6-o-substituted benzoxazole and benzothiazole compounds and methods of inhibiting csf-1r signaling

DESCRIPTION “BENZOTIAZOLE AND BENZOXAZOLE 6-O-REPLACED COMPOUNDS AND SIGNALING INHIBIT METHODS CSF-1R”

Field of the Invention

The present invention relates to 6-0 substituted benzoxazole and benzothiazole compounds as defined herein, their tautomers, stereoisomers, solvates, oxides and their pharmaceutically acceptable salts. The invention also relates to compositions of the compounds combined with pharmaceutically acceptable carriers. In another aspect, this invention relates to the uses of the compounds, alone or in combination with at least one additional therapeutic agent, for the prophylaxis or treatment of cancer.

State of the Art

CSF-1R is the receptor for M-CSF (macrophage colony stimulating factor, also called CSF1) and mediates the biological effects of this cytokine (Sherr 1985). Colony-stimulating factor 1 receptor cloning (also called c-fms) was first described in Roussel et al., Nature 325: 549-552 (1987). In this publication, it was shown that CSF-1R had transformation potential dependent on

ΡΕ2010528 C-terminus of protein, including loss of inhibition of Cbl-binding tyrosine 969 phosphorylation and thus down-regulating receptor (Lee 1999).

CSF-1R is a single stranded transmembrane tyrosine kinase (RTK) receptor and a member of the immunoglobulin (Ig) motif family containing RTKs characterized by repeated Ig domains in the extracellular portion of the receptor. 0 The intracellular protein tyrosine kinase domain is disrupted by a single inserted domain that is also present in the other related class III RTK family members that include platelet-derived growth factor receptors (PDGFR), the growth factor receptor stem cells (c-Kit) and the fms-like cytokine receptor (FLT3). Despite the structural homology between this family of growth factor receptors, they have distinct tissue specific functions. CSF-1R is mainly expressed in monocytic lineage cells and in the female reproductive tract and placenta. In addition, CSF-1R expression has been reported in skin Langerhans cells, a subset of smooth muscle cells (Inaba, 1992), B cells (Baker, 1993) and microglia (Sawada, 1990).

The main biological effects of CSF-1R signaling are the differentiation, proliferation, migration and survival of macrophage and osteoclasts precursors of the monocyte lineage. Activation of CSF-1R is mediated by its single ligand, M-CSF. Binding of M-CSF to CSF-1R induces

102010528 homodimer formation and kinase activation by tyrosine phosphorylation (Stanley, 1997). Additional signaling is mediated by the p85 subunit of PI3K and Grb2 which binds to the PI3K / AKT and Ras / MAPK pathways, respectively. These two important signaling pathways can regulate proliferation, survival and apoptosis. Other signaling molecules that bind to the intracellular domain

<td>CSF-1R phosphorylated (Bourette 2000).</td><td>include STATl,</td><td>STAT3,</td><td>PLCy and Cbl</td>

<td>The signage</td><td>of CSF-1R has</td><td>A paper</td><td>physiological</td>

<td>in immune responses,</td><td>in remodeling</td><td>bone and</td><td>in the system</td>

reproductive. M-CSF-1 knockout animals (op / op mouse; Pollard 1996) or CSF-1R (Dai 2002) knockouts have been shown to have osteoporotic, hematopoietic, tissue macrophage, and reproductive phenotypes consistent with a role for CSF-1R in their cell types.

The recent success of targeted therapies such as Herceptin® and Avastin® has emphasized the importance of developing cleaner less promiscuous drugs with a more specific mechanism of action. These drugs can minimize side effects, have greater predictability, give doctors more flexibility in their treatments, and give researchers a better understanding of a specific target. In addition, targeted therapy may allow the treatment of multiple indications affected by the same signaling pathway with less toxicities and potentially easier to manage.

102010528 (BioCentury, V. 14 (10) Feb, 2006) Inhibition of an individual kinase such as CSF-1R, which is integrated into a pathway associated with cancer or other diseases, can also effectively modulate downstream kinases, thereby affecting the entire pathway. However, the active sites of 491 human protein kinase domains are highly conserved, which makes designing selective inhibitors a formidable challenge (Cohen, 2005). For example (WO2005 / 073224) discloses compounds which are effective for the prophylaxis and treatment of HGF-mediated diseases, for example diseases and other conditions or conditions involving cancer and the like.

SUMMARY OF THE INVENTION

There is a continuing need for compounds that inhibit cell proliferation, that inhibit tumor growth, that treat cancer, that modulate cell cycle arrest, and / or that specifically inhibit molecules such as CSF-1R and for pharmaceutical and drug formulations. containing such compounds. There is also a need for selective CSF-1R inhibitor compounds. There is also a need for compounds for use in methods of administering such compounds, pharmaceutical formulations and medicaments to patients or individuals in need thereof.

One embodiment is directed to compounds selected from the group defined in claim 1, stereoisomers, tautomers, solvates, oxides or salts thereof.

102010528 pharmaceutically acceptable.

Another embodiment is directed to the compound defined in claim 2, or to a pharmaceutically acceptable solvate, oxide or salts thereof.

Another embodiment is directed to the compound defined in claim 3, or to a pharmaceutically acceptable solvate, oxide or salts thereof.

Another embodiment is directed to the compound defined in claim 4, or a pharmaceutically acceptable solvate, oxide or salts thereof.

Also disclosed is a CSR-1R inhibitor compound selected from the group defined in claim 1, or a stereoisomer, tautomer, solvate or oxide, or a pharmaceutically acceptable salt thereof for use in treating a CSF-1R mediated disorder as defined herein.

In a more particular embodiment said compound does not substantially inhibit Raf kinase. In a more particular embodiment said compound preferably inhibits CSF-1R over Raf kinase. In a more particular embodiment, said compound inhibits Raf kinase at an ICso greater than about 1 μΜ. In a more particular embodiment said compound inhibits CSF-1R at an ICso of less than about 1 μΜ. More particularly, said compound inhibits CSF-1R

102010528 at an IC50 of less than about 0.1 µM.

DETAILED DESCRIPTION

Throughout this application, the text refers to various embodiments of the present compounds, compositions and uses.

Definitions

Unless specifically defined otherwise, terms used herein are defined below.

Oxide refers to products resulting from the oxidation of one or more heteroatoms. Examples include oxides, sulfoxides and sulfones. Solvate or solvates refer to compounds or a salt thereof that are bound in a stoichiometric or non-stoichiometric amount of a solvent. Preferred solvents are volatile, non-toxic and / or acceptable for administration to humans in trace amounts. Suitable solvates include water.

Stereoisomers or stereoisomers refer to compounds that differ in chirality from one or more stereocenters. Stereoisomers include enantiomers and diastereomers.

Tautomer refers to alternative forms of a

2010528 compounds which differ in the position of a proton, such as enol keto and imine enamine tautomers, or tautomeric forms of heteroaryl groups containing a ring atom attached to a -NH- moiety and a ring having a moiety. = Ntal as pyrazoles, imidazoles, benzimidazoles, triazoles and tetrazoles.

Pharmaceutically acceptable salt refers to pharmaceutically acceptable salts of a compound, the salts of which are derived from a variety of organic and inorganic counterions well known in the art and include, by way of example only, sodium, potassium, calcium, magnesium, ammonium and tetraalkylammonium; and when the molecule contains basic functionality, salts of organic or inorganic acids, such as hydrochloride, hydrobromide, tartrate, mesylate, acetate, maleate and oxalate. 0 The term also includes pharmaceutically acceptable salts of stereoisomers, tautomers, of the compound.

mammals

Patient refers to human mammals and nonhuman mammals.

and includes

Treating patient refers to patient who is developing disease symptoms; or disease.

or treating a disease in a to 1) prevent the disease from occurring in a predisposed or not yet present condition; 2) inhibit the disease or stop your

3) improve or cause regression of

102010528

Selective inhibition refers to a compound, composition or chemotype that preferentially inhibits a particular target or class of targets. Reference to selective CSF-1R inhibition indicates preferential inhibition of CSF1R and optionally as kinase receptors such as PDGFR. In some embodiments, selective inhibition of CSF-1R refers to preferential inhibition of CSF-1R over Raf kinase. Selective, targeted, specific or preferential inhibition is not intended to mean complete absence of inhibitory activity with respect to all other kinases or receptors.

CSF-1R inhibitor refers to a compound that can inhibit CSF-1R. Preferably, a CSF1R inhibitor is CSF-1R selective over other targets. In one embodiment, a CSF-1R inhibitor has selective inhibition of CSF-1R relative to Raf kinase. In a preferred embodiment, such selective inhibition relates to at least one 2: 1 binding preference of a compound of this invention to CSF-1R over Raf kinase, more preferably at least 5: 1, and even more preferably. at least 10: 1.

One embodiment is directed to compounds selected from the group defined in claim 1, stereoisomers, tautomers, solvates, or oxides, or pharmaceutically acceptable salts thereof.

Another embodiment is directed to the compound defined in claim 2, or to a solvate, oxide or derivatives thereof.

102010528 pharmaceutically acceptable salts.

Another embodiment is directed to the compound defined in claim 3, or to a solvate, oxide, or pharmaceutically acceptable salts thereof.

Another embodiment is directed to the compound defined in claim 4, or a solvate, oxide, or pharmaceutically acceptable salts thereof.

In other embodiments, a compound of Table 2 or 3, or a stereoisomer, tautomer, solvate, oxide, or a pharmaceutically acceptable salt thereof is provided.

It will also be apparent to those skilled in the art that the compounds of the invention according to any one of claims 1 to 4 or their stereoisomers as well as the pharmaceutically acceptable salts may be subject to tautomerization and may therefore exist in various tautomeric forms in which a proton of one atom of a molecule shifts to another atom and the chemical bonds between the atoms of the molecules are consequently rearranged. See, for example, March, Advanced Organic Chemistry: Reactions, Mechanisms and Structures, Fourth Edition, John Wiley &amp; Sons, pages 69-74 (1992).

Preferred embodiments, including the compounds according to any one of claims 1 to 4, or their tautomers, as well as the pharmaceutically salts thereof.

102010528, may comprise asymmetrically substituted carbon atoms. Such asymmetrically substituted carbon atoms may result in compounds of preferred embodiments existing in enantiomers, diastereomers and other stereoisomeric forms which may be defined in terms of absolute stereochemistry, such as (R) - or (S) - forms. As a result, all possible isomers, individual stereoisomers in their optically pure forms, mixtures thereof, racemic mixtures (or racemates), mixtures of diastereomers, as well as individual diastereomers of the compounds of the preferred embodiments are contemplated. Terms with an S and R configuration as used herein are as defined by IUPAC 1974 RECOMMENDATIONS FOR SECTION E, STEREOCHEMISTRY FOUNDATION,

Pure Appl. Chem. 45: 13-30 (1976). The terms α and β are used for ring positions of cyclic compounds. The α side of the reference plane is the side where the preferred substituent is at the lowest numbered position. Those substituents on the opposite side of the reference plane are assigned to the descriptor β. It should be noted that this use differs from that for cyclic stereoparents, where it means below the plane and denotes absolute configuration. The terms of configuration α and β as used herein are as defined by CHEMICAL ABSTRACTS INDEZ GUIDE - APPENDIX IV (1987), paragraph 203.

There are 3 distinct mechanisms by which CSF-1R signaling is probably involved in

102010528 tumor growth and metastasis. The first is that CSF ligand and receptor expression was found in tumor cells originating in the female reproductive system (breast, ovary, endometrium, cervical) (Scholl 1994; Kacinski 1997; Nagan 199; Kirma 2007) breast cancer xenograft growth, as well as a poor prognosis in breast cancer patients. Two point mutations in CSF-1R were observed in about 1020% of patients with acute myelocytic leukemia, chronic myelocytic leukemia and myelodysplasia tested in one study, and one of the mutations was found to disrupt receptor production (Ridge 1990). However, the incidence of mutations could not be confirmed in subsequent studies (Abu-Duhier, 2003). Mutations have also been found in some cases of hepatocellular cancer (Yang 2004) and idiopathic myelofibrosis (Abu-Duhier, 2003).

Pigmented villonodular synovitis (PVNS) and giant cell tenosynovial tumors (TGCT) may occur as a result of translocation that fuses the M-CSF gene into a collagen COL6A3 gene and results in overexpression of M-CSF (West 2006). A landscaping effect is proposed to be responsible for the resulting tumor mass consisting of monocytic cells attracted by cells expressing M-CSF. TGCTs are smaller tumors that can easily be removed from the fingers where they usually occur. PVNS is more aggressive because it can recur in large joints and is not as easily surgically controlled.

102010528 second mechanism is based on M-CSF / CSF-1R blocking signaling at metastatic sites in bone that induces osteoclastogenesis, bone resorption and osteolytic bone lesions. Breast, kidney and lung cancers are examples of cancers that have been found to metastasize to bone and cause osteolytic bone disease, resulting in bone complications. 0 M-CSF released by tumor cells and stroma induces differentiation of hematopoietic myeloid monocyte progenitors in mature osteoclasts in collaboration with the kapa-B nuclear factor activating receptor ligand - RANKL. During this process, M-CSF acts as a permissive factor in giving the survival signal to osteoclasts (Tanaka, 1993). Inhibition of CSF-1R kinase activity during osteoclast differentiation and maturation with a small molecular inhibitor is likely to prevent the unbalanced activity of osteoclasts that cause osteolytic disease and bone-related events in metastatic disease. Since breast cancer, lung cancer and multiple myeloma usually result in osteolytic lesions, bone metastasis in prostate cancer initially has an osteoblastic appearance in which increased bone-forming activity results in fibrous bone that is different from lamellar structure typical of normal bone. During disease progression, bone lesions exhibit a significant osteolytic component as well as high serum bone resorption levels and suggest that anti-resorption therapy may be helpful. Bisphosphonates have been shown to inhibit the formation of

ΡΕ2010528 Osteolytic lesions and reducing the number of skeletal-related events only in men with hormone-refractory metastatic prostate cancer, but at present their effect on osteoblastic lesions is controversial and bisphosphonates have not been beneficial to date in preventing bone metastasis. or hormone-responsive prostate cancer. 0 The effect of anti-resorptive agents on mixed osteolytic / osteoblastic prostate cancer is still being clinically studied (Choueiri 2006; Vessella

2006).

The third mechanism is based on the recent observation that tumor-associated macrophages (TAM) found in solid breast, prostate, ovarian and cervical cancer tumors correlate with a poor prognosis (Bingle 2002, Pollard, 2004). Macrophages are recruited into the tumor by M-CSF and other chemokines. Macrophages may then contribute to tumor progression by secreting angiogenic factors, proteases and other growth factors and cytokines and may be blocked by inhibiting CSF-1R signaling. Recently, it has been shown by Zins et al (Zins 2007) that expression of tumor necrosis factor alpha (TNFα) siRNA, M-CSF or a combination of both would reduce tumor growth in a mouse xenograft model by 34% to 50%. % after intratumoral injection of the respective xenograft siRNA. The siRNA directed to TNFalpha secreted by human SW620 cells reduced mouse M-CSF and led to reduction of tumor macrophages. In addition, the treatment of xenografts

MCF7 tumor Δ2010528 with an antigen-binding fragment directed against the M-CSF antibody resulted in 40% inhibition of tumor growth, reversed chemotherapeutic resistance and improved survival of mice when administered in combination with chemotherapeutic agents (Paulus 2006).

TAMs are just one example of an emerging link between chronic inflammation and cancer. There is additional evidence of a link between inflammation and cancer, as many chronic diseases are associated with an increased risk of cancer, cancers arise at sites of chronic inflammation, many chemical mediators of inflammation are found in many cancers; Deletion of cellular or chemical mediators of inflammation inhibits the development of experimental cancers and prolonged use of anti-inflammatory agents reduces the risk of some cancers. There is a cancer link to a number of inflammatory conditions including H. pylori-induced gastritis for gastric cancer, schistosomiasis for bladder cancer, HHV8 for Kaposi's sarcoma, ovarian cancer endometriosis and prostate cancer prostatitis (Balkwill 2005 ). Macrophages are key cells in chronic inflammation and respond differently to their microenvironment. There are two types of macrophages that are considered extreme in a continuum of functional states: M1 macrophages are involved in Type 1 reactions. These reactions involve activation by microbial products and consequent death of pathogenic microorganisms that result.

102010528 in reactive oxygen intermediates. At the other extreme, M2 macrophages are involved in Type 2 reactions that promote cell proliferation, adjust inflammation and adaptive immunity, and promote tissue remodeling, angiogenesis and maintenance (Mantovani, 2004). Chronic inflammation that results in established neoplasia is usually associated with M2 macrophages. A key cytokine mediating inflammatory reactions is TNF-α that lives up to its name and can stimulate anti-tumor immunity and hemorrhagic necrosis at high doses, but it has also recently been found to be expressed by tumor cells and acts as a promoter. of tumor (Zins 2007; Balkwill 2006). The specific role of macrophages in relation to the tumor still needs to be better understood, including the potential spatial and temporal dependence of their function and relevance to specific tumor types.

Also disclosed is a compound as defined in any one of claims 1 to 4 for use in the treatment of periodontitis, histiocytosis X, osteoporosis, Paget bone disease (PDB), cancer therapy bone loss, periprosthetic osteolysis, glucocorticoid-induced osteoporosis, rheumatoid arthritis, psoriatic arthritis, osteoarthritis, inflammatory arthritis and inflammation.

Rabello 2006 demonstrated that CSF1 SNPs were positively associated with aggressive periodontitis: an inflammatory disease of periodontal tissues that causes tooth loss due to bone resorption.

102010528 alveolar.

Histiocytosis X (also called Langerhans cell histiocytosis, LCH) is a proliferative disease of Langerhans dendritic cells that appear to differentiate into osteoclasts in LCH bone and extraosseous lesions.

Langerhans cells are derived from circulating monocytes (Ginoux 2006). The increase in M-CSF levels that was measured in sera and lesions correlated with disease severity (da Costa 2005). The disease occurs mainly in a pediatric patient population and has to be treated with chemotherapy when the disease becomes systemic or recurs.

The pathophysiology of osteoporosis is mediated by loss of bone-forming osteoblasts and increased osteoclast-dependent bone resorption. Supporting data have been described by Cenci et al showing that an injection of anti-M-CSF antibodies preserves bone density and inhibits bone resorption in ovariectomized mice (Cenci 2000). A potential link between postmenopausal bone loss due to estrogen deficiency has recently been identified and the presence of TNF alpha producing T cells was observed to affect bone metabolism (Roggia 2004). A possible mechanism could be the induction of M-CSF by TNF alpha in vivo. An important role for M-CSF in TNF-alpha induced osteoclastogenesis has been confirmed by the effect of an antibody directed against the M-CSF inhibitor that blocked osteolysis induced by

102010528

TNF alpha in mice and thereby making CSF-1R signaling inhibitors potential targets for inflammatory arthritis (Kitaura 2005).

Paget's bone disease (PDB) is the 2nd most common bone metabolism disorder after osteoporosis, in which focal abnormalities of increased bone production lead to complications such as bone pain, deformation, pathological fractures and deafness. Mutations have been identified in four genes that regulate normal osteoclast function and predispose individuals to PDB and related disorders: insertion mutations in TNFRSF11A, which codes for nuclear factor (NF) receptor activator kappaB (RANK) is a critical regulator of osteoclast function, inactivating mutations of TNFRSF11B, which codes for osteoprotegerin (a decoy receptor for RANK ligand) ), sequestosome 1 gene mutations (SQSTM1), which codes for an important structural protein in the NFkappaB pathway, and mutations in the valosine protein (VCP) -containing gene. This gene codes for PCV, which plays a role in NFkappaB inhibitor targeting for proteasome degradation (Daroszewska, 2006). Specific CSF-1R inhibitors provide an opportunity to block RANKL signaling disruption indirectly and add an additional treatment option to the currently used bisphosphonates.

Cancer therapy-induced bone loss, especially in breast and prostate cancer patients, is

ΡΕ2010528 an additional indication that a targeted CSF1R inhibitor may prevent bone loss (Lester, 2006). With the better prognosis for early breast cancer, the long-term consequences of adjunctive therapies become more important as some of the therapies, including chemotherapy, radiation, aromatase inhibitors and ovarian ablation, affect bone metabolism, decreasing bone mineral density, resulting in a higher risk of osteoporosis and associated fractures (Lester, 2006). The equivalent of adjuvant aromatase inhibitor therapy in breast cancer is androgen ablation therapy in prostate cancer, which leads to loss of bone mineral density and significantly increases the risk of osteoporosis-related fractures (Stoch, 2001). .

Targeted inhibition of CSF-1R signaling is likely to be beneficial in other indications also when target cell types include osteoclasts and macrophages, for example treating specific complications in response to joint replacement as a consequence of rheumatoid arthritis. Implant failure due to periprosthetic bone loss and consequent loss of prostheses is an important complication of joint replacement and requires repeated surgery with high socioeconomic burden for the individual patient and the healthcare system. To date, there is no approved pharmacological therapy to prevent or inhibit periprosthetic osteolysis (Drees, 2007).

Glucocorticoid-induced osteoporosis (GIOP)

102010528 is another indication that a CSF-1R inhibitor can prevent bone loss following prolonged glucocorticoid use that is given as a result of several states between chronic obstructive pulmonary disease, asthma and rheumatoid arthritis (Guzman-Clark, 2007 Feldstein 2005 ).

Rheumatoid arthritis, psoriatic arthritis, and inflammatory arthritis are, by themselves, potential indications for CSF-1R signaling inhibitors, as they are a macrophage component of varying bone degradation (Ritchlin 2003). Osteoarthritis and rheumatoid arthritis are autoimmune inflammatory diseases caused by macrophage accumulation in connective tissue and macrophage infiltration into synovial fluid, which is at least partially mediated by M-CSF. Campbell et al. (2000) demonstrated that M-CSF is produced by human joint tissue cells (chondrocytes, synovial fibroblasts) in vitro and is found in the synovial fluid of rheumatoid arthritis patients, suggesting that it contributes to the proliferation of synovial tissue and infiltration of macrophages that is associated with the pathogenesis of the disease. Inhibition of CSF-1R signaling is likely to control the number of macrophages in the joint and alleviate the pain of associated bone destruction. In order to minimize adverse effects and to better understand the impact of CSF-1R signaling in these states, one method is to specifically inhibit CSF-1R without reaching a myriad of other kinases, such as Raf kinase.

Recent literature reports correlate the

102010528 increased circulating M-CSF with a poor prognosis and atherosclerotic progression in chronic coronary artery disease (Saitoh 2000; Ikonomidis 2005); M-CSF influences the atherosclerotic process by helping the formation of foam cells (macrophages with ingested oxidized LDL) that express CSF-1R and represent the initial plaque (Murayama, 1999).

M-CSF and CSF-1R expression and signaling are found in activated microglyocytes. Microglyocytes, which are resident macrophages of the central nervous system, can be activated by various insults, including infections and traumatic injuries. M-CSF is considered a key regulator of inflammatory responses in the brain and M-CSF levels increase HIV-1 encephalitis, Alzheimer's disease (AD) and brain tumors. Microgliosis as a result of autocrine signaling by M-CSF / CSF-1R results in the induction of inflammatory cytokines and released nitric oxides, as demonstrated by, for example, using an experimental model of neuronal damage (Hao 2002; Murphy, 1998). It was found that microglyocytes that increased CSF-1R expression surround plaques in AD and in the AD V717F amyloid precursor protein transgenic mouse model (Murphy 2000). In contrast, op / op mice with fewer brain microglyocytes resulted in fibrillar deposition of Αβ and neuronal loss compared to normal control suggesting that microglyocytes have a neuroprotective function in AD development absent in op / op mice (Kaku,

2003).

102010528

In one aspect, the disclosure provides a compound according to any one of claims 1 to 4 for use in treating CSF1R-related disorders in an amount effective to reduce or prevent the disorder. In a preferred embodiment, the disorder is tumor growth and / or metathesis in the subject.

In other aspects, the disclosure provides a compound according to any one of claims 1 to 4 for use in treating CSF-1R related disorders in an amount effective to reduce or prevent osteoclastogenesis, bone resorption and / or bone damage in the subject. .

In yet other aspects, the disclosure provides a compound according to any one of claims 1 to 4 for use in treating CSF-1R related disorders in an amount effective to treat the disorder in the individual, in combination with at least one additional agent. . In a more particular embodiment, the additional agent is a bisphosphonate. In one embodiment, the disorder is tumor growth and / or metastasis, osteoclastogenesis, bone resorption, and / or bone lesions.

In yet other aspects, the disclosure provides compounds according to any one of claims 1 to 4 which are capable of selectively or preferably inhibiting CSF-1R. In embodiments

Preferred CSF-1R inhibitors are capable of inhibiting CSF-1R greater than about 5-fold, or about 10-fold, or about 20-fold, or about 30-fold, or about 50-fold, or about. 100 times, or about 250 times, or about 500 times, or about 750 times, or about

1,000 fold, or about 2,000 fold, inhibitory activity (relative to ICso values, for example) in Raf kinase.

In other aspects, the disclosure provides a compound according to any one of claims 1 to 4 for use in inhibiting CSF-1R comprising contacting a cell with said compound.

In one aspect, the inhibitory effect of the compounds on Raf is determined using the following biotinylated assay. Raf kinase activity is measured by providing ATP, an inactive MEK substrate for recombinant kinase and analyzing the transfer of the phosphate moiety to the MEK residue. Full length recombinant MEK with an inactivating K97R mutation at the ATP binding site (rendering the kinase inactive) is expressed in E. coli and labeled with biotin after purification. MEK cDNA is subcloned with an N-end (His) label 6 and expressed in E. coli and the recombinant MEK substrate is purified from E. coli lysate by nickel affinity chromatography followed by anion exchange. The final MEK substrate preparation is biotinylated (Pierce EZ-Link Sulfo-NHS-LC-Biotin) and concentrated to about 11.25 μΜ. Recombinant Raf (including c-Raf and mutant B-Raf isoforms) is obtained by

102010528 purification from sf9 insect cells infected with the corresponding recombinant human Rat expression vectors. Recombinant Rat isoforms are purified by Glu antibody interaction or by metal ion chromatography.

For each assay, the compound is serially diluted, for example, from 25 μΜ with 3-fold dilutions in DMSO and then mixed with various Rat isoforms (about 0.50 nM each). Kinase-inactive MEK-biotin substrate (50 nM) is added in the reaction buffer plus ATP (1 μΜ). The reaction buffer contains 30 mM Tris-HCl₂ pH 7.5, 10 mM MgCl₂MT DTT, 4 mM EDTA, 25 mM betaglycerophosphate, 5 mM MnCl₂, and 0.01% BSA / PBS. The reactions are subsequently incubated for about 2 hours at room temperature and quenched by the addition of 0.5 M EDTA. The quenched reaction mixture is transferred to a neutravidine coated plate and incubated for about 1 hour. The phosphorylated product is measured with the time resolved fluorescence system DELFIA using a rabbit antip-MEK (Cell Signaling) as the primary antibody and europium-labeled anti-rabbit as the secondary antibody. Time resolved fluorescence can be read on a Wallac 1232 DELFIA fluorometer. Compound concentration for 50% inhibition (IC 50) is calculated by nonlinear regression using XL Fit data analysis software.

In yet other aspects, preferred embodiments provide a compound, tautomer, solvate,

Oxide or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 4, in an amount effective to reduce or prevent tumor growth in the individual in combination with at least one additional cancer treatment agent. In a more particular embodiment, the additional agent is a bisphosphonate.

A number of suitable anticancer agents to be used as combination therapy are contemplated for use as defined herein. In fact, preferred embodiments include administration of various additional anticancer agents such as but not limited to apoptosis-inducing agents; polynucleotides (e.g. ribozymes); polypeptides (e.g. enzymes); drugs; biological mimetics; alkaloids; alkylating agents; antitumor antibiotics; antimetabolites; hormones; platinum compounds; monoclonal antibodies conjugated with anticancer drugs, toxins and / or radionuclides; biological response modifiers (e.g., interferons [e.g. IFN-Î ±, etc.] and interleukins [e.g. IL-2, etc.], etc.); adoptive immunotherapy agents; hematopoietic growth factors; agents that induce tumor cell differentiation (e.g., all-trans-retinoic acid, etc.); gene therapy reagents; antisense therapy reagents and nucleotides; tumor vaccines; angiogenesis inhibitors and the like.

Numerous other examples of compounds

Chemotherapeutic and anticancer therapies suitable for co-administration with the disclosed compounds according to any one of claims 1 to 4 are known to those skilled in the art.

In preferred embodiments, additional anticancer agents to be used in combination with compounds as defined in any one of claims 1 to 4 comprise agents that induce or stimulate apoptosis. Apoptosis-inducing agents include, but are not limited to, radiation (e.g., ω); kinase inhibitors (e.g., epidermal growth factor receptor kinase inhibitor [EGFR], vascular endothelial growth factor receptor kinase inhibitor [VEGFR], fibroblast growth factor receptor kinase inhibitor [FGFR] , platelet-derived growth factor receptor kinase I [PDGFR] and Bcr-Abl kinase inhibitors such as STI-571, Gleevec and Glivec]); antisense molecules; antibodies [e.g. Herceptin and Rituximab]; antiestrogens [e.g. raloxifene and tamoxifen]; antiandrogens [e.g. flutamide, bicalutamide, finasteride, aminoglutetemide, ketoconazole and corticosteroids]; cyclooxygenase 2 (COX-2) inhibitors [e.g., Celecoxib, meloxicam, NS-398 and non-steroidal antiinflammatory drugs (NSAIDs)]; and cancer chemotherapeutic drugs (e.g., irinotecan (Camptosar), CPT-11, fludarabine (Fludara), dacarbazine (DTIC), dexamethasone, mitoxantrone, Milotarg, VP-16,

102010528 cisplatin, 5-FU, Doxrubicin, Taxotere or taxol; cell signaling molecules; ceramides and cytokines; and staurosporine, and the like.

Compounds of preferred embodiments are useful in vitro or in vivo in inhibiting cancer cell growth. The compounds may be used alone or in compositions in conjunction with a pharmaceutically acceptable carrier or excipient.

In other aspects, the disclosure provides a pharmaceutical composition comprising at least one compound according to any one of claims 1 to 4, together with a pharmaceutically acceptable carrier, suitable for administration to a human or animal individual, alone or in conjunction with other agents. anticancer drugs.

In other aspects, the disclosure provides methods of manufacturing compounds of the invention as described above.

In other aspects, there is provided a pharmaceutical composition comprising an effective amount of a compound, stereoisomer, tautomer, solvate or oxide, or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 4, and a pharmaceutically acceptable carrier. In some aspects, the compound preferably inhibits CSF-1R over Raf kinase. More particularly, said compound inhibits Raf kinase from

102010528 more than about 1 μΜ.

Other aspects further comprise an additional agent. More particularly, said additional agent is a bisphosphonate.

Other aspects provide compounds as defined in any one of claims 1 to 4 effective to inhibit CSF-1R activity in a human or animal subject when administered to it. More particularly, said compound has an ICso value of CSF-1R inhibition of less than about 1 μ 1. More particularly, said compound has an ICso value for Raf inhibition of greater than about 1 μΜ.

The disclosure provides for the use of a compound as defined herein to inhibit CSF-1R, wherein said compound selectively inhibits CSF-1R.

The compounds of the embodiments are useful in vitro or in vivo in inhibiting cancer cell growth. The compounds may be used alone or in compositions in conjunction with a pharmaceutically acceptable carrier or excipient. Suitable pharmaceutically acceptable carriers or excipients include, for example, drug delivery agents and modifiers and enhancers such as, for example, calcium phosphate, magnesium stearate, talc, monosaccharides, disaccharides, starch, gelatin, cellulose, methylcellulose ,

102010528 sodium carboxymethylcellulose, dextrose, hydroxypropyl-pciclodextrin, polyvinylpyrrolidinone, low melting waxes, ion exchange resins and the like, as well as combinations of any two or more thereof. Other suitable pharmaceutically acceptable excipients are described in Remington's Pharmaceutical Sciences, Mack Pub. Co., New Jersey (1991).

Pharmaceutical Composition and Administration

In general, the compounds of the disclosure will be administered in a therapeutically effective amount by any of the accepted modes of administration for agents with similar utilities. The effective amount of the compound of preferred embodiments, i.e. the active ingredient, will depend on numerous factors such as the severity of the disease being treated, the age and relative health of the individual, the potency of the compound used, the route and the form of administration, and other factors. 0 The drug may be administered more than once a day, preferably once or twice a day. All these factors are within the competence of the attending clinician.

Effective amounts of the compounds of the disclosure generally include any amount sufficient to detectably inhibit CSF-1R activity by any of the assays described herein, by other CSF-1R kinase activity assays known to those skilled in the art or to the art. detection of an inhibition or

102010528 of a relief of cancer symptoms.

The amount of active ingredient that may be combined with the carrier materials to produce a single dosage form will vary depending upon the host treated and the particular mode of administration. It will be understood, however, that the specific dose level for any specific patient will depend on a variety of factors, including the activity of the specific compound employed, age, body weight, general health, gender, diet, time of administration, route of administration. , excretion rate, drug combination and disease severity, in particular subjected to therapy. The therapeutically effective amount for a given condition can be readily determined by routine experimentation and is within the competence and judgment of the ordinary practitioner.

For purposes of disclosure, a therapeutically effective dose may generally be a total daily dose administered to a host in single or divided doses, may be in amounts, for example, from about 0.001 to about 1000 mg / kg body weight daily and More preferably from about 1.0 to about 30 mg / kg body weight daily. Dosage unit compositions may contain such amounts of their submultiples to form the daily dose.

The choice of formulation depends on several factors, such as the mode of drug administration and the

102010528 bioavailability of the drug substance. In general, the compounds of the disclosure may be administered as pharmaceutical compositions by any of the following routes: oral, systemic (e.g. transdermal, intranasal or suppository) administration, or parenteral (e.g. intramuscular, intravenous or subcutaneous) administration. The preferred mode of administration is oral, using a convenient daily dosage regimen that can be adjusted according to the degree of distress. The compositions may take the form of tablets, pills, capsules, semisolids, powders, sustained release formulations, solutions, suspensions, elixirs, aerosols or any other suitable compositions. Another preferred way to administer the compounds of the preferred embodiments is inhalation. This is an effective method for administering a therapeutic agent directly to the respiratory tract (see US Patent 5,607,915).

Suitable pharmaceutically acceptable carriers or excipients include, for example, processing agents and drug release modifying and enhancing agents, such as, for example, calcium phosphate, magnesium stearate, talc, monosaccharides, disaccharides, starch, gelatin, cellulose, methylcellulose, sodium carboxymethylcellulose, dextrose, hydroxypropyl-pciclodextrin, polyvinylpyrrolidinone, low melting waxes, ion exchange resins and the like, as well as combinations of any two or more of them. The excipients

Liquid and semi-solid may be selected from glycerol, propylene glycol, water, ethanol and various oils, including those of petroleum, animal, vegetable or synthetic origin, for example peanut oil, soybean oil, mineral oil, sesame oil, etc. . Preferred liquid carriers, particularly for injectable solutions, include water, saline, aqueous dextrose and glycols. Other suitable pharmaceutically acceptable excipients are described in Remington's Pharmaceutical Sciences, Mack Pub. Co., New Jersey (1991).

As used herein, the term pharmaceutically acceptable salts refers to the non-toxic alkaline earth metal or acid salts of the compounds as defined in any one of claims 1 to 4. These salts may be prepared in situ during isolation and final purification. of the compounds as defined in any one of claims 1 to 4, or by separate reaction of the base or acid functions with a suitable organic or inorganic acid or base, respectively. Representative salts include, but are not limited to the following: acetate, adipate, alginate, citrate, aspartate, benzoate, benzenesulfonate, bisulfate, butyrate, camphor, camphorsulfonate, digluconate, cyclopentanopropionate, dodecyl sulfate, ethanesulfonate, glucoheptanoate, glycerophosphate heptanoate, hexanoate, fumarate, hydrochloride, hydrobromide, hydroiodide, 2-hydroxyethanesulfonate, lactate, maleate, methanesulfonate, nicotinate, 2-naphthalenesulfonate, oxalate, pamoate,

102010528 pectinate, persulfate, 3-phenylproionate, picrate, pivalate, propionate, succinate, sulfate, tartrate, thiocyanate , ptoluenesulfonate and undecanoate. In addition, basic nitrogen-containing groups may be quaternized with agents such as alkyl halides such as methyl, ethyl, propyl and butyl chloride, bromides and iodides; di-alkyl sulfates such as dimethyl, diethyl, dibutyl and diamyl sulfates, long chain halides such as decyl, lauryl, myristyl and stearyl chlorides, bromides and iodides, aralkyl halides such as benzyl and phenethyl bromides and others. Thus, aqueous or water dispersible or oil dispersible products are obtained.

Examples of acids that may be used to form pharmaceutically acceptable acid addition salts include inorganic acids such as hydrochloric acid, sulfuric acid and phosphoric acid and organic acids such as oxalic acid, maleic acid, methanesulfonic acid, succinic acid and citric acid. The basic addition salts may be prepared in situ during the final isolation and purification of the compounds as defined in any one of claims 1 to 4, or separately by reacting portions of carboxylic acid with a suitable base such as hydroxide, carbonate or bicarbonate. a pharmaceutically acceptable or ammoniacal metal cation or a primary, secondary or tertiary organic amine. Pharmaceutically acceptable salts include, but are not limited to, alkali metal and alkaline earth metal cations such as sodium, lithium, potassium,

102010528 calcium, magnesium, aluminum and the like, as well as non-toxic ammonium, quaternary ammonia, and amine cations, including, but not limited to, ammonia, tetramethylammonium, tetraethylammonium, methylamine, dimethylamine, trimethylamine, triethylamine, ethylamine. Other representative organic amines useful for the formation of basic addition salts include diethylamine, ethylenediamine, ethanolamine, diethanolamine, piperazine and the like.

The compounds of the disclosure may be administered orally, parenterally, sublingually, by aerosolization, or by inhalation by spray, rectally or topically in dosage unit formulations containing conventional non-toxic pharmaceutically acceptable carriers, adjuvants and vehicles as desired. . Topical administration may also involve the use of transdermal administration, such as transdermal patches or iontophoresis devices. 0 The term parenteral as used herein includes subcutaneous, intravenous, intrathecal, intramuscular injections, intrasternal injection or infusion techniques.

Injectable 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 or suspension in a non-toxic parenterally acceptable diluent or solvent,

ΡΕ2010528 for example as a solution in 1,3-propanediol. Among the acceptable vehicles and solvents that may be employed are water, Ringer's solution 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 may be employed including synthetic mono- or diglycerides. In addition, fatty acids such as oleic acid find use in the preparation of injectables.

Suppositories for rectal administration of the drug may be prepared by mixing the drug with a suitable non-irritating excipient such as cocoa butter and polyethylene glycols, which are solid at normal temperatures but liquid at rectal temperature and therefore melt in the rectum and release. the drug.

Solid dosage forms for oral administration may include capsules, tablets, pills, powders and granules. In such solid dosage forms, the active compound may be mixed 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, for example lubricating agents, such as magnesium stearate. In the case of capsules, tablets and pills, dosage forms may also comprise buffering agents. Tablets and tablets may also be prepared with enteric coatings.

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Liquid dosage forms for oral administration may include pharmaceutically acceptable emulsions, solutions, suspensions, syrups and elixirs containing inert diluents commonly used in the art, such as water. Such compositions may also comprise adjuvants such as wetting agents, emulsifying and suspending agents, cyclodextrins and sweetening, flavoring and perfuming agents.

The compounds of the disclosure may also be administered as liposomes. As is known in the art, liposomes are generally derived from phospholipids or other lipid substances. Liposomes are formed by mono- or multilamellar hydrated liquid crystals which are dispersed in an aqueous medium. Any non-toxic, physiologically acceptable and metabolizable lipid capable of forming liposomes may be used. The present liposome form compositions may contain, in addition to a compound of preferred embodiments, stabilizers, preservatives, excipients and the like. Preferred lipids are natural and synthetic phospholipids and phosphatidylcholines (lecithins). Methods for forming liposomes are known in the art. See, for example, Prescott, Ed., Methods in Cell Biology, Volume XIV, Academic Press, New York, NW, p. 33 et seq. (1976).

Compressed gases may be used to disperse a compound of the disclosure in aerosol form. The

Suitable inert gases for this purpose are nitrogen, carbon dioxide, etc. Other suitable pharmaceutical excipients and their formulations are described in Remington's Pharmaceutical Sciences, edited by EW Martin (Mack Publishing Company, 18^The ed., 1990)).

For administration by inhalation, the compound may be formulated as a liquid solution, suspensions, aerosol propellants or dry powder and loaded into a dispenser suitable for administration. There are several types of pharmaceutical inhalation devices - nebulizer inhalers, metered dose inhalers (MDI) and dry powder inhalers (DPI). Nebulizer devices produce a high-velocity airflow that causes therapeutic agents (which are formulated in liquid form) to spray like a mist that is carried into the patient's respiratory tract. MDIs are typically formulated with a compressed gas. Upon actuation, the device discharges a metered amount of therapeutic agent by compressed gas, thereby providing a reliable method of administering a fixed amount of agent. DPI dispenses therapeutic agents in the form of a free-flowing powder that can be dispersed in the patient's inspiratory stream while breathing through the device. To achieve a free flowing powder, the therapeutic agent is formulated with an excipient such as lactose. A measured amount of the therapeutic agent is stored in a capsule form and is dispensed with each actuation.

102010528

Recently, pharmaceutical formulations have been developed especially for drugs that have low bioavailability based on the principle that bioavailability can be increased by increasing surface area, i.e. decreasing particle size. For example, U.S. Pat. No. 4,107,288 describes a pharmaceutical formulation with particles in the size range from about 10 to about 1000 nm, wherein the active material is supported on a cross-linked macromolecule matrix. US Patent No.

5,145,684 describes the production of a pharmaceutical formulation in which the drug substance is sprayed onto nanoparticles (average particle size of about 400 nm) in the presence of a surface modifier and then dispersed in a liquid medium to give a pharmaceutical formulation having a pharmaceutical composition. remarkably high bioavailability.

Combination Therapies

It is disclosed that the compounds of preferred embodiments may be administered as the sole active pharmaceutical agent and may also be used in combination with one or more other agents used in the treatment of cancer. Compounds of preferred embodiments are also useful in combination with known therapeutic agents and anticancer agents, and combinations of the presently described compounds with other anticancer or chemotherapeutic agents are disclosed. Examples of such agents can be found in Cancer.

102010528

Principles and Practice of Oncology, VT Devita and S. Hellman (editors), 6^The edition (February 15, 2001), Lippincott Williams &amp; Wilkins Publishers. One of ordinary skill in the art would be able to discern which combinations of agents would be useful based on the particular characteristics of the drugs and the cancer involved. Such anticancer agents include, but are not limited to the following: estrogen receptor modulators, androgen receptor modulators, retinoid receptor modulators, cytotoxic / cytostatic agents, antiproliferative agents, prenyl protein transferase inhibitors, HMG-CoA reductase inhibitors and other angiogenesis inhibitors, proliferation inhibitors survival and signaling, apoptosis inducing agents and agents that interfere with cell cycle control points. Compounds of preferred embodiments are also useful when co-administered with radiation therapy.

Accordingly, in one embodiment, the compounds are also used in combination with known anticancer agents including, for example, estrogen receptor modulators, androgen receptor modulators, retinoid receptor modulators, cytotoxic agents, antiproliferative agents, prenyl protein transferase, HMG-CoA reductase inhibitors, HIV protease inhibitors, reverse transcriptase inhibitors and other angiogenesis inhibitors.

102010528

Estrogen receptor modulators are compounds that can interfere with or inhibit estrogen binding to the receptor, regardless of the mechanism. Examples of estrogen receptor modulators include, but are not limited to, tamoxifen, raloxifene, oxyphene, LY353381, LY117081, toremifene, fulvestrant, 4- [7- (2,2-dimethyl-1-oxopropoxy-4-methyl-2- Methyl [4- [2- (1-piperidinyl) ethoxy] phenyl] -2H-1-benzopyran-3-yl] phenyl-2,2-dimethylpropanoate, 4,4'-dihydroxybenzophenone2,4-dinitrophenylhydrazone and SH646.

Androgen receptor modulators are compounds that can interfere with or inhibit androgen binding to an androgen receptor. Representative examples of androgen receptor modulators include finasteride and other 5α-reductase inhibitors, nilutamide, flutamide, bicalutamide, liarozole and abiraterone acetate. Retinoid receptor modulators are compounds that interfere with or inhibit retinoid binding to a retinoid receptor. Examples of retinoid receptor modulators include bexarotene, tretinoin, 13-cis retinoic acid, 9-cis retinoic acid, adifluoromethylornithine, LX23-7553, trans-N- (4'hydroxyphenyl) retinamide and N4-carboxyphenyl retinamide.

Cytotoxic and / or cytostatic agents are compounds that can cause cell death or inhibit cell proliferation primarily by directly interfering with cell function or by inhibiting or

102010528 interference with cellular mitosis, including alkylating agents, tumor necrosis factors, intercalators, hypoxia activatable compounds, microtubule inhibitors / microtubule stabilizing agents, mitotic kinesin inhibitors, kinase inhibitors involved in mitotic progression, anti-metabolites; biological response modifiers; hormonal / antihormonal therapeutic agents, hematopoietic growth factors, monoclonal antibody targeted therapeutic agents, topoisomerase inhibitors, proteasome inhibitors and ubiquitin ligase inhibitors. Examples of cytotoxic agents include, but are not limited to, sertenef, catechin, ifosfamide, tasonermin, lonidamine, carboplatin, altretamine, prednimustine, dibromodulcitol, ranimustine, fotemustine, nedaplatin, oxaliplatin, temozolomide, heptaplatin, impramphosphosphosphate, troptaplatin nimustine, dibrospidium chloride, pumitepa, lobaplatin, satraplatin, profiromycin, cisplatin, irofulvene, dexiphosphamide, cisaminadichloro (2-methylpyridine) platinum, benzylguanine, glufosfamide, GPX100, (trans, trans, trans) bis-mu- (hexane-1,6-diamide) mu- [diamineplatin (II) bis [diamine (chlorine) platinum (II)], diarizidinilspermine, trioxide tetrachloride arsenic, 1- (11dodecylamino-10-hydroxidencil) -3,7-dimethylxanthine, zorubicin, idarubicin, daunorubicin, bisanthrene, mitoxantrone, pirarubicin, pinafide, valrubicin, amrubicin, antineoplaston, 3'-diaoxino-3'-diamino-amino Hydroxycarminomycin, anamycin, galarubicin,

Δ2010528 elinafide, MEN10755, and 4-methoxy-3-deamino-3-aziridinyl4-methylsulfonyl-daunorubicin (see WO 00/50032). A representative example of a hypoxia activatable compound is tirapazamine. Proteasome inhibitors include, but are not limited to, lactacystin and bortezomib. Examples of microtubule inhibitors / microtubule stabilizing agents include paclitaxel, vindesine sulfate, 3 ', 4'-didehydro-4'-deoxy-8'-norvincaleucoblastine, docetaxol, rhizoxin, dolastatin, mivobulin isothionate, auristatin, cemadotine, R10988, R10988 , BMS184476, vinflunine, cryptophycin, 2,3,4,5,6-pentafluoro-N- (3-fluoro4methoxyphenyl) benzene sulfonamide, anhydrovinblastine, N, Ndimethyl-L-valyl-L-valyl-N-methyl-L-valyl -L-prolyl-L-prolinat-butylamide, TDX258, epothilones (see for example US Pat. Nos. 6,284,781 and 6,288,237) and BMS188797. Representative examples of topoisomerase inhibitors include topotecan, hapaptama, irinotecan, rubitecan, 6-ethoxypropionyl-3 ', 4'-O-exo-benzylidene carbonose, 9-methoxyN, N-dimethyl-5-nitropyrazolo [3,4,5-kl ] acridine-2 (6H) propanamine, 1-amino-9-ethyl-5-fluoro-2,3-dihydro-9hydroxy-4-methyl-1H, 12H-benzo [de] pyran [3'4 ': b, 7] indolizino [1,2b] quinoline-10,13 (9H, 15H) dione, lurtotecan, 7- [2- (Nisopropylamino) ethyl] - (20S) camptothecin, BNP1350, BNPI1100,

BN80915, BN80942, etoposide phosphate, teniposide, sobuzoxane, 2'-dimethylamino-2'-deoxy etoposide, GL331, N [2- (dimethylamino) ethyl] -9-hydroxy-5,6-dimethyl-6H-pyrido [ 4,3-b] carbazole-1-carboxamide, asulacrine, (5a, 5aB, 8aa, 9b) 9- [2- [N- [2- (dimethylamino) ethyl] -N-methylamino] ethyl] 5- [4hydroxy -3,5-dimethoxyphenyl] -5,5a, 6,8,8a, 9-hexahydrofuro (3 ',

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4 ': 6.7) naphtho (2,3-d) -1,3-dioxol-6-one, 2,3- (methylene dioxy) -5-methyl-7-hydroxy-8-methoxybenzo [c] -phenanthridinium, 6,9-bis [(2-aminoethyl) amino] benzo [g] isogininoline-5,10-dione, 5- (3aminopropylamino) -7,10-dihydroxy-2- (2hydroxyethylaminomethyl) -6H- pyrazolo [4,5,1'-de] acridin-6-one, N- [1- [2- (diethylamino) ethylamino] -7-methoxy-9-oxo-9Hthioxanten-4-ylmethyl] formamide, N- ( 2- (dimethylamino) ethyl) acridine-4-carboxamide, 6 - [[2- (dimethylamino) ethyl] amino] -3-hydroxy-7H-indene [2,1c] quinolin-7-one and dimesna. Examples of mitotic kinesin inhibitors such as human mitotic kinesin KSP are described in PCT Publications WO 01/30768 and WO 01/98278, WO 03 / 050,064 (Jun 19, 2003), WO 03 / 050,122 (Jun 19 ., 2003), WO 03 / 049,527 (Jun 19, 2003), WO 03 / 049,679 (Jun 19, 2003), WO 03/049, 678 (Jun 19, 2003) and WO

03/39460 (May 15, 2003) and PCT applications pending Nos. US03 / 06403 (submitted Mar. 4, 2003), US03 / 15861 (submitted May 19, 2003), US03 / 15810 (submitted May 19, 2003), US03 / 18482 (submitted 12 Jun., 2003) and US03 / 18694 (submitted Jun. 12, 2003). Mitotic kinesin inhibitors include, but are not limited to, KSP inhibitors, MKLP1 inhibitors, CENPE inhibitors, MCAK inhibitors, Kifl4 inhibitors, Mphosphl inhibitors, and Rab6-KIFL inhibitors.

Kinase inhibitors involved in mitotic progression include, but are not limited to, aurora kinase inhibitors, Polo-like kinase inhibitors (PLK) (e.g., PLK-1 inhibitors), bubΡΕ2010528 inhibitors, and bub-lR inhibitors. . Antiproliferative agents include antisense DNA and DNA oligonucleotides such as G3139, ODN698, RVASKRAS, GEM231 and INX3001, and antimetabolites such as enocytabine, carmofur, tegafur, pentostatin, doxifluridine, trimetrexate, capuditabine, fludarabine, fludarabine, cytarabine ocphosphate, sodium phostoabine hydrate, raltitrexed, paltitrexide, emitefur, thiazofurin, decitabine, nolatrexed, pemetrexed, nelzarabine, 2'-deoxy-2'-methylidenecitidine, 2'fluoromethylene-2'-deoxycytidine, N- [5- (2,3-dihydrobenzofuryl) sulfonyl] -N '- (3,4-dichlorophenyl) urea, N6 - [4-deoxy-4- [N2- [2 (E), 4 (E) -tetradecadienoyl] glycylamino] -glycerol-BL-manoheptopyranosyl] adenine, aplidine, ecteinascidine, troxacytabine, 4- [2-amino-4oxo4,6,7,8-tetrahydro- 3H-pyrimidin [5,4-b] [1,4] thiazin-6yl- (S) -ethyl] -2,5-thienyl-L-glutamic, aminopterin, 5-fluorouracil, alanosine, 11-Acetyl-8- (carbamoyloxymethyl) -4-formyl-6-methoxy-14-oxa-1,1-diazatetracyclo (7.4.1.0.0) -tetradeca-2,4,6-trien-9-ylacetic acid ester, swainsonine , lometrexol, dexrazoxane, methioninase, 2'-cyano-2'-deoxy-N4-palmitoyl-1BD-arabino furanosyl cytosine and 3-aminopyridine-2-carboxaldehyde thiosemicarbazone. Examples of monoclonal antibody directed therapeutic agents include those therapeutic agents having cytotoxic agents or radioisotopes attached to a specific monoclonal antibody of cancer cells or target cells. Examples include, for example, Bexxar. HMG-CoA reductase inhibitors are 3-hydroxy-3-methylglutaryl-CoA reductase inhibitors. The

102010528 compounds having inhibitory activity for HMG-CoA reductase can be readily identified using assays well known in the art, such as those described or cited in U.S. Pat. No. 4,231,938 and WO 84/02131. Examples of HMG-CoA reductase inhibitors that may be used include, but are not limited to, lovastatin (MEVACOR®;

see Pat. US Nos. 4,231,938, 4,294,926 and 4,319,039), simvastatin (ZOCOR®; see US Pat. Nos. 4,444,784, 4,820,850 and 4,916,239), pravastatin (PRAVACHOL®; see US Pat. Nos. 4,346,227, 4,537,859, 4,410, 629, 5, 030,447 and 5, 180,589), fluvastatin (LESCOL®; see US Pat. Nos. 5,354,772, 4, 911,165, 4, 929, 437, 5, 189, 164, 5, 118,853, 5,290, 946 and

5,356,896) and atorvastatin (LIPITOR®; see US Pat. Nos. 5,273,995, 4,681,893, 5,489,691 and 5,342,952). It is disclosed that the structural formulas of these and additional HMGCoA reductase inhibitors that can be used in the instantaneous methods are described on page 87 of M. Yalpani, Cholesterol Lowering Drugs, Chemistry &amp; Industry, pp. 8589 (Feb. 5, 1996) and Pat. US Nos. 4,782,084 and 4,885,314. In one embodiment, the HMG-CoA reductase inhibitor is selected from lovastatin or simvastatin.

Prenyl protein transferase inhibitors are compounds that inhibit any or any combination of prenyl protein transferase enzymes, including farnesyl protein transferase (FPTase), geranylgeranyl protein transferase type I (GGPTase-I) and geranylgeranyl protein transferase type II ( GGPTase-II, also called Rab GGPTase). Examples of prenylΡΕ2010528 protein transferase inhibitor compounds include (±) -6- [amino (4-chlorophenyl) (1-methyl-1H-imidazol-5-yl) methyl] -4- (3-chlorophenyl) -1-methyl2 (1H ) -quinolinone, (-) - 6- [amino (4-chlorophenyl) (1-methyl-1H-imidazol-5-yl) methyl] -4- (3-chlorophenyl) -1-methyl-2 (1H) quinolinone, (+) - 6- [amino (4-chlorophenyl) (1-methyl-1H-imidazol-5-yl) methyl] -4- (3-chlorophenyl) -1-methyl-2 (1H) quinolinone, 5 (S) -n-butyl-1- (2,3-dimethylphenyl) -4- [1- (4-cyanobenzyl) -5-imidazolylmethyl-2-piperazinone, (S) -1- (3-chlorophenyl) -4- [1- (4-cyanobenzyl) -5-imidazolylmethyl] -5- [2- (ethanesulfonyl) methyl) -2-piperazinone, 5 (S) -n-butyl- 1- (2-methylphenyl) -4- [1- (4-cyanobenzyl) -5-imidazolylmethyl] -2piperazinone, 1- (3-chlorophenyl) -4- [1- (4-cyanobenzyl) -2-methyl-5-imidazolylmethyl ] -2-piperazinone, 1- (2,2-diphenylethyl) -3- [N- (1- (4-cyanobenzyl) -1H-imidazol-5ethylyl) carbamoyl] piperidine, 4- {- [4-hydroxymethyl-4- (4-chloropyridin-2-ylmethyl) -piperidin-1-ylmethyl] -2-methylimidazol-1-ylmethyl} benzonitrile, 4 - {- 5- [4-hydroxymethyl-4- (3-chlorobenzyl) piperidin-1-ylmethyl] -2methylimidazol-1-ylmethyl} benzonitrile, 4- {3- [4- (2-oxo-2H pyridin-1 -yl) benzyl] -3H-imidazol-4-ylmethyl} benzonitrile, - {3- [4- (5-chloro-2-oxo-2H- [1,2 '] bipyridin-5'-ylmethyl] -3H imidazole -4-ylmethyl} benzonitrile, 4- {3- [4- (2-oxo-2H [1,2 '] bipyridin-5'-ylmethyl] -3H-imidazol4-ylmethyl} benzonitrile, 4- [3- (2-oxo-1-phenyl-1,2-dihydropyridin-4-ylmethyl) -3H-midazol-4-ylmethyl} benzonitrile 18,19-dihydro-19-oxo-5H, 17H -6.10: 12,16-dimethen-1-Himidazo [4,3-c] [1,11,4] dioxaazacyclo-nonadecin-9carbonitrile, (±) -19,20-dihydro-19-oxo-5H- 18,21-ethane

12,14-etene-6,10-metene-22H-benzo [d] imidazo [4,3-k] Δ2010528 [1,6,9,12] oxatriaza-cyclooctadecin-9-carbonitrile, 19,20dihydro 19-oxo-5H, 17H-18,21-ethane-6,10: 12,16-dimethene-22Himidazo [3,4-h] [1,8,11,14] oxatriazacyclocenosine-9carbonitrile, and (±) - 19,20-dihydro-3-methyl-19-oxo-5H-18,21ethane-12,14-etene-6,10-metene-22H-benzo [d] imidazo [4,3k] [1,6 9,12] oxa-1riazacyclocytadecine-9-carbonitrile. Other examples of prenyl protein transferase inhibitors can be found in the following publications and patents: WO 96/30343, WO 97/18813, WO 97/21701, WO 97/23478, WO 97/38665, WO 98/28980, WO 98 / 29119, WO 95/32987, Pat. No. 5,420,245, Pat. No. 5,523,430, Pat. US No.

5,532,359, Pat. No. 5,510,510, Pat. No. 5,589,485, Pat. No. 5,602,098, Publ. European Patent Application 0 618 221, Publ. European Patent Application 0 675 112, Publ. European Patent Application 0 604 181, Publ. European Patent Application 0 696 593, WO 94/19357, WO 95/08542, WO 95/11917, WO 95/12612, WO 95/12572, WO 95/10514, Pat. No. 5,661,152, WO 95/10515, WO

95/10516, WO 95/24612, WO 95/34535, WO 95/25086, WO 96/05529, WO 96/06138, WO 96/06193, WO 96/16443, WO 96/21701, WO

96/21456, WO 96/22278, WO 96/24611, WO 96/24612, WO 96/05168, WO 96/05169, WO 96/00736, Pat. US No. 5,571,792, WO

96/17861, WO 96/33159, WO 96/34850, WO 96/34851, WO 96/30017, WO 96/30018, WO 96/30362, WO 96/30363, WO 96/31111, WO

WO 96/31478, WO 96/31501, WO 97/00252, WO 97/03047, WO 97/03050, WO 97/04785, WO 97/02920, WO 97/17070, WO

97/23478, WO 97/26246, WO 97/30053, WO 97/44350, WO 98/02436, and US Patent No. 5,532,359. For an example of the role of a prenyl protein transferase inhibitor in angiogenesis, see European J. of Cancer 35 (9): 1394-1401 (1999).

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Angiogenesis inhibitors refer to compounds which may inhibit the formation of new blood vessels, regardless of the mechanism. Examples of angiogenesis inhibitors include, but are not limited to, tyrosine kinase inhibitors, such as Flt-1 (VEGFR1) and Flk-1 / KDR (VEGFR2) tyrosine receptor inhibitors, epidermis-derived growth factor inhibitors , fibroblast derivatives, or plaguette derivatives, MMP (matrix metalloprotease) inhibitors, integrin bloggers, interferon-alpha, interleukin-12, pentosan polysulfate, cyclooxygenase inhibitors, including non-steroidal anti-inflammatory drugs (NSAIDs) such as aspirin and ibuprofen as well as selective cyclooxygenase-2 inhibitors such as celecoxib and rofecoxib (PNAS 89: 7384

<td> (1992);</td><td>JNCI</td><td> 69:475</td><td> (1982);</td><td>Arch.</td><td>Ophthalmol. 108: 573</td>

<td> (1990);</td><td>Anat.</td><td>Rec.,</td><td> (238):68</td><td> (1994);</td><td>FEBS Letters 372: 83</td>

<td> (1995);</td><td>Clin,</td><td>Orthop</td><td> . 313:76</td><td> (1995);</td><td>J. Mol. Endocrinol.</td>

16: 107 (1996); Jpn. J. Pharmacol. 75: 105 (1997); Cancer Res. 57: 1625 (1997); Cell 93: 705 (1998); Intl. J. Mol. Med. 2: 115 (1998); J. Biol. Chem. 274: 9116 (1999)), steroidal anti-inflammatory drugs (such as corticosteroids, mineralocorticoids, dexamethasone, prednisone, prednisolone, methylpred, betamethasone), carboxyamidotriazole, combretastatin A4, esgualamine, 6-O-chloroacetylcarbonyl) -fumagilol, thalidatidol , troponin-1, angiotensin II antagonists (see Fernandez et al., J. Lab. Clin. 105: 141-145 (1985)) and antibodies to VEGF (see, Nature Biotechnology, 17: 963-968 (October 1999); Kim et al.,

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Nature, 362: 841-844 (1993); WO 00/44777; and WO 00/61186). Other therapeutic agents that modulate or inhibit angiogenesis, and which may also be used in combination with compounds of preferred embodiments, include agents that modulate or inhibit coagulation and fibrinolysis systems (see review in Clin. Chem. La. Med. 38: 679692 (2000)). Examples of such agents that modulate or inhibit coagulation and fibrinolysis pathways include, but are not limited to, heparin (see Thromb. Haemost. 80: 10-23 (1998)), low molecular weight heparins, and carboxypeptidase U inhibitors. (also known as active thrombin activatable fibrinolysis inhibitor inhibitors [TAFIa]) (see Thrombosis Res. 101: 329-354 (2001)). TAFIa inhibitors have been described in PCT publication WO 03/013, 526 and US Ser. 60/349,925 (filed Jan. 18, 2002). The disclosure also encompasses combinations of compounds of preferred embodiments with NSAIDs which are selective COX-2 inhibitors (generally defined as those having a specificity to inhibit COX2 over COX-1 by at least about 100-fold).

<td>as i</td><td>measured</td><td>for the reason</td><td>from IC5C</td><td colspan="2">j of COX-2 relative to</td>

<td>IC50 of</td><td>COX-1</td><td>evaluated</td><td>per</td><td colspan="2">cellular tests or</td>

<td colspan="2">microsomes).</td><td colspan="2">Such compounds</td><td>include but</td><td>are not</td>

<td>limited</td><td>to</td><td>described</td><td colspan="2">in U.S. Pat. US No.</td><td> 5,474,995,</td>

<td>granted</td><td>in 12</td><td>from Dec.</td><td> 1995,</td><td>Pat. US No.</td><td> 5,861,419,</td>

<td>granted</td><td>in 19</td><td>from Jan of</td><td> 1999,</td><td>Pat. US No.</td><td> 6,001,843,</td>

<td>granted</td><td>in 14</td><td>from Dec.</td><td> 1999,</td><td>Pat. US No.</td><td> 6, 020,343,</td>

<td>granted</td><td>in 1</td><td>from Feb.</td><td> 2000,</td><td>Pat. US No.</td><td> 5,409,944,</td>

issued Apr. 25, 1995, Pat. No. 5,436,265,

102010528

<td>granted</td><td>in</td><td>25 of</td><td>Jul</td><td>in</td><td>1995, Pat.</td><td>US</td><td>No. 5,536,752,</td>

<td>granted</td><td>in</td><td colspan="2">16 Jul.</td><td>in :</td><td>L996, Pat.</td><td>US</td><td>No. 5,550,142,</td>

<td>granted</td><td>in</td><td>27 of</td><td>Ag o</td><td>. in</td><td>1996, Pat.</td><td>US</td><td>No. 5,604,260,</td>

<td>granted</td><td>in</td><td>18 of</td><td>Feb</td><td>. in</td><td>1997, Pat.</td><td>US</td><td>No. 5,698,584,</td>

<td>granted</td><td>in</td><td>16 of</td><td>Ten</td><td>. in</td><td>1997, Pat.</td><td>US</td><td>No. 5,710,140,</td>

<td>granted</td><td>in</td><td>2 0 of</td><td>Jan.</td><td>in</td><td colspan="3">1998, WO 94/15932, published in</td>

<td>Jul 21</td><td>in</td><td> 1994,</td><td>Pat</td><td>. U.</td><td>S. No. 5,344</td><td> , 991,</td><td>granted on 6</td>

<td>from Jun. 6</td><td>in</td><td> 1994,</td><td>Pat.</td><td>US</td><td>. No. 5,134,</td><td> 142,</td><td>granted on 28</td>

July 1992, Pat. No. 5,380,738, issued October 10,

<td>Jan.</td><td>in</td><td> 1995,</td><td>Pat.</td><td>US</td><td>At the.</td><td> 5,393,790,</td><td>granted</td><td>in</td><td> 20</td><td>in</td>

<td>Feb.</td><td>in</td><td> 1995,</td><td>Pat.</td><td>US</td><td>At the.</td><td> 5,466,823,</td><td>granted</td><td>in</td><td> 14</td><td>in</td>

<td>Nov.</td><td>in</td><td> 1995,</td><td>Pat.</td><td>US</td><td>At the.</td><td> 5,633,272,</td><td>granted</td><td>in</td><td> 27</td><td>in</td>

<td>May</td><td>in</td><td> 1997,</td><td>and Pat</td><td>. US</td><td>At the.</td><td> 5,932,598,</td><td>granted</td><td>in</td><td colspan="2">3 Aug</td>

Representative COX-2 inhibitors that are useful include 3-phenyl-4- (4- (methylsulfonyl) phenyl) -2- (5H) -furanone;

and 5-chloro-3- (4-methylsulfonyl) phenyl-2- (2-methyl-5-pyridinyl) pyridine. Compounds which are described as specific COX-2 inhibitors and are therefore useful in preferred embodiments and their syntheses may be found in the following patents, pending applications and

<td>publications:</td><td>WO 94/15932,</td><td>published</td><td>in</td><td> 21</td><td>in</td><td>Jul</td><td>in</td><td> 1994,</td>

<td>Pat. US No</td><td> . 5,344,991,</td><td>granted</td><td>in</td><td> 6</td><td>in</td><td>Jun</td><td>in</td><td> 1994,</td>

<td>Pat. US No</td><td> . 5,134,142,</td><td>granted</td><td>in</td><td> 28</td><td>in</td><td>Jul</td><td>in</td><td> 1992,</td>

<td>Pat. US No</td><td> . 5,380,738,</td><td>granted</td><td>in</td><td> 10</td><td>in</td><td>Jan.</td><td>in</td><td> 1995,</td>

<td>Pat. US No</td><td> . 5,393,790,</td><td>granted</td><td>in</td><td> 20</td><td>in</td><td>Feb.</td><td>in</td><td> 1995,</td>

<td>Pat. US No</td><td> . 5,466, 823,</td><td>granted</td><td>in</td><td> 14</td><td>in</td><td>Nov.</td><td>in</td><td> 1995,</td>

<td>Pat. US No</td><td> . 5,633,272,</td><td>granted</td><td>in</td><td> 27</td><td>in</td><td>May</td><td>in</td><td> 1997,</td>

<td>Pat. US No</td><td> . 5,932,598,</td><td>granted</td><td>in</td><td> 3</td><td>in</td><td>Aug</td><td>in</td><td> 1999,</td>

<td>Pat. US No</td><td> . 5,474,995,</td><td>granted</td><td>in</td><td> 12</td><td>in</td><td>Ten .</td><td>in</td><td> 1995,</td>

102010528

<td>Pat.</td><td>US</td><td>At the .</td><td> 5,861,419,</td><td>granted</td><td>in</td><td> 19</td><td>in</td><td>Jan.</td><td>in</td><td> 1999,</td>

<td>Pat.</td><td>US</td><td>At the .</td><td> 6,001,843,</td><td>granted</td><td>in</td><td> 14</td><td>in</td><td>Ten .</td><td>in</td><td> 1999,</td>

<td>Pat.</td><td>US</td><td>At the.</td><td> 6, 020,343,</td><td>granted</td><td>in</td><td> 1</td><td>in</td><td>Feb.</td><td>in</td><td> 2000,</td>

<td>Pat.</td><td>US</td><td>At the.</td><td> 5,409,944,</td><td>granted</td><td>in</td><td> 15</td><td>in</td><td>Apr</td><td>in</td><td> 1995,</td>

<td>Pat.</td><td>US</td><td>At the .</td><td> 5,436,265,</td><td>granted</td><td>in</td><td> 25</td><td>in</td><td>Jul</td><td>in</td><td> 1995,</td>

<td>Pat.</td><td>US</td><td>At the .</td><td> 5,536,752,</td><td>granted</td><td>in</td><td> 16</td><td>in</td><td>Jul</td><td>in</td><td> 1996,</td>

<td>Pat.</td><td>US</td><td>At the .</td><td> 5,550,142,</td><td>granted</td><td>in</td><td> 27</td><td>in</td><td>Aug</td><td>in</td><td> 1996,</td>

<td>Pat.</td><td>US</td><td>At the .</td><td> 5,604,260,</td><td>granted</td><td>in</td><td> 18</td><td>in</td><td>Feb.</td><td>in</td><td> 1997,</td>

<td>Pat.</td><td>US</td><td>At the.</td><td> 5,698,584,</td><td colspan="7">issued Dec. 16, 1997, and</td>

<td>Pat.</td><td>US</td><td>At the .</td><td> 5,710,140,</td><td colspan="2">granted on 20</td><td>in</td><td colspan="2">Jan. 20</td><td>in</td><td> 1998 .</td>

Other examples of angiogenesis inhibitors include, but are not limited to, endostatin, ukrain, ranpirnase, IM862, 5-methoxy-4- [2-methyl-3- (3-methyl-2-butenyl) oxiranyl] 1-oxaspiro [ 2,5] oct-6-yl (chloroacetyl) carbamate, acetylidinanine, 5-amino-1 - [[3,5-dichloro-4- (4-chlorobenzoyl) phenyl] methyl] -1H-1,2,3-one triazole-4-carboxamide, CM101, squalamine, combretastatin, RPI4610, NX31838, sulfated mannopentaose phosphate, 7,7- (carbonyl-bis [imino-N-methyl4,2-pyrrolocarbonylimino [N-methyl-4,2-pyrrol] carbonylimino] -bis (1,3-naphthalene disulfonate) and 3 - [(2,4 - dimethylpyrrol-5-yl) methylene] -2-indolinone (SU5416).

Agents that interfere with cell cycle checkpoints are compounds that can inhibit protein kinases that transduce cell cycle checkpoint signals, thereby sensitizing the cancer cell to DNA-damaging agents. Such agents include ATR, ATM, Chkl and Chk2 kinase inhibitors and cdk and cdc kinase inhibitors and are specifically exemplified by 7ΡΕ2010528 hydroxy restaurantsporine, flavopyridol, CYC202 (Cyclacel) and

BMS-387032.

Cell proliferation and survival signaling inhibitors may be pharmaceutical agents that can inhibit cell surface receptors and signal transduction cascades downstream of these surface receptors. Such agents include EGFR inhibitors (e.g. gefitinib and erlotinib), ERB-2 inhibitors (e.g. trastuzumab), IGFR inhibitors, cytokine receptor inhibitors, MET inhibitors, PI3K inhibitors (e.g. LY294002) , serine / threonine kinases (including but not limited to Akt inhibitors such as described in WO 02/083064, WO 02/083139, WO 02/083140 and WO 02/083138), Raf kinase inhibitors (e.g. BAY- 439006), MEK inhibitors (e.g., CI-1040 and PD-098059) and mTOR inhibitors (e.g. Wyeth CCI-779). Such agents include small molecule inhibitory compounds and antibody antagonists.

Apoptosis-inducing agents include activators of TNF receptor family members (including TRAIL receptors).

The disclosure provides representative agents useful in combination with compounds of preferred embodiments for the treatment of cancer include, for example, irinotecan, topotecan, gemcitabine, 5-fluorouracil, carboplatin leucovorin, cisplatin, taxanes, tezacitabine,

102010528 cyclophosphamide, vinca alkaloids, imatinib (Gleevec), anthracyclines, rituximab, trastuzumab, as well as other cancer chemotherapeutic agents.

The foregoing compounds to be used in combination with the compounds of preferred embodiments may be used in therapeutic amounts as indicated in the Physicians' Desk Reference (PDR).^The Edition (1993), or such therapeutically useful amounts, as would be known to one skilled in the art.

Compounds of preferred embodiments and other anticancer agents may be administered at the maximum recommended clinical dosage, or at lower doses. Dosage levels of the active compounds in the compositions of the preferred embodiments may be varied to obtain a desired therapeutic response, depending upon the route of administration, the severity of the disease and the patient's response. The combination may be administered as separate compositions, or as a single dosage form containing both agents. When administered as a combination, the therapeutic agents may be formulated as separate compositions which are administered at the same time or at different times or the therapeutic agents may be administered as a single composition.

General Synthesis Methods

102010528

Compounds of preferred embodiments may be prepared from readily available starting materials using the following general methods and procedures. It will be appreciated that when typical or preferred process conditions are given (i.e. reaction temperatures, times, reactant molar ratios, solvents, pressures, etc.) other process conditions may also be used unless otherwise indicated. Optimal reaction conditions may vary with the particular reagents or solvents used, but such conditions may be determined by one of ordinary skill in the art by routine optimization procedures.

In addition, as will be apparent to those skilled in the art, conventional protecting groups may be required to prevent certain functional groups from being subjected to undesired reactions. Suitable protecting groups for various functional groups, as well as the conditions suitable for protecting and deprotecting particular functional groups, are well known in the art. For example, numerous protection groups are described in TW Greene and GM. Wuts, Protecting Groups in Organic Synthesis, 3^The Edition, Wiley, New York, 1999, and cited references.

In addition, the compounds of preferred embodiments contain one or more chiral centers. Accordingly, if desired, such compounds may be

102010528 prepared or isolated as pure stereoisomers, i.e. as individual enantiomers or diastereomers, or as mixtures enriched with stereoisomers. All such stereoisomers (and enriched mixtures) are included within the scope of the embodiments unless otherwise indicated. Pure stereoisomers (or enriched mixtures) may be prepared using, for example, optically active starting materials or stereoselective reagents well known in the art.

Alternatively, racemic mixtures of such compounds may be separated using, for example, chiral column chromatography, chiral resolving agents and the like.

Starting materials for the following reactions are generally known compounds or may be prepared by known procedures or obvious modifications thereof. For example, many of the starting materials are available from commercial suppliers, such as Aldrich Chemical Co. (Milwaukee, Wisconsin, USA), Bachem (Torrance, California, USA), Emka-Chemce, or Sigma (St. Louis, Missouri, USA). ). Others may be prepared by procedures, or their obvious modifications, described in reference texts, such as Reacents for Organic Synthesis, Volumes 1-15 (John Wiley and Sons, 1991) by Fieser and Fieser, Organic Reactions, Volumes 1-40 ( John Wiley and Sons, 1991), Rodd's Chemistry of Carbon Compounds, Volumes 1-5 and Supplements (Elsevier Science Publishers, 1989), Advanced Organic Chemistry, (John Wiley and Sons, 4^The March Edition and Comprehensive Organic Transformations (VCH Publishers Inc., 1989) by Larock.

102010528

The various starting materials, intermediates and compounds of preferred embodiments may be isolated and purified where appropriate using conventional techniques such as precipitation, filtration, crystallization, evaporation, distillation and chromatography. The characterization of these compounds can be performed using conventional methods such as melting point, mass spectrum, nuclear magnetic resonance and various other spectroscopic analyzes.

The compounds of the embodiments can generally be prepared using a number of methods familiar to one skilled in the art, such as for example the methods described in US patent publication Nos. US20040087626 A1 and US20040122237 A1. The compounds of embodiments may generally be prepared according to the following Reaction Reference Schemes 1-8, which are described in detail in the Examples and Reference Examples below.

Reference Schemes 1-8 illustrate general methods for preparing intermediates and compounds of the Examples and Reference Examples. These compounds are prepared from starting materials known in the art or are commercially available. Specific compounds are for illustrative purposes only.

Reference Scheme 1

102010528

<img file="PT2010528T_D0001.tif" />

In Reference Scheme 1, 2-hydroxyaniline or a derivative thereof reacts with ethylxantic acid to give a thiolbenzoxazole. Thiol-benzoxazole is converted to a chlorobenzoxazole with reaction with thionyl chloride. Alternatively, thiol-benzoxazole may be converted to halogenated benzoxazole with a set of halogenating agents, such as but not limited to phosphorus trichloride, phosphorus tribromide, phosgene, or oxalyl chloride. 0 Chloro-benzoxazole is then reacted with a benzylamine, such as 2-chlorobenzylamine to give a benzylamino-benzoxazole. Benzylamino-benzoxazole is coupled with chloro-pyridine in the presence of a base such as cesium carbonate to give a compound of the Examples or Reference Examples. Alternatively, a halogenated pyridine may be used for coupling.

102010528

Reference Scheme 2

<img file="PT2010528T_D0002.tif" />

<img file="PT2010528T_D0003.tif" />

In Reference Scheme 2, a thiol-benzoxazole, or a derivative thereof, is alkylated at the thiol moiety. The alkylated thiolbenzoxazole is coupled with the corresponding halopyridine such as chloropyridine in the presence of a base such as cesium carbonate to give a compound of the Examples or Reference Examples. The resulting benzoxazolyloxypyridine is oxidized, for example, with mCPBA. Other oxidizing agents may be used to oxidize the thiol to a sulfoxide. Other oxidizing agents include, but are not limited to, hydrogen peroxide, sodium periodate, pyridinium chlorochromate, or chromium trioxide. Benzoxazolyloxy pyridine sulfoxide is subjected to nucleophilic attack with an amine to give a compound of the Examples or Reference Examples.

Reference Scheme 3

102010528

<img file="PT2010528T_D0004.tif" />

In Reference Scheme 3, cyanoacetate and 1,5-dibromopentane are coupled to form 1-cyano-cyclohexanecarboxylic acid ethyl ester after cyclization. This product is reduced with Raney hydrogen and nickel. Other reducing agents may be used to reduce the nitrile group to an amine. Other reducing agents include, but are not limited to, catalytic hydrogenation using platinum oxide, or Raney nickel, or lithium aluminum hydride, diisobutyl aluminum hydride, sodium borohydride, or lithium triethylborohydride. The reduced product is coupled with sulfoxobenzoxazolyloxypyridine. The resulting coupling reaction product may further be functionalized or derivatized. For example, in Reference Scheme 3, an ester group may be converted to a carboxylic acid group by hydrolysis and then converted to an amide by reaction with an amine. These reactions are conversions well known to one skilled in the art.

Reference Scheme 4

102010528

<img file="PT2010528T_D0005.tif" />

In Reference Scheme 4, 3- (2-piperidin-1-ylethyl) phenylamine is an example of an amine which may be used to form a compound of the Examples or Reference Examples. 3- (2-Piperidin-1-yl-ethyl) phenylamine is formed from the sulfonation of 2- (3-nitrophenyl) -ethanol, then amination of the resulting methanesulfonic acid 2- (3-nitrophenyl) -ethyl ester and subsequently reduced to the resulting 1- [2- (3-nitrophenyl) ethyl] piperidine.

Reference Scheme 5

<img file="PT2010528T_D0006.tif" />

In Reference Scheme 5, 4- (2- (methylsulfinyl)

102010528 benzo [d] oxazol-6-yloxy) -N-methylpyridine-2-carboxamide is aminated with cyclohexyl methanamine. The resulting 4- (2- (cyclohexylmethylamino) benzo [d] oxazol-6-yloxy) -N-methylpyridine-2carboxamide is then hydrolyzed to form 4- [2- (cyclohexylmethyl-amino) -benzooxazol-6-acid yloxy] pyridine-2-carboxylic acid. 4- [2- (Cyclohexylmethylamino) -benzooxazol-6-yloxy] -pyridine-2-carboxylic acid then reacts with benzotriazol-1-yloxy-tris (dimethylamino) -phosphonate, tert-butyl-carbazate and triethylamine hexafluorophosphate to form 4- (2- (cyclohexylmethylamino) benzo [d] oxazol-6-yloxy) pyridine-2-carbohydrazide. 4- (2- (Cyclohexylmethylamino) benzo [d] oxazol-6-yloxy) pyridine-2carbohydrazide then reacts with trimethyl orthoformate to form a compound of the Examples or Examples of

Reference.

Reference Scheme 6

<img file="PT2010528T_D0007.tif" />

<img file="PT2010528T_D0008.tif" />

In Reference Scheme 6, a compound of the Examples or Reference Examples may be further functionalized. For example, 4- (2- (cyclohexylmethylamino) benzo [d] oxazol-6-yloxy) pyridine-2carboxylic acid is reduced to {4- (2- (cyclohexylmethylamino)

102010528 benzo [d] oxazol-6-yloxy) pyridin-2-yl} methanol with borane. Other suitable reducing agents include, but are not limited to, lithium aluminum hydride, aluminum hydride, diisobultyl aluminum hydride, sodium borohydride, or lithium triethylborohydride. {4- (2- (Cyclohexylmethylamino) benzo [d] oxazol-6-yloxy) pyridin-2-yl} methanol is then oxidized to 4- (2- (cyclohexylmethylamino) benzo [d] oxazol-6- yloxy) pyridine-2-carbaldehyde with Dess-Martin reagent. Other suitable oxidizing agents include, but are not limited to, pyridinium chlorochromate, SO3 pyridine in DMSO, or conditions commonly referred to as a Swern or Moffet oxidation. 4- (2- (Cyclohexylmethylamino) benzo [d] oxazol-6-yloxy) pyridin-2-carbaldehyde is then converted to cyclohexylmethyl- {6- [2- (5-methyl-1H-imidazol2-yl ) -pyridin-4-yloxy] -benzooxazol-2-yl} -amine by reaction with pyruvic aldehyde.

Reference Scheme 7

<img file="PT2010528T_D0009.tif" />

OO

102010528

In Reference Scheme 7, a compound of the Examples or Reference Examples is synthesized from the reaction of 4- (2-methanesulfinylbenzooxazol-6-yloxy) -pyridine-2-carboxylic acid methylamide (from Example 2) with 1- (2,3-dihydro-benzo [1,4] dioxin-5-yl) -ethylamine. In one example, 1- (2,3-dihydro-benzo [1,4] dioxin-5-yl) -ethylamine is synthesized through a resin.

A resin-bound amine is reacted with 2,3dihydro-benzo [1,4] dioxin-5-carbaldehyde, thereby giving C (2,3-dihydro-benzo [1,4] dioxin-5-yl ) -methylenamine. AC (2,3-dihydro-benzo [1,4] dioxin-5-yl) methylenamine is derivatized with alkylation at the imino site, such as methylmagnesium bromide. 1- (2,3-Dihydrobenzo [1,4] dioxin-5-yl) methylenamine is cleaved from the resin. An example of a resin cleavage agent is trifluoroacetic acid (TFA). The resulting 1- (2,3-dihydro-benzo [1,4] dioxin-5-yl) ethylamine may be used for the synthesis of a compound of the Examples or Reference Examples. For example, 1- (2,3-dihydro-benzo [1,4] dioxin-5-yl) -ethylamine may be used to react with 4- (2-methanesultin-1-benzooxazol-6-yloxy) -pyridine-1-one. Carboxylic acid 2-methylamide to form 4- {2- [1- (2,3-dihydro-benzo [1,4] dioxin-5-yl) ethylamino] -benzooxazol-6-yloxy} -pyridine-2-methylamide 2-carboxylic acid.

Reference Scheme 8

102010528

<img file="PT2010528T_D0010.tif" />

In Reference Scheme 8, 2mercapto-benzothiazol-6-ol was prepared according to United States Patent No. 4,873,346. 2-Mercapto-benzothiazol-6-ol is then converted to 2-methylsulfanyl-benzothiazol-6-ol by conventional procedures to remove an ether protecting group. Reaction of 2-methylsulfanyl-benzothiazol-6-ol with methyl iodide provides alkylation at the thiol position. Reaction of 2-methylsulfanyl-benzothiazol-6-ol with 4-chloro-pyridine-2-carboxylic acid methylamide gives 4- (2-methylsulfanyl-benzothiazol-6-yloxy) -pyridine-2-carboxylic acid methylamide. Subsequent oxidation of 4- (2-methylsulfanyl-benzothiazol-6-yloxy) -pyridine-2-carboxylic acid methylamide yields 4- (2-methanesulfinyl-benzothiazol-6-yloxy) -pyridine-2-carboxylic acid methylamide. 4- (2-Methanesulfinylbenzothiazol-6-yloxy) -pyridine-2-carboxylic acid methylamide may be a substrate for the reaction with various amines. For example, 4- (2-methanesulfinyl-benzothiazol-6yloxy) -pyridine-2-carboxylic acid methylamide may be reacted with cyclohexylmethylamine to give 4- [2ΡΕ2010528 (cyclohexylmethyl-amino) -benzothiazol-6-yloxy-acid methylamide ] -pyridine-2carboxylic.

Reference Scheme 9

<img file="PT2010528T_D0011.tif" />

Y = F, Ciamine

<img file="PT2010528T_D0012.tif" />

<img file="PT2010528T_D0013.tif" />

<img file="PT2010528T_D0014.tif" />

S.7

In Reference Scheme 9, benzothiazoles or benzoxazoles of formula 9.1 may be reacted with a substituted amine to provide intermediates of formula 9.2. Treatment of intermediates of formula 9.2 with a reagent such as BBr3 provides phenols of formula 9.3. 0 Subsequent treatment of intermediates of formula 9.3 with 4-halopyridines of formula 9.4 at temperatures generally ranging from, but not limited to, room temperature to 130 ° C provides, in the presence of a base such as, for example, potassium carbonate or of cesium, compounds of formula 9.5. Further treatment with boronic acids or stananes under conditions known to those skilled in the art such as Suzuki or Stille reactions provides compounds of formula 9.6. In addition, treating a compound of formula 9.5 with an amine

Substituted ΡΕ2010528, under conditions known to those skilled in the art as Buchwald reaction or SnAr reaction, provide compounds of formula 9.7.

Reference Scheme 10

<img file="PT2010528T_D0015.tif" />

In Reference Scheme 10, benzoxazoles or benzothiazoles of formula 10.5 and 10.6 may be prepared starting with a 4-halopyridine of formula 10.1 which may be (1) treated with boronic acids or stannans under conditions known to those skilled in the art as reactions. Suzuki or Stille, provides intermediates of formula 10.2 or (2) reacted with a substituted amine under conditions known to those skilled in the art as a Buchwald or SnAr reaction to provide intermediates of formula 10.3. Subsequent reaction of intermediates of formula 10.2 or 10.3 with a phenolic intermediate of formula 10.4, in the presence of a base, such as for example potassium or cesium carbonate in a solvent such as dimethylformamide, acetonitrile or dioxane, provides compounds. of formula 10.5 and 10.6.

EXAMPLES

102010528

With reference to the following examples and reference examples, compounds of preferred embodiments have been synthesized using the methods described herein, or other methods that are known in the art.

The compounds and / or intermediates were characterized by high performance liquid chromatography (HPLC) using a Waters Millenium chromatography system with a 2695 separation module (Milford, MA). Analytical columns were Phenomenex Luna 08 -5 μ, 4.6 x 50 mm from Alltech (Deerfield, IL). Gradient elution (flow rate 2.5 mL / min) was used, typically starting with 5% acetonitrile / 95% water and progressing to 100% acetonitrile over a 10 minute period. All solvents contained 0.1% trifluoroacetic acid (TFA). Compounds were detected by ultraviolet light (UV) absorption at 220 or 254 nm. HPLC solvents were from Burdick and Jackson (Muskegan, MI), or Fisher Scientific (Pittsburgh, PA).

In some cases, purity was assessed by thin layer chromatography (TLC) using glass or plastic supported silica gel plates, such as Baker-Flex Silica Gel 1B2-F flexible sheets. TLC results were readily detected visually under ultraviolet light, or employing the well-known iodine vapor and other various staining techniques.

102010528

Mass spectrometric analysis was performed on one of two LCMS instruments: a Waters system (HPLC Alliance HT and a Micromass ZQ mass spectrometer; Column: Eclipse XDB-C18, 2.1 x 50 mm; gradient: 5-95% ( or 35-95%, or 65-95% or 95-95%) acetonitrile in water with 0.05% TFA over a period of 4 min; flow rate 0.8 mL / min; molecular weight range 200-1500, Cone voltage 20 V; column temperature 40 ° C) or a Hewlett Packard system (1100 HPLC Series; Column: Eclipse XDB-C18, 2.1 x 50 mm; gradient: 5-95% acetonitrile in water with 0.05% TFA over a period of 4 min; flow rate 0.8 mL / min; molecular weight range 150-850; Cone voltage 50 V; column temperature 30 ° C). All masses were recorded as those of the corresponding protonated ions.

GCMS analysis is performed on a Hewlett instrument

Packard (HP6890 Series Gas Chromatograph with Selective Mass Detector 5973; injector volume: 1 µl, initial column temperature: 50 ° C; final column temperature: 250 ° C; ramp time: 20 minutes, gas flow rate) : 1 mL / min column: 5% phenylmethylsiloxane, model No. HP 190915-443; dimensions: 30.0 mx 25 mx 0.25 m).

Nuclear magnetic resonance (NMR) analysis was performed on some of the compounds with a Varian 300 MHz NMR instrument (Paio Alto, CA). The spectral reference was TMS or the known chemical shift of the solvent. Some samples of the compound were run at temperatures

102010528 (eg 75 ° C) to promote increased sample solubility.

The purity of some of the compounds is assessed by elemental analysis (Desert Analytics, Tucson, AZ).

Melting points are determined on a Mel-Temp laboratory device apparatus (Holliston, MA).

Preparative separations are performed using a Flash 40 and KP-Sil, 60A chromatography system (Biotage, Charlottesville, VA), or by flash column chromatography using silica gel as a filler (230-400 mesh), or by HPLC using a Waters 2767 Sample Manager, C-18 reverse phase column, 30X50 mm, flow rate 75 mL / min. Typical solvents used for the Flash 40 Biotage system and flash column chromatography are dichloromethane, methanol, ethyl acetate, hexane, acetone, aqueous ammonia (or ammonium hydroxide) and triethylamine. Typical solvents employed for reverse phase HPLC have varying concentrations of acetonitrile and water with 0.1% trifluoroacetic acid.

It should be understood that organic compounds according to preferred embodiments may exhibit the phenomenon of tautomerism. Since the chemical structures within this description may only represent one of the possible tautomeric forms, it should be understood that preferred embodiments include any form.

Tautomeric da2010528 of the drawn structure.

The examples below, as well as throughout the order, the following abbreviations have the following meanings. If

<td>is not defined, the</td><td>terms have their meanings</td>

<td>generally accepted.</td><td></td>

<td>Abbreviations</td><td></td>

<td>ACN</td><td>Acetonitrile</td>

<td>BINAP</td><td>2,2'-bis (diphenylphosphine) - 1,1'-binaftyl</td>

<td>DCM</td><td>Dichloromethane</td>

<td>DIEA</td><td>diisopropylethylamine</td>

<td>DIPEA</td><td>N, N-diisopropylethylamine</td>

<td>DME</td><td>1,2-dimethoxyethane</td>

<td>DMF</td><td>N, N-dimethylformamide</td>

<td>DMSO</td><td>dimethyl sulfoxide</td>

<td>DPPF</td><td> 1,1' -</td>

<td>EtOAc</td><td>bis (diphenylphosphine) ferrocene ethyl acetate</td>

<td>EtOH</td><td>ethanol</td>

<td>HATU</td><td>2- (7- Hexafluorophosphate)</td>

<td>HPLC</td><td>aza-1H-benzotriazol-1-yl) - 1,1,3,3-tetramethyluronium High light chromatography</td>

<td>MC PB A</td><td>efficiency metha-</td>

<td>MeOH</td><td>chloroperoxybenzoic acid methanol</td>

102010528

<td>NBS</td><td>N-bromosuccinimide</td>

<td>NMP</td><td>N-methyl-2-pyrrolidone</td>

<td>OK</td><td>Room temperature</td>

<td>THF</td><td>Tetrahydrofuran</td>

Compounds according to any one of Claims 1 to 4

Reference Example 2

4- [2 - ((IS, 2R) -2-Hydroxy-indan-lilamino) -benzoxazol-6-yloxy] -pyridine-2-carboxylic acid methylamide (Table

<img file="PT2010528T_D0016.tif" />

<img file="PT2010528T_D0017.tif" />

Step 1. Synthesis of 2- (methylthio) benzo [d] oxazol-6-ol

<img file="PT2010528T_D0018.tif" />

102010528

Triethylamine (1.87 g, 18.56 mmol, 2.0 eq) and methyl iodide (1.77 g, 13.92 mmol, 1.5 eq) were added to a solution of 2-mercaptobenzo [d] oxazol-6-ol (1.55 g, 9.28 mmol, 1.0 eq) in 20 mL of methylene chloride at room temperature. The reaction mixture was stirred at room temperature for 3 hours. The mixture was diluted with 100 mL methylene chloride. The resulting mixture was washed with water (10 mL), brine (10 mL) and then dried over MgSO 4, filtered and evaporated under reduced pressure to give crude product, which was purified by silica gel column eluted with ethyl acetate and hexane to give the title compound. MH + = 182.

Step 2. Synthesis of 4- (2- (methylthio) benzo [d] oxazol-6-yloxy) -N-methylpyridine-2-carboxamide

<img file="PT2010528T_D0019.tif" />

4-Chloro-N-methylpyridine-2carboxamide (16.0 g, 93.4 mmol, 2.0 eq) and cesium carbonate (45.7 g, 140.1 mmol, 3.0 eq) were added to a solution of 2- (methylthio) benzo [d] oxazol-6-ol (8.5 g, 46.7 mmol, 1 eq) in 80 mL of N, W-dimethylformamide. The reaction mixture was stirred at 75 ° C for 6 hours. After the mixture was cooled to room temperature, 120 mL of water was added to the mixture. After filtration, the solid was purified by silica gel column eluted with acetate.

102010528 ethyl and hexane to give the title compound. MH + =

316.

Step 3. Synthesis of 4- (2- (methylsulfinyl) benzo [d] oxazol-6yloxy) -N-methylpyridine-2-carboxamide

<img file="PT2010528T_D0020.tif" />

3-chloroceroxybenzoic acid (70%, 989 mg, 4.4 mmol, 1.1 eq) was added to a solution of 4- (2- (methylthio) benzo [d] oxazol-6-yloxy) -N-methylpyridine -2-carboxamide (1.26 g, 4.0 mmol, 1.0 eq) in 40 mL of methylene chloride. The reaction mixture was stirred at room temperature for 5 hours and then diluted with 200 mL of methylene chloride. The resulting mixture was washed with aqueous sodium bicarbonate and brine and then dried over MgSO 4, filtered and evaporated under reduced pressure to give crude product, which was used for the next step without further purification. MH + = 332.

Step 4. 4- [2 - ((IS, 2R) -2-Hydroxy-indan-1-ylamino) -benzooxazol-6-yloxy] -pyridine-2-carboxylic acid methylamide

102010528

A solution of 4- (2- (methylsulfinyl) benzo [d] oxazol-6-yloxy) -77-methylpyridine-2-carboxamide (17 mg, 0.05 mmol, 1.0 eq) and (IS, 2R ) -1-amino-2,3-dihydro-1Hinden-2-ol (30 mg, 0.2 mmol, 4.0 eq) in 1 mL of N, N-dimethylacetamide in the microwave at 90 ° C for 600 seconds The crude product was purified by reverse phase preparative HPLC to give the title compound. MH = 417.0.

Reference Example 15 (Reference Scheme 8)

Preparation of 4- [2- (Cyclohexylmethylamino) -benzothiazol-6-yloxy] -pyridine-2-carboxylic acid methylamide (Table

<img file="PT2010528T_D0021.tif" />

Step 1. Preparation of 2-Mercapto-benzothiazol-6-ol

<img file="PT2010528T_D0022.tif" />

According to the United States Patent

4,873,346 - Substituted Benzothiazoles, Benzimidazoles and

102010528 benzoxazoles; Anderson, David J .; The Upjohn Company, Kalamazoo, Michigan; Oct 10, 1989. M + H = 184.0.

Step 2. Preparation of 2-Methylsulfanyl-benzothiazol-6-ol

<img file="PT2010528T_D0023.tif" />

<img file="PT2010528T_D0024.tif" />

Triethylamine (7.29 mL, 51.91 mmol, 2.5 eq) was added to the 2-mercapto-benzothiazol-6-ol solution from step 1 (3.80 g, 20.76 mmol, 1.0 eq). ) in ice-cold DCM (40 mL, 0.5 M) at 0 ° C, followed by iodomethane (1.93 mL, 31.14 mmol, 1.5 eq). The reaction was stirred at 0 ° C to -10 ° C for hours. The solvent was removed under vacuum. Water (ca. 200 mL) was added and the aqueous layer was extracted with ethyl acetate (3 x 150 mL). The organic layer was dried over sodium sulfate, filtered and evaporated under vacuum to give 2-methylsulfanyl-benzothiazol-6-ol as a light green powder (3.76 g, 92%). The crude product was used for the next step without purification. M + H = 198.0

Step 3. Preparation of 4- (2-Methylsulfanyl-benzothiazol-6-yloxy) -pyridine-2-carboxylic acid methylamide

102010528

<img file="PT2010528T_D0025.tif" />

CSCO3 (15.54 g, 47.70 mmol, 2.5 eq) was added to the solution of 2-methylsulfanyl benzothiazole-6-oi (3.76 g, 19.08 mmol, 1.0 eq) in DMF (25 mL) at room temperature. After stirring for a while, 4-chloro-pyridine-2-carboxylic acid methylamide (4.86 g, 28, 62 mmol, 1.5 eq) was added to the mixture and the mixture was stirred at 70 ° C. under a reflux condenser overnight. After cooling the reaction mixture in an ice bath, water (100 mL) was added and the aqueous layer was extracted with ethyl acetate (3 x 150 mL). The organic layer was dried over sodium sulfate, filtered and evaporated in vacuo. The crude product was purified using 20 g of ISCO silica gel column (0% -50% -80% -100% ethyl acetate-hexane mixture over a 45 min run, 40 mL / min) to yield methylamide. 4- (2-Methylsulfanyl-benzothiazole-6-yloxy) -pyridine-2-carboxylic acid (3.88 g, 62%) as a white solid. M + H = 332.1

Step 4. Preparation of 4- (2-Methanesulfinyl-benzothiazol-6-yloxy) -pyridine-2-carboxylic acid methylamide

<img file="PT2010528T_D0026.tif" />

<img file="PT2010528T_D0027.tif" />

102010528

MCPBA (77%, 2.88 g, 1.1 eq) was added to the 4- (2-methylsulfanyl-benzothiazol-6yloxy) -pyridine-2-carboxylic acid methylamide solution from step 3 (3.88 g, 11.72 mmol, 1.0 eq) in DCM (20 mL) at 0 ° C. The mixture was stirred at this temperature for one hour. Saturated sodium bicarbonate solution (100 mL) was added. The aqueous layer was extracted with DCM (3X150 mL). The organic layer was dried over sodium sulfate, filtered and evaporated under vacuum to afford 4- (2-methanesulfinylbenzothiazol-6-yloxy) -pyridine-2-carboxylic acid methylamide as a white powder in quantitative yields. The crude product was used for the next step without purification. M + H = 348.0.

Step 5. Preparation of 4- [2- (Cyclohexylmethyl-amino) -benzothiazol-6-yloxy] -pyridine-2-carboxylic acid methylamide

<img file="PT2010528T_D0028.tif" />

O

102010528

Cyclohexylmethylamine (18.7 µl, 0.144 mmol, 2.0 eq) was added to 4- (2-methanesulfinyl-benzothiazol-6-yloxy) -pyridine-2-carboxylic acid methylamide solution (25 mg, 0.072 mmol, 1.0 eq) in DMF (500 µl) and the reaction was stirred at 70 ° C overnight. The single reaction mixture was purified by preparative reverse phase HPLC. Pure fractions were lyophilized as TFA salts. M + H = 397.1

Example 16

Preparation of 4- (2 - ((IS, 2S) -2-hydroxy-cyclohexylamino) benzo [d] thiazol-6-yloxy) -N-methylpicolinamide (Table 2, Compound 137)

<img file="PT2010528T_D0029.tif" />

<img file="PT2010528T_D0030.tif" />

(IS, 2S) -2-Aminocyclohexanol Hydrochloride (92 mg, 0.606 mmol, 3.0 eq) was added to 4- (2-methanesulfinyl-benzothiazol-6-yloxy) pyridine-2-carboxylic acid methylamide solution (70 mg, 0.202 mmol, 1.0 eq, Reference Example 15-step 4) in DMA (600 µL), followed by diisopropylethylamine (0.21 mL, 1.21 mmol). The reaction was heated at 110 ° C for 24 hours. The single reaction mixture was purified by preparative reverse phase HPLC. Pure fractions were lyophilized as TFA salts. M + H

ΡΕ2010528 = 398

The compounds in Table 2 below were prepared by the general procedures described above.

Compound

Table 2 '' '1 M + H; .......

[tR (min)

Compound Name

137

Structure

<img file="PT2010528T_D0031.tif" />

] 4- [2] ((IS, 2S) -2-hydroxy-cyclo, 94] hexylamino) -benzooxazol-6] yloxy] -pyridine-2] carboxylic acid] methylamide [(2) (( IS, 2S) -2-hydroxy-cyclo, 94] hexylamino) -benzothiazol-6 [yloxy] -pyridine-2 [carboxylic acid]

<img file="PT2010528T_D0032.tif" />

) 4- [2] ((IR, 2R) -2-hydroxy-cyclo1,94] hexylamino) -benzothiazol-6] yloxy] -pyridine-2] carboxylic acid methylamide

Reference Example 162

Preparation of 4- (2 - ((IR, 2S) -2-hydroxy-2,3-dihydro-1Hinden-1-ylamino) benzo [d] thiazol-6-yloxy) -N-methylpicolinamide as a compound was prepared according to the general squad below

Step 1

Hn

<img file="PT2010528T_D0033.tif" />

<img file="PT2010528T_D0034.tif" />

DIPEA to NMP

<img file="PT2010528T_D0035.tif" />

102010528

(IR, 2S) -1-Amino-2,3-dihydro-1Hinden-2-ol (597 mg, 4 mmol) and DIPEA (300 µL, 1.73 mmol) were added to the N-methyl solution. -4- (2- (methylsulfinyl) benzo [d] thiazol-6yloxy) picolinamide (300 mg, 0.86 mmol) in 5 mL of NMP. The reaction solution was stirred at 105 ° C for 24 hours. The crude reaction solution was purified by preparative HPLC and evaporated under vacuum to give 4- (2 - ((1R, 2S) -2-hydroxy-2,3-dihydro-1H-indenylamino) benzo [d ] thiazol-6-yloxy) -N-methylpicolinamide (347 mg, 0.63 mmol) as a TFA salt. ES / MS m / z 433.1 (MH⁺) .

Reference Example 170

Preparation of N- (cyclohexylmethyl) -6- (2- (ethylamino) pyridin4-yloxy) benzo [d] thiazol-2-amine

Step 1

Cl o

<img file="PT2010528T_D0036.tif" />

DIPEA (9 µL, 0.05 mmol) and 70% ethylamine in water (200 µL, 2.51 mmol) were added to the 6- (2-chloropyridin-4-yloxy) -N- (cyclohexylmethyl) reaction solution benzo [d] thiazol-2-amine (12 mg, 0.03 mmol) in 400 µl NMP. The reaction mixture was stirred at 110 ° C for 96 hours, or until prepared by LC. The crude reaction mixture was filtered, purified by preparative HPLC and evaporated under vacuum to give N- (cyclohexylmethyl) -6- (2ΡΕ2010528 (ethylamino) pyridin-4-yloxy) benzo [d] thiazol-2-amine as salt of TFA (1.8 mg). ES / MS m / z 383.1 (MH +).

Example 171

N-cyclopropyl-4- (2 - ((1R, 2R) -2-hydroxy-cyclohexylamino) benzo [d] thiazol-6-yloxy) picolinamide

The compound in question was prepared according to the general scheme below: Step 1

<img file="PT2010528T_D0037.tif" />

Step 2

<img file="PT2010528T_D0038.tif" />

<img file="PT2010528T_D0039.tif" />

<img file="PT2010528T_D0040.tif" />

N

Step 3

<img file="PT2010528T_D0041.tif" />

Step 5

O πο η,

N

<img file="PT2010528T_D0042.tif" />

HATU DIEA in NMP

H_sN “&lt;3

<img file="PT2010528T_D0043.tif" />

<img file="PT2010528T_D0044.tif" />

102010528

Step 1 . Synthesis of tert-Butyl 4- (2- (methylthio) benzo [d] thiazol-6-yloxy) picolinate

Tert-Butyl 4-chloropicolinate (8.13 g, 38.07 mmol, 1.5 eq) and cesium carbonate (20.67 g, 63.45 mmol, 2.5 eq) were added to a solution of 2- (methylthio) benzo [d] thiazol-6-ol (5.0 g, 25.38 mmol, 1.0 eq) in 25 mL of N, W-dimethylformamide. The reaction mixture was stirred at 75 ° C for 6 hours. After the mixture was cooled to room temperature, 120 mL of water was added and the aqueous phase was extracted with ethyl acetate (3 x 150 mL), the combined organic layers were dried over sodium sulfate. After filtration, the solid was purified by silica gel column eluted with 0% to 50% ethyl acetate-hexane mixture to give 5.84 g of the title compound as brown powder (62%). MH + = 375.

Step 2. Synthesis of tert-Butyl 4- (2- (methylsulfinyl) benzo [d] thiazol-6yloxy) picolinate

3-Chloroperoxybenzoic acid (77%, 3.84 g, 17.17 mmol, 1.1 eq) was added to a solution of tert 4- (2- (methylthio) benzo [d] thiazol-6-yloxy) picolinate -butyl (5.84 g, 15.61 mmol, 1.0 eq) in 25 mL of methylene chloride. The reaction mixture was stirred at room temperature for 1.5 hours and then diluted with 200 mL of methylene chloride. The resulting mixture was washed with aqueous sodium bicarbonate and brine and then dried over MgSO4.

102010528 filtered and evaporated under reduced pressure to give crude product, which was used for the next step without further purification. MH + = 391.0.

Step 3. Preparation of tert-Butyl 4- (2 - ((IR, 2R) -2-hydroxy-cyclohexylamino) -benzo [d] thiazol-6-yloxy) picolinate

(IR, 2R) -cyclohexane-1,2-diamine (581 mg, 3.84 mmol) and DIPEA (0.955 mL, 5.76 mmol) were added to the solution of 4- (2- (methylsulfinyl) benzo tert-Butyl [d] thiazol-6-yloxy) picolinate (500 mg, 1.25 mmol) in 10 mL of NMP. The reaction solution was stirred at 100 ° C for 3 days. The crude reaction solution was purified by preparative HPLC and evaporated under vacuum to give tert-butyl 4- (2 - ((IR, 2R) -2-hydroxycyclohexylamino) benzo [d] thiazol-6-yloxy) picolinate (240 mg 0.544 mmol) as a white powder. ES / MS m / z 442.5 (MH +).

Step 4. Preparation of 4- (2 - ((IR, 2R) -2-hydroxy-cyclohexylamino) benzo [d] thiazol-6-yloxy) picolinic acid

Hydrochloric acid (1 mL, 6 mmol) was added to the solution of tert-butyl 4- (2 - ((1R, 2R) -2-hydroxy-cyclohexylamino) benzo [d] thiazol-6-yloxy) picolinate (250 mg, 0566 mmol) in 10 mL of acetonitrile. The reaction solution was stirred at room temperature for 1 hour and then at 60 ° C for 2 hours. The crude reaction solution was concentrated and redissolved with 10 mL of

102010528 acetonitrile. The resulting solution was evaporated under vacuum to give a light brown oily product 4- (2 ((IR, 2R) -2-hydroxycyclohexylamino) benzo [d] thiazol-6yloxy) picolinic acid (215 mg, 0.56 mmol ). ES / MS m / z 386.5 (MH⁺) .

Step 5. Preparation of N-Cyclopropyl-4- (2 - ((IR, 2R) -2hydroxy-cyclohexylamino) -benzo [d] thiazol-6yloxy) picolinamide

Cyclopropylamine (7 µL mg, 30 pmol) was added to the 4- (2 - ((IR, 2R) -2-hydroxy-cyclohexylamino) benzo [d] thiazol-6-yloxy) picolinic acid reaction solution (5 mg, 39 pmol), HATU (15 mg, 39 pmol) and DIPEA (14 µL, 78 pmol) in 1 mL of NMP. The reaction solution was stirred at room temperature for 12 hours. The crude reaction solution was purified by preparative HPLC and evaporated under vacuum to give N-cyclopropyl-4- (2 - ((IR, 2R) -2-hydroxy-cyclohexylamino) benzo [d] thiazol-6-yloxy) picolinamide (1 mg, 2.3 pmol) as white powder. ES / MS m / z 425.2 (MH +).

Example 173

Preparation of (IR, 2R) -2- (6- (2-chloropyridin-4-yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol

The compound under management was prepared according to the general scheme below:

102010528

Step 1

<img file="PT2010528T_D0045.tif" />

Step 3

<img file="PT2010528T_D0046.tif" />

<img file="PT2010528T_D0047.tif" />

Step 1.

Preparation of (IR, 2R) -2- (6-methoxybenzo [d] thiazol-2ylamino) cyclohexanol.

(IR, 2R) -2-Aminocyclohexanol hydrochloride (910 mg, 6 mmol) and DIPEA (2.44 mL, 14 mmol) were added to the solution of 2-chloro-6-methoxybenzo [d] thiazole (1.0 g, 5 mmol) in 5.5 mL of NMP. The reaction solution was stirred at 115 ° C for 96 hours. The crude reaction solution was purified by preparative HPLC to give purified fractions which were combined and neutralized with solid NaHCO3. The resulting solution was extracted with ethyl acetate (2 x 300 mL). The combined organic layers were washed with water (60 mL) and brine (60 mL) and then dried over Na 2 SO 4.₄ and evaporated under vacuum to give (IR, 2R) -2- (6-methoxybenzo [d] thiazol-2-ylamino) cyclohexanol (1.06 g, 3.81 mmol) as an ivory solid. ES / MS m / z 279.1 (MH⁺) .

102010528

Step 2. Preparation of 2 - ((IR, 2R) -2-hydroxy-cyclohexylamino) benzo [d] thiazol-6-ol.

1M Boron tribromide in DCM (8 mL, 8 mmol) was slowly added to the solution of (IR, 2R) -2- (6-methoxybenzo [d] thiazol-2-ylamino) cyclohexanol (1.06 g, 3 , 81 mmol) in 16 mL of DCM at 0 ° C. The reaction solution was stirred at room temperature for 2 hours. Removal of all solvents under vacuum followed by quenching with water (ca. 30 mL) and dilute NaHCO 3 solution and extraction of the aqueous phase with ethyl acetate (3 x 100 mL) and drying of combined organic extracts over Na₂SO4 and subsequent removal of ethyl acetate under vacuum afforded the desired product (1.16 g) as a pink solid. The residue was purified by flash column chromatography to give 2 - ((IR, 2R) -2-hydroxy-cyclohexylamino) benzo [d] thiazol-6-ol (1.0 g, 3.78 mmol) as a brown solid. ES / MS m / z 265.1 (MH +).

Step 3. Preparation of (IR, 2R) -2- (6- (2-chloropyridin-4yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol.

2-Chloro-4-fluoropyridine (263 mg, 2 mmol) was added to the mixture of 2 - ((IR, 2R) -2-hydroxy-cyclohexylamino) benzo [d] thiazol-6-ol (265 mg, 1 mmol) and cesium carbonate (651 mg, 2 mmol) in 3 mL of NMP. The reaction mixture was stirred at 60 ° C for 20 hours. The crude reaction mixture was filtered and then purified by preparative HPLC to give (IR, 2R) -2- (6- (2-chloropyridin-4-yloxy) benzo

102010528 [d] thiazol-2-ylamino) cyclohexanol as powder (341 mg, 0.9 mmol). ES / MS m / z 376.0 (MH +).

Example 174

Preparation of (IR, 2R) -2- (6- (2- (1-methyl-1H-pyrazol-4yl) pyridin-4-yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol

<img file="PT2010528T_D0048.tif" />

<img file="PT2010528T_D0049.tif" />

Step 4. Preparation of (IR, 2R) -2- (6- (2- (1-methyl-1H-pyrazol-4-yl) pyridin-4-yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol.

1-Methyl-4- (4,4,5,5-tetramethyl-1,2,2dioxaborolan-2-yl) -1H-pyrazole (21 mg, 100 pmol), Pd (dppfHC® (4 mg, 5 pmol) and 2M Na 2 CO 3 (100 µL, 200 pmol) to the reaction mixture of (IR, 2R) -2- (6- (2-chloropyridin-4yloxy) benzo [d] thiazol-2-ylamino) cyclohexane. hexanol (20 mg, 40 pmol) in 400 µl of DME The reaction mixture was stirred at

90 ° C for 24 hours. The reaction mixture was poured into mL of saturated NaHCO 3 solution and extracted with ethyl acetate (2 x 30 mL). The combined organic layers were washed with water (2 x 10 mL) and brine (20 mL), then dried over Na 2 SO 4 and evaporated under vacuum to give a brown solid (65 mg) which was purified on preparative HPLC to

102010528 give (1R, 2R) -2- (6- (2- (1-methyl-1H-pyrazol-4-yl) pyridin4-yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol as powder (6.4 mg). ES / MS m / z 422.2 (MH⁺) .

Example 175

Preparation of (IR, 2R) -2- (6- (2- (1-methyl-1H-imidazol-5yl) pyridin-4-yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol compound was prepared according to the general scheme below:

<img file="PT2010528T_D0050.tif" />

Preparation of (IR, 2R) -2- (6- (2- (1-methyl-1H-imidazol-5yl) pyridin-4-yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol.

Pd (dppf) was added₂Cl₂ (70.2 mg, 0.0088 mmol), LiCl (19 mg, 0.44 mmol) and then 1-methyl-5- (tributylstannyl) -1H-imidazole (44 mg, 0.117 mmol) to the reaction mixture of (IR, 2R) -2- (6- (2-chloropyridin-4-yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol (11 mg, 0.029 mmol) in 0.5 mL of DMF. The reaction solution was stirred at 105110 ° C for 18 hours or prepared by LC. The crude reaction mixture was filtered, purified by preparative HPLC and lyophilized to give (IR, 2R) -2- (6- (2- (1ΡΕ2010528 methyl-1H-imidazol-5-yl) pyridin-4-yloxy) benzo [ d] thiazol-2ylamino) cyclohexanol as TFA salt (3.5 mg). ES / MS m / z 422.1 (MH +).

Example 176

Preparation of (IR, 2R) -2- (6- (2- (1- (2,2-difluoroethyl) -1H-pyrazol-4-yl) pyridin-4-yloxy) benzo [d] thiazol-2-ylamino) cyclo -hexanol compound in management was prepared from the general squirt below:

a deal with

<img file="PT2010528T_D0051.tif" />

<img file="PT2010528T_D0052.tif" />

Preparation of (IR, 2R) -2- (6- (2- (1- (2,2-difluoroethyl) -1H-pyrazol-4-yl) pyridin-4-yloxy) benzo [d] thiazol-2-ylamino) cyclo -hexanol.

Cesium carbonate (672 mg, 2.06 mmol) was added to the reaction mixture of 4- (4,4,5,5-tetramethyl-1,2,2dioxaborolan-2-yl) -1H-pyrazole (210 mg, 1.08 mmol) in 2.0 mL NMP. The reaction mixture was stirred for 5 minutes and then 1,1-difluoro-2-iodoethane (197 mg, 1.03 ml) was added.

102010528 mmol) and stirred at RT for 40 hours. The above crude reaction mixture was removed (0.8 mL, 0.432 mol) for use. (The remaining 1.2 mL were stored in the freezer). (IR, 2R) -2- (6- (2-chloropyridin-4yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol (20 mg, 0.053 mmol), Pd (dppf) 2Cl2 (150 mL, 2 mg, 0.019 mmol) and 2 M Na 2 CO 3 (0.150 mL, 0.3 mmol) at 0.8 mL of the reaction mixture. The reaction mixture was microwaved at 140 ° C for 720 seconds. The crude reaction mixture was filtered, purified by preparative HPLC and lyophilized to give (IR, 2R) -2- (6- (2- (1- (2,2-difluoroethyl) -1H-pyrazol-4-yl) pyridin-4 -yloxy) benzo [d] thiazol-2ylamino) cyclohexanol as a TFA salt (4.6 mg). ES / MS m / z 472.0 (MH +).

Example 177

Preparation of (IR, 2R) -2- (6- (2- (4-methyl-1H-imidazol-2yl) pyridin-4-yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol

The compound in question was prepared according to the general scheme below:

102010528

Step 1

<img file="PT2010528T_D0053.tif" />

Step 1. Preparation of 4- (2 - ((IR, 2R) -2-hydroxycyclohexylamino) benzo [d] thiazol-6-yloxy) picolinaldehyde.

Cesium carbonate (232 mg, 0.71 mmol) and 4-chloropicolinaldehyde (125 mg, 0.883 mmol) were added to the reaction mixture of 2 - ((IR, 2R) -2-hydroxy-cyclohexylamino) benzo [d] thiazole -6-ol (90 mg, 0.34 mmol) in 1.9 mL of NMP. The reaction mixture was stirred at RT for 10 minutes and then incubated in a microwave at 150 ° C for 750 seconds. The crude reaction mixture was filtered, purified by preparative HPLC and lyophilized to give 4- (2 - ((1R, 2R) -2-hydroxy-cyclohexylamino) benzo [d] thiazol-6yloxy) picolinaldehyde as TFA salt ( 88 mg). ES / MS m / z 388.1 (MH⁺) as the hydrate (+ 18).

Step 2. Preparation of (IR, 2R) -2- (6- (2- (4-methyl-1H-imidazol-2-yl) pyridin-4-yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol.

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Ammonium acetate (32 mg, 0.41 mmol) and 40 wt.% 2-oxopropanal solution of water (0.037 mL, 0.21 mmol) were added to the reaction mixture of 4- (2 - ((IR, 2R) -2-hydroxycyclohexylamino) benzo [d] thiazol-6-yloxy) picolinaldehyde (16 mg, 0.041 mmol) in 0.75 mL of MeOH. The reaction mixture was stirred at 70 ° C for 2 hours. The crude reaction mixture was concentrated, redissolved in 0.8 mL DMF, filtered, purified by preparative HPLC and lyophilized to give (1R, 2R) -2- (6- (2- (4-methyl-1H -imidazol-2-yl) pyridin-4yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol as TFA salt (3.2 mg). ES / MS m / z 422.1 (MH +).

Example 178

Preparation of (IR, 2R) -2- (6- (3-bromopyridin-4-yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol

The compound in question was prepared according to the general scheme below:

<img file="PT2010528T_D0054.tif" />

Preparation of (IR, 2R) -2- (6- (3-bromopyridin-4-yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol.

Cesium carbonate (39 mg, 0.18 mmol) was added to the reaction mixture of 2 - ((IR, 2R) -2-hydroxycycloΡΕ2010528 hexylamino) benzo [d] thiazol-6-ol (12.5 mg, 0.047 mmol). ) in 0.4 mL NMP and stirred at RT for 1-3 minutes. To this mixture was added 3-bromo-4-chloropyridine (18.2 mg, 0.094 mmol). The reaction mixture was stirred at 90 ° C for 4 hours or until prepared by LC. The crude reaction mixture was filtered, purified by preparative HPLC and lyophilized to afford (IR, 2R) -2- (6- (3-bromopyridin-4yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol salt. TFA (9.2 mg). ES / MS m / z 420.1 / 422.0 (MH +).

EXAMPLE 179

Preparation of (IR, 2R) -2- (6- (3- (1-methyl-1H-pyrazol-4yl) pyridin-4-yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol

The compound in question was prepared according to the general scheme below:

/

<img file="PT2010528T_D0055.tif" />

NMP

Preparation of (IR, 2R) -2- (6- (3- (1-methyl-1H-pyrazol-4yl) pyridin-4-yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol.

Pd (dppf) 2Cl2 (80.8 mg, 0.0107 mmol), 1-methyl-4- (4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl) - 1H pyrazole (30 mg, 0.143 mmol) and 2M Na₂CO3 (0.12 mL, 0.24

102010528 mmol) to the reaction mixture of (1R, 2R) -2- (6- (3-bromopyridin4-yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol (15 mg, 0.036 mmol) in 0.5 mL from NMP. The reaction solution was stirred at 105-110 ° C for 2 hours or until processed by LC. The crude reaction mixture was filtered, purified by preparative HPLC and lyophilized to give (1R, 2R) -2- (6- (3- (1-methyl-1H-pyrazol-4-yl) pyridin-4-yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol as a TFA salt (5.5 mg). ES / MS m / z 422.1 (MH +).

Reference Example 180

Preparation of 4- (2- (cyclohexylmethylamino) benzo [d] thiazol-6yloxy) picolinonitrile

The compound in question was prepared according to the general scheme below: Step 1

<img file="PT2010528T_D0056.tif" />

Step 1. Preparation of N- (cyclohexylmethyl) -6-methoxybenzo [d] thiazol-2-amine.

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Cyclohexyl methanamine (865 mg, 7.65 mmol) and DIPEA (1.57 mL, 9.0 mmol) were added to the solution of 2-chloro-6-methoxybenzo [d] thiazole (900 mg, 4.5 mmol) in 4.5 mL of NMP. The reaction solution was stirred at 105-110 ° C for 66 hours. The reaction was treated by adding 250 mL of ethyl acetate and washed with 2 x 60 mL of saturated NaHCO 3, 3 x 60 mL of water, 1 x 60 mL of saturated NaCl, dried over sodium sulfate, filtered and concentrated in vacuo to give N- (cyclohexylmethyl) -6-methoxybenzo [d] thiazol-2-amine as solid (1.18 grams). ES / MS m / z 277.1 (MH⁺) .

Step 2. Preparation of 2- (cyclohexylmethylamino) benzo [d] thiazol-6-ol.

1M Boron tribromide in DCM (10.6 mL, 10.6 mmol) was added slowly to the solution of N- (cyclohexylmethyl) 6-methoxybenzo [d] thiazol-2-amine (1.40 g, 5 µM, 05 mmol) in 12 mL of DCM for about 3 minutes at 0 ° C. The reaction solution was stirred at 0 ° C for 20 min and then at RT for 2 h.

The reaction mixture was concentrated to a solid. To the residual solids is added 200 mL of ethyl acetate and mL of water and stirred at RT for 10 minutes. With stirring, excess solid NaHCCb is carefully added until a basic solution is obtained. Stir at RT for about 1 hour to dissolve the solids. The aqueous layer is removed and extracted with 100 mL of ethyl acetate. The organic layers are combined and washed with 1 x 30 mL water, 1 x 25 mL saturated NaCl solution and dried with sulfur.

102010528 sodium. This mixture was filtered through a silica gel bed (1.25 in. X 3 in.) And washed with ethyl acetate. The filtrate was concentrated under reduced pressure to give 2- (cyclohexylmethylamino) benzo [d] thiazol-6-ol as solid (1.32 grams). ES / MS m / z 263.1 (MH⁺) .

Step 3. Preparation of 4- (2- (cyclohexylmethylamino) benzo [d] thiazol-6-yloxy) picolinonitrile.

Cesium carbonate (56 mg, 0.171 mmol) was added to the reaction mixture of 2- (cyclohexylmethylamino) benzo [d] thiazol-6-ol (18 mg, 0.068 mmol) in 0.4 mL of NMP and stirred. at RT for 1-3 minutes. To this mixture was added 4-chloropicolinonitrile (19 mg, 0.136 mmol). The reaction mixture was stirred at 60 ° C for 5 hours or until prepared for LC. The crude reaction mixture was filtered, purified by preparative HPLC and lyophilized to give 4- (2- (cyclohexylmethylamino) benzo [d] thiazol-6yloxy) picolinonitrile as TFA salt (9.8 mg). ES / MS m / z 365.1 (MH +).

EXAMPLE 183

Preparation of (IR, 2R) -2- (6- (2-morpholinopyridin-4-yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol

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<img file="PT2010528T_D0057.tif" />

<img file="PT2010528T_D0058.tif" />

DIPEA, NMP

<img file="PT2010528T_D0059.tif" />

<img file="PT2010528T_D0060.tif" />

<img file="PT2010528T_D0061.tif" />

Preparation of (IR, 2R) -2- (6- (2-morpholinopyridin-4-yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol.

(DIPEA) diisopropylethylamine (13 µL, 0.074 mmol) and morpholine (49 mg, 0.558 mmol) were added to the (IR, 2R) -2- (6- (2-chloropyridin-4-yloxy) benzo) reaction mixture [d] thiazol-2-ylamino) cyclohexanol (14 mg, 0.037 mmol) in 0.4 mL of NMP. The reaction mixture was stirred at 110 ° C for 48 hours or until prepared by LC. The crude reaction mixture was filtered, purified by preparative HPLC and lyophilized to give (1R, 2R) -2- (6- (2-morpholinopyridin4-yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol salt. TFA (3.7 mg). ES / MS m / z 427.1 (MH +).

EXAMPLE 187

Preparation of (IR, 2R) -2- (6- (2- (1H-imidazol-1-yl) pyridin-4yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol

<img file="PT2010528T_D0062.tif" />

Preparation of (IR, 2R) -2- (6- (2- (1H-imidazol-1-yl) pyridin-4-2010528 yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol.

(DIPEA) diisopropylethylamine (17 µL, 0.096 mmol) and 1H-imidazole (180 mg, 2.64 mmol) were added to the (IR, 2R) -2- (6- (2-chloropyridin) reaction mixture -4-yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol (12 mg, 0.032 mmol) in 0.55 mL of NMP. The reaction mixture was followed by LC, LCMS and microwaved as follows: (150 ° C for 750 seconds, 230 ° C for 750 seconds, 250 ° C for 1000 seconds, again at 250 ° C for 1000 seconds) . The crude reaction mixture was filtered, purified by preparative HPLC and lyophilized to give (IR, 2R) -2- (6- (2- (1H-imidazol-1-yl) pyridin-4-yloxy) benzo [d] thiazol-2-one. 2ylamino) cyclohexanol as a TFA salt (1.4 mg). ES / MS m / z 40 8.2 (MH +).

Example 188

Preparation of (IR, 2R) -2- (6- (2- (1,2,3,6-tetrahydropyridin4-yl) pyridin-4-yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol

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STEPS

Step 1 ci

<img file="PT2010528T_D0063.tif" />

<img file="PT2010528T_D0064.tif" />

4M HCI in Dioxane

Hf

<img file="PT2010528T_D0065.tif" />

PH

Step 1. Preparation of 4- (4- (2 - ((IR, 2R) -2-hydroxycyclohexylamino) benzo [d] thiazol-6-yloxy) pyridin-2-yl) -5,6-dihydropyridine-1 (2H tert-Butyl carboxylate

Added

Tert-Butyl 4- (4,4,5,5-tetramethyl-1,3,2dioxaborolan-2-yl) -5,6-dihydropyridine-1 (2H) -carboxylate (36 mg, 0.117 mmol), Pd (dppf) 2 Cl 2 (70.2 mg, 0.0088 mmol) and 2 M Na₂CO3 (0.125 mL, 0.25 mmol) to the reaction mixture of (IR, 2R) -2- (6- (2-chloropyridin-4-yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol (11 mg 0.029 mmol) in 0.5 mL DME. The reaction solution was stirred at 105-110 ° C for 24 hours or until processed by LC. The crude reaction mixture was concentrated to a solid, redissolved in 0.8 mL DMF, filtered, preparative HPLC purified and lyophilized to give 4- (4- (2 - ((IR, 2R) -2hydroxycyclohexylamino) benzo [d] thiazol-6-yloxy) pyridin-2yl) -5,6-dihydropyridine-1 (2H) carboxylate as a TFA salt (2.5 mg). ES / MS m / z 523.1 (MH +).

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Step 2. Preparation of (IR, 2R) -2- (6- (2- (1,2,3,6-tetrahydropyridin-4-yl) pyridin-4-yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol.

4M HCL in Dioxane (1 mL, 4.0 mmol) was added to 4- (4- (2 - ((IR, 2R) -2-hydroxycyclohexylamino) benzo [d] thiazol-6-yloxy) pyridin solid Tert-Butyl -2-yl) -5,6-dihydropyridin-1- (2H) -carboxylate (2.5 mg, 0.0039 mmol). The reaction mixture was stirred at RT for minutes. The crude reaction mixture was concentrated to a solid and lyophilized to give (IR, 2R) -2- (6- (2- (1,2,3,6-tetrahydropyridin-4-yl) pyridin-4-yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol as an HCl salt (1.4 mg). ES / MS m / z 423.1 (MH +).

Reference Example 192

Preparation of (IR, 2R) -2- (6- (6-fluoroquinolin-4-yloxy) benzo [d] thiazol-2-ylamino) cyclohexane

The compound in question was prepared according to the general scheme below:

F

<img file="PT2010528T_D0066.tif" />

Preparation of (IR, 2R) -2- (6- (6-fluoroquinolin-4-yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol.

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100

Cesium carbonate (47 mg, 0.143 mmol) was added to the reaction mixture of 2 - ((IR, 2R) -2-hydroxy-cyclohexylamino) benzo [d] thiazol-6-ol (15.1 mg, 0.057 mmol) in 0.4 mL NMP and stirred at RT for 1-3 minutes. To this mixture was added 4-chloro-6-fluoroquinoline (21 mg, 0.114 mmol). The reaction mixture was stirred at 105-110 ° C for 18 hours or until prepared by LC. The crude reaction mixture was filtered, purified by preparative HPLC and lyophilized to give (IR, 2R) -2- (6- (6-fluoroquinolin-4yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol salt. TFA (9.2 mg). ES / MS m / z 410.1 (MH +).

Example 195

4- (7-bromo-2 - ((1R, 2R) -2-hydroxy-cyclohexylamino) benzo [d] thiazol-6-yloxy) -N-methylpicolinamide

Br

HO,

<img file="PT2010528T_D0067.tif" />

Step 1. Synthesis of 7-bromo-2-chloro-6-methoxybenzo [d] thiazole

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101

N-Bromosuccinimide (213 mg, 1.20 mmol, 1.2 eq) was added to the solution of 2-chloro-6-methoxy-benzothiazole (200 mg, 1.0 mmol, 1.0 eq) in 5 mL of NMP at room temperature. The reaction mixture was stirred at 75 ° C for&gt; 24 hours with subsequent addition of NBS in small batches for reaction progress, after which the mixture was diluted with water (ca. 100 mL) and the aqueous layer was extracted with ethyl acetate. ethyl (about 150mLX3). The combined organic layers were dried over sodium sulfate, filtered and condensed under reduced pressure. Purification with ISCO using a 0% -50% gradient of ethyl acetate-hexane gave 336 mg of product as a 60% yield white off-white powder, the structure of which was confirmed by H + NMR. LC / MS (m / z) [279.9] (MH +)

Step 2. Synthesis of (IR, 2R) -2- (7-bromo-6-methoxybenzo [d] thiazol-2ylamino) cyclohexanol

Br Br

<img file="PT2010528T_D0068.tif" />

(IR, 2R) -2-Aminocyclohexanol hydrochloride (0.123 mg, 0.809 mmol, 1.5 eq) and DIPEA (263 µL, 1.50 mmol, 2.8 eq) were added to the 7-bromo-2 solution -chloro-6-methoxybenzo [d] thiazole (150 mg, 0.539 mmol, 1.0 eq) in 1 mL of NMP at room temperature. The reaction mixture was stirred at 125 ° C for 12 hours, after which the mixture was diluted.

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102 with saturated sodium bicarbonate solution (about 100 mL) and the aqueous layer was extracted with ethyl acetate (about 200mLX3). The combined organic layers were dried over sodium sulfate, filtered and condensed under reduced pressure to afford the crude product as a brown oil that was sufficiently pure and was carried on to the next step without further purification. LC / MS (m / z) [359.0] (MH +)

Step 3. Synthesis of 7-Bromo-2 - ((IR, 2R) -2-hydroxycyclohexylamino) -benzo [d] thiazol-6-ol

<img file="PT2010528T_D0069.tif" />

1M Boron tribromide solution (about 3.0 mL, 2.68 mmol, 5.0 eq) was added to (1R, 2R) 2- (7-bromo-6-methoxybenzo [d] thiazole solution) 2-ylamino) cyclohexanol (191 mg, 0.537 mmol, 1.0 eq) in 10 mL of DCM at room temperature. The reaction mixture was refluxed for 12 hours, after which the mixture was diluted with saturated sodium bicarbonate (about 100 mL) to pH = 7 and the aqueous layer was extracted with ethyl acetate (about 150mLX3). The combined organic layers were dried over sodium sulfate, filtered and condensed under reduced pressure to give a sufficiently pure crude product which was taken to the next step without purification.

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103 additional LC / MS (m / z) [345.0] (MH⁺) .

Step 4. Synthesis of 4- (7-bromo-2 - ((IR, 2R) -2-hydroxy-cyclohexylamino) benzo [d] thiazol-6-yloxy) -N-methylpicolinamide

Br

<img file="PT2010528T_D0070.tif" />

4-Chloro-N-methylpicolinamide (29 mg, 0.174 mmol, 1.2 eg) and cesium carbonate (165 mg, 0.507 mmol, 3.5 eg) were added to the 7-bromo-2 - ((IR 2R) -2-hydroxy-cyclohexylamino) benzo [d] thiazol-6-ol (50 mg, 0.145 mmol, 1.0 eg) in 1 mL of NMP at room temperature. The reaction mixture was stirred at 80 ° C for approximately 12 hours, then the mixture was purified by reverse phase HPLC. LC / MS (m / z) [479.0] (MH +)

Example 196

4- (2 - ((1R, 2R) -2-hydroxy-cyclohexylamino) -7methylbenzo [d] thiazol-6-yloxy) -N-methylpicolinamide

<img file="PT2010528T_D0071.tif" />

Step 1. Synthesis of 2 - ((IR, 2R) -2-hydroxy-cyclohexylamino) 7-methylbenzo [d] thiazol-6-ol

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104

<img file="PT2010528T_D0072.tif" />

Trimethylboraxine (128 mg, 1.02 mmol, 3.0 eq), Pd (Cl2) dPPf (27 mg, 0.034mmol, 0.1 eq) and 1 ml of 2M Na2 CO3 solution were added to the 7- bromo-2 - ((IR, 2R) -2hydroxy-cyclohexylamino) benzo [d] thiazol-6-ol (117 mg, 0.34 mmol, 1.0 eq) in 1 mL of DMF in a vial waves at room temperature. Then, the reaction mixture was heated in the microwave at 120 ° C for 15 minutes. The reaction was quenched with saturated NaHCO 3 solution (25 mL) and the aqueous phase was extracted with ethyl acetate (50 mL x 3), the combined organic layers were dried over Na 2 SO 4, filtered and condensed under reduced pressure. Purification with ISCO using a 0% - 18% methanol-DCM gradient yielded 17 mg of product as brownish powder in 17% yield. LC / MS (m / z) [279.1] (MH⁺)

Step 2. Synthesis of 4- (2 - ((IR, 2R) -2-hydroxy-cyclohexylamino) -7-methylbenzo [d] thiazol-6-yloxy) -Nmethylpicolinamide

<img file="PT2010528T_D0073.tif" />

Prepared by following the procedure in example 543,

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105 step 4. LC / MS (m / z) [413.1] (MH +)

Example 197

4- (7-chloro-2 - ((1R, 2R) -2-hydroxy-cyclohexylamino) benzo [d] thiazol-6-yloxy) -N-methylpicolinamide

<img file="PT2010528T_D0074.tif" />

Step 1. Synthesis of 7-chloro-2- (methylthio) benzo [d] thiazol-6ol

Cl

<img file="PT2010528T_D0075.tif" />

N-Chlorosuccinimide (507 mg, 3.80 mmol, 1.5 eq) was added to the 2- (methylthio) benzo [d] thiazol-6-ol solution (500 mg, 2.53 mmol, 1.0 eq). ) in 10 mL of NMP at room temperature. The reaction mixture was stirred at room temperature for 1 hour, after which the mixture was diluted with saturated sodium bicarbonate solution (about 100 mL) and the aqueous layer extracted with DCM (about 150mLX3). The combined organic layers were dried over sodium sulfate, filtered and condensed under reduced pressure. Purification with ISCO using 0% 100% ethyl acetate-hexane gradient yielded 283.39 mg of product in 48% yield, structure confirmed by

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106

H + NMR. LC / MS (m / z) [232.0] (MH +)

Step 2. Synthesis of 4- (7-chloro-2- (methylthio) benzo [d] thiazol6-yloxy) -N methyl picolinamide

<img file="PT2010528T_D0076.tif" />

4-Chloro-N-methylpicolinamide (88 mg, 0.519 mmol, 1.5 eg) and cesium carbonate (281 mg, 0.865 mmol, 2.5 eg) were added to the 7-chloro-2- (methylthio) solution. benzo [d] thiazol-6ol (80 mg, 0.346 mmol, 1.0 eg) in 1 mL of NMP at room temperature. The reaction mixture was stirred at 85 ° C for&gt; 48 hours to about 75% to 80% of the complete reaction, after which the mixture was diluted with water (about 50 mL) and the aqueous layer was extracted with ethyl acetate ( about 50mLX3). The combined organic layers were dried over sodium sulfate, filtered and condensed under reduced pressure to give crude product which was purified with ISCO using 0% to 100% gradient of ethyl acetate-hexane mixture to give 64 mg of product. with 50% yield. LC / MS (m / z) [366.0] (MH⁺)

Step 3. Synthesis of 4- (7-chloro-2- (methylsulfinyl) benzo [d] thiazol-6-yloxy) -N-methylpicolinamide

The Cl The Cl

<img file="PT2010528T_D0077.tif" />

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107

MCPBA (33 mg, 0.192 mmol, 1.1 eq) was added to the solution of 4- (7-chloro-2- (methylthio) benzo [d] thiazol-6-yloxy) N-methylpicolinamide (64 mg, 0.175 mmol 1.0 eq) in 5 mL of DCM was added at 0 ° C and the reaction mixture was stirred for 30 to 45 min. Then the reaction was quenched with water (10 mL) and the aqueous phase was extracted with ethyl acetate (25 mLX5), the combined organic layers were dried over sodium sulfate, filtered and condensed under reduced pressure to yield crude product. which was sufficiently pure and was taken to the next step without further purification. LC / MS (m / z) [382.0] (MH⁺)

Step 4. Synthesis of 4- (7-chloro-2 - ((IR, 2R) -2-hydroxycyclohexylamino) benzo [d] thiazol-6-yloxy) -N-methylpicolinamide

The Cl The Cl

<img file="PT2010528T_D0078.tif" />

(1R, 2R) -2-Aminocyclohexanol hydrochloride (6 mg, 0.039 mmol, 1.5 eq) and DIPEA (13 pL, 0.078 mmol, 3.0 eq) were added to the solution of 4- (7 -chloro-2- (methylsulfinyl) benzo [d] thiazol-6-yloxy) -N-methylpicolinamide (10 mg, 0.026 mmol), 1.0 eq) in NMP and the reaction mixture was heated to 160 ° C in microwaves for 15 min. Subsequently, the product was purified by reverse phase HPLC. LC / MS (m / z) [433.1] (MH⁺)

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108

Example 201

4- (2 - ((IR, 2R) -2-hydroxy-cyclohexylamino) benzo [d] oxazol-6yloxy) -N-methylpicolinamide

<img file="PT2010528T_D0079.tif" />

(IR, 2R) -2-Amino-cyclohexanol hydrochloride (17 mg, 0.112 mmol, 1.5 eq) and DIPEA (40 µL, 0.225 mmol, 3.0 eq) were added to the N-methyl-4 solution. - (2- (methylsulfinyl) benzo [d] oxazol-6-yloxy) picolinamide (25 mg, 0.075 mmol, 1.0 eq, described in step 3 of reference example 2) in 1 mL of NMP and the reaction mixture was stirred at room temperature for 48 hours. Subsequently, the product was purified by reverse phase HPLC. LC / MS (m / z) [383.1] (MH +)

Intermediates

Synthesis of 4-Chloro-N-methylpyridine-3-carboxamide

<img file="PT2010528T_D0080.tif" />

Step 1. Thionyl chloride (1.8 mL,

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109

25.0 mmol, 2.5 eq) to a suspension of 4-chloronicotinic acid (1.57 g, 10.0 mmol, 1.0 eq) in 25 mL of toluene at room temperature. The reaction mixture was stirred at 100 ° C for 3 hours. The mixture was concentrated under reduced pressure, dissolved in 25 mL of toluene and concentrated again to give crude 4-chloronicotinoyl chloride hydrochloride salt, which was used for the next step without further purification.

Step 2. Methylamine solution (2M in THF, 20 mL, 40 mmol, 4.0 eq) was added to a suspension of crude 4-chloronicotinoyl chloride hydrochloride in 25 mL of THF at

0 ° C. The reaction mixture was stirred at room temperature for 1 hour and concentrated under reduced pressure. The crude material was dissolved in ethyl acetate (75 mL) and water / brine / saturated sodium bicarbonate solution (1/1/1, 75 mL). The separated aqueous layer was extracted with

EtOAc. The combined organic layers were washed with water / brine / saturated sodium bicarbonate solution (1/1/1, 25 mL) and brine (25 mL) and dried over sodium sulfate. Removal of solvent under reduced pressure afforded the title compound as an orange solid (400 mg, 24%), which was used without further purification. MH + = 171.0, t R = 0.55 min.

Synthesis of 4-chloro-N ', N'-dimethylpyridine-2-carboxyhydrazide

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110

<img file="PT2010528T_D0081.tif" />

N, N-dimethylhydrazine (120 mg, 2.0 mmol, 1.0 eg) and N, N-diisopropylethylamine (383 µL, 2.2 mmol, 1.1 eg) were added to a suspension of 4chloropicolinoyl chloride hydrochloride (352 mg, 2.0 mmol, 1.0 eg) in 10 mL of THF at room temperature. The reaction mixture was stirred for 15 minutes and diluted with water (25 mL) and EtOAc (50 mL). The separated organic layer was washed with brine (25 mL), saturated sodium bicarbonate solution (25 mL) and dried over sodium sulfate. Concentration under reduced pressure gave the title compound as a colorless solid (223 mg, 56%), which was used without further purification. MH + = 200, t R = 1.42 min.

Synthesis of 4-chloro-N-methylpicolinamide and 4chloropicolinoyl chloride

OO

<img file="PT2010528T_D0082.tif" />

Prepared following the method described in A Scaleable Synthesis of BAY 43-9006: A Potent Raf Kinase Inhibitor for the treatment of cancer. Donald Bankston, Jacgues Dumas, Reina Natero, Bernd Riedl, Mary-Katherine Monahan, Robert Sibley .; Bayer Research Center. Pharmaceutical Division. Organic Process Research and

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111

Development 2002 (6) 777-781.

(IR, 2R) -2-Amino-cyclohexanol hydrochloride synthesis nh₂ hci oh

A solution of 4.0 M HCl in dioxane (49 mL, 195 mmol) was slowly added via syringe to a (IR, 2R) - (-) - 2benciloxy-cyclohexylamine ice bath ( 20 g, 97.4 mmol) in dry MeOH (390 mL). The ice bath was removed and the resulting solution was bubbled with N₂ for 10 min. 10% Pd / C (3 g, 28 mmol) was added to the solution and the reaction was purged with H₂ and kept under an atmosphere of H₂. After 4 h, an additional 10 mL of 4.0 M HCl solution in dioxane was added and the reaction was kept under an atmosphere of H₂ during the night. Upon completion (followed by LCMS), the reaction was filtered through a thin, well-packed celite bed and the collected solids were washed successively with MeOH and EtOAc. The combined organic filtrates were evaporated and dried under vacuum to give (IR, 2R) -2-aminocyclohexanol hydrochloride as a pale solid (13.8 g, 91 mmol, 93%). LCMS m / z 116.0 (MH⁺), Br = 0.37 min.

(IS, 2S) -2-Aminocyclohexanol hydrochloride was prepared in the same manner.

102010528

112

Reference Example 202

4-chloro-W-isobutoxypicolinamide

<img file="PT2010528T_D0083.tif" />

Added

O-isobutyl and N, W-hydroxylamine dihydrochloride (785 mg, 6.25 mmol, 1.1 eq) isopropylethylamine (2.97 ml, 17.0 mmol, 3.0 eq) to a suspension of hydrochloride hydrochloride. 4-chloropicolinyloyl (1.0 g, 5.68 mmol, 1.0 eq) in 25 mL of THF at room temperature. The reaction mixture was stirred for 30 min and diluted with water (25 mL) and EtOAc (50 mL). The separated organic layer was washed with brine (25 mL), saturated sodium bicarbonate solution (2 x 25 mL) and dried over sodium sulfate. Concentration under reduced pressure afforded the title compound as a colorless solid (870 mg, 67%) which was used without further purification. ES / MS m / z 229.0 (MH +), t R = 2.61 min.

Reference Example 203

4- (2- (cyclohexylmethylamino) benzo [d] thiazol-6-yloxy) -Nisobutoxypicolinamide

<img file="PT2010528T_D0084.tif" />

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113

4-Chloro-N-isobutoxypicolinamide (40 mg, 0.171 mmol) was added to the reaction mixture 2- (cyclohexylmethylamino) benzo [d] thiazol-6-ol (30 mg, 0.114 mmol; reference example 180-step 2) and carbonate of cesium (326 mg, 0.228 mmol) in 1.2 mL of DMF. The reaction mixture was stirred at room temperature for 10 min and then microwaved at 130 ° C for 3 x 20 min. The crude reaction mixture was filtered, purified by preparative HPLC and lyophilized to give the title compound as its TFA salt as a white solid (15 mg, 23%). ES / MS m / z 455.1 (MH⁺), t R = 2.90 min.

Example 204

4- (2 - ((IR, 2R) -2-hydroxy-cyclohexylamino) benzo [d] thiazol-6yloxy) -N-isobutoxypicolinamide

<img file="PT2010528T_D0085.tif" />

Prepared as in the previous example. ES / MS m / z 457.0 (MH⁺), t R = 2.35 min.

yloxy) benzo [d] thiazol-2-amine as TFA salt (3.0 mg). ES / MS m / z 434.2 (MH +).

The compounds in Table 3 were prepared according to the above examples and in particular according to the

102010528

114 example noted in the Ex Prep column (Preparation Example)

Table 3 [ ^Ex í [Comp [[Structure [[Prep [

Name (M + H) +, t R (min)

201

<img file="PT2010528T_D0086.tif" />

383, 1.96

171

H, C,

<img file="PT2010528T_D0087.tif" />

171

<img file="PT2010528T_D0088.tif" />

171

<img file="PT2010528T_D0089.tif" />

4- (2 - ((IR, 2R) -2 [hydroxycyclohexylamino) benzo [d]

7oxazol-6-yloxy, -N [methylpicolinamide 4- (2 - ((IR, 2R) -2- [hydroxycyclo- [jhexylamino) benzo [d] thiazole [413.1, 1.87

-6-yloxy) -N, N- [dimethylpicolinamide

N-cyclopropyl-4- (2 ((IR, 2R) -2-hydroxycyclohexylamino) benzo [d] thiazole

-6-yloxy) picolinamide

4- (2 - ((IR, 2R) -2hydroxycyclohexylamino) benzo [d] thiazole

-6-yloxy) -N- (tetrahydro5 2H-pyran-4-yl) picolinamide

425,2, 2,13

469,2, 2,09

H, 171.1 4- (2 - ((IR, 2R) -2 [482.2, 1.85

<img file="PT2010528T_D0090.tif" />

hexylamino) benzo [d] thiazole

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115 [ 171

<img file="PT2010528T_D0091.tif" />

S 171

<img file="PT2010528T_D0092.tif" />

I 171 lo.

<img file="PT2010528T_D0093.tif" />

ST

171

<img file="PT2010528T_D0094.tif" />

] -6-yloxy) -N- (1] methylpiperidin-4yl) picolinamide

4- (2- ((IR, 2R) -2hydroxycyclohexylamino) benzo [d] thiazole

-6-yloxy) -N- (1-methylpiperidin-3yl) picolinamide

N- (2-acetamidoethyl) -4 (2 - ((IR, 2R) -2hydroxycyclohexylamino) benzo [d] thiazol-6-yloxy) picolinamide

4- (2 - ((IR, 2R) -2] hydroxycyclohexylamino) benzo [d] thiazole

-6-yloxy) -N- (2 [(pyrrolidin-1] yl) ethyl) picolinamide

4- (2 - ((IR, 2R) -2] hydroxycyclohexylamino) benzo [d] thiazole

-6-yloxy) -N - ((tetra [hydrofuran-2 (yl) methyl) picolinamide

4- (2- ((IR, 2R) -2f hydroxycyclo482.2, 1.87

470,2, 1,86

482,2, 1,84

469,2, 2,16

171

483,2, 2,12

102010528

116

171

171

171

171 ο

<img file="PT2010528T_D0095.tif" />

<img file="PT2010528T_D0096.tif" />

<img file="PT2010528T_D0097.tif" />

hexylamino) benzo [d] thiazole

-6-yloxy) -N - ((tetrahydro-2H-pyran-4yl) methyl) picolinamide

4- (2 - ((IR, 2R) -2hydroxycyclohexylamino) benzo [d] thiazole

-6-yloxy) -N - ((1 [methylpiperidin-4 [yl) methyl) picolinamide [N- (((S) -1-ethylpyrrolidin-2-yl) methyl) -4- (2 ((IR, 2R) -2-hydroxycyclohexylamino) benzo [d] thiazol] -6-yloxy) picolinamide

N-ethyl-4- (2 - ((1R, 2R) -2hydroxycyclohexylamino) benzo [d] thiazole

-6-yloxy) picolinamide

4- (2 - ((IR, 2R) -2 - [[hydroxycyclo- [θ hexylamino) benzo [d] thiazole [

-6-yloxy) -N- (2,2,2 - [; trifluoroethyl) picolinamid [

I a

496,2, 1,84

496.2, 1.88] H, C Ν

<img file="PT2010528T_D0098.tif" />

FI μ

<img file="PT2010528T_D0099.tif" />

413,2, 2,09

467,1, 2,37

S KC '

S 4- (2- ((IR, 2R) -2O 'hydroxycyclo [427.1, 2.25

<img file="PT2010528T_D0100.tif" />

I 171 hexylamino) benzo [d] thiazole

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117 ί 171

<img file="PT2010528T_D0101.tif" />

ϊ 171

<img file="PT2010528T_D0102.tif" />

156 175

125 [ 162

<img file="PT2010528T_D0103.tif" />

155 175

<img file="PT2010528T_D0104.tif" />

<img file="PT2010528T_D0105.tif" />

] -6-yloxy) -Ν] propylpicolinamide] Ν- (cyclopropylmethyl) -4- [ ^ξ (2 - ((IR, 2R) -2'hydroxycyclo-439.2, 2.32 hexylamino) benzo [d] thiazole §] -6-yloxy) picolinamide ^ xxxxxxxxxxxxxxxxxxxxxxxxxxxxxxxxxxxxxxxxxxxxxxxxxx -........... ........

4- (2 - ((IR, 2R) -2- [ ⁵ hydroxycyclo- [Î ± (hexylamino) benzo [d] thiazol] 427.2, 2.24

-6-yloxy) -N-] isopropylpicolinamide

4- (2- ((1r, 4r) -4hydroxycyclo] hexylamino) benzo [d] thiazol] 399.1, 1.83 -6-yloxy) -N-] methylpicolinolinamide (IR, 2R) -2- (6 - (2- (1-methylH-imidazol-2-yl) pyridin4-yloxy) benzo [d] thiazol-2ylamino) cyclohexanol (IR, 2R) -2- (6- (2- (pyrazin2-yl) pyridin -4yloxy) benzo [d] thiazole-2_s ylamino) cyclohexanol

422,1, 1,83

420,1, 1,91

<img file="PT2010528T_D0106.tif" />

| (R5-yl) pyridin-4^! (IR, 2R) -2- (6- (2- (thiazol157.175

425.0, 2.05 yloxy) benzo [d] thiazol-2ΡΕ2010528

118

161 § 188

162 § 177

164 § 174

158 § 175

<img file="PT2010528T_D0107.tif" />

159 i 175

Z = N

<img file="PT2010528T_D0108.tif" />

160 § 175

<img file="PT2010528T_D0109.tif" />

I hn

<img file="PT2010528T_D0110.tif" />

H, H, C

<img file="PT2010528T_D0111.tif" />

H, C.

<img file="PT2010528T_D0112.tif" />

ylamino) cyclohexanol (IR, 2R) -2- (6- (2- (1-methylH-imidazol-5-yl) pyridin4-yloxy) benzo [d] thiazol-2ylamino) cyclohexanol (IR, 2R) -2- (6- (2- (thiazol4-yl) pyridin-4-yloxy) benzo [d] thiazol-2ylamino) cyclohexanol (IR, 2R) -2- (6- (2- (thiazol2-yl) pyridin-4-yloxy) benzo [d] thiazol-2ylamino) cyclohexanol (IR, 2R) -2- (6- (2- (1,2,3,6tetrahydropyridin-4yl) pyridin-4yloxy) benzo [d] thiazol-2ylamino) cyclohexanol (IR, 2R) -2- (6- (2- (5-ethyl4-methyl-1H-imidazol-2-yl) pyridin-4yloxy) benzo [d] thiazol-2ylamino) cyclohexanol (IR, 2R) -2- (6- (6 '- ( 4methylpiperazin-1-yl)₀ 2,3'-bipyridin-4yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol

422,1, 1,84

425,0, 1,85

425,0, 2,28

423,1, 1,38

450,1, 1,82

517,2, 1,75

102010528

119

165 i 174

167 ]174

169 174

170 i174

166 § 174

<img file="PT2010528T_D0113.tif" />

(IR, 2R) -2- (6- (6'morpholin-2,3'-bipyridin-1'4-yloxy) benzo [d] thiazol-2ylamino) cyclohexanol (IR, 2R) -2- (6- ( 2- (3 (morpholinomethyl) phenyl) pyrimidin-4-yloxy) benzo

N&gt;

[d] thiazol-2-ylamino)] cyclohexanol (1R, 2R) -2- (6- (2-cyclohexenylpyridin-4yloxy) benzo [d] thiazol-2ylamino) cyclohexanol (IR, 2R) -2- (6- (2- (4 (morpholinomethyl) phenyl) pyr

<img file="PT2010528T_D0114.tif" />

168 )174

<img file="PT2010528T_D0115.tif" />

<img file="PT2010528T_D0116.tif" />

idin-4-yloxy) benzo [d] thiazol-2-ylamino)] cyclohexanol (IR, 2R) -2- (6- (2-cyclopropylpyridin-4yloxy) benzo [d] thiazol-2ylamino) cyclohexanol (IR, 2R) -2- (6- (6'-methoxy2,3'-bipyridin-4 ' ¹ yloxy) benzo [d] thiazol-25-ylamino) cyclohexanol (IR, 2R) -2- (6- (2'-fluoro [2,4'-bipyridin-4

<img file="PT2010528T_D0117.tif" />

504,1, 1,93

517,1, 1,78

422,1, 2,07

517,2, 1,76

382,1, 1,84

449,1, 1,98

171 § 174

437,1, 2,22

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120

<img file="PT2010528T_D0118.tif" />

yloxy) benzo [d] thiazol-2ylamino) cyclohexanol

172 I174

<img file="PT2010528T_D0119.tif" />

173 1 174

<img file="PT2010528T_D0120.tif" />

437,1, 2,04

174 I183

<img file="PT2010528T_D0121.tif" />

175 1183

<img file="PT2010528T_D0122.tif" />

176 I183

<img file="PT2010528T_D0123.tif" />

[(IR, 2R) -2- (6- (3'-fluoro- [ ^! 2'-morpholin-2,4'- [Â''bipyridin-4- [522.2, 2.17 yloxy) benzo [d] thiazol-2 - [[ylamino) cyclohexanol (IR, 2R) -2- (6- (6'-fluoro2,3'-bipyridin-4-yloxy) benzo [d] thiazol-2_s ylamino) cyclohexanol (1R, 2R) -2- (6- (2- (piperidin-1-yl) pyridin4-yloxy) benzo [d] thiazol-2ylamino) cyclohexanol (1R, 2R) -2- (6 - (2morpholinopyridin-4yloxy) benzo [d] thiazole-2₍ ylamino) cyclohexanol (IR, 2R) -2- (6- (2- (4- [methylpiperazin-1- [yl) pyridin-4- [440.1, 1.66 yloxy) benzo [d] thiazole 2- ([ylamino) cyclohexanol

N - ((R) -1- (4- (2 - ((IR, 2R) 2-hydroxycyclo

425,1, 1,98

427.1, 1.80 o

H, C

Tl η) h hexylamino) benzo [d] thiazole I

M ✓JU „!

-6-yloxy) pyriclin-2-yl ·) [468.1, 1.78

177 S 183

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121

178 I 183

179 j 183

180 i 175

181 I175

HC (pyrrolidin-3-yl)

N - ((S) -1- (4- (2 - ((IR, 2R) - 2-hydroxycyclohexylamino) benzo [d] thiazole |

'] (p-6-yloxy) pyridin-2-yl) pyrrolidin-3-yl) acetamide

4- (4- (2 - ((IR, 2R) -2hydroxycyclo | 468.1, 1.77

ΙηΛ

I hexylamino) benzo [d] thiazole 440, 1.70

<img file="PT2010528T_D0124.tif" />

419,0, 1,87

<img file="PT2010528T_D0125.tif" />

182 i 174

<img file="PT2010528T_D0126.tif" />

-6-yloxy) pyridin-2-yl)

Piperazin-2-one (IR, 2R) -2- (6- (2,2'-bipyridin-4-yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol (IR, 2R) -2- (6- (2- (1-Methyl 3- (trifluoromethyl) -1H-pyrazol-S-ylpyridin-liloxy) benzo [d] thiazol-2ylamino) cyclohexanol (IR, 2R) -2- (6- (2- (1 - (2morpholinoethyl) -1Hpyrazol-4-yl) pyridin-4490.1, 2.60

521,1, 1,71

(V-yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol

184 I 192 (IR, 2R) -2- (6- (6,7) 452.1, 2.00

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122

ΤΑ] HgC-O ^ Á. dimethoxyquinolin-4I-yl (N-tyloxy) benzo [d] thiazol-2-ylamino) cyclohexanol

185 I176

187 )176

188 )179

189 I179 ((IR, 2R) -2- (6- (2- (1-ethylH-pyrazol-4-yl) pyridinyl) [14-yloxy) benzo [d] thiazol-2ylamino) cyclohexanol (IR, 2R) -2- (6- (2- (1- (2 (diethylamino) ethyl) -1Hpyrazol-4-yl) pyridin-4-yloxy) benzo [d] thiazol-2Sylamino) cyclohexanol (IR, 2R) -2- (6- (2- (1- (2,2-difluoroethyl) -1H-pyrazol-4-yl) pyridin-4yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol (IR, 2R) -2- (6- (3- (1H-pyrazol-4-yl) pyridin-4-yloxy) benzo [d] thiazol-20-N'-ylamino) cyclohexanol (IR, 2R) -2- (6- (3- ( 1-methylH-pyrazol-4-yl) pyridin-2-one

<img file="PT2010528T_D0127.tif" />

436,0, 1,89

186 S 176

<img file="PT2010528T_D0128.tif" />

507,1, 1,75

<img file="PT2010528T_D0129.tif" />

472,0, 1,90

<img file="PT2010528T_D0130.tif" />

H, C | (ΐ ^ ΓθΙι ^ Τ '³), 4-yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol (IR, 2R) -2- (6- (3,3 '[bipyridin-4-yloxy) benzoTdltiazol-2408,1, 1, 70

422,1, 1,79

190 ]179

419,1, 1,66

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123

191

179

192

179

193 179

194

179

<img file="PT2010528T_D0131.tif" />

] (IR, 2R) -2- (6- (3- (1-methyl₃ç

<img file="PT2010528T_D0132.tif" />

1H-pyrazol-5-yl) pyridins

-yloxy) benzo [d] thiazol-2N-ylamino) cyclohexanol

<img file="PT2010528T_D0133.tif" />

(IR, 2R) -2- (6- (3,4'bipyridin-4yloxy) benzo [d] thiazol-2ylamino) cyclohexanol

<img file="PT2010528T_D0134.tif" />

(IR, 2R) -2- (6- (6'-amino3,3'-bipyridin-4yloxy) benzo [d] thiazol-2ylamino) cyclohexanol

422,1, 1,83

419,1,

1,61

434,1, 1,60

<img file="PT2010528T_D0135.tif" />

(IR, 2R) -2- (6- (6 '- (4methylpiperazin-1-yl) 3,3'-bipyridin-4yloxy) benzo [d] thiazol-2ylamino) cyclohexanol

517,2

1,64

<img file="PT2010528T_D0136.tif" />

382,1

505, 1

1,97

2,03

102010528

124

197

174

200

177

<img file="PT2010528T_D0137.tif" />

IH, C,

<img file="PT2010528T_D0138.tif" />

SF

<img file="PT2010528T_D0139.tif" />

yloxy) benzo [d] thiazol-2ylamino) cyclohexanol (IR, 2R) -2- (6- (2- (4- (4methylpiperazin-1-] _çyl) phenyl) pyridin-4-] 516.2, 1.85 yloxy) benzo [d] thiazol-2-] ylamino) cyclohexanol (IR, 2R) -2- (6- (2- (4-) (trifluoromethyl) -1H-] imidazol-2-yl) pyridin-4-] 476.1, 2.35 yloxy) benzo [d] thiazol-2-]) ylamino) cyclohexanol I

<img file="PT2010528T_D0140.tif" />

] (IR, 2R) -2- (6- (2-methylpyridin-4-]] 356.1, 1.73 yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol]

209

192 | (IR, 2R) -2- (6- (3F g fluoropyridin-4Nyloxy) benzo [d] thiazol-2ylamino) cyclohexanol

<img file="PT2010528T_D0141.tif" />

360.1, 1.83] (IR, 2R) -2- (6- (3224

178

<img file="PT2010528T_D0142.tif" />

Br g bromopyridin-4Nyloxy) benzo [d] thiazol-2420.1 / 42

2,0, 1,81

225

173 ylamino) cyclohexanol

<img file="PT2010528T_D0143.tif" />

(ylamino) cyclohexanol

376,1, 2,13

102010528

125

258

177

259 [176

260 176

226 I173

<img file="PT2010528T_D0144.tif" />

231 [177

IH₃Ç

<img file="PT2010528T_D0145.tif" />

232 I177

H, C

<img file="PT2010528T_D0146.tif" />

hexylamino) benzo [d] thiazole

-6-yloxy] picolinonitrile (IR, 2R) -2- (6- (2- (4-methylH-imidazol-2-yl) pyridin4-yloxy) benzo [d] thiazol-2ylamino) cyclohexanol (IR, 2R ) -2- (6- (2- (4,5-dimethyl-1H-imidazol-2yl) pyridin-4yloxy) benzo [d] thiazol-2 [ylamino) cyclohexanol

<img file="PT2010528T_D0147.tif" />

367,1, 2,03

422,1, 1,

436,1, 1,93

<img file="PT2010528T_D0148.tif" />

H₃(IR, 2R) -2- (6- (2- (4,5,6,7] tetrahydro-β-] benzo [d] imidazol-2-] δ 462.2, 2.03 yl) pyridin-4-] _s yloxy) benzo [d] thiazol-2-]] ylamino) cyclohexanol] (IR, 2R) -2- (6- (2- (1- (2-fluoroethyl) -1H-pyrazol-4] yl) pyridin-2-one 4-] 454.2, 1.89 yloxy) benzo [d] thiazol-2-] [ylamino) cyclohexanol [] (IR, 2R) -2- (6- (2- (1- (2-methoxyethyl) - 1H-pyrazol-4yl) pyridin-4466.2, 1.89

<img file="PT2010528T_D0149.tif" />

iloxy) benzo [d] thiazol-2ΡΕ2010528

126

262 i 187

265 § 174

266 § 177

267 [177

280 § 174

261 § 174

<img file="PT2010528T_D0150.tif" />

<img file="PT2010528T_D0151.tif" />

H, C

<img file="PT2010528T_D0152.tif" />

H C

<img file="PT2010528T_D0153.tif" />

IN

<img file="PT2010528T_D0154.tif" />

[ylamino) cyclohexanol [(IR, 2R) -2- (6- (6'-amino2,3'-bipyridin-4yloxy) benzo [d] thiazol-25 ylamino) cyclohexanol j (IR, 2R) - 2- (6- (2- (1Himidazol-1-yl) pyridin-4X4-yloxy) benzo [d] thiazol-2_s ylamino) cyclohexanol (IR, 2R) -2- (6- (2- (1-methylH-pyrazol-5-yl) pyridin4-yloxy) benzo [d] thiazol-2ylamino) cyclohexanol (IR, 2R) -2- (6- (2- (4,5-diethyl-1H-imidazol-2yl) pyridin-4yloxy) benzo [d] thiazol-2ylamino) cyclohexanol (IR, 2R) -2- (6- (2 - (5-methyl4-propyl-1H-imidazol-2yl) pyridin-4yloxy) benzo [d] thiazol-2ylamino) cyclohexanol (IR, 2R) -2- (6- (2,4'bipyridin-4yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol

434,2, 1,78

408,2, 1,85

422,1, 1,93

464,2, 2,10

464,2, 2,10

419,0, 1,68

281 1774 (IR, 2R) -2- (6- (2- (1450.0, 2.02

102010528

127

285

174

SH, C

I?

<img file="PT2010528T_D0155.tif" />

282 |174

<img file="PT2010528T_D0156.tif" />

283 ϊ 174

IH C

<img file="PT2010528T_D0157.tif" />

284 |174

<img file="PT2010528T_D0158.tif" />

propyl-1H-pyrazol-4, yl) pyridin-4yloxy) benzo [d] thiazol-2ylamino) cyclohexanol] (IR, 2R) -2- (6- (6'-amino^! 5 '- (trifluoromethyl) -2,3' '(bipyridin-4-502.0, 2.02 yloxy) benzo [d] thiazol-2] ylamino) cyclohexanol (IR, 2R) -2- (6 - (2- (1,3,5-] trimethyl-1H-pyrazol-4-] yl) pyridin-4-] 450.0, 1.86 yloxy) benzo [d] thiazol-2-] ylamino) cyclohexane hexanol (IR, 2R) -2- (6- (2- (1H] pyrrolo [2,3-b] pyridin-5-yl) pyridin-4-] 458.0, 1.91) H 5 * yloxy ) benzo [d] thiazole-2-]

Ylamino) cyclohexanol]

<img file="PT2010528T_D0159.tif" />

] (IR, 2R) -2- (6- (2-] (pyrimidin-5-yl) pyridin] -420.1, 1.90

4-yloxy) benzo [d] thiazol-2-] ylamino) cyclohexanol |

<img file="PT2010528T_D0160.tif" />

(IR, 2R) -2- (6- (2- (2-])₃ (dimethylamino) pyrimidin-]

5-yl) pyridin-4-] 463.1, 1.96 yloxy) benzo [d] thiazol-2-]

Iylamino) cyclohexanol I

102010528

128

287 I174

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288

174

<img file="PT2010528T_D0162.tif" />

291 1195

<img file="PT2010528T_D0163.tif" />

] 4- (2 - ((IR, 2R) -2-]] hydroxycyclo] hexylamino) benzo [d] thiazol] x, 444.0, 2.18 l [-6-yloxy) -2,3 '- ] bipyridine-6'-]] carbonitrile] (IR, 2R) -2- (6- (2- (4-fluorophenyl) pyridin-4-yloxy) benzo [d] thiazol-2ylamino) cyclohexanol

4- (2 - ((IR, 2R) -2-] hydroxycyclo-] hexylamino) benzo [d] thiazol] 399.1, 1.72 -6-yloxy) -N-] methylnicotinamide]

4- (2- (3-hydroxycyclo436.0, 2.06

293 1162

<img file="PT2010528T_D0164.tif" />

hexylamino) benzo [d] thiazol] [399.1, 1.76

-6-yloxy) -N] methylpicolinamide

4- (2- ((IS, 2S) -2hydroxycyclopentylamino) b]

320 1 162

<img file="PT2010528T_D0165.tif" />

322 I 162

Hn

<img file="PT2010528T_D0166.tif" />

] 385.1, 1.74 enzo [d] thiazol-6-yloxy) -N-] methylpicolinamide

4- (2- ((IS, 2R) -2hydroxycyclopentylamino) b]

385.1, 1.75 enzo [d] thiazol-6-yloxy) -N-] methylpicolinolinamide [4- (2 - ((IR, 2R) -2-] 385.1, 1.75

324 I 162

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129

344 § 174

345 S 174

346 § 174

360 § 174

I ΗΝ

<img file="PT2010528T_D0167.tif" />

I HC-N

<img file="PT2010528T_D0168.tif" />

| Ν

<img file="PT2010528T_D0169.tif" />

<img file="PT2010528T_D0170.tif" />

359 I174

<img file="PT2010528T_D0171.tif" />

ΗΝ

<img file="PT2010528T_D0172.tif" />

<img file="PT2010528T_D0173.tif" />

q. (hydroxycyclopentylamino) b'-enzo [d] thiazol-6-yloxy) -methylpicolinolinamide (IR, 2R) -2- (6- (2- (1-methylH-pyrazol-4-yl) pyridin4-yloxy) benzo [d] thiazole-2_s ylamino) cyclohexanol (IR, 2R) -2- (6- (2,3'bipyridin-4yloxy) benzo [d] thiazol-2ylamino) cyclohexanol (IR, 2R) -2- (6- (2- (2-aminopyrimidin-5-yl) pyridin-4yloxy) benzo [d] thiazol-2] ylamino) cyclohexanol (IR, 2R) -2- (6- (2- (1isobutyl-1H-pyrazol-4CRyl) pyridin-4yloxy ) benzo [d] thiazol-2-ylamino) cyclohexanol (1R, 2R) -2- (6- (2- (1H-pyrazol-4-yl) pyridin-4-yloxy) benzo [d] thiazol-2_s ylamino) cyclohexanol ^§ (IR, 2R) -2- (6- (2 (methylamino) pyridin-4422.2, 1.68

419,1, 1,69

435,2, 1,61

464,4, 1,95

408,1, 1,63 ] 371,1, 1,79

368 ί 170 yloxy) benzo [d] thiazol-2ΡΕ2010528

130

369 170

<img file="PT2010528T_D0174.tif" />

371 i 175

<img file="PT2010528T_D0175.tif" />

372 [ 175

<img file="PT2010528T_D0176.tif" />

] ylamino) cyclohexanol (IR, 2R) -2- (6- (2 (ethylamino) pyridin-4yloxy) benzo [d] thiazol-2'-ylamino) cyclohexanol (IR, 2R) -2- (6- (2- (Furan-2yl) pyridin-4yloxy) benzo [d] thiazol-2ylamino) cyclohexanol (IR, 2R) -2- (6- (2- (oxazol2-yl) pyridin-4yloxy) benzo [d] thiazol-2-ylamino) cyclohexanol] 4- (7-bromo-2 - ((IR, 2R) [2-hydroxycyclo385.1, 1.74

408,1, 1,94

409,1, 1,98

376 195

377 [196

<img file="PT2010528T_D0177.tif" />

378 ί 197

<img file="PT2010528T_D0178.tif" />

HN 2 O 4 (3 O. ^ x ^ s (hexylamino) benzo [d] thiazole 479, 2.37 CH₃ (Χ-6-yloxy) -N-] methylmethylcholinamide

4- (2 - ((IR, 2R) -2] hydroxycyclohexylamino) -]

ÇH |

7-methylbenzo [d] thiazol-6-] 413, 2.09 yloxy) -N-] [methylpicolinamide-4- (7-chloro-2 - ((IR, 2R) - [§ 2-hydroxycyclo-]

Cl | hexylamino) benzo [d] thiazol] 433, 2,33 -6-yloxy) -N - [[methylpicolinamide

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4- (2 - ((IR, 2R) -2hydroxycyclo381

<img file="PT2010528T_D0179.tif" />

(Hexylamino) benzo [d] thiazole

457,0, 2,35

-6-yloxy) -Nisobutoxypicolinamide

Each of the compounds of the invention listed in

Table 2 has been shown to have CSF-1R inhibition activity with an ICso of less than about 10 μΜ. Many of the compounds exhibited activity with an ICso of less than about 1 μΜ, or less than about 0.1 μ ou, or less than about 0.01 μΜ relative to CSF-1R inhibition. The compounds of the invention in Tables 3 were found to have an activity of less than 1 μΜ. As such, each of the compounds of Tables 2 and 3 is preferred individually and as a member of a group.

In addition to CSF-1R inhibitory activity many of the compounds of Tables 2 and 3 were also screened for Raf inhibition (according to biochemical screenings described in US 10 / 405,945) as well as other kinases and shown to inhibit CSF-1R. significantly larger (about 2 to about 1000 times larger) than Raf and other screened kinases. More particularly, many of the screened compounds had activity greater than about 1 μΜ in relation to Raf inhibition, while many of the same compounds showed CSF-1R activity less than about 0.1 μΜ. As such, many of the compounds in Tables 2 and 3 are

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132 potent and selective CSF-1R inhibitors.

BIOLOGICAL EXAMPLES

Biological Example 1

In vitro Colony Stimulation Factor Receptor Kinase Assays (CSF-1R)

Kinase activity of various tyrosine kinases proteins can be measured by providing ATP and a suitable peptide or substrate containing the tyrosine protein, and by analyzing the transfer of the phosphate moiety to the tyrosine residue. The recombinant protein corresponding to the human CSF-1R cytoplasmic domain was purchased from Invitrogen Corporation, Carlsbad, CA USA (# PV3249). For each assay, test compounds were serially diluted starting at 25 μΜ with 3 dilutions in DMSO in 384-well plates, then mixed with an appropriate kinase reaction buffer consisting of 50 mM Hepes, 5 mM MgCl 2, 10 mM MnCl 2, 0.1% BSA, pH 7.5, 1.0 mM dithiothreitol, 0.01% Tween 80 plus 1 μΜ ATP. Protein kinase and an appropriate 50 nM biotinylated peptide substrate were added to give a final volume of 20 pL, reactions were incubated for 2 hours at room temperature and stopped by the addition of 10 pL 45 mM EDTA, 50 mM Hepes, pH 7.5. 30 µl of PThas Alphascreen beads (Perkin Elmer, Boston, MA, USA) were added to the mixture with the reaction stopped. The reaction was incubated overnight and read on the apparatus.

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133

Envision (Perkin Elmer). Phosphorylated peptide product was measured with the AlphaScreen system (Perkin Elmer) using PT66 antiphosphotyrosine antibody coated acceptor beads and streptavidin coated donor beads that emit a fluorescent signal at the emission wavelength of 520-620 nM if they are in close proximity. The concentration of each compound for 50% inhibition (ICso) was calculated by nonlinear regression using XL Fit data analysis software.

CSF-1R kinase was analyzed in 50 mM Hepes pH 7.0, MgCl₂ 5 mM, MnCl₂ 10 mM, 1 mM DTT, 1 mg / mL BSA, 1.0 μΜ ATP and biotin-GGGGRPRAATF-NH peptide substrate₂ 0.05 μΜ (SEQ ID NO: 2). CSF-1R kinase was added at a final concentration of 4 nM.

Biological Example 2

In vitro inhibition of receptor tyrosine phosphorylation

CSF-1R

To test for inhibition of CSF-1R receptor tyrosine phosphorylation, HEK293H purchased from Invitrogen Cat. # 11631017 cells transfected with the fully cloned human CSF-1R receptor internally cloned into a mammalian episomal transfection vector, were incubated for 1h with dilutions in series of compounds starting at 10 μΜ

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134 at 3-fold dilutions and then stimulated for 8 minutes with 50 ng / mL MCSF. After the supernatant was removed, cells were lysed on ice with lysis buffer (150 mM NaCl, 20 mM Tris, pH 7.5, 1 mM EDTA, 1 mM EGTA, 1% Triton X100 and NaF, protease inhibitors and phosphatase) and then stirred for 15-20 min at 4 ° C. The lysate was then transferred to total CSF-1R antibody coated 96-well plates that had already been blocked with 3% Mesoscale discovery Blocker A (MSD) for 2 hours and then washed. Used were incubated overnight at 4 ° C and the plates were then washed 4x with MSD Tris Wash Buffer. The anti-pTyr SULFO-TAG MSD antibody was diluted to final 20nM in 1% Blocker A (MSD) solution and added to the washed plates and incubated for 1.5-2 h before addition of the reading buffer (MSD). . Plates were read on the Sector 6000 (MSD) instrument. The raw data were imported into Abase and ECsos were calculated with XL-fit data analysis software.

Biological Example 3

CSF-1R Inhibitors on MNFS-60 Pk / Pd Model

Five million MNFS-60 cells were implanted in a subcutaneous HBSS / matrigel solution on the right flank. Approximately 3 weeks after tumor cell injection, tumors were measured and selected mice were randomized (n = 3, except for vehicle group, where n = 6) into size-based groups.

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135 of the tumor.

Compounds that inhibited M-CSF-mediated proliferation in MNFS-60 cells and CSF-1R phosphorylation with CE50S &lt;100 nM were tested in the MNFS-60 synergic tumor model (5 X 10⁶ with subcutaneous implantation in matrigel and growth for 3-4 weeks until approximately 150 mm²). A single 100 mg / kg dose of representative compounds disclosed herein was administered to animals with MNFS-60 tumors; Plasma and tumor samples were taken at various time points after dosing, starting at 1 h to 24 h.

Several of the compounds described herein have been found to inhibit CSF-1R Tyr723 phosphorylation in&gt; 50% tumor lysates compared to vehicle control 4 h after administration as determined by

Western blot.

Additionally, several of the compounds described herein were tested in a rapid onset severe arthritis mouse model (Terato, K. et al., Journal of Immunology 148: 2103-2108; 1992) and treatment began on the third day after injection of the anti-collagen antibody cocktail followed by LPS stimulation. Over the 12 days of treatment with CSF-1R inhibitors, the extent of foot swelling and severity of bone resorption were classified. Significant attenuation of swelling was not observed in the treated group compared to the control group; at the

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136 However, there was a tendency to improve the severity of bone resorption. There are no reports to date that CSF-1R inhibitors are effective in this arthritis model.

The only successful reduction in disease progression has been reported for inhibition of CSF-1R signaling with an anti-MCSF antibody in a slower onset, less severe arthritis mouse model (Campbell et al J. Leukoc. Biol. 68: 144-150; 2000).

Biological Example 4

Raf Kinase Signaling Inhibition in a Biochemical Assay

In Vi

The inhibitory effect of the compounds on Raf was determined using the following biotinylated assay. Raf kinase activity was measured by providing ATP, a recombinant kinase inactive MEK substrate, and analysis of the transfer of the phosphate moiety to the MEK residue. Recombinant full-length MEK with an inactivating K97R mutation of the ATP binding site (rendering the kinase inactive) was expressed in E. coli and labeled with post biotin purification. MEK cDNA was subcloned with an N-terminal (His) tag 6 and expressed in E. coli and the recombinant MEK substrate was purified from E. coli lysate by niguel affinity chromatography followed by anion exchange. The final MEK substrate preparation was biotinylated (Pierce EZ-Link Sulfo-NHS-LC-Biotin) and concentrated to 11.25 μΜ. Recombinant Raf (including cΡΕ2010528

137

Raf and mutant B-Raf isoforms) was obtained by purification from sf9 insect cells infected with the corresponding human recombinant Raf expression vectors.

Recombinant Raf isoforms were purified by a Glu antibody interaction or by metal ion chromatography.

For each assay, the compound was serially diluted starting at 25 μΜ with 3-fold dilutions in DMSO and then mixed with various Raf isoforms (0.50 nM each). Kinase-inactive biotin-MEK substrate (50 nM) was added in the reaction buffer plus AIP (1 μ). The reaction buffer contained 30 mM Tris-HCl pH 7.5, 10 mM MgCl2, 2 mM Dll, 4 mM EDTA, 25 mM beta-glycerophosphate, 5 mM MnCl2, and 0.01% BSA / PBS. The reactions were subsequently incubated for 2 hours at room temperature and quenched by the addition of 0.5 Μ EDTA. The quenched reaction mixture was transferred to a neutradavine coated plate (Pierce) and incubated for 1 hour. Phosphorylated product was measured with the time resolved fluorescence system DELFIA (Wallac) using a rabbit anti-p-MEK (Cell Signaling) as the primary antibody and Europium-labeled anti-rabbit as the secondary antibody. Time resolved fluorescence can be read on a Wallac 1232 DELFIA fluorometer. Compound concentration for 50% inhibition (IC 50) was calculated by nonlinear regression using XL Fit data analysis software.

Biological Example 5

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Inhibition of cKIT and PDGFRb Kinase Signaling in an In Vitro Biochemical Assay

IC50 values ​​for inhibition of RTKs were determined in alphascreen format by measuring compound inhibition of phosphate transfer to a substrate by the respective enzyme. Briefly, the respective RTK domain acquired as human recombinant protein (CKIT Upstate # 14-559, PDGFRb Invitrogen # P3082) was incubated with serial dilutions of the compound in the presence of substrate and ATP concentrations within 3 times the Km of the enzyme.

cKIT kinase domain was analyzed in 50 mM Hepes, pH = 7.5, MgCl₂ 5 mM, MnCl₂ 10 mM, 1 mM DTT, 0.1% BSA with 0.06 æM biotinylated peptide substrate (GGLFDDPSYVNVQNL-NH2) and 15 æM ATP (apparent ATP KM = 15 æM). PDGFR3 kinase domain was analyzed in 50 mM Hepes, pH = 7.5, 20 mM MgCl₂, 1 mM DTT, 0.1% BSA with 0.1 µM biotinylated peptide substrate (GGLFDDPSYVNVQNL-NH2) and 10 µM ATP (apparent ATP KM = 25 µM). Reactions were incubated at room temperature for 3 to 4h and quenched with buffer (20 mM EDTA, 0.01% Tween-20 for PDGFRb and cKIT). Alphascreen PY20 beads were added to the cKIT Stopped Reactions and P320 Ab / Protein A beads were added to the PDGFRP Stopped Reactions. Both reactions were incubated overnight and read in the Alphascreen reader. Compound concentration for 50% inhibition (ICso) was calculated using unregulated regression.

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139 using the XL-Fit data analysis software. As a control comparou844, staurosporine is read in all assays and a Z '&gt; 0.5 is required to validate the results.

Biological Example 6

Cell Viability Assay in MCSF-Dependent MNFS60 Cells

Cell viability was assessed by Cell Titer Glo, Promega. MNFS60 (murine AML cells) were seeded into TC-treated 96-well plates at a density of 5000 cells per well in RPMI-1640, 10% FBS and 1% Penicillin Streptomycin prior to addition of compound. Test compounds were serially diluted (3-fold) in DMSO to 500x final concentration. For each concentration of test compound, 2 µl (500x) aliquots of compound or 100% DMSO (control) were diluted in 500 µl of culture medium containing 2x the final concentration of MCSF growth factor by 2x the concentration and then dilute 1x in the cells. The final concentration of MCSF is 10 ng / mL. Cells were incubated for 72 hours at 37 ° C, 5% CO 2. After incubation, 100 µl Cell Titer Glo is added to each well to determine viable cells. The assay was performed according to the manufacturer's instructions (Promega Corporation, Madison, WI, USA). Each experimental condition was performed in triplicate. Raw data were imported into Abase and EC50S calculated with XLfit data analysis software. The relative light units of wells that contained

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140 cells without MCSF in the medium and as a result did not grow were defined as 100% inhibited.

Biological Example 7

Tumor Induced Osteolysis Model

Tumor-induced osteolysis (TIO) models have revealed the recapitulation of the gross bone destruction observed in cancer patients with osteolytic tumor metastasis and have been widely reported in both the bisphosphonate literature and in conjunction with testing for new anti-osteolytic agents. The results of these studies correlate well with human clinical activity (Kim SJ et al., 2005, Canc. Res., 65 (9): 3707; Corey, E et al., 2003, Clin. Canc. Res., 9: 295; Alvarez, E. et al. , 2003,

Clin. Cancel Res., 9: 5705). The procedure includes injection of tumor cells directly into the proximal tibia. Once cells are established, they proliferate and secrete factors that potentiate osteoclast activity, resulting in trabecular and cortical bone resorption. Animals are treated with anti-desorptive agents after tumor cell implantation and bone destruction is measured in various ways at the end of the study.

The tumor cell lines used in this protocol are of human origin and represent tumor lines that have been previously modified to now express the Luciferase enzyme to screen tumor cells in the animal.

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141 using the Xenogen system. The strength of the light signal also gives an indication of approximately how many tumor cells are located at a given site.

Rats are injected subcutaneously with 2.5 mg / kg flunixin meglumine 30 minutes prior to cell inoculation to provide post-procedural analgesia. Mice are then anesthetized by isoflurane inhalation (ketamine / xylazine injection may be used if isoflurane is not available). Anesthetized animals are placed in the supine position and after aspiration of the tumor cells in a 50 or 100 µL micro-syringe equipped with a 26 or 27 gauge needle, the needle will be inserted through the cortex of the right tibial anterior tuberosity with a rotary movement of the drill to minimize the chance of cortical fracture. Successful passage of the needle through the cortex and into the medulla is indicated by loss of resistance against forward movement of the needle. Once the bone cortex is traversed, it will be injected into the tibia bone marrow 10-20 pL of cell suspension (6Χ10^Λ5th MDA-MB-231 Luc or 3Χ10 breast carcinoma^Λ5th PC-3MLuc prostate carcinoma cells). Animals will be observed to ensure uneventful recovery (heating pad or lamp) until they recover from anesthesia.

The progression of tumor growth in bone can be divided into five stages (Steps 0-4). The stages are defined as follows and can be monitored at

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142 Comparison with uninjected mouse leg (left): Stage 0: Normal, no sign of any changes in bone.

Stage 1: Minimal or equivocal injury; cortex / normal architecture.

Stage 2: Definitive Injury; minimal cortex / architecture disorder.

Stage 3: Large Injury; cortex disorder / architecture. Stage 4: Severe Destruction; no preservation of architecture, late stage. Animals that reach this stage will be removed from the study and euthanized.

Photon imaging of the legs is used to assess tumor growth at injection sites and remote sites during the study using the Xenogen system to quantify tibial tumor cells and confirm the absence of effusion in other areas. Radiographs of the legs are performed up to once a week until the end of the study using a Faxitron X-ray unit to assess the destruction of cortical bone at the injection site. While using more invasive cell lines such as PC-3M-Luc, bone damage is monitored one to two weeks after injection and weekly thereafter. For cell lines that form lesions at a slower rate, such as MDA-MB-231Luc, which show no bone damage until 4 to 5 weeks post-implantation, the first radiographic images are taken approximately 4 weeks after intratibial implantation. animals with cells to establish control baseline and then once

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143 per week to measure bone injury starting at a time when the lesions begin to develop based on pilot model development studies. For example, in mice injected with MDA-MB-231Luc, an image would be taken approximately 4 weeks after implantation, with weekly images thereafter.

Animals may be dosed with small molecules, monoclonal antibodies or proteins once or twice a day by conventional routes.

The end point of this study is the time when most untreated animals (negative control) reached the final stage of the disease (stage 4) and were euthanized. At this point, the remaining animals in the study are euthanized regardless of the stage of their tumors. The studies last about 5-10 weeks, depending on the cell line. After completion of the final X-ray, blood is drawn from the animals by cardiac puncture (to assess serum bone markers; see below). Endpoint x-ray images are then distributed to 5 volunteers who mark each image according to the scoring system detailed above. The scores for each mouse are average and expressed as the average osteolytic score or percentage of animals with severe osteolysis (animals with score greater than 2).

Biological Example 8

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Mouse Trap5b Assay (IDS Inc., Fountain Hills, AZ)

This assay is a solid phase assay of immunofixed enzyme activity for the determination of osteoclast-derived tartrate-resistant acid phosphatase 5b in mouse serum samples. Trap5b is expressed by bone resorption osteoclasts and secreted into the circulation. Thus, serum Trap5b is considered a useful marker of osteoclast activity, number and bone resorption.

Mouse Trap5b assay utilizes a polyclonal antibody prepared using recombinant mouse Trap5b as antigen. In the test, the antibody is incubated in anti-rabbit IgG coated microplate wells. After washing, the standard, controls and diluted serum samples are incubated in the wells, and Trap5b binding activity is determined with a chromogenic substrate to develop color. The reaction is stopped and the absorbance of the reaction mixture is read in a 405 nm microplate reader. The color intensity is directly proportional to the amount and activity of Trap5b present in the sample. By plotting the mean absorbance for each standard in the ordinate relative to the abscissa concentration, values ​​for unknown samples can be read from the calibration curve and expressed in U / L Trap5b. The analytical sensitivity of the assay is 0.1 U / L and the inter and intra-assay variation is below 10%. Trap5b levels correlated well with average osteolytic outcome.

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145 (assessed by x-ray).

While a number of preferred embodiments of the invention and variations thereof have been described in detail, other modifications and methods of use will be readily apparent to those skilled in the art. Accordingly, it is to be understood that various applications, modifications and substitutions may be made of equivalents without departing from the spirit of the invention or the scope of the claims.

The percent inhibition activities of the compounds of Tables 2 and 3, when tested at about 1 μΜ in the assay indicated as described in the Biological Examples, are shown respectively in Tables 5, 6 and 7. It is contemplated that compounds with 0% inhibition 1 μΜ will exhibit inhibitory activities at a higher concentration. An N / A means that the compound was not tested in that particular assay.

Table 5. Activities of the compounds of Table 2 | Comp | PDGFRp | CSF-1R | cKit | M-NFS-60 CP pCSFlR |

<img file="PT2010528T_D0180.tif" />

Table 6. Activities of the compounds of Table 3

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<td></td><td>—G ..........................</td>

<td>Comp | PDGFRp [CSF-1R [cKit | m-NFS-60</td><td>CP | pCSFlR</td>

<td> 7 | 9 | 100 | 12 | 35</td><td>N / A</td>

<td>10 (9 (67 [0 (N / A</td><td>N / A</td>

<td> 11 | 60 | 100 | 0 | 69</td><td> [ 92</td>

<td>12 [1 [95 | 0 [N / A</td><td>N / A</td>

<td>13 | 0 | 93 | 9 | N / A</td><td>N / A</td>

<td>14 (0 (96 3 (N / A</td><td>N / A</td>

<td>15 J 7 | 99 | 15 | 4</td><td>N / A</td>

<td> 16 [ 9 [ 99 [ 12 [ 30</td><td>N / A</td>

<td>17 12 | 99 j 19 (23</td><td>[N / A</td>

<td>18 [23 l 99 [80 [22</td><td>N / A</td>

<td> 19 ( 0 | 96 | 8 ( 23</td><td>N / A</td>

<td>20 | 0 | 76 | 0 | N / A</td><td>N / A</td>

<td> 47 | 4 | 100 | 0 | 55</td><td> 1 83</td>

<td>48 | 16 | 99 l 14 [26</td><td> ———</td>

<td> 49 [ 12 | 100 | 21 [ 33</td><td> [ 77</td>

<td> 50 ( 14 ( 99 | 13 ( 18</td><td>N / A</td>

<td>51 | 0 | 95 | 10 | N / A</td><td>j N / A</td>

<td> 125 [9(99[2[ 17</td><td>N / A</td>

<td> 155 | 18 | 99 | 34 | 0</td><td>N / A</td>

<td> 156 | 3 | 100 1 36 | 33</td><td>N / A</td>

<td> 157 | 17 | 100 | 43 | 42</td><td>N / A</td>

<td>158 [l (99 | 27 [0</td><td>| N / A</td>

<td> 159 | 23 | 100 | 29 | 98</td><td>l 96</td>

<td> 160 ( 19 ( 100 ( 95 [ 30</td><td>j N / A</td>

<td>161 | 26 | 54 | 20 | N / A</td><td>N / A</td>

<td> 162 30 100 20 100</td><td> [ ₉₉</td>

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<td></td><td>—-Η ..........................</td>

<td>Comp | PDGFRp [CSF-1R [cKit | m-NFS-60</td><td>CP | pCSFlR</td>

<td> 164 | 87 | 100 | 79 | 100</td><td> [ 100</td>

<td> 165 83 [ 100 [ 83 100</td><td> | 98</td>

<td> 166 | 0 | 100 | 13 | 49</td><td> | 78</td>

<td>167 0 [100 | 16 l 24</td><td>[N / A</td>

<td> 168 | 43 | 100 | 40 [ 100</td><td> [ 99</td>

<td>169 | 0 | 75 | 12 | N / A</td><td>[N / A</td>

<td> 170 | 23 | 100 | 28 | 100</td><td> 1 96</td>

<td> 171 18 [ 100 [ 25 [ 96</td><td> | 95</td>

<td> 172 [ 18 | 100 | 22 [ 100</td><td> [ 97</td>

<td>173 [8 l 100 [18 [52</td><td> [ 81</td>

<td>174 | 3 | 86 | 18 | N / A</td><td>[N / A</td>

<td>175 | 1 | 100 (11 l 17</td><td>[N / A</td>

<td>176 | 0 | 63 | 12 | N / A</td><td>[N / A</td>

<td>177 | 0 [67 [15 [N / A</td><td>[N / A</td>

<td>178 [-3 | 82 | 17 [N / A</td><td>[N / A</td>

<td> 179 [ 16 [ 98 | 10 [ 27</td><td>[N / A</td>

<td> 180 | 12 | 100 | 23 | 96</td><td> } 93</td>

<td>181 [17 [72 [27] N / A</td><td>[N / A</td>

<td> 182 | 4 [ 100 [ 16 | 100</td><td> 1 98</td>

<td> 184 | 84 [ 100 [ 45 [ 100</td><td> [ 95</td>

<td> 185 | 29 | 100 | 32 | 100</td><td> | 94</td>

<td> 186 14 [ 100 | 13 100</td><td> | 99</td>

<td> 187 | 25 | 100 | 32 | 100</td><td> [ 98</td>

<td> 188 35 [ 100 [ 55 [ 38</td><td>[N / A</td>

<td> 189 | 23 [ 100 | 31 | 26</td><td>[N / A</td>

<td> 190 19 98 22 5</td><td>N / A</td>

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<td></td><td>—-Η ..........................</td>

<td>Comp | PDGFRp [CSF-1R [cKit | m-NFS-60</td><td>CP | pCSFlR</td>

<td>191 | 15 | 45 | 19 | N / A</td><td>N / A</td>

<td> 192 ( 22 [ 99 [ 56 |_ 14</td><td>N / A</td>

<td> 193 | 15 | 95 | 27 | 0</td><td>| N / A</td>

<td>194 16 77 [20 [N / A</td><td>N / A</td>

<td>195 | 25 | 81 | 90 | N / A</td><td>N / A</td>

<td> 196 | 23 | 100 | 29 [ 100</td><td> | 98</td>

<td> 197 | 89 | 100 | 93 | 100</td><td> 1 98</td>

<td> 200 19 100 [ 29 [ 69</td><td> | 83</td>

<td>203 15! 98 I 22 [20</td><td>[N / A</td>

<td> 209 [ 1 [ 100 | 17 [ 36</td><td>N / A</td>

<td> 224 | 0 | 100 | 26 | 33</td><td>N / A</td>

<td>225 | 0 | 90 | 21 | N / A</td><td>N / A</td>

<td>226 | 0 | 80 | 20 | N / A</td><td>N / A</td>

<td> 231 | 49 | 100 | 9 [ 100</td><td> | ₉₉</td>

<td> 232 [ 64 | 100 | 34 [ 100</td><td> [ 99</td>

<td> 258 62 100 | 44 100</td><td> | 99</td>

<td> 259 | 6 | 100 | 24 | 100</td><td> } 98</td>

<td> 260 [ 0 100 [ 25 100</td><td> | 98</td>

<td> 261 | 30 | 100 | 25 | 100</td><td> 1 98</td>

<td> 262 | 3 | 100 [ 20 [ 46</td><td> | 48</td>

<td> 265 | 12 | 100 | 9 | 32</td><td>N / A</td>

<td> 266 21 100 | 18 100</td><td> | 94</td>

<td> 267 | 26 | 100 | 16 | 100</td><td> [ 96</td>

<td>280 22 | _ 100 j 11 100</td><td>j 65</td>

<td> 281 | 36 | 100 | 22 | 100</td><td> 1 96</td>

<td>282 | 7 99 í 23 22</td><td>N / A</td>

102010528

149 [Comp | PDGFRp [CSF-1R [cKit | m-NFS-60 CP | pCSFlR |

<td> ] 283 [ 20 99</td><td>38 [0 N / A</td>

<td> 284 53 100</td><td>[61 100 N / A</td>

<td>| 285 f 8 | 99</td><td>| 33 [0 | N / A</td>

<td> [ 286 [ 0 100</td><td>| 23 l 71 | 94 j</td>

<td>1 287 | 20 I 100</td><td>| 57 | 35 [N / A |</td>

<td> [ 288 | 24 | 100</td><td>78 70 N / A</td>

<td> | 291 | 0 | 48</td><td>4 [N / DN / D</td>

<td> 293 0 [ 100</td><td> [ 22 [ 67 | 82 |</td>

<td> | 320 20 | 100</td><td>1 16 25 [N / A</td>

<td> 322 [ 11 [ 99</td><td>[18 [21 N / A |</td>

<td> | 324 | 19 | 99</td><td>| 21 | 15 [N / A |</td>

<td> [ 344 | 65 | 100</td><td> | 45 | 100 | 95 |</td>

<td> | 345 | 13 | 100</td><td> 1 19 | 97 | 93 |</td>

<td> [ 346 | 0 | 100</td><td>I 13 | 67 | 84 j</td>

<td> | 359 [ 19 1 100</td><td> | 31 [ 95 [ 93 |</td>

<td> [ 360 16 100</td><td> | 29 100 98</td>

<td> | 368 | 8 | 100</td><td>| 24 | 12 j N / A |</td>

<td> 369 12 100</td><td>26 44 N / A</td>

<td> | 371 | 9 | 100</td><td> 1 29 | 69 | 79 |</td>

<td> [ 372 | 8 | 100</td><td>I 21 | 75 | 88 j</td>

<td> | 376 | 28 | 100</td><td> | 72 | 87 | 77 |</td>

<td>[377 49,100</td><td>| 75 93 | N / A |</td>

<td> | 378 | 43 | 100</td><td> | 62 | 90 [ 87 |</td>

<td> 381 20 |_ 100</td><td>j 30 100 j 99 |</td>

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