl₃) δ 10.19 (s, 1 H), 7.23 (d, <i>J=</i> 5.6, 1 H), 6.63 (d, <i>J</i>= 9.6, 1 H), 6.08 (s, 2 H). <chemistry id="chem0163" num="0163"><img file="EP2007756B1_D0189.tif" /></chemistry>

(6-Fluoro-benzo[1,3]dioxol-5-yl)-methanol

0242To a stirred solution of 6-fluoro-benzo[1,3]dioxole-5-carbaldehyde (700 mg, 4.2 mmol) in MeOH (50 mL) was added NaBH₄ (320 mg, 8.4 mmol) in portions at 0 °C. The mixture was stirred at this temperature for 30 min and was then concentrated <i>in vacuo</i> to give a residue. The residue was dissolved in EtOAc and the organic layer was washed with water, dried over Na₂SO4, and concentrated in <i>vacuo</i> to afford (6-fluoro-benzo[1,3]dioxol-5-yl)-methanol (650 mg, 92%), which was directly used in the next step. <chemistry id="chem0164" num="0164"><img file="EP2007756B1_D0190.tif" /></chemistry>

5-Chloromethyl-6-fluoro-benzo[1,3]dioxole

0243(6-Fluoro-benzo[1,3]dioxol-5-yl)-methanol (650 mg, 3.8 mmol) was added to SOCl₂ (20 mL) in portions at 0 °C. The mixture was warmed to room temperature for 1 h and then heated at reflux for 1 h. The excess SOCl₂ was evaporated under reduced pressure to give the crude product, which was basified with sat. NaHCO₃ solution to pH ~ 7. The aqueous phase was extracted with EtOAc (50 mL × 3). The combined organic layers were dried over Na₂SO₄ and evaporated under reduced pressure to give 5-chloromethyl-6-fluoro-benzo[1,3]dioxole (640 mg, 90%), which was directly used in the next step. <chemistry id="chem0165" num="0165"><img file="EP2007756B1_D0191.tif" /></chemistry>

(6-Fluoro-benzo[1,3]dioxol-5-yl)-acetonitrile

0244A mixture of 5-chloromethyl-6-fluoro-benzo[1,3]dioxole (640 mg, 3.4 mmol) and NaCN (340 mg, 6.8 mmol) in DMSO (20 mL) was stirred at 30 °C for 1 h and then poured into water. The mixture was extracted with EtOAc (50 mL × 3). The combined organic layers were washed with water (50 mL) and brine (50 mL), dried over Na₂SO₄, and evaporated under reduced pressure to give the crude product, which was purified by column chromatography on silica gel (5-10% ethyl acetate/petroleum ether) to afford (6-fluoro-benzo[1,3]dioxol-5-yl)-acetonitrile (530 mg, 70%). ¹H-NMR (300 MHz, CDCl₃) δ 6.82 (d, <i>J</i> = 4.8, 1 H), 6.62 (d, <i>J</i>= 5.4, 1 H), 5.99 (s, 2 H), 3.65 (s, 2 H). <chemistry id="chem0166" num="0166"><img file="EP2007756B1_D0192.tif" /></chemistry>

1-(6-Fluoro-benzo[1,3]dioxol-5-yl)-cyclopropanecarbonitrile

0245A flask was charged with water (10 mL), followed by a rapid addition of NaOH (10 g, 0.25 mol) in three portions over a 5 min period. The mixture was allowed to cool to room temperature. Subsequently, the flask was charged with toluene (6 mL), tetrabutylammonium bromide (50 mg, 0.12 mmol), (6-fluoro-benzo[1,3]dioxol-5-yl)-acetonitrile (600 mg, 3.4 mmol) and 1-bromo-2-chloroethane (1.7 g, 12 mmol). The mixture stirred vigorously at 50 °C overnight. The cooled flask was charged with additional toluene (20 mL). The organic layer was separated and washed with water (30 mL) and brine (30 mL). The organic layer was removed in <i>vacuo</i> to give the crude product, which was purified by column chromatography on silica gel (5-10% ethyl acetate/petroleum ether) to give 1-(6-fluoro-benzo[1,3]dioxol-5-yl)-cyclopropanecarbonitrile (400 mg, 60%). ¹H NMR (300 MHz, CDCl₃) δ 6.73 (d, <i>J</i> = 3.0 Hz, 1 H), 6.61 (d, <i>J</i> = 9.3 Hz, 1 H), 5.98 (s, 2 H), 1.67-1.62 (m, 2 H), 1.31-1.27 (m, 2 H). <chemistry id="chem0167" num="0167"><img file="EP2007756B1_D0193.tif" /></chemistry>

1-(6-Fluoro-benzo[1,3]-dioxol-5-yl)-cyclopropanecarboxylic acid

0246A mixture of 1-(6-fluoro-benzo[1,3]dioxol-5-yl)-cyclopropanecarbonitrile (400 mg, 0.196 mmol) and 10% NaOH (10 mL) was stirred at 100 °C overnight. After the reaction was cooled, 5% HCl was added until the pH &lt; 5 and then EtOAc (30 mL) was added to the reaction mixture. The layers were separated and combined organic layers were evaporated <i>in vacuo</i> to afford 1-(6-fluoro-benzo[1,3]dioxol-5-yl)-cyclopropanecarboxylic acid (330 mg, 76%). ¹H NMR (400 MHz, DMSO) δ 12.2 (s, 1 H), 6.87-6.85 (m, 2 H), 6.00 (s, 1 H), 1.42-1.40 (m, 2 H), 1.14-1.07 (m, 2 H).

Example 7: 1-(Benzofuran-5-yl)cyclopropanecarboxylic acid

0247<chemistry id="chem0168" num="0168"><img file="EP2007756B1_D0194.tif" /></chemistry>

1-[4-(2,2-Diethoxy-ethoxy)-phenyl]-cyclopropanecarboxylic acid

0248To a stirred solution of 1-(4-hydroxy-phenyl)-cyclopropanecarboxylic acid methyl ester (15.0 g, 84.3 mmol) in DMF (50 mL) was added sodium hydride (6.7 g, 170 mmol, 60% in mineral oil) at 0 °C. After hydrogen evolution ceased, 2-bromo-1,1-diethoxy-ethane (16.5 g, 84.3 mmol) was added dropwise to the reaction mixture. The reaction was stirred at 160 °C for 15 hours. The reaction mixture was poured onto ice (100 g) and was extracted with CH₂Cl₂. The combined organics were dried over Na₂SO₄. The solvent was evaporated under <i>vacuum</i> to give 1-[4-(2,2-diethoxy-ethoxy)-phenyl]-cyclopropanecarboxylic acid (10 g), which was used directly in the next step without purification. <chemistry id="chem0169" num="0169"><img file="EP2007756B1_D0195.tif" /></chemistry>

1-Benzofuran-5-yl-cyclopropanecarboxylic acid

0249To a suspension of 1-[4-(2,2-diethoxy-ethoxy)-phenyl]-cyclopropanecarboxylic acid (20 g, ~65 mmol) in xylene (100 mL) was added PPA (22.2 g, 64.9 mmol) at room temperature. The mixture was heated at reflux (140 °C) for 1 hour before it was cooled to room temperature and decanted from the PPA. The solvent was evaporated under <i>vacuum</i> to obtain the crude product, which was purified by preparative HPLC to provide 1-(benzofuran-5-yl)cyclopropanecarboxylic acid (1.5 g, 5%). ¹H NMR (400 MHz, DMSO-<i>d₆</i>) δ 12.25 (br s, 1 H), 7.95 (d, <i>J</i> = 2.8 Hz, 1 H), 7.56 (d, <i>J</i>= 2.0 Hz, 1 H), 7.47 (d, <i>J</i>= 11.6 Hz, 1 H), 7.25 (dd, <i>J</i>= 2.4, 11.2 Hz, 1 H), 6.89 (d, <i>J</i> = 1.6 Hz, 1 H), 1.47-1.44 (m, 2 H), 1.17-1.14 (m, 2 H).

Example 8: 1-(2,3-Dihydrobenzofuran-6-yl)cyclopropanecarboxylic acid

0250<chemistry id="chem0170" num="0170"><img file="EP2007756B1_D0196.tif" /></chemistry>

0251To a solution of 1-(benzofuran-6-yl)cyclopropanecarboxylic acid (370 mg, 1.8 mmol) in MeOH (50 mL) was added PtO₂ (75 mg, 20%) at room temperature. The reaction mixture was stirred under hydrogen atmosphere (1 atm) at 20 °C for 3 d. The reaction mixture was filtered and the solvent was evaporated <i>in vacuo</i> to afford the crude product, which was purified by prepared HPLC to give 1-(2,3-dihydrobenzofuran-6-yl)cyclopropanecarboxylic acid (155 mg, 42%). ¹H NMR (300 MHz, MeOD) δ 7.13 (d, <i>J</i>= 7.5 Hz, 1 H), 6.83 (d, <i>J</i>= 7.8 Hz, 1 H), 6.74 (s, 1 H), 4.55 (t, <i>J</i>= 8.7 Hz, 2 H), 3.18 (t, <i>J</i> = 8.7 Hz, 2 H), 1.56-1.53 (m, 2 H), 1.19-1.15 (m, 2 H).

Example 9: 1-(3,3-Dimethyl-2,3-dihydrobenzofuran-5-yl)cyclopropanecarboxylic acid.

0252<chemistry id="chem0171" num="0171"><img file="EP2007756B1_D0197.tif" /></chemistry>

1-(4-Hydroxy-phenyl)-cyclopropanecarboxylic acid methyl ester

0253To a solution of methyl 1-(4-methoxyphenyl)cyclopropanecarboxylate (10.0 g, 48.5 mmol) in dichloromethane (80 mL) was added EtSH (16 mL) under ice-water bath. The mixture was stirred at 0 °C for 20 min before AlCl₃ (19.5 g, 0.15 mmol) was added slowly at 0 °C. The mixture was stirred at 0 °C for 30 min. The reaction mixture was poured into ice-water, the organic layer was separated, and the aqueous phase was extracted with dichloromethane (50 mL × 3). The combined organic layers were washed with H₂O, brine, dried over Na₂SO₄ and evaporated under <i>vacuum</i> to give 1-(4-hydroxy-phenyl)-cyclopropanecarboxylic acid methyl ester (8.9 g, 95%). ¹H NMR (400 MHz, CDCl₃) δ 7.20-7.17 (m, 2 H), 6.75-6.72 (m, 2 H), 5.56 (s, 1 H), 3.63 (s, 3 H), 1.60-1.57 (m, 2 H), 1.17-1.15 (m, 2 H). <chemistry id="chem0172" num="0172"><img file="EP2007756B1_D0198.tif" /></chemistry>

1-(4-Hydroxy-3,5-diiodo-phenyl)-cyclopropanecarboxylic acid methyl ester

0254To a solution of 1-(4-hydroxy-phenyl)-cyclopropanecarboxylic acid methyl ester (8.9 g, 46 mmol) in CH₃CN (80 mL) was added NIS (15.6 g, 69 mmol). The mixture was stirred at room temperature for 1 hour. The reaction mixture was concentrated and the residue was purified by column chromatography on silica gel (petroleum ether/ethyl acetate 10:1) to give 1-(4-hydroxy-3,5-diiodo-phenyl)-cyclopropanecarboxylic acid methyl ester (3.5 g, 18%). ¹H NMR (400 MHz, CDCl₃) δ 7.65 (s, 2 H), 5.71 (s, 1 H), 3.63 (s, 3 H), 1.59-1.56 (m, 2 H), 1.15-1.12 (m, 2 H). <chemistry id="chem0173" num="0173"><img file="EP2007756B1_D0199.tif" /></chemistry>

1-[3,5-Diiodo-4-(2-methyl-allyloxy)-phenyl]-cyclopropanecarboxylic acid methyl ester

0255A mixture of 1-(4-hydroxy-3,5-diiodo-phenyl)-cyclopropanecarboxylic acid methyl ester (3.2 g, 7.2 mmol), 3-chloro-2-methyl-propene (1.0 g, 11 mmol), K₂CO₃ (1.2 g, 8.6 mmol), NaI (0.1 g, 0.7 mmol) in acetone (20 mL) was stirred at 20 °C overnight. The solid was filtered off and the filtrate was concentrated under <i>vacuum</i> to give 1-[3,5-diiodo-4-(2-methyl-allyloxy)-phenyl]-cyclopropane-carboxylic acid methyl ester (3.5 g, 97%). ¹H NMR (300 MHz, CDCl₃) δ 7.75 (s, 2 H), 5.26 (s, 1 H), 5.06 (s, 1 H), 4.38 (s, 2 H), 3.65 (s, 3 H), 1.98 (s, 3H), 1.62-1.58 (m, 2 H), 1.18-1.15 (m, 2 H). <chemistry id="chem0174" num="0174"><img file="EP2007756B1_D0200.tif" /></chemistry>

1-(3,3-Dimethyl-2,3-dihydro-benzofuran-5-yl)-cyclopropanecarboxylic acid methyl ester

0256To a solution of 1-[3,5-diiodo-4-(2-methyl-allyloxy)-phenyl]-cyclopropanecarboxylic acid methyl ester (3.5 g, 7.0 mmol) in toluene (15 mL) was added Bu₃SnH (2.4 g, 8.4 mmol) and AIBN (0.1 g, 0.7 mmol). The mixture was heated at reflux overnight. The reaction mixture was concentrated under <i>vacuum</i> and the residue was purified by column chromatography on silica gel (petroleum ether/ethyl acetate 20:1) to give 1-(3,3-dimethyl-2,3-dihydro-benzofuran-5-yl)-cyclopropanecarboxylic acid methyl ester (1.05 g, 62%). ¹H NMR (400 MHz, CDCl₃) δ 7.10-7.07 (m, 2 H), 6.71 (d, <i>J</i> = 8 Hz, 1 H), 4.23 (s, 2 H), 3.62 (s, 3 H), 1.58-1.54 (m, 2 H), 1.34 (s, 6 H), 1.17-1.12 (m, 2 H). <chemistry id="chem0175" num="0175"><img file="EP2007756B1_D0201.tif" /></chemistry>

1-(3,3-Dimethyl-2,3-dihydrobenzofuran-5-yl)cyclopropanecarboxylic acid

0257To a solution of 1-(3,3-dimethyl-2,3-dihydro-benzofuran-5-yl)-cyclopropanecarboxylic acid methyl ester (1.0 g, 4.0 mmol) in MeOH (10 mL) was added LiOH (0.40 g, 9.5 mmol). The mixture was stirred at 40 °C overnight. HCl (10%) was added slowly to adjust the pH to 5. The resulting mixture was extracted with ethyl acetate (10 mL × 3). The extracts were washed with brine and dried over Na₂SO₄. The solvent was removed under <i>vaccum</i> and the crude product was purified by preparative HPLC to give 1-(3,3-dimethyl-2,3-dihydrobenzofuran-5-yl)cyclopropanecarboxylic acid (0.37 g, 41%). ¹H NMR (400 MHz, CDCl₃) δ 7.11-7.07 (m, 2 H), 6.71 (d, <i>J</i> = 8 Hz, 1 H), 4.23 (s, 2 H), 1.66-1.63 (m, 2 H), 1.32 (s, 6 H), 1.26-1.23 (m, 2 H).

Example 10: 2-(7-Methoxybenzo[d][1,3]dioxol-5-yl)acetonitrile.

0258<chemistry id="chem0176" num="0176"><img file="EP2007756B1_D0202.tif" /></chemistry>

3,4-Dihydroxy-5-methoxybenzoate

0259To a solution of 3,4,5-trihydroxy-benzoic acid methyl ester (50 g, 0.27 mol) and Na₂B₄O₇ (50 g) in water (1000 mL) was added Me₂SO₄ (120 mL) and aqueous NaOH solution (25%, 200 mL) successively at room temperature. The mixture was stirred at room temperature for 6 h before it was cooled to 0 °C. The mixture was acidified to pH ~ 2 by adding conc. H₂SO₄ and then filtered. The filtrate was extracted with EtOAc (500 mL × 3). The combined organic layers were dried over anhydrous Na₂SO₄ and evaporated under reduced pressure to give methyl 3,4-dihydroxy-5-methoxybenzoate (15.3 g 47%), which was used in the next step without further purification. <chemistry id="chem0177" num="0177"><img file="EP2007756B1_D0203.tif" /></chemistry>

Methyl 7-methoxybenzo[d][1,3]dioxole-5-carboxylate

0260To a solution of methyl 3,4-dihydroxy-5-methoxybenzoate (15.3 g, 0.0780 mol) in acetone (500 mL) was added CH₂BrCl (34.4 g, 0.270 mol) and K₂CO₃ (75.0 g, 0.540 mol) at 80 °C. The resulting mixture was heated at reflux for 4 h. The mixture was cooled to room temperature and solid K₂CO₃ was filtered off. The filtrate was concentrated under reduced pressure, and the residue was dissolved in EtOAc (100 mL). The organic layer was washed with water, dried over anhydrous Na₂SO₄, and evaporated under reduced pressure to give the crude product, which was purified by column chromatography on silica gel (petroleum ether/ethyl acetate = 10:1) to afford methyl 7-methoxybenzo[d][1,3]dioxole-5-carboxylate (12.6 g, 80%). ¹H NMR (400 MHz, CDCl₃) δ 7.32 (s, 1 H), 7.21 (s, 1 H), 6.05 (s, 2 H), 3.93 (s, 3 H), 3.88 (s, 3 H). <chemistry id="chem0178" num="0178"><img file="EP2007756B1_D0204.tif" /></chemistry>

(7-Methoxybenzo[d][1,3]dioxol-5-yl)methanol

0261To a solution of methyl 7-methoxybenzo[d][1,3]dioxole-5-carboxylate (14 g, 0.040 mol) in THF (100 mL) was added LiAlH₄ (3.1 g, 0.080 mol) in portions at room temperature. The mixture was stirred for 3 h at room temperature. The reaction mixture was cooled to 0 °C and treated with water (3.1 g) and NaOH (10%, 3.1 mL) successively. The slurry was filtered off and washed with THF. The combined filtrates were evaporated under reduced pressure to give (7-methoxy-benzo[d][1,3]dioxol-5-yl)methanol (7.2 g, 52%). ¹H NMR (400 MHz, CDCl₃) δ 6.55 (s, 1H), 6.54 (s, 1H), 5.96 (s, 2 H), 4.57 (s, 2 H), 3.90 (s, 3 H). <chemistry id="chem0179" num="0179"><img file="EP2007756B1_D0205.tif" /></chemistry>

6-(Chloromethyl)-4-methoxybenzo[d][1,3]dioxole

0262To a solution of SOCl₂ (150 mL) was added (7-methoxybenzo[d][1,3]dioxol-5-yl)methanol (9.0 g, 54 mmol) in portions at 0 °C. The mixture was stirred for 0.5 h. The excess SOCl₂ was evaporated under reduced pressure to give the crude product, which was basified with sat. aq. NaHCO₃ to pH ~ 7. The aqueous phase was extracted with EtOAc (100 mL × 3). The combined organic layers were dried over anhydrous Na₂SO₄ and evaporated to give 6-(chloromethyl)-4-methoxybenzo[d][1,3]dioxole (10 g 94%), which was used in the next step without further purification. ¹H NMR (400 MHz, CDCl₃) δ 6.58 (s, 1 H), 6.57 (s, 1 H), 5.98 (s, 2 H), 4.51 (s, 2 H), 3.90 (s, 3 H). <chemistry id="chem0180" num="0180"><img file="EP2007756B1_D0206.tif" /></chemistry>

2-(7-Methoxybenzo[d][1,3]dioxol-5-yl)acetonitrile

0263To a solution of 6-(chloromethyl)-4-methoxybenzo[d][1,3]dioxole (10 g, 40 mmol) in DMSO (100 mL) was added NaCN (2.4 g, 50 mmol) at room temperature. The mixture was stirred for 3 h and poured into water (500 mL). The aqueous phase was extracted with EtOAc (100 mL × 3). The combined organic layers were dried over anhydrous Na₂SO₄ and evaporated to give the crude product, which was washed with ether to afford 2-(7-methoxybenzo[d][1,3]dioxol-5-yl)acetonitrile (4.6 g, 45%). ¹H NMR (400 MHz, CDCl₃) δ 6.49 (s, 2 H), 5.98 (s, 2 H), 3.91 (s, 3 H), 3.65 (s, 2 H). ¹³C NMR (400 MHz, CDCl₃) δ 148.9, 143.4, 134.6, 123.4, 117.3, 107.2, 101.8, 101.3, 56.3, 23.1.

Example 11: 2-(3-(Benzyloxy)-4-methoxyphenyl)acetonitrile.

0264<chemistry id="chem0181" num="0181"><img file="EP2007756B1_D0207.tif" /></chemistry>

0265To a suspension of t-BuOK (20.2 g, 0.165 mol) in THF (250 mL) was added a solution of TosMIC (16.1 g, 82.6 mmol) in THF (100 mL) at -78 °C. The mixture was stirred for 15 minutes, treated with a solution of 3-benzyloxy-4-methoxy-benzaldehyde (10.0 g, 51.9 mmol) in THF (50 mL) dropwise, and continued to stir for 1.5 hours at -78 °C. To the cooled reaction mixture was added methanol (50 mL). The mixture was heated at reflux for 30 minutes. Solvent was removed to give a crude product, which was dissolved in water (300 mL). The aqueous phase was extracted with EtOAc (100 mL × 3). The combined organic layers were dried and evaporated under reduced pressure to give crude product, which was purified by column chromatography (petroleum ether/ethyl acetate 10:1) to afford 2-(3-(benzyloxy)-4-methoxyphenyl)- acetonitrile (5.0 g, 48%). ¹H NMR (300 MHz, CDCl₃) δ 7.48-7.33 (m, 5 H), 6.89-6.86 (m, 3 H), 5.17 (s, 2 H), 3.90 (s, 3 H), 3.66 (s, 2 H). ¹³C NMR (75 MHz, CDCl₃) δ 149.6, 148.6, 136.8, 128.8, 128.8, 128.2, 127.5, 127.5, 122.1, 120.9, 118.2, 113.8, 112.2, 71.2, 56.2, 23.3.

Example 12: 2-(3-(Benzyloxy)-4-chlorophenyl)acetonitrile.

0266<chemistry id="chem0182" num="0182"><img file="EP2007756B1_D0208.tif" /></chemistry>

(4-Chloro-3-hydroxy-phenyl)acetonitrile

0267BBr₃ (17 g, 66 mmol) was slowly added to a solution of 2-(4-chloro-3-methoxyphenyl)acetonitrile (12 g, 66 mmol) in dichloromethane (120 mL) at -78 °C under N₂. The reaction temperature was slowly increased to room temperature. The reaction mixture was stirred overnight and then poured into ice and water. The organic layer was separated, and the aqueous layer was extracted with dichloromethane (40 mL × 3). The combined organic layers were washed with water, brine, dried over Na₂SO₄, and concentrated under vacuum to give (4-chloro-3-hydroxy-phenyl)-acetonitrile (9.3 g, 85%). ¹H NMR (300 MHz, CDCl₃) δ 7.34 (d, J = 8.4 Hz, 1 H), 7.02 (d, J = 2.1 Hz, 1 H), 6.87 (dd, J = 2.1, 8.4 Hz, 1 H), 5.15 (brs, 1H), 3.72 (s, 2 H). <chemistry id="chem0183" num="0183"><img file="EP2007756B1_D0209.tif" /></chemistry>

2-(3-(Benzyloxy)-4-chlorophenyl)acetonitrile

0268To a solution of (4-chloro-3-hydroxy-phenyl)acetonitrile (6.2 g, 37 mmol) in CH₃CN (80 mL) was added K₂CO₃ (10 g, 74 mmol) and BnBr (7.6 g, 44 mmol). The mixture was stirred at room temperature overnight. The solids were filtered off and the filtrate was evaporated under vacuum. The residue was purified by column chromatography on silica gel (petroleum ether/ethyl acetate 50:1) to give 2-(3-(benzyloxy)-4-chlorophenyl)-acetonitrile (5.6 g, 60%). ¹H NMR (400 MHz, CDCl₃) δ 7.48-7.32 (m, 6 H), 6.94 (d, J = 2 Hz, 2 H), 6.86 (dd, J = 2.0, 8.4 Hz, 1 H), 5.18 (s, 2 H), 3.71 (s, 2 H).

Example 13: 2-(3-(Benzyloxy)-4-methoxyphenyl)acetonitrile.

0269<chemistry id="chem0184" num="0184"><img file="EP2007756B1_D0210.tif" /></chemistry>

0270To a suspension of t-BuOK (20.2 g, 0.165 mol) in THF (250 mL) was added a solution of TosMIC (16.1 g, 82.6 mmol) in THF (100 mL) at -78 °C. The mixture was stirred for 15 minutes, treated with a solution of 3-benzyloxy-4-methoxy-benzaldehyde (10.0 g, 51.9 mmol) in THF (50 mL) dropwise, and continued to stir for 1.5 hours at -78 °C. To the cooled reaction mixture was added methanol (50 mL). The mixture was heated at reflux for 30 minutes. Solvent of the reaction mixture was removed to give a crude product, which was dissolved in water (300 mL). The aqueous phase was extracted with EtOAc (100 mL × 3). The combined organic layers were dried and evaporated under reduced pressure to give crude product, which was purified by column chromatography (petroleum ether/ethyl acetate 10:1) to afford 2-(3-(benzyloxy)-4-methoxyphenyl)acetonitril (5.0 g, 48%). ¹H NMR (300 MHz, CDCl₃) δ 7.48-7.33 (m, 5 H), 6.89-6.86 (m, 3 H), 5.17 (s, 2 H), 3.90 (s, 3 H), 3.66 (s, 2 H). ¹³C NMR (75 MHz, CDCl₃) δ 149.6, 148.6, 136.8, 128.8, 128.8, 128.2, 127.5, 127.5, 122.1, 120.9, 118.2, 113.8, 112.2, 71.2, 56.2, 23.3.

Example 14: 2-(3-Chloro-4-methoxyphenyl)acetonitrile.

0271<chemistry id="chem0185" num="0185"><img file="EP2007756B1_D0211.tif" /></chemistry>

0272To a suspension of t-BuOK (4.8 g, 40 mmol) in THF (30 mL) was added a solution of TosMIC (3.9 g, 20 mmol) in THF (10 mL) at -78 °C. The mixture was stirred for 10 minutes, treated with a solution of 3-chloro-4-methoxy-benzaldehyde (1.7 g, 10 mmol) in THF (10 mL) dropwise, and continued to stir for 1.5 hours at -78 °C. To the cooled reaction mixture was added methanol (10 mL). The mixture was heated at reflux for 30 minutes. Solvent of the reaction mixture was removed to give a crude product, which was dissolved in water (20 mL). The aqueous phase was extracted with EtOAc (20 mL × 3). The combined organic layers were dried and evaporated under reduced pressure to give crude product, which was purified by column chromatography (petroleum ether/ethyl acetate 10:1) to afford 2-(3-chloro-4-methoxyphenyl)acetonitrile (1.5 g, 83%). ¹H NMR (400 MHz, CDCl₃) δ 7.33 (d, <i>J</i>= 2.4 Hz, 1 H), 7.20 (dd, <i>J</i>= 2.4, 8.4 Hz, 1 H), 6.92 (d, <i>J</i>= 8.4 Hz, 1 H), 3.91 (s, 3 H), 3.68 (s, 2 H). ¹³C NMR (100 MHz, CDCl₃) δ 154.8, 129.8, 127.3, 123.0, 122.7; 117.60, 112.4,56.2,22.4.

Example 15: 2-(3-Fluoro-4-methoxyphenyl)acetonitrile.

0273<chemistry id="chem0186" num="0186"><img file="EP2007756B1_D0212.tif" /></chemistry>

0274To a suspension of t-BuOK (25.3 g, 0.207 mol) in THF (150 mL) was added a solution of TosMIC (20.3 g, 0.104 mol) in THF (50 mL) at -78 °C. The mixture was stirred for 15 minutes, treated with a solution of 3-fluoro-4-methoxy-benzaldehyde (8.00 g, 51.9 mmol) in THF (50 mL) dropwise, and continued to stir for 1.5 hours at -78 °C. To the cooled reaction mixture was added methanol (50 mL). The mixture was heated at reflux for 30 minutes. Solvent of the reaction mixture was removed to give a crude product, which was dissolved in water (200 mL). The aqueous phase was extracted with EtOAc (100 mL × 3). The combined organic layers were dried and evaporated under reduced pressure to give crude product, which was purified by column chromatography (petroleum ether/ethyl acetate 10:1) to afford 2-(3-fluoro-4-methoxyphenyl)acetonitrile (5.0 g, 58%). ¹H NMR (400 MHz, CDCl₃) δ 7.02-7.05 (m, 2 H), 6.94 (t, J= 8.4 Hz, 1 H), 3.88 (s, 3 H), 3.67 (s, 2 H). ¹³C NMR (100 MHz, CDCl₃) δ 152.3, 147.5, 123.7, 122.5, 117.7, 115.8, 113.8, 56.3, 22.6.

Example 16: 2-(4-Chloro-3-methoxyphenyl)acetonitrile.

0275<chemistry id="chem0187" num="0187"><img file="EP2007756B1_D0213.tif" /></chemistry>

Chloro-2-methoxy-4-methyl-benzene

0276To a solution of 2-chloro-5-methyl-phenol (93 g, 0.65 mol) in CH₃CN (700 mL) was added CH₃I (110 g, 0.78 mol) and K₂CO₃ (180 g, 1.3 mol). The mixture was stirred at 25 °C overnight. The solid was filtered off and the filtrate was evaporated under <i>vacuum</i> to give 1-chloro-2-methoxy-4-methyl-benzene (90 g, 89%). ¹H NMR (300 MHz, CDCl₃) δ 7.22 (d, <i>J</i> = 7.8 Hz, 1 H), 6.74-6.69 (m, 2 H), 3.88 (s, 3 H), 2.33 (s, 3 H). <chemistry id="chem0188" num="0188"><img file="EP2007756B1_D0214.tif" /></chemistry>

4-Bromomethyl-1-chloro-2-methoxy-benzene

0277To a solution of 1-chloro-2-methoxy-4-methyl-benzene (50 g, 0.32 mol) in CCl₄ (350 mL) was added NBS (57 g, 0.32 mol) and AIBN (10 g, 60 mmol). The mixture was heated at reflux for 3 hours. The solvent was evaporated under <i>vacuum</i> and the residue was purified by column chromatography on silica gel (petroleum ether/ethyl acetate = 20:1) to give 4-bromomethyl-1-chloro-2-methoxy-benzene (69 g, 92%). ¹H NMR (400 MHz, CDCl₃) δ 7.33-7.31 (m, 1 H), 6.95-6.91 (m, 2 H), 4.46 (s, 2 H), 3.92 (s, 3 H). <chemistry id="chem0189" num="0189"><img file="EP2007756B1_D0215.tif" /></chemistry>

2-(4-Chloro-3-methoxyphenyl)acetonitrile

0278To a solution of 4-bromomethyl-1-chloro-2-methoxy-benzene (68.5 g, 0.290 mol) in C₂H₅OH (90%, 500 mL) was added NaCN (28.5 g, 0.580 mol). The mixture was stirred at 60 °C overnight. Ethanol was evaporated and the residue was dissolved in H₂O. The mixture was extracted with ethyl acetate (300 mL × 3). The combined organic layers were washed with brine, dried over Na₂SO₄ and purified by column chromatography on silica gel (petroleum ether/ethyl acetate 30:1) to give 2-(4-chloro-3-methoxyphenyl)acetonitrile (25 g, 48%). ¹H NMR (400 MHz, CDCl₃) δ 7.36 (d, <i>J</i> = 8 Hz, 1 H), 6.88-6.84 (m, 2 H), 3.92 (s, 3 H), 3.74 (s, 2 H). ¹³C NMR (100 MHz, CDCl₃) δ 155.4, 130.8, 129.7, 122.4, 120.7, 117.5, 111.5, 56.2, 23.5.

Example 17: 1-(3-(Hydroxymethyl)-4-methoxyphenyl)cyclopropanecarboxylic acid.

0279<chemistry id="chem0190" num="0190"><img file="EP2007756B1_D0216.tif" /></chemistry><chemistry id="chem0191" num="0191"><img file="EP2007756B1_D0217.tif" /></chemistry>

1-(4-Methoxy-phenyl)-cyclopropanecarboxylic acid methyl ester

0280To a solution of 1-(4-methoxy-phenyl)-cyclopropanecarboxylic acid (50 g, 0.26 mol) in MeOH (500 mL) was added toluene-4-sulfonic acid monohydrate (2.5 g, 13 mmol) at room temperature. The reaction mixture was heated at reflux for 20 hours. MeOH was removed by evaporation under <i>vacuum</i> and EtOAc (200 mL) was added. The organic layer was washed with sat. aq. NaHCO₃ (100 mL) and brine, dried over anhydrous Na₂SO₄ and evaporated under <i>vacuum</i> to give 1-(4-methoxy-phenyl)-cyclopropanecarboxylic acid methyl ester (53 g, 99%). ¹H NMR (CDCl₃, 400 MHz) δ 7.25-7.27 (m, 2 H), 6.85 (d, <i>J</i>= 8.8 Hz, 2 H), 3.80 (s, 3 H), 3.62 (s, 3 H), 1.58 (m, 2 H), 1.15 (m, 2 H). <chemistry id="chem0192" num="0192"><img file="EP2007756B1_D0218.tif" /></chemistry>

1-(3-Chloromethyl-4-methoxy-phenyl)-cyclopropanecarboxylic acid methyl ester

0281To a solution of 1-(4-methoxy-phenyl)-cyclopropanecarboxylic acid methyl ester (30.0 g, 146 mmol) and MOMCl (29.1 g, 364 mmol) in CS₂ (300 mL) was added TiCl₄ (8.30 g, 43.5 mmol) at 5 °C. The reaction mixture was heated at 30 °C for 1 d and poured into ice-water. The mixture was extracted with CH₂Cl₂ (150 mL × 3). The combined organic extracts were evaporated under <i>vacuum</i> to give 1-(3-chloromethyl-4-methoxy-phenyl)-cyclopropanecarboxylic acid methyl ester (38.0 g), which was used in the next step without further purification. <chemistry id="chem0193" num="0193"><img file="EP2007756B1_D0219.tif" /></chemistry>

1-(3-Hydroxymethyl-4-methoxy-phenyl)-cyclopropanecarboxylic acid methyl

ester

0282To a suspension of 1-(3-chloromethyl-4-methoxy-phenyl)-cyclopropanecarboxylic acid methyl ester (20 g) in water (350 mL) was added Bu₄NBr (4.0 g) and Na₂CO₃ (90 g, 0.85 mol) at room temperature. The reaction mixture was heated at 65 °C overnight. The resulting solution was acidified with aq. HCl (2 mol/L) and extracted with EtOAc (200 mL × 3). The organic layer was washed with brine, dried over anhydrous Na₂SO₄ and evaporated under <i>vacuum</i> to give crude product, which was purified by column (petroleum ether/ethyl acetate 15:1) to give 1-(3-hydroxymethyl-4-methoxy-phenyl)-cyclopropanecarboxylic acid methyl ester (8.0 g, 39%). ¹H NMR (CDCl₃, 400 MHz) δ 7.23-7.26 (m, 2 H), 6.83 (d, <i>J</i>= 8.0 Hz, 1 H), 4.67 (s, 2 H), 3.86 (s, 3 H), 3.62 (s, 3 H), 1.58 (q, <i>J</i> = 3.6 Hz, 2 H), 1.14-1.17 (m, 2 H). <chemistry id="chem0194" num="0194"><img file="EP2007756B1_D0220.tif" /></chemistry>

1-[3-(<i>tert</i>-Butyl-dimethyl-silanyloxymethyl)-4-methoxy-phenyl]cyclopropane carboxylic acid methyl ester

0283To a solution of 1-(3-hydroxymethyl-4-methoxy-phenyl)-cyclopropanecarboxylic acid methyl ester (8.0 g, 34 mmol) in CH₂Cl₂ (100 mL) were added imidazole (5.8 g, 85 mmol) and TBSCI (7.6 g, 51 mmol) at room temperature. The mixture was stirred overnight at room temperature. The mixture was washed with brine, dried over anhydrous Na₂SO₄ and evaporated under <i>vacuum</i> to give crude product, which was purified by column (petroleum ether/ethyl acetate 30:1) to give 1-[3-(<i>tert</i>-butyl-dimethyl-silanyloxymethyl)-4-methoxy-phenyl]-cyclopropanecarboxylic acid methyl ester (6.7 g, 56%). ¹H NMR (CDCl₃, 400 MHz) δ 7.44-7.45 (m, 1 H), 7.19 (dd, <i>J</i> = 2.0, 8.4 Hz, 1 H), 6.76 (d, <i>J</i> = 8.4 Hz, 1 H), 4.75 (s, 2 H), 3.81 (s, 3 H), 3.62 (s, 3 H), 1.57-1.60 (m, 2 H), 1.15- 1.18 (m, 2 H), 0.96 (s, 9 H), 0.11 (s, 6 H). <chemistry id="chem0195" num="0195"><img file="EP2007756B1_D0221.tif" /></chemistry>

1-(3-Hydroxymethyl-4-methoxy-phenyl)-cyclopropanecarboxylic acid

0284To a solution of 1-[3-(<i>tert</i>-butyl-dimethyl-silanyloxymethyl)-4-methoxy-phenyl]-cyclopropane carboxylic acid methyl ester (6.2 g, 18 mmol) in MeOH (75 mL) was added a solution of LiOH.H₂O (1.5 g, 36 mmol) in water (10 mL) at 0 °C. The reaction mixture was stirred overnight at 40 °C. MeOH was removed by evaporation under <i>vacuum.</i> AcOH (1 mol/L, 40 mL) and EtOAc (200 mL) were added. The organic layer was separated, washed with brine, dried over anhydrous Na₂SO₄ and evaporated under <i>vacuum</i> to provide 1-(3-hydroxymethyl-4-methoxy-phenyl)-cyclopropanecarboxylic acid (5.3 g).

Example 18: 2-(7-Chlorobenzo[d][1,3]dioxol-5-yl)acetonitrile.

0285<chemistry id="chem0196" num="0196"><img file="EP2007756B1_D0222.tif" /></chemistry><chemistry id="chem0197" num="0197"><img file="EP2007756B1_D0223.tif" /></chemistry>

3-Chloro-4,5-dihydroxybenzaldehyde

0286To a suspension of 3-chloro-4-hydroxy-5-methoxy-benzaldehyde (10 g, 54 mmol) in dichloromethane (300 mL) was added BBr₃ (26.7 g, 107 mmol) dropwise at -40 °C under N₂. After addition, the mixture was stirred at this temperature for 5 h and then was poured into ice water. The precipitated solid was filtered and washed with petroleum ether. The filtrate was evaporated under reduced pressure to afford 3-chloro-4,5-dihydroxybenzaldehyde (9.8 g, 89%), which was directly used in the next step. <chemistry id="chem0198" num="0198"><img file="EP2007756B1_D0224.tif" /></chemistry>

7-Chlorobenzo[d][1,3]dioxole-5-carbaldehyde

0287To a solution of 3-chloro-4,5-dihydroxybenzaldehyde (8.0 g, 46 mmol) and BrClCH₂ (23.9 g, 185 mmol) in dry DMF (100 mL) was added Cs₂CO₃ (25 g, 190 mmol). The mixture was stirred at 60 °C overnight and was then poured into water. The resulting mixture was extracted with EtOAc (50 mL × 3). The combined extracts were washed with brine (100 mL), dried over Na₂SO₄ and concentrated under reduced pressure to afford 7-chlorobenzo[d][1,3]dioxole-5-carbaldehyde (6.0 g, 70%). ¹H NMR (400 MHz, CDCl₃) δ 9.74 (s, 1 H), 7.42 (d, <i>J</i> = 0.4 Hz, 1 H), 7.26 (d, <i>J</i> = 3.6 Hz, 1 H), 6.15 (s, 2 H). <chemistry id="chem0199" num="0199"><img file="EP2007756B1_D0225.tif" /></chemistry>

(7-Chlorobenzo[d][1,3]dioxol-5-yl)methanol

0288To a solution of 7-chlorobenzo[d][1,3]dioxole-5-carbaldehyde (6.0 g, 33 mmol) in THF (50 mL) was added NaBH₄ (2.5 g, 64 mmol)) in portions at 0 °C. The mixture was stirred at this temperature for 30 min and then poured into aqueous NH₄Cl solution. The organic layer was separated, and the aqueous phase was extracted with EtOAc (50 mL × 3). The combined extracts were dried over Na₂SO₄ and evaporated under reduced pressure to afford (7-chlorobenzo[d][1,3]dioxol-5-yl)methanol, which was directly used in the next step. <chemistry id="chem0200" num="0200"><img file="EP2007756B1_D0226.tif" /></chemistry>

4-Chloro-6-(chloromethyl)benzo[d][1,3]dioxole

0289A mixture of (7-chlorobenzo[d][1,3]-dioxol-5-yl)methanol (5.5 g, 30 mmol) and SOCl₂ (5.0 mL), 67 mmol) in dichloromethane (20 mL) was stirred at room temperature for 1 h and was then poured into ice water. The organic layer was separated and the aqueous phase was extracted with dichloromethane (50 mL × 3). The combined extracts were washed with water and aqueous NaHCO₃ solution, dried over Na₂SO₄ and evaporated under reduced pressure to afford 4-chloro-6-(chloromethyl)benzo[d][1,3]dioxole, which was directly used in the next step. <chemistry id="chem0201" num="0201"><img file="EP2007756B1_D0227.tif" /></chemistry>

2-(7-Chlorobenzo[d][1,3]dioxol-5-yl)acetonitrile

0290A mixture of 4-chloro-6-(chloromethyl)benzo[d][1,3]dioxole (6.0 g, 29 mmol) and NaCN (1.6 g, 32 mmol) in DMSO (20 mL) was stirred at 40 °C for 1 h and was then poured into water. The mixture was extracted with EtOAc (30 mL × 3). The combined organic layers were washed with water and brine, dried over Na₂SO₄ and evaporated under reduced pressure to afford 2-(7-chlorobenzo[d][1,3]dioxol-5-yl)acetonitrile (3.4 g, 58%). ¹H NMR δ 6.81 (s, 1 H), 6.71 (s, 1 H), 6.07 (s, 2 H), 3.64 (s, 2 H). ¹³C-NMR δ149.2, 144.3, 124.4, 122.0, 117.4, 114.3, 107.0, 102.3, 23.1.

Example 19: 1-(Benzo[d]oxazol-5-yl)cyclopropanecarboxylic acid.

0291<chemistry id="chem0202" num="0202"><img file="EP2007756B1_D0228.tif" /></chemistry><chemistry id="chem0203" num="0203"><img file="EP2007756B1_D0229.tif" /></chemistry>

1-Benzooxazol-5-yl-cyclopropanecarboxylic acid methyl ester

0292To a solution of 1-(3-amino-4-hydroxyphenyl)cyclopropanecarboxylic acid methyl ester (3.00 g, 14.5 mmol) in DMF were added trimethyl orthoformate (5.30 g, 14.5 mmol) and a catalytic amount of <i>p</i>-tolueneslufonic acid monohydrate (0.3 g) at room temperature. The mixture was stirred for 3 hours at room temperature. The mixture was diluted with water and extracted with EtOAc (100 mL × 3). The combined organic layers were dried over anhydrous Na₂SO₄ and evaporated under <i>vacuum</i> to give 1-benzooxazol-5-yl-cyclopropanecarboxylic acid methyl ester (3.1 g), which was directly used in the next step. ¹H NMR (CDCl₃ 400 MHz) δ 8.09 (s, 1), 7.75 (d, <i>J</i>= 1.2 Hz, 1 H), 7.53-7.51 (m, 1 H), 7.42-7.40 (m, 1 H), 3.66 (s, 3 H), 1.69-1.67 (m, 2 H), 1.27-1.24 (m, 2 H). <chemistry id="chem0204" num="0204"><img file="EP2007756B1_D0230.tif" /></chemistry>

1-(Benzo[d]oxazol-5-yl)cyclopropanecarboxylic acid

0293To a solution of 1-benzooxazol-5-yl-cyclopropanecarboxylic acid methyl ester (2.9 g) in EtSH (30 mL) was added AlCl₃ (5.3 g, 40 mmol) in portions at 0 °C. The reaction mixture was stirred for 18 hours at room temperature. Water (20 mL) was added dropwise at 0 °C. The resulting mixture was extracted with EtOAc (100 mL × 3). The combined organic layers were dried over anhydrous Na₂SO₄ and evaporated under <i>vacuum</i> to give the crude product, which was purified by column chromatography on silica gel (petroleum ether/ethyl acetate 1:2) to give 1-(benzo[d]oxazol-5-yl)cyclopropanecarboxylic acid (280 mg, 11% over two steps). ¹H NMR (DMSO, 400 MHz) δ 12.25 (brs, 1 H), 8.71 (s, 1 H), 7.70-7.64 (m, 2 H), 7.40 (dd, <i>J</i>= 1.6, 8.4 Hz, 1 H), 1.49-1.46 (m, 2 H), 1.21-1.18 (m, 2 H). MS (ESI) m/e (M+H⁺) 204.4.

Example 20: 2-(7-Fluorobenzo[d][1,3]dioxol-5-yl)acetonitrile

0294<chemistry id="chem0205" num="0205"><img file="EP2007756B1_D0231.tif" /></chemistry><chemistry id="chem0206" num="0206"><img file="EP2007756B1_D0232.tif" /></chemistry>

3-Fluoro-4,5-dihydroxy-benzaldehyde

0295To a suspension of 3-fluoro-4-hydroxy-5-methoxy-benzaldehyde (1.35 g, 7.94 mmol) in dichloromethane (100 mL) was added BBr₃ (1.5 mL, 16 mmol) dropwise at -78 °C under N₂. After addition, the mixture was warmed to - 30 °C and it was stirred at this temperature for 5 h. The reaction mixture was poured into ice water. The precipitated solid was collected by filtration and washed with dichloromethane to afford 3-fluoro-4,5-dihydroxy-benzaldehyde (1.1 g, 89%), which was directly used in the next step. <chemistry id="chem0207" num="0207"><img file="EP2007756B1_D0233.tif" /></chemistry>

7-

Fluoro-benzo[1,3]dioxole-5-carbaldehyde

0296To a solution of 3-fluoro-4,5-dihydroxy-benzaldehyde (1.5 g, 9.6 mmol) and BrClCH₂ (4.9 g, 38.5 mmol) in dry DMF (50 mL) was added Cs₂CO₃ (12.6 g, 39 mmol). The mixture was stirred at 60 °C overnight and was then poured into water. The resulting mixture was extracted with EtOAc (50 mL × 3). The combined organic layers were washed with brine (100 mL), dried over Na₂SO₄ and evaporated under reduced pressure to give the crude product, which was purified by column chromatography on silica gel (petroleum ether/ethyl acetate = 10/1) to afford 7-fluoro-benzo[1,3]dioxole-5-carbaldehyde (0.80 g, 49%). ¹H NMR (300 MHz, CDCl₃) δ 9.78 (d, <i>J</i> = 0.9 Hz, 1 H), 7.26 (dd, <i>J</i>= 1.5, 9.3 Hz, 1H), 7.19 (d, <i>J</i>= 1.2 Hz, 1 H), 6.16 (s, 2 H). <chemistry id="chem0208" num="0208"><img file="EP2007756B1_D0234.tif" /></chemistry>

(7-Fluoro-benzo[1,3]dioxol-5-yl)-methanol

0297To a solution of 7-fluoro-benzo[1,3]dioxole-5-carbaldehyde (0.80 g, 4.7 mmol) in MeOH (50 mL) was added NaBH₄ (0.36 g, 9.4 mmol) in portions at 0 °C. The mixture was stirred at this temperature for 30 min and was then concentrated to dryness. The residue was dissolved in EtOAc. The EtOAc layer was washed with water, dried over Na₂SO₄ and concentrated to dryness to afford (7-fluoro-benzo[1,3]dioxol-5-yl)-methanol (0.80 g, 98%), which was directly used in the next step. <chemistry id="chem0209" num="0209"><img file="EP2007756B1_D0235.tif" /></chemistry>

6-Chloromethyl-4-fluoro-benzo[1,3]dioxole

0298To SOCl₂ (20 mL) was added (7-fluoro-benzo[1,3]dioxol-5-yl)-methanol (0.80 g, 4.7 mmol) in portions at 0 °C. The mixture was warmed to room temperature over 1 h and then was heated at reflux for 1 h. The excess SOCl₂ was evaporated under reduced pressure to give the crude product, which was basified with saturated aqueous NaHCO₃ to pH ~ 7. The aqueous phase was extracted with EtOAc (50 mL × 3). The combined organic layers were dried over Na₂SO₄ and evaporated under reduced pressure to give 6-chloromethyl-4-fluoro-benzo[1,3]dioxole (0.80 g, 92%), which was directly used in the next step. <chemistry id="chem0210" num="0210"><img file="EP2007756B1_D0236.tif" /></chemistry>

2-(7-Fluorobenzo[d][1,3]dioxol-5-yl)acetonitrile

0299A mixture of 6-chloromethyl-4-fluoro-benzo[1,3]dioxole (0.80 g, 4.3 mmol) and NaCN (417 mg, 8.51 mmol) in DMSO (20 mL) was stirred at 30 °C for 1 h and was then poured into water. The mixture was extracted with EtOAc (50 mL × 3). The combined organic layers were washed with water (50 mL) and brine (50 mL), dried over Na₂SO₄ and evaporated under reduced pressure to give the crude product, which was purified by column chromatography on silica gel (petroleum ether/ethyl acetate = 10/1) to afford 2-(7-fluorobenzo[d][1,3]dioxol-5-yl)acetonitrile (530 mg, 70%). ¹H NMR (300 MHz, CDCl₃) δ 6.68-6.64 (m, 2 H), 6.05 (s, 2 H), 3.65 (s, 2 H). ¹³C-NMR δ151.1, 146.2, 134.1, 124.2, 117.5, 110.4, 104.8, 102.8, 23.3.

Example 21: 1-(1H-Indol-5-yl)cyclopropanecarboxylic acid

0300<chemistry id="chem0211" num="0211"><img file="EP2007756B1_D0237.tif" /></chemistry><chemistry id="chem0212" num="0212"><img file="EP2007756B1_D0238.tif" /></chemistry>

Methyl 1-phenylcyclopropanecarboxylate

0301To a solution of 1-phenylcyclopropanecarboxylic acid (25 g, 0.15 mol) in CH₃OH (200 mL) was added TsOH (3 g, 0.1 mol) at room temperature. The mixture was refluxed overnight. The solvent was evaporated under reduced pressure to give crude product, which was dissolved into EtOAc. The EtOAc layer was washed with aq. sat. NaHCO₃. The organic layer was dried over anhydrous Na₂SO₄ and evaporated under reduced pressure to give methyl 1-phenylcyclopropanecarboxylate (26 g, 96%), which was used directly in the next step. ¹H NMR (400 MHz, CDCl₃) δ 7.37-7.26 (m, 5 H), 3.63 (s, 3 H), 1.63-1.60 (m, 2 H), 1.22-1.19 (m, 2 H). <chemistry id="chem0213" num="0213"><img file="EP2007756B1_D0239.tif" /></chemistry>

Methyl 1-(4-nitrophenyl)cyclopropanecarboxylate

0302To a solution of 1-phenylcyclopropanecarboxylate (20.62 g, 0.14 mol) in H₂SO₄/CH₂Cl₂ (40 mL/40 mL) was added KNO₃ (12.8 g, 0.13 mol) in portion at 0 °C. The mixture was stirred for 0.5 hr at 0 °C. Ice water was added and the mixture was extracted with EtOAc (100 mL × 3). The organic layers were dried with anhydrous Na₂SO₄ and evaporated to give methyl 1-(4-nitrophenyl)cyclopropanecarboxylate (21 g, 68%), which was used directly in the next step. ¹H NMR (300 MHz, CDCl₃) δ 8.18 (dd, <i>J</i> = 2.1, 6.9 Hz, 2 H), 7.51 (dd, <i>J</i> = 2.1, 6.9 Hz, 2 H), 3.64 (s, 3 H), 1.72-1.69 (m, 2 H), 1.25-122 (m, 2 H). <chemistry id="chem0214" num="0214"><img file="EP2007756B1_D0240.tif" /></chemistry>

Methyl 1-(4-aminophenyl)cyclopropanecarboxylate

0303To a solution of methyl 1-(4-nitrophenyl)cyclopropanecarboxylate (20 g, 0.09 mol) in MeOH (400 mL) was added Ni (2 g) under nitrogen atmosphere. The mixture was stirred under hydrogen atmosphere (1 atm) at room temperature overnight. The catalyst was filtered off through a pad of Celite and the filtrate was evaporated under <i>vacuum</i> to give crude product, which was purified by chromatography column on silica gel (petroleum ether/ethyl acetate=10:1) to give methyl 1-(4-aminophenyl)cyclopropanecarboxylate (11.38 g, 66%). ¹H NMR (300 MHz, CDCl₃) δ 7.16 (d, <i>J</i> = 8.1 Hz, 2 H), 6.86 (d, <i>J</i> = 7.8 Hz, 2 H), 4.31 (br, 2 H), 3.61 (s, 3 H), 1.55-1.50 (m, 2 H), 1.30-1.12 (m, 2 H). <chemistry id="chem0215" num="0215"><img file="EP2007756B1_D0241.tif" /></chemistry>

Methyl 1-(4-amino-3-bromophenyl)cyclopropanecarboxylate

0304To a solution of methyl 1-(4-aminophenyl)cyclopropanecarboxylate (10.38 g, 0.05 mol) in acetonitrile (200 mL) was added NBS (9.3 g, 0.05 mol) at room temperature. The mixture was stirred overnight. Water (200 mL) was added. The organic layer was separated and the aqueous phase was extracted with EtOAc (80 mL ×3). The organic layers were dried with anhydrous Na₂SO₄ and evaporated to give methyl 1-(4-amino-3-bromophenyl)cyclopropanecarboxylate (10.6 g, 78%), which was used directly in the next step. ¹H NMR (400 MHz, CDCl₃) δ 7.38 (d, <i>J</i>= 2.0 Hz, 1 H), 7.08 (dd, <i>J</i>= 1.6, 8.4 Hz, 1 H), 6.70 (d, <i>J</i> = 8.4 Hz, 1 H), 3.62 (s, 3 H), 1.56-1.54 (m, 2 H), 1.14-1.11(m, 2 H). <chemistry id="chem0216" num="0216"><img file="EP2007756B1_D0242.tif" /></chemistry>

Methyl 1-(4-amino-3-((trimethylsilyl)ethynyl)phenyl)cyclopropane carboxylate

0305To a degassed solution of methyl 1-(4-amino-3-bromophenyl)cyclopropane carboxylate (8 g, 0.03 mol) in Et₃N (100 mL) was added ethynyl-trimethyl-silane (30 g, 0.3 mol), DMAP (5% mol) and Pd(PPh₃)₂Cl₂ (5% mol) under N₂. The mixture was refluxed at 70 °C overnight. The insoluble solid was filtered off and washed with EtOAc (100 mL × 3). The filtrate was evaporated under reduced pressure to give a residue, which was purified by chromatography column on silica gel (petroleum ether/ethyl acetate =20:1) to give methyl 1-(4-amino-3-((trimethylsilyl)ethynyl)phenyl)cyclopropanecarboxylate (4.8 g, 56%). ¹H NMR (300 MHz, CDCl₃) δ7.27 (s, 1 H), 7.10 (dd, <i>J</i> = 2.1, 8.4 Hz, 1 H), 6.64 (d, <i>J</i>= 8.4 Hz, 1 H), 3.60 (s, 3 H), 1.55-1.51 (m, 2 H), 1.12-1.09 (m, 2 H), 0.24 (s, 9 H). <chemistry id="chem0217" num="0217"><img file="EP2007756B1_D0243.tif" /></chemistry>

Methyl 1-(1H-indol-5-yl)cyclopropanecarboxylate

0306To a degassed solution of methyl 1-(4-amino-3-((trimethylsilyl)ethynyl)phenyl) cyclopropanecarboxylate (4.69 g, 0.02 mol) in DMF (20 mL) was added CuI (1.5 g, 0.008 mol) under N₂ at room temperature. The mixture was stirred for 3 hr at room temperature. The insoluble-solid was filtered off and washed with EtOAc (50 mL × 3). The filtrate was evaporated under reduced pressure to give a residue, which was purified by chromatography column on silica gel (petroleum ether/ethyl acetate =20:1) to give methyl 1-(1H-indol-5-yl)cyclopropanecarboxylate (2.2 g, 51%). ¹H NMR (400 MHz, CDCl₃) δ 7.61 (s, 1 H), 7.33 (d, <i>J</i>= 8.4 Hz, 1 H), 7.23-7.18 (m, 2 H), 6.52-6.51 (m, 1 H) 3.62 (s, 3 H), 1.65-1.62 (m, 2 H), 1.29-1.23(m, 2 H). <chemistry id="chem0218" num="0218"><img file="EP2007756B1_D0244.tif" /></chemistry>

1-(1H-Indol-5-yl)cyclopropanecarboxylic acid

0307To a solution of methyl 1-(1H-indol-5-yl)cyclopropanecarboxylate (1.74 g, 8 mmol) in CH₃OH (50 m L) and water (20 mL) was added LiOH (1.7 g, 0.04 mol). The mixture was heated at 45 °C for 3 hr. Water was added and the mixture was acidified with concentrated HCl to pH ~3 before being extracted with EtOAc (20 mL × 3). The organic layers were dried over anhydrous Na₂SO₄ and evaporated to give 1-(1H-indol-5-yl)cyclopropanecarboxylic acid (1.4 g, 87%). ¹H NMR (300 MHz, DMSO-<i>d</i>₆) 7.43 (s, 1 H), 7.30-7.26 (m, 2 H), 7.04 (dd, <i>J</i>= 1.5, 8.4 Hz, 1 H), 6.35 (s, 1 H), 1.45-1.41 (m, 2 H), 1.14-1.10 (m, 2H).

Example 22: 1-(4-Oxochroman-6-yl)cyclopropanecarboxylic acid

0308<chemistry id="chem0219" num="0219"><img file="EP2007756B1_D0245.tif" /></chemistry><chemistry id="chem0220" num="0220"><img file="EP2007756B1_D0246.tif" /></chemistry>

1-[4-(2-<i>tert</i>-Butoxycarbonyl-ethoxy)-phenyl]-cyclopropanecarboxylic methyl

ester

0309To a solution of 1-(4-hydroxy-phenyl)-cyclopropanecarboxylic methyl ester (7.0 g, 3.6 mmol) in acrylic <i>tert</i>-butyl ester (50 mL) was added Na (42 mg, 1.8 mmol) at room temperature. The mixture was heated at 110 °C for 1 h. After cooling to room temperature, the resulting mixture was quenched with water and extracted with EtOAc (100 mL × 3). The combined organic extracts were dried over anhydrous Na₂SO₄ and evaporated under <i>vacuum</i> to give the crude product, which was purified by column chromatography on silica gel (petroleum ether/ethyl acetate 20:1) to give 1-[4-(2-<i>tert</i>-butoxycarbonyl-ethoxy)-phenyl]-cyclopropanecarboxylic methyl ester (6.3 g, 54%) and unreacted start material (3.0 g). ¹H NMR (300 MHz, CDCl₃) δ 7.24 (d, <i>J</i> = 8.7 Hz, 2 H), 6.84 (d, <i>J</i> = 8.7 Hz, 2 H), 4.20 (t, <i>J</i> = 6.6 Hz, 2 H), 3.62 (s, 3 H), 2.69 (t, <i>J</i>= 6.6 Hz, 2 H), 1.59-1.56 (m, 2 H), 1.47 (s, 9 H), 1.17-1.42 (m, 2H). <chemistry id="chem0221" num="0221"><img file="EP2007756B1_D0247.tif" /></chemistry>

1-[4-(2-Carboxy-ethoxy)-phenyl]-cyclopropanecarboxylic methyl ester

0310A solution of 1-[4-(2-<i>tert</i>-butoxycarbonyl-ethoxy)-phenyl]-cyclopropanecarboxylic methyl ester (6.3 g, 20 mmol) in HCl (20%, 200 mL) was heated at 110 °C for 1 h. After cooling to room temperature, the resulting mixture was filtered. The solid was washed with water and dried under <i>vacuum</i> to give 1-[4-(2-carboxy-ethoxy)-phenyl]-cyclopropanecarboxylic methyl ester (5.0 g, 96%). ¹H NMR (300 MHz, DMSO) δ 7.23-7.19 (m, 2 H), 6.85-6.81 (m, 2 H), 4.13 (t, <i>J</i> = 6.0 Hz, 2 H), 3.51 (s, 3 H), 2.66 (t, <i>J</i> = 6.0 Hz, 2 H), 1.43-1.39 (m, 2 H), 1.14-1.10 (m, 2 H). <chemistry id="chem0222" num="0222"><img file="EP2007756B1_D0248.tif" /></chemistry>

1-(4-Oxochroman-6-yl)cyclopropanecarboxylic acid

0311To a solution of 1-[4-(2-carboxy-ethoxy)-phenyl]-cyclopropanecarboxylic methyl ester (5.0 g, 20 mmol) in CH₂Cl₂ (50 mL) were added oxalyl chloride (4.8 g, 38 mmol) and two drops of DMF at 0 °C. The mixture was stirred at 0∼5 °C for 1 h and then evaporated under <i>vacuum.</i> To the resulting mixture was added CH₂Cl₂ (50 mL) at 0 °C and stirring was continued at 0∼5 °C for 1 h. The reaction was slowly quenched with water and was extracted with EtOAc (50 mL × 3). The combined organic extracts were dried over anhydrous Na₂SO₄ and evaporated under <i>vacuum</i> to give the crude product, which was purified by column chromatography on silica gel (petroleum ether/ethyl acetate 20:1-2:1) to give 1-(4-oxochroman-6-yl)cyclopropanecarboxylic acid (830 mg, 19%) and methyl 1-(4-oxochroman-6-yl)cyclopropanecarboxylate (1.8 g, 38%). 1-(4-Oxochroman-6-yl)cyclopropane-carboxylic acid: ¹H NMR (400 MHz, DMSO) δ 12.33 (br s, 1 H), 7.62 (d, <i>J=</i> 2.0 Hz, 1 H), 7.50 (dd, <i>J=</i> 2.4, 8.4 Hz, 1 H), 6.95 (d, <i>J=</i> 8.4 Hz, 1 H), 4.50 (t, <i>J =</i> 6.4 Hz, 2 H), 2.75 (t, <i>J=</i> 6.4 Hz, 2 H), 1.44-1.38 (m, 2 H), 1.10-1.07 (m, 2H). MS (ESI) m/z (M+H⁺) 231.4. 1-(4-Oxochroman-6-yl)cyclopropanecarboxylate: ¹H NMR (400 MHz, CDCl₃) δ 7.83 (d, <i>J</i> = 2.4 Hz, 1 H), 7.48 (dd, <i>J</i> = 2.4, 8.4 Hz, 1 H), 6.93 (d, <i>J</i> = 8.4 Hz, 1 H), 4.55-4.52 (m, 2 H), 3.62 (s, 3 H), 2.80 (t, <i>J</i> = 6.4 Hz, 2 H), 1.62-1.56 (m, 2 H), 1.18-1.15 (m, 2H).

Example 23: 1-(4-Hydroxy-4-methoxychroman-6-yl)cyclopropanecarboxylic acid

0312<chemistry id="chem0223" num="0223"><img file="EP2007756B1_D0249.tif" /></chemistry>

1-(4-Hydroxy-4-methoxychroman-6-yl)cyclopropanecarboxylic acid

0313To a solution of methyl 1-(4-oxochroman-6-yl)cyclopropanecarboxylate (1.0 g, 4.1 mmol) in MeOH (20 mL) and water (20 mL) was added LiOH·H₂O (0.70 g, 16 mmol) in portions at room temperature. The mixture was stirred overnight at room temperature before the MeOH was removed by evaporation under <i>vacuum.</i> Water and Et₂O were added to the residue and the aqueous layer was separated, acidified with HCl and extracted with EtOAc (50 mL × 3). The combined organic extracts dried over anhydrous Na₂SO₄ and evaporated under <i>vacuum</i> to give 1-(4-hydroxy-4-methoxychroman-6-yl)cyclopropanecarboxylic acid (480 mg, 44%). ¹H NMR (400 MHz, CDCl₃) δ 12.16 (s, 1 H), 7.73 (d, <i>J=</i> 2.0 Hz, 1 H), 7.47 (dd, <i>J=</i> 2.0, 8.4 Hz, 1 H), 6.93 (d, <i>J=</i> 8.8 Hz, 1 H), 3.83-3.80 (m, 2 H), 3.39 (s, 3 H), 3.28-3.25 (m, 2 H), 1.71-1.68 (m, 2 H), 1.25-1.22 (m, 2H). MS (ESI) m/z (M+H⁺) 263.1.

Example 24: 1-(4-Hydroxy-4-methoxychroman-6-yl)cyclopropanecarboxylic

acid

0314<chemistry id="chem0224" num="0224"><img file="EP2007756B1_D0250.tif" /></chemistry>

1-Chroman-6-yl-cyclopropanecarboxylic methyl ester

0315To trifluoroacetic acid (20 mL) was added NaBH₄ (0.70 g, 130 mmol) in portions at 0 °C under N₂ atmosphere. After stirring for 5 min, a solution of 1-(4-oxo-chroman-6-yl)-cyclopropanecarboxylic methyl ester (1.6 g, 6.5 mmol) was added at 15 °C. The reaction mixture was stirred for 1 h at room temperature before being slowly quenched with water. The resulting mixture was extracted with EtOAc (50 mL × 3). The combined organic extracts dried over anhydrous Na₂SO₄ and evaporated under <i>vacuum</i> to give 1-chroman-6-yl-cyclopropanecarboxylic methyl ester. (1.4 g, 92%), which was used directly in the next step. ¹H NMR (300 MHz, CDCl₃) δ 7.07-7.00 (m, 2 H), 6.73 (d, <i>J</i> = 8.4 Hz, 1 H), 4.17 (t, <i>J</i> = 5.1 Hz, 2 H), 3.62 (s, 3 H), 2.79-2.75 (m, 2 H), 2.05-1.96 (m, 2 H), 1.57-1.54 (m, 2 H), 1.16-1.13 (m, 2H). <chemistry id="chem0225" num="0225"><img file="EP2007756B1_D0251.tif" /></chemistry>

1-(4-Hydroxy-4-methoxychroman-6-yl)cyclopropanecarboxylic acid

0316To a solution of 1-chroman-6-yl-cyclopropanecarboxylic methyl ester (1.4 g, 60 mmol) in MeOH (20 mL) and water (20 mL) was added LiOH·H₂O (1.0 g, 240 mmol) in portions at room temperature. The mixture was stirred overnight at room temperature before the MeOH was removed by evaporation under <i>vacuum.</i> Water and Et₂O were added and the aqueous layer was separated, acidified with HCl and extracted with EtOAc (50 mL × 3). The combined organic extracts dried over anhydrous Na₂SO₄ and evaporated under <i>vacuum</i> to give 1-(4-Hydroxy-4-methoxychroman-6-yl)cyclopropanecarboxylic acid (1.0 g, 76%). ¹H NMR (400 MHz, DMSO) δ 12.10 (br s, 1 H), 6.95 (d, <i>J</i> = 2.4 Hz, 2 H), 6.61-6.59 (m, 1 H), 4.09-4.06 (m, 2 H), 2.70-2.67 (m, 2 H), 1.88-1.86 (m, 2 H), 1.37-1.35 (m, 2 H), 1.04-1.01 (m, 2H). MS (ESI) m/z (M+H⁺) 217.4.

Example 25: 1-(3-Methylbenzo[d]isoxazol-5-yl)cyclopropanecarboxylic acid

0317<chemistry id="chem0226" num="0226"><img file="EP2007756B1_D0252.tif" /></chemistry><chemistry id="chem0227" num="0227"><img file="EP2007756B1_D0253.tif" /></chemistry>

1-(3-Acetyl-4-hydroxy-phenyl)-cyclopropanecarboxylic methyl ester

0318To a stirred suspension of AlCl₃ (58 g, 440 mmol) in CS₂ (500 mL) was added acetyl chloride (7.4 g, 95 mmol) at room temperature. After stirring for 5 min, methyl 1-(4-methoxyphenyl)cyclopropanecarboxylate (15 g, 73 mmol) was added. The reaction mixture was heated at reflux for 2 h before ice water was added carefully to the mixture at room temperature. The resulting mixture was extracted with EtOAc (150 mL × 3). The combined organic extracts were dried over anhydrous Na₂SO₄ and evaporated under reduced pressure to give 1-(3-acetyl-4-hydroxy-phenyl)-cyclopropanecarboxylic methyl ester (15 g, 81%), which was used in the next step without further purification. ¹H NMR (CDCl₃, 400 MHz) δ 12.28 (s, 1 H), 7.67 (d, <i>J</i> = 2.0 Hz, 1 H), 7.47 (dd, <i>J</i> = 2.0, 8.4 Hz, 1 H), 6.94 (d, <i>J</i> = 8.4 Hz, 1 H), 3.64 (s, 3 H), 2.64 (s, 3 H), 1.65-1.62 (m, 2 H), 1.18-1.16 (m, 2 H). <chemistry id="chem0228" num="0228"><img file="EP2007756B1_D0254.tif" /></chemistry>

1-[4-Hydroxy-3-(1-hydroxyimino-ethyl)-phenyl]-cyclopropanecarboxylic methyl ester

0319To a stirred solution of 1-(3-acetyl-4-hydroxy-phenyl)-cyclopropanecarboxylic methyl ester (14.6 g, 58.8 mmol) in EtOH (500 mL) were added hydroxylamine hydrochloride (9.00 g, 129 mmol) and sodium acetate (11.6 g, 141 mmol) at room temperature. The resulting mixture was heated at reflux overnight. After removal of EtOH under <i>vacuum,</i> water (200 mL) and EtOAc (200 mL) were added. The organic layer was separated and the aqueous layer was extracted with EtOAc (100 mL × 3). The combined organic layers were dried over anhydrous Na₂SO₄ and evaporated under <i>vacuum</i> to give 1-[4-hydroxy-3-(1-hydroxyimino-ethyl)-phenyl]-cyclopropanecarboxylic methyl ester (14.5 g, 98%), which was used in the next step without further purification. ¹H NMR (CDCl₃, 400 MHz) δ 11.09 (s, 1 H), 7.39 (d, <i>J=</i> 2.0 Hz, 1 H), 7.23 (d, <i>J=</i> 2.0 Hz, 1 H), 7.14 (s, 1 H), 6.91 (d, <i>J=</i> 8.4 Hz, 1 H), 3.63 (s, 3 H), 2.36 (s, 3 H), 1.62-1.59 (m, 2 H), 1.18-1.15 (m, 2 H). <chemistry id="chem0229" num="0229"><img file="EP2007756B1_D0255.tif" /></chemistry>

(<i>E</i>)<i>-</i>Methyl 1-(3-(1-(acetoxyimino)ethyl)-4-hydroxyphenyl)cyclopropane carboxylate

0320The solution of 1-[4-hydroxy-3-(1-hydroxyimino-ethyl)-phenyl]-cyclopropanecarboxylic methyl ester (10.0 g, 40.1 mmol) in Ac₂O (250 mL) was heated at 45 °C for 4 h. The Ac₂O was removed by evaporation under <i>vacuum</i> before water (100 mL) and EtOAc (100 mL) were added. The organic layer was separated and the aqueous layer was extracted with EtOAc (100 mL × 2). The combined organic layers were dried over anhydrous Na₂SO₄ and evaporated under <i>vacuum</i> to give (<i>E</i>)-methyl 1-(3-(1-(acetoxyimino)ethyl)-4-hydroxyphenyl)cyclopropanecarboxylate (10.5 g, 99%), which was used in the next step without further purification. <chemistry id="chem0230" num="0230"><img file="EP2007756B1_D0256.tif" /></chemistry>

Methyl 1-(3-methylbenzo[d]isoxazol-5-yl)cyclopropanecarboxylate

0321A solution of (<i>E</i>)-methyl 1-(3-(1-(acetoxyimino)ethyl)-4-hydroxyphenyl)cyclopropane carboxylate (10.5 g, 39.6 mmol) and pyridine (31.3 g, 396 mmol) in DMF (150 mL) was heated at 125 °C for 10 h. The cooled reaction mixture was poured into water (250 mL) and was extracted with EtOAc (100 mL × 3). The combined organic layers were dried over anhydrous Na₂SO₄ and evaporated under <i>vacuum</i> to give the crude product, which was purified by column chromatography on silica gel (petroleum ether/ethyl acetate 50:1) to give methyl 1-(3-methylbenzo[d]isoxazol-5-yl)cyclopropanecarboxylate (7.5 g, 82%). ¹H NMR (CDCl₃ 300 MHz) δ 7.58-7.54 (m, 2 H), 7.48 (dd, <i>J</i> = 1.5, 8.1 Hz, 1 H), 3.63 (s, 3 H), 2.58 (s, 3 H), 1.71-1.68 (m, 2 H), 1.27-1.23 (m, 2 H). <chemistry id="chem0231" num="0231"><img file="EP2007756B1_D0257.tif" /></chemistry>

1-(3-Methylbenzo[d]isoxazol-5-yl)cyclopropanecarboxylic acid

0322To a solution of methyl 1-(3-methylbenzo[d]isoxazol-5-yl)cyclopropanecarboxylate (1.5 g, 6.5 mmol) in MeOH (20 mL) and water (2 mL) was added LiOH·H₂O (0.80 g, 19 mmol) in portions at room temperature. The reaction mixture was stirred at room temperature overnight before the MeOH was removed by evaporation under <i>vacuum.</i> Water and Et₂O were added and the aqueous layer was separated, acidified with HCl and extracted with EtOAc (50 mL × 3). The combined organic extracts were dried over anhydrous Na₂SO₄ and evaporated under <i>vacuum</i> to give 1-(3-methylbenzo[d]isoxazol-5-yl)cyclopropanecarboxylic acid (455 mg, 32%). ¹H NMR (400 MHz, DMSO) δ 12.40 (br s, 1 H), 7.76 (s, 1 H), 7.60-7.57 (m, 2 H), 2.63 (s, 3 H), 1.52-1.48 (m, 2 H), 1.23-1.19 (m, 2H). MS (ESI) m/z (M+H⁺) 218.1.

Example 26: 1-(Spiro[benzo[d][1,3]dioxole-2,1'-cyclobutane]-5-yl)cyclopropane carboxylic acid

0323<chemistry id="chem0232" num="0232"><img file="EP2007756B1_D0258.tif" /></chemistry><chemistry id="chem0233" num="0233"><img file="EP2007756B1_D0259.tif" /></chemistry>

1-(3,4-Dihydroxy-phenyl)-cyclopropanecarboxylic methyl ester

0324To a solution of 1-(3,4-dihydroxyphenyl)cyclopropanecarboxylic acid (4.5 g) in MeOH (30 mL) was added TsOH (0.25 g, 1.3 mmol). The stirring was continued at 50 °C overnight before the mixture was cooled to room temperature. The mixture was concentrated under <i>vacuum</i> and the residue was purified by column chromatography on silica gel (petroleum ether/ethyl acetate 3:1) to give 1-(3,4-dihydroxy-phenyl)-cyclopropanecarboxylic methyl ester (2.1 g). ¹H NMR (DMSO 300 MHz) δ 8.81 (brs, 2 H), 6.66 (d, <i>J</i> = 2.1 Hz, 1 H), 6.61 (d, <i>J</i> = 8.1 Hz, 1 H), 6.53 (dd, <i>J =</i> 2.1, 8.1 Hz, 1 H), 3.51 (s, 3 H), 1.38-1.35 (m, 2 H), 1.07-1.03 (m, 2 H). <chemistry id="chem0234" num="0234"><img file="EP2007756B1_D0260.tif" /></chemistry>

Methyl 1-(spiro[benzo[d][1,3]dioxole-2,1'-cyclobutane]-5-yl)cyclopropane carboxylate

0325To a solution of 1-(3,4-dihydroxy-phenyl)-cyclopropanecarboxylic methyl ester (1.0 g, 4.8 mmol) in toluene (30 mL) was added TsOH (0.10 g, 0.50 mmol) and cyclobutanone (0.70 g, 10 mmol). The reaction mixture was heated at reflux for 2 h before being concentrated under <i>vacuum.</i> The residue was purified by chromatography on silica gel (petroleum ether/ethyl acetate 15:1) to give methyl 1-(spiro[benzo[d][1,3]dioxole-2,1'-cyclobutane]-5-yl)cyclopropanecarboxylate (0.6 g, 50%). ¹H NMR (CDCl₃ 300 MHz) δ 6.78-6.65 (m, 3 H), 3.62 (s, 3 H), 2.64-2.58 (m, 4 H), 1.89-1.78 (m, 2 H), 1.56-1.54 (m, 2 H), 1.53-1.12(m, 2 H). <chemistry id="chem0235" num="0235"><img file="EP2007756B1_D0261.tif" /></chemistry>

1-(Spiro[benzo[d][1,3]dioxole-2,1'-cyclobutane]-5-yl)cyclopropane carboxylic acid

0326To a mixture of methyl 1-(spiro[benzo[d][1,3]dioxole-2,1'-cyclobutane]-5-yl)cyclopropanecarboxylate (0.60 g, 2.3 mmol) in THF/H₂O (4:1, 10 mL) was added LiOH (0.30 g, 6.9 mmol). The mixture was stirred at 60 °C for 24 h. HCl (0.5 N) was added slowly to the mixture at 0 °C until pH 2-3. The mixture was extracted with EtOAc (10 mL × 3). The combined organic phases were washed with brine, dried over anhydrous MgSO₄, and washed with petroleum ether to give 1-(spiro[benzo[d][1,3]-dioxole-2,1'-cyclobutane]-5-yl)cyclopropane carboxylic acid (330 mg, 59%). ¹HNMR (400 MHz, CDCl₃) δ 6.78-6.65 (m, 3 H), 2.65-2.58 (m, 4 H), 1.86-1.78 (m, 2 H), 1.63-1.60 (m, 2 H), 1.26-1.19 (m, 2 H).

Example 27: 2-(2,3-Dihydrobenzo[b][1,4]dioxin-6-yl)acetonitrile

0327<chemistry id="chem0236" num="0236"><img file="EP2007756B1_D0262.tif" /></chemistry><chemistry id="chem0237" num="0237"><img file="EP2007756B1_D0263.tif" /></chemistry>

2,3-Dihydro-benzo[1,4]dioxine-6-carboxylic acid ethyl ester

0328To a suspension of Cs₂CO₃ (270 g, 1.49 mol) in DMF (1000 mL) were added 3,4-dihydroxybenzoic acid ethyl ester (54.6 g, 0.3 mol) and 1,2-dibromoethane (54.3 g, 0.29 mol) at room temperature. The resulting mixture was stirred at 80 °C overnight and then poured into ice-water. The mixture was extracted with EtOAc (200 mL × 3). The combined organic layers were washed with water (200 mL × 3) and brine (100 mL), dried over Na₂SO₄ and concentrated to dryness. The residue was purified by column (petroleum ether/ethyl acetate 50:1) on silica gel to obtain 2,3-dihydro-benzo[1,4]dioxine-6-carboxylic acid ethyl ester (18 g, 29%). ¹H NMR (300 MHz, CDCl₃) δ 7.53 (dd, J = 1.8, 7.2 Hz, 2 H), 6.84-6.87 (m, 1 H), 4.22-4.34 (m, 6 H), 1.35 (t, J = 7.2 Hz, 3 H). <chemistry id="chem0238" num="0238"><img file="EP2007756B1_D0264.tif" /></chemistry>

(2,3-Dihydro-benzo[1,4]dioxin-6-yl)-methanol

0329To a suspension of LiAlH₄ (2.8 g, 74 mmol) in THF (20 mL) was added dropwise a solution of 2,3-dihydro-benzo[1,4]dioxine-6-carboxylic acid ethyl ester (15 g, 72 mmol) in THF (10 mL) at 0 °C under N₂. The mixture was stirred at room temperature for 1 h and then quenched carefully with addition of water (2.8 mL) and NaOH (10%, 28 mL) with cooling. The precipitated solid was filtered off and the filtrate was evaporated to dryness to obtain (2,3-dihydro-benzo[1,4]dioxin-6-yl)-methanol (10.6 g). ¹H NMR (300 MHz, DMSO-d₆) δ 6.73-6.78 (m, 3 H), 5.02 (t, J = 5.7 Hz, 1 H), 4.34 (d, J = 6.0 Hz, 2 H), 4.17-4.20 (m, 4 H). <chemistry id="chem0239" num="0239"><img file="EP2007756B1_D0265.tif" /></chemistry>

6-Chloromethyl-2,3-dihydro-benzo[1,4]dioxine

0330A mixture of (2,3-dihydro-benzo[1,4]dioxin-6-yl)methanol (10.6 g) in SOCl₂ (10 mL) was stirred at room temperature for 10 min and then poured into ice-water. The organic layer was separated and the aqueous phase was extracted with dichloromethane (50 mL × 3). The combined organic layers were washed with NaHCO₃ (sat solution), water and brine, dried over Na₂SO₄ and concentrated to dryness to obtain 6-chloromethyl-2,3-dihydro-benzo[1,4]dioxine (12 g, 88% over two steps), which was used directly in next step. <chemistry id="chem0240" num="0240"><img file="EP2007756B1_D0266.tif" /></chemistry>

2-(2,3-Dihydrobenzo[b][1,4]dioxin-6-yl)acetonitrile

0331A mixture of 6-chloromethyl-2,3-dihydro-benzo[1,4]dioxine (12.5 g, 67.7 mmol) and NaCN (4.30 g, 87.8 mmol) in DMSO (50 mL) was stirred at rt for 1 h. The mixture was poured into water (150 mL) and then extracted with dichloromethane (50 mL × 4). The combined organic layers were washed with water (50 mL × 2) and brine (50 mL), dried over Na₂SO₄ and concentrated to dryness. The residue was purified by column (petroleum ether/ethyl acetate 50:1) on silica gel to obtain 2-(2,3-dihydrobenzo[b][1,4]dioxin-6-yl)acetonitrile as a yellow oil (10.2 g, 86%). ¹H-NMR (300 MHz, CDCl₃) δ 6.78-6.86 (m, 3 H), 4.25 (s, 4 H), 3.63 (s, 2 H).

0332The following Table 2 contains a list of carboxylic acid building blocks that were commercially available, or prepared by one of the three methods described above: <tables id="tabl0027" num="0027"><table frame="all"><title>Table 2: Carboxylic acid building blocks.</title><tgroup cols="2"><colspec colnum="1" colname="col1" colwidth="117mm" /><colspec colnum="2" colname="col2" colwidth="49mm" /><thead><row><entry align="center" valign="middle"><b>Name</b></entry><entry align="center" valign="middle"><b>Structure</b></entry></row></thead><tbody><row><entry align="center" valign="middle">1-benzo[1,3]dioxol-5-ylcyclopropane-1-carboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0241" num="0241"><img file="EP2007756B1_D0267.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(2,2-difluorobenzo[1,3]dioxol-5-yl)cyclopropane-1-carboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0242" num="0242"><img file="EP2007756B1_D0268.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(3,4-dihydroxyphenyl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0243" num="0243"><img file="EP2007756B1_D0269.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(3-methoxyphenyl)cyclopropane-1-carboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0244" num="0244"><img file="EP2007756B1_D0270.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(2-methoxyphenyl)cyclopropane-1-carboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0245" num="0245"><img file="EP2007756B1_D0271.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-[4-(trifluoromethoxy)phenyl]cyclopropane-1-carboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0246" num="0246"><img file="EP2007756B1_D0272.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(2,2-dimethylbenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0247" num="0247"><img file="EP2007756B1_D0273.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">tetrahydro-4-(4-methoxyphenyl)-2<i>H-</i>pyran-4-carboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0248" num="0248"><img file="EP2007756B1_D0274.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-phenylcyclopropane-1-carboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0249" num="0249"><img file="EP2007756B1_D0275.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(4-methoxyphenyl)cyclopropane-1-carboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0250" num="0250"><img file="EP2007756B1_D0276.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(4-chlorophenyl)cyclopropane-1-carboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0251" num="0251"><img file="EP2007756B1_D0277.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(3-hydroxyphenyl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0252" num="0252"><img file="EP2007756B1_D0278.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-phenylcyclopentanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0253" num="0253"><img file="EP2007756B1_D0279.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(2-oxo-2,3-dihydrobenzo[d]oxazol-5-yl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0254" num="0254"><img file="EP2007756B1_D0280.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(benzofuran-5-yl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0255" num="0255"><img file="EP2007756B1_D0281.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(4-methoxyphenyl)cyclohexanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0256" num="0256"><img file="EP2007756B1_D0282.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(4-chlorophenyl)cyclohexanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0257" num="0257"><img file="EP2007756B1_D0283.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(2,3-dihydrobenzofuran-5-yl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0258" num="0258"><img file="EP2007756B1_D0284.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(3,3-dimethyl-2,3-dihydrobenzofuran-5-yl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0259" num="0259"><img file="EP2007756B1_D0285.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(7-methoxybenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0260" num="0260"><img file="EP2007756B1_D0286.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(3-hydroxy-4-methoxyphenyl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0261" num="0261"><img file="EP2007756B1_D0287.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(4-chloro-3-hydroxyphenyl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0262" num="0262"><img file="EP2007756B1_D0288.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(3-(benzyloxy)-4-chlorophenyl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0263" num="0263"><img file="EP2007756B1_D0289.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(4-chlorophenyl)cyclopentanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0264" num="0264"><img file="EP2007756B1_D0290.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(3-(benzyloxy)-4-methoxyphenyl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0265" num="0265"><img file="EP2007756B1_D0291.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(3-chloro-4-methoxyphenyl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0266" num="0266"><img file="EP2007756B1_D0292.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(3-fluoro-4-methoxyphenyl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0267" num="0267"><img file="EP2007756B1_D0293.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(4-methoxy-3-methylphenyl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0268" num="0268"><img file="EP2007756B1_D0294.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(4-(benzyloxy)-3-methoxyphenyl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0269" num="0269"><img file="EP2007756B1_D0295.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(4-chloro-3-methoxyphenyl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0270" num="0270"><img file="EP2007756B1_D0296.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(3-chloro-4-hydroxyphenyl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0271" num="0271"><img file="EP2007756B1_D0297.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(3-(hydroxymethyl)-4-methoxyphenyl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0272" num="0272"><img file="EP2007756B1_D0298.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(4-methoxyphenyl)cyclopentanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0273" num="0273"><img file="EP2007756B1_D0299.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-phenylcyclohexanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0274" num="0274"><img file="EP2007756B1_D0300.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(3,4-dimethoxyphenyl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0275" num="0275"><img file="EP2007756B1_D0301.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(7-chlorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0276" num="0276"><img file="EP2007756B1_D0302.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(benzo[djoxazol-5-yl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0277" num="0277"><img file="EP2007756B1_D0303.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(7-fluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0278" num="0278"><img file="EP2007756B1_D0304.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(3,4-difluorophenyl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0279" num="0279"><img file="EP2007756B1_D0305.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(1H-indol-5-yl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0280" num="0280"><img file="EP2007756B1_D0306.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(1H-benzo[d]imidazol-5-yl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0281" num="0281"><img file="EP2007756B1_D0307.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(2-methyl-1H-benzo[d]imidazol-5-yl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0282" num="0282"><img file="EP2007756B1_D0308.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(-methyl-1H-benzo[d]imidazol-5-yl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0283" num="0283"><img file="EP2007756B1_D0309.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(3-methylbenzo[d]isoxazol-5-yl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0284" num="0284"><img file="EP2007756B1_D0310.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(spiro[benzo[d][1,3]dioxole-2,1'-cyclobutane]-5-yl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0285" num="0285"><img file="EP2007756B1_D0311.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(1H-benzo[d][1,2,3]triazol-5-yl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0286" num="0286"><img file="EP2007756B1_D0312.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(1-methyl-1H-benzo[d][1,2,3]triazol-5-yl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0287" num="0287"><img file="EP2007756B1_D0313.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(1,3-dihydroisobenzofuran-5-yl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0288" num="0288"><img file="EP2007756B1_D0314.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(6-fluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0289" num="0289"><img file="EP2007756B1_D0315.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(2,3-dihydrobenzofuran-6-yl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0290" num="0290"><img file="EP2007756B1_D0316.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(chroman-6-yl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0291" num="0291"><img file="EP2007756B1_D0317.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(4-hydroxy-4-methoxychroman-6-yl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0292" num="0292"><img file="EP2007756B1_D0318.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(4-oxochroman-6-yl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0293" num="0293"><img file="EP2007756B1_D0319.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(3,4-dichlorophenyl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0294" num="0294"><img file="EP2007756B1_D0320.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(2,3-dihydrobenzo[b][1,4]dioxin-6-yl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0295" num="0295"><img file="EP2007756B1_D0321.tif" /></chemistry></entry></row><row><entry align="center" valign="middle">1-(benzofuran-6-yl)cyclopropanecarboxylic acid</entry><entry align="center" valign="middle"><chemistry id="chem0296" num="0296"><img file="EP2007756B1_D0322.tif" /></chemistry></entry></row></tbody></tgroup></table></tables>

Specific Procedures: Synthesis of aminoindole building blocks

Example 28: 3-Methyl-1H-indol-6-amine

0333<chemistry id="chem0297" num="0297"><img file="EP2007756B1_D0323.tif" /></chemistry><chemistry id="chem0298" num="0298"><img file="EP2007756B1_D0324.tif" /></chemistry>

(3-Nitro-phenyl)-hydrazine hydrochloride salt

03343-Nitro-phenylamine (27.6 g, 0.2 mol) was dissolved in the mixture of H₂O (40 mL) and 37% HCl (40 mL). A solution of NaNO₂ (13.8 g, 0.2 mol) in H₂O (60 mL) was added to the mixture at 0 °C, and then a solution of SnCl₂.H₂O (135.5 g, 0.6 mol) in 37% HCl (100 mL) was added at that temperature. After stirring at 0 °C for 0.5 h, the insoluble material was isolated by filtration and was washed with water to give (3-nitrophenyl)hydrazine hydrochloride (27.6 g, 73%). <chemistry id="chem0299" num="0299"><img file="EP2007756B1_D0325.tif" /></chemistry>

<i>N</i>

-

(3-Nitro-phenyl)-<i>N'</i>-propylidene-hydrazine

0335Sodium hydroxide solution (10%, 15 mL) was added slowly to a stirred suspension of (3-nitrophenyl)hydrazine hydrochloride (1.89 g, 10 mmol) in ethanol (20 mL) until pH 6. Acetic acid (5 mL) was added to the mixture followed by propionaldehyde (0.7 g, 12 mmol). After stirring for 3 h at room temperature, the mixture was poured into ice-water and the resulting precipitate was isolated by filtration, washed with water and dried in air to obtain (E)-1-(3-nitrophenyl)-2-propylidenehydrazine, which was used directly in the next step. <chemistry id="chem0300" num="0300"><img file="EP2007756B1_D0326.tif" /></chemistry>

3-Methyl-4-nitro-1H-indole 3 and 3-methyl-6-nitro-1H-indole

0336A mixture of (E)-1-(3-nitrophenyl)-2-propylidenehydrazine dissolved in 85 % H₃PO₄ (20 mL) and toluene (20 mL) was heated at 90-100 °C for 2 h. After cooling, toluene was removed under reduced pressure. The resultant oil was basified to pH 8 with 10 % NaOH. The aqueous layer was extracted with EtOAc (100 mL x 3). The combined organic layers were dried, filtered and concentrated under reduced pressure to afford the mixture of 3-methyl-4-nitro-1H-indole and 3-methyl-6-nitro-1H-indole [1.5 g in total, 86 %, two steps from (3-nitrophenyl)hydrazine hydrochloride] which was used to the next step without further purification. <chemistry id="chem0301" num="0301"><img file="EP2007756B1_D0327.tif" /></chemistry>

3-Methyl-1H-indol-6-amine

0337The crude mixture from previous steps (3 g, 17 mmol) and 10% Pd-C (0.5 g) in ethanol (30 mL) was stirred overnight under H₂ (1 atm) at room temperature. Pd-C was filtered off and the filtrate was concentrated under reduced pressure. The solid residue was purified by column to give 3-methyl-1H-indol-6-amine (0.6 g, 24%). ¹H NMR (CDCl₃) δ 7.59 (br s. 1H), 7.34 (d, <i>J</i> = 8.0 Hz, 1H), 6.77 (s, 1H), 6.64 (s, 1H), 6.57 (m, 1H), 3.57 (brs, 2H), 2.28 (s, 3H); MS (ESI) m/e (M+H⁺) 147.2.

Example 29: 3

<b>-tert</b>-

Butyl-1H-indol-5-amine

0338<chemistry id="chem0302" num="0302"><img file="EP2007756B1_D0328.tif" /></chemistry><chemistry id="chem0303" num="0303"><img file="EP2007756B1_D0329.tif" /></chemistry>

3-<i>tert</i>-Butyl-5-nitro-1H-indole

0339To a mixture of 5-nitro-1H-indole (6.0 g, 37 mmol) and AlCl₃ (24 g, 0.18 mol) in CH₂Cl₂ (100 mL) at 0 °C was added 2-bromo-2-methyl-propane (8.1 g, 37 mmol) dropwise. After being stirred at 15 °C overnight, the mixture was poured into ice (100 mL). The precipitated salts were removed by filtration and the aqueous layer was extracted with CH₂Cl₂ (30 mL × 3). The combined organic layers were washed with water, brine, dried over Na₂SO₄ and concentrated under <i>vacuum</i> to obtain the crude product, which was purified by column chromatography on silica gel (petroleum ether/ethyl acetate = 20: 1) to give 3<i>-tert-</i>butyl-5-nitro-1H-indole (2.5 g, 31%). ¹H NMR (CDCl₃, 400 MHz) δ 8.49 (d, <i>J</i> = 1.6 Hz, 1 H), 8.31 (brs, 1 H), 8.05 (dd, <i>J</i> = 2.0, 8.8 Hz, 1 H), 7.33 (d, <i>J</i> = 8.8 Hz, 1 H), 6.42 (d, <i>J</i> = 1.6 Hz, 1 H), 1.42 (s, 9 H). <chemistry id="chem0304" num="0304"><img file="EP2007756B1_D0330.tif" /></chemistry>

3-<i>tert</i>-Butyl-1H-indol-5-amine

0340To a solution of 3-<i>tert</i>-butyl-5-nitro-1H-indole (2.5 g, 12 mmol) in MeOH (30 mL) was added Raney Nickel (0.2 g) under N₂ protection. The mixture was stirred under hydrogen atmosphere (1 atm) at 15 °C for 1 h. The catalyst was filtered off and the filtrate was concentrated to dryness under <i>vacuum.</i> The residue was purified by preparative HLPC to afford 3-<i>tert</i>-butyl-1H-indol-5-amine (0.43 g, 19%). ¹H NMR (CDCl₃, 400 MHz) δ 7.72 (br.s, 1 H), 7.11 (d, <i>J</i> = 8.4 Hz, 1 H), 6.86 (d, <i>J</i> = 2.0 Hz, 1 H), 6.59 (dd, <i>J</i> = 2.0, 8.4 Hz, 1 H), 6.09 (d, <i>J</i> = 1.6 Hz, 1 H), 1.37 (s, 9 H); MS (ESI) m/e (M+H⁺) 189.1.

Example 30: 2-<i>tert</i>-Butyl-6-fluoro-1H-indol-5-amine and 6-<i>tert</i>-butoxy-2-<i>tert-</i>butyl-1H-indol-5-amine

0341<chemistry id="chem0305" num="0305"><img file="EP2007756B1_D0331.tif" /></chemistry>

2-Bromo-5-fluoro-4-nitroaniline

0342To a mixture of 3-fluoro-4-nitroaniline (6.5 g, 42.2 mmol) in AcOH (80 mL) and chloroform (25 mL) was added dropwise Br₂ (2.15 mL, 42.2 mmol) at 0 °C. After addition, the resulting mixture was stirred at room temperature for 2 h and then poured into ice water. The mixture was basified with aqueous NaOH (10%) to pH ∼ 8.0-9.0 under cooling and then extracted with EtOAc (50 mL × 3). The combined organic layers were washed with water (80 mL × 2) and brine (100 mL), dried over Na₂SO₄ and concentrated under reduced pressure to give 2-bromo-5-fluoro-4-nitroaniline (9 g, 90%). ¹H-NMR (400 MHz, DMSO-<i>d</i>₆) δ 8.26 (d, <i>J</i> = 8.0, Hz, 1H), 7.07 (brs, 2H), 6.62 (d, <i>J</i> = 9.6 Hz, 1H). <chemistry id="chem0306" num="0306"><img file="EP2007756B1_D0332.tif" /></chemistry>

2-(3,3-Dimethylbut-1-ynyl)-5-fluoro-4-nitroaniline

0343A mixture of 2-bromo-5-fluoro-4-nitroaniline (9.0 g, 38.4 mmol), 3,3-dimethyl-but-1-yne (9.95 g, 121 mmol), CuI (0.5 g 2.6 mmol), Pd(PPh₃)₂Cl₂ (3.4 g, 4.86 mmol) and Et₃N (14 mL, 6.9 mmol) in toluene (100 mL) and water (50 mL) was heated at 70 °C for 4 h. The aqueous layer was separated and the organic layer was washed with water (80 mL × 2) and brine (100 mL), dried over Na₂SO₄ and concentrated under reduced pressure to dryness. The residue was recrystallized with ether to afford 2-(3,3-dimethylbut-1-ynyl)-5-fluoro-4-nitroaniline (4.2 g, 46%). ¹H-NMR (400 MHz, DMSO-<i>d</i>₆) δ 7.84 (d, <i>J</i> = 8.4 Hz, 1H), 6.84 (brs, 2H), 6.54 (d, <i>J</i> = 14.4 Hz, 1H), 1.29 (s, 9H). <chemistry id="chem0307" num="0307"><img file="EP2007756B1_D0333.tif" /></chemistry>

<i>N</i>-(2-(3,3-Dimethylbut-1-ynyl)-5-fluoro-4-nitrophenyl)butyramide

0344To a solution of 2-(3,3-dimethylbut-1-ynyl)-5-fluoro-4-nitroaniline (4.2 g, 17.8 mmol) in dichloromethane (50 mL) and Et₃N (10.3 mL, 71.2 mmol) was added butyryl chloride (1.9 g, 17.8 mmol) at 0 °C. The mixture was stirred at room temperature for 1 h and then poured into water. The aqueous phase was separated and the organic layer was washed with water (50 mL × 2) and brine (100 mL), dried over Na₂SO₄ and concentrated under reduced pressure to dryness. The residue was washed with ether to give N-(2-(3,3-dimethylbut-1-ynyl)-5-fluoro-4-nitrophenyl)butyramide (3.5 g, 67%), which was used in the next step without further purification. <chemistry id="chem0308" num="0308"><img file="EP2007756B1_D0334.tif" /></chemistry>

2-<i>tert</i>-Butyl-6-fluoro-5-nitro-1H-indole

0345A solution of N-(2-(3,3-dimethylbut-1-ynyl)-5-fluoro-4-nitrophenyl)butyramide (3.0 g, 9.8 mmol) and TBAF (4.5 g, 17.2 mmol) in DMF (25 mL) was heated at 100 °C overnight. The mixture was poured into water and then extracted with EtOAc (80 mL × 3). The combined extracts were washed with water (50 mL) and brine (50 mL), dried over Na₂SO₄ and concentrated under reduced pressure to dryness. The residue was purified by column chromatography on silica gel (petroleum ether/ethyl acetate 20:1) to give compound 2-tert-butyl-6-fluoro-5-nitro-1H-indole (1.5 g, 65%). ¹H-NMR (400 MHz, CDCl₃) δ 8.30 (d, <i>J</i> = 7.2 Hz, 1H), 7.12 (d, <i>J</i> = 11.6 Hz, 1H), 6.3 5 (d, <i>J</i> = 1.2 Hz, 1H), 1.40 (s, 9H). <chemistry id="chem0309" num="0309"><img file="EP2007756B1_D0335.tif" /></chemistry>

2-<i>tert</i>-Butyl-6-fluoro-1H-indol-5-amine

0346A suspension of 2-tert-butyl-6-fluoro-5-nitro-1H-indole (1.5 g, 6.36 mmol) and Ni (0.5 g) in MeOH (20 mL) was stirred under H₂ atmosphere (1 atm) at the room temperature for 3 h. The catalyst was filtered off and the filtrate was concentrated under reduced pressure to dryness. The residue was recrystallized in ether to give 2-tert-butyl-6-fluoro-1H-indol-5-amine (520 mg, 38%). ¹H-NMR (300 MHz, DMSO-<i>d</i>₆) δ 10.46 (brs, 1H), 6.90 (d, <i>J</i> = 8.7 Hz, 1H), 6.75 (d, <i>J</i> = 9.0 Hz, 1H), 5.86 (s, 1H), 4.37 (brs, 2H), 1.29 (s, 9H); MS (ESI) m/e 206.6. <chemistry id="chem0310" num="0310"><img file="EP2007756B1_D0336.tif" /></chemistry>

6-<i>tert</i>-Butoxy-2-tert-butyl-5-nitro-1H-indole

0347A solution of N-(2-(3,3-dimethylbut-1-ynyl)-5-fluoro-4-nitrophenyl)butyramide (500 mg, 1.63 mmol) and <i>t</i>-BuOK (0.37 g, 3.26 mmol) in DMF (10 mL) was heated at 70 °C for 2 h. The mixture was poured into water and then extracted with EtOAc (50 mL × 3). The combined extracts were washed with water (50 mL) and brine (50 mL), dried over Na₂SO₄ and concentrated under reduced pressure to give 6-tert-butoxy-2-tert-butyl-5-nitro-1H-indole (100 mg, 21%). ¹H-NMR (300 MHz, DMSO-<i>d</i>₆) δ 11.35 (brs, 1H), 7.99 (s, 1H), 7.08 (s, 1H), 6.25 (s, 1H), 1.34 (s, 9H), 1.30 (s, 9H). <chemistry id="chem0311" num="0311"><img file="EP2007756B1_D0337.tif" /></chemistry>

6-<i>tert</i>-Butoxy-2-<i>tert</i>-butyl-1H-indol-5-amine

0348A suspension of 6-<i>tert</i>-butoxy-2-tert-butyl-5-nitro-1H-indole (100 mg, 0.36 mmol) and Raney Ni (0.5 g) in MeOH (15 mL) was stirred under H₂ atmosphere (1 atm) at the room temperature for 2.5 h. The catalyst was filtered off and the filtrate was concentrated under reduced pressure to dryness. The residue was recrystallized in ether to give 6-<i>tert</i>-butoxy-2-<i>tert</i>-butyl-1H-indol-5-amine (30 mg, 32%). ¹H-NMR (300 MHz, MeOD) 6.98 (s, 1H), 6.90 (s, 1H), 5.94 (d, J= 0.6 Hz, 1H), 1.42 (s, 9H), 1.36 (s, 9H); MS (ESI) m/e 205.0.

Example 31: 1-<i>tert</i>-Butyl-1H-indol-5-amine

0349<chemistry id="chem0312" num="0312"><img file="EP2007756B1_D0338.tif" /></chemistry><chemistry id="chem0313" num="0313"><img file="EP2007756B1_D0339.tif" /></chemistry>

<i>N-tert</i>-Butyl-4-nitroaniline

0350A solution of 1-fluoro-4-nitro-benzene (1 g, 7.1 mmol) and tert-butylamine (1.5 g, 21 mmol) in DMSO (5 mL) was stirred at 75 °C overnight. The mixture was poured into water (10 mL) and extracted with EtOAc (7 mL × 3). The combined organic layers were washed with water, brine, dried over Na₂SO₄ and concentrated under <i>vacuum</i> to dryness. The residue was purified by column chromatography on silica gel (petroleum ether/ethyl acetate 30:1) to afford <i>N-tert</i>-butyl-4-nitroaniline (1 g, 73%). ¹H NMR (CDCl₃, 400 MHz) δ 8.03-8.00 (m, 2H), 6.61-6.57 (m, 2H), 4.67 (brs, 1H), 1.42 (s, 9H). <chemistry id="chem0314" num="0314"><img file="EP2007756B1_D0340.tif" /></chemistry>

(2-Bromo-4-nitro-phenyl)-<i>tert</i>-butyl-amine

0351To a solution of <i>N-tert</i>-butyl-4-nitroaniline (1 g, 5.1 mmol) in AcOH (5 mL) was added Br₂ (0.86 g, 54 mmol) dropwise at 15 °C. After addition, the mixture was stirred at 30 °C for 30 min and then filtered. The filter cake was basified to pH 8-9 with aqueous NaHCO₃. The aqueous layer was extracted with EtOAc (10 mL × 3). The combined organic layers were washed with water, brine, dried over Na₂SO₄ and concentrated under <i>vacuum</i> to give (2-bromo-4-nitro-phenyl)-tert-butyl-amine (0.6 g, 43%). ¹H-NMR (CDCl₃, 400 MHz) δ 8.37 (dd, <i>J</i> = 2.4 Hz, 1H), 8.07 (dd, J= 2.4, 9.2 Hz, 1H), 6.86 (d, <i>J</i> = 9.2 Hz, 1H), 5.19 (brs, 1H), 1.48 (s, 9H). <chemistry id="chem0315" num="0315"><img file="EP2007756B1_D0341.tif" /></chemistry>

<i>tert</i>-Butyl-(4-nitro-2-trimethylsilanylethynyl-phenyl)-amine

0352To a solution of (2-bromo-4-nitro-phenyl)-tert-butyl-amine (0.6 g, 2.2 mmol) in Et₃N (10 mL) was added Pd(PPh₃)₂Cl₂ (70 mg, 0.1 mmol), CuI (20.9 mg, 0.1 mmol) and ethynyl-trimethyl-silane (0.32 g, 3.3 mmol) successively under N₂ protection. The reaction mixture was heated at 70 °C overnight. The solvent was removed under <i>vacuum</i> and the residue was washed with EtOAc (10 mL × 3). The combined organic layers were washed with water, brine, dried over Na₂SO₄ and concentrated under <i>vacuum</i> to dryness. The residue was purified by column chromatography on silica gel (petroleum ether/ethyl acetate 20:1) to afford tert-butyl-(4-nitro-2-trimethylsilanylethynyl-phenyl)-amine (100 mg, 16%). ¹H-NMR (CDCl₃, 400 MHz) δ 8.20 (d, <i>J</i> = 2.4, Hz, 1H), 8.04 (dd, <i>J</i> = 2.4, 9.2 Hz, 1H), 6.79 (d, <i>J</i> = 9.6 Hz, 1H), 5.62 (brs, 1H), 1.41 (s, 9H), 0.28 (s, 9H). <chemistry id="chem0316" num="0316"><img file="EP2007756B1_D0342.tif" /></chemistry>

1-<i>tert</i>-Butyl-5-nitro-1H-indole

0353To a solution of tert-butyl-(4-nitro-2-trimethylsilanylethynyl-phenyl)-amine (10 mg, 0.035 mmol) in DMF (2 mL), was added CuI (13 mg, 0.07 mmol) under N₂ protection. The reaction mixture was stirred at 100 °C overnight. At this time, EtOAc (4 mL) was added to the mixture. The mixture was filtered and the filtrate was washed with water, brine, dried over Na₂SO₄ and concentrated under <i>vacuum</i> to obtain 1-tert-butyl-5-nitro-1H-indole (7 mg, 93%). ¹H-NMR (CDCl₃, 300 MHz) δ 8.57 (d, <i>J</i> = 2.1 Hz, 1H), 8.06 (dd, <i>J</i> = 2.4, 9.3 Hz, 1H), 7.65 (d, <i>J</i> = 9.3 Hz, 1H), 7.43 (d, <i>J</i> = 3.3 Hz, 1H), 6.63 (d, <i>J</i> = 3.3 Hz, 1H), 1.76 (s, 9H). <chemistry id="chem0317" num="0317"><img file="EP2007756B1_D0343.tif" /></chemistry>

1-<i>tert</i>-Butyl-1H-indol-5-amine

0354To a solution of 1-<i>tert</i>-butyl-5-nitro-1H-indole (6.5 g, 0.030 mol) in MeOH (100 mL) was added Raney Nickel (0.65 g, 10%) under N₂ protection. The mixture was stirred under hydrogen atmosphere (1 atm) at 30 °C for 1 h. The catalyst was filtered off and the filtrate was concentrated under <i>vacuum</i> to dryness. The residue was purified by column chromatography on silica gel (PE/EtOAc 1:2) to give 1-<i>tert</i>-butyl-1H-indol-5-amine (2.5 g, 45%). ¹H-NMR (CDCl₃, 400 MHz) δ 7.44 (d, <i>J</i> = 8.8 Hz, 1H), 7.19 (dd, <i>J</i> = 3.2 Hz, 1H), 6.96 (d, <i>J</i> = 2.0 Hz, 1H), 6.66 (d, <i>J</i> = 2.0, 8.8 Hz, 1H), 6.26 (d, <i>J</i> = 3.2 Hz, 1H), 1.67 (s, 9H). MS (ESI) m/e (M+H⁺) 189.2.

Example 32: 2<i>-tert</i>-Butyl-1-methyl-1H-indol-5-amine

0355<chemistry id="chem0318" num="0318"><img file="EP2007756B1_D0344.tif" /></chemistry><chemistry id="chem0319" num="0319"><img file="EP2007756B1_D0345.tif" /></chemistry>

(2-Bromo-4-nitro-phenyl)-methyl-amine

0356To a solution of methyl-(4-nitro-phenyl)-amine (15.2 g, 0.1 mol) in AcOH (150 mL) and CHCl₃ (50 mL) was added Br₂ (16.0 g, 0.1 mol) dropwise at 5 °C. The mixture was stirred at 10 °C for 1h and then basified with sat. aq. NaHCO₃. The resulting mixture was extracted with EtOAc (100 mL × 3), and the combined organics were dried over anhydrous Na₂SO₄ and evaporated under vacuum to give (2-bromo-4-nitro-phenyl)-methyl-amine (2-bromo-4-nitro-phenyl)-methyl-amine (23.0 g, 99%), which was used in the next step without further purification. ¹H NMR (300 MHz, CDCl₃) δ 8.37 (d, J = 2.4 Hz, 1 H), 8.13 (dd, J = 2.4, 9.0 Hz, 1 H), 6.58 (d, J = 9.0 Hz, 1 H), 5.17 (brs, 1 H), 3.01 (d, J = 5.4 Hz, 3 H). <chemistry id="chem0320" num="0320"><img file="EP2007756B1_D0346.tif" /></chemistry>

[2-(3,3-Dimethyl-but-1-ynyl)-4-nitro-phenyl]-methyl-amine

0357To a solution of (2-bromo-4-nitro-phenyl)-methyl-amine (22.5 g, 97.4 mmol) in toluene (200 mL) and water (100 mL) were added Et₃N (19.7 g, 195 mmol), Pd(PPh₃)₂Cl₂ (6.8 g, 9.7 mmol), CuI (0.7 g, 3.9 mmol) and 3,3-dimethyl-but-1-yne (16.0 g, 195 mmol) successively under N₂ protection. The mixture was heated at 70 °C for 3 hours and then cooled down to room temperature. The resulting mixture was extracted with EtOAc (100 mL × 3). The combined organic extracts were dried over anhydrous Na₂SO₄ and evaporated under vacuum to give [2-(3,3-dimethyl-but-1-ynyl)-4-nitro-phenyl]-methyl-amine (20.1 g, 94%), which was used in the next step without further purification. ¹H NMR (400 MHz, CDCl₃) δ 8.15 (d, J = 2.4 Hz, I H), 8.08 (dd, J = 2.8, 9.2 Hz, 1H), 6.50 (d, J = 9.2 Hz, 1H), 5.30 (brs, 1H), 3.00 (s, 3H), 1.35 (s, 9H). <chemistry id="chem0321" num="0321"><img file="EP2007756B1_D0347.tif" /></chemistry>

2-<i>tert</i>-Butyl-1-methyl-5-nitro-1H-indole

0358A solution of [2-(3,3-dimethyl-but-1-ynyl)-4-nitro-phenyl]-methyl-amine (5.0 g, 22.9 mmol) and TBAF (23.9 g, 91.6 mmol) in THF (50 mL) was heated at reflux overnight. The solvent was removed by evaporation under <i>vacuum</i> and the residue was dissolved in brine (100 mL) and EtOAc (100 mL). The organic phase was separated, dried over Na₂SO₄ and evaporated under vacuum to give 2-tert-butyl-1-methyl-5-nitro-1H-indole (5.0 g, 99%), which was used in the next step without further purification. ¹H NMR (CDCl₃, 400 MHz) δ 8.47 (d, J = 2.4 Hz, 1H), 8.07 (dd, J = 2.4, 9.2 Hz, 1H), 7.26-7.28 (m, 1H), 6.47 (s, 1H), 3.94 (s, 3H), 1.50 (s, 9H). <chemistry id="chem0322" num="0322"><img file="EP2007756B1_D0348.tif" /></chemistry>

2-<i>tert-</i>Butyl-1-methyl-1H-indol-5-amine

0359To a solution of 2-tert-butyl-1-methyl-5-nitro-1H-indole (3.00 g, 13.7 mmol) in MeOH (30 mL) was added Raney Ni (0.3 g) under nitrogen atmosphere. The mixture was stirred under hydrogen atmosphere (1 atm) at room temperature overnight. The mixture was filtered through a Celite pad and the filtrate was evaporated under <i>vacuum.</i> The crude residue was purified by column chromatography on silica gel (P.E/EtOAc 20:1) to give 2-tert-butyl-1-methyl-1H-indol-5-amine (1.7 g, 66%). ¹H NMR (300 MHz, CDCl₃) δ 7.09 (d, J = 8.4 Hz, 1H), 6.89-6.9 (m, 1H), 6.66 (dd, J = 2.4, 8.7 Hz, 1H), 6.14 (d, J = 0.6 Hz, 1H), 3.83 (s, 3H), 3.40 (brs, 2H), 1.45 (s, 9H); MS (ESI) m/e (M+H⁺) 203.1.

Example 33: 2-Cyclopropyl-1H-indol-5-amine

0360<chemistry id="chem0323" num="0323"><img file="EP2007756B1_D0349.tif" /></chemistry><chemistry id="chem0324" num="0324"><img file="EP2007756B1_D0350.tif" /></chemistry>

2-Bromo-4-nitroaniline

0361To a solution of 4-nitro-aniline (25 g, 0.18 mol) in HOAc (150 mL) was added liquid Br₂ (30 g, 0.19 mol) dropwise at room temperature. The mixture was stirred for 2 hours. The solid was collected by filtration and poured into water (100 mL), which was basified with sat. aq. NaHCO₃ to pH 7 and extracted with EtOAc (300 mL × 3). The combined organic layers were dried over anhydrous Na₂SO₄ and evaporated under reduced pressure to give 2-bromo-4-nitroaniline (30 g, 80%), which was directly used in the next step. <chemistry id="chem0325" num="0325"><img file="EP2007756B1_D0351.tif" /></chemistry>

2-(Cyclopropylethynyl)-4-nitroaniline

0362To a deoxygenated solution of 2-bromo-4-nitroaniline (2.17 g, 0.01 mmol), ethynyl-cyclopropane (1 g, 15 mmol) and CuI (10 mg, 0.05 mmol) in triethylamine (20 mL) was added Pd(PPh₃)₂Cl₂ (210 mg, 0.3 mmol) under N₂. The mixture was heated at 70 °C and stirred for 24 hours. The solid was filtered off and washed with EtOAc (50 mL x 3). The filtrate was evaporated under reduced pressure, and the residue was purified by column chromatography on silica gel (petroleum ether/ethyl acetate = 10/1) to give 2-(cyclopropylethynyl)-4-nitroaniline (470 mg, 23%). ¹H NMR (300 MHz, CDCl₃) δ 8.14 (d, <i>J</i> = 2.7 Hz, 1H), 7.97 (dd, <i>J</i> = 2.7, 9.0 Hz, 1H), 6.63 (d, <i>J</i> = 9.0 Hz, 1H), 4.81 (brs, 2H), 1.55-1.46 (m, 1H), 0.98-0.90 (m, 2H), 0.89-0.84 (m, 2H). <chemistry id="chem0326" num="0326"><img file="EP2007756B1_D0352.tif" /></chemistry>

<i>N</i>-(2-(Cyclopropylethynyl)phenyl)-4-nitrobutyramide

0363To a solution of 2-(cyclopropylethynyl)-4-nitroaniline (3.2 g, 15.8 mmol) and pyridine (2.47 g, 31.7 mmol) in CH₂Cl₂ (60 mL) was added butyryl chloride (2.54 g, 23.8 mmol) at 0 °C. The mixture was warmed to room temperature and stirred for 3 hours. The resulting mixture was poured into ice-water. The organic layer was separated. The aqueous phase was extracted with CH₂Cl₂ (30 m L × 3). The combined organic layers were dried over anhydrous Na₂SO₄ and evaporated under reduced pressure to give the crude product, which was purified by column chromatography on silica gel (petroleum ether/ethyl acetate = 10/1) to give N-(2-(cyclopropylethynyl)phenyl)-4-nitrobutyramide (3.3 g, 76%). ¹H NMR (400 MHz, CDCl₃) δ 8.61 (d, <i>J</i> = 9.2 Hz, 1H), 8.22 (d, <i>J</i> = 2.8 Hz, 1H), 8.18 (brs, 1H), 8.13 (dd, <i>J</i> = 2.4, 92 Hz, 1H), 2.46 (t, <i>J</i> = 7.2 Hz, 2H), 1.83-1.76 (m, 2H), 1.59-1.53 (m, 1H), 1.06 (t, <i>J</i> = 7.2 Hz, 3H), 1.03-1.01 (m, 2H), 0.91-0.87 (m, 2H). <chemistry id="chem0327" num="0327"><img file="EP2007756B1_D0353.tif" /></chemistry>

2-Cyclopropyl-5-nitro-1H-indole

0364A mixture of N-(2-(cyclopropylethynyl)phenyl)-4-nitrobutyramide (3.3 g, 0.01 mol) and TBAF (9.5 g, 0.04 mol) in THF (100 mL) was heated at reflux for 24 hours. The mixture was cooled to the room temperature and poured into ice water. The mixture was extracted with CH₂Cl₂ (50 m L × 3). The combined organic layers were dried over anhydrous Na₂SO₄ and evaporated under reduced pressure. The residue was purified by column chromatography on silica gel (petroleum ether/ethyl acetate = 10/1) to give 2-cyclopropyl-5-nitro-1H-indole (1.3 g, 64%). ¹H NMR (400 MHz, CDCl₃) δ 8.44 (d, <i>J</i> = 2.0 Hz, 1H), 8.40 (brs, 1H), 8.03 (dd, <i>J</i> = 2.0, 8.8 Hz, 1H), 7.30 (d, <i>J</i> = 8.8 Hz, 1H), 6.29 (d, <i>J</i> = 0.8 Hz, 1H), 2.02-1.96 (m, 1H) 1.07-1.02 (m, 2H), 0.85-0.81 (m, 2H). <chemistry id="chem0328" num="0328"><img file="EP2007756B1_D0354.tif" /></chemistry>

2-Cyclopropyl-1H-indol-5-amine

0365To a solution of 2-cyclopropyl-5-nitro-1H-indole (1.3 g, 6.4 mmol) in MeOH (30 mL) was added Raney Nickel (0.3 g) under nitrogen atmosphere. The mixture was stirred under hydrogen atmosphere (1 atm) at room temperature overnight. The catalyst was filtered through a Celite pad and the filtrate was evaporated under <i>vacuum</i> to give the crude product, which was purified by column chromatography on silica gel (petroleum ether/ethyl acetate = 5/1) to give 2-cyclopropyl-1H-indol-5-amine (510 mg, 56%). ¹H NMR (400 MHz, CDCl₃) δ 6.89 (d, <i>J</i> = 8.4 Hz, 1H), 6.50 (d, <i>J</i> = 1.6 Hz, 1H), 6.33 (dd, <i>J</i> = 2.0, 8.4 Hz, 1H), 5.76 (s, 1H), 4.33 (brs, 2H), 1.91-1.87 (m, 1H), 0.90-0.85(m, 2H), 0.70-0.66 (m, 2H); MS (ESI) m/e (M+H⁺) 173.2.

Example 34: 3

<b>-tert</b>-

Butyl-1H-indol-5-amine

0366<chemistry id="chem0329" num="0329"><img file="EP2007756B1_D0355.tif" /></chemistry><chemistry id="chem0330" num="0330"><img file="EP2007756B1_D0356.tif" /></chemistry>

3-<i>tert</i>-Butyl-5-nitro-1H-indole

0367To a mixture of 5-nitro-1H-indole (6 g, 36.8 mmol) and AlCl₃ (24 g, 0.18 mol) in CH₂Cl₂ (100 mL) was added 2-bromo-2-methyl-propane (8.1 g, 36.8 mmol) dropwise at 0 °C. After being stirred at 15 °C overnight, the reaction mixture was poured into ice (100 mL). The precipitated salts were removed by filtration and the aqueous layer was extracted with CH₂Cl₂ (30 mL × 3). The combined organic layers were washed with water, brine, dried over Na₂SO₄ and concentrated under <i>vacuum</i> to obtain the crude product, which was purified by column chromatography on silica gel (petroleum ether/ethyl acetate 20:1) to give 3-tert-butyl-5-nitro-1H-indole (2.5 g, 31%). ¹H NMR (CDCl₃, 400 MHz) δ 8.49 (d, <i>J</i> = 1.6 Hz, 1H), 8.31 (brs, 1H), 8.05 (dd, <i>J</i> = 2.0, 8.8 Hz, 1H), 7.33 (d, <i>J</i> = 8.8 Hz, 1H), 6.42 (d, <i>J</i> =1.6 Hz, 1H), 1.42 (s, 9H). <chemistry id="chem0331" num="0331"><img file="EP2007756B1_D0357.tif" /></chemistry>

3

<b>-tert</b>-

Butyl-1H-indol-5-amine

0368To a solution of 3-tert-butyl-5-nitro-1H-indole (2.5 g, 11.6 mmol) in MeOH (30 mL) was added Raney Nickel (0.2 g) under N₂ protection. The mixture was stirred under hydrogen atmosphere (1 atm) at 15 °C for 1 hr. The catalyst was filtered off and the filtrate was concentrated under <i>vacuum</i> to dryness. The residue was purified by preparative HLPC to afford 3-tert-butyl-1H-indol-5-amine (0.43 g, 19%). ¹H NMR (CDCl₃, 400 MHz) δ 7.72 (brs, 1H), 7.11 (d, <i>J</i> = 8.4 Hz, 1H), 6.86 (d, <i>J</i> = 2.0 Hz, 1H), 6.59 (dd, <i>J</i> = 2.0, 8.4 Hz, 1H), 6.09 (d, <i>J</i> = 1.6 Hz, 1H), 1.37 (s, 9H); MS (ESI) m/e (M+H⁺) 189.1.

Example 35

:

2-Phenyl-1H-indol-5-amine

0369<chemistry id="chem0332" num="0332"><img file="EP2007756B1_D0358.tif" /></chemistry><chemistry id="chem0333" num="0333"><img file="EP2007756B1_D0359.tif" /></chemistry>

2-Bromo-4-nitroaniline

0370To a solution of 4-nitroaniline (50 g, 0.36 mol) in AcOH (500 mL) was added liquid Br₂ (60 g, 0.38 mol) dropwise at 5 °C. The mixture was stirred for 30 min at that temperature. The insoluble solid was collected by filtration and poured into EtOAc (200 mL). The mixture was basified with saturated aqueous NaHCO₃ to pH 7. The organic layer was separated. The aqueous phase was extracted with EtOAc (300 mL x 3). The combined organic layers were dried and evaporated under reduced pressure to give 2-bromo-4-nitroaniline (56 g, 72%), which was directly used in the next step. <chemistry id="chem0334" num="0334"><img file="EP2007756B1_D0360.tif" /></chemistry>

4-Nitro-2-(phenylethynyl)aniline

0371To a deoxygenated solution of 2-bromo-4-nitroaniline (2.17g, 0.01 mmol), ethynyl-benzene (1.53 g, 0.015 mol) and Cu<b>I</b> (10 mg, 0.05 mmol) in triethylamine (20 mL) was added Pd(PPh₃)₂Cl₂ (210 mg, 0.2 mmol) under N₂. The mixture was heated at 70°C and stirred for 24 hours. The solid was filtered off and washed with EtOAc (50 mL × 3). The filtrate was evaporated under reduced pressure and the residue was purified by column chromatography on silica gel (petroleum ether/ethyl acetate = 10/1) to give 4-nitro-2-(phenylethynyl)aniline (340 mg, 14%). ¹H NMR (300 MHz, CDCl₃) δ 8.37-8.29 (m, 1H), 8.08-8.00 (m, 1H), 7.56-7.51 (m, 2H), 7.41-7.37 (m, 3H), 6.72 (m, 1H), 4.95 (brs, 2H). <chemistry id="chem0335" num="0335"><img file="EP2007756B1_D0361.tif" /></chemistry>

<i>N</i>

-

(2-(Phenylethynyl)phenyl)-4-nitrobutyramide

0372To a solution of 4-nitro-2-(phenylethynyl)aniline (17 g, 0.07mmol) and pyridine (11.1 g, 0.14 mol) in CH₂Cl₂ (100 mL) was added butyryl chloride (11.5 g, 0.1 mol) at 0 °C. The mixture was warmed to room temperature and stirred for 3 hours. The resulting mixture was poured into ice-water. The organic layer was separated. The aqueous phase was extracted with CH₂Cl₂ (30 m L x 3). The combined organic layers were dried over anhydrous Na₂SO₄ and evaporated under reduced pressure. The residue was purified by column chromatography on silica gel (petroleum ether/ethyl acetate = 10/1) to give N-(2-(phenylethynyl)phenyl)-4-nitrobutyramide (12 g, 55%). ¹H NMR (400 MHz, CDCl₃) δ 8.69 (d, <i>J</i>=9.2 Hz, 1H), 8.39 (d, <i>J</i>=2.8 Hz, 1H), 8.25-8.20 (m, 2H), 7.58-7.55 (m, 2H), 7.45-7.42 (m, 3H), 2.49 (t, <i>J</i>=7.2Hz, 2H), 1.85-1.79 (m, 2H), 1.06 (t, <i>J=</i>7.2 Hz, 3H). <chemistry id="chem0336" num="0336"><img file="EP2007756B1_D0362.tif" /></chemistry>

5-Nitro-2-phenyl-1H-indole

0373A mixture of N-(2-(phenylethynyl)phenyl)-4-nitrobutyramide (5.0 g, 0.020 mol) and TBAF (12.7 g, 0.050 mol) in THF (30 mL) was heated at reflux for 24 h. The mixture was cooled to room temperature and poured into ice water. The mixture was extracted with CH₂Cl₂ (50 m L × 3). The combined organic layers were dried over anhydrous Na₂SO₄ and evaporated under reduced pressure. The residue was purified by column chromatography on silica gel (petroleum ether/ethyl acetate^-= 10/l} to give 5-nitro-2-phenyl-1H-indole (3.3 g, 69%). ¹H NMR (400 MHz, CDCl₃) δ 8.67 (s, 1H), 8.06 (dd, <i>J</i>= 2.0, 8.8 Hz, 1H), 7.75 (d, <i>J</i> =7.6 Hz, 2H), 7.54 (d, <i>J=8.8</i>Hz, 1 H), 7.45 (t, <i>J</i>=7.6Hz, 2H), 7.36 (t, <i>J=</i>7.6 Hz, 1H). 6.95 (s, 1 H). <chemistry id="chem0337" num="0337"><img file="EP2007756B1_D0363.tif" /></chemistry>

2-Phenyl-1H-indol-5-amine

0374To a solution of 5-nitro-2-phenyl-1H-indole (2.83 g, 0.01 mol) in MeOH (30 mL) was added Raney Ni (510 mg) under nitrogen atmosphere. The mixture was stirred under hydrogen atmosphere (1 atm) at room temperature overnight. The catalyst was filtered through a Celite pad and the filtrate was evaporated under <i>vacuum</i> to give the crude product, which was purified by column chromatography on silica gel (petroleum ether/ethyl acetate = 5/l) to give 2-phenyl-1H-indol-5-amine (1.6 g, 77%). ¹H NMR (400 MHz, CDCl₃) δ 7.76 (d, <i>J</i>=7.6Hz, 2H), 7.39 (t, <i>J</i> = 7.6 Hz, 2H), 7.24 (t, <i>J</i> = 7.6 Hz, 1 H), 7.07 (d, <i>J</i> = 8.4 Hz, 1 H), 6.64 (d, <i>J</i>=1.6 Hz, 1 H), 6.60 (d, <i>J</i>=1.2 Hz, 1H), 6.48 (dd, <i>J</i>=2.0, 8.4 Hz, 1 H), 4.48 (brs, 2H); MS (ESI) m/e (M+H⁺) 209.0.

Example 36: 2

<b>-tert</b>-

Butyl-4-fluoro-1H-indol-5-amine

0375<chemistry id="chem0338" num="0338"><img file="EP2007756B1_D0364.tif" /></chemistry><chemistry id="chem0339" num="0339"><img file="EP2007756B1_D0365.tif" /></chemistry>

2-Bromo-3-fluoroaniline

0376To a solution of 2-bromo-l -fluoro-3-nitrobenzene (1.0 g, 5.0 mmol) in CH₃OH (50 mL) was added NiCl₂ (2.2 g 10 mmol) and NaBH₄ (0.50 g 14 mmol) at 0°C. After the addition, the mixture was stirred for 5 min. Water (20 mL) was added and the mixture was extracted with EtOAc (20 mL × 3). The organic layers were dried over anhydrous Na₂SO₄ and evaporated under vacuum to give 2-bromo-3-fluoroaniline (600 mg, 70%). ¹H NMR (400MHz, CDCl₃) δ 7.07-7.02 (m, 1 H), 6.55-6.49(m, 1 H), 4.22 (br s, 2 H). <chemistry id="chem0340" num="0340"><img file="EP2007756B1_D0366.tif" /></chemistry>

<i>N</i>-(2-Bromo-3-fluorophenyl)butyramide

0377To a solution of 2-bromo-3-fluoroaniline (2.0 g, 11 mmol) in CH₂Cl₂ (50 mL) was added butyryl chloride (1.3 g, 13 mmol) and pyridine (1.7 g, 21 mmol) at 0°C. The mixture was stirred at room temperature for 24 h. Water (20 mL) was added and the mixture was extracted with CH₂Cl₂ (50 mL × 3). The organic layers were dried anhydrous over Na₂SO₄ and evaporated under vacuum to give <i>N-</i>(2-bromo-3-fluorophenyl)butyramide (2.0 g, 73%), which was directly used in the next step. <chemistry id="chem0341" num="0341"><img file="EP2007756B1_D0367.tif" /></chemistry>

<i>N</i>-(2-(3,3-Dimethylbut-1-ynyl)-3-fluorophenyl)butyramide

0378To a solution of <i>N</i>-(2-bromo-3-fluorophenyl)butyramide (2.0 g, 7.0 mmol) in Et₃N (100 mL) was added 4,4-dimethylpent-2-yne(6.0 g, 60 mmol), CuI (70 mg, 3.8 mmol), and Pd(PPh₃)₂Cl₂ (500 mg) successively at room temperature under N₂. The mixture was heated at 80°C overnight. The cooled mixture was filtered and the filtrate was extracted with EtOAc (40 mL × 3). The organic layers were washed with sat. NaCl, dried over anhydrous Na₂SO₄, and evaporated under vacuum. The crude compound was purified by column chromatography on silica gel (10% EtOAc in petroleum ether) to give N-(2-(3,3-dimethylbut-1-ynyl)-3-fluorophenyl)butyramide (1.1 g, 55%). ¹H NMR (400 MHz, CDCl₃) δ 8.20 (d, J<i>=</i> 7.6, 1 H), 7.95 (s, 1 H), 7.21 (m, 1 H), 6.77 (t, <i>J</i>= 7.6 Hz, 1 H), 2.39 (t, <i>J</i>= 7.6 Hz, 2 H), 1.82-1.75 (m, 2 H), 1.40 (s, 9 H), 1.12 (t, <i>J</i>= 7.2 Hz, 3 H). <chemistry id="chem0342" num="0342"><img file="EP2007756B1_D0368.tif" /></chemistry>

2<i>-tert</i>-Butyl-4-fluoro-1H-indole

0379To a solution of N-(2-(3,3-dimethylbut-i-ynyl)-3-fluorophenyl)butyrarnide (6.0 g, 20 mmol) in DMF (100 mL) was added <i>t</i>-BuOK(5.0 g, 50 mmol) at room temperature. The mixture was heated at 90°C overnight before it was poured into water and extracted with EtOAc (100 mL × 3). The organic layers were washed with sat. NaCl and water, dried over anhydrous Na₂SO₄, and evaporated under vacuum to give 2-<i>tert</i>-butyl-4-fluoro-1H-indole (5.8 g, 97%). ¹H NMR (400 MHz, CDCl₃) δ 8.17(br s, 1 H), 7.11 (d, <i>J</i>=7.2 Hz, 1 H), 7.05-6.99 (m, 1 H), 6.76-6.71 (m, 1 H), 6.34 (m, 1 H), 1.41 (s, 9 H). <chemistry id="chem0343" num="0343"><img file="EP2007756B1_D0369.tif" /></chemistry>

2<i>-tert</i>-Butyl-4-fluoro-5-nitro-1H-indole

0380To a solution of 2-<i>tert</i>-butyl-4-fluoro-1H-indole (2.5 g, 10 mmol) in H₂SO₄ (30 mL) was added KNO₃ (1.3 g, 10 mmol) at 0°C. The mixture was stirred for 0.5 h at -10°C. The mixture was poured into water and extracted with EtOAc (100 mL × 3). The organic layers were washed with sat. NaCl and water, dried over anhydrous Na₂SO₄, and evaporated under vacuum. The crude compound was purified by column chromatography on silica gel (10% EtOAc in petroleum ether) to give 2-<i>tert</i>-butyl-4-fluoro-5-nitro-1H-indole (900 mg, 73%). ¹H NMR (400 MHz, CDCl₃) δ 8.50 (br s, 1 H), 7.86 (dd, <i>J</i> = 7.6, 8.8 Hz, 1 H), 7.13 (d, <i>J</i>=8.8 Hz, 1 H), 6.52 (dd, <i>J</i>= 0.4, 2.0 Hz, 1 H), 1.40 (s, 9 H). <chemistry id="chem0344" num="0344"><img file="EP2007756B1_D0370.tif" /></chemistry>

2<i>-tert-</i>Butyl<i>-4-</i>fluoro-1H-indol-5-amine

0381To a solution of 2-<i>t</i>ert-butyl-4-fluoro-5-nitro-1H-indole (2.1 g, 9.0 mmol) in methanol (50 mL) was added NiCl₂ (4.2 g, 18 mmol) and NaBH₄ (1.0 g, 27 mmol) at 0°C. After the addition, the mixture was stirred for 5 min. Water (20 mL) was added and the mixture was extracted with EtOAc (30 mL × 3). The organic layers were washed with sat. NaCl and water, dried over anhydrous Na₂SO₄, evaporated under vacuum to give 2-tert-butyl-4-fluoro-1H-indol-5-amine (900 mg, 50%). ¹H NMR (300 MHz, CDCl₃) δ 7.80 (brs, 1 H), 6.91 (d, <i>J</i>= 8.4 Hz, 1 H), 6.64 (dd, <i>J</i>= 0.9, 2.4 Hz, 1 H), 6.23 (s, 1 H), 1.38 (s, 9 H).

Example 37: 2,3,4,9-Tetrahydro-1H-carbazol-6-amine

0382<chemistry id="chem0345" num="0345"><img file="EP2007756B1_D0371.tif" /></chemistry>

2,3,4,9-Tetrahydro-1<i>H</i>-carbazol-6-amine

03836-Nitro-2,3,4,9-tetrahydro-1<i>H</i>-carbazole (0.100 g, 0.462 mmol) was dissolved in a 40 mL scintillation vial containing a magnetic stir bar and 2 mL of ethanol. Tin(II) chloride dihydrate (1.04 g, 4.62 mmol) was added to the reaction mixture and the resulting suspension was heated at 70°C for 16 h. The crude reaction mixture was then diluted with 15 mL of a saturated aqueous solution of sodium bicarbonate and extracted three times with an equivalent volume of ethyl acetate. The ethyl acetate extracts were combined, dried over sodium sulfate, and evaporated to dryness to yield 2,3,4,9-tetrahydro-1H-carbazol-6-amine (82 mg, 95%) which was used without further purification.

Example 38: 2-<i>tert</i>-Butyl-7-fluoro-1H-indol-5-amine

0384<chemistry id="chem0346" num="0346"><img file="EP2007756B1_D0372.tif" /></chemistry><chemistry id="chem0347" num="0347"><img file="EP2007756B1_D0373.tif" /></chemistry>

2-Bromo-6-fluoro-4-nitro-phenylamine

0385To a solution of 2-fluoro-4-nitro-phenylamine (12 g, 77 mmol) in AcOH (50 mL) was added Br₂ (3.9 mL, 77 mmol) dropwise at 0 °C. The mixture was stirred at 20 °C for 3 h. The reaction mixture was basified with sat. aq. NaHCO₃ and extracted with EtOAc (100 mL × 3). The combined organics were dried over anhydrous Na₂SO₄ and evaporated under vacuum to give 2-bromo-6-fluoro-4-nitro-phenylamine (18 g, 97%). ¹H NMR (400 MHz, CDCl₃) δ 8.22 (m, 1 H), 7.90 (dd, <i>J</i>= 2.4, 10.8 Hz, 1 H), 4.88 (brs, 2 H). <chemistry id="chem0348" num="0348"><img file="EP2007756B1_D0374.tif" /></chemistry>

2-(3,3-Dimethyl-but-1-ynyl)-6-fluoro-4-nitro-phenylamine

0386To a solution of 2-bromo-6-fluoro-4-nitro-phenylamine (11 g, 47 mmol) in dry Et₃N (100 mL) was added CuI (445 mg, 5% mol), Pd(PPh₃)₂Cl₂ (550 mg, 5% mol) and 3,3-dimethyl-but-1-yne (9.6 g, 120 mmol) under N₂ protection. The mixture was stirred at 80 °C for 10 h. The reaction mixture was filtered, poured into ice (100 g), and extracted with EtOAc (50 mL × 3). The combined organic extracts were dried over anhydrous Na₂SO₄ and evaporated under vacuum to give the crude product, which was purified by column chromatography on silica gel (petroleum ether/ethyl acetate 50:1) to give 2-(3,3-dimethyl-but-1-ynyl)-6-fluoro-4-nitro-phenylamine (4.0 g, 36%). ¹HNMR(400 MHz, CDCl₃) δ 8.02 (d, <i>J</i>=1.2 Hz, 1 H), 7.84 (dd, <i>J</i>= 2.4, 10.8 Hz, 1 H), 4.85 (brs, 2 H), 1.36 (s, 9 H). <chemistry id="chem0349" num="0349"><img file="EP2007756B1_D0375.tif" /></chemistry>

N

-[2-(3,3-Dimethyl-but-1-ynyl)-6-fluoro-4-nitro-phenyl]-butyramide

0387To a solution of 2-(3,3-dimethyl-but-1-ynyl)-6-fluoro-4-nitro-phenylamine (4.0 g, 17 mmol) and pyridine (2.7 g, 34 mmol) in anhydrous CH₂Cl₂ (30 mL) was added and butyryl chloride (1.8 g, 17 mmol) dropwise at 0 °C. After stirring for 5 h at 0 °C, the reaction mixture was poured into ice (50 g) and extracted with CH₂Cl₂ (30 mL × 3). The combined organic extracts were dried over anhydrous Na₂SO₄ and evaporated under <i>vacuum</i> to give <i>N-</i>[2-(3,3-dimethyl- but-1-ynyl)-6-fluoro-4-nitro-phenyl]-butyramide (3.2 g, 62%), which was used in the next step without further purification. ¹HNMR (300 MHz, DMSO) δ 8.10 (dd, <i>J</i> = 1.5, 2.7 Hz, 1 H), 7.95 (dd, <i>J=</i> 2.4, 9.6 Hz, 1 H), 7.22 (brs, 1 H), 2.45 (t, <i>J=</i> 7.5 Hz, 2 H), 1.82 (m, 2 H), 1.36 (s, 9 H), 1.06 (t, <i>J =</i> 7.5 Hz, 3 H). <chemistry id="chem0350" num="0350"><img file="EP2007756B1_D0376.tif" /></chemistry>

2-<i>tert</i>-Butyl-7-fluoro-5-nitro-1H-indole

0388To a solution of <i>N</i>-[2-(3,3-dimethyl-but-1-ynyl)- 6-fluoro-4-nitro-phenyl]-butyramide (3.2 g, 10 mmol) in DMF (20 mL) was added t-BuOK (2.3 g, 21 mmol) at room temperature. The mixture was heated at 120 °C for 2 g before being cooled down to room temperature. Water (50 mL) was added to the reaction mixture and the resulting mixture was extracted with CH₂Cl₂ (30 mL × 3). The combined organic extracts were dried over anhydrous Na₂SO₄ and evaporated under <i>vacuum</i> to give 2-tert-butyl-7-fluoro- 5-nitro-1H-indole (2.0 g, 81%), which was used in the next step without further purification. ¹H NMR (300 MHz, CDCl₃) δ 9.95 (brs, 1 H), 8.30 (d, <i>J</i>=2.1 Hz, 1 H), 7.74 (dd, <i>J</i>=1.8, 11.1 Hz, 1 H), 6.43 (dd, <i>J =</i> 2.4, 3.3 Hz, 1 H), 1.43 (s, 9 H). <chemistry id="chem0351" num="0351"><img file="EP2007756B1_D0377.tif" /></chemistry>

2<i>-tert</i>-Butyl-7-fluoro-1H-indol-5-amine

0389To a solution of 2-tert-butyl-7-fluoro- 5-nitro-1H-indole (2.0 g, 8.5 mmol) in MeOH (20 mL) was added Ni (0.3 g) under nitrogen atmosphere. The reaction mixture was stirred under hydrogen atmosphere (1 atm) at room temperature overnight. The catalyst was filtered off through the celite pad and the filtrate was evaporated under <i>vacuum.</i> The crude product was purified by column chromatography on silica gel (petroleum ether/ethyl acetate 100:1) to give 2-<i>tert</i>-butyl-7-fluoro-1H-indol-5-amine (550 mg, 24%). ¹HNMR (300 MHz, CDCl₃) δ 7.87 (brs, 1 H), 6.64 (d, <i>J=</i> 1.5 Hz, 1 H), 6.37 (dd, <i>J=</i> 1.8, 12.3 Hz, 1 H), 6.11 (dd, <i>J=</i> 2.4, 3.6 Hz, 1 H), 1.39 (s, 9 H). MS (ESI) m/z (M+H⁺) 207.

Example 39: 5-Amino-2-<i>tert</i>-butyl-1H-indole-7-carbonitrile

0390<chemistry id="chem0352" num="0352"><img file="EP2007756B1_D0378.tif" /></chemistry><chemistry id="chem0353" num="0353"><img file="EP2007756B1_D0379.tif" /></chemistry>

2-Amino-3-(3,3-dimethylbut-1-ynyl)

-

5

-nitrobenzonitrile

0391To a stirred solution of 2-amino-3-bromo-5-nitrobenzonitrile (2.4 g, 10 mmol) in dry Et₃N (60 mL) was added CuI (380 mg, 5% mol) and Pd(PPh₃)₂Cl₂ (470 mg, 5% mol) at room temperature. 3,3-dimethyl-but-1-yne (2.1 g, 25 mmol) was added dropwise to the mixture at room temperature. The reaction mixture was stirred at 80 °C for 10 h. The reaction mixture was filtered and the filtrate was poured into ice (60 g), extracted with ethyl acetate. The phases were separated and the organic phase was dried over Na₂SO₄. The solvent was removed under <i>vacuum</i> to obtain the crude product, which was purified by column chromatography (2-10% EtOAc in petroleum ether) to obtain 2-amino-3-(3,3-dimethylbut-1-ynyl)- 5-nitrobenzonitrile (1.7 g, 71%). ¹HNMR (300 MHz, CDCl₃) δ 8.28 (d, <i>J=</i> 2.7 Hz, 1 H), 8.27 (d, <i>J=</i> 2.7 Hz, 1 H), 5.56 (br s, 2 H), 1.37 (s, 9 H). <chemistry id="chem0354" num="0354"><img file="EP2007756B1_D0380.tif" /></chemistry>

2<i>-tert</i> Butyl-5-nitro-1H-indole-7-carbonitrile

0392To a solution of 2-amino-3-(3,3-dimethylbut-1-ynyl)- 5-nitrobenzonitrile (1.7 g, 7.0 mmol) in THF (35 mL) was added TBAF (9.5 g, 28 mmol) at room temperature. The mixture was heated at reflux overnight. The reaction mixture was cooled and the THF was removed under reduced pressure. Water (50ml) was added to the residue and the mixture was extracted with EtOAc. The organics were dried over Na₂SO₄ and the solvent was evaporated under <i>vacuum</i> to obtain 0.87 g of crude product 2-tert-butyl-5-nitro-1H-indole-7-carbonitrile which was used directly in the next step without purification. <chemistry id="chem0355" num="0355"><img file="EP2007756B1_D0381.tif" /></chemistry>

5-Amino-2-<i>tert</i>-butyl-1H-indol-7-carbonitrile

0393To a solution of crude product <i>2-tert-</i>butyl-5-nitro-1H-indole-7-carbonitrile (0.87 g, 3.6mmol) in MeOH (10 mL) was added NiCl₂.6H₂O (1.8 g, 7.2 mmol) at -5 °C. The reaction mixture was stirred for 30 min, then NaBH₄ (0.48g, 14.32 mmol) was added to the reaction mixture at 0°C. After 5 min, the reaction mixture was quenched with water, filtered and extracted with EtOAc. The combined organic layers were dried over Na₂SO₄ and concentrated under <i>vacuum</i> to obtain the crude product, which was purified by column chromatography (5-20% EtOAc in petroleum ether) to obtain 5-amino-2-<i>tert</i>-butyl-1H-indol-7-carbonitrile (470 mg, 32% over two steps). ¹HNMR (400MHz, CDCl₃) δ 8.25 (s, 1 H), 7.06 (d, <i>J</i>=2.4Hz, 1 H), 6.84 (d, <i>J</i>=2.4 Hz, 1 H), 6.14 (d, <i>J</i>=2.4 Hz, 1 H), 3.57 (br s, 2 H), 1.38 (s, 9 H). MS (ESI) m/z: 214 (M+H⁺).

Example 40: Methyl 5-ammo-2-<i>tert</i>-butyl-1H-indole-7-carboxylate

0394<chemistry id="chem0356" num="0356"><img file="EP2007756B1_D0382.tif" /></chemistry><chemistry id="chem0357" num="0357"><img file="EP2007756B1_D0383.tif" /></chemistry>

2<i>-tert-</i>Butyl-5-nitro-1H-indole-7-carboxylic

acid

03952-<i>tert</i>-Butyl-5-nitro-1H-indole-7-carbonitrile (4.6 g, 19 mmol) was added to a solution of KOH in EtOH (10%, 100 mL) and the mixture was heated at reflux overnight. The solution was evaporated to remove alcohol, a small amount of water was added, and then the mixture was acidified with dilute hydrochloric acid. Upon standing in the refrigerator, an orange-yellow solid precipitated, which was purified by chromatography on silica gel (15% EtOAc in petroleum ether) to afford 2-<i>tert</i>-butyl-5-nitro-1H-indole-7-carboxylic acid (4.0 g, 77%). ¹HNMR (CDCl₃, 300MHz) δ 10.79 (brs, 1 H), 8.66 (s, 1 H), 8.45(s, 1 H), 6.57 (s, 1 H), 1.39 (s, 9 H). <chemistry id="chem0358" num="0358"><img file="EP2007756B1_D0384.tif" /></chemistry>

Methyl 2<i>-tert</i> butyl-5-nitro-1H-indole-7-carboxylate

0396SOCl₂ (3.6 g, 30mol) was added dropwise to a solution of 2-<i>tert</i>-butyl-5-nitro-1H-indole-7-carboxylic acid (4.0 g, 15 mol) and methanol (30 mL) at 0°C. The mixture was stirred at 80 °C for 12 h. The solvent was evaporated under vacuum and the residue was purified by column chromatography on silica gel (5% EtOAc in petroleum ether) to afford methyl 2-<i>tert</i>-butyl-5-nitro-1H-indole-7-carboxylate (2.95g, 70%). ¹HNMR (CDCl₃, 300 MHz) δ 9.99 (brs, 1 H), 8.70 (d, <i>J</i>=2.1 Hz, 1 H), 8.65 (d, <i>J</i>=2.1 Hz, 1 H), 6.50 (d, <i>J=</i>2.4 Hz, 1 H), 4.04 (s, 3H), 1.44(s, 9H). <chemistry id="chem0359" num="0359"><img file="EP2007756B1_D0385.tif" /></chemistry>

Methyl 5-amino-2<i>-tert-</i>butyl-1H-indole-7-carboxylate

0397A solution of 2-<i>tert</i>-butyl-5-nitro-1H-indole-7-carboxylate (2.0 g, 7.2 mmol) and Raney Nickel (200 mg) in CH₃OH (50 mL) was stirred for 5h at the room temperature under H₂ atmosphere. The catalyst was filtered off through a celite pad and the filtrate was evaporated under vacuum to give methyl 5-amino-2-tert-butyl-1H-indole-7-carboxylate (1.2 g, 68%) ¹HNMR (CDCl₃, 400 MHz) δ 9.34 (brs, 1H), 7.24 (d, <i>J=</i> 1.6 Hz, 1H), 7.10 (s, 1H), 6.12 (d, <i>J</i>=1.6 Hz, 1H), 3.88 (s, 3H), 1.45 (s, 9H).

Example 41: (5-Amino-2-<i>tert</i>-butyl-1H-indol-7-yl)methanol

0398<chemistry id="chem0360" num="0360"><img file="EP2007756B1_D0386.tif" /></chemistry><chemistry id="chem0361" num="0361"><img file="EP2007756B1_D0387.tif" /></chemistry>

(2<i>-tert</i>-Butyl-5-nitro-1H-indol-7-yl) methanol

0399To a solution of methyl 2-<i>tert</i>-butyl-5-nitro-1H-indole-7-carboxylate (6.15 g, 22.3 mmol) and dichloromethane (30ml) was added DIBAL-H (1.0 M, 20 mL, 20 mmol) at 78°C. The mixture was stirred for 1 h before water (10 mL) was added slowly. The resulting mixture was extracted with EtOAc (120 mL × 3). The combined organic extracts were dried over anhydrous Na₂SO₄ and evaporated under vacuum to give (2-<i>tert</i>-butyl-5-nitro-1H-indol-7-yl)methanol (4.0 g, 73%), which was used in the next step directly. <chemistry id="chem0362" num="0362"><img file="EP2007756B1_D0388.tif" /></chemistry>

(5-Amino-2-<i>tert</i>-butyl-1H-indol-7-yl)methanol

0400A mixture of (2-<i>tert</i>-butyl-5-nitro-1H-indol-7-yl)methanol (4.0 g, 16 mmol) and Raney Nickel (400 mg) in CH₃OH (100 mL) was stirred for 5 g at room temperature under H₂. The catalyst was filtered off through a celite pad and the filtrate was evaporated under vacuum to give (5-amino-2-<i>tert</i>-butyl-1H-indol-7-yl)methanol (3.4g, 80%). ¹H NMR (CDCl₃,400 MHz) δ 8.53 (br s, 1H), 6.80 (d, <i>J</i>=2.0 Hz, 1 H), 6.38 (d, <i>J</i>=1.6Hz, 1 H), 4.89 (s, 2 H), 1.37 (s, 9H).

Example 42: 2-(1-Methylcyclopropyl)-1H-indol-5-amine

0401<chemistry id="chem0363" num="0363"><img file="EP2007756B1_D0389.tif" /></chemistry><chemistry id="chem0364" num="0364"><img file="EP2007756B1_D0390.tif" /></chemistry><chemistry id="chem0365" num="0365"><img file="EP2007756B1_D0391.tif" /></chemistry>

Trimethyl-(1-methyl-cyclopropylethynyl)-silane

0402To a solution of cyclopropylethynyl-trimethyl-silane (3.0 g, 22 mmol) in ether (20 mL) was added dropwise <i>n</i>-BuLi (8.6 mL, 21.7 mol, 2.5 M solution in hexane) at 0 °C. The reaction mixture was stirred at ambient temperature for 24 h before dimethyl sulfate (6.85 g, 54.3 mmol) was added dropwise at -10°C. The resulting solution was stirred at 10 °C and then at 20 °C for 30 min each. The reaction was quenched by adding a mixture of sat. aq. NH₄Cl and 25% aq. ammonia (1:3, 100 mL). The mixture was then stirred at ambient temperature for 1 h. The aqueous phase was extracted with diethyl ether (3 × 50 mL) and the combined organic layers were washed successively with 5% aqueous hydrochloric acid (100 mL), 5% aq. NaHCO₃ solution (100 mL), and water (100 mL). The organics were dried over anhydrous NaSO₄ and concentrated at ambient pressure. After fractional distillation under reduced pressure, trimethyl-(1-methyl-cyclopropylethynyl)-silane (1.7 g, 52%) was obtained as a colorless liquid. ¹HNMR (400 MHz, CDCl₃) δ1.25 (s, 3 H), 0.92-0.86 (m, 2 H), 0.58-0.56(m,2H),0.15(s,9H). <chemistry id="chem0366" num="0366"><img file="EP2007756B1_D0392.tif" /></chemistry>

1-Ethynyl-1-methyl-cyclopropane

0403To a solution of trimethyl-(1-methyl-cyclopropylethynyl)-silane (20 g, 0.13 mol) in THF (250 mL) was added TBAF (69 g, 0.26 mol). The mixture was stirred overnight at 20 °C. The mixture was poured into water and the organic layer was separated. The aqueous phase was extracted with THF (50 mL). The combined organic layers were dried over anhydrous Na₂SO₄ and distilled under atmospheric pressure to obtain 1-ethynyl-1-methyl-cyclopropane (7.0 g, contained 1/2 THF, 34%). ¹HNMR (400 MHz, CDCl₃) δ 1.82 (s, 1 H), 1.26 (s, 3 H), 0.90-0.88 (m, 2 H), 0.57-0.55 (m, 2 H). <chemistry id="chem0367" num="0367"><img file="EP2007756B1_D0393.tif" /></chemistry>

2-Bromo-4-nitroaniline

0404To a solution of 4-nitro-phenylamine (50 g, 0.36 mol) in AcOH (500 mL) was added Br₂ (60 g, 0.38 mol) dropwise at 5 °C. The mixture was stirred for 30 min at that temperature. The insoluble solid was collected by filtration and basified with saturated aqueous NaHCO₃ to pH 7. The aqueous phase was extracted with EtOAc (300 mL × 3). The combined organic layers were dried and evaporated under reduced pressure to obtain compound 2-bromo-4-nitroaniline (56 g, 72%), which was directly used in the next step. <chemistry id="chem0368" num="0368"><img file="EP2007756B1_D0394.tif" /></chemistry>

2-((1-Methylcyclopropyl)ethynyl)-4-nitroaniline

0405To a deoxygenated solution of 2-bromo-4-nitroaniline (430 mg, 2.0 mmol) and 1-ethynyl-1-methyl-cyclopropane (630 mg, 8.0 mmol) in triethylamine (20 mL) was added CuI (76 mg, 0.40 mmol) and Pd(PPh₃)₂Cl₂ (140 mg, 0.20 mmol) under N₂. The mixture was heated at 70°C and stirred for 24 h. The solid was filtered off and washed with EtOAc (50 mL × 3). The filtrate was evaporated under reduced pressure and the residue was purified by column chromatography on silica gel (petroleum ether/ethyl acetate = 10/1) to give 2-((1-methylcyclopropyl)ethynyl)-4-nitroaniline (340 mg, 79%). ¹HNMR (300 MHz, CDCl₃) δ 8.15-8.14 (m, 1 H), 7.98-7.95 (m, 1 H), 6.63 (d, <i>J</i>=6.9 Hz, 1 H), 4.80 (brs, 2 H), 1.38 (s, 3 H), 1.04-1.01 (m, 2 H), 0.76-0.73 (m, 2 H). <chemistry id="chem0369" num="0369"><img file="EP2007756B1_D0395.tif" /></chemistry>

N-[2-(1-Methyl-cyclopropylethynyl)-4-nitro-phenyl]-butyramide

0406To a solution of 2-((1-methylcyclopropyl)ethynyl)-4-nitroaniline (220 mg, 1.0 mmol) and pyridine (160 mg, 2.0 mol) in CH₂Cl₂ (20 mL) was added butyryl chloride (140 mg, 1.3 mmol) at 0 °C. The mixture was warmed to room temperature and stirred for 3 h. The mixture was poured into ice-water. The organic layer was separated and the aqueous phase was extracted with CH₂Cl₂ (30 mL × 3). The combined organic layers were dried over anhydrous Na₂SO₄ and evaporated under reduced pressure to obtain <i>N</i>-[2-(1-methyl-cyclopropyl-ethynyl)-4-nitro-phenyl]-butyramide (230 mg, 82%), which was directly used in the next step. <chemistry id="chem0370" num="0370"><img file="EP2007756B1_D0396.tif" /></chemistry>

2-(1-Methylcyclopropyl)-5-nitro-1H-indole

0407A mixture of <i>N</i>-[2-(1 -methyl-cylopropylethynyl)-4-nitro-phenyl]-butyramide (1.3 g, 4.6 mmol) and TBAF (2.4 g, 9.2 mmol) in THF (20 mL) was heated at reflux for 24 h. The mixture was cooled to room temperature and poured into ice water. The mixture was extracted with CH₂Cl₂ (30 mL × 3). The combined organic layers were dried over anhydrous Na₂SO₄ and evaporated under reduced pressure. The residue was purified by column chromatography on silica gel (petroleum ether/ethyl acetate = 10/1) to afford 2-(1-methylcyclopropyl)-5-nitro-1H-indole (0.70 g, 71%). ¹HNMR (400 MHz, CDCl₃) δ 8.56 (brs, 1H), 8.44 (d, <i>J</i>=2.0 Hz, 1 H), 8.01 (dd, <i>J</i>=2.4, 8.8 Hz, 1 H), 7.30 (d, <i>J=</i> 8.8 Hz, 1 H), 6.34 (d, <i>J=</i> 1.6 Hz, 1 H), 1.52 (s, 3 H), 1.03-0.97 (m, 2 H), 0.89-0.83 (m, 2 H). <chemistry id="chem0371" num="0371"><img file="EP2007756B1_D0397.tif" /></chemistry>

2-(1-Methyl-cyclopropyl)-1H-indol-5-ylamine

0408To a solution of 2-(1-methylcyclopropyl)-5-nitro-1H-indole (0.70 g, 3.2 mmol) in EtOH (20 mL) was added Raney Nickel (100 mg) under nitrogen atmosphere. The mixture was stirred under hydrogen atmosphere (1 atm) at room temperature overnight. The catalyst was filtered off through a celite pad and the filtrate was evaporated under <i>vacuum.</i> The residue was purified by column chromatography on silica gel (petroleum ether/ethyl acetate = 5/1) to afford 2-(1-methyl-cyclopropyl)-1H -indol-5-ylamine (170 mg, 28%). ¹HNMR (400 MHz, CDCl₃) δ 7.65 (brs, 1 H), 7.08 (d, <i>J=</i> 8.4 Hz, 1 H), 6.82 (s, 1 H), 6.57 (d, <i>J=</i> 8.4 Hz, 1 H), 6.14 (s, 1 H), 3.45 (brs, 2 H), 1.47 (s, 3 H), 0.82-0.78 (m, 2 H), 0.68-0.63 (m, 2 H).

Example 43: Methyl 2-(5-amino-1H-indol-2-yl)-2-methylpropanoate

0409<chemistry id="chem0372" num="0372"><img file="EP2007756B1_D0398.tif" /></chemistry><chemistry id="chem0373" num="0373"><img file="EP2007756B1_D0399.tif" /></chemistry><chemistry id="chem0374" num="0374"><img file="EP2007756B1_D0400.tif" /></chemistry>

Methyl 2,2-dimethyl-3-oxobutanoate

0410To a suspension of NaH (42 g, 1.1 mol, 60%) in THF (400 mL) was added dropwise a solution of methyl 3-oxobutanoate (116 g, 1.00 mol) in THF (100mL) at 0 °C. The mixture was stirred for 0.5 h at that temperature before Mel (146 g, 1.1 mol) was added dropwise at 0 °C. The resultant mixture was warmed to room temperature and stirred for 1 h. NaH (42 g, 1.05 mol, 60%) was added in portions at 0 °C and the resulting mixture was continued to stir for 0.5 h at this temperature. Mel (146 g, 1.05 mol) was added dropwise at 0 °C. The reaction mixture was warmed to room temperature and stirred overnight. The mixture was poured into ice water and the organic layer was separated. The aqueous phase was extracted with EtOAc (500 mL × 3). The combined organic layers were dried and evaporated under reduced pressure to give methyl 2,2-dimethyl-3-oxobutanoate (85 g), which was used directly in the next step. <chemistry id="chem0375" num="0375"><img file="EP2007756B1_D0401.tif" /></chemistry>

Methyl 3-chloro-2,2-dimethylbut-3-enoate

0411To a suspention of PCl₅ (270 g, 1.3 mol) in CH₂Cl₂ (1000 mL) was added dropwise methyl 2,2-dimethyl-3-oxobutanoate (85 g) at 0 °C, following by addition of approximately 30 drops of dry DMF. The mixture was heated at reflux overnight. The reaction mixture was cooled to ambient temperature and slowly poured into ice water. The organic layer was separated and the aqueous phase was extracted with CH₂Cl₂ (500 mL × 3). The combined organic layers were washed with saturated aqueous NaHCO₃ and dried over anhydrous Na₂SO₄. The solvent was evaporated and the residue was distilled under reduced pressure to give methyl 3-chloro-2,2-dimethylbut-3-enoate (37 g, 23%). ¹HNMR (400 MHz, CDCl₃) δ 5.33 (s, 1 H), 3.73 (s, 3 H), 1.44 (s, 6 H). <chemistry id="chem0376" num="0376"><img file="EP2007756B1_D0402.tif" /></chemistry>

3-Chloro-2,2-dimethylbut-3-enoic acid

0412A mixture of methyl 3-chloro-2,2-dimethylbut-3-enoate (33 g, 0.2 mol) and NaOH (9.6 g, 0.24 mol) in water (200 mL) was heated at reflux for 5 h. The mixture was cooled to ambient temperature and extracted with ether. The organic layer was discarded. The aqueous layer was acidified with cold 20% HCl solution and extracted ether (200 mL × 3). The combined organic layers were dried and evaporated under reduced pressure to give 3-chloro-2,2-dimethyl-but-3-enoic acid (21 g, 70%), which was used directly in the next step. ¹HNMR (400 MHz, CDCl₃) δ 7.90 (brs, 1 H), 5.37 (dd, <i>J</i>=2.4, 6.8 Hz, 2 H), 1.47 (s, 6 H). <chemistry id="chem0377" num="0377"><img file="EP2007756B1_D0403.tif" /></chemistry>

2,2-Dimethyl-but-3-ynoic acid

0413Liquid NH₃ was condensed in a 3-neck, 250 mL round bottom flask at -78 °C. Na (3.98 g, 0.173 mol) was added to the flask in portions. The mixture was stirred for 2 h at -78 °C before anhydrous DMSO (20 mL) was added dropwise at -78 °C. The mixture was stirred at room temperature until no more NH₃ was given off. A solution of 3-chloro-2,2-dimethyl-but-3-enoic acid (6.5 g, 43 mmol) in DMSO (10 mL) was added dropwise at -40 °C. The mixture was warmed and stirred at 50 °C for 5 h, then stirred at room temperature overnight. The cloudy, olive green solution was poured into cold 20% HCl solution and then extracted three times with ether. The ether extracts were dried over anhydrous Na₂SO₄ and concentrated to give crude 2,2-dimethyl-but-3-ynoic acid (2 g), which was used directly in the next step. ¹HNMR (400 MHz, CDCl₃) δ 2.30 (s, 1 H), 1.52 (s, 6 H). <chemistry id="chem0378" num="0378"><img file="EP2007756B1_D0404.tif" /></chemistry>

Methyl 2,2-d

i

methylbut-3-ynoate

0414To a solution of diazomethane (∼10g) in ether (400 mL) was added dropwise 2,2-dimethyl-but-3-ynoic acid (10.5 g, 93.7 mmol) at 0°C. The mixture was warmed to room temperature and stirred overnight. The mixture was distilled under atmospheric pressure to give crude methyl 2,2-dimethylbut-3-ynoate (14 g), which was used directly in the next step. ¹HNMR (400 MHz, CDCl₃) δ 3.76 (s, 3 H), 2.28 (s, 1 H), 1.50 (s, 6 H). <chemistry id="chem0379" num="0379"><img file="EP2007756B1_D0405.tif" /></chemistry>

Methyl 4-(2-amino-5-nitrophenyl)-2,2-dimethylbut-3-ynoate

0415To a deoxygenated solution of compound 2-bromo-4-nitroaniline (9.43 g, 43.7 mmol), methyl 2,2-dimethylbut-3-ynoate (5.00 g, 39.7 mmol), CuI (754 mg, 3.97 mmol) and triethylamine (8.03 g, 79.4 mmol) in toluene/H₂O (100/30mL) was added Pd(PPh₃)₄ (6.17 g, 3.97 mmol) under N₂. The mixture was heated at 70 °C and stirred for 24 h. After cooling, the solid was filtered off and washed with EtOAc (50 mL × 3). The organic layer was separated and the aqueous phase was washed with EtOAc (50 mL × 3). The combined organic layers were dried and evaporated under reduced pressure to give a residue, which was purified by column chromatography on silica gel (petroleum ether/ethyl acetate = 10/1) to obtain methyl 4-(2-amino-5-nitrophenyl)-2,2-dimethylbut-3-ynoate (900 mg, 9%). ¹H NMR (400 MHz, CDCl₃) δ 8.17 (d, <i>J</i>=2.8 Hz, 1 H), 8.01 (dd, <i>J</i>= 2.8, 9.2 Hz, 1 H), 6.65 (d, <i>J=</i> 9.2 Hz, 1 H), 5.10 (brs, 2 H), 3.80 (s, 3 H), 1.60 (s, 6 H). <chemistry id="chem0380" num="0380"><img file="EP2007756B1_D0406.tif" /></chemistry>

Methyl 4-(2-butyramido-5-nitrophenyl)-2,2-dimethylbut-3-ynoate

0416To a solution of methyl 4-(2-amino-5-nitrophenyl)-2,2-dimethylbut-3-ynoate (260 mg, 1.0 mmol) and pyridine (160 mg, 2.0 mol) in CH₂Cl₂ (20 mL) was added butyryl chloride (140 mg, 1.3 mmol) at 0 °C. The reaction mixture was warmed to room temperature and stirred for 3 h before the mixture was poured into ice-water. The organic layer was separated and the aqueous phase was extracted with CH₂Cl₂ (30 mL × 3). The combined organic layers were dried over anhydrous Na₂SO₄ and evaporated under reduced pressure to obtain methyl 4-(2-butyramido-5-nitrophenyl)-2,2-dimethylbut-3-ynoate (150 mg, 45%), which was used directly in the next step. ¹HNMR (400 MHz, CDCl₃) δ 8.79 (brs, 1 H), 8.71 (d, <i>J</i>=9.2 Hz, 1 H), 8.24 (d, <i>J</i>=2.8 Hz, 1 H), 8.17(dd, <i>J</i>=2.8, 9.2 Hz, 1 H), 3.82 (s, 3 H), 2.55 (t, <i>J</i>=7.2 Hz, 2 H), 1.85-1.75 (m, 2 H), 1.63 (s, 6 H), 1.06 (t, <i>J</i>=6.8 Hz, 3 H). <chemistry id="chem0381" num="0381"><img file="EP2007756B1_D0407.tif" /></chemistry>

Methyl 2-methyl-2-(5-nitro-1H-indol-2-yl)propanoate

0417To a deoxygenated solution of methyl 4-(2-butyramido-5-nitrophenyl)-2,2-dimethylbut-3-ynoate (1.8 g, 5.4 mmol) in acetonitrile (30 mL) was added Pd(CH₃CN)₂Cl₂ (0.42 g, 1.6= mmol) under N₂. The mixture was heated at reflux for 24 h. After cooling the mixture to ambient temperature, the solid was filtered off and washed with EtOAc (50 mL × 3). The filtrate was evaporated under reduced pressure to give a residue, which was purified by column chromatography on silica gel (petroleum ether/ethyl acetate = 30/1) to give methyl 2-methyl-2-(5-nitro-1H-indol-2-yl)propanoate (320 mg, 23%). ¹HNMR (400 MHz, CDCl₃) δ 9.05 (brs, 1 H), 8.52 (d, <i>J=</i> 2.0 Hz, 1 H), 8.09 (dd, <i>J</i>=2.0, 8.8 Hz, 1 H), 7.37 (d, <i>J=</i> 8.8 Hz, 1 H), 6.54 (d, <i>J</i>=1.6 Hz, 1 H), 3.78 (d, <i>J=</i> 9.6 Hz, 3 H), 1.70 (s, 6 H). <chemistry id="chem0382" num="0382"><img file="EP2007756B1_D0408.tif" /></chemistry>

Methyl 2-(5-amino-1H-indol-2-yl)-2-methylpropanoate

0418A suspension of methyl 2-methyl-2-(5-nitro-1H-indol-2-yl)propanoate (60 mg, 0.23 mmol) and Raney Nickel (10 mg) in MeOH (5 mL) was hydrogenated under hydrogen (1 atm) at room temperature overnight. The catalyst was filtered off through a celite pad and the filtrate was evaporated under <i>vacuum</i> to give a residue, which was purified by column chromatography on silica gel (petroleum ether/ethyl acetate = 5/1) to give methyl 2-(5-amino-1H-indol-2-yl)-2-methylpropanoate (20 mg, 38%). ¹HNMR (400 MHz, CDCl₃) δ 8.37 (br s, 1 H), 7.13 (d, <i>J=</i> 8.4 Hz, 1 H), 6.87 (d, <i>J=</i> 2.0 Hz, 1 H), 6.63 (dd, <i>J=</i>2.0, 8.4 Hz, 1 H), 6.20 (d, <i>J</i>=1.2 Hz, 1 H), 3.72 (d, <i>J=</i> 7.6 Hz, 3 H), 3.43 (br s, 1 H), 1.65 (s, 6 H); MS (ESI) m/e (M+H⁺) 233.2.

Example 44: 2-Isopropyl-1H-indol-5-amine

0419<chemistry id="chem0383" num="0383"><img file="EP2007756B1_D0409.tif" /></chemistry><chemistry id="chem0384" num="0384"><img file="EP2007756B1_D0410.tif" /></chemistry>

2-Isopropyl-5-nitro-1H-indole

0420A mixture of methyl 4-(2-butyramido-5-nitrophenyl)-2,2-dimethylbut-3-ynoate (0.50 g, 1.5 mmol) and TBAF (790 mg, 3.0 mmol) in DMF (20 mL) was heated at 70°C for 24 h. The reaction mixture was cooled to room temperature and poured into ice water. The mixture was extracted with ether (30 mL × 3). The combined organic layers were dried over anhydrous Na₂SO₄ and evaporated under reduced pressure to give a residue, which was purified by column chromatography on silica gel (petroleum ether/ethyl acetate = 20/1) to give 2-isopropyl-5-nitro-1H-indole (100 mg, 33%). ¹HNMR (400 MHz, CDCl₃) δ 8.69 (s, 1 H), 8.25 (br s, 1 H), 8.21 (dd, <i>J=</i> 2.4, 10.0 Hz, 1 H), 7.32 (d, <i>J=</i> 8.8 Hz, 1 H), 6.41 (s, 1 H), 3.07-3.14 (m, 1 H), 1.39 (d, <i>J=</i> 6.8 Hz, 6 H). <chemistry id="chem0385" num="0385"><img file="EP2007756B1_D0411.tif" /></chemistry>

2-Isopropyl-1H-indol-5-amine

0421A suspension of 2-isopropyl-5-nitro-1H-indole (100 mg, 0.49 mmol) and Raney Nickel (10 mg) in MeOH (10 mL) was hydrogenated under hydrogen (1 atm) at the room temperature overnight. The catalyst was filtered off through a celite pad and the filtrate was evaporated under <i>vacuum</i> to give a residue, which was purified by column (petroleum ether/ethyl acetate = 5/1) to give 2-isopropyl-1H-indol-5-amine (35 mg, 41%). ¹HNMR(400 MHz, CDCl₃) δ 7.69 (br s, 1 H), 7.10 (d, <i>J</i>=8.4 Hz, 1 H), 6.86 (d, <i>J=</i> 2.4Hz, 1 H), 6.58 (dd, <i>J=</i> 2.4, 8.8 Hz, 1 H), 6.07 (t, <i>J=</i> 1.2 Hz, 1 H), 3.55 (br s, 2 H), 3.06-2.99 (m, 1 H), 1.33 (d, <i>J</i> = 7.2 Hz, 6 H); MS (ESI) m/e (M+H⁺) 175.4.

Example 45: 1-(Benzo[d](1,3]dioxol-5-yl)-<i>N-</i>(2-(1-hydroxy-2-methylpropan-2-yl)-1H-indol-5-yl)cyclopropanecarboxamide

0422<chemistry id="chem0386" num="0386"><img file="EP2007756B1_D0412.tif" /></chemistry><chemistry id="chem0387" num="0387"><img file="EP2007756B1_D0413.tif" /></chemistry>

Triphenyl(2-aminobenzyl)phosphonium bromide

04232-Aminobenzyl alcohol (60.0 g, 0.487 mol) was dissolved in acetonitrile (2.5 L) and brought to reflux. Triphenylphosphine hydrobromide (167 g, 0.487 mol) was added and the mixture was heated at reflux for 3 h. The reaction mixture was concentrated to approximately 500 mL and left at room temperature for 1 h. The precipitate was filtered and washed with cold acetonitrile followed by hexane. The solid was dried overnight at 40°C under vacuum to give triphenyl(2-aminoberizyl)phosphonium bromide (193 g, 88%). <chemistry id="chem0388" num="0388"><img file="EP2007756B1_D0414.tif" /></chemistry>

Triphenyl((ethyl(2-carbamoyl)acetate)-2-benzyl)phosphonium bromide

0424To a suspension of triphenyl(2-aminobenzyl)phosphonium bromide (190 g, 0.43 mol) in anhydrous dichloromethane (1 L) was added ethyl malonyl chloride (55 ml, 0.43 mol). The reaction was stirred for 3 h at room temperature. The mixture was evaporated to dryness before ethanol (400 mL) was added. The mixture was heated at reflux until a clear solution was obtained. The solution was left at room temperature for 3 h. The precipitate was filtered, washed with cold ethanol followed by hexane and dried. A second crop was obtained from the mother liquor in the same way. In order to remove residual ethanol both crops were combined and dissolved in dichloromethane (approximately 700 mL) under heating and evaporated. The solid was dried overnight at 50 °C under vacuum to give triphenyl((ethyl(2-carbamoyl)acetate)-2-benzyl)-phosphonium bromide (139 g, 58%). <chemistry id="chem0389" num="0389"><img file="EP2007756B1_D0415.tif" /></chemistry>

Ethyl 2-

(1H-indol-2-yl)acetate

0425Triphenyl((ethyl(2-carbamoyl)acetate)-2-benzyl)phosphonium bromide (32.2 g, 57.3 mmol) was added to anhydrous toluene (150 mL) and the mixture was heated at reflux. Fresh potassium tert-butoxide (7.08 g, 63.1 mmol) was added in portions over 15 minutes. Reflux was continued for another 30 minutes. The mixture was filtered hot through a plug of celite and evaporated under reduced pressure. The residue was purified by column chromatography on silica gel (0-30% ethyl acetate in hexane over 45 min) to give ethyl 2-(1 H-indol-2-yl)acetate (9.12 g, 78%). <chemistry id="chem0390" num="0390"><img file="EP2007756B1_D0416.tif" /></chemistry>

<i>tert-</i>Butyl 2-((ethoxycarbonyl)methyl)-1H-indole-1-carboxylate

0426To a solution of ethyl 2-(1H-indol-2-yl)acetate (14.7 g, 72.2 mmol) in dichloromethane (150 mL) was added 4-dimethylaminopyridine (8.83 g, 72.2 mmol) and ditert-butyl carbonate (23.7 g, 108 mmol) in portions. After stirring for 2 h at room temperature, the mixture was diluted with dichloromethane, washed with water, dried over magnesium sulfate and purified by silica gel chromatography (0 to 20% EtOAc in hexane) to give <i>tert</i>-butyl 2-((ethoxycarbonyl)methyl)-1H-indole-1-carboxylate (20.0 g, 91%). <chemistry id="chem0391" num="0391"><img file="EP2007756B1_D0417.tif" /></chemistry>

<i>tert</i>-Butyl 2-(2-(ethoxycarbonyl)propan-2-yl)-1H-indole-1-carboxylate

0427<i>tert</i>-Butyl 2-((ethoxycarbonyl)methyl)-1H-indole-1-carboxylate (16.7 g, 54.9 mmol) was added to anhydrous THF (100 mL) and cooled to -78°C. A 0.5M solution of potassium hexamethyldisilazane (165 mL, 82 mmol) was added slowly such that the internal temperature stayed below -60 °C. Stirring was continued for 30 minutes at -78°C. To this mixture, methyl iodide (5.64 mL, 91 mmol) was added. The mixture was stirred for 30 min at room temperature and then cooled to -78°C. A 0.5M solution of potassium hexamethyldisilazane (210 mL, 104 mmol) was added slowly and the mixture was stirred for another 30 minutes at -78°C. More methyl iodide (8.6 mL, 137 mmol) was added and the mixture was stirred for 1.5 h at room temperature. The reaction was quenched with sat. aq. ammonium chloride and partitioned between water and dichloromethane. The aqueous phase was extracted with dichloromethane and the combined organic phases were dried over magnesium sulfate and evaporated under reduced pressure. The residue was purified by column chromatography on silica gel (0 to 20% ethylacetate in hexane) to give <i>tert</i>-butyl 2-(2-(ethoxycarbonyl)propan-2-yl)-1H-indole-1-carboxylate (17.1 g, 94%). <chemistry id="chem0392" num="0392"><img file="EP2007756B1_D0418.tif" /></chemistry>

Ethyl

2-(1H-indol-2-yl)-2-methylpropanoate

0428<i>tert</i>-Butyl 2-(2-(ethoxycarbonyl)propan-2-yl)-1H-indole-1-carboxylate (22.9 g, 69.1 mmol) was dissolved in dichloromethane (200 mL) before TFA (70 mL) was added. The mixture was stirred for 5 h at room temperature. The mixture was evaporated to dryness, taken up in dichloromethane and washed with saturated sodium bicarbonate solution, water, and brine. The product was purified by column chromatography on silica gel (0-20% EtOAc in hexane) to give ethyl 2-(1H-indol-2-yl)-2-methylpropanoate (12.5 g, 78%). <chemistry id="chem0393" num="0393"><img file="EP2007756B1_D0419.tif" /></chemistry>

Ethyl2-methyl-2-(5-nitro-1H-indol-2-yl)propanoate

0429Ethyl 2-(1H-indol-2-yl)-2-methylpropanoate (1.0 g, 4.3 mmol) was dissolved in concentrated sulfuric acid (6 mL) and cooled to -10°C (salt/ice-mixture). A solution of sodium nitrate (370 mg, 4.33 mmol) in concentrated sulfuric acid (3 mL) was added dropwise over 30 min. Stirring was continued for another 30 min at -10°C. The mixture was poured into ice and the product was extracted with dichloromethane. The combined organic phases were washed with a small amount of sat. aq. sodium bicarbonate. The product was purified by column chromatography on silica gel (5-30% EtOAc in hexane) to give ethyl 2-methyl-2-(5-nitro-1H-indol-2-yl)propanoate (0.68 g, 57%). <chemistry id="chem0394" num="0394"><img file="EP2007756B1_D0420.tif" /></chemistry>

2-Methyl-2-(5-nitro-1H-indol-2-yl)propan-1-ol

0430To a cooled solution of LiAlH₄ (1.0 M in THF, 1.1 mL, 1.1 mmol) in THF (5 mL) at 0 °C was added a solution of ethyl 2-methyl-2-(5-nitro-1H-indol-2-yl)propanoate (0.20 g, 0.72 mmol) in THF (3.4 mL) dropwise. After addition, the mixture was allowed to warm up to room temperature and was stirred for 3 h. The mixture was cooled to 0 °C before water (2 mL) was slowly added followed by careful addition of 15% NaOH (2 mL) and water (4 mL). The mixture was stirred at room temperature for 0.5 h and was filtered through a short plug of celite using ethyl acetate. The organic layer was separated from the aqueous layer, dried over Na₂SO₄, filtered and evaporated under reduced pressure. The residue was purified by column chromatography on silica gel (ethyl acetate/hexane = 1/1) to give 2-methyl-2-(5-nitro-1H-indol-2-yl)propan-1-ol (0.098 g, 58%). <chemistry id="chem0395" num="0395"><img file="EP2007756B1_D0421.tif" /></chemistry>

2-(5-Amino-1H-indol-2-yl)-2-methylpropan-1-ol

0431To a solution of 2-methyl-2-(5-nitro-1H-indol-2-yl)propan-1-ol (0.094 g, 0.40 mmol) in ethanol (4 mL) was added tin chloride dihydrate (0.451 g, 2.0 mmol). The mixture was heated in the microwave at 120 °C for 1 h. The mixture was diluted with ethyl acetate and water before being quenched with saturated aqueous NaHCO₃. The reaction mixture was filtered through a plug of celite using-ethyl acetate. The organic layer was separated from the aqueous layer, dried over Na₂SO₄, filtered and evaporated under reduced pressure to give 2-(5-amino-1H-indol-2-yl)-2-methylpropan-1-ol (0.080 g, 98%).

Example 46: 2-(Pyridin-2-yl)-1H-indol-5-amine

0432<chemistry id="chem0396" num="0396"><img file="EP2007756B1_D0422.tif" /></chemistry><chemistry id="chem0397" num="0397"><img file="EP2007756B1_D0423.tif" /></chemistry>

4-Nitro-2-(pyridin-2-ylethynyl)aniline

0433To the solution of 2-iodo-4-nitroaniline (3.0 g, 11 mmol) in DMF (60mL) and Et₃N (60 mL) was added 2-ethynylpyridine (3.0 g, 45 mmol), Pd(PPh₃)₂Cl₂ (600 mg) and CuI (200 mg) under N₂. The reaction mixture was stirred at 60 °C for 12 h. The mixture was diluted with water and extracted with dichloromethane (3 × 100 mL). The combined organic layers were washed with brine, dried over anhydrous Na₂SO₄ and concentrated in vacuum. The residue was purified by chromatography on silica gel (5-10% ethyl acetate/petroleum ether) to afford 4-nitro-2-(pyridin-2-ylethynyl)aniline (1.5 g, 60%). ¹HNMR (300 MHz, CDCl₃) δ 8.60 (s, 1 H), 8.13 (d, <i>J</i>=2.1 Hz, 1 H), 7.98 (d, <i>J=</i> 1.8, 6.9 Hz, 1 H), 7.87-7.80 (m, 2 H), 7.42-7.39 (m, 1 H), 7.05 (brs, 2 H), 6.80 (d, J= 6.9 Hz, 1 H). <chemistry id="chem0398" num="0398"><img file="EP2007756B1_D0424.tif" /></chemistry>

5-Nitro-2-(pyridin-2-yl)-1H-indole

0434To the solution of 4-nitro-2-(pyridin- 2-ylethynyl)aniline (1.5 g, 6.3 mmol) in DMF (50 mL) was added t-BuOK (1.5 g, 13 mmol). The reaction mixture was stirred at 90 °C for 2 h. The mixture was diluted with water and extracted with dichloromethane (3 × 50 mL). The combined organic layers were washed with brine, dried over anhydrous Na₂SO₄ and concentrated in vacuum. The residue was purified by chromatography on silica gel (5-10% ethyl acetate/petroleum ether) to afford 5-nitro-2-(pyridin-2-yl)-1H-indole (1.0 g, 67%yield). ¹HNMR (300 MHz, d-DMSO) δ 12.40 (s, 1H), 8.66 (d, J= 2.1 Hz, 1 H), 8.58 (d, J= 1.8 Hz, 1 H), 8.07-7.91 (m, 3 H), 7.59 (d, J= 6.6 Hz, 1 H), 7.42-7.37 (m, 2 H). <chemistry id="chem0399" num="0399"><img file="EP2007756B1_D0425.tif" /></chemistry>

2-(Pyridin-2-yl)-1H-indol-5-amine

0435To a solution of 5-nitro-2-(pyridin-2-yl)-1H-indole (700 mg, 2.9 mmol) in EtOH (20 mL) was added SnCl₂ (2.6 g, 12 mmol). The mixture was heated at reflux for 10 h. Water was added and the mixture was extracted with EtOAc (50 mL × 3). The combined organic layers were washed with brine, dried over anhydrous Na₂SO₄ and concentrated in vacuum. The residue was purified by chromatography on silica gel (5-10% ethyl acetate/petroleum ether) to afford 2-(pyridin-2-yl)-1H-indol-5-amine (120 mg, 20%). ¹H NMR (400 MHz, CDCl₃) δ 9.33 (brs, 1 H), 8.55 (dd, <i>J=</i>1.2, 3.6 Hz, 1 H), 7.76-7.67 (m, 2 H), 7.23 (d, <i>J=</i> 6.4 Hz, 1 H), 7.16-7.12 (m, 1 H), 6.94 (d, J= 2.0 Hz, 1 H), 6.84 (d, <i>J=</i> 2.4 Hz, I H), 6.71-6.69 (dd, <i>J</i>=2.0, 8.4 Hz, 1 H).

Example 47: 2-(Pyridin-2-yl)-1H-indol-5-amine

0436<chemistry id="chem0400" num="0400"><img file="EP2007756B1_D0426.tif" /></chemistry><chemistry id="chem0401" num="0401"><img file="EP2007756B1_D0427.tif" /></chemistry>

[2-(<i>tert</i>-Butyl-dimethyl-silanyloxy)-ethyl]-(2-iodo-4-nitro-phenyl)-amine

0437To a solution of 2-iodo-4-nitroaniline (2.0 g, 7.6 mmol) and 2<i>-</i>(<i>tert-</i>butyldimethylsilyloxy)-acetaldehyde (3.5 g, 75% purity, 15 mmol) in methanol (30 mL) was added TFA (1.5 mL) at 0 °C. The reaction mixture was stirred at this temperature for 30 min before NaCNBH₃ (900 mg, 15 mmol) was added in portions. The mixture was stirred for 2 h and was then quenched with water. The resulting mixture was extracted with EtOAc (30 mL × 3), the combined organic extracts were dried over anhydrous Na₂SO₄ and evaporated under vacuum, and the residue was purified by chromatography on silica gel (5 % ethyl acetate/petroleum) to afford [2-(tert-butyl-dimethyl-silanyloxy)-ethyl]-(2-iodo-4-nitrophenyl)-amine (800 mg, 25 %). ¹HNMR (300 MHz, CDCl₃) δ 8.57 (d, <i>J</i>=2.7 Hz, 1 H), 8.12 (dd, <i>J=</i> 2.4 ,9.0 Hz, 1 H), 6.49 (d, <i>J</i>=9.3 Hz, 1 H), 5.46 (br s, 1 H), 3.89 (t, <i>J=</i> 5.4 Hz, 2 H), 3.35 (q, <i>J=</i> 5.4 Hz, 2 H), 0.93 (s, 9 H), 0.10 (s, 6 H). <chemistry id="chem0402" num="0402"><img file="EP2007756B1_D0428.tif" /></chemistry>

5-{2-[2-(<i>tert</i>-Butyl-dimethyl-silanyloxy)-ethylamino]-5-nitro-phenyl}-3,3-dimethyl-pent-4-ynoic acid ethyl ester

0438To a solution of [2-(<i>tert</i>-butyl-dimethyl-silanyloxy)-ethyl]-(2-iodo-4-nitro-phenyl)-amine (800 mg, 1.9 mmol) in Et₃N (20 mL) was added Pd(PPh₃)₂Cl₂ (300 mg, 0.040 mmol), CuI (76 mg, 0.040 mmol) and 3,3-dimethyl-but-1-yne (880 mg, 5.7 mmol) successively under N₂ protection. The reaction mixture was heated at 80°C for 6 h and allowed to cool down to room temperature. The resulting mixture was extracted with EtOAc (30 mL × 3). The combined organic extracts were dried over anhydrous Na₂SO₄ and evaporated under vacuum to give 5-{2-[2-(<i>tert-b</i>utyl-dimethyl-silanyloxy)-ethylamino]-5-nitro-phenyl}-3,3-dimethyl-pent-4- ynoic acid ethyl ester (700 mg, 82 %), which was used in the next step without further purification. ¹HNMR (400 MHz, CDCl₃) δ 8.09 (s, 1 H), 8.00 (d, <i>J=</i> 9.2 Hz, 1 H), 6.54 (d, <i>J=</i> 9.2 Hz, 1 H), 6.45 (brs, 1 H), 4.17-4.10 (m, 4 H), 3.82 (t, <i>J</i>=5.6 Hz, 2 H), 3.43 (q, <i>J=</i> 5.6 Hz, 2 H), 2.49 (s, 2 H), 1.38 (s, 6 H), 1.28 (t, <i>J</i>=7.2 Hz, 3 H), 0.84 (s, 9 H), 0.00 (s, 6 H). <chemistry id="chem0403" num="0403"><img file="EP2007756B1_D0429.tif" /></chemistry>

3-[1-(2-Hydroxy-ethyl)-5-nitro-1H-indol-2-yl]-3-methyl-butyric acid ethyl ester

0439A solution of 5- {2-[2-(<i>tert</i>-butyl-dimethyl-silanyloxy)-ethylamino]-5-nitrophenyl}-3,3-dimethyl-pent-4- ynoic acid ethyl ester (600 mg, 1.34 mmol) and PdCl₂(650 mg) in CH₃CN (30 mL) was heated at reflux overnight. The resulting mixture was extracted with EtOAc (30 mL × 3). The combined organic extracts were dried over anhydrous Na₂SO₄ and evaporated under vacuum. The residue was dissolved in THF (20 mL) and TBAF (780 mg, 3.0 mmol) was added. The mixture was stirred at room temperature for 1 h, the solvent was removed under vaccum, and the residue was purified by chromatography on silica gel (10% ethyl acetate/petroleum) to afford 3-[1-(2-hydroxy-ethyl)-5-nitro-1H-indol-2-yl]-3-methyl-butyric acid ethyl ester (270 mg, 60 %). ¹HNMR (300 MHz, CDCl₃) δ 8.45 (d, <i>J=</i> 2.1 Hz, 1 H), 8.05 (dd, <i>J=</i> 2.1, 9.0 Hz, 1 H), 6.36 (d, <i>J=</i> 9.0 Hz, 1 H), 6.48 (s, 1 H), 4.46 (t, <i>J</i> =6.6Hz, 2 H), 4.00-3.91 (m, 4 H), 2.76 (s, 2 H), 1.61 (s, 6 H), 0.99 (t, <i>J=</i> 7.2 Hz, 1 H), 0.85 (s, 9 H), 0.03 (s, 6 H). <chemistry id="chem0404" num="0404"><img file="EP2007756B1_D0430.tif" /></chemistry>

3-[1-(2-Hydroxy-ethyl)-5-nitro-1H-indol-2-yl],3-methyl-butan-1-ol

0440To a solution of 3-[1-(2-hydroxy-ethyl)-5-nitro-1H-indol-2-yl]-3-methyl-butyric acid ethyl ester (700 mg, 2.1 mmol) in THF (25 mL) was added DIBAL-H (1.0 M, 4.2 mL, 4.2 mmol) at -78 °C. The mixture was stirred at room temperature for 1 h. Water (2 mL) was added and the resulting mixture was extracted with EtOAc (15 mL × 3). The combined organic layers were dried over anhydrous Na₂SO₄ and evaporated under vacuum. The residue was purified by chromatography on silica gel (15 % ethyl acetate/petroleum) to afford 3-[1-(2-hydroxy-ethyl)-5-nitro-1H-indol-2-yl]-3-methyl-butan-1-ol (300 mg, 49%). ¹H NMR (300 MHz, d-DMSO) δ 8.42 (d, <i>J =</i> 1.5 Hz, 1 H), 7.95 (dd, <i>J</i> = 1.2, 8.7 Hz, 1 H), 6.36 (d, <i>J =</i> 9.3 Hz, 1 H), 6.50 (s, 1 H), 5.25 (br s, 1 H), 4.46-4.42 (m, 4 H), 3.69-3.66 (m ,2 H), 3.24-3.21 (m, 2 H), 1.42(s,6H). <chemistry id="chem0405" num="0405"><img file="EP2007756B1_D0431.tif" /></chemistry>

3-(5-Amino-1-(2-hydroxy-ethyl)-1H-indol-2-yl]-3-methyl-butan-1-ol

0441A solution of 3-[1-(2-hydroxy-ethyl)-5-nitro-1H-indol-2-yl]-3-methyl-butan-1-ol (300 mg, 1.03 mmol) and Raney Nickel (200 mg,) in CH₃OH (30 mL) was stirred for 5 h at room temperature under a H₂ atmosphere. The catalyst was filtered through a celite pad and the filtrate was evaporated under vacuum to give a residue, which was purified by preparative TLC to afford 3-[5-amino-1-(2-hydroxy-ethyl)-1H-indol-2-yl]-3-methyl-butan-1-ol (70 mg, 26%). ¹H NMR (300 MHz, CDCl₃) δ 7.07 (d, <i>J=</i> 8.7 Hz, 1 H), 6.83 (d, <i>J=</i> 2.1 Hz, 1 H), 6.62 (dd, <i>J=</i> 2.1, 8.4 Hz, 1 H), 6.15 (s, 1 H), 4.47 (t, <i>J=</i> 5.4 Hz, 2 H), 4.07 (t, <i>J=</i> 5.4 Hz, 2 H), 3.68 (t, <i>J</i> = 5.7 Hz, 2 H), 2.16 (t, <i>J</i> = 5.7 Hz, 2 H), 4.00-3.91 (m, 4 H), 2.76 (s, 2 H), 1.61 1 (s, 6 H), 1.42 (s, 6 H).

Example 48: <i>tert-</i>Butyl 2-(5-amino-1H-indol-2-yl)piperidine-1-carboxylate

0442<chemistry id="chem0406" num="0406"><img file="EP2007756B1_D0432.tif" /></chemistry><chemistry id="chem0407" num="0407"><img file="EP2007756B1_D0433.tif" /></chemistry>

2-(Piperidin-2-yl)-1H-indol-5-amine

04435-Nitro-2-(pyridin-2-yl)-1H-indole (1.0 g, 4.2 mmol) was added to HCl/MeOH (2 M, 50 mL). The reaction mixture was stirred at room temperature for 1 h and the solvent was evaporated under vacuum. PtO₂ (200 mg) was added to a solution of the residue in MeOH (50 mL) and the reaction mixture was stirred under hydrogen atmosphere (1 atm) at room temperature for 2 h. The catalyst was filtered through a celite pad and the solvent was evaporated under vacuum to afford 2-(piperidin-2-yl)-1H-indol-5-amine (1.0 g), which was directly used in the next step. <chemistry id="chem0408" num="0408"><img file="EP2007756B1_D0434.tif" /></chemistry>

<i>tert</i>-Butyl 2-(5-amino-1H-indol-2-yl)piperidine-1-carboxylate

0444To a solution of 2-(piperidin-2-yl)-1H-indol-5-amine (1.0 g) in Et₃N (25 mL) and THF (25mL) was added Boc₂O (640 mg, 2.9 mmol). The reaction mixture was stirred at room temperature overnight. The mixture was diluted with water and extracted with dichloromethane (3 × 25 mL). The combined organic layers were washed with brine, dried over anhydrous Na₂SO₄ and concentrated in vacuum. The residue was purified by chromatography on silica gel (5-10% ethyl acetate/petroleum ether) followed by preparative HPLC to afford <i>tert</i>-butyl 2-(5-amino-1H-indol-2-yl)piperidine-1-carboxylate (15 mg, 1% over 2 steps). ¹H NMR (400 MHz, CDCl₃) δ 8.82 (s, 1 H), 7.58 (s, 1 H), 7.22 (d, <i>J</i> = 8.8 Hz, 1 H), 7.02 (d, <i>J =</i> 1.6, 8.0 Hz, 1 H), 6.42 (s, 1H), 6.25 (s, 1 H), 3.91-3.88 (m, 1 H), 3.12-3.10 (m, 1 H), 2.81-2.76 (m, 1 H), 2.06-1.97 (m, 4 H), 1.70-1.58 (m, 2H), 1.53 (s, 9 H).

Example 49: 6-amino-1H-indole-2-carbonitrile

0445<chemistry id="chem0409" num="0409"><img file="EP2007756B1_D0435.tif" /></chemistry><chemistry id="chem0410" num="0410"><img file="EP2007756B1_D0436.tif" /></chemistry>

(3-Nitrophenyl)hydrazine hydrochloride

04463-Nitroaniline (28 g, 0.20 mol) was dissolved in a mixture of H₂O (40 mL) and 37% HCl (40 mL). A solution of NaNO₂ (14 g, 0.20 mol) in H₂O (60 mL) was added to the mixture at 0 °C, and then a solution of SnCl₂.H₂O (140 g, 0.60 mol) in 37% HCl (100 mL) was added. After stirring at 0 °C for 0.5 h, the insoluble material was isolated by filtration and was washed with water to give (3-nitrophenyl)hydrazine hydrochloride (28 g, 73%). <chemistry id="chem0411" num="0411"><img file="EP2007756B1_D0437.tif" /></chemistry>

(<i>E</i>)-Ethyl 2-(2-(3-nitrophenyl)hydrazono)propanoate

0447(3-Nitrophenyl)hydrazine hydrochloride (30 g, 0.16 mol) and 2-oxo-propionic acid ethyl ester (22 g, 0.19 mol) were dissolved in ethanol (300 mL). The mixture was stirred at room temperature for 4 h before the solvent was evaporated under reduced pressure to give (<i>E</i>)-ethyl 2-(2-(3-nitrophenyl)hydrazono)propanoate, which was used directly in the next step. <chemistry id="chem0412" num="0412"><img file="EP2007756B1_D0438.tif" /></chemistry>

Ethyl 4-nitro-1H-indole-2-carboxylate and ethyl 6-nitro-1H-indole-2-carboxylate

0448(<i>E</i>)-Ethyl 2-(2-(3-nitrophenyl)hydrazono)propanoate was dissolved in toluene (300 mL) and PPA (30 g) was added. The mixture was heated at reflux overnight and then was cooled to room temperature. The solvent was decanted and evaporated to obtain a crude mixture that was taken on to the next step without purification (15 g, 40%). <chemistry id="chem0413" num="0413"><img file="EP2007756B1_D0439.tif" /></chemistry>

4-Nitro-1H-indole-2-carboxylic acid and 6-nitro-1H-indole-2-carboxylic acid

0449A mixture of ethyl 6-nitro-1H-indole-2-carboxylate (0.5 g) and 10 % NaOH (20 mL) was heated at reflux overnight and then was cooled to room temperature. The mixture was extracted with ether and the aqueous phase was acidified with HCl to pH 1∼2. The insoluble solid was isolated by filtration to give a crude mixture that was taken on to the next step without purification (0.3 g, 68%). <chemistry id="chem0414" num="0414"><img file="EP2007756B1_D0440.tif" /></chemistry>

4-Nitro-1H-indole-2-carboxamide and 6-nitro-1H-indole-2-carboxamide

0450A mixture of 6-nitro-1H-indole-2-carboxylic acid (12 g, 58 mmol) and SOCl₂ (50 mL, 64 mmol) in benzene (150 mL) was heated at reflux for 2 h. The benzene and excess SOCl₂ was removed under reduced pressure. The residue was dissolved in anhydrous CH₂Cl₂ (250 mL) and NH3.H₂O (22 g, 0.32 mol) was added dropwise at 0 °C. The mixture was stirred at room temperature for 1 h. The insoluble solid was isolated by filtration to obtain crude mixture (9.0 g, 68%), which was used directly in the next step. <chemistry id="chem0415" num="0415"><img file="EP2007756B1_D0441.tif" /></chemistry>

4-Nitro-1H-indole-2-carbonitrile

and

6-nitro-1H-indole-2-carbonitrile

04516-Nitro-1H-indole-2-carboxamide (5.0 g, 24 mmol) was dissolved in CH₂Cl₂ (200 mL). Et₃N (24 g, 0.24 mol) and (CF₃CO)₂O (51 g, 0.24 mol) were added dropwise to the mixture at room temperature. The mixture was continued to stir for 1 h and was then poured into water (100 mL). The organic layer was separated and the aqueous layer was extracted with EtOAc (100 mL × 3). The combined organic layers were dried over Na₂SO_4, filtered and concentrated under reduced pressure to obtain crude product which was purified by column chromatography on silica gel to give a impure sample of 4-nitro-1H-indole-2-carbonitrile (2.5 g, 55%). <chemistry id="chem0416" num="0416"><img file="EP2007756B1_D0442.tif" /></chemistry>

6-Amino-1H-indole-2-carbonitrile

0452A mixture of 6-nitro-1H-indole-2-carbonitrile (2.5 g, 13 mmol) and Raney Nickel (500 mg) in EtOH (50 mL) was stirred at room temperature under H₂ (1 atm) for 1 h. Raney Nickel was removed via filtration and the filtrate was evaporated under reduced pressure to give a residue, which was purified by column chromatograpy on silica get to give 6-amino-1H-indole-2-carbonitrile (1.0 g, 49 %). ¹H NMR (DMSO-<i>d</i>₆) δ 12.75 (br s, 1 H), 7.82 (d, <i>J=</i> 8 Hz, 1 H), 7.57 (s, 1H), 7.42 (s, 1 H), 7.15 (d, <i>J =</i> 8 Hz, 1 H); MS (ESI) m/e (M+H⁺) 158.2.

Example 50: 6-Amino-1H-indole-3-carbonitrile

0453<chemistry id="chem0417" num="0417"><img file="EP2007756B1_D0443.tif" /></chemistry><chemistry id="chem0418" num="0418"><img file="EP2007756B1_D0444.tif" /></chemistry>

6-Nitro-1H-indole-3-carbonitrile

0454To a solution of 6-nitroindole (4.9 g 30 mmol) in DMF (24 mL) and CH₃CN (240 mL) was added dropwise a solution of ClSO₂NCO (5.0 mL) in CH₃CN (39 mL) at 0 °C. After addition, the reaction was allowed to warm to room temperature and was stirred for 2 h. The mixture was then poured into ice-water and basified with sat. NaHCO₃ solution to pH 7∼8. The mixture was extracted with ethyl acetate. The organics were washed with brine, dried over Na₂SO₄ and concentrated to give 6-nitro-1H-indole-3-carbonitrile (4.6 g, 82%). <chemistry id="chem0419" num="0419"><img file="EP2007756B1_D0445.tif" /></chemistry>

6-Amino-1H-indole-3-carbonitrile

0455A suspention of 6-nitro-1H-indole-3-carbonitrile (4.6 g, 25 mmol) and 10% Pd-C (0.46 g) in EtOH (50 mL) was stirred under H₂ (1 atm) at room temperature overnight. After filtration, the filtrate was concentrated and the residue was purified by column chromatography on silica gel (petroleum ether/ethyl acetate = 3/1) to give 6-amino-1H-indole-3-carbonitrile (1.0 g, 98%) as a pink solid. ¹H NMR (DMSO-<i>d</i>₆) δ 11.51 (s, 1 H), 7.84 (d, <i>J =</i> 2.4 Hz, 1 H), 7.22 (d, <i>J=</i> 8.4 Hz, 1 H), 6.62 (s, 1H), 6.56 (d, <i>J =</i> 8.4 Hz, 1 H), 5.0 (s, 2H); MS (ESI) m/e (M+H⁺) 157.1.

Example 51: 2-

tert

-Butyl-1H-indol-6-amine

0456<chemistry id="chem0420" num="0420"><img file="EP2007756B1_D0446.tif" /></chemistry><chemistry id="chem0421" num="0421"><img file="EP2007756B1_D0447.tif" /></chemistry>

N-o-Tolylpivalamide

0457To a solution of o-tolylamine (21 g, 0.20 mol) and Et₃N (22 g, 0.22 mol) in CH₂Cl₂ was added 2,2-dimethyl-propionyl chloride (25 g, 0.21 mol) at 10 °C. After addition, the mixture was stirred overnight at room temperature. The mixture was washed with aq. HCl (5%, 80 mL), saturated aq. NaHCO₃ and brine. The organic layer was dried over Na₂SO₄ and concentrated under <i>vacuum</i> to give <i>N</i>-o-tolylpivalamide (35 g, 91%). ¹H NMR (300 MHz, CDCl₃) δ 7.88 (d, <i>J=</i> 7.2 Hz, 1 H), 7.15-7.25 (m, 2 H), 7.05 (t, <i>J=</i> 7.2 Hz, 1 H), 2.26 (s, 3 H), 1.34 (s, 9 H). <chemistry id="chem0422" num="0422"><img file="EP2007756B1_D0448.tif" /></chemistry>

2-<i>tert</i>-Butyl-1H-indole

0458To a solution of <i>N</i>-o-tolylpivalamide (30.0 g, 159 mmol) in dry THF (100 mL) was added dropwise n-BuLi (2.5 M in hexane, 190 mL) at 15 °C. After addition, the mixture was stirred overnight at 15 °C. The mixture was cooled in an ice-water bath and treated with saturated NH₄Cl. The organic layer was separated and the aqueous layer was extracted with ethyl acetate. The combined organic layers were dried over anhydrous Na₂SO_4, filtered, and concentrated in <i>vacuum.</i> The residue was purified by column chromatography on silica gel to give 2-<i>tert</i>-butyl-1H-indole (24 g, 88%). ¹H NMR (300 MHz, CDCl₃) δ 7.99 (br. s, 1 H), 7.54 (d, <i>J</i> = 7.2 Hz, 1 H), 7.05 (d, <i>J=</i> 7.8 Hz, 1 H), 7.06 -7.13 (m, 2 H), 6.26 (s, 1 H), 1.39 (s, 9 H). <chemistry id="chem0423" num="0423"><img file="EP2007756B1_D0449.tif" /></chemistry>

2-tert-Butylindoline

0459To a solution of 2-<i>tert</i>-butyl-1H-indole (10 g, 48 mmol) in AcOH (40 mL) was added NaBH₄ at 10 °C. The mixture was stirred for 20 minutes at 10 °C before being treated dropwise with H₂O under ice cooling. The mixture was extracted with ethyl acetate. The combined organic layers were dried over anhydrous Na₂SO_4, filtered, and concentrated under <i>vacuum</i> to give 2-<i>tert</i>-butylindoline (9.8 g), which was used directly in the next step. <chemistry id="chem0424" num="0424"><img file="EP2007756B1_D0450.tif" /></chemistry>

2-<i>tert</i>-butyl-6-nitroindoline and 2-<i>tert</i>-butyl-5-nitro-1H-indole

0460To a solution of 2-tert-butylindoline (9.7 g) in H₂SO₄ (98%, 80 mL) was slowly added KNO₃ (5.6 g, 56 mmol) at 0 °C. After addition, the reaction mixture was stirred at room temperature for 1 h. The mixture was carefully poured into cracked ice, basified with Na₂CO₃ to pH 8 and extracted with ethyl acetate. The combined extracts were washed with brine, dried over anhydrous Na₂SO₄ and concentrated under <i>vacuum.</i> The residue was purified by column chromatography to give 2-<i>tert</i>-butyl-6-nitroindoline (4.0 g, 31 % over two steps). ¹H NMR (300 MHz, CDCl₃) δ 7.52 (dd, <i>J=</i> 1.8, 8.1 Hz, 1 H), 7.30 (s, 1 H), 7.08 (d, <i>J</i> = 7.8 Hz, 1 H), 3.76 (t, <i>J</i> = 9.6 Hz, 1 H), 2.98 - 3.07 (m, 1 H), 2.82 - 2.91 (m, 1 H), 0.91 (s, 9 H). <chemistry id="chem0425" num="0425"><img file="EP2007756B1_D0451.tif" /></chemistry>

2-<i>tert</i>-Butyl-6-nitro-1H-indole

0461To a solution of 2-<i>tert</i>-butyl-6-nitroindoline (2.0 g, 9.1 mmol) in 1,4-dioxane (20 mL) was added DDQ (6.9 g, 30 mmol) at room temperature. The mixture was heated at reflux for 2.5 h before being filtered and concentrated under <i>vacuum.</i> The residue was purified by column chromatography to give 2-<i>tert</i>-butyl-6-nitro-1H-indole (1.6 g, 80%). ¹H NMR (300 MHz, CDCl₃) δ 8.30 (br. s, 1 H), 8.29 (s, 1 H), 8.00 (dd, <i>J=</i> 2.1, 8.7 Hz, 1 H), 7.53 (d, <i>J =</i> 9.3 Hz, 1 H), 6.38 (s, 1 H), 1.43 (s, 9 H). <chemistry id="chem0426" num="0426"><img file="EP2007756B1_D0452.tif" /></chemistry>

2-<i>tert</i>-Butyl-1H-indol-6-amine

0462To a solution of 2-<i>tert</i>-butyl-6-nitro-1H-indole (1.3 g, 6.0 mmol) in MeOH (10 mL) was added Raney Nickel (0.2 g). The mixture was hydrogenated under 1 atm of hydrogen at room temperature for 3 h. The reaction mixture was filtered and the filtrate was concentrated. The residue was washed with petroleum ether to give 2-<i>tert</i>-butyl-1H-indol-6-amine (1.0 g, 89%). ¹H NMR (300 MHz, DMSO-<i>d</i>₆) δ 10.19 (s, 1 H), 6.99 (d, <i>J =</i> 8.1 Hz, 1 H), 6.46 (s, 1 H), 6.25 (dd, <i>J=</i> 1.8, 8.1 Hz, 1 H), 5.79 (d, <i>J=</i> 1.8 Hz, 1 H), 4.52 (s, 2 H), 1.24 (s, 9 H); MS (ESI) m/e (M+H⁺) 189.1.

Example 52:

3-<i>tert</i>-Butyl-1H-indol-6-amine

0463<chemistry id="chem0427" num="0427"><img file="EP2007756B1_D0453.tif" /></chemistry>

3-<i>tert</i> Butyl-6-nitro-1H-indole

0464To a mixture of 6-nitroindole (1.0 g, 6.2 mmol), zinc triflate (2.1 g, 5.7 mmol), and TBAI (1.7 g, 5.2 mmol) in anhydrous toluene (11 mL) was added DIEA (1.5 g, 11 mmol) at room temperature under nitrogen. The reaction mixture was stirred for 10 min at 120 °C, followed by the addition of <i>t</i>-butyl bromide (0.71 g, 5.2 mmol). The resulting mixture was stirred for 45 min at 120 °C. The solid was filtered off and the filtrate was concentrated to dryness. The residue was purified by column chromatography on silica gel (petroleum ether/ethyl acetate = 20:1) to give 3-<i>tert</i>-butyl-6-nitro-1H-indole (0.25 g, 19%) as a yellow solid. ¹H-NMR (CDCl₃) δ 8.32 (d, <i>J=</i> 2.1 Hz, 1H), 8.00 (dd, <i>J =</i> 2.1, 14.4 Hz, 1H), 7.85 (d, <i>J</i> = 8.7 Hz, 1H), 7.25 (s, 1H), 1.46 (s, 9H). <chemistry id="chem0428" num="0428"><img file="EP2007756B1_D0454.tif" /></chemistry>

3-<i>tert</i>-Butyl-1H-indol-6-amine

0465A suspension of 3-<i>tert</i>-butyl-6-nitro-1H-indole (3.0 g, 14 mmol) and Raney Nickel (0.5 g) was hydrogenated under H₂ (1 atm) at room temperature for 3 h. The catalyst was filtered off and the filtrate was concentrated to dryness. The residue was purified by column on silica gel (petroleum ether/ethyl acetate = 4:1) to give 3-<i>tert</i>-butyl- 1H-indol-6-amine (2.0 g, 77%) as a gray solid. ¹HNMR (CDCl₃) δ 7.58 (m, 2H), 6.73 (d, <i>J =</i> 1.2 Hz, 1H), 6.66 (s, 1H), 6.57(dd, <i>J</i> = 0.8, 8.6 Hz, 1H), 3.60 (br, 2H), 1.42 (s, 9H).

Example 53: 5-(Trifluoromethyl]-1H-indol-6-amine

0466<chemistry id="chem0429" num="0429"><img file="EP2007756B1_D0455.tif" /></chemistry><chemistry id="chem0430" num="0430"><img file="EP2007756B1_D0456.tif" /></chemistry>

1-Methyl-2,4-dinitro-5-(trifluoromethyl)benzene

0467To a mixture of HNO₃ (98%, 30 mL) and H₂SO₄ (98%, 30 mL) was added dropwise 1-methyl-3-trifluoromethyl-benzene (10 g, 63 mmol) at 0 °C. After addition, the mixture was stirred at rt for 30 min and was then poured into ice-water. The precipitate was filtered and washed with water to give 1-methyl-2,4-dinitro-5-trifluoromethyl-benzene (2.0 g, 13%). <chemistry id="chem0431" num="0431"><img file="EP2007756B1_D0457.tif" /></chemistry>

(<i>E</i>)-2-(2,4-Dinitro-5-(trifluoromethyl)phenyl)-<i>N,N</i>-dimethylethenamine

0468A mixture of 1-methyl-2,4-dinitro-5-trifluoromethyl-benzene (2.0 g, 8.0 mmol) and DMA (1.0 g, 8.2 mmol) in DMF (20 mL) was stirred at 100 °C for 30 min. The mixture was poured into ice-water and stirred for 1 h. The precipitate was filtered and washed with water to give (<i>E</i>)-2-(2,4-dinitro-5-(trifluoromethyl)phenyl)-<i>N,N</i>-dimethylethenamine (2.1 g, 86%). <chemistry id="chem0432" num="0432"><img file="EP2007756B1_D0458.tif" /></chemistry>

5-(Trifluoromethyl)-1H-indol-6-amine

0469A suspension of (<i>E</i>)-2-(2,4-dinitro-5-(trifluoromethyl)phenyl)-<i>N,N-</i>dimethylethenamine (2.1 g, 6.9 mmol) and Raney Nickel (1 g) in ethanol (80 mL) was stirred under H₂ (1 atm) at room temperature for 5 h. The catalyst was filtered off and the filtrate was concentrated to dryness. The residue was purified by column on silica gel to give 5-(trifluoromethyl)-1H-indol-6-amine (200 mg, 14%). ¹H NMR (DMSO-<i>d</i>₆) δ 10.79 (br s, 1 H), 7.55 (s, 1 H), 7.12 (s, 1 H), 6.78 (s, 1 H), 6.27(s, 1 H), 4.92 (s, 2 H); MS (ESI) m/e (M+H⁺): 200.8.

Example 54: 5-Ethyl-1H-indol-6-amine

0470<chemistry id="chem0433" num="0433"><img file="EP2007756B1_D0459.tif" /></chemistry><chemistry id="chem0434" num="0434"><img file="EP2007756B1_D0460.tif" /></chemistry>

1-(Phenylsulfonyl)indoline

0471To a mixture of DMAP (1.5 g), benzenesulfonyl chloride (24.0 g, 136 mmol) and indoline (14.7 g, 124 mmol) in CH₂Cl₂ (200 mL) was added dropwise Et₃N (19.0 g, 186 mmol) at 0 °C. The mixture was stirred at room temperature overnight. The organic layer was washed with water (2x), dried over Na₂SO₄ and concentrated to dryness under reduced pressure to obtain 1-(phenylsulfonyl)indoline (30.9 g, 96%). <chemistry id="chem0435" num="0435"><img file="EP2007756B1_D0461.tif" /></chemistry>

1-(1-(Phenylsulfonyl)indolin-5-yl)ethanone

0472To a suspension of AlCl₃ (144 g, 1.08 mol) in CH₂Cl₂ (1070 mL) was added acetic anhydride (54 mL). The mixture was stirred for 15 minutes before a solution of 1-(phenylsulfonyl)indoline (46.9 g, 0.180 mol) in CH₂Cl₂ (1070 mL) was added dropwise. The mixture was stirred for 5 h and was quenched by the slow addition of crushed ice. The organic layer was separated and the aqueous layer was extracted with CH₂Cl₂. The combined organics were washed with saturated aqueous NaHCO₃ and brine, dried over Na₂SO_4, and concentrated under <i>vacuum</i> to obtain 1-(1-(phenylsulfonyl)indolin-5-yl)ethanone (42.6 g). <chemistry id="chem0436" num="0436"><img file="EP2007756B1_D0462.tif" /></chemistry>

5-Ethyl-1-(phenylsulfonyl)indoline

0473To TFA (1600 mL) at 0 °C was added sodium borohydride (64.0 g, 1.69 mol) over 1 h. To this mixture was added dropwise a solution of 1-(1-(phenylsulfonyl)indolin-5-yl)ethanone (40.0 g, 0.133 mol) in TFA (700 mL) over 1 h. The mixture was then stirred overnight at 25 °C. After dilution with H₂O (1600 mL), the mixture was made basic by the addition of sodium hydroxide pellets at 0 °C. The organic layer was separated and the aqueous layer was extracted with CH₂Cl₂. The combined organic layers were washed with brine, dried over Na₂SO₄ and concentrated under reduced pressure. The residue was purified by silica column to give 5-ethyl-l-(phenylsulfonyl)indoline (16.2 g, 47% over two steps). <chemistry id="chem0437" num="0437"><img file="EP2007756B1_D0463.tif" /></chemistry>

5-Ethylindoline

0474A mixture of 5-ethyl-1-(phenylsulfonyl)indoline (15 g, 0.050 mol) in HBr (48%, 162 mL) was heated at reflux for 6 h. The mixture was basified with sat. NaOH to pH 9 and then it was extracted with ethyl acetate. The organic layer was washed with brine, dried over Na₂SO_4, and concentrated under reduced pressure. The residue was purified by silica column to give 5-ethylindoline (2.5 g, 32%). <chemistry id="chem0438" num="0438"><img file="EP2007756B1_D0464.tif" /></chemistry>

5-Ethyl-6-nitroindoline

0475To a solution of 5-ethylindoline (2.5 g, 17 mmol) in H₂SO₄ (98%, 20 mL) was slowly added KNO₃ (1.7 g, 17 mmol) at 0 °C. The mixture was stirred at 0 - 10 °C for 10 minutes. The mixture was then carefully poured into ice, basified with NaOH solution to pH 9, and extracted with ethyl acetate. The combined extracts were washed with brine, dried over Na₂SO₄ and concentrated to dryness. The residue was purified by silica column to give 5-ethyl-6-nitroindoline (1.9 g, 58%). <chemistry id="chem0439" num="0439"><img file="EP2007756B1_D0465.tif" /></chemistry>

5-Ethyl-6-nitro-1H-indole

0476To a solution of 5-ethyl-6-nitroindoline (1.9 g, 9.9 mmol) in CH₂Cl₂ (30 mL) was added MnO₂ (4.0 g, 46 mmol). The mixture was stirred at ambient temperature for 8 h. The solid was filtered off and the filtrate was concentrated to dryness to give 5-ethyl-6-nitro-1H-indole (1.9 g). <chemistry id="chem0440" num="0440"><img file="EP2007756B1_D0466.tif" /></chemistry>

5-Ethyl-1H-indol-6-amine

0477A suspension of 5-ethyl-6-nitro-1H-indole (1.9 g, 10 mmol) and Raney Nickel (1 g) was hydrogenated under H₂ (1 atm) at room temperature for 2 h. The catalyst was filtered off and the filtrate was concentrated to dryness. The residue was purified by silica gel column to give 5-ethyl-1H-indol-6-amine (760 mg, 48% over two steps). ¹H NMR (CDCl₃) δ 7.90 (br s, 1H), 7.41 (s, 1H), 7.00 (s, 1H), 6.78 (s, 2H), 6.39 (s, 1H), 3.39 (br s, 2H), 2.63 (q, <i>J</i> = 7.2 Hz, 2H), 1.29 (t, <i>J</i> = 6.9 Hz, 3H); MS (ESI) m/e (M+H⁺) 161.1.

Example 55: Ethyl 6-amino-1H-indole-4-carboxylate

0478<chemistry id="chem0441" num="0441"><img file="EP2007756B1_D0467.tif" /></chemistry><chemistry id="chem0442" num="0442"><img file="EP2007756B1_D0468.tif" /></chemistry>

2-Methyl-3,5-dinitrobenzoic acid

0479To a mixture of HNO₃ (95%, 80 mL) and H₂SO₄ (98%, 80 mL) was slowly added 2-methylbenzic acid (50 g, 0.37 mol) at 0 °C. After addition, the reaction mixture was stirred below 30 °C for 1.5 h. The mixture then was poured into ice-water and stirred for 15 min. The precipitate was filtered and washed with water to give 2-methyl-3,5-dinitrobenzoic acid (70 g, 84%). <chemistry id="chem0443" num="0443"><img file="EP2007756B1_D0469.tif" /></chemistry>

Ethyl 2-methyl-3,5-dinitrobenzoate

0480A mixture of 2-methyl-3,5-dinitrobenzoic acid (50 g, 0.22 mol) in SOCl₂ (80 mL) was heated at reflux for 4 h and then was concentrated to dryness. The residue was dissolved in CH₂Cl₂ (50 mL), to which EtOH (80 mL) was added and the mixture was stirred at room temperature for 1 h. The mixture was poured into ice-water and extracted with EtOAc (3 x 100 mL). The combined extracts were washed sat. Na₂CO₃ (80 mL), water (2 × 100 mL) and brine (100 mL), dried over Na₂SO₄ and concentrated to dryness to give ethyl 2-methyl-3,5-dinitrobenzoate (50 g, 88%) <chemistry id="chem0444" num="0444"><img file="EP2007756B1_D0470.tif" /></chemistry>

(<i>E</i>)-Ethyl 2-(2-(dimethylamino)vinyl)-3,5-dinitrobenzoate

0481A mixture of ethyl 2-methyl-3,5-dinitrobenzoate (35 g, 0.14 mol) and DMA (32 g, 0.27 mol) in DMF (200 mL) was heated at 100 °C for 5 h. The mixture was poured into ice-water and the precipitated solid was filtered and washed with water to give (<i>E</i>)-ethyl 2-(2-(dimethylamino)vinyl)-3,5-dinitrobenzoate (11 g, 48%) <chemistry id="chem0445" num="0445"><img file="EP2007756B1_D0471.tif" /></chemistry>

Ethyl 6-amino-1H-indole-4-carboxylate

0482A mixture of (<i>E</i>)-ethyl 2-(2-(dimethylamino)vinyl)-3,5-dinitrobenzoate (11 g, 0.037 mol) and SnCl₂ (83 g, 0.37 mol) in ethanol was heated at reflux for 4 h. The mixture was concentrated to dryness and the residue was poured into water and basified using sat. aq. Na₂CO₃ to pH 8. The precipitated solid was filtered and the filtrate was extracted with ethyl acetate (3 × 100 mL). The combined extracts were washed with water (2 × 100 mL) and brine (150 mL), dried over Na₂SO_4, and concentrated to dryness. The residue was purified by column on silica gel to give ethyl 6-amino-1H-indole-4-carboxylate (3.0 g, 40%). ¹HNMR (DMSO-<i>d</i>₆) δ 10.76 (br s, 1 H), 7.11-7.14 (m, 2 H), 6.81-6.82 (m, 1 H), 6.67-6.68 (m, 1 H), 4.94 (br s, 2 H), 4.32-4.25 (q, <i>J =</i> 7.2 Hz, 2 H), 1.35-1.31 (t, <i>J=</i> 7.2, 3 H); MS (ESI) m/e (M+H⁺) 205.0.

Example 56: 5-Fluoro-1H-indol-6-amine

0483<chemistry id="chem0446" num="0446"><img file="EP2007756B1_D0472.tif" /></chemistry><chemistry id="chem0447" num="0447"><img file="EP2007756B1_D0473.tif" /></chemistry>

1-Fluoro-5-methyl-2,4-dinitrobenzene

0484To a stirred solution of HNO₃ (60 mL) and H₂SO₄ (80 mL) was added dropwise 1-fluoro-3-methylbenzene (28 g, 25 mmol) under ice-cooling at such a rate that the temperature did not rise above 35 °C. The mixture was allowed to stir for 30 min at rt and was then poured into ice water (500 mL). The resulting precipitate (a mixture of 1-fluoro-5-methyl-2,4-dinitrobenzene and 1-fluoro-3-methyl-2,4-dinitrobenzene, 32 g, ca. 7:3 ratio) was collected by filtration and purified by recrystallization from 50 mL isopropyl ether to give pure 1-fluoro-5-methyl-2,4-dinitro-benzene as a white solid (18 g, 36%). <chemistry id="chem0448" num="0448"><img file="EP2007756B1_D0474.tif" /></chemistry>

(<i>E</i>)-2-(5-Fluoro-2,4-dinitrophenyl)-<i>N-N</i>-dimethylethenamine

0485A mixture of 1-fluoro-5-methyl-2,4-dinitro-benzene (10 g, 50 mmol), DMA (12 g, 100 mmol) and DMF (50 mL) was heated at 100 °C for 4h. The solution was cooled and poured into water. The precipitated red solid was collected, washed with water, and dried to give (<i>E</i>)-2-(5-fluoro-2,4-dinitrophenyl)-<i>N,N</i>-dimethylethenamine (8.0 g, 63%). <chemistry id="chem0449" num="0449"><img file="EP2007756B1_D0475.tif" /></chemistry>

5-Fluoro-1H-indol-6-amine

0486A suspension of (<i>E</i>)-2-(5-fluoro-2,4-dinitrophenyl)-<i>N,N</i>-dimethylethenamine (8.0 g, 31 mmol) and Raney Nickel (8 g) in EtOH (80 mL) was stirred under H₂ (40 psi) at room temperature for 1 h. After filtration, the filtrate was concentrated and the residue was purified by column chromatography (petroleum ether/ethyl acetate = 5/1) to give 5-fluoro-1H-indol-6-amine (1.0 g, 16%) as a brown solid. ¹HNMR (DMSO-<i>d</i>₆) δ 10.56 (br s, 1 H), 7.07 (d, <i>J</i> = 12 Hz, 1 H), 7.02 (m, 1H), 6.71 (d, <i>J</i> = 8 Hz, 1H), 6.17 (s, 1H), 3.91 (br s , 2H); MS (ESI) m/e (M+H⁺) 150.1.

Example 57: 5-Chloro-1H-indol-6-amine

0487<chemistry id="chem0450" num="0450"><img file="EP2007756B1_D0476.tif" /></chemistry><chemistry id="chem0451" num="0451"><img file="EP2007756B1_D0477.tif" /></chemistry>

.1-Chloro-5-methyl-2,4-dinitrobenzene

0488To a stirred solution of HNO₃ (55 mL) and H₂SO₄ (79 mL) was added dropwise 1-chloro-3-methylbenzene (25.3 g, 200 mmol) under ice-cooling at such a rate that the temperature did not rise above 35 °C. The mixture was allowed to stir for 30 min at ambient temperature and was then poured into ice water (500 mL). The resulting precipitate was collected by filtration and purified by recrystallization to give 1-chloro-5-methyl-2,4-dinitrobenzene (26 g, 60%). <chemistry id="chem0452" num="0452"><img file="EP2007756B1_D0478.tif" /></chemistry>

(<i>E</i>)-2-(5-Chloro-2,4-dinitrophenyl)-<i>N,N</i>-dimethylethenamine

0489A mixture of 1-chloro-5-methyl-2,4-dinitro-benzene (11.6 g, 50.0 mmol), DMA (11.9 g, 100 mmol) in DMF (50 mL) was heated at 100 °C for 4 h. The solution was cooled and poured into water. The precipitated red solid was collected by filtration, washed with water, and dried to give (<i>E</i>)-2-(5-chloro-2,4-dinitrophenyl)-<i>N,N</i> dimethylethenamine (9.84 g, 72%). <chemistry id="chem0453" num="0453"><img file="EP2007756B1_D0479.tif" /></chemistry>

5-Chloro-1H-indol-6-amine

0490A suspension of (<i>E</i>)-2-(5-chloro-2,4-dinitrophenyt)-<i>N,N</i>-dimethylethenamine (9.8 g, 36 mmol) and Raney Nickel (9.8 g) in EtOH (140 mL) was stirred under H₂ (1 atm) at room temperature for 4 h. After filtration, the filtrate was concentrated and the residue was purified by column chromatograph (petroleum ether/ethyl acetate = 10:1) to give 5-chloro-1H-indol-6-amine (0.97 g, 16%) as a gray powder. ¹HNMR (CDCl₃) δ 7.85 (br s, 1 H), 7.52 (s, 1 H), 7.03 (s, 1H), 6.79 (s, 1H), 6.34 (s, 1H), 3.91 (br s, 1H); MS (ESI) m/e (M+H⁺) 166.0.

Example 58: Ethyl 6-amino-1H-indole-7-carboxylate

0491<chemistry id="chem0454" num="0454"><img file="EP2007756B1_D0480.tif" /></chemistry><chemistry id="chem0455" num="0455"><img file="EP2007756B1_D0481.tif" /></chemistry>

3-Methyl-2,6-dinitrobenzoic acid

0492To a mixture of HNO₃ (95%, 80 mL) and H₂SO₄ (98%, 80 mL) was slowly added 3-methylbenzic acid (50 g, 0.37 mol) at 0 °C. After addition, the mixture was stirred below 30 °C for 1.5 hours. The mixture was then poured into ice-water and stirred for 15 min. The precipitate solid was filtered and washed with water to give a mixture of 3-methyl-2,6-dinitro-benzoic acid and 5-methyl-2,4-dinitrobenzoic acid (70 g, 84%). To a solution of this mixture (70 g, 0.31 mol) in EtOH (150 mL) was added dropwise SOCl₂ (54 g, 0.45 mol). The mixture was heated at reflux for 2 h before being concentrated to dryness under reduced pressure. The residue was partitioned between EtOAc (100 mL) and aq. Na₂CO₃ (10%, 120 mL). The organic layer was washed with brine (50 mL), dried over Na₂SO_4, and concentrated to dryness to obtain ethyl 5-methyl-2,4-dinitrobenzoate (20 g), which was placed aside. The aqueous layer was acidified by HCl to pH 2 ∼ 3 and the precipitated solid was filtered, washed with water, and dried in air to give 3-methyl-2,6-dinitrobenzoic acid (39 g, 47%). <chemistry id="chem0456" num="0456"><img file="EP2007756B1_D0482.tif" /></chemistry>

Ethyl 3-methyl-2,6-dinitrobenzoate

0493A mixture of 3-methyl-2,6-dinitrobenzoic acid (39 g, 0.15 mol) and SOCl₂ (80 mL) was heated at reflux 4 h. The excess SOCl₂ was evaporated off under reduced pressure and the residue was added dropwise to a solution of EtOH (100 mL) and Et₃N (50 mL). The mixture was stirred at 20 °C for 1 h and then concentrated to dryness. The residue was dissolved in EtOAc (100 mL), washed with Na₂CO₃ (10 %, 40 mL × 2), water (50 mL × 2) and brine (50 mL), dried over Na₂SO₄ and concentrated to give ethyl 3-methyl-2,6-dinitrobenzoate (20 g, 53%). <chemistry id="chem0457" num="0457"><img file="EP2007756B1_D0483.tif" /></chemistry>

(<i>E</i>)-Ethyl 3-(2-(dimethylamino)vinyl)-2,6-dinitrobenzoate

0494A mixture of ethyl 3-methyl-2,6-dinitrobenzoate (35 g, 0.14 mol) and DMA (32 g, 0.27 mol) in DMF (200 mL) was heated at 100 °C for 5 h. The mixture was poured into ice water. The precipitated solid was filtered and washed with water to give (<i>E</i>)-ethyl 3-(2-(dimethylamino)vinyl)-2,6-dinitrobenzoate (25 g, 58%). <chemistry id="chem0458" num="0458"><img file="EP2007756B1_D0484.tif" /></chemistry>

Ethyl 6-amino-1H-indole-7-carboxylate

0495A mixture of (<i>E</i>)-ethyl 3-(2-(dimethylamino)vinyl)-2,6-dinitrobenzoate (30 g, 0.097 mol) and Raney Nickel (10 g) in EtOH (1000 mL) was hydrogenated at room temperature under 50 psi for 2 h. The catalyst was filtered off and the filtrate was concentrated to dryness. The residue was purified by column on silica gel to give ethyl 6-amino-1H-indole-7-carboxylate as an off-white solid (3.2 g, 16%). ¹H NMR (DMSO-<i>d</i>₆) δ 10.38 (s, 1 H), 7.42 (d, <i>J =</i> 8.7 Hz, 1 H), 6.98 (t, <i>J</i> = 3.0 Hz, 1 H), 6.65 (s, 2 H), 6.48 (d, <i>J =</i> 8.7 Hz, 1 H), 6.27-6.26 (m, 1 H), 4.38 (q, <i>J=</i> 7.2 Hz, 2 H), 1.35 (t, <i>J=</i> 7.2 Hz, 3 H).

Example 59: Ethyl 6-amino-1H-indole-5-carboxylate

0496<chemistry id="chem0459" num="0459"><img file="EP2007756B1_D0485.tif" /></chemistry><chemistry id="chem0460" num="0460"><img file="EP2007756B1_D0486.tif" /></chemistry>

(<i>E</i>)-Ethyl 5-(2-(dimethylamino)vinyl)-2,4-dinitrobenzoate

0497A mixture of ethyl 5-methyl-2,4-dinitrobenzoate (39 g, 0.15 mol) and DMA (32 g, 0.27 mol) in DMF (200 mL) was heated at 100 °C for 5 h. The mixture was poured into ice water and the precipitated solid was filtered and washed with water to afford (<i>E</i>)-ethyl 5-(2-(dimethylamino)vinyl)-2,4-dinitrobenzoate (15 g, 28%). <chemistry id="chem0461" num="0461"><img file="EP2007756B1_D0487.tif" /></chemistry>

Ethyl 6-amino-1H-indole-5-carboxylate

0498A mixture of (<i>E</i>)-ethyl 5-(2-(dimethylamino)vinyl)-2,4-dinitrobenzoate (15 g, 0.050 mol) and Raney Nickel (5 g) in EtOH (500 mL) was hydrogenated at room temperature under 50 psi of hydrogen for 2 h. The catalyst was filtered off and the filtrate was concentrated to dryness. The residue was purified by column on silica gel to give ethyl 6-amino-1H-indole-5-carboxylate (3.0 g, 30%). ¹H NMR (DMSO-<i>d</i>₆) δ 10.68 (s, 1 H), 7.99 (s, 1 H), 7.01-7.06 (m, 1 H), 6.62 (s, 1 H), 6.27-6.28 (m, 1 H), 6.16 (s, 2 H), 4.22 (q, <i>J=</i> 7.2 Hz, 2 H), 1.32-1.27 (t, <i>J</i> = 7.2 Hz, 3 H).

Example 60

:

5-<i>tert</i>-Butyl-1H-indol-6-amine

0499<chemistry id="chem0462" num="0462"><img file="EP2007756B1_D0488.tif" /></chemistry><chemistry id="chem0463" num="0463"><img file="EP2007756B1_D0489.tif" /></chemistry>

2-<i>tert</i>-Butyl-4-methylphenyl diethyl phosphate

0500To a suspension of NaH (60% in mineral oil, 8.4 g, 0.21 mol) in THF (200 mL) was added dropwise a solution of 2-<i>tert</i>-butyl-4-methylphenol (33 g, 0.20 mol) in THF (100 mL) at 0 °C. The mixture was stirred at 0 °C for 15 min and then phosphorochloridic acid diethyl ester (37 g, 0.21 mol) was added dropwise at 0 °C. After addition, the mixture was stirred at ambient temperature for 30 min. The reaction was quenched with sat. NH₄Cl (300 mL) and then extracted with Et₂O (350 mL × 2). The combined organic layers were washed with brine, dried over anhydrous Na₂SO_4, and then evaporated under <i>vacuum</i> to give 2<i>-tert-</i>butyl-4-methylphenyl diethyl phosphate (contaminated with mineral oil) as a colorless oil (60 g, ∼100%), which was used directly in the next step. <chemistry id="chem0464" num="0464"><img file="EP2007756B1_D0490.tif" /></chemistry>

1-<i>tert</i>-Butyl-3-methylbenzene

0501To NH₃ (liquid, 1000 mL) was added a solution of 2-<i>tert</i>-butyl-4-methylphenyl diethyl phosphate (60 g, crude from last step, about 0.2 mol) in Et₂O (anhydrous, 500 mL) at -78 °C under N₂ atmosphere. Lithium metal was added to the solution in small pieces until the blue color persisted. The reaction mixture was stirred at -78 °C for 15 min and then was quenched with sat. NH₄Cl until the mixture turned colorless. Liquid NH₃ was evaporated and the residue was dissolved in water. The mixture was extracted with Et₂O (400 mL × 2). The combined organics were dried over Na₂SO₄ and evaporated to give 1-<i>tert</i>-butyl-3-methylbenzene (contaminated with mineral oil) as a colorless oil (27 g, 91%), which was used directly in next step. <chemistry id="chem0465" num="0465"><img file="EP2007756B1_D0491.tif" /></chemistry>

1-<i>tert</i>-Butyl-5-methyl-2,4-dinitrobenzene and 1-<i>tert</i>-butyl-3-methyl-2,4-dinitro-benzene

.

0502To HNO₃ (95%, 14 mL) was added H₂SO₄ (98 %, 20 mL) at 0 °C and then 1<i>-tert-</i>butyl-3-methylbenzene (7.4 g, ∼50 mmol, crude from last step) dropwise to the with the temperature being kept below 30 °C. The mixture was stirred at ambient temperature for 30 min, poured onto crushed ice (100 g), and extracted with EtOAc (50 mL × 3). The combined organic layers were washed with water and brine, before being evaporated to give a brown oil, which was purified by column chromatography to give a mixture of 1-<i>tert</i>-butyl-5-methyl-2,4-dinitrobenzene and 1-<i>tert</i>-butyl-3-methyl-2,4-dinitrobenzene (2:1 by NMR) as a yellow oil (9.0 g, 61%). <chemistry id="chem0466" num="0466"><img file="EP2007756B1_D0492.tif" /></chemistry>

<i>(E)-</i>2-(5<i>-tert-</i>Butyl<i>-</i> 2,4-dinitrophenyl)-<i>N,N</i>-dimethylethenamine

0503A mixture of 1-<i>tert</i>-butyl-5-methyl-2,4-dinitrobenzene and 1-<i>tert</i>-butyl-3-methyl-2,4-dinitrobenzene (9.0 g, 38 mmol, 2:1 by NMR) and DMA (5.4 g, 45 mmol) in DMF (50 mL) was heated at reflux for 2 h before being cooled to room temperature. The reaction mixture was poured into water-ice and extracted with EtOAc (50 mL × 3). The combined organic layers were washed with water and brine, before being evaporated to give a brown oil, which was purified by column to give (<i>E</i>)-2-(5-<i>tert</i>-butyl-2,4-dinitrophenyl)-<i>N,N-</i>dimethylethen-amine (5.0 g, 68%). <chemistry id="chem0467" num="0467"><img file="EP2007756B1_D0493.tif" /></chemistry>

5-<i>tert-</i>Butyl-1H-indol-6-amine

0504A solution of (<i>E</i>)-2-(5-<i>tert</i>-butyl-2,4-dinitrophenyl)-<i>N,N</i>-dimethylethen-amine (5.3 g, 18 mmol) and tin (II) chloride dihydrate (37 g, 0.18 mol) in ethanol (200 mL) was heated at reflux overnight. The mixture was cooled to room temperature and the solvent was removed under <i>vacuum.</i> The residual slurry was diluted with water (500 mL) and was basifed with 10 % aq. Na₂CO₃ to pH 8. The resulting suspension was extracted with ethyl acetate (3 × 100 mL). The ethyl acetate extract was washed with water and brine, dried over Na₂SO_4, and concentrated. The residual solid was washed with CH₂Cl₂ to afford a yellow powder, which was purified by column chromatography to give 5-<i>tert</i>-butyl-1H-indol-6-amine (0.40 g, 12%). ¹H NMR (DMSO_<i>d</i>₆) δ 10.34 (br s, 1 H), 7.23 (s, 1 H), 6.92 (s, 1 H), 6.65 (s, 1H), 6.14 (s, 1 H), 4.43 (br s, 2 H), 2.48 (s, 9 H); MS (ESI) m/e (M+H⁺) 189.1.

General Procedure IV: Synthesis of acylaminoindoles

0505<chemistry id="chem0468" num="0468"><img file="EP2007756B1_D0494.tif" /></chemistry>

0506One equivalent of the appropriate carboxylic acid and one equivalent of the appropriate amine were dissolved in <i>N,N</i>-dimethylformamide (DMF) containing triethylamine (3 equivalents). <i>O</i>-(7-Azabenzotriazol-1-yl)-<i>N,N,N',N'</i>-tetramethyluronium hexafluorophosphate (HATU) was added and the solution was allowed to stir. The crude product was purified by reverse-phase preparative liquid chromatography to yield the pure product.

Example 61:

N-

<i>(</i>2<i>-tert-</i> Butyl-1H-indol-5-yl)-1-(4-methoxyphenyl)-cyclopropanecarboxamide

0507<chemistry id="chem0469" num="0469"><img file="EP2007756B1_D0495.tif" /></chemistry>

05082-<i>tert</i>-Butyl-1H-indol-5-amine (19 mg, 0.10 mmol) and 1-(4-methoxyphenyl)-cyclopropanecarboxylic acid (19 mg, 0.10 mmol) were dissolved in <i>N,N-</i>dimethylformamide (1.00 mL) containing triethylamine (28 µL, 0.20 mmol). <i>O</i>-(7-Azabenzotriazol-1-yl)-<i>N,N,N'N</i>'-tetramethyluronium hexafluorophosphate (42 mg, 0.11 mmol) was added to the mixture and the resulting solution was allowed to stir for 3 hours. The crude reaction mixture was filtered and purified by reverse phase HPLC. ESI-MS m/z calc. 362.2, found 363.3 (M+1)⁺; Retention time 3.48 minutes.

General Procedure V: Synthesis of acylaminoindoles

0509<chemistry id="chem0470" num="0470"><img file="EP2007756B1_D0496.tif" /></chemistry>

0510One equivalent of the appropriate carboxylic acid was placed in an oven-dried flask under nitrogen. A minimum (3 equivalents) of thionyl chloride and a catalytic amount of and <i>N,N-</i>dimethylformamide were added and the solution was allowed to stir for 20 minutes at 60 °C. The excess thionyl chloride was removed under vacuum and the resulting solid was suspended in a minimum of anhydrous pyridine. This solution was slowly added to a stirred solution of one equivalent the appropriate amine dissolved in a minimum of anhydrous pyridine. The resulting mixture was allowed to stir for 15 hours at 110°C. The mixture was evaporated to dryness, suspended in dichloromethane, and then extracted three times with 1N HCl. The organic layer was then dried over sodium sulfate, evaporated to dryness, and then purified by column chromatography.

Example 62: Ethyl 5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indole-2-carboxylate (Compd. 28)

0511<chemistry id="chem0471" num="0471"><img file="EP2007756B1_D0497.tif" /></chemistry>

05121-Benzo[1,3]dioxol-5-yl-cyclopropanecarboxylic acid (2.07 g, 10.0 mmol) was dissolved in thionyl chloride (2.2 mL) under N₂. <i>N,N-</i>dimethylformamide (0.3 mL) was added and the solution was allowed to stir for 30 minutes. The excess thionyl chloride was removed under vacuum and the resulting solid was dissolved in anhydrous dichloromethane (15 mL) containing triethylamine (2.8 mL, 20.0 mmol). Ethyl 5-amino-1H-indole-2-carboxylate (2.04 g, 10.0 mmol) in 15 mL of anhydrous dichloromethane was slowly added to the reaction. The resulting solution was allowed to stir for 1 hour. The reaction mixture was diluted to 50 mL with dichloromethane and washed three times with 50 mL of 1N HCl, saturated aqueous sodium bicarbonate, and saturated aqueous sodium chloride. The organic layer was dried over sodium sulfate and evaporated to dryness to yield ethyl 5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indole-2-carboxylate as a gray solid (3.44 g, 88 %). ESI-MS <i>m</i>/<i>z</i> calc. 392.4; found 393.1 (M+1)⁺ Retention time 3.17 minutes. ¹H NMR (400 MHz, DMSO-d6) δ 11.80 (s, 1H), 8.64 (s, 1H), 7.83 (m, 1H), 7.33-7.26 (m, 2H), 7.07 (m, 1H), 7.02 (m, 1H), 6.96-6.89 (m, 2H), 6.02 (s, 2H), 4.33 (q, <i>J =</i> 7.1 Hz, 2H), 1.42-1.39 (m, 2H), 1.33 (t, <i>J=</i> 7.1 Hz, 3H), 1.06-1.03 (m, 2H).

Example 63: 1-(Benzo[<i>d</i>][1,3]dioxol-5-yl)-N-(2-<i>tert-</i>butyl-1<i>H-</i>indol-5-yl)cyclopropanecarboxamide

0513<chemistry id="chem0472" num="0472"><img file="EP2007756B1_D0498.tif" /></chemistry>

05141-Benzo[1,3]dioxol-5-yl-cyclopropanecarboxylic acid (1.09 g, 5.30 mmol) was dissolved in 2 mL of thionyl chloride under nitrogen. A catalytic amount (0.3 mL) <i>of N,N-</i>dimethylformamide (DMF) was added and the reaction mixture was stirred for 30 minutes. The excess thionyl chloride was evaporated and the resulting residue was dissolved in 15 mL of dichloromethane. This solution was slowly added to a solution of 2-<i>tert</i>-butyl-1<i>H</i>-indol-5-amine (1.0 g, 5.3 mmol) in 10 mL of dichloromethane containing triethylamine (1.69 mL, 12.1 mmol). The resulting solution was allowed to stir for 10 minutes. The solvent was evaporated to dryness and the crude reaction mixture was purified by silica gel column chromatography using a gradient of 5-50 % ethyl acetate in hexanes. The pure fractions were combined and evaporated to dryness to yield a pale pink powder (1.24 g 62%). ESI-MS <i>m</i>/z calc. 376.18, found 377.3 (M+1)⁺. Retention time of 3.47 minutes. ¹H NMR (400 MHz, DMSO) δ 10.77 (s, 1H), 8.39 (s, 1H), 7.56 (d, <i>J=</i> 1.4 Hz, 1H), 7.15 (d, <i>J=</i> 8.6 Hz, 1H), 7.05 - 6.87 (m, 4H), 6.03 (s, 3H), 1.44 - 1.37 (m, 2H), 1.33 (s, 9H), 1.05-1.00 (m, 2H).

Example 64: 1-(Benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N-</i>(1-methyl-2-(1-methylcyclopropyl)-1<i>H</i>-indol-5-yl)cyclopropanecarboxamide

0515<chemistry id="chem0473" num="0473"><img file="EP2007756B1_D0499.tif" /></chemistry>

05161-Methyl-2-(1-methylcyclopropyl)-1<i>H</i>-indol-5-amine (20.0 mg, 0.100 mmol) and 1-(benzo[<i>d</i>][1,3]dioxol-5-yl)cyclopropanecarboxylic acid (20.6 mg, 0.100 mmol) were dissolved in <i>N,N-</i> dimethylformamide (1 mL) containing triethylamine (42.1 µL, 0.300 mmol) and a magnetic stir bar. <i>O</i>-(7-Azabenzotriazol-1-yl)-<i>N,N,N,N'</i>-tetramethyluronium hexafluorophosphate (42 mg, 0.11 mmol) was added to the mixture and the resulting solution was allowed to stir for 6 h at 80 °C. The crude product was then purified by preparative HPLC utilizing a gradient of 0-99% acetonitrile in water containing 0.05% trifluoroacetic acid to yield 1-(benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N-</i>(1-methyl-2-(1-methylcyclopropyl)-1<i>H-</i>indol-5-yl)cyclopropanecarboxamide. ESI-MS <i>m</i>/z calc. 388.2, found 389.2 (M+1)⁺. Retention time of 3.05 minutes.

Example 65: 1-(Benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N</i>-(1,1-dimethyl-2,3-dihydro-1<i>H-</i>pyrrolo[1,2-a]indol-7-yl)cyclopropanecarboxamide

0517<chemistry id="chem0474" num="0474"><img file="EP2007756B1_D0500.tif" /></chemistry>

05181,1-Dimethyl-2,3-dihydro-1<i>H</i>-pyrrolo [1,2-<i>α</i>]indol-7-amine (40.0 mg, 0.200 mmol) and 1-(benzo[<i>d</i>][1,3]dioxol-5-yl)cyclopropanecarboxylic acid (41.2 mg, 0.200 mmol) were dissolved in <i>N,N-</i>dimethylformamide (1 mL) containing triethylamine (84.2 µL, 0.600 mmol) and a magnetic stir bar. <i>O</i>-(7-Azabenzotriazol-1-yl)-<i>N,N,N',N</i>'-tetramethyluronium hexafluorophosphate (84 mg, 0.22 mmol) was added to the mixture and the resulting solution was allowed to stir for 5 minutes at room temperature. The crude product was then purified by preparative HPLC utilizing a gradient of 0-99% acetonitrile in water containing 0.05% trifluoroacetic acid to yield 1-(benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N</i>-(1,1-dimethyl-2,3-dihydro-1<i>H-</i>pyrrolo[1,2-a]-indol-7-yl)cyclopropanecarboxamide. ESI-MS <i>m</i>/z calc. 388.2, found 389.2 (M+1)⁺. Retention time of 2.02 minutes. ¹H NMR (400 MHz, DMSO-<i>d</i>₆) δ 8.41 (s, 1H), 7.59 (d, <i>J=</i> 1.8 Hz, 1H), 7.15 (d, <i>J=</i> 8.6 Hz, 1H), 7.06 - 7.02 (m, 2H), 6.96 - 6.90 (m, 2H), 6.03 (s, 2H), 5.98 (d, <i>J=</i> 0.7 Hz, 1H), 4.06 (t, <i>J=</i> 6.8 Hz, 2H), 2.35 (t, <i>J =</i> 6.8 Hz, 2H), 1.42-1.38 (m, 2H), 1.34 (s, 6H), 1.05-1.01 (m, 2H).

Example 66: Methyl 5-(1-(benzo[<i>d</i>][1,3]dioxol-5-yl)cyclopropanecarboxamido)-2-<i>tert</i> butyl-1<i>H</i> indole-7-carboxylate

0519<chemistry id="chem0475" num="0475"><img file="EP2007756B1_D0501.tif" /></chemistry>

05201-(Benzo[<i>d</i>][1,3]dioxol-5-yl)cyclopropanecarbonyl chloride (45 mg, 0.20 mmol) and methyl 5-amino-2-<i>tert</i>-butyl-1<i>H</i>-indole-7-carboxylate (49.3 mg, 0.200 mmol) were dissolved in <i>N,N-</i> dimethylformamide (2 mL) containing a magnetic stir bar and triethylamine (0.084 mL, 0.60 mmol). The resulting solution was allowed to stir for 10 minutes at room temperature. The crude product was then purified by preparative HPLC using a gradient of 0-99% acetonitrile in water containing 0.05% trifluoroacetic acid to yield methyl 5-(1-(benzo[<i>d</i>][1,3]dioxol-5-yl)cyclopropanecarbox-amido)-2-<i>tert</i>-butyl-1<i>H</i> indole-7-carboxylate. ESI-MS <i>m</i>/<i>z</i> calc. 434.2, found 435.5. (M+1)⁺. Retention time of 2.12 minutes.

Example 67: 1-(Benzo[d)[1,3]dioxol-5-yl)-<i>N</i>-(2-(1-hydroxy-2-methylpropan-2-yl)-1H-indol-5-yl)cyclopropanecarboxamide

0521<chemistry id="chem0476" num="0476"><img file="EP2007756B1_D0502.tif" /></chemistry>

0522To a solution of 1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylic acid (0.075 g, 0.36 mmol) in acetonitrile (1.5 mL) were added HBTU (0.138 g, 0.36 mmol) and Et₃N (152 µL, 1.09 mmol) at room temperature. The mixture was stirred at room temperature for 10 minutes before a solution of 2-(5-amino-1H-indol-2-yl)-2-methylpropan-1-ol (0.074 g, 0.36 mmol) in acetonitrile (1.94 mL) was added. After addition, the reaction mixture was stirred at room temperature for 3 h. The solvent was evaporated under reduced pressure and the residue was dissolved in dichloromethane. The organic layer was washed with 1 N HCl (1 x 3 mL) and saturated aqueous NaHCO₃ (1 x 3 mL). The organic layer was dried over Na₂SO_4, filtered and evaporated under reduced pressure. The crude material was purified by column chromatography on silica gel (ethyl acetate/hexane = 1/1) to give 1-(benzo[d][1,3]dioxol-5-yl)-<i>N-</i>(2-(1-hydroxy-2-methylpropan-2-yl)-1H-indol-5-yl)cyclopropanecarboxamide (0.11 g, 75%). ¹H NMR (400 MHz, DMSO-d6) δ 10.64 (s, 1H), 8.38 (s, 1H), 7.55 (s, 1H), 7.15 (d, J = 8.6 Hz, 1H), 7.04-6.90 (m, 4H), 6.06 (s, 1H), 6.03 (s, 2H), 4.79 (t, J = 2.7 Hz, 1H), 3.46 (d, J = 0.0 Hz, 2H), 1.41-1.39 (m, 2H), 1.26 (s, 6H), 1.05-1.02 (m, 2H).

Example 67: 1-(Benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N</i>-(2,3,4,9-tetrahydro-1<i>H-</i>carbazol-6-yl)cyclopropanecarboxamide

0523<chemistry id="chem0477" num="0477"><img file="EP2007756B1_D0503.tif" /></chemistry>

05242,3,4,9-Tetrahydro-1<i>H</i>-carbazol-6-amine (81.8 mg, 0.439 mmol) and 1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylic acid (90.4 mg, 0.439 mmol) were dissolved in acetonitrile (3 mL) containing diisopropylethylamine (0.230 mL, 1.32 mmol) and a magnetic stir bar. <i>O</i>-(7-Azabenzotriazol-1-yl)-<i>N,N,N;N'</i>-tetramethyluronium hexafluorophosphate (183 mg, 0.482 mmol) was added to the mixture and the resulting solution was allowed to stir for 16 h at 70 °C. The solvent was evaporated and the crude product was then purified on 40 g of silica gel utilizing a gradient of 5-50% ethyl acetate in hexanes to yield 1-(benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N</i>-(2,3,4,9-tetrahydro-1<i>H</i>-carbazol-6-yl)cyclopropanecarboxamide as a beige powder (0.115 g, 70%) after drying. ESI-MS <i>m</i>/z calc. 374.2, found 375.3 (M+1)⁺. Retention time of 3.43 minutes. ¹H NMR (400 MHz, DMSO-<i>d</i>₆) δ 10.52 (s, 1H), 8.39 (s, 1H), 7.46 (d, <i>J=</i> 1.8 Hz, 1H), 7.10 - 6.89 (m, 5H), 6.03 (s, 2H), 2.68 - 2.65 (m, 2H), 2.56 - 2.54 (m, 2H), 1.82 - 1.77 (m, 4H), 1.41 - 1.34 (m, 2H), 1.04 - 0.97 (m, 2H).

Example 69: <i>tert</i>-Butyl 4-(5-(1-(benzo[<i>d</i>][1,3]dioxol-5-yl)cyclopropanecarbox-amido)-1<i>H</i>-indol-2-yl)piperidine-1-carboxylate

0525<chemistry id="chem0478" num="0478"><img file="EP2007756B1_D0504.tif" /></chemistry>

05261-(Benzo[<i>d</i>][1,3]dioxol-5-yl)cyclopropanecarbonyl chloride (43 mg, 0.19 mmol) and <i>tert</i>-butyl 4-(5-amino-1<i>H</i>-indol-2-yl)piperidine-1-carboxylate (60 mg, 0.19 mmol) were dissolved in dichloromethane (1 mL) containing a magnetic stir bar and triethylamine (0.056 mL, 0.40 mmol). The resulting solution was allowed to stir for two days at room temperature. The crude product was then evaporated to dryness, dissolved in a minimum of <i>N,N</i>-dimethylformamide, and then purified by preparative HPLC using a gradient of 0-99% acetonitrile in water containing 0.05% trifluoroacetic acid to yield <i>tert</i>-butyl 4-(5-(1-(benzo[<i>d</i>][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1<i>H</i>-indol-2-yl)piperidine-1-carboxylate. ESI-MS <i>m</i>/z calc. 503.2, found 504.5. (M+1)⁺. Retention time of 1.99 minutes.

Example 70: Ethyl 2-(5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-2-yl)propanoate

0527<chemistry id="chem0479" num="0479"><img file="EP2007756B1_D0505.tif" /></chemistry><chemistry id="chem0480" num="0480"><img file="EP2007756B1_D0506.tif" /></chemistry>

<i>tert</i>-Butyl 2-(1-ethoxy-1-oxopropan-2-yl)-1H-indole-1-carboxylate

0528<i>tert</i>-Butyl 2-(2-ethoxy-2-oxoethyl)-1H-indole-1-carboxylate (3.0 g, 9.9 mmol) was added to anhydrous THF (29 mL) and cooled to -78 °C. A 0.5M solution of potassium hexamethyldisilazane (20 mL, 9.9 mmol) was added slowly such that the internal temperature stayed below -60 °C. Stirring was continued for 1 h at -78 °C. Methyl iodide (727 µL, 11.7 mmol) was added to the mixture. The mixture was stirred for 30 minutes at room temperature. The mixture was quenched with sat. aq. ammonium chloride and partitioned between water and dichloromethane. The aqueous phase was extracted with dichloromethane and the combined organic phases were dried over Na₂SO₄ and evaporated under reduced pressure. The residue was purified by column chromatography on silica gel (ethylacetate/hexane = 1/9) to give <i>tert</i>-butyl 2-(1-ethoxy-1-oxopropan-2-yl)-1H-indole-1-carboxylate (2.8 g, 88%). <chemistry id="chem0481" num="0481"><img file="EP2007756B1_D0507.tif" /></chemistry>

Ethyl 2-(1H-indol-2-yl)propanoate

0529<i>tert</i>-Butyl 2-(1-ethoxy-1-oxopropan-2-yl)-1H-indole-1-carboxylate (2.77 g, 8.74 mmol) was dissolved in dichloromethane (25 mL) before TFA (9.8 mL) was added. The mixture was stirred for 1.5 h at room temperature. The mixture was evaporated to dryness, taken up in dichloromethane and washed with sat. aq. sodium bicarbonate, water, and brine. The product was purified by column chromatography on silica gel (0-20% EtOAc in hexane) to give ethyl 2-(1H-indol-2-yl)propanoate (0.92 g, 50%). <chemistry id="chem0482" num="0482"><img file="EP2007756B1_D0508.tif" /></chemistry>

Ethyl 2-(5-nitro-1H-indol-2-yl)propanoate

0530Ethyl 2-(1H-indol-2-yl)propanoate (0.91 g, 4.2 mmol) was dissolved in concentrated sulfuric acid (3.9 mL) and cooled to -10°C. (salt/ice-mixture). A solution of sodium nitrate (0.36 g, 4.2 mmol) in concentrated sulfuric acid (7.8 mL) was added dropwise over 35 min. Stirring was continued for another 30 min at -10 °C. The mixture was poured into ice and the product was extracted with ethyl acetate. The combined organic phases were washed with a small amount of sat. aq. sodium bicarbonate. The product was purified by column chromatography on silica gel (5-30% EtOAc in hexane) to give ethyl 2-(5-nitro-1H-indol-2-yl)propanoate (0.34 g, 31%). <chemistry id="chem0483" num="0483"><img file="EP2007756B1_D0509.tif" /></chemistry>

Ethyl 2-(5-amino-1H-indol-2-yl)propanoate

0531To a solution of ethyl 2-(5-nitro-1H-indol-2-yl)propanoate (0.10 g, 0.38 mmol) in ethanol (4 mL) was added tin chloride dihydrate (0.431 g, 1.91 mmol). The mixture was heated in the microwave at 120 °C for 1 h. The mixture was diluted with ethyl acetate before water and saturated aqueous NaHCO₃ were added. The reaction mixture was filtered through a plug of celite using ethyl acetate. The organic layer was separated from the aqueous layer. The organic layer was dried over Na₂SO₄, filtered and evaporated under reduced pressure to give ethyl 2-(5-amino-1H-indol-2-yl)propanoate (0.088 g, 99%). <chemistry id="chem0484" num="0484"><img file="EP2007756B1_D0510.tif" /></chemistry>

Ethyl 2-(5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-2-yl)propanoate

0532To a solution of 1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylic acid (0.079 g, 0.384 mmol) in acetonitrile (1.5 mL) were added HBTU (0.146 g, 0.384 mmol) and Et₃N (160 µL, 1.15 mmol) at room temperature. The mixture was allowed to stir at room temperature for 10 min before a solution of ethyl 2-(5-amino-1H-indol-2-yl)propanoate (0.089 g, 0.384 mmol) in acetonitrile (2.16 mL) was added. After addition, the reaction mixture was stirred at room temperature for 2 h. The solvent was evaporated under reduced pressure and the residue was dissolved in dichloromethane. The organic layer was washed with 1 N HCl (1 x 3 mL) and then saturated aqueous NaHCO₃ (1 x 3 mL). The organic layer was dried over Na₂SO₄, filtered and evaporated under reduced pressure. The crude material was purified by column chromatography on silica gel (ethyl acetate/hexane = 1/1) to give ethyl 2-(5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-2-yl)propanoate (0.081 g, 50%). ¹H NMR (400 MHz, CDCl₃) δ 8.51 (s, 1H), 7.67 (s, 1H), 7.23-7.19 (m, 2H), 7.04-7.01 (m, 3H), 6.89 (d, J = 0.0 Hz, 1H), 6.28 (s, 1H), 6.06 (s, 2H), 4.25-4.17 (m, 2H), 3.91 (q, J = 7.2 Hz, 1H), 1.72-1.70 (m, 2H), 1.61 (s, 2H), 1.29 (t, J = 7.1 Hz, 4H), 1.13-1.11 (m, 2H).

Example 71: <i>tert</i>-Butyl 2-(5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarbox-amido)-1H-indol-2-yl)-2-methylpropylcarbamate

0533<chemistry id="chem0485" num="0485"><img file="EP2007756B1_D0511.tif" /></chemistry><chemistry id="chem0486" num="0486"><img file="EP2007756B1_D0512.tif" /></chemistry>

2-Methyl-2-(5-nitro-1H-indol-2-yl)propanoic acid

0534Ethyl 2-methyl-2-(5-nitro-1H-indol-2-yl)propanoate (4.60 g, 16.7 mmol) was dissolved in THF/water (2:1, 30 mL). LiOH˙H₂O (1.40 g, 33.3 mmol) was added and the mixture was stirred at 50 °C for 3 h. The mixture was made acidic by the careful addition of 3N HCl. The product was extracted with ethylacetate and the combined organic phases were washed with brine and dried over magnesium sulfate to give 2-methyl-2-(5-nitro-1H-indol-2-yl)propanoic acid (4.15 g, 99%). <chemistry id="chem0487" num="0487"><img file="EP2007756B1_D0513.tif" /></chemistry>

2-Methyl-2-(5-nitro-1H-indol-2-yl)propanamide

05352-Methyl-2-(5-nitro-1H-indol-2-yl)-propanoic acid (4.12 g, 16.6 mmol) was dissolved in acetonitrile (80 mL). EDC (3.80 g, 0.020 mmol), HOBt (2.70 g, 0.020 mmol), Et₃N (6.9 mL, 0.050 mmol) and ammonium chloride (1.34 g, 0.025 mmol) were added and the mixture was stirred overnight at room temperature. Water was added and the mixture was extracted with ethylacetate. Combined organic phases were washed with brine, dried over magnesium sulfate and dried to give 2-methyl-2-(5-nitro-1H-indol-2-yl)propanamide (4.3 g, 99%). <chemistry id="chem0488" num="0488"><img file="EP2007756B1_D0514.tif" /></chemistry>

2-Methyl-2-(5-nitro-1H-indol-2-yl)propan-1-amine

05362-Methyl-2-(5-nitro-1H-indol-2-yl)propanamide (200 mg, 0.81 mmol) was suspended in THF (5 ml) and cooled to 0 °C. Borane-THF complex solution (1.0 M, 2.4 mL, 2.4 mmol) was added slowly and the mixture was allowed to stir overnight at room temperature. The mixture was cooled to 0 °C and carefully acidified with 3 N HCl. THF was evaporated off, water was added and the mixture was washed with ethylacetate. The aqueous layer was made alkaline with 50% NaOH and the mixture was extracted with ethylacetate. The combined organic layers were dried over magnesium sulfate, filtered and evaporated to give 2-methyl-2-(5-nitro-1H-indol-2-yl)propan-1-amine (82 mg, 43%). <chemistry id="chem0489" num="0489"><img file="EP2007756B1_D0515.tif" /></chemistry>

<i>tert</i>-Butyl 2-methyl-2-(5-nitro-1H-indol-2-yl)propylcarbamate

05372-Methyl-2-(5-nitro-1H-indol-2-yl)propan-1-amine (137 mg, 0.587 mmol) was dissolved in THF (5 mL) and cooled to 0 °C. Et₃N (82 µL, 0.59 mmol) and di-<i>tert</i>-butyl dicarbonate (129 mg, 0.587 mmol) were added and the mixture was stirred at room temperature overnight. Water was added and the mixture was extracted with ethylacetate. The residue was purified by silica gel chromatography (10-40% ethylacetate in hexane) to give <i>tert</i>-butyl 2-methyl-2-(5-nitro-1H-indol-2-y1)propylcarbamate (131 mg, 67%). <chemistry id="chem0490" num="0490"><img file="EP2007756B1_D0516.tif" /></chemistry>

<i>tert</i>-Butyl 2-(5-amino-1H-indol-2-yl)-2-methylpropylcarbamate

0538To a solution of tert-butyl 2-methyl-2-(5-nitro-1H-indol-2-yl)propylcarbamate (80 mg, 0.24 mmol) in THF (9 mL) and water (2 mL) was added ammonium formate (60 mg, 0.96 mmol) followed by 10% Pd/C (50 mg). The mixture was stirred at room temperature for 45 minutes. Pd/C was filtered off and the organic solvent was removed by evaporation. The remaining aqueous phase was extracted with dichloromethane. The combined organic phases were dried over magnesium sulfate and evaporated to give <i>tert</i>-butyl 2-(5-amino-1H-indol-2-yl)-2-methylpropylcarbamate (58 mg, 80%). <chemistry id="chem0491" num="0491"><img file="EP2007756B1_D0517.tif" /></chemistry>

<i>tert</i>-Butyl 2-(5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-2-yl)-2-methylpropylcarbamate

0539<i>tert</i>-Butyl 2-(5-amino-1H-indol-2-yl)-2-methylpropylcarbamate (58 mg, 0.19 mmol), 1-(benzo[d][1,3]dioxol-6-yl)cyclopropanecarboxylic acid (47 mg, 0.23 mmol), EDC (45 mg, 0.23 mmol), HOBt (31 mg, 0.23 mmol) and Et₃N (80 µL, 0.57 mmol) were dissolved in DMF (4 mL) and stirred overnight at room temperature. The mixture was diluted with water and extracted with ethylacetate. The combined organic phases were dried over magnesium sulfate and evaporated to dryness. The residue was purified by silica gel chromatography (10-30% ethylacetate in hexane) to give tert-butyl 2-(5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-2-yl)-2-methylpropyl-carbamate (88 mg, 94%). ¹H NMR (400 MHz, CDCl₃) δ 8.32 (s, 1H), 7.62 (d, J = 1.5 Hz, 1H), 7.18 - 7.16 (m, 2H), 7.02 - 6.94 (m, 3H), 6.85 (d, J = 7.8 Hz, 1H), 6.19 (d, J = 1.5 Hz, 1H), 6.02 (s, 2H), 4.54 (m, 1H), 3.33 (d, J = 6.2 Hz, 2H), 1.68 (dd, J = 3.7, 6.8 Hz, 2H), 1.36 (s, 9H), 1.35 (s, 6H), 1.09 (dd, J = 3.7, 6.8 Hz, 2H).

Example 72: (<i>R</i>)-<i>N</i>-(2-<i>tert</i>-Butyl-1-(2,3-dihydroxypropyl)-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

0540<chemistry id="chem0492" num="0492"><img file="EP2007756B1_D0518.tif" /></chemistry><chemistry id="chem0493" num="0493"><img file="EP2007756B1_D0519.tif" /></chemistry>

(<i>R</i>)-2-<i>tert</i>-Butyl-1-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-5-nitro-1H-indole

0541To a stirred solution of (S)-(2,2-dimethyl-1,3-dioxolan-4-yl)methyl 4-methylbenzenesulfonate (1.58 g, 5.50 mmol) in anhydrous DMF (10 mL) under nitrogen gas was added 2-<i>tert</i>-butyl-5-nitro-1H-indole (1.00 g, 4.58 mmol) followed by Cs₂CO₃ (2.99 g, 9.16 mol). The mixture was stirred and heated at 80 °C under nitrogen gas. After 20 hours, 50% conversion was observed by LCMS. The reaction mixture was re-treated with Cs₂CO₃ (2.99 g, 9.16 mol) and (<i>S</i>)-(2,2-dimethyl-1,3-dioxolan-4-yl)methyl 4-methylbenzenesulfonate (1.58 g, 5.50 mmol) and heated at 80 °C for 24 hours. The reaction mixture was cooled to room temperature. The solids were filtered and washed with ethyl acetate and hexane (1:1). The layers were separated and the organic layer was washed with water (2 x 10 mL) and brine (2 x 10 mL). The organic layer was dried over Na₂SO₄, filtered and evaporated under reduced pressure. The residue was purified by column chromatography on silica gel (dichloromethane /hexane = 1.5/1) to give (<i>R</i>)-2-<i>tert</i>-butyl-1-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-5-nitro-1H-indole (1.0 g, 66%). ¹H NMR (400 MHz, CDCl₃) δ 8.48 (d, J = 2.2 Hz, 1H), 8.08 (dd, J = 2.2, 9.1 Hz, 1H), 7.49 (d, J = 9.1 Hz, 1H), 6.00 (s, 1H), 4.52-4.45 (m, 3H), 4.12 (dd, J = 6.0, 8.6 Hz, 1H), 3.78 (dd, J = 6.0, 8.6 Hz, 1H), 1.53 (s, 3H), 1.51 (s, 9H), 1.33 (s, 3H). <chemistry id="chem0494" num="0494"><img file="EP2007756B1_D0520.tif" /></chemistry>

(<i>R</i>)-2-<i>tert</i>-Butyl-1-((2,2-dimethyl-1,3-dioxolan4-yl)methyl-1H-indol-5-amine

0542To a stirred solution of (<i>R</i>)-2-<i>tert</i>-butyl-1-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-5-nitro-1H-indole (1.0 g, 3.0 mmol) in ethanol (20 mL) and water (5 mL) was added ammonium formate (0.76 g, 12 mmol) followed by slow addition of 10 % palladium on carbon (0.4 g). The mixture was stirred at room temperature for 1 h. The reaction mixture was filtered through a plug of celite and rinsed with ethyl acetate. The filtrate was evaporated under reduced pressure and the crude product was dissolved in ethyl acetate. The organic layer was washed with water (2 x 5 mL) and brine (2 x 5 mL). The organic layer was dried over Na₂SO₄, filtered and evaporated under reduced pressure to give (<i>R</i>)-2-<i>tert</i>-butyl-1-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl-1<i>H</i>-indol-5-amine (0.89 g, 98%). ¹H NMR (400 MHz, CDCl₃) δ 7.04 (d, J = 4 Hz, 1H), 6.70 (d, J = 2.2 Hz, 1H), 6.48 (dd, <i>J</i> = 2.2, 8.6 Hz, 1H), 6.05 (s, 1H,), 4.38-4.1 (m, 2H), 4.21 (dd, J = 7.5, 16.5 Hz, 1H), 3.87 (dd, J = 6.0, 8.6 Hz, 1H), 3.66 (dd, J = 6.0, 8.6 Hz, 1H), 3.33 (br s, 2H), 1.40 (s, 3H), 1.34 (s, 9H), 1.25 (s, 3H). <chemistry id="chem0495" num="0495"><img file="EP2007756B1_D0521.tif" /></chemistry>

<i>N</i>-((<i>R</i>)-2-<i>tert</i>-Butyl-1-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-1H-indo1-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

0543To 1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylic acid (0.73 g, 3.0 mmol) was added thionyl chloride (660 µL, 9.0 mmol) and DMF (20 µL) at room temperature. The mixture was stirred for 30 minutes before the excess thionyl chloride was evaporated under reduced pressure. To the resulting acid chloride, dichloromethane (6.0 mL) and Et₃N (2.1 mL, 15 mmol) were added. A solution of (R)-2-<i>tert</i>-butyl-1-((2,2-dimethyl-1,3-dioxotan-4-yl)methyl-1H-indol-5-amine (3.0 mmol) in dichloromethane (3.0 mL) was added to the cooled acid chloride solution. After addition, the reaction mixture was stirred at room temperature for 45 minutes. The reaction mixture was filtered and the filtrate was evaporated under reduced pressure. The residue was purified by column chromatography on silica gel (ethyl acetate/hexane = 3/7) to give <i>N</i>-((<i>R</i>)-2-<i>tert</i>-butyl-1-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-1<i>H</i>-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide (1.33 g, 84%). ¹H NMR (400 MHz, CDCl₃) δ 7.48 (d, J = 2 Hz, 1H,), 7.31 (dd, J = 2, 8 Hz, 1H), 7.27 (dd, J = 2, 8 Hz, 1H), 7.23 (d, J = 8 Hz, 1H), 7.14 (d, J = 8 Hz, 1H), 7.02 (dd, J = 2, 8 Hz, 1H), 6.92 (br s, 1H), 6.22 (s, 1H), 4.38-4.05 (m, 3H), 3.91 (dd, J = 5, 8 Hz, 1H), 3.75 (dd, J = 5, 8 Hz, 1H), 2.33 (q, J = 8 Hz, 2H), 1.42 (s, 3H), 1.37 (s, 9H), 1.22 (s, 3H), 1.10 (q, J = 8 Hz, 2H). <chemistry id="chem0496" num="0496"><img file="EP2007756B1_D0522.tif" /></chemistry>

<i>N</i>-((<i>R</i>)-2-<i>tert</i>-Butyl-1-((2,3-dihydroxypropyl)-1<i>H</i>-indol-5-yl)-1-(2,2-difluorobenzo-[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

0544To a stirred solution of <i>N</i>-(2-tert-butyl-1-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide (1.28 g, 2.43 mmol) in methanol (34 mL) and water (3.7 mL) was added para-toluenesulfonic acid-hydrate (1.87 g, 9.83 mmol). The reaction mixture was stirred and heated at 80 °C for 25 minutes. The solvent was evaporated under reduced pressure. The crude product was dissolved in ethyl acetate. The organic layer was washed with saturated aqueous NaHCO₃ (2 x 10 mL) and brine (2 x 10 mL). The organic layer was dried over Na₂SO₄, filtered and evaporated under reduced pressure. The residue was purified by column chromatography on silica gel (ethyl acetate/hexane = 13/7) to give <i>N</i>-((R)-2-tert-butyl-1-((2,3-dihydroxypropyl)-1<i>H</i>-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide (0.96 g, 81%). ¹H NMR (400 MHz, CDCl₃) δ 7.50 (d, J = 2 Hz, 1H), 7.31 (dd, J = 2, 8 Hz, 1H), 7.27 (dd, J = 2, 8 Hz, 1H), 7.23 (d, J = 8 Hz, 1H), 7.14 (d, J = 8 Hz, 1H), 7.02 (br s, 1H,), 6.96 (dd, J = 2, 8 Hz, 1H), 6.23 (s, 1H), 4.35 (dd, J = 8, 15 Hz, 1H), 4.26 (dd, J = 4, 15 Hz, 1H,), 4.02-3.95 (m, 1H), 3.60 (dd, J = 4, 11 Hz, 1H), 3.50 (dd, J = 5, 11 Hz, 1H), 1.75 (q, J = 8 Hz, 3H), 1.43 (s, 9H), 1.14 (q, J = 8 Hz, 3H).

Example 73: 3-(2-<i>tert</i>-Butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-1-yl)-2-hydroxypropanoic acid

0545<chemistry id="chem0497" num="0497"><img file="EP2007756B1_D0523.tif" /></chemistry><chemistry id="chem0498" num="0498"><img file="EP2007756B1_D0524.tif" /></chemistry>

3-(2-<i>tert</i>-Butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropaneearbox-amido)-1H-indol-1-yl)-2-oxopropanoic acid

0546To a solution of <i>N</i>-(2-<i>tert</i>-butyl-1-(2,3-dihydroxypropyl)-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropane-carboxamide (97 mg, 0.20 mmol) in DMSO (1 mL) was added Dess-Martin periodinane (130 mg, 0.30 mmol). The mixture was stirred at room temperature for 3 h. The solid was filtered off and washed with EtOAc. The filtrate was partitioned between EtOAc and water. The aqueous layer was extracted with EtOAc twice and the combined organic layers were washed with brine and dried over MgSO₄. After the removal of solvent, the residue was purified by preparative TLC to yield 3-(2-<i>tert</i>-butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-1-yl)-2-oxopropanoic acid that was used without further purification. <chemistry id="chem0499" num="0499"><img file="EP2007756B1_D0525.tif" /></chemistry>

3-(2-<i>tert</i>-Butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarbox-amido)-1H-indol-1-yl)-2-hydroxypropanoic acid

0547To a solution of 3-(2-<i>tert</i>-butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-1-yl)-2-oxopropanoic acid (50 mg, 0.10 mmol) in MeOH (1 mL) was added NaBH₄ (19 mg, 0.50 mmol) at 0 °C. The mixture was stirred at room temperature for 15 min. The resulting mixture was partitioned between EtOAc and water. The aqueous layer was extracted with EtOAc twice and the combined organic layers were washed with brine and dried over anhydrous MgSO₄. After the removal of the solvent, the residue was taken up in DMSO and purified by preparative LC/MS to give 3-(2-<i>tert-</i>butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-1-yl)-2-hydroxypropanoic acid. ¹H NMR (400 MHz, CDCl₃) δ 7.36 (s), 7.27-7.23 (m, 2H), 7.15-7.11 (m, 2H), 6.94 (d, J = 8.5 Hz, 1H), 6.23 (s, 1H), 4.71 (s, 3H), 4.59 (q, J = 10.3 Hz, 1H), 4.40-4.33 (m, 2H), 1.70 (d, J = 1.9 Hz, 2H), 1.15 (q, J = 4.0 Hz, 2H). ¹³C NMR (400 MHz, CDCl₃) δ 173.6, 173.1, 150.7, 144.1, 143.6, 136.2, 135.4, 134.3, 131.7, 129.2, 129.0, 127.6, 126.7, 116.6, 114.2, 112.4, 110.4, 110.1, 99.7, 70.3, 48.5, 32.6, 30.9, 30.7, 16.8. MS (ESI) m/e (M+H⁺) 501.2.

Example 74: (<i>R</i>)-<i>N</i>-(2-<i>tert</i>-Butyl-1-(2,3-dihydroxypropyl)-1H-indol-5-yl)-1-(2,2-dideuteriumbenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

0548<chemistry id="chem0500" num="0500"><img file="EP2007756B1_D0526.tif" /></chemistry><chemistry id="chem0501" num="0501"><img file="EP2007756B1_D0527.tif" /></chemistry>

Methyl 1-(3,4-dihydroxyphenyl)cyclopropanecarboxylate

0549To a solution of 1-(3,4-dihydroxyphenyl)cyclopropanecarboxylic acid (190 mg, 1.0 mmol) in MeOH (3 mL) was added 4-methylbenzenesulfonic acid (19 mg, 0.10 mmol). The mixture was heated at 80°C overnight. The reaction mixture was concentrated in <i>vacuo</i> and partitioned between EtOAc and water. The aqueous layer was extracted with EtOAc twice and the combined organic layers were washed with sat. NaHCO₃ and brine and dried over MgSO₄. After the removal of solvent, the residue was dried in <i>vacuo</i> to yield methyl 1-(3,4-dihydroxyphenyl)cyclopropanecarboxylate (190 mg, 91%) that was used without further purification. ¹H NMR (400 MHz, DMSO-d⁶) δ 6.76-6.71 (m, 2H), 6.66 (d, J = 7.9 Hz, 1H), 3.56 (s, 3H), 1.50 (q, J = 3.6 Hz, 2H), 1.08 (q, J = 3.6 Hz, 2H). <chemistry id="chem0502" num="0502"><img file="EP2007756B1_D0528.tif" /></chemistry>

Methyl 1-(2,2-dideuteriumbenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylate

0550To a solution of methyl 1-(3,4-dihydroxyphenyl)cyclopropanecarboxylate (21 mg, 0.10 mmol) and CD₂Br₂ (35 mg, 0.20 mmol) in DMF (0.5 mL) was added Cs₂CO₃ (19 mg, 0.10 mmol). The mixture was heated at 120°C for 30 min. The reaction mixture was partitioned between EtOAc and water. The aqueous layer was extracted with EtOAc twice and the combined organic layers were washed with 1N NaOH and brine before being dried over MgSO₄. After the removal of solvent, the residue was dried in <i>vacuo</i> to yield methyl 1-(2,2-dideuteriumbenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylate (22 mg) that was used without further purification. ¹H NMR (400 MHz, CDCl₃) δ 6.76-6.71 (m, 2H), 6.66 (d, J = 7.9 Hz, 1H), 3.56 (s, 3H), 1.50 (q, J = 3.6 Hz, 2H), 1.08 (q, J = 3.6 Hz, 2H). <chemistry id="chem0503" num="0503"><img file="EP2007756B1_D0529.tif" /></chemistry>

1-(2,2-Dideuteriumbenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylic acid

0551To a solution of methyl 1-(2,2-dideuteriumbenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylate (22 mg, 0.10 mmol) in THF (0.5 mL) was added NaOH (IN, 0.25 mL, 0.25 mmol). The mixture was heated at 80°C for 2 h. The reaction mixture was partitioned between EtOAc and 1N NaOH. The aqueous layer was extracted with EtOAc twice, neutralized with 1N HCl and extracted with EtOAc twice. The combined organic layers were washed with brine and dried over MgSO₄. After the removal of solvent, the residue was dried in <i>vacuo</i> to yield 1-(2,2-dideuteriumbenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylic acid (21 mg) that was used without further purification. <chemistry id="chem0504" num="0504"><img file="EP2007756B1_D0530.tif" /></chemistry>

(<i>R</i>)-<i>N</i>-(2-<i>tert</i>-Butyl-1-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-1H-indo1-5-yl)-1-(2,2-dideuteriumbenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

0552To a solution of 1-(2,2-dideuteriumbenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylic acid (21 mg, 0.10 mmol), (<i>R</i>)-2-<i>tert</i>-butyl-1-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-1H-indol-5-amine (30 mg, 0.10 mmol), HATU (42 mg, 0.11 mol) in DMF (1 mL) was added triethylamine (0.030 mL, 0.22 mmol). The mixture was heated at room temperature for 5 min. The reaction mixture was partitioned between EtOAc and water. The aqueous layer was extracted with EtOAc twice and the combined organic layers were washed with 1N NaOH, 1N HCl, and brine before being dried over MgSO₄. After the removal of solvent, the residue was purified by column chromatography (20-40% ethyl acetate/hexane) to yield (<i>R</i>)-<i>N</i>-(2-<i>tert</i>-butyl-1-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-1H-indol-5-yl)-1-(2,2-dideuteriumbenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide (24 mg, 49% from methyl 1-(3,4-dihydroxyphenyl)cyclopropanecarboxylate). MS (ESI) m/e (M+H⁺) 493.5. <chemistry id="chem0505" num="0505"><img file="EP2007756B1_D0531.tif" /></chemistry>

(<i>R</i>)-<i>N</i>-(2-<i>tert</i>-Butyl-1-(2,3-dihydroxypropyl)-1H-indol-5-yl)-1-(2,2-dideuterium-benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

0553To a solution of (<i>R</i>)-<i>N</i>-(2-<i>tert</i>-butyl-1-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-1H-indol-5-yl)-1-(2,2-dideutenum-benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide (24 mg, 0.050 mmol), in methanol (0.5 mL) and water (0.05 mL) was added 4-methylbenzenesulfonic acid (2.0 mg, 0.010 mmol). The mixture was heated at 80°C for 30 min. The reaction mixture was partitioned between EtOAc and water. The aqueous layer was extracted with EtOAc twice and the combined organic layers were washed with sat. NaHCO₃ and brine before being dried over MgSO₄. After the removal of solvent, the residue was purified by preparative HPLC to yield (<i>R</i>)-<i>N</i>-(2-<i>tert</i>-butyl-1-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-1H-indol-5-yl)-1-(2,2-dideuteriumbenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide (12 mg, 52%). ¹H NMR (400 MHz, CDCl₃) δ 7.44 (d, J = 2.0 Hz, 1H), 7.14 (dd, J = 22.8, 14.0 Hz, 2H), 6.95-6.89 (m, 2H), 6.78 (d, J = 7.8 Hz, 1H), 6.14 (s, 1H), 4.28 (dd, J = 15.1, 8.3 Hz, 1H), 4.19 (dd, J = 15.1, 4.5 Hz, 1H), 4.05 (q, J = 7.1 Hz, 1H), 3.55 (dd, J = 11.3, 4.0 Hz, 1H), 3.45 (dd, J = 11.3, 5.4 Hz, 1H), 1.60 (q, J = 3.5 Hz, 2H), 1.35 (s, 9H), 1.02 (q, J = 3.5 Hz, 2H). ¹³C NMR (400 MHz, CDCl₃) δ 171.4, 149.3, 147.1, 146.5, 134.8, 132.3, 129.2, 126.5, 123.6, 114.3, 111.4, 110.4, 109.0, 107.8, 98.5, 70.4, 63.1, 46.6, 31.6, 30.0, 29.5, 15.3. MS (ESI) m/e (M+H⁺) 453.5.

0554It is further noted that the mono-deuterated analogue for this compound can be synthesized by substitution the reagent CHDBR₂ for CD₂BR₂ and following the procedures described in example 74. Furthermore, deuterated analogues of the compounds as described herein such as of formula I can be produced using known synthesitc methods as well as the methodology described herein. The deuterated analogues include both di and mono-deuterated analogues of the compounds of the present invention. The di and mono deuterated analoges of the compounds exhibit measurable acitivity when tested using the assays described below.

Example 75: 4-(5-(1-(Benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-2-yl)-4-methylpentanoic acid

0555<chemistry id="chem0506" num="0506"><img file="EP2007756B1_D0532.tif" /></chemistry><chemistry id="chem0507" num="0507"><img file="EP2007756B1_D0533.tif" /></chemistry>

1-(Benzo[d][1,3]dioxol-5-yl)-<i>N</i>-(2-(4-cyano-2-methylbutan-2-yl)-1H-indol-5-yl)cyclopropanecarboxamide

0556To 1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylic acid (0.068 g, 0.33 mmol) was added thionyl chloride (72 µL, 0.99 mmol) and DMF (20 µL) at room temperature. The mixture was stirred for 30 minutes before the excess thionyl chloride was evaporated under reduced pressure. To the resulting acid chloride, dichloromethane (0.5 mL) and Et₃N (230 µL, 1.7 mmol) were added. A solution of 4-(5-amino-1H-indol-2-yl)-4-methylpentanenitrile (0.33 mmol) in dichloromethane (0.5 mL) was added to the acid chloride solution and the mixture was stirred at room temperature for 1.5 h. The resulting mixture was diluted with dichloromethane and washed with 1 N HCl (2 x 2 mL), saturated aqueous NaHCO₃ (2 x 2 mL) and brine (2 x 2 mL). The organic layer was dried over anhydrous Na₂SO₄ and evaporated under reduced pressure to give 1-(benzo[d][1,3]dioxol-5-yl)-<i>N</i>-(2-(4-cyano-2-methylbutan-2-yl)-1 H-indol-5-yl)cyclopropanecarboxamide. <chemistry id="chem0508" num="0508"><img file="EP2007756B1_D0534.tif" /></chemistry>

4-(5-(1-(Benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-2-yl)-4-methylpentanoic acid

0557A mixture of 1-(benzo[d][1,3]dioxol-5-yl)-<i>N</i>-(2-(4-cyano-2-methylbutan-2-yl)-1H-indol-5-yl)cyclopropanecarboxamide (0.060 g, 0.15 mmol) and KOH (0.081 g, 1.5 mmol) in 50% EtOH/water (2 mL) was heated in the microwave at 100 °C for 1 h. The solvent was evaporated under reduced pressure. The crude product was dissolved in DMSO (1 mL), filtered, and purified by reverse phase preparative HPLC to give 4-(5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1 H-indol-2-yl)-4-methylpentanoic acid. ¹H NMR (400 MHz, DMSO-d6) δ 11.98 (s, 1H), 10.79 (s, 1H), 8.44 (s, 1H), 7.56 (s, 1H), 7.15 (d, J = 8.6 Hz, 1H), 7.03-6.90 (m, 4H), 6.05 (s, 1H), 6.02 (s, 2H), 1.97-1.87 (m, 4H), 1.41-1.38 (m, 2H), 1.30 (s, 6H), 1.04-1.02 (m, 2H).

Example 76: 1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-(1-hydroxypropan-2-yl)-1H-indol-5-yl)cyclopropanecarboxamide

0558<chemistry id="chem0509" num="0509"><img file="EP2007756B1_D0535.tif" /></chemistry>

2-(5-Nitro-1H-indol-2-yl)propan-1-ol

0559To a cooled solution of LiAlH₄ (1.0 M in THF, 1.2 mL, 1.2 mmol) in THF (5.3 mL) at 0 °C was added a solution of ethyl 2-(5-nitro-1H-indol-2-yl)propanoate (0.20 g, 0.76 mmol) in THF (3.66 mL) dropwise. After addition, the mixture was allowed to warm up to room temperature and was stirred at room temperature for 3 h. The mixture was cooled to 0 °C. Water (2 mL) was slowly added followed by careful addition of 15% NaOH (2 mL) and water (4 mL). The mixture was stirred at room temperature for 0.5 h and was then filtered through a short plug of celite using ethyl acetate. The organic layer was separated from the aqueous layer, dried over Na₂SO₄, filtered and evaporated under reduced pressure. The residue was purified by column chromatography on silica gel (ethyl acetate/hexane = 1/1) to give 2-(5-nitro-1H-indol-2-yl)propan-1-ol (0.14 g, 81%). <chemistry id="chem0510" num="0510"><img file="EP2007756B1_D0536.tif" /></chemistry>

2-(5-Amino-1H-indol-2-yl)propan-1-ol

0560To a solution of 2-(5-nitro-1H-indol-2-yl)propan-1-ol (0.13 g, 0.60 mmol) in ethanol (5 mL) was added tin chloride dihydrate (0.67 g, 3.0 mmol). The mixture was heated in the microwave at 120 °C for 1 h. The mixture was diluted with ethyl acetate before water and saturated aqueous NaHCO₃ were added. The reaction mixture was filtered through a plug of celite using ethyl acetate. The organic layer was separated from the aqueous layer, dried over Na₂SO₄, filtered and evaporated under reduced pressure to give 2-(5-amino-1H-indol-2-yl)propan-1-ol (0.093 g, 82%). <chemistry id="chem0511" num="0511"><img file="EP2007756B1_D0537.tif" /></chemistry>

1-(Benzo[d][1,3]dioxol-5-yl)-<i>N</i>-(2-(1-hydroxypropan-2-yl)-1H-indol-5-yl)cyclopropanecarboxamide

0561To a solution of 1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylic acid (0.10 g, 0.49 mmol) in acetonitrile (2.0 mL) were added HBTU (0.185 g, 0.49 mmol) and Et₃N (205 µL, 1.47 mmol) at room temperature. The mixture was allowed to stir at room temperature for 10 minutes before a slurry of 2-(5-amino-1H-indol-2-yl)propan-1-ol (0.093 g, 0.49 mmol) in acetonitrile (2.7 mL) was added. After addition, the reaction mixture was stirred at room temperature for 5.5 h. The solvent was evaporated under reduced pressure and the residue was dissolved in dichloromethane. The organic layer was washed with 1 N HCl (1 x 3 mL) and saturated aqueous NaHCO₃ (1 x 3 mL). The organic layer was dried over Na₂SO₄, filtered and evaporated under reduced pressure. The crude material was purified by column chromatography on silica gel (ethyl.acetate/hexane = 13/7) to give 1-(benzo[d][1,3]dioxol-5-yl)-<i>N</i>-(2-(1-hydroxypropan-2-yl)-1H-indol-5-yl)cyclopropanecarboxamide (0.095 g, 51%). ¹H NMR (400 MHz, DMSO-d6) δ 10.74 (s, 1H), 8.38 (s, 1H), 7.55 (s, 1H), 7.14 (d, J = 8.6 Hz, 1H), 7.02-6.90 (m, 4H), 6.06 (s, 1H), , 6.02 (s, 2H), 4.76 (t, J = 5.3 Hz, 1H), 3.68-3.63 (m, 1H), 3.50-3.44 (m, 1H), 2.99-2.90 (m, 1H), 1.41-1.38 (m, 2H), 1.26 (d, J = 7.0 Hz, 3H), 1.05-1.02 (m, 2H).

Example 77: 1-(Benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N</i>-(2-tert-butyl-1<i>H</i>-indol-5-yl)-<i>N-</i>methylcyclopropanecarboxamide

0562<chemistry id="chem0512" num="0512"><img file="EP2007756B1_D0538.tif" /></chemistry>

1-(Benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N</i>-(2-tert-butyl-1<i>H</i>-indol-5-yl)-<i>N-</i>methylcyclopropanecarboxamide

05632-<i>tert</i>-Butyl-<i>N</i>-methyl-1<i>H</i>-indol-5-amine (20.2 mg, 0.100 mmol) and 1-(benzo[<i>d</i>][1,3]dioxol-5-yl)cyclopropanecarboxylic acid (20.6 mg, 0.100 mmol) were dissolved in <i>N,N</i>-dimethylformamide (1 mL) containing triethylamine (42.1 µL, 0.300 mmol) and a magnetic stir bar. <i>O</i>-(7-Azabenzotriazol-1-yl)-<i>N,N,N',N'</i>-tetramethyluronium hexafluorophosphate (42 mg, 0.11 mmol) was added to the mixture and the resulting solution was allowed to stir for 16 h at 80 °C. The crude product was then purified by preparative HPLC utilizing a gradient of 0-99% acetonitrile in water containing 0.05% trifluoroacetic acid to yield 1-(benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butyl-1<i>H</i>-indol-5-yl)-<i>N-</i>methylcyclopropanecarboxamide. ESI-MS <i>m</i>/<i>z</i> calc. 390.2, found 391.3 (M+1)⁺. Retention time of 3.41 minutes.

Example 78: <i>N</i>-(2-<i>tert</i>-Butyl-1-methyl-1H-indol-5-yl)-1-(benzo[d][1,3]dioxol-6-yl)-N-methylcyclopropanecarboxamide

0564<chemistry id="chem0513" num="0513"><img file="EP2007756B1_D0539.tif" /></chemistry>

0565Sodium hydride (0.028 g, 0.70 mmol, 60% by weight dispersion in oil) was slowly added to a stirred solution of N-(2-tert-butyl-1H-indol-5-yl)-1-(benzo[d][1,3]dioxol-6-yl)cyclopropanecarboxamide (0.250 g, 0.664 mmol) in a mixture of 4.5 mL of anhydrous tetrahydrofuran (THF) and 0.5 mL of anhydrous <i>N,N</i>-dimethylformamide (DMF). The resulting suspension was allowed to stir for 2 minutes and then iodomethane (0.062 mL, 1.0 mmol) was added to the reaction mixture. Two additional aliquots of sodium hydride and iodomethane were required to consume all of the starting material which was monitored by LC / MS. The crude reaction product was evaporated to dryness, redissolved in a minimum of DMF and purified by preparative LC / MS chromatography to yield the pure product (0.0343 g, 13%) ESI-MS <i>m</i>/<i>z</i> calc. 404.2, found 405.3 (M+1)⁺. Retention time of 3.65 minutes.

Example 79: 1-(Benzo[d][1,3]dioxol-5-yl)-N-(2-(hydroxymethyl)-1H-indol-5-yl)cyclopropanecarboxamide

0566<chemistry id="chem0514" num="0514"><img file="EP2007756B1_D0540.tif" /></chemistry>

0567Ethyl 5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indole-2-carboxylate (1.18 g, 3.0 mmol) was added to a solution of LiBH₄ (132 mg, 6.0 mmol) in THF (10 mL) and water (0.1 mL). The mixture was allowed to stir for 16h at 25 °C before it was quenched with water (10 mL) and slowly made acidic by addition of 1 N HCl. The mixture was extracted with three 50-mL portions of ethyl acetate. The organic extracts were dried over Na₂SO₄ and evaporated to yield 1-(benzo[d][1,3]dioxol-5-yl)-N-(2-(hydroxymethyl)-1H-indol-5-yl)cyclopropanecarboxamide (770 mg, 73%). A small amount was further purified by reverse phase HPLC. ESI-MS <i>m</i>/<i>z</i> calc. 350.4, found 351.3 (M+1)⁺; retention time 2.59 minutes.

Example 80: 5-(1-(Benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-<i>N-tert</i>-butyl-1H-indole-2-carboxamide

0568<chemistry id="chem0515" num="0515"><img file="EP2007756B1_D0541.tif" /></chemistry><chemistry id="chem0516" num="0516"><img file="EP2007756B1_D0542.tif" /></chemistry>

5-(1-(Benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indole-2-carboxylic acid

0569Ethyl 5-(1-(benzo[d][1,3]dioxol-5-yl)cydopropanecarboxamido)-1H-indole-2-carboxylate (392 mg, 1.0 mmol) and LiOH (126 mg, 3 mmol) were dissolved in H₂O (5 mL) and 1,4-dioxane (3 mL). The mixture was heated in an oil bath at 100 °C for 24 hours before it was cooled to room temperature. The mixture was acidified with 1N HCl and it was extracted with three 20 mL portions of dichloromethane. The organic extracts were dried over Na₂SO₄ and evaporated to yield 5-(1-(benzo[d][1,3]-dioxol-5-yl)cyclopropanecarboxamido)-1H-indole-2-carboxylic acid (302 mg, 83%). A small amount was further purified by reverse phase HPLC. ESI-MS <i>m</i>/<i>z</i> calc. 364.1, found 365.1 (M+1)⁺; retention time 2.70 minutes. <chemistry id="chem0517" num="0517"><img file="EP2007756B1_D0543.tif" /></chemistry>

5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-<i>N</i>-<i>tert</i>-butyl-1H-indole-2-carboxamide

05705-(1-(Benzo[d][1,3]dioxol-5-yl)cyclopropane-carboxamido)-1H-indole-2-carboxylic acid (36 mg, 0.10 mmol) and 2-methylpropan-2-amine (8.8 mg, 0.12 mmol) were dissolved in <i>N,N</i>-dimethylformamide (1.0 mL) containing triethylamine (28 µL, 0.20 mmol). <i>O</i>-(7-Azabenzotriazol-1-yl)-<i>N,N,N',N'</i>-tetramethyluronium hexafluorophosphate (46 mg, 0.12 mmol) was added to the mixture and the resulting solution was allowed to stir for 3 hours. The mixture was filtered and purified by reverse phase HPLC to yield 5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-<i>N</i>-<i>tert</i>-butyl-1H-indole-2-carboxamide. ESI-MS <i>m</i>/<i>z</i> calc. 419.2, found 420.3 (M+1)⁺; retention time 3.12 minutes.

Example 81: <i>N</i>-(3-Amino-2-<i>tert</i>-butyl-1H-indol-5-yl)-1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

0571<chemistry id="chem0518" num="0518"><img file="EP2007756B1_D0544.tif" /></chemistry>

0572A solution of 1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-1H-indol-5-yl)cyclopropane carboxamide (50 mg, 0.13 mmol) was dissolved in AcOH (2 mL) and warmed to 45 °C. To the mixture was added a solution of NaNO₂ (9 mg) in H₂O (0.03 mL). The mixture was allowed to stir for 30 min at 45 °C before the precipitate was collected and washed with Et₂O. This material was used in the next step without further purification. To the crude material, 1-(benzo[d][1,3]dioxol-5-yl)-N-(2-tert-butyl-3-nitroso-1H-indol-5-yl)cyclopropanecarboxamide, was added AcOH (2 mL) and Zn dust (5 mg). The mixture was allowed to stir for 1h at ambient temperature. EtOAc and H₂O were added to the mixture. The layers were separated and the organic layer was washed with sat. aq. NaHCO₃, dried over MgSO₄, and concentrated <i>in vacuo.</i> The residue was taken up in DMF (1 mL) and was purified using prep-HPLC. LCMS: <i>m</i>/<i>z</i> 392.3; retention time of 2.18 min.

Example 82: 1-(Benzo[d][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butyl-3-(methylsulfonyl)-1H-indol-5-yl)cyclopropanecarboxamide

0573<chemistry id="chem0519" num="0519"><img file="EP2007756B1_D0545.tif" /></chemistry>

1-(Benzo[d][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butyl-3-(methylsulfonyl)-1H-indol-5-yl)cyclopropanecarboxamide

0574To a solution of 1-(benzo[d][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butyl-1H-indol-5-yl)cyclopropanecarboxamide (120 mg, 0.31 mmol) in anhydrous DMF-THF (3.3 mL, 1:9) was added NaH (60% in mineral oil, 49 mg, 1.2 mmol) at room temperature. After 30 min under N₂, the suspension was cooled down to -15 °C and a solution of methanesulfonyl chloride (1.1 eq.) in DMF (0.5 mL) was added dropwise. The reaction mixture was stirred for 30 min at -15 °C then for 6 h at room temperature. Water (0.5 mL) was added at 0 °C, solvent was removed, and the residue was diluted with MeOH, filtrated and purified by preparative HPLC to give 1-(Benzo[d][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butyl-3-(methylsulfonyl)-1H-indol-5-yl)cyclopropanecarboxamide. ¹H NMR (400 MHz, DMSO) δ 11.6 (s, 1H), 8.7 (s, 1H), 7.94 (d, <i>J</i> =1.7 Hz, 1H), 7.38 (d, <i>J</i> =8.7 Hz, 1H), 7.33 (dd, <i>J1</i> =1.9 Hz, <i>J2</i> =8.7 Hz, 1H), 7.03 (d, <i>J</i> =1.7 Hz, 1H), 6.95 (dd, <i>J1</i> =1.7 Hz, <i>J2</i> =8.0 Hz, 1H), 6.90 (d, <i>J</i> =8.0 Hz, 1H), 6.02 (s, 2H), 3.07 (s, 3H), 1.56-1.40 (m, 9H), 1.41 (dd, <i>J1</i> =4.0 Hz, <i>J2</i> =6.7 Hz, 2H), 1.03 (dd, <i>J1</i> =4.0 Hz, <i>J2</i> =6.7 Hz, 2H). MS (ESI) m/e (M+H⁺) 455.5.

Example 83: 1-(Benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N</i>-(3-phenyl-1<i>H</i>-indol-5-yl)cyclopropane carboxamide

0575<chemistry id="chem0520" num="0520"><img file="EP2007756B1_D0546.tif" /></chemistry><chemistry id="chem0521" num="0521"><img file="EP2007756B1_D0547.tif" /></chemistry>

1-(Benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N</i>-(3-bromo-1<i>H</i>-indol-5-yl)cyclopropanecarboxamide

0576Freshly recrystallized <i>N</i>-bromosuccinimde (0.278 g, 1.56 mmol) was added portionwise to a solution of 1-(benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N</i>-(1<i>H</i>-indol-5-yl)cyclopropanecarboxamide (0.500 g, 1.56 mmol) in <i>N,N</i>-dimethylformamide (2 mL) over 2 minutes. The reaction mixture was protected from light and was stirred bar for 5 minutes. The resulting green solution was poured into 40 mL of water. The grey precipitate which formed was filtered and washed with water to yield 1-(benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N</i>-(3-bromo-1<i>H</i>-indol-5-yl)cyclopropapecarboxamide (0.564 g, 91%). ESI-MS <i>m</i>/<i>z</i> calc. 398.0, found 399.3 (M+1)⁺. Retention time of 3.38 minutes. ¹H NMR (400 MHz, DMSO-<i>d</i>6) 11.37 (s, 1H), 8.71 (s, 1H), 7.67 (d, <i>J</i> = 1.8 Hz, 1H), 7.50 (d, <i>J</i> = 2.6 Hz, 1H), 7.29 (d, <i>J</i> = 8.8 Hz, 1H), 7.22 (dd, <i>J</i> = 2.0, 8.8 Hz, 1H), 7.02 (d, <i>J</i> = 1.6 Hz, 1H), 6.96 - 6.88 (m, 2H), 6.03 (s, 2H), 1.43 - 1.40 (m, 2H), 1.09 - 1.04 (m, 2H). <chemistry id="chem0522" num="0522"><img file="EP2007756B1_D0548.tif" /></chemistry>

1-(Benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N</i>-(3-phenyl-1<i>H</i>-indol-5-yl)cyclopropanecarboxamide

0577Phenyl boronic acid (24.6 mg, 0.204 mmol) was added to a solution of 1-(benzo[<i>d</i>][1,3]-dioxol-5-yl)-<i>N</i>-(3-bromo-1<i>H</i>-indol-5-yl)cyclopropanecarboxamide (39.9 mg, 0.100 mmol) in ethanol (1 mL) containing FibreCat 1001 (6 mg) and 1M aqueous potassium carbonate (0.260 mL). The reaction mixture was then heated at 130 °C in a microwave reactor for 20 minutes. The crude product was then purified by preparative HPLC utilizing a gradient of 0-99% acetonitrile in water containing 0.05% trifluoroacetic acid to yield 1-(benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N</i>-(3-phenyl-1<i>H</i>-indol-5-yl)cyclopropane carboxamide. ESI-MS <i>m</i>/<i>z</i> calc. 396.2, found 397.3 (M+1)⁺. Retention time of 3.52 minutes. ¹H NMR (400 MHz, DMSO-<i>d</i>6) δ 11.27 (d, <i>J</i> = 1.9 Hz, 1H), 8.66 (s, 1H), 8.08 (d, <i>J</i> = 1.6 Hz, 1H), 7.65-7.61 (m, 3H), 7.46-7.40 (m, 2H), 7.31 (d, <i>J</i> = 8.7 Hz, 1H), 7.25-7.17 (m, 2H), 7.03 (d, <i>J</i> = 1.6 Hz, 1H), 6.98-6.87 (m, 2H), 6.02 (s, 2H), 1.43-1.39 (m, 2H), 1.06-1.02 (m, 2H).

Example 84: 1-(Benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butyl-3-cyano-1<i>H</i>-indol-5-yl)cyclopropanecarboxamide

0578<chemistry id="chem0523" num="0523"><img file="EP2007756B1_D0549.tif" /></chemistry><chemistry id="chem0524" num="0524"><img file="EP2007756B1_D0550.tif" /></chemistry>

1-(Benzo[d][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butyl-3-formyl-1H-indol-5-yl)cyclopropane-carboxamide

0579POCl₃ (12 g, 80 mmol) was added dropwise to DMF (40 mL) held at -20 °C. After the addition was complete, the reaction mixture was allowed to warm to 0 °C and was stirred for 1 h. 1-(Benzo[d][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butyl-1H-indol-5-yl)cyclopropanecarboxamide (3.0 g, 8.0 mmol) was added and the mixture was warmed to 25 °C. After stirring for 30 minutes the reaction mixture was poured over ice and stirred for 2 h. The mixture was then heated at 100 °C for 30 min. The mixture was cooled and the solid precipitate was collected and washed with water. The solid was then dissolved in 200 mL dichloromethane and washed with 200 mL of a saturated aq. NaHCO₃. The organics were dried over Na₂SO₄ and evaporated to yield 1-(benzo[d][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butyl-3-formyl-1H-indol-5-yl)cyclopropane-carboxamide (2.0 g, 61%). ESI-MS <i>m</i>/<i>z</i> calc. 404.5, found 405.5 (M+1)⁺; retention time 3.30 minutes. ¹H NMR (400 MHz, DMSO-d6) δ 11.48 (s, 1H), 10.39 (s, 1H), 8.72 (s, 1H), 8.21 (s, 1H), 7.35-7.31 (m, 2H), 7.04-7.03 (m, 1H), 6.97-6.90 (m, 2H), 6.03 (s, 2H), 1.53 (s, 9H), 1.42-1.39 (m, 2H), 1.05-1.03 (m, 2H). <chemistry id="chem0525" num="0525"><img file="EP2007756B1_D0551.tif" /></chemistry>

(<i>Z</i>)-1-(Benzo[d][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butyl-3-((hydroxyimino)methyl)-1H-indol-5-yl)cyclopropanecarboxamide

0580To a solution of 1-(benzo[d][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butyl-3-formyl-1H-indol-5-yl)cyclopropanecarboxamide (100 mg, 0.25 mmol) in dichloromethane (5 mL) was added hydroxylamine hydrochloride (21 mg, 0.30 mmol). After stirring for 48 h, the mixture was evaporated to dryness and purified by column chromatography (0-100% ethyl acetate/hexanes) to yield (<i>Z</i>)-1-(benzo[d][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butyl-3-((hydroxyimino)methyl)-1H-indol-5-yl)cyclopropanecarboxamide (81 mg, 77%). ESI-MS <i>m</i>/<i>z</i> calc. 419.5, found 420.5 (M+1)⁺; retention time 3.42 minutes. ¹H NMR (400 MHz, DMSO-d6) δ 10.86 (s, 0.5H), 10.55 (s, 0.5H), 8.56-8.50 (m, 2H), 8.02 (m, 1H), 7.24-7.22 (m, 1H), 7.12-7.10 (m, 1H), 7.03 (m, 1H), 6.96-6.90 (m, 2H), 6.03 (s, 2H), 1.43 (s, 9H), 1.40-1.38 (m, 2H), 1.04-1.01 (m, 2H). <chemistry id="chem0526" num="0526"><img file="EP2007756B1_D0552.tif" /></chemistry>

1-(Benzo[d][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butyl-3-cyano-1H-indol-5-yl)cyclopropane-carboxamide

0581(<i>Z</i>)-1-(Benzo[d][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butyl-3-((hydroxyimino)-methyl)-1H-indol-5-yl)cyclopropanecarboxamide (39 mg, 0.090 mmol) was dissolved in acetic anhydride (1 mL) and heated at reflux for 3 h. The mixture was cooled in an ice bath and the precipitate was collected and washed with water. The solid was further dried under high vacuum to yield 1-(benzo[d][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butyl-3-cyano-1H-indol-5-yl)cyclopropanecarboxamide. ESI-MS <i>m</i>/<i>z</i> calc. 401.5, found 402.5 (M+1)⁺; retention time 3.70 minutes. ¹H NMR (400 MHz, DMSO-d6) δ 11.72 (s, 1H), 8.79 (s, 1H), 7.79 (s, 1H), 7.32 (m, 2H), 7.03-7.02 (m, 1H), 6.95-6.89 (m, 2H), 6.03 (s, 2H), 1.47 (s, 9H), 1.43-1.41 (m, 2H), 1.06-1.04 (m, 2H).

Example 85: 1-(Benzo[d][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butyl-3-methyl-1H-indol-5-yl)cyclopropanecarboxamide

0582<chemistry id="chem0527" num="0527"><img file="EP2007756B1_D0553.tif" /></chemistry>

0583A solution of 1-(benzo[d][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butyl-1H-indol-5-yl)cyclopropanecarboxamide (75 mg, 0.20 mmol) and iodomethane (125 µL, 2.0 mmol) in <i>N,N</i>-dimethylformamide (1 mL) was heated at 120 °C in a sealed tube for 24 h. The reaction was filtered and purified by reverse phase HPLC. ESI-MS <i>m</i>/<i>z</i> calc. 390.5, found 391.3 (M+1)⁺; retention time 2.04 minutes. ¹H NMR (400 MHz, DMSO-d6) δ 10.30 (s, 1H), 8.39 (s, 1H), 7.51 (m, 1H), 7.13-7.11 (m, 1H), 7.03-6.90 (m, 4H), 6.03 (s, 2H), 2.25 (s, 3H), 1.40-1.38 (m, 11H), 1.03-1.01 (m, 2H).

Example 86: 1-(Benzo[d][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butyl-3-(2-hydroxyethyl)-1H-indol-5-yl)cyclopropanecarboxamide

0584<chemistry id="chem0528" num="0528"><img file="EP2007756B1_D0554.tif" /></chemistry>

0585Approximately 100 µL of ethylene dioxide was condensed in a reaction tube at -78 °C. A solution of 1-(benzo[d][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butyl-1H-indol-5-yl)cyclopropanecarboxamide (200 mg, 0.50 mmol) and indium trichloride (20 mg, 0.10 mmol) in dichloromethane (2 mL) was added and the reaction mixture was irradiated in the microwave for 20 min at 100 °C. The volatiles were removed and the residue was purified by column chromatography (0-100 % ethyl acetate/hexanes) to give 1-(benzo[d][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butyl-3-(2-hydroxyethyl)-1H-indol-5-yl)cyclopropanecarboxamide (5 mg, 3%). ESI-MS <i>m</i>/<i>z</i> calc. 420.5, found 421.3 (M+1)⁺; retention time 1.67 minutes. ¹H NMR (400 MHz, CD₃CN) δ 8.78 (s, 1H), 7.40 (m, 1H), 7.33 (s, 1H), 7.08 (m, 1H), 6.95 - 6.87 (m, 3H), 6.79 (m, 1H), 5.91 (s, 2H), 3.51 (dd, J = 5.9, 7.8 Hz, 2H), 2.92 - 2.88 (m, 2H), 2.64 (t, J = 5.8 Hz, 1H), 1.50 (m, 2H), 1.41 (s, 9H), 1.06 (m, 2H).

Example 87: 2-(5-(1-(Benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-2-yl)acetic acid

0586<chemistry id="chem0529" num="0529"><img file="EP2007756B1_D0555.tif" /></chemistry>

0587To a solution of ethyl 2-(5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-2-yl)acetate (0.010 g, 0.025 mmol) in THF (0.3 mL) were added LiOH.H₂O (0.002 g, 0.05 mmol) and water (0.15 mL) were added. The mixture was stirred at room temperature for 2 h. dichloromethane (3 mL) was added to the reaction mixture and the organic layer was washed with 1 N HCl (2 x 1.5 mL) and water (2 x 1.5 mL). The organic layer was dried over Na₂SO₄ and filtered. The filtrate was evaporated under reduced pressure to give 2-(5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-2-yl)-acetic acid. ¹H NMR (400 MHz, DMSO-d6) δ 12.53 (s, 1H), 10.90 (s, 1H), 8.42 (s, 1H), 7.57 (s, 1H), 7.17 (d, J = 8.6 Hz, 1H), 7.05-6.90 (m, 4H), 6.17 (s, 1H), 6.02 (s, 2H), 3.69 (s, 2H), 1.41-1.39 (m, 2H), 1.04-1.02 (m, 2H).

Example 88: 5-(1-(Senzo[<i>d</i>][1,3]dioxol-5-yl)cyclopropanecarboxamido)-2-<i>tert-</i>butyl-1<i>H</i>-indole-7-carboxylic acid

0588<chemistry id="chem0530" num="0530"><img file="EP2007756B1_D0556.tif" /></chemistry>

0589Methyl 5-(1-(benzo[<i>d</i>][1,3]dioxol-5-yl)cyclopropanecarboxamido)-2-<i>tert</i>-butyl-1<i>H</i>-indole-7-carboxylate (30 mg, 0.069 mmol) was dissolved in a mixture of 1,4-dioxane (1.5 mL) and water (2 mL) containing a magnetic star bar and lithium hydroxide (30 mg, 0.71 mmol). The resulting solution was stirred at 70 °C for 45 minutes. The crude product was then acidified with 2.6 M hydrochloric acid and extracted three times with an equivalent volume of dichloromethane. The dichloromethane extracts were combined, dried over sodium sulfate, filtered, and evaporated to dryness. The residue was dissolved in a minimum of <i>N,N</i>-dimethylformamide and then purified by preparative HPLC using a gradient of 0-99% acetonitrile in water containing 0.05% trifluoroacetic acid to yield 5-(1-(benzo[<i>d</i>][1,3]dioxol-5-yl)cyclopropanecarboxamido)-2-<i>tert</i>-butyl-1<i>H</i>-indole-7-carboxylic acid. ESI-MS <i>m</i>/<i>z</i> calc. 434.2, found 435.5. Retention time of 1.85 minutes. ¹H NMR (400 MHz, DMSO-<i>d</i>6) δ 13.05 (s, 1H), 9.96 (d, <i>J</i> = 1.6 Hz, 1H), 7.89 (d, <i>J</i> = 1.9 Hz, 1H), 7.74 (d, <i>J</i> = 2.0 Hz, 1H), 7.02 (d, <i>J</i> = 1.6 Hz, 1H), 6.96-6.88 (m, 2H), 6.22 (d, <i>J</i> = 2.3 Hz, 1H), 6.02 (s, 2H), 1.43 - 1.40 (m, 2H), 1.37 (s, 9H), 1.06-1.02 (m, 2H).

Example 89: 1-(Benze[<i>d</i>][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butyl-1-(1,3-dihydroxypropan-2-yl)-1<i>H</i>-indol-5-yl)cyclopropanecarboxamide

0590<chemistry id="chem0531" num="0531"><img file="EP2007756B1_D0557.tif" /></chemistry>

1-(Benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butyl-1-(1,3-dihydroxypropan-2-yl)indolin-5-yl)cyclopropanecarboxamide

05911-(Benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butylindolin-5-yl)cyclopropanecarboxamide (50 mg, 0.13 mmol) was dissolved in dichloroethane (0.20 mL) and 2,2-dimethyl-1,3-dioxan-5-one (0.20 mL). Trifluoroacetic acid was added (0.039 mL) and the resulting solution was allowed to stir for 20 minutes. Sodium triacetoxyborohydride was added (55 mg, 0.26 mmol) and the reaction mixture was stirred for 30 minutes. The crude reaction mixture was then evaporated to dryness, dissolved in <i>N,N</i>-dimethylformamide and purified by preparative HPLC using a gradient of 0-99% acetonitrile in water containing 0.05% trifluoroacetic acid. <chemistry id="chem0532" num="0532"><img file="EP2007756B1_D0558.tif" /></chemistry>

1-(Benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butyl-1-(1,3-dihydroxypropan-2-yl)-1H-indol-5-yl)cyclopropanecarboxamide

05921-(Benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butyl-1-(1,3-dihydroxypropan-2-yl)indolin-5-yl)cyclopropanecarboxamide (40.3 mg, 0.0711 mmol as the trifluoracetic acid salt) was dissolved in toluene (1 mL). To the resulting solution was added 2,3,5,6-tetrachlorocyclohexa-2,5-diene-1,4-dione (35 mg, 0.14 mmol). The resulting suspension was heated at 100 °C in an oil bath for 10 minutes. The crude product was then evaporated to dryness, dissolved in a 1 mL of <i>N,N</i>-dimethylformamide and purified by purified by preparative HPLC using a gradient of 0-99% acetonitrile in water containing 0.05% trifluoroacetic acid to yield 1-(benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butyl-1-(1,3-dihydroxypropan-2-yl)-1<i>H</i>-indol-5-yl)cyclopropanecarboxamide. ESI-MS <i>m</i>/<i>z</i> calc. 450.2, found 451.5 (M+1)⁺. Retention time of 1.59 minutes.

Example 90: <i>N</i>-(7-(Aminomethyl)-2-<i>tert</i>-butyl-1<i>H</i>-indol-5-yl)-1-(benzo[<i>d</i>][1,3]-dioxol-5-yl)cyclopropanecarboxamide

0593<chemistry id="chem0533" num="0533"><img file="EP2007756B1_D0559.tif" /></chemistry>

<i>N</i>-(7-(Aminomethyl)-2-<i>tert</i>-butyl-1<i>H</i>-indol-5-yl)-1-(benzo[<i>d</i>][1,3]dioxol-5-yl)cyclopropanecarboxamide

05941-(Benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butyl-7-cyano-1<i>H</i>-indol-5-yl)cyclopropanecarboxamide (375 mg, 0.934 mmol) was dissolved in 35 mL of ethyl acetate. The solution was recirculated through a continuous flow hydrogenation reactor containing 10% palladium on carbon at 100 °C under 100 bar of hydrogen for 8 h. The crude product was then evaporated to dryness and purified on 12 g of silica gel utilizing a gradient of 0-100% ethyl acetate (containing 0.5% triethylamine) in hexanes to yield <i>N</i>-(7-(aminomethyl)-2-<i>tert</i>-butyl-1<i>H</i>-indol-5-yl)-1-(benzo[<i>d</i>][1,3]-dioxol-5-yl)-cyclopropanecarboxamide (121 mg, 32%). ESI-MS <i>m</i>/<i>z</i> calc. 405.2, found 406.5 (M+1)⁺. Retention time of 1.48 minutes.

Example 91: 5-(1-(Benzo[<i>d</i>][1,3]dioxol-5-yl)cyclopropanecarboxamido)-2-<i>tert-</i>butyl-1<i>H</i>-indole-7-carboxamide

0595<chemistry id="chem0534" num="0534"><img file="EP2007756B1_D0560.tif" /></chemistry>

5-(1-(Benzo[<i>d</i>][1,3]dioxol-5-yl)cyclopropanecarboxamido)-2-<i>tert</i>-butyl-1<i>H-</i>indole-7-carboxamide

05961-(Benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butyl-7-cyano-1<i>H</i>-indol-5-yl)-cyclopropanecarboxamide (45 mg, 0.11 mmol) was suspended in a mixture of methanol (1.8 mL), 30% aqueous hydrogen peroxide (0.14 mL, 4.4 mmol) and 10% aqueous sodium hydroxide (0.150 mL). The resulting suspension was stirred for 72 h at room temperature. The hydrogen peroxide was then quenched with sodium sulfite. The reaction mixture was diluted with 0.5 mL of <i>N,N</i>-dimethylformamide, filtered, and purified by preparative HPLC using a gradient of 0-99% acetonitrile in water containing 0.05% trifluoroacetic acid to yield 5-(1-(benzo[<i>d</i>][1,3]dioxol-5-yl)cyclopropane-carboxamido)-2-<i>tert</i>-butyl-1<i>H</i>-indole-7-carboxamide. ESI-MS <i>m</i>/<i>z</i> calc. 419.2, found 420.3 (M+1)⁺. Retention time of 1.74 minutes.

Example 92: 1-(Benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butyl-7-(methylsulfonamido-methyl)-1<i>H</i>-indol-5-yl)cyclopropanecarboxamide

0597<chemistry id="chem0535" num="0535"><img file="EP2007756B1_D0561.tif" /></chemistry>

1-(Benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N-</i>(2-<i>tert-</i>butyl-7-(methylsulfonamidomethyl)-1<i>H-</i>indol-5-yl)cyclopropanecarboxamide

0598<i>N</i>-(7-(Aminomethyl)-2-<i>tert-</i>butyl-1<i>H</i>-indol-5-yl)-1-(benzo[<i>d</i>][1,3]dioxol-5-yl)cyclopropanecarboxamide (20 mg, 0.049 mmol) was dissolved in DMF (0.5 mL) containing triethylamine (20.6 µL, 0.147 mmol) and a magnetic stir bar. Methanesulfonyl chloride (4.2 µL, 0.054 mmol) was then added to the reaction mixture. The reaction mixture was allowed to stir for 12 h at room temperature. The crude product was purified by preparative HPLC using a gradient of 0-99% acetonitrile in water containing 0.05% trifluoroacetic acid to yield 1-(benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N-</i>(2-<i>tert</i>-butyl-7-(methylsulfonamidomethyl)-1<i>H</i>-indol-5-yl)cyclopropanecarboxamide. ESI-MS <i>m</i>/<i>z</i> calc. 483.2, found 484.3 (M+1)⁺. Retention time of 1.84 minutes.

Example 93: <i>N</i>-(7-(Acetamidomethyl)-2-<i>tert</i>-butyl-1<i>H</i>-indol-5-yl)-1-(benzo[<i>d</i>][1,3]-dioxol-5-yl)cyclopropanecarboxamide

0599<chemistry id="chem0536" num="0536"><img file="EP2007756B1_D0562.tif" /></chemistry>

0600<i>N</i>-(7-(Aminomethyl)-2-<i>tert</i>-butyl-1<i>H</i>-indol-5-yl)-1-(benzo[<i>d</i>][1,3]dioxol-5-yl)cyclopropanecarboxamide (20 mg, 0.049 mmol) was dissolved in DMF (0.5 mL) containing triethylamine (20.6 µL, 0.147 mmol) and a magnetic stir bar. Acetyl chloride (4.2 µL, 0.054 mmol) was then added to the reaction mixture. The reaction mixture was allowed to stir for 16 h at room temperature. The crude product was purified by preparative HPLC using a gradient of 0-99% acetonitrile in water containing 0.05% trifluoroacetic acid to yield <i>N-</i>(7-(acetamidomethyl)-2-<i>tert</i>-butyl-1<i>H-</i>indol-5-yl)-1-(benzo[<i>d</i>][1,3]dioxol-5-yl)cyclopropanecarboxamide. ESI-MS <i>m</i>/<i>z</i> calc. 447.2, found 448.3 (M+1)⁺. Retention time of 1.76 minutes.

Example 94: <i>N</i>-(1-Acetyl-2-<i>tert</i>-butyl-1<i>H</i>-butyl-5-yl)-1-(benzo[d][1,3]dioxol-5-yl)-cyclopropanecarboxamide

0601<chemistry id="chem0537" num="0537"><img file="EP2007756B1_D0563.tif" /></chemistry>

0602To a solution of 1-(benzo[d][1,3]dioxol-5-yl)-<i>N-</i>(2-<i>tert</i>-butyl-1H-indol-5-yl)cyclopropanecarboxamide (120 mg, 0.31 mmol) in anhydrous DMF-THF (3.3 mL, 1:9) was added NaH (60% in mineral oil, 49 mg, 1.2 mmol) at room temperature. After 30 min under N₂, the suspension was cooled down to -15 °C and a solution of acetyl chloride (1.1 eq.) in DMF (0.5 mL) was added dropwise. The reaction mixture was stirred for 30 min at-15 °C then for 6 h at room temperature. Water (0.5 mL) was added at 0 °C, solvent was removed, and the residue was diluted with MeOH, filtrated and purified by preparative HPLC to give <i>N-</i>(1-acetyl-2-<i>tert</i>-butyl-1H-indol-5-yl)-1-(benzo[d][1,3]dioxol-5-yl)cyclo-propanecarboxamide. ¹H NMR (400 MHz, DMSO) δ 8.9 (s, 1H), 7.74 (d, <i>J</i> =2.1 Hz, 1H), 7.54 (d, <i>J</i> =9.0 Hz, 1H), 7.28 (dd, <i>J1</i> =2.1 Hz, <i>J2</i> =9.0 Hz, 1H), 7.01 (d, <i>J</i> =1.5 Hz, 1H), 6.93 (dd, <i>J1</i> =1.7 Hz, <i>J2</i> =8.0 Hz, 1H), 6.89 (d, J=8.0 Hz, 1H), 6.54 (bs, 1H), 6.02 (s, 2H), 2.80 (s, 3H), 1.42-1.40 (m, 11H), 1.06-1.05 (m, 2H). MS (ESI) m/e (M+H⁺) 419.3.

Example 95: <i>N</i>-(1-(2-Acetamidoethyl)-2-<i>tert</i>-butyl-6-fluoro-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

0603<chemistry id="chem0538" num="0538"><img file="EP2007756B1_D0564.tif" /></chemistry><chemistry id="chem0539" num="0539"><img file="EP2007756B1_D0565.tif" /></chemistry>

<i>N</i>-(1-(2-Aminoethyl)-2-<i>tert</i>-butyl-6-fluoro-1H-indol-5-yl)-1-(2,2-difluorobenzo-[d] [1,3]dioxol-5-yl)cyclopropanecarboxamide

0604To a solution of <i>tert</i>-butyl 2-(2-<i>tert</i>-butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-6-fluoro-1H-indol-1-yl)ethylcarbamate (620 mg, 1.08 mmol) in CH₂Cl₂ (8 mL) was added TFA (2 mL). The reaction was stirred at room temperature for 1.5 h before being neutralized with solid NaHCO₃. The solution was partitioned between H₂O and CH₂Cl₂. The organic layer was dried over MgSO₄ filtered and concentrated to yield the product as a cream colored solid (365 mg, 71%). ¹H NMR (400 MHz, DMSO-d6) δ 8.38 (s, 1H), 7.87 (br s, 3H, NH₃⁺), 7.52 (s, 1H), 7.45-7.38 (m, 3H), 7.32 (dd, J = 8.3, 1.5 Hz, 1H), 6.21 (s, 1H), 4.46 (m, 2H), 3.02 (m, 2H), 1.46 (m, 2H), 1.41 (s, 9H), 1.14 (m, 2H). HPLC ret. time 1.66 min, 10-99 % CH₃CN, 3 min run; ESI-MS 474.4 m/z (M+H⁺). <chemistry id="chem0540" num="0540"><img file="EP2007756B1_D0566.tif" /></chemistry>

<i>N</i>-(1-(2-Acetamidoethyl)-2-<i>tert</i>-butyl-6-fluoro-1H-indol-5-yl)-1-(2,2-difluorobenzo [d][1,3]dioxol-5-yl)cyclopropanecarboxamide

0605To a solution of <i>N</i>-(1-(2-aminoethyl)-2-<i>tert</i>-butyl-6-fluoro-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropane-carboxamide (47 mg, 0.10 mmol) and Et₃N (28 µL, 0.20 mmol) in DMF (1 mL) was added acetyl chloride (7.1 µL, 0.10 mmol). The mixture was stirred at room temperature for 1 h before being filtered and purified by reverse phase HPLC (10 - 99 % CH₃CN/ H₂O) to yield <i>N-</i>(1-(2-acetamidoethyl)-2-<i>tert</i>-butyl-6-fluoro-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide. ¹H NMR (400 MHz, DMSO-d6). δ 8.35 (s, 1H), 8.15 (t, J = 5.9 Hz, 1H), 7.53 (s, 1H), 7.43-7.31 (m, 4H), 6.17 (s, 1H), 4.22 (m, 2H), 3.30 (m, 2H), 1.85 (s, 3H), 1.47 (m, 2H), 1.41 (s, 9H), 1.13 (m, 2H). HPLC ret. time 2.06 min, 10-99 % CH₃CN, 3 min run; ESI-MS 516.4 m/z (M+H⁺).

Example 96: 1-(Benzo[d][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butyl-1-(2-hydroxy-3-methoxy-propyl)-1H-indol-5-yl)cyclopropenecarboxamide

0606<chemistry id="chem0541" num="0541"><img file="EP2007756B1_D0567.tif" /></chemistry>

06071-(Benzo[d][1,3]dioxol-5-yl)-<i>N-</i>(2-<i>tert</i>-butyl-1H-indol-5-yl)cyclopropanecarboxamide (320 mg, 0.84 mmol) was dissolved in a mixture composed of anhydrous DMF (0.5 mL) and anhydrous THF (5 mL) under N₂. NaH (60% in mineral oil, 120 mg, 3.0 mmol) was added at room temperature. After 30 min of stirring, the reaction mixture was cooled to -15 °C before a solution of epichlorohydrin (79 µL, 1.0 mmol) in anhydrous DMF (1 mL) was added dropwise. The reaction mixture was stirred for 15 min at -15°C, then for 8 h at room temperature. MeOH (1 mL) was added and the mixture was heated for 10 min at 105 °C in the microwave oven. The mixture was cooled, filtered and purified by preparative HPLC to give 1-(benzo[d][1,3]dioxol-5-yl)-<i>N-</i>(2-<i>tert</i>-butyl-1-(2-hydroxy-3-methoxy-propyl)-1H-indol-5-yl)cyclopropanecarboxamide. ¹H NMR (400 MHz, DMSO-d6) δ 8.44 (s, 1H), 7.59 (d, <i>J</i> = 1.9 Hz, 1H), 7.31 (d, J = 8.9 Hz, 1H), 7.03 (dd, <i>J</i> = 8.7, 1.9 Hz, 2H), 6.95 (dd, <i>J</i> = 8.0, 1.7 Hz, 1H), 6.90 (d, <i>J =</i> 8.0 Hz, 1H), 6.16 (s, 1H), 6.03 (s, 2H), 4.33 (dd, <i>J =</i> 15.0, 4.0 Hz, 1H), 4.19 (dd, <i>J =</i> 15.0, 8.1 Hz, 1H), 4.02 (ddd, <i>J</i> = 8.7, 4.8 Hz, 1H), 3.41-3.32 (m, 2H), 3.30 (s, 3H), 1.41 (s, 9H), 1.41-1.38 (m, 2H), 1.03 (dd, <i>J =</i> 6.7, 4.0 Hz, 2H). MS (ESI) m/e (M+H⁺) 465.0.

Example 97: 1-(Benzo[d][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert-</i>butyl-1-(2-hydroxy-3-(methyl-amino)propyl)-1H-indol-5-yl)cyclopropanecarboxamide

0608<chemistry id="chem0542" num="0542"><img file="EP2007756B1_D0568.tif" /></chemistry>

06091-(Benzo[d][1,3]dioxol-5-yl)-<i>N-</i>(2-<i>tert</i>-butyl-1H-indol-5-yl)cyclopropanecarboxamide (320 mg, 0.84 mmol) was dissolved in a mixture composed of anhydrous DMF (0.5 mL) and anhydrous THF (5 mL) under N₂. NaH (60% in mineral oil, 120 mg, 3.0 mmol) was added at room temperature. After 30 min of stirring, the reaction mixture was cooled to -15 °C before a solution of epichlorohydrin (79 µL, 1.0 mmol) in anhydrous DMF (1 mL) was added dropwise. The reaction mixture was stirred for 15 min at -15 °C, then for 8 h at room temperature. MeNH₂ (2.0 M in MeOH, 1.0 mL) was added and the mixture was heated for 10 min at 105 °C in the microwave oven. The mixture was cooled, filtered and purified by preparative HPLC to give 1-(benzo[d][1,3]dioxol-5-yl)-<i>N-</i>(2-<i>tert</i>-butyl-1-(2-hydroxy-3-(methylamino)propyl)-1H-indol-5-yl)cyclopropanecarboxamide. ¹H NMR (400 MHz, DMSO-d6) δ 8.50 (s, 1H), 7.60-7.59 (m, 1H), 7.35 (dd, J = 14.3, 8.9 Hz, 1H), 7.10 (d, J = 8.8 Hz, 1H), 1H), 6.94 (dd, J = 8.0, 1.6 Hz, 1H), 6.91 (d, J = 7.9 Hz, 1H), 6.20 (d, J = 2.3 Hz, 1H), 6.03 (s, 2H), 2.82 (d, J = 4.7 Hz, 1H), 2.72 (d, J = 4.7 Hz, 1H), 2.55 (dd, J = 5.2, 5.2 Hz, 1H), 2.50 (s, 3H), 1.43 (s, 9H), 1.39 (dd, J = 6.4, 3.7 Hz, 2H), 1.04 (dd, J = 6.5, 3.9 Hz, 2H). MS (ESI) m/e (M+H⁺) 464.0.

Example 98: (<i>S</i>)-<i>N</i>-(1-(3-Amino-2-hydroxypropyl)-2-<i>tert</i>-butyl-1H-indol-5-yl)-1-(2,2-difluorobenzo[d] [1,3]dioxol-5-yl)cyclopropanecarboxamide

0610<chemistry id="chem0543" num="0543"><img file="EP2007756B1_D0569.tif" /></chemistry><chemistry id="chem0544" num="0544"><img file="EP2007756B1_D0570.tif" /></chemistry>

(<i>R</i>)-3-(2-<i>tert</i>-Butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarbox-amido)-1H-indol-1-yl)-2-hydroxypropyl-4-methylbenzenesulfonate

0611To a stirred solution of (<i>R</i>)-<i>N-</i>(2-<i>tert</i>-butyl-1-(2,3-dihydroxypropyl)-1H-indol-5-yl)-1-(2,2-difluoro-benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide (3.0 g, 6.1 mmol) in dichloromethane (20 mL) was added triethylamine (2 mL) and para-toluenesulfonylchloride (1.3 g, 7.0 mmol). After 18 hours, the reaction mixture was partitioned between 10 mL of water and 10 mL of ethyl acetate. The organic layer was dried over magnesium sulfate, filtered and evaporated. The residue was purified using column chromatography on silica gel (0-60% ethyl acetate/hexane) providing (<i>R</i>)-3-(2-<i>tert</i>-butyl-5-(1-(2,2-difluorobenzo[d][1,3]-dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-1-yl)-2-hydroxypropyl-4-methyl-benzenesulfonate (3.21 g, 86%). LC/MS (M + 1) = 641.2. ¹H NMR (400 MHz, CDCl₃) δ 7.77 (d, 2H, <i>J</i> = 16 Hz), 7.55 (d, 1H, <i>J</i> = 2 Hz), 7.35 (d, 2H, J = 16 Hz), 7.31 (m, 3H), 6.96 (s, 1H), 6.94 (dd, 1H, <i>J</i> = 2, 8 Hz), 6.22 (s, 1H), 4.33 (m, 1H), 4.31 (dd, 1H, <i>J =</i> 6, 15 Hz), 4.28 (dd, 1H, <i>J =</i> 11<i>,</i> 15 Hz), 4.18 (m, 1H), 3.40 (dd, 1H, <i>J</i> = 3, 6 Hz), 3.36 (dd, 1H<i>, J =</i> 3, 6 Hz<i>),</i> 2.46 (s, 3H), 2.40 (br s, 1H), 1.74 (m, 2H), 1.40 (s, 9H), 1.11 (m, 2 H). <chemistry id="chem0545" num="0545"><img file="EP2007756B1_D0571.tif" /></chemistry>

(<i>R</i>)-<i>N</i>-(1-(3-Azido-2-hydroxypropyl)-2-<i>tert</i>-butyl-1H-indol-5-yl)-1-(2,2-difluorobenzo [d][1,3]dioxol-5-yl)cyclopropanecarboxamide

0612To a stirred solution (<i>R</i>)-3-(2-<i>tert</i>-butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1 H-indol-1-yl)-2-hydroxypropyl-4-methylbenzenesulfonate (3.2 g, 5.0 mmol) in DMF (6 mL) was added sodium azide (2.0 g, 30 mmol). The reaction was heated at 80 °C for 2 h. The mixture was partitioned between 20 mL ethyl acetate and 20 mL water. The layers were separated and the organic layer was evaporated. The residue was purified using column chromatography (0-85% ethyl acetate/hexane) to give (<i>R</i>)-<i>N-</i>(1-(3-azido-2-hydroxypropyl)-2-<i>tert</i>-butyl-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)-cyclopropanecarboxamide (2.48 g). LC/MS (M + 1) = 512.5. ¹H NMR (400 MHz, CDCl₃) δ 7.55 (d, 1H, <i>J =</i> 2 Hz), 7.31 (m, 3H), 6.96 (s, 1H), 6.94 (dd, 1H, <i>J =</i> 2, 8 Hz), 6.22 (s, 1H), 4.33 (m, 1H), 4.31 (dd, 1H, <i>J =</i> 6, 15 Hz), 4.28 (dd, 1H, <i>J =</i> 11, 15 Hz), 4.18 (m, 1H), 3.40 (dd, 1H, J = 3, 6 Hz), 3.36 (dd, 1H, <i>J =</i> 3, 6 Hz), 2.40 (br s, 1H), 1.74 (m, 2H), 1.40 (s, 9H), 1.11 (m, 2H). <chemistry id="chem0546" num="0546"><img file="EP2007756B1_D0572.tif" /></chemistry>

(<i>S</i>)-<i>N</i>-(1-(3-Amino-2-hydroxypropyl)-2-<i>tert</i>-butyl-1H-indol-5-yl)-1-(2,2-difluoro-benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxanude

0613To a stirred solution (<i>R</i>)-<i>N</i>-(1-(3-azido-2-hydroxypropyl)-2-<i>tert</i>-butyl-1H-indol-5-yl)-1-(2,2-difluorobenzo [d][1,3]dioxol-5-yl)cyclopropanecarboxamide (2.4 g, 4.0 mmol) in MeOH (25 mL) was added 5 % Pd/C (2.4 g) under a Hydrogen gas filled balloon. After 18 h, the reaction mixture was filtered through celite and rinsed with 300 mL ethyl acetate. The organic layer was washed with 1 N HCl and evaporated to give (<i>S</i>)<i>-N</i>-(1-(3-amino-2-hydroxypropyl)-2-<i>tert</i>-butyl-1H-indol-5-yl)-1-(2,2-difluoro-benzo[d][1,3]-dioxol-5-yl)cyclopropane-carboxamide (1.37 g). MS (M + 1) = 486.5.

Example 99: (<i>S</i>)-Methyl 3-(2-<i>tert</i>-butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-1-yl)-2-hydroxypropylcarbamate

0614<chemistry id="chem0547" num="0547"><img file="EP2007756B1_D0573.tif" /></chemistry>

0615To a stirred solution (<i>R</i>)-<i>N-</i>(1-(3-amino-2-hydroxypropyl)-2-<i>tert</i>-butyl-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide (0.10 g, 0.20 mmol) in methanol (1 mL) was added 2 drops of triethylamine and methylchloroformyl chloride (0.020 mL, 0.25 mmol). After 30 min, the reaction mixture was filtered and purified using reverse phase HPLC providing (<i>S</i>)-methyl 3-(2-<i>tert</i>-butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclo-propanecarboxamido)-1H-indol-1-yl)-2-hydroxypropylcarbamate. The retention time on a three minute run is 1.40 min. LC/MS (M + 1) = 544.3. ¹H NMR (400 MHz, CDCl₃) δ 7.52 (d, 1H, <i>J =</i> 2Hz), 7.30 (dd, 1H, <i>J =</i> 2, 8 Hz), 7.28(m, 1H), 7.22 (d, 1H, <i>J =</i> 8 Hz), 7.14 (d, 1H, <i>J =</i> 8 Hz), 7.04 (br s, 1H), 6.97 (dd, 1H, <i>J</i> = 2, 8 Hz), 6.24 (s, 1H), 5.19 (1H, br s), 4.31 (dd, 1H, <i>J =</i> 6, 15 Hz), 4.28 (dd, 1H, <i>J =</i> 11, 15 Hz), 4.18 (m, 1H), 3.70 (s, 3H), 3.40 (dd, 1H, <i>J</i> = 3, 6 Hz), 3.36 (dd, 1H, <i>J =</i> 3, 6 Hz), 3.26 (m, 1H), 1.74 (m, 2H), 1.40 (s, 9 H), 1.11 (m, 2 H).

Example 100: 4-(5-(1-(Benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-2-<i>tert-</i>butyl-1H-indol-1-yl)butanoic acid

0616<chemistry id="chem0548" num="0548"><img file="EP2007756B1_D0574.tif" /></chemistry><chemistry id="chem0549" num="0549"><img file="EP2007756B1_D0575.tif" /></chemistry>

1-(Benzo[d][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butylindolin-5-yl)cyclopropanecarboxamide

0617To a solution of 1-(benzo[d][1,3]dioxol-5-yl)-<i>N-</i>(2-<i>tert</i>-butyl-1H-indol-5-yl)cyclo-propanecarboxamide (851 mg, 2.26 mmol) in acetic acid (60 mL) was added NaBH₃CN (309 mg, 4.91 mmol) at 0 °C. The reaction mixture was stirred for 5 min at room temperature after which no starting material could be detected by LCMS. The solvent was evaporated under reduced pressure and the residue was purified by column chromatography on silica gel (5-40% ethyl acetate/hexanes) to give 1-(benzo[d][1,3]dioxol-5-yl)-<i>N-</i>(2-<i>tert</i>-butylindolin-5-yl)cyclopropanecarboxamide (760 mg, 89%). <chemistry id="chem0550" num="0550"><img file="EP2007756B1_D0576.tif" /></chemistry>

4-(5-(1-(Benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-2-<i>tert-</i>butylindolin-1-yl)butanoic acid

0618To a solution of 1-(benzo[d][1,3]dioxol-5-yl)-<i>N-</i>(2-<i>tert</i>-butylindolin-5-yl)cyclopropanecarboxamide (350 mg, 0.93 mmol, 1 eq) in anhydrous methanol (6.5 mL) and AcOH (65 µL) was added 4-oxobutanoic acid (15% in water, 710 mg, 1.0 mmol) at room temperature. After 20 min of stirring, NaBH₃CN (130 mg, 2.0 mmol) was added in one portion and the reaction mixture was stirred for another 4 h at room temperature. The reaction mixture was quenched by addition of AcOH (0.5 mL) at 0 °C and the solvent was removed under reduced pressure. The residue was purified by column chromatography on silica gel (5-75% ethyl acetate/hexanes) to give 4-(5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-2-<i>tert</i>-butylindolin-1-yl)butanoic acid (130 mg, 30%). <chemistry id="chem0551" num="0551"><img file="EP2007756B1_D0577.tif" /></chemistry>

4-(5-(1-(Benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-2-<i>tert</i>-butyl-1H-indol-1-yl)butanoic acid

06194-(5-(1-(Benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-2-<i>tert</i>-butylindolin-1-yl)butanoic acid (130 mg, 0.28 mmol) was taken up in a mixture of acetonitrile-H₂O-TFA. The solvent was removed under reduced pressure and the residue obtained was dissolved in CDCl₃. After a brief exposition to daylight (5-10 min), the solution turned purple. The mixture was stirred open to the atmosphere at room temperature until complete disappearance of the starting material (8 h). Solvent was removed under reduced pressure and the residue was purified by reverse pharse HPLC to give 4-(5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-2-tert-butyl-1H-indol-1-yl)butanoic acid. ¹H NMR (400 MHz, CDCl₃) δ 7.52 (d, J = 1.9 Hz, 1H), 7.18 (d, J = 2.1 Hz, 1H), 7.16 (s, 1H), 7.03 (dd, J = 9.4, 1.9 Hz, 1H), 7.00-6.98 (m, 2H), 6.85 (d, J = 7.9 Hz, 1H), 6.16 (s, 1H), 6.02 (s, 2H), 4.29-4.24 (m, 2H), 2.48 (dd, J = 6.9, 6.9 Hz, 2H), 2.12-2.04 (m, 2H), 1.69 (dd, J = 6.8, 3.7 Hz, 2H), 1.43 (s, 9H), 1.09 (dd, J = 6.8, 3.7 Hz, 2H). MS (ESI) m/e (M+H⁺) 463.0.

Example 101: 1-(Benzo[d][1,3]dioxol-5-yl)-<i>N-</i>(2-<i>tert-</i>butyl-1-(4-(2-hydroxyethyl-amino)-4-oxobutyl)-1H-indol-5-yl)cyclopropanecarboxamide

0620<chemistry id="chem0552" num="0552"><img file="EP2007756B1_D0578.tif" /></chemistry>

0621To a solution of 4-(5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-2-tert-butyl H-indol-1-yl)butanoic acid (10 mg) in anhydrous DMF (0.25 mL) were successively added Et₃N (9.5 mL, 0.069 mmol) and HBTU (8.2 mg, 0.022 mmol). After stirring for 10 min at 60 °C, ethanolamine (1.3 µL, 0.022 mmol) was added, and the mixture was stirred for another 4 h at 60 °C. 1-(Benzo[d][1,3]dioxol-5-yl)-<i>N-</i>(2-<i>tert</i>-butyl-1-(4-(2-hydroxyethyl-amino)-4-oxobutyl)-1H-indol-5-yl)cyclopropanecarboxamide (5.8 mg, 64%) was obtained after purification by preparative HPLC. MS (ESI) m/e (M+H⁺) 506.0.

Example 102: 1-(Benzo[d][1,3]dioxol-5-yl)-<i>N-</i>(2-<i>tert</i>-butyl-1-(2-(dimethylamino)-2-oxoethyl)-1H-indol-5-yl)cyclopropanecarboxamide

0622<chemistry id="chem0553" num="0553"><img file="EP2007756B1_D0579.tif" /></chemistry>

0623To a solution of 1-(benzo[d][1,3]dioxol-5-yl)-<i>N-</i>(2-<i>tert</i>-butylindolin-5-yl)cyclopropanecarboxamide (62 mg, 0.16 mmol) in anhydrous DMF (0.11 mL) and THF (1 mL) was added NaH (60% in mineral oil, 21 mg, 0.51 mmol) at room temperature under N₂. After 30 min of stirring, the reaction mixture was cooled to 0 °C and 2-chloro-N,N-dimethylacetamide (11 mL, 0.14 mmol,) was added. The reaction mixture was stirred for 5 min at 0 °C and then for 10 h at room temperature. The mixture was purified by preparative HPLC and the resultant solid was dissolved in DMF (0.6 mL) in the presence of Pd-C (10 mg). The mixture was stirred open to the atmosphere overnight at room temperature. The reaction mixture was filtrated and purified by preparative HPLC providing 1-(benzo[d][1,3]dioxol-5-yl)-<i>N-</i>(2-<i>tert</i>-butyl-1-(2-(dimethylamino)-2-oxoethyl)-1H-indol-5-yl)cyclopropanecarboxamide. MS (ESI) m/e (M+H⁺) 462.0.

Example 103: 3-(2-<i>tert</i>-Butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclo-propanecarboxamido)-1H-indol-1-yl)propanoic acid

0624<chemistry id="chem0554" num="0554"><img file="EP2007756B1_D0580.tif" /></chemistry><chemistry id="chem0555" num="0555"><img file="EP2007756B1_D0581.tif" /></chemistry>

<i>N-</i>(2-<i>tert-</i>Butyl-1-(2-chloroethyl)indolin-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

0625To a solution of <i>N</i>-(2-<i>tert</i>-butyl-1-(2-cyanoethyl)indolin-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide (71 mg, 0.17 mmol) in anhydrous dichloromethane (1 mL) was added chloroacetaldehyde (53 µL, 0.41 mmol) at room temperature under N₂. After 20 min of stirring, NaBH(OAc)₃ (90 mg, 0.42 mmol) was added in two portions. The reaction mixture was stirred overnight at room temperature. The product was purified by column chromatography on silica gel (2-15% ethyl acetate/hexanes) providing <i>N-</i>(2-<i>tert</i>-butyl-1-(2-chloroethyl)indolin-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide (51 mg, 63%). <chemistry id="chem0556" num="0556"><img file="EP2007756B1_D0582.tif" /></chemistry>

N-

<i>(</i>2<i>-tert-</i>Butyl-1-(2-cyanoethyl)indolin-5-yl)-1-(2,2-difluorobenzo[d] [1,3]dioxol-5-yl)cyclopropanecarboxamide

0626<i>N</i>-(2-<i>tert</i>-butyl-1-(2-chloroethyl)indolin-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide (51 mg), NaCN (16 mg, 0.32 mmol) and KI (cat) in EtOH (0.6 mL) and water (0.3 mL) were combined and heated at 110 °C for 30 min in the microwave. The solvent was removed under reduced pressure and the residue was purified by column chromatography on silica gel (2-15% ethyl acetate/hexanes) providing <i>N-</i>(2<i>-tert-</i>butyl-1-(2-cyanoethyl)indolin-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide (24 mg, 48%). <chemistry id="chem0557" num="0557"><img file="EP2007756B1_D0583.tif" /></chemistry>

3-(2-<i>tert</i> Butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclo-propanecarbox-amido)-1H-indol-1-yl)propanoic acid

0627<i>N</i>-(2-<i>tert</i>-butyl-1-(2-cyanoethyl)indolin-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropane-carboxamide (24 mg, 0.050 mmol) was taken up in 50% aq. KOH (0.5 mL) and 1,4-dioxane (1 mL). The mixture was heated at 125 °C for 2 h. The solvent was removed and the residue was purified by preparative HPLC. The residue was dissolved in CDCl₃ (1 mL) then briefly exposed to daylight. The purple solution that formed was stirred until complete disappearance of the starting material (1 h). The solvent was removed under reduced pressure and the residue was purified by preparative HPLC providing 3-(2-tert-butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclo-propanecarboxamido)-1H-indol-1-yl)propanoic acid. MS (ESI) m/e (M+H⁺) 485.0.

Example 104: 1-(Benzo[d][1,3]dioxol-5-yl)-<i>N-</i>(2-<i>tert-</i>butyl-6-fluoro-1-(2-hydroxy-ethyl)-1H-indol-5-yl)cyclopropenecarboxamide

0628<chemistry id="chem0558" num="0558"><img file="EP2007756B1_D0584.tif" /></chemistry>

0629To a solution of 1-(benzo[d][1,3]dioxol-5-yl)-<i>N-</i>(2-<i>tert</i>-butyl-6-fluoroindolin-5-yl)cyclopropanecarboxamide (340 mg, 0.86 mmol) in anhydrous MeOH (5.7 mL) containing 1% of acetic acid was added glyoxal 40% in water (0.60 mL, 5.2 mmol) at room temperature under N₂. After 20 min of stirring, NaBH₃CN (120 mg, 1.9 mmol) was added in one portion and the reaction mixture was stirred overnight at room temperature. The solvent was removed under reduced pressure and the residue obtained was purified by column chromatography on silica gel (10-40% ethyl acetate/hexanes) providing a pale yellow oil which was treated with 50/50 CH₃CN-H₂O containing 0.05% TFA and CDCl₃. Solvent was removed under reduced pressure and the residue was purified by column chromatography on silica gel (20-35% ethyl acetate/hexanes) to give 1-(benzo[d][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert-</i>butyl-6-fluoro-1-(2-hydroxyethyl)-1H-indol-5-yl)cyclopropanecarboxamide. ¹H NMR (400 MHz, CDCl₃) δ 8.02 (d, J = 7.7 Hz, 1H), 7.30 (d, J = 2.1 Hz, 1H), 6.93 (dd, J = 1.6, 7.9 Hz, 1H), 6.90 (d, J = 1.6 Hz, 1H), 6.90 (d, J = 1.6 Hz, 1H), 6.78 (d, J = 7.9 Hz, 1H), 6.08 (s, 1H), 5.92 (s, 2H), 4.21 (dd, J = 6.9, 6.9 Hz, 2H), 3.68 (m, 2H), 2.28 (s, 1H), 1.60 (dd, J = 3.7, 6.7 Hz, 2H), 1.35 - 1.32 (m, 9H), 1.04 (dd, J = 3.7, 6.8 Hz, 2H). MS (ESI) m/e (M+H⁺) 439.0.

Example 105: 1-(Benzo[d][1,3]dioxol-5-yl)-<i>N-</i>(2-<i>tert-</i>butyl-6-fluoro-1-(3-hydroxy-propyl)-1H-indol-5-yl)cyclopropanecarboxamide

0630<chemistry id="chem0559" num="0559"><img file="EP2007756B1_D0585.tif" /></chemistry><chemistry id="chem0560" num="0560"><img file="EP2007756B1_D0586.tif" /></chemistry>

3-(Benzyloxy)propanal

0631To a suspension of PCC (606 mg, 2.82 mmol) in anhydrous dichloromethane (8 mL) at room temperature under N₂ was added a solution of 3-benzyloxy-1-propanol (310 mg, 1.88 mmol) in anhydrous dichloromethane. The reaction mixture was stirred overnight at room temperature, filtrated through Celite, and concentrated. The residue was purified by column chromatography on silica gel (1-10% ethyl acetate/hexanes) to give 3-(benzyloxy)propanal (243 mg, 79%). <chemistry id="chem0561" num="0561"><img file="EP2007756B1_D0587.tif" /></chemistry>

1-(Benzo[d][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butyl-6-fluoro-1-(3-hydroxypropyl)-1H-indol-5-yl)cyclopropanecarboxamide

0632To a solution of 1-(benzo[d][1,3]dioxol-5-yl)-<i>N-</i>(2-<i>tert</i>-butyl-6-fluoroindolin-5-yl)cyclopropanecarboxamide (160 mg, 0.50 mmol) in anhydrous dichloromethane (3.4 mL) was added 3-(benzyloxy)propanal (160 mg, 0.98 mmol) at room temperature. After 10 min of stirring, NaBH(OAc)₃ (140 mg, 0.65 mmol) was added in one portion and the reaction mixture was stirred for 4 h at room temperature. The solvent was removed under reduced pressure and the residue was taken-up in a mixture of 50/50 CH₃CN-H₂O containing 0.05% TFA. The mixture was concentrated to dryness and the residue was dissolved in CDCl₃ (5 mL) and briefly exposed to daylight. The purple solution was stirred open to the atmosphere at room temperature for 2 h. The solvent was removed under reduced pressure and the residue was treated with Pd-C (10 mg) in MeOH (2 mL) under 1 atm of H₂ for 2 h. The catalyst was filtered through Celite and the solvent was removed under reduced pressure. The residue was purified by preparative TLC 30% ethyl acetate/hexanes to provide 1-(benzo[d][1,3]dioxol-5-yl)-<i>N-</i>(2-<i>tert</i>-butyl-6-fluoro-1-(3-hydroxypropyl)-1H-indol-5-yl)cyclopropanecarboxamide (18 mg, 8% from 1-(benzo[d][1,3]dioxol-5-yl)-<i>N-</i>(2-<i>tert</i>-butyl-6-fluoroindolin-5-yl)cyclopropane-carboxamide). ¹H NMR (400 MHz, CDCl₃) δ 8.11 (d, J = 7.8 Hz, 1H), 7.31 (d, J = 2.2 Hz, 1H), 6.94 (dd, J = 7.9, 1.7 Hz, 1H), 6.91 (d, J = 1.6 Hz, 1H), 6.85 (d, J = 11.7 Hz, 1H), 6.79 (d, J = 7.9 Hz, 1H), 6.10 (s, 1H), 5.94 (s, 2H), 4.25-4.21 (m, 2H), 3.70 (dd, J = 5.7, 5.7 Hz, 2H), 1.93-1.86 (m, 2H), 1.61 (dd, J = 6.8, 3.7 Hz, 2H), 1.35 (s, 9H), 1.04 (dd, J = 6.8, 3.7 Hz, 2H). MS (ESI) m/e (M+H⁺) 453.0.

Example 106: <i>N</i>-(1-(2-Acetamidoethyl)-2-<i>tert</i>-butyl-1H-indol-5-yl)-1-(benzo[d] [1,3]-dioxol-5-yl)cyclopropanecarboxamide

0633<chemistry id="chem0562" num="0562"><img file="EP2007756B1_D0588.tif" /></chemistry><chemistry id="chem0563" num="0563"><img file="EP2007756B1_D0589.tif" /></chemistry>

<i>N</i>-(1-(2-azidoethyl)-2-<i>tert</i>-butyl-1H-indol-5-yl)-1-(benzo[d][1,3]dioxol-5-yl)-cyclopropanecarboxamide

0634To a solution of 1-(benzo[d][1,3]dioxol-5-yl)-<i>N-</i>(2-<i>tert</i>-butylindolin-5-yl)cyclopropane-carboxamide (73 mg, 0.19 mmol) in anhydrous dichloromethane (1.2 mL) was added chloroacetaldehyde (60 µL, 0.24 mmol) at room temperature. After 10 min of stirring, NaBH(OAc)₃ (52 mg, 0.24 mmol) was added in one portion and the reaction mixture was stirred for another 30 min at room temperature. The solvent was removed under reduced pressure and the residue was purified by preparative HPLC to give the indoline, which oxidized to the corresponding indole when taken-up in CDCl₃. The resulting indole was treated with NaN₃ (58 mg, 0.89 mmol) and NaI (cat) in anhydrous DMF (0.8 mL) for 2 h at 85 °C. The reaction mixture was purified by preparative HPLC providing <i>N</i>-(1-(2-azidoethyl)-2-<i>tert</i>-butyl-1H-indol-5-yl)-1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide (15 mg, 18% from 1-(benzo[d][1,3]dioxol-5-yl)-<i>N-</i>(2-<i>tert-</i>butylindolin-5-yl)cyclopropane-carboxamide). <chemistry id="chem0564" num="0564"><img file="EP2007756B1_D0590.tif" /></chemistry>

<i>N</i>-(1-(2-Acetamidoethyl)-2-<i>tert</i>-butyl-1H-indol-5-yl)-1-(benzo[d][1,3]-dioxol-5-yl)cyclopropanecarboxamide

0635A solution of <i>N</i>-(1-(2-azidoethyl)-2-<i>tert</i>-butyl-1H-indol-5-yl)-1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide (13 mg, 0.029 mmol) in MeOH-AcOH (0.2 mL, 99:1) in the presence of Pd-C (2 mg) was stirred at room temperature under 1 atm of H₂ for 2 h, filtered through Celite, and concentrated under reduced pressure. The crude product was treated with AcCl (0.05 mL) and Et₃N (0.05 mL) in anhydrous THF (0.2 mL) at 0 °C for 30 min and then 1 h at room temperature. The mixture was purified by preparative HPLC providing <i>N-</i>(1-(2-acetamidoethyl)-2-<i>tert</i>-butyl-1H-indol-5-yl)-1-(benzo[d][1,3]-dioxol-5-yl)cyclopropanecarboxamide. MS (ESI) m/e (M+H⁺) 462.0.

Example 107: <i>N</i>-(2-<i>tert</i>-Butyl-1-(3-cyano-2-hydroxypropyl)-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

0636<chemistry id="chem0565" num="0565"><img file="EP2007756B1_D0591.tif" /></chemistry><chemistry id="chem0566" num="0566"><img file="EP2007756B1_D0592.tif" /></chemistry>

3-(2-<i>tert-</i>Butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarbox-amido)-1H-indol-1-yl)-2-hydroxypropyl-4-methylbenzenesulfonate

0637To a solution of <i>N</i>-(2-<i>tert-</i>butyl-1-(2,3-dihydroxypropyl)-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]-dioxol-5-yl)cyclopropanecarboxamide (172 mg, 0.35 mmol) in anhydrous dichloromethane (1.4 mL) at 0 °C in the presence of Et₃N (56 µL, 0.40 mmol) was added TsCl (71 mg, 0.37 mmol). The reaction mixture was stirred for 2 h at room temperature before being cooled to 0 °C and another portion of TsCl (71 mg, 0.37 mmol) was added. After 1 h of stirring at room temperature, the mixture was purified by column chromatography on silica gel (10-30% ethyl acetate/hexanes) providing 3-(2-tert-butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-1-yl)-2-hydroxypropyl-4-methylbenzene-sulfonate (146 mg, 64%). <chemistry id="chem0567" num="0567"><img file="EP2007756B1_D0593.tif" /></chemistry>

<i>N</i>-(2-<i>tert</i>-Butyl-1-(3-cyano-2-hydroxypropyl)-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

0638<i>N</i>-(2-<i>tert</i>-Butyl-1-(3-cyano-2-hydroxypropyl)-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)-cyclopropanecarboxamide (145 mg, 0.226 mmol) was treated with powdered NaCN (34 mg, 0.69 mmol) in anhydrous DMF (1.5 mL) at 85 °C for 2 h. The reaction mixture was cooled down to room temperature before it was diluted with dichloromethane (10 mL) and aq. sat. NaHCO₃ (10 mL). The organic phase was separated and the aqueous phase was extracted with dichloromethane (2 × 10 mL). The organic phases were combined, washed with brine, dried with sodium sulfate, filtered then concentrated. The residue was purified by column chromatography on silica gel (25-55% ethyl acetate/hexanes) providing <i>N-</i>(2-<i>tert</i>-butyl-1-(3-cyano-2-hydroxypropyl)-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide (89 mg, 79%). ¹H NMR (400 MHz, CDCl₃) δ 7.43 (d, J = 1.9 Hz, 1H), 7.20-7.16 (m, 2H), 7.08 (d, J = 8.8 Hz, 1H), 7.04 (d, J = 8.2 Hz, 1H), 6.94 (s, 1H), 6.88 (dd, J = 8.7, 2.0 Hz, 1H), 6.16 (s, 1H), 4.32-4.19 (m, 3H), 2.83 (s, 1H), 2.40 (dd, J = 5.2, 5.2 Hz, 2H), 1.62 (dd, J = 6.6, 3.6 Hz, 2H), 1.35 (s, 9H), 1.04 (dd, J = 6.9, 3.9 Hz, 2H). MS (ESI) m/e (M+H⁺) 496.0.

Example 108: <i>N</i>-(2-<i>tert</i>-Butyl-1-(2-hydroxy-3-(2H-tetrazol-5-yl)propyl)-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

0639<chemistry id="chem0568" num="0568"><img file="EP2007756B1_D0594.tif" /></chemistry>

0640To a solution of <i>N</i>-(2-<i>tert</i>-butyl-1-(3-cyano-2-hydroxypropyl)-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide (27 mg, 0.054 mmol) in anhydrous DMF (1.2 mL) were successively added NH₄Cl (35 mg, 0.65 mmol) and NaN₃ (43 mg, 0.65 mmol) at room temperature. The reaction mixture was stirred for 4 h at 110 °C in the microwave, at which stage 50% of the starting material was converted to the desired product. The reaction mixture was purified by preparative HPLC to provide <i>N</i>-(2-<i>tert</i>-butyl-1-(2-hydroxy-3-(2H-tetrazol-5-yl)propyl)-1H-indol-5-yl)-1-(2,2-difluorobenzo-[d][1,3]dioxol-5-yl)cyclopropanecarboxamide. MS (ESI) m/e (M+H⁺) 539.0.

Example 109: 4-(2-<i>tert-</i>Butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclo-propanecarboxamido)-1H-indol-1-yl)-3-hydroxybutanoic acid

0641<chemistry id="chem0569" num="0569"><img file="EP2007756B1_D0595.tif" /></chemistry>

0642A solution of <i>N</i>-(2-<i>tert</i>-butyl-1-(3-cyano-2-hydroxypropyl)-1H-indol-5-yl)-1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide (14 mg, 0.028 mmol) in methanol (0.8 mL) and 4 M NaOH (0.8 mL) was stirred at 60 °C for 4 h. The reaction mixture was neutralized with 4 M HCl and concentrated. The residue was purified by preparative HPLC to provide 4-(2-<i>tert</i>-butyl-5-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1<i>H</i>-indol-1-yl)-3-hydroxybutanoic acid. MS (ESI) m/e (M+H⁺) 515.0.

Example 110: <i>N-</i>(1-(2-(2H-Tetrazol-5-yl)ethyl)-2-<i>tert-</i>butyl-1H-indol-5-yl)-1-(benzo[d] [1,3] dioxol-5-yl)cyclopropanecarboxamide

0643<chemistry id="chem0570" num="0570"><img file="EP2007756B1_D0596.tif" /></chemistry><chemistry id="chem0571" num="0571"><img file="EP2007756B1_D0597.tif" /></chemistry>

1-(Benzo[d][1,3]dioxol-5-yl)-<i>N-</i>(2-<i>tert-</i>butyl-1-(2-cyanoethyl)indolin-5-yl)-cyclopropanecarboxamide

0644To a solution of 1-(benzo[d][1,3]dioxol-5-yl)-<i>N-</i>(2-<i>tert</i>-butyl-1-(2-chloroethyl)indolin-5-yl)cyclopropanecarboxamide (66 mg, 0.15 mmol) in ethanol (0.8 mL) and water (0.4 mL) were added NaCN (22 mg, 0.45 mmol) and KI (cat) at room temperature. The reaction mixture was stirred for 30 min at 110 °C in the microwave before being purified by column chromatography on silica gel (5-15% ethyl acetate/hexanes) to provide 1-(benzo[d][1,3]dioxol-5-yl)-<i>N</i>-(2-<i>tert</i>-butyl-1-(2-cyano-ethyl)indolin-5-yl)cyclopropanecarboxamide (50 mg, 77%). <chemistry id="chem0572" num="0572"><img file="EP2007756B1_D0598.tif" /></chemistry>

<i>N</i>-(1-(2-(2H-Tetrazol-5-yl)ethyl)-2-<i>tert</i>-butyl-1H-indol-5-yl)-1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide

0645To a solution of 1-(benzo[d][1,3]dioxol-5-yl)-<i>N-</i>(2-<i>tert</i>-butyl-1-(2-cyano-ethyl)indolin-5-yl)cyclopropanecarboxamide (50 mg, 0.12 mmol) in anhydrous DMF (2.6 mL) was added NH₄Cl (230 mg, 4.3 mmol) and NaN₃ (280 mg, 4.3 mmol). The reaction mixture was stirred for 30 min at 110 °C in the microwave, filtrated, and purified by preparative HPLC. The solid residue was dissolved in CDCl₃ (3 mL) and briefly (2 to 4 min) exposed to daylight, which initiated a color change (purple). After 2 h of stirring open to the atmosphere at room temperature, the solvent was removed and the residue was purified by preparative HPLC to.give <i>N</i>-(1-(2-(2H-tetrazol-5-yl)ethyl)-2-<i>tert</i>-butyl-1H-indol-5-yl)-1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide. MS (ESI) m/e (M+H⁺) 473.0.

Example 111: 1-(Benzo[d][1,3]dioxol-5-yl)-<i>N-</i>(2-<i>tert-</i>butyl-6-fluoro-1-((tetrahydro-2H-pyran-3-yl)methyl)-1H-indol-5-yl)cyclopropanecarboxamide

0646<chemistry id="chem0573" num="0573"><img file="EP2007756B1_D0599.tif" /></chemistry>

0647To a solution of 1-(benzo[d][1,3]dioxol-5-yl)-<i>N-</i>(2-<i>tert</i>-butyl-6-fluoroindolin-5-yl)cyclopropane-carboxamide (150 mg, 0.38 mmol) in anhydrous dichloromethane (2.3 mL) at room temperature under N₂ was added tetrahydropyran-3-carbaldehyde (54 mg, 0.47 mmol). After 20 min of stirring, NaBH(OAc)₃ (110 mg, 0.51 mmol) was added in one portion at room temperature. The reaction mixture was stirred for 6 h at room temperature before being purified by column chromatography on silica gel (5-20% ethyl acetate/hexanes) to provide 1-(benzo[d][1,3]dioxol-5-yl)-<i>N-</i>(2-<i>tert</i>-butyl-6-fluoro-1-((tetrahydro-2H-pyran-3-yl)methyl)indolin-5-yl)cyclopropanecarboxamide (95 mg, 50%). CDCl₃ was added to the indoline and the solution was allowed to stir overnight at ambient temperature. The solution was concentrated to give 1-(benzo[d][1,3]dioxol-5-yl)-<i>N-</i>(2-tert-butyl-6-fluoro-1-((tetrahydro-2H-pyran-3-yl)methyl)-1H-indol-5-yl)cyclopropanecarboxamide. MS (ESI) m/e (M+H⁺) 493.0.

Example 112: 1-(Benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N</i>-(2-(2-hydroxypropan-2-yl)-1<i>H-</i>indol-5-yl)cyclopropanecarboxamide

0648<chemistry id="chem0574" num="0574"><img file="EP2007756B1_D0600.tif" /></chemistry>

0649Methyl 5-(1-(benzo[<i>d</i>][1,3]dioxol-5-yl)cyclopropane-carboxamido)-1<i>H</i>-indole-2-carboxylate (100 mg, 0.255 mmol) was dissolved in anhydrous tetrahydrofuran (2 mL) under an argon atmosphere. The solution was cooled to 0 °C in an ice water bath before methyllithium (0.85 mL, 1.6 M in diethyl ether) was added by syringe. The mixture was allowed to warm to room temperature. The crude product was then partitioned between a saturated aqueous solution of sodium chloride (5 mL) and dichloromethane (5 mL). The organic layers were combined, dried over sodium sulfate, filtered, evaporated to dryness, and purified on 12 g of silica gel utilizing a gradient of 20-80% ethyl acetate in hexanes to yield 1-(benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N-</i>(2-(2-hydroxypropan-2-yl)-1<i>H</i>-indol-5-yl)cyclopropanecarboxamide (35 mg, 36%) as a white solid. -ESI-MS <i>m</i>/<i>z</i> calc. 378.2, found 379.1 (M+1)⁺. Retention time of 2.18 minutes. ¹H NMR (400 MHz, DMSO-<i>d</i>6) δ 10.78 (s, 1H), 8.39 (s, 1H), 7.57 (d, <i>J</i> = 1.7 Hz, 1H), 7.17 (d, <i>J =</i> 8.6 Hz, 1H), 7.03 - 6.90 (m, 4H), 6.12 (d, <i>J =</i> 1.5 Hz, 1H), 6.03 (s, 2H), 5.18 (s, 1H), 1.50 (s, 6H), 1.41 - 1.38 (m, 2H), 1.05-0.97 (m, 2H).

Example 113: <i>N</i>-(2-(1-Amino-2-methylpropan-2-yl)-1H-indol-5-yl)-1-(benzo[d] [1,3]-dioxol-5-yl)cyclopropanecarboxamide

0650<chemistry id="chem0575" num="0575"><img file="EP2007756B1_D0601.tif" /></chemistry>

0651Trifluoroacetic acid (0.75 mL) was added to a solution of <i>tert</i>-butyl 2-(5-(1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamido)-1H-indol-2-yl)-2-methylpropylcarbamate (77 mg, 0.16 mmol) in dichloromethane (3 mL) and the mixture was stirred at room temperature for 1.5 h. The mixture was evaporated, dissolved in dichloromethane, washed with saturated sodium bicarbonate solution, dried over magnesium sulfate and evaporated to dryness to give <i>N</i>-(2-(1-amino-2-methylpropan-2-yl)-1H-indol-5-yl)-1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide (53 mg, 86%). ¹H NMR (400 MHz, CDCl₃) δ 9.58 (s, 1H), 7.60 (d, J = 1.6 Hz, 1H), 7.18 - 7.15 (m, 2H), 7.02 - 6.94 (m, 3H), 6.85 (d, J = 7.8 Hz, 1H), 6.14 (d, J = 1.2 Hz, 1H), 6.02 (s, 2H), 2.84 (s, 2H), 1.68 (dd, J = 3.6, 6.7 Hz, 2H), 1.32 (s, 6H), 1.08 (dd, J = 3.7, 6.8 Hz, 2H).

Example 114: 1-(Benzo[d][1,3]dioxol-5-yl)-<i>N</i>-(2-(1-(dimethylamino)-2-methyl-propan-2-yl)-1H-indol-5-yl)cyclopropanecarboxamide

0652<chemistry id="chem0576" num="0576"><img file="EP2007756B1_D0602.tif" /></chemistry>

0653To a solution of <i>N</i>-(2-(1-amino-2-methylpropan-2-yl)-1H-indol-5-yl)-1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide (20 mg, 0.051 mmol) in DMF (1 mL) was added potassium carbonate (35 mg, 0.26 mmol) and iodomethane (7.0 µL, 0.11 mmol). The mixture was stirred for 2 h. Water was added and the mixture was extracted with dichloromethane. Combined organic phases were dried over magnesium sulfate, evaporated, coevaporated with toluene (3x) and purified by silica gel chromatography (0-30% EtOAc in hexane) to give 1-(benzo[d][1,3]dioxol-5-yl)-<i>N-</i>(2-(1-(dimethylamino)-2-methylpropan-2-yl)-1H-indol-5-yl)cyclopropanecarboxamide (7 mg, 33%). ¹H NMR (400 MHz, CDCl₃) δ 9.74 (s, 1H), 7.58 (d, J = 1.9 Hz, 1H), 7.20 (d, J = 8.6 Hz, 1H), 7.15 (s, 1H), 7.01 - 6.95 (m, 3H), 6.85 (d, J = 7.9 Hz, 1H), 6.10 (d, J = 0.9 Hz, 1H), 6.02 (s, 2H), 2.43 (s, 2H), 2.24 (s, 6H), 1.68 (dd, J = 3.7, 6.7 Hz, 2H), 1.33 (s, 6H), 1.08 (dd, J = 3.7, 6.8 Hz, 2H).

Example 115: <i>N</i>-(2-(1-Acetamido-2-methylpropan-2-yl)-1H-indol-5-yl)-1-(benzo[d][1,3]-dioxol-5-yl)cyclopropanecarboxamide

0654<chemistry id="chem0577" num="0577"><img file="EP2007756B1_D0603.tif" /></chemistry>

0655To a solution of <i>N</i>-(2-(1-amino-2-methylpropan-2-yl)-1H-indol-5-yl)-1-(benzo[d][1,3]dioxol-5-yl)cyclopropanecarboxamide (21 mg, 0.054 mmol) in dichloromethane (1 mL) was added pyridine (14 µL, 0.16 mmol) followed by acetic anhydride (6.0 µL, 0.059 mmol). The mixture was stirred for 2 h. Water was added and the mixture was extracted with dichloromethane, evaporated, coevaporated with toluene (3x) and purified by silica gel chromatography (60-100% ethylacetate in hexane) to give <i>N-</i>(2-(1-acetamido-2-methylpropan-2-yl)-1H-indol-5-yl)-1-(benzo[d][1,3]-dioxol-5-yl)cyclopropanecarboxamide (17 mg, 73%). ¹H NMR (400 MHz, DMSO) δ 10.79 (s, 1H), 8.39 (s, 1H), 7.66 (t, J = 6.2 Hz, 1H), 7.56 (d, J = 1.7 Hz, 1H), 7.18 - 7.14 (m, 1H), 7.02-6.89 (m, 4H), 6.08 (d, J = 1.5 Hz, 1H), 6.03 (s, 2H), 3.31 (d, J = 6.2 Hz, 2H), 1.80 (s, 3H), 1.41 - 1.38 (m, 2H), 1.26 (s, 6H), 1.04 - 1.01 (m, 2H).

Example 116: 1-(Benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N</i>-(2-(2-methyl-4-(1<i>H</i>-tetrazol-5-yl)butan-2-yl)-1<i>H</i>-indol-5-yl)cyclopropanecarboxamide

0656<chemistry id="chem0578" num="0578"><img file="EP2007756B1_D0604.tif" /></chemistry>

06571-(Benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N-</i>(2-(4-cyano-2-methylbutan-2-yl)-1<i>H-</i>indol-5-yl)cyclopropanecarboxamide (83 mg, 0.20 mmol) was dissolved in <i>N,N</i>-dimethylformamide (1 mL) containing ammonium chloride (128 mg, 2.41 mmol), sodium azide (156 mg, 2.40 mmol), and a magnetic stir bar. The reaction mixture was heated at 110 °C for 40 minutes in a microwave reactor. The crude product was filtered and then purified by preparative HPLC using a gradient of 0-99% acetonitrile in water containing 0.05% trifluoroacetic acid to yield 1-(benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N-</i>(2-(2-methyl-4-(1<i>H-</i>tetrazol-5-yl)butan-2-yl)-1<i>H-</i>indol-5-yl)cyclopropanecarboxamide. ESI-MS m/z calc. 458.2, found 459.2 (M+1)⁺. Retention time of 1.53 minutes. ¹H NMR (400 MHz, CD₃CN) 9.23 (s, 1H), 7.51 - 7.48 (m, 2H), 7.19 (d, <i>J =</i> 8.6 Hz, 1H), 7.06 - 7.03 (m, 2H), 6.95 - 6.89 (m, 2H), 6.17 (dd, <i>J =</i> 0.7, 2.2 Hz, 1H), 6.02 (s, 2H), 2.61 - 2.57 (m, 2H), 2.07 - 2.03 (m, 2H), 1.55-1.51 (m, 2H), 1.39 (s, 6H), 1.12-1.09 (m, 2H).

Example 117: 1-(Benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N-</i>(2-(piperidin-2-yl)-1<i>H-</i>indol-5-yl)cyclopropanecarboxamide

0658<chemistry id="chem0579" num="0579"><img file="EP2007756B1_D0605.tif" /></chemistry>

0659<i>tert</i>-Butyl 2-(5-(1-(benzo[<i>d</i>][1,3]dioxol-5-yl)cyclo-propanecarboxamido)-1<i>H-</i>indol-2-yl)piperidine-1-carboxylate (55 mg, 0.11 mmol) was dissolved in dichloromethane (2.5 mL) containing trifluoroacetic acid (1 mL). The reaction mixture was stirred for 6 h at room temperature. The crude product was purified by preparative HPLC using a gradient of 0-99% acetonitrile in water containing 0.05% trifluoroacetic acid to yield 1-(benzo[<i>d</i>][1,3]dioxol-5-yl)-<i>N</i>-(2-(piperidin-2-yl)-1<i>H</i>-indol-5-yl)cyclopropanecarboxamide. ESI-MS m/z calc. 403.2, found 404.4 (M+1)⁺. Retention time of 0.95 minutes.

Example 118: 5-<i>tert-</i>Butyl-1H-indol-6-ylamine

0660<chemistry id="chem0580" num="0580"><img file="EP2007756B1_D0606.tif" /></chemistry><chemistry id="chem0581" num="0581"><img file="EP2007756B1_D0607.tif" /></chemistry>

2-Bromo-4-<i>tert</i>-butyl-phenylamine

0661To a solution of 4-<i>tert</i>-Butyl-phenylamine (447 g, 3.00 mol) in DMF (500 mL) was added dropwise NBS (531 g, 3.00 mol) in DMF (500 mL) at room temperature. Upon completion, the reaction mixture was diluted with water and extracted with EtOAc. The organic layer was washed with water, brine, dried over Na₂SO₄ and concentrated. The crude product was directly used in the next step without further purification. <chemistry id="chem0582" num="0582"><img file="EP2007756B1_D0608.tif" /></chemistry>

2-Bromo-4-<i>tert-</i>butyl-5-nitro-phenylamine

06622-Bromo-4-<i>tert</i>-butyl-phenylamine (160 g, 0.71 mol) was added dropwise to H₂SO₄ (410 mL) at room temperature to yield a clear solution. This clear solution was then cooled down to -5 to -10 °C. A solution of KNO₃ (83 g, 0.82 mol) in H₂SO₄ (410 mL) was added dropwise while the temperature was maintained between -5 to -10 °C. Upon completion, the reaction mixture was poured into ice / water and extracted with EtOAc. The combined organic layers were washed with 5% Na₂CO₃ and brine, dried over Na₂SO₄ and concentrated. The residue was purified by a column chromatography (ethyl acetate/petroleum ether 1:10) to give 2-bromo-4-<i>tert</i>-butyl-5-nitro-phenylamine as a yellow solid (150 g, 78%). <chemistry id="chem0583" num="0583"><img file="EP2007756B1_D0609.tif" /></chemistry>

4-<i>tert</i>-Butyl-5-nitro-2-trimethylsilanylethynyl-phenylamine

0663To a mixture of 2-bromo-4-<i>tert</i>-butyl-5-nitro-phenylamine (27.3 g, 100 mmol) in toluene (200 mL) and water (100 mL) was added Et₃N (27.9 mL, 200 mmol), Pd(PPh₃)₂Cl₂ (2.11 g, 3.00 mmol), CuI (950 mg, 0.500 mmol) and trimethylsilyl acetylene (21.2 mL, 150 mmol) under a nitrogen atmosphere. The reaction mixture was heated at 70 °C in a sealed pressure flask for 2.5 h., cooled down to room temperature and filtered through a short plug of Celite. The filter cake was washed with EtOAc. The combined filtrate was washed with 5% NH₄OH solution and water, dried over Na₂SO₄ and concentrated. The crude product was purified by column chromatography (0 -10 % ethyl acetate/petroleum ether) to provide 4-<i>tert</i>-butyl-5-nitro-2-trimethylsilanylethynyl-phenylamine as a brown viscous liquid (25 g, 81 %). <chemistry id="chem0584" num="0584"><img file="EP2007756B1_D0610.tif" /></chemistry>

5-<i>tert-</i>Butyl-6-nitro-1H-indole

0664To a solution of 4-<i>tert</i>-butyl-5-nitro-2-trimethylsilanylethynyl-phenylamine (25 g, 86 mmol) in DMF (100 mL) was added CuI (8.2 g, 43 mmol) under a nitrogen atmosphere. The mixture was heated at 135 °C in a sealed pressure flask overnight, cooled down to room temperature and filtered through a short plug of Celite. The filter cake was washed with EtOAc. The combined filtrate was washed with water, dried over Na₂SO₄ and concentrated. The crude product was purified by column chromatography (10 - 20 % ethyl aetate/hexane) to provide 5-<i>tert</i>-butyl-6-nitro-1H-indole as a yellow solid (13 g, 69 %). <chemistry id="chem0585" num="0585"><img file="EP2007756B1_D0611.tif" /></chemistry>

5-<i>tert</i>-Butyl-1H-indol-6-ylamine

0665Raney Nickel (3 g) was added to 5-<i>tert</i>-butyl-6-nitro-1H-indole (15 g, 67 mmol) in methanol (100 mL). The mixture was stirred under hydrogen (1 atm) at 30 °C for 3 h. The catalyst was filtered off. The filtrate was dried over Na₂SO₄ and concentrated. The crude dark brown viscous oil was purified by column chromatography (10 - 20 % ethyl acetate/petroleum ether) to give 5-<i>tert</i>-butyl-1H-indol-6-ylamine as a gray solid (11 g, 87 %). ¹H NMR (300 MHz, DMSO-d6) δ 10.3 (br s, 1H), 7.2 (s, 1H), 6.9 (m, 1H), 6.6 (s, 1H), 6.1 (m, 1H), 4.4 (br s, 2H), 1.3 (s, 9H).

0666A person skilled in the chemical arts can use the examples and schemes along with known synthetic methodologies to synthesize compounds of the present invention, including the compounds in Table 3, below. <tables id="tabl0028" num="0028"><img file="EP2007756B1_D0612.tif" /></tables><tables id="tabl0029" num="0029"><img file="EP2007756B1_D0613.tif" /></tables><tables id="tabl0030" num="0030"><img file="EP2007756B1_D0614.tif" /></tables><tables id="tabl0031" num="0031"><img file="EP2007756B1_D0615.tif" /></tables><tables id="tabl0032" num="0032"><img file="EP2007756B1_D0616.tif" /></tables><tables id="tabl0033" num="0033"><img file="EP2007756B1_D0617.tif" /></tables><tables id="tabl0034" num="0034"><img file="EP2007756B1_D0618.tif" /></tables><tables id="tabl0035" num="0035"><img file="EP2007756B1_D0619.tif" /></tables>

VII. ASSAYS FOR DETECTING AND MEASURING ΔF508-CFTR CORRECTION PROPERTIES OF COMPOUNDS

Membrane potential optical methods for assaying ΔF508-CFTR modulation properties of compounds

0667The optical membrane potential assay utilized voltage-sensitive FRET sensors described by Gonzalez and Tsien (See, <nplcit id="ncit0032" npl-type="s"><text>Gonzalez, J. E. and R. Y. Tsien (1995) "Voltage sensing by fluorescence resonance energy transfer in single cells" Biophys J 69(4): 1272-80</text></nplcit>, and <nplcit id="ncit0033" npl-type="s"><text>Gonzalez, J. E. and R. Y. Tsien (1997) "Improved indicators of cell membrane potential that use fluorescence resonance energy transfer" Chem Biol 4(4): 269-77</text></nplcit>) in combination with instrumentation for measuring fluorescence changes such as the Voltage/Ion Probe Reader (VIPR) (See, <nplcit id="ncit0034" npl-type="s"><text>Gonzalez, J. E., K. Oades, et al. (1999) "Cell-based assays and instrumentation for screening ion-channel targets" Drug Discov Today 4(9): 431-439</text></nplcit>).

0668These voltage sensitive assays are based on the change in fluorescence resonant energy transfer (FRET) between the membrane-soluble, voltage-sensitive dye, DiSBAC₂(3), and a fluorescent phospholipid, CC2-DMPE, which is attached to the outer leaflet of the plasma membrane and acts as a FRET donor. Changes in membrane potential (V_m) cause the negatively charged DiSBAC₂(3) to redistribute across the plasma membrane and the amount of energy transfer from CC2-DMPE changes accordingly. The changes in fluorescence emission were monitored using VIPR™ II, which is an integrated liquid handler and fluorescent detector designed to conduct cell-based screens in 96- or 384-well microtiter plates.

Identification of Correction Compounds

0669To identify small molecules that correct the trafficking defect associated with ΔF508-CFTR; a single-addition HTS assay format was developed. The cells were incubated in serum-free medium for 16 hrs at 37 °C in the presence or absence (negative control) of test compound. As a positive control, cells plated in 384-well plates were incubated for 16 hrs at 27 °C to "temperature-correct" ΔF508-CFTR. The cells were subsequently rinsed 3X with Krebs Ringers solution and loaded with the voltage-sensitive dyes. To activate ΔF508-CFTR, 10 µM forskolin and the CFTR potentiator, genistein (20 µM), were added along with Cl^--free medium to each well. The addition of Cl^--free medium promoted Cl^- efflux in response to ΔF508-CFTR activation and the resulting membrane depolarization was optically monitored using the FRET-based voltage-sensor dyes.

Identification of Potentiator Compounds

0670To identify potentiators of ΔF508-CFTR, a double-addition HTS assay format was developed. During the first addition, a Cl^--free medium with or without test compound was added to each well. After 22 sec, a second addition of Cl^--free medium containing 2 - 10 µM forskolin was added to activate ΔF508-CFTR. The extracellular Cl^- concentration following both additions was 28 mM, which promoted Cl^- efflux in response to ΔF508-CFTR activation and the resulting membrane depolarization was optically monitored using the FRET-based voltage-sensor dyes.<u>Solutions</u>Bath Solution #1: (in mM) NaCl 160, KCl 4.5, CaCl₂ 2, MgCl₂ 1, HEPES 10, pH 7.4 with NaOH.

0671Chloride-free bath solution: Chloride salts in Bath Solution #1 are substituted with gluconate salts.

0672CC2-DMPE: Prepared as a 10 mM stock solution in DMSO and stored at-20°C.

0673DiSBAC₂(3): Prepared as a 10 mM stock in DMSO and stored at -20°C.

Cell Culture

0674NIH3T3 mouse fibroblasts stably expressing ΔF508-CFTR are used for optical measurements of membrane potential. The cells are maintained at 37 °C in 5% CO₂ and 90 % humidity in Dulbecco's modified Eagle's medium supplemented with 2 mM glutamine, 10 % fetal bovine serum, 1 X NEAA, β-ME, 1 X pen/strep, and 25 mM HEPES in 175 cm² culture flasks. For all optical assays, the cells were seeded at 30,000/well in 384-well matrigel-coated plates and cultured for 2 hrs at 37 °C before culturing at 27 °C for 24 hrs. for the potentiator assay. For the correction assays, the cells are cultured at 27 °C or 37 °C with and without compounds for 16 - 24 hours <u>Electrophysiological Assays for assaying ΔF508-CFTR modulation properties of compounds Ussing Chamber Assay</u> Ussing chamber experiments were performed on polarized epithelial cells expressing ΔF508-CFTR to further characterize the ΔF508-CFTR modulators identified in the optical assays. FRT^ΔF508-CFTR epithelial cells grown on Costar Snapwell cell culture inserts were mounted in an Ussing chamber (Physiologic Instruments, Inc., San Diego, CA), and the monolayers were continuously short-circuited using a Voltage-clamp System (Department of Bioengineering, University of Iowa, IA, and, Physiologic Instruments, Inc., San Diego, CA). Transepithelial resistance was measured by applying a 2-mV pulse. Under these conditions, the FRT epithelia demonstrated resistances of 4 KΩ/ cm² or more. The solutions were maintained at 27 °C and bubbled with air. The electrode offset potential and fluid resistance were corrected using a cell-free insert. Under these conditions, the current reflects the flow of Cl^- through ΔF508-CFTR expressed in the apical membrane. The I_SC was digitally acquired using an MP100A-CE interface and AcqKnowledge software (v3.2.6; BIOPAC Systems, Santa Barbara, CA).

Identification of Correction Compounds

0675Typical protocol utilized a basolateral to apical membrane Cl^- concentration gradient. To set up this gradient, normal ringer was used on the basolateral membrane, whereas apical NaCl was replaced by equimolar sodium gluconate (titrated to pH 7.4 with NaOH) to give a large Cl^- concentration gradient across the epithelium. All experiments were performed with intact monolayers. To fully activate ΔF508-CFTR, forskolin (10 µM) and the PDE inhibitor, IBMX (100 µM), were applied followed by the addition of the CFTR potentiator, genistein (50 µM).

0676As observed in other cell types, incubation at low temperatures of FRT cells stably expressing ΔF508-CFTR increases the functional density of CFTR in the plasma membrane. To determine the activity of correction compounds, the cells were incubated with 10 µM of the test compound for 24 hours at 37°C and were subsequently washed 3X prior to recording. The cAMP- and genistein-mediated I_SC in compound-treated cells was normalized to the 27°C and 37°C controls and expressed as percentage activity. Preincubation of the cells with the correction compound significantly increased the cAMP- and genistein-mediated I_SC compared to the 37°C controls.

Identification of Potentiator Compounds

0677Typical protocol utilized a basolateral to apical membrane Cl^- concentration gradient. To set up this gradient, normal ringers was used on the basolateral membrane and was permeabilized with nystatin (360 µg/ml), whereas apical NaCl was replaced by equimolar sodium gluconate (titrated to pH 7.4 with NaOH) to give a large Cl^- concentration gradient across the epithelium. All experiments were performed 30 min after nystatin permeabilization. Forskolin (10 µM) and all test compounds were added to both sides of the cell culture inserts. The efficacy ot the putative ΔF508-CFTR potentiators was compared to that of the known potentiator, genistein.

Solutions

0678Basolateral solution (in mM): NaCl (135), CaCl₂(1.2), MgCl₂ (1.2), K₂HPO₄ (2.4), KHPO₄ (0.6), N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid (HEPES) (10), and dextrose (10). The solution was titrated to pH 7.4 with NaOH.

0679Apical solution (in mM): Same as basolateral solution with NaCl replaced with Na Gluconate (135).

Cell Culture

0680Fisher rat epithelial (FRT) cells expressing ΔF508-CFTR(FRT^ΔF508-CFTR) were used for Ussing chamber experiments for the putative ΔF508-CFTR modulators identified from our optical assays. The cells were cultured on Costar Snapwell cell culture inserts and cultured for five days at 37 °C and 5% CO₂ in Coon's modified Ham's F-12 medium supplemented with 5% fetal calf serum, 100 U/ml penicillin, and 100 µg/ml streptomycin. Prior to use for characterizing the potentiator activity of compounds, the cells were incubated at 27 °C for 16 - 48 hrs to correct for the ΔF508-CFTR. To determine the activity of corrections compounds, the cells were incubated at 27 °C or 37 °C with and without the compounds for 24 hours.

0681The macroscopic ΔF508-CFTR current (I_ΔF508) in temperature- and test compound-corrected NIH3T3 cells stably expressing ΔF508-CFTR were monitored using the perforated-patch, whole-cell recording. Briefly, voltage-clamp recordings of I_ΔF508 were performed at room temperature using an Axopatch 200B patch-clamp amplifier (Axon Instruments Inc., Foster City, CA). All recordings were acquired at a sampling frequency of 10 kHz and low-pass filtered at 1 kHz. Pipettes had a resistance of 5 - 6 MΩ when filled with the intracellular solution. Under these recording conditions, the calculated reversal potential for Cl^- (E_Cl) at room temperature was -28 mV. All recordings had a seal resistance > 20 GΩ and a series resistance < 15 MΩ. Pulse generation, data acquisition, and analysis were performed using a PC equipped with a Digidata 1320 A/D interface in conjunction with Clampex 8 (Axon Instruments Inc.). The bath contained < 250 µl of saline and was continuously perifused at a rate of 2 ml/min using a gravity-driven perfusion system.

Identification of Correction Compounds

0682To determine the activity of correction compounds for increasing the density of functional ΔF508-CFTR in the plasma membrane, we used the above-described perforated-patch-recording techniques to measure the current density following 24-hr treatment with the correction compounds. To fully activate ΔF508-CFTR, 10 µM forskolin and 20 µM genistein were added to the cells. Under our recording conditions, the current density following 24-hr incubation at 27°C was higher than that observed following 24-hr incubation at 37 °C. These results are consistent with the known effects of low-temperature incubation on the density of ΔF508-CFTR in the plasma membrane. To determine the effects of correction compounds on CFTR current density, the cells were incubated with 10 µM of the test compound for 24 hours at 37°C and the current density was compared to the 27°C and 37°C controls (% activity). Prior to recording, the cells were washed 3X with extracellular recording medium to remove any remaining test compound. Preincubation with 10 µM of correction compounds significantly increased the cAMP- and genistein-dependent current compared to the 37°C controls.

Identification of Potentiator Compounds

0683The ability of ΔF508-CFTR potentiators to increase the macroscopic ΔF508-CFTR Cl^- current (I_ΔF508) in NIH3T3 cells stably expressing ΔF508-CFTR was also investigated using perforated-patch-recording techniques. The potentiators identified from the optical assays evoked a dose-dependent increase in I_ΔF508 with similar potency and efficacy observed in the optical assays. In all cells examined, the reversal potential before and during potentiator application was around -30 mV, which is the calculated E_Cl (-28 mV).

Solutions

0684Intracellular solution (in mM): Cs-aspartate (90), CsCl (50), MgCl₂ (1), HEPES (10), and 240 µg/ml amphotericin-B (pH adjusted to 7.35 with CsOH).

0685Extracellular solution (in mM): <i>N</i>-methyl-D-glucamine (NMDG)-Cl (150), MgCl₂ (2), CaCl₂ (2), HEPES (10) (pH adjusted to 7.35 with HCl).

Cell Culture

0686NIH3T3 mouse fibroblasts stably expressing ΔF508-CFTR are used for whole-cell recordings. The cells are maintained at 37 °C in 5% CO₂ and 90 % humidity in Dulbecco's modified Eagle's medium supplemented with 2 mM glutamine, 10 % fetal bovine serum, 1 X NEAA, β-ME, 1 X pen/strep, and 25 mM HEPES in 175 cm² culture flasks. For whole-cell recordings, 2,500 - 5,000 cells were seeded on poly-L-lysine-coated glass coverslips and cultured for 24 - 48 hrs at 27 °C before use to test the activity of potentiators; and incubated with or without the correction compound at 37 °C for measuring the activity of correctors.

0687The single-channel activities of temperature-corrected ΔF508-CFTR stably expressed in NIH3T3 cells and activities of potentiator compounds were observed using excised inside-out membrane patch. Briefly, voltage-clamp recordings of single-channel activity were performed at room temperature with an Axopatch 200B patch-clamp amplifier (Axon Instruments Inc.). All recordings were acquired at a sampling frequency of 10 kHz and low-pass filtered at 400 Hz. Patch pipettes were fabricated from Coming Kovar Sealing #7052 glass (World Precision Instruments, Inc., Sarasota, FL) and had a resistance of 5 - 8 MΩ when filled with the extracellular solution. The ΔF508-CFTR was activated after excision, by adding 1 mM Mg-ATP, and 75 nM of the cAMP-dependent protein kinase, catalytic subunit (PKA; Promega Corp. Madison, WI). After channel activity stabilized, the patch was perifused using a gravity-driven microperfusion system. The inflow was placed adjacent to the patch, resulting in complete solution exchange within 1 - 2 sec. To maintain ΔF508-CFTR activity during the rapid perifusion, the nonspecific phosphatase inhibitor F^- (10 mM NaF) was added to the bath solution. Under these recording conditions, channel activity remained constant throughout the duration of the patch recording (up to 60 min). Currents produced by positive charge moving from the intra- to extracellular solutions (anions moving in the opposite direction) are shown as positive currents. The pipette potential (V_p) was maintained at 80 mV.

0688Channel activity was analyzed from membrane patches containing ≤ 2 active channels. The maximum number of simultaneous openings determined the number of active channels during the course of an experiment. To determine the single-channel current amplitude, the data recorded from 120 sec of ΔF508-CFTR activity was filtered "off-line" at 100 Hz and then used to construct all-point amplitude histograms that were fitted with multigaussian functions using Bio-Patch Analysis software (Bio-Logic Comp. France). The total microscopic current and open probability (P_o) were determined from 120 sec of channel activity. The P_o was determined using the Bio-Patch software or from the relationship P_o = I/i(N), where I = mean current, i = single-channel current amplitude, and N = number of active channels in patch.

Solutions

0689Extracellular solution (in mM): NMDG (150), aspartic acid (150), CaCl₂ (5), MgCl₂ (2), and HEPES (10) (pH adjusted to 7.35 with Tris base).

0690Intracellular solution (in mM): NMDG-Cl (150), MgCl₂ (2), EGTA (5), TES (10), and Tris base (14) (pH adjusted to 7.35 with HCl).

Cell Culture

0691NIH3T3 mouse fibroblasts stably expressing ΔF508-CFTR are used for excised-membrane patch-clamp recordings. The cells are maintained at 37 °C in 5% CO₂ and 90 % humidity in Dulbecco's modified Eagle's medium supplemented with 2 mM glutamine, 10 % fetal bovine serum, 1 X NEAA, β-ME, 1 X pen/strep, and 25 mM HEPES in 175 cm² culture flasks. For single channel recordings, 2,500 - 5,000 cells were seeded on poly-L-lysine-coated glass coverslips and cultured for 24 - 48 hrs at 27 °C before use.

0692Compounds of the invention are useful as modulators of ATP binding cassette transporters. Using the procedures described above, the activities, i.e., EC50s, of compounds of the present invention have been measured to be from about 3.8 nM to about 13.5 µM. Furthermore, using those methods described above, the efficacies of compounds of the present invention have been measured to be from about 35 % to about 110 %.