Treatment of hair loss disorders with deuterated jak inhibitors

RELATED APPLICATIONS This application claims the benefit of U.S. Provisional Application Nos. 62/331,827, filed May 4, 2016, 62/338,869, filed May 19, 2016, 62/418,774, filed November 7, 2016, 62/419,237, filed November 8, 2016, 62/434,404, filed December 14, 2016, 62/466,358, filed March 2, 2017, and 62/492,758, filed May 1, 2017. The entire teachings of the above applications are incorporated herein by reference.

The present invention relates to the treatment of hair loss disorders with deuterated JAK inhibitors.

Many current pharmaceuticals suffer from poor absorption, distribution, metabolism and/or excretion (ADME) properties that prevent the drug from being widely used or limit its use in certain indications. Poor ADME properties are also a major reason for drug candidates being dropped in clinical trials. In some cases, formulation techniques and prodrug strategies can be utilized to improve certain ADME properties, but these approaches often fail to address the underlying ADME problems that exist for many drugs and drug candidates. One such problem is rapid metabolism, which causes many drugs that would otherwise be very effective in treating diseases to be cleared too quickly from the body. A possible solution to rapid drug clearance is frequent or high dosing to achieve sufficiently high plasma levels of the drug. However, this results in many potential treatment problems (e.g., poor patient compliance with dosing regimens, more severe side effects with higher doses, and increased treatment costs). Rapidly metabolized drugs can also expose patients to undesirable toxic or reactive metabolites.

Another ADME limitation that affects many pharmaceuticals is the formation of toxic or biologically reactive metabolites.As a result, some patients to whom the drug is administered may experience toxicity, or the safe administration of such drugs may be restricted to administering suboptimal amounts of active agent to patients.In certain cases, changing the administration interval or formulation approach may help reduce clinical side effects, but the formation of such undesirable metabolites is often inherent to the metabolism of the compound.

In some selected cases, metabolic inhibitors are co-administered with drugs that are cleared too quickly. This is the case for drugs of the protease inhibitor class used to treat HIV infection. The FDA generally recommends co-administering this drug with ritonavir, an inhibitor of the cytochrome P450 enzyme 3A4 (CYP3A4), the enzyme responsible for the metabolism of this drug (see Non-Patent Document 1). However, ritonavir causes side effects and increases the pill burden of HIV patients who already have to take a combination of different drugs. Similarly, the CYP2D6 inhibitor quinidine has been added to dextromethorphan in order to reduce the rapid CYP2D6 metabolism of dextromethorphan in the treatment of emotion dysregulation. However, quinidine has undesirable side effects that greatly limit its use in potential combination therapy (see Non-Patent Document 2; and Non-Patent Document 3).

In general, combining drugs with cytochrome P450 (CYP) inhibitors is not a sufficient strategy to reduce drug clearance. Inhibition of the activity of CYP enzymes may affect the metabolism and clearance of other drugs metabolized by the same enzymes. Inhibition of CYP may cause the accumulation of other drugs in the body to toxic levels.

A potentially attractive strategy for improving the metabolic properties of drugs is deuterium modification. This approach attempts to delay the CYP-mediated metabolism of drugs or reduce the formation of undesirable metabolites by replacing one or more hydrogen atoms with deuterium atoms. Deuterium is a safe, stable, non-radioactive isotope of hydrogen. Compared to hydrogen, deuterium forms stronger bonds with carbon. In selected cases, the increased bond strength imparted by deuterium can positively affect the ADME properties of drugs, potentially improving their efficacy, safety, and/or tolerability. At the same time, since the size and shape of deuterium are essentially identical to those of hydrogen, the replacement of hydrogen with deuterium would not be expected to affect the biochemical potency and selectivity of drugs compared to the original chemical entity that contains only hydrogen.

Over the past 35 years, the effect of deuterium substitution on metabolic rate has been reported for only a small fraction of approved drugs (see, e.g., 4; 5 ("Foster"); 6; 7 ("Fisher"). Results have been variable and unpredictable. For some compounds, deuteration caused a decrease in metabolic clearance in vivo. In others, metabolism was unchanged. Still others showed an increase in metabolic clearance. The variability of deuterium effects has also led experts to question or dismiss deuterium modification as a viable drug design strategy to inhibit deleterious metabolism (see Foster at p. 35 and Fisher at p. 101).

The effect of deuterium modification on the metabolic properties of a drug is not predictable, even when the deuterium atom is incorporated at a known metabolic site. Only by actually preparing and testing a deuterated drug can one determine whether and how much its metabolic rate differs from that of its non-deuterated counterpart. See, for example, Fukuto et al. (Non-Patent Document 8). Many drugs have multiple sites at which they can be metabolized. The site at which deuterium substitution is required and the degree of deuteration required to see an effect on metabolism, if any, will be different for each drug.

Ruxolitinib phosphate is a heteroaryl-substituted pyrrolo[2,3-d]pyrimidine (also known as 3(R)-cyclopentyl-3-[4-(7H-pyrrolo[2,3-d]pyrimidin-4-yl)-1H-pyrazol-1-yl]propanenitrile phosphate and (R)-3-(4-(7H-pyrrolo[2,3-d]pyrimidin-4-yl)-1H-pyrazol-1-yl)-3-cyclopentylpropanenitrile phosphate) that inhibits the Janus Associated Kinases (JAKs) JAK1 and JAK2. These kinases mediate the signal transduction of many cytokines and growth factors important for hematopoiesis and immune function. JAK signaling involves the recruitment of STATs (signal transducers and activators of transcription) to cytokine receptors, followed by STAT activation and nuclear localization, where gene expression is regulated.

Ruxolitinib phosphate is currently approved for the treatment of patients with intermediate- or high-risk myelofibrosis (e.g., primary myelofibrosis, myelofibrosis evolving from polycythemia vera, and myelofibrosis evolving from essential thrombocythemia). Ruxolitinib phosphate is also currently in clinical trials for the treatment of additional conditions.

<p><nplcit><text>Kempf, DJ et al., Antimicrobial agents and chemotherapy, 1997, 41(3):654-60</text></nplcit><nplcit><text>Wang, L et al., Clinical Pharmacology and Therapeutics, 1994, 56(6Pt1):659-67</text></nplcit><nplcit><text>FDA label for quinidine at www.accessdata.fda.gov</text></nplcit><nplcit><text>Blake, MI et al, J Pharm Sci,1975,64:367-91</text></nplcit><nplcit><text>Foster, AB, Adv Drug Res, 1985, 14:1-40.</text></nplcit><nplcit><text>Kushner, DJ et al, Can J Physiol Pharmacol, 1999, 79-88</text></nplcit><nplcit><text>Fisher,MB et al,Curr Opin Drug Discov Devel,2006,9:101-09</text></nplcit><nplcit><text>J. Med. Chem., 1991, 34, 2871-76</text></nplcit></p>

<p>Despite the beneficial activity of ruxolitinib, there is a continuing need for new compounds to treat the above-mentioned diseases and conditions.</p>

<p>Deuterated analogs of ruxolitinib (e.g., Compound (I), (R)-3-(4-(7H-pyrrolo[2,3-d]pyrimidin-4-yl)-1H-pyrazol-1-yl)-3-(cyclopentyl-2,2,3,3,4,4,5,5-d₈It has now been discovered that compound (I), also referred to as propanenitrile or D8-ruxolitinib, is useful in the treatment of hair loss disorders (e.g., alopecia areata). <img file="JP7586868B2_D0001.tif" /> It is expressed as:</p><p>In certain embodiments, compound (I) is administered as a pharma- ceutically acceptable salt (e.g., phosphate salt). Compound (I) may be administered at a dose ranging from about 4 mg to about 50 mg per day (or the equivalent weight based on a salt such as compound (I) phosphate salt), administered as a single dose or divided doses (e.g., twice per day). Based on these discoveries, novel therapeutic methods using compound (I) or a pharma- ceutically acceptable salt thereof for treating hair loss disorders in mammalian subjects are disclosed herein.</p><p>One aspect of the present invention is a method for treating hair loss disorders that can be treated by a compound that modulates the activity of Janus-associated kinase 1 (JAK1) and/or Janus-associated kinase 2 (JAK2). The method comprises administering to a subject (e.g., a mammalian subject) an effective amount of Compound (I) or a pharma- ceutically acceptable salt thereof (i.e., an equivalent amount of a pharma- ceutically acceptable salt (e.g., a phosphate salt)) once or twice per day, wherein the amount of Compound (I) or a pharma- ceutically acceptable salt thereof ranges from about 4 mg/day to about 50 mg/day, e.g., about 5 mg/day, about 10 mg/day, about 20 mg/day, about 30 mg/day, about 40 mg/day, or about 50 mg/day. In certain embodiments, the amount of Compound (I) or a pharma- ceutically acceptable salt thereof is about 4 mg/day, 8 mg/day, 16 mg/day, 32 mg/day, or 48 mg/day. In certain embodiments, the amount of compound (I) or its pharma- ceutically acceptable salt is 8 mg/day, 16 mg/day, 24 mg/day or 32 mg/day. In certain embodiments, the amount of compound (I) or its pharma- ceutically acceptable salt is 8 mg/day, 16 mg/day, 24 mg/day or 32 mg/day. In certain embodiments, the amount of compound (I) or its pharma- ceutically acceptable salt is 4 mg, 8 mg, 12 mg or 16 mg twice per day. In certain embodiments, the hair loss disorder is alopecia areata. In certain embodiments, the subject is a human. Preferably, compound (I) or its pharma- ceutically acceptable salt (e.g., phosphate) is orally administered in any of the above dosages. Preferably, compound (I) or its pharma- ceutically acceptable salt is orally administered in any of the above dosages in a pharmaceutical formulation that is a tablet.</p><p>In an alternative aspect, the present invention provides a method of treating a hair loss disorder, comprising topically administering to a subject (e.g., a mammalian subject) an effective amount of Compound (I) or a pharma- ceutically acceptable salt thereof (i.e., an equivalent amount of a pharma- ceutically acceptable salt, e.g., a phosphate salt). In certain embodiments, the compound is administered in a pharmaceutical composition (e.g., a cream, ointment, lotion, foam, or the like) that is formulated for topical administration.</p><p>In another aspect, the present invention provides a method for inducing hair growth in a subject. The method comprises administering to a mammalian subject an effective amount of Compound (I) or a pharma- ceutically acceptable salt thereof (i.e., an equivalent amount of a pharma- ceutically acceptable salt, such as a phosphate salt), once or twice per day, wherein the amount of Compound (I) or a pharma- ceutically acceptable salt thereof ranges from about 4 mg/day to about 50 mg/day, such as about 5 mg/day, about 10 mg/day, about 20 mg/day, about 30 mg/day, about 40 mg/day, or about 50 mg/day. In certain embodiments, the amount of Compound (I) or a pharma- ceutically acceptable salt thereof is about 4 mg/day, 8 mg/day, 16 mg/day, 32 mg/day, or 48 mg/day. In certain embodiments, the amount of Compound (I) or a pharma- ceutically acceptable salt thereof is 8 mg/day, 16 mg/day, 24 mg/day, or 32 mg/day. In certain embodiments, the subject suffers from hair loss disorder, and in further embodiments, the hair loss disorder is alopecia areata.In certain embodiments, the subject is a human.In one embodiment, the subject is a human aged 6 years or older.Preferably, compound (I) or its pharmaceutically acceptable salt (e.g., phosphate) is orally administered in any of the above dosages.Preferably, compound (I) or its pharmaceutically acceptable salt is orally administered in any of the above dosages in a pharmaceutical formulation that is a tablet.</p><p>Another aspect of the present invention is a method for treating an autoimmune skin disorder that can be treated with a compound that modulates the activity of Janus-associated kinase 1 (JAK1) and/or Janus-associated kinase 2 (JAK2). The method comprises administering to a subject (e.g., a mammalian subject) an effective amount of Compound (I) or a pharma- ceutically acceptable salt thereof (i.e., an equivalent amount of a pharma- ceutically acceptable salt (e.g., a phosphate salt)) once or twice per day, wherein the amount of Compound (I) or a pharma- ceutically acceptable salt thereof ranges from about 4 mg/day to about 50 mg/day, e.g., about 5 mg/day, about 10 mg/day, about 20 mg/day, about 30 mg/day, about 40 mg/day, or about 50 mg/day. In certain embodiments, the amount of Compound (I) or a pharma- ceutically acceptable salt thereof is about 4 mg/day, 8 mg/day, 16 mg/day, 32 mg/day, or 48 mg/day. In certain embodiments, the amount of compound (I) or its pharma- ceutically acceptable salt is 8 mg/day, 16 mg/day, 24 mg/day or 32 mg/day. In certain embodiments, the amount of compound (I) or its pharma- ceutically acceptable salt is 8 mg/day, 16 mg/day, 24 mg/day or 32 mg/day. In certain embodiments, the amount of compound (I) or its pharma- ceutically acceptable salt is 4 mg, 8 mg, 12 mg or 16 mg twice per day. In certain embodiments, the autoimmune skin disorder is alopecia areata, vitiligo, atopic dermatitis (ezcema) or psoriasis. In certain embodiments, the subject is a human. Preferably, compound (I) or its pharma- ceutically acceptable salt (e.g., phosphate) is orally administered in any of the above dosages. Preferably, compound (I) or its pharma- ceutically acceptable salt is orally administered in any of the above dosages in a pharmaceutical formulation that is a tablet.</p><p>Another aspect of the present invention is compound (I) or its pharmaceutically acceptable salt (i.e., an equivalent amount of pharmaceutically acceptable salt (e.g., phosphate)) for use in treating hair loss disorder that can be treated by compound that modulates the activity of Janus kinase 1 (JAK1) and/or Janus kinase 2 (JAK2).The compound can be administered in the dosage regimen disclosed herein.In certain embodiments, the hair loss disorder is alopecia areata.</p><p>Yet another aspect of the present invention is the use of compound (I) or its pharmaceutically acceptable salt (i.e., an equivalent amount of pharmaceutically acceptable salt (e.g., phosphate)) for the manufacture of a medicament for the method of treating hair loss disorder that can be treated by a compound that regulates the activity of Janus-associated kinase 1 (JAK1) and/or Janus-associated kinase 2 (JAK2).The compound can be administered in the dosage regimen disclosed herein.In certain embodiments, the hair loss disorder is alopecia areata.</p><p>Another aspect of the present invention is a pharmaceutical composition comprising Compound (I) in the range of about 4 mg to about 50 mg (e.g., about 5 mg, about 10 mg, about 20 mg, about 30 mg, about 40 mg, or about 50 mg) or an equivalent amount of a pharma- ceutically acceptable salt thereof (i.e., an equivalent amount of a pharma- ceutically acceptable salt (e.g., a phosphate salt)), together with a pharma- ceutically acceptable carrier or diluent. In certain embodiments, the amount of Compound (I) or a pharma- ceutically acceptable salt thereof is about 4 mg, 8 mg, 16 mg, 24 mg, 32 mg, or 48 mg. In certain embodiments, the amount of Compound (I) or a pharma- ceutically acceptable salt thereof is 4 mg, 8 mg, 12 mg, or 16 mg. In certain embodiments, the pharmaceutical composition is a tablet.</p><p>That is, the gist of the present invention is as follows.</p><p>[1] A method for treating a hair loss disorder in a mammalian subject, comprising administering to the subject an amount ranging from 4 mg to 50 mg per day of a compound having the following structural formula: <img file="JP7586868B2_D0002.tif" /> or a pharma- ceutically acceptable salt thereof.</p><p>[2] The method according to [1], wherein the hair loss disorder is alopecia areata.</p><p>[3] The method of [1] or [2], comprising administering to the subject about 5 mg/day, about 10 mg/day, about 20 mg/day, about 30 mg/day, about 40 mg/day or about 50 mg/day of Compound (I) or a pharma- ceutically acceptable salt thereof.</p><p>[4] The method according to any one of [1] to [3], comprising administering to the subject about 8 mg/day, about 12 mg/day, about 16 mg/day or about 24 mg/day of compound (I) or a pharma- ceutically acceptable salt thereof.</p><p>[5] The method according to any one of [1] to [4], wherein the compound is administered orally.</p><p>[6] The method according to any one of [1] to [5], wherein the compound is administered in a pharmaceutical formulation that is a tablet.</p><p>[7] The method according to any one of [1] to [6], wherein the compound is administered once per day.</p><p>[8] The method according to any one of [1] to [6], wherein the compound is administered twice a day.</p><p>[9] The method according to any one of [1] to [8], wherein in compound (I), any atom not designated as deuterium is present at the natural isotopic abundance of that atom.</p><p>[10] A pharmaceutical composition comprising a pharma- ceutically acceptable carrier or diluent and 4 to 50 mg of a compound having the following structural formula: <img file="JP7586868B2_D0003.tif" /> or a pharma- ceutically acceptable salt thereof.</p><p>[11] The pharmaceutical composition according to [10], comprising about 5 mg, about 10 mg, about 20 mg, about 30 mg, about 40 mg or about 50 mg of Compound (I) or a pharma- ceutically acceptable salt thereof.</p><p>[12] The pharmaceutical composition according to [10] or [11], wherein the pharmaceutical composition is suitable for oral administration.</p><p>[13] The pharmaceutical composition described in [10] or [11], wherein the composition is a tablet.</p><p>[14] The pharmaceutical composition according to any one of [10] to [13], wherein the composition is administered once per day.</p><p>[15] The pharmaceutical composition of [10], wherein the composition is a tablet comprising: (i) about 5.2% by weight of Compound (I) phosphate, (ii) about 90.8% by weight of microcrystalline cellulose, (iii) about 2.5% by weight of povidone, (iv) about 0.5% by weight of colloidal silicon dioxide, and (v) about 1.0% by weight of magnesium stearate.</p><p>[16] The pharmaceutical composition of [15], wherein the composition is a 200 mg tablet containing about 10.5 mg of Compound (I) phosphate.</p><p>[17] The pharmaceutical composition of [10], wherein the composition is a tablet comprising: (i) about 5.3% by weight of Compound (I) phosphate, (ii) about 88.2% by weight of microcrystalline cellulose, (iii) about 5.0% by weight of povidone, (iv) about 0.5% by weight of colloidal silicon dioxide, and (v) about 1.0% by weight of magnesium stearate.</p><p>[18] The pharmaceutical composition of [17], wherein the composition is a 200 mg tablet containing about 10.6 mg of Compound (I) phosphate.</p><p>[19] The pharmaceutical composition of [10], wherein the composition is a tablet comprising: (i) about 10.6% by weight of Compound (I) phosphate, (ii) about 82.9% by weight of microcrystalline cellulose, (iii) about 5.0% by weight of povidone, (iv) about 0.5% by weight of colloidal silicon dioxide, and (v) about 1.0% by weight of magnesium stearate.</p><p>[20] The pharmaceutical composition of [19], wherein the composition is a 200 mg tablet containing about 21.1 mg of Compound (I) phosphate.</p><p>[21] A method of treating a hair loss disorder in a mammalian subject, comprising administering to the subject about 4 mg of a compound having the following structural formula: <img file="JP7586868B2_D0004.tif" /> or a pharma- ceutically acceptable salt thereof twice daily.</p><p>[22] The method according to [21], wherein compound (I) is administered as about 5.3 mg of phosphate twice a day.</p><p>[23] A method of treating a hair loss disorder in a mammalian subject, comprising administering to the subject about 8 mg of a compound having the following structural formula: <img file="JP7586868B2_D0005.tif" /> or a pharma- ceutically acceptable salt thereof twice daily.</p><p>[24] The method according to [23], wherein compound (I) is administered as about 10.5 mg of phosphate twice daily.</p><p>[25] A method of treating a hair loss disorder in a mammalian subject, comprising administering to the subject about 12 mg of a compound having the following structural formula: <img file="JP7586868B2_D0006.tif" /> or a pharma- ceutically acceptable salt thereof twice daily.</p><p>[26] The method according to [25], wherein compound (I) is administered as about 15.8 mg of phosphate twice a day.</p><p>[27] A method of treating a hair loss disorder in a mammalian subject, comprising administering to the subject about 16 mg of a compound having the following structural formula: <img file="JP7586868B2_D0007.tif" /> or a pharma- ceutically acceptable salt thereof twice daily.</p><p>[28] The method according to [27], wherein compound (I) is administered twice a day in the form of about 21.1 mg of phosphate.</p><p>[29] The method according to any one of [21] to [28], wherein the hair loss disorder is alopecia areata.</p><p>[30] The method according to any one of [21] to [29], wherein the compound is administered orally.</p><p>[31] The method according to any one of [21] to [30], wherein the compound is administered in a pharmaceutical formulation that is a tablet.</p>

<p>The present invention may provide for the treatment of hair loss disorders with deuterated JAK inhibitors.</p>

<figref num="1">Figure 1 shows the design and results of a single ascending dose (SAD) study in healthy volunteers. Figure 1A shows the design of the SAD study, Figure 1B is a graph showing the plasma concentrations of CTP-543 (compound (I)) from 0 to 48 hours after dosing, and Figure 1C is a table showing the mean PK parameters of CTP-543 (compound (I)) in the SAD study.</figref><figref num="2">Figure 22 shows the design and results of a multiple ascending dose (MAD) study in healthy volunteers. Figure 22A shows the design of the MAD study, Figure 2B is a graph showing the plasma concentrations of CTP-543 (compound (I)) from 0 to 24 hours on days 1 and 7 of the MAD study, and Figure 2C is a table showing the mean PK parameters of CTP-543 (compound (I)) in the MAD study.</figref>

The term "treat" refers to reducing, inhibiting, attenuating, reducing, stopping or stabilizing the onset or progression of a disease (e.g., a disease or disorder detailed herein), reducing the severity of the disease, or improving the symptoms associated with the disease.For example, treating hair loss disorder includes hair regrowth, preventing further hair loss, or reducing the rate of hair loss.

"Hair loss disorder" refers to any condition or disorder that results in hair loss in one or more areas of the body. Hair loss disorders include, but are not limited to, androgenetic alopecia, alopecia areata, telogen effluvium, alopecia areata, alopecia totalis, and alopecia universalis.

The term "mammal", as used herein, includes humans as well as non-human mammals, such as cats, dogs, sheep, cows, pigs, goats, non-human primates (e.g., monkeys and apes), and the like.

It will be recognized that some natural isotopic abundance variations occur in synthesized compounds depending on the origin of chemicals used in synthesis.Therefore, preparations of ruxolitinib will essentially contain a small amount of deuterated isotopologues.Despite this variation, the concentration of naturally abundant stable hydrogen and carbon isotopes is low and insignificant compared to the degree of stable isotopic substitution of compounds of the present invention.See, for example, Wada, E et al., Seikagaku, 1994, 66:15; Gannes, LZ et al., Comp Biochem Physiol Mol Integr Physiol, 1998, 119:725.

In compound (I), any atom not specifically designated to a particular isotope is intended to represent any stable isotope of that atom. Unless otherwise stated, when a position is specifically designated as "H" or "hydrogen", this position is understood to have hydrogen as the isotopic composition of hydrogen's natural abundance. Similarly, unless otherwise stated, when a position is specifically designated as "D" or "deuterium", this position is understood to have deuterium at an abundance at least 3000 times greater than the natural abundance of deuterium, which is 0.015% (i.e., at least 45% incorporation of deuterium).

The term "isotopic enrichment factor" as used herein means the ratio between the abundance of an isotope and the natural abundance of a particular isotope.

In other embodiments, the compounds of the invention have the following isotopic enrichment factors for each designated deuterium atom: at least 3500 (52.5% deuterium incorporation at each designated deuterium atom), at least 4000 (60% deuterium incorporation), at least 4500 (67.5% deuterium incorporation), at least 5000 (75% deuterium), at least 5500 (82.5% deuterium incorporation), at least 6000 (90% deuterium incorporation), at least 6333.3 (95% deuterium incorporation), at least 6466.7 (97% deuterium incorporation), at least 6600 (99% deuterium incorporation) or at least 6633.3 (99.5% deuterium incorporation).

The term "isotopologue" refers to a species that differs in chemical structure from Compound (I) only in isotopic composition.

The term "compound" when referring to a compound of the present invention refers to a collection of molecules having the same chemical structure, although there may be isotopic variations among the constituent atoms of these molecules. Therefore, it will be clear to one skilled in the art that a compound represented by a particular chemical structure containing a deuterium atom as shown also includes lesser amount isotopologues having hydrogen atoms at one or more of the designated deuterium positions in the structure. The relative amounts of such isotopologues in the compounds of the present invention will depend on many factors, such as the isotopic purity of the deuteration reagent used to make the compound, and the efficiency of deuterium incorporation in the various synthetic steps used to prepare the compound.

The present invention also provides a salt of compound (I).The salt of the compound of the present invention is formed between an acid group and a basic group of the compound (e.g., an amino functional group), or between a basic group and an acid group of the compound (e.g., a carboxyl functional group).According to another embodiment, the compound is a pharma-ceutically acceptable acid addition salt (e.g., a phosphate salt).

The term "pharmaceutically acceptable" as used herein refers to a component that is, within the scope of sound medical judgment, suitable for use in contact with the tissues of humans and other mammals without undue toxicity, irritation, allergic response, and the like, and is commensurate with a reasonable benefit/risk ratio. A "pharmaceutically acceptable salt" refers to any non-toxic salt that is capable of providing, directly or indirectly, a compound of the present invention upon administration to a recipient. A "pharmaceutically acceptable counterion" refers to an ionic portion of a salt that is non-toxic when released from the salt upon administration to a recipient.

Acids commonly utilized to form pharma-ceutically acceptable salts include inorganic acids such as hydrogen disulfide, hydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuric acid, and phosphoric acid, and organic acids such as para-toluenesulfonic acid, salicylic acid, tartaric acid, bitartaric acid, ascorbic acid, maleic acid, besylic acid, fumaric acid, gluconic acid, glucuronic acid, formic acid, glutamic acid, methanesulfonic acid, ethanesulfonic acid, benzenesulfonic acid, lactic acid, oxalic acid, para-bromophenylsulfonic acid, carbonic acid, succinic acid, citric acid, benzoic acid, and acetic acid, and related inorganic and organic acids. Such pharma- ceutically acceptable salts include, for example, sulfate, pyrosulfate, bisulfate, sulfite, bisulfite, phosphate, monohydrogen phosphate, dihydrogen phosphate, metaphosphate, pyrophosphate, chloride, bromide, iodide, acetate, propionate, decanoate, caprylate, acrylate, formate, isobutyrate, caprate, heptanoate, propionate, oxalate, malonate, succinate, suberate, sebacate, fumarate, maleate, butylene-1,4-dioate, hexaphosphate ... San-1,6-dioate, benzoate, chlorobenzoate, methylbenzoate, dinitrobenzoate, hydroxybenzoate, methoxybenzoate, phthalate, terephthalate, sulfonate, xylenesulfonate, phenylacetate, phenylpropionate, phenylbutyrate, citrate, lactate, β-hydroxybutyrate, glycolate, maleate, tartrate, methanesulfonate, propanesulfonate, naphthalene-1-sulfonate, naphthalene-2-sulfonate, mandelate and other salts. In one embodiment, pharma-ceutically acceptable acid addition salts include those formed with mineral acids, such as hydrochloric acid and hydrobromic acid, and especially those formed with organic acids, such as maleic acid.

The term "stable compound," as used herein, refers to a compound that has sufficient stability to permit the manufacture of the compound and maintains its integrity for a period of time sufficient to be useful for the purposes detailed herein (e.g., therapeutic products, intermediates used in the manufacture of therapeutic compounds, formulation into isolatable or storable intermediate compounds, treatment of a disease or condition that responds to a therapeutic agent).

"D" and "d" both refer to deuterium. "Stereoisomer" refers to both enantiomers and diastereomers. "Ter" and "t-" refer to tertiary, respectively. "US" refers to the United States of America.

"Deuterium substituted" refers to the replacement of one or more hydrogen atoms with a corresponding number of deuterium atoms.

In one aspect, the present invention provides a method for treating hair loss disorders that can be treated with a compound that modulates (e.g., inhibits) the activity of JAK (JAK1, JAK2, and/or JAK3). The method comprises administering to a mammalian subject an effective amount of Compound (I) or a pharma- ceutically acceptable salt thereof (i.e., an equivalent amount of a pharma- ceutically acceptable salt (e.g., a phosphate salt)) once or twice per day, wherein the amount of Formula (I) or a pharma- ceutically acceptable salt thereof ranges from about 4 mg/day to about 50 mg/day (e.g., 4 mg/day to 50 mg/day), such as about 5 mg/day (e.g., 5 mg/day), about 10 mg/day (e.g., 10 mg/day), about 20 mg/day (e.g., 20 mg/day), about 30 mg/day (e.g., 30 mg/day), about 40 mg/day (e.g., 40 mg/day), or about 50 mg/day (e.g., 50 mg/day).

In certain embodiments, the amount of Compound (I) or a pharma- ceutically acceptable salt thereof (i.e., an equivalent amount of a pharma- ceutically acceptable salt (e.g., a phosphate salt) administered in a method for treating a hair loss disorder is about 4 mg/day (e.g., 4 mg/day), about 8 mg/day (e.g., 8 mg/day), about 16 mg/day (e.g., 16 mg/day), about 32 mg/day (e.g., 32 mg/day), or about 48 mg/day (e.g., 48 mg/day).

In certain embodiments, the amount of compound (I) or its pharmaceutically acceptable salt administered in the method for treating hair loss disorder is about 8mg/day (e.g., 8mg/day), about 16mg/day (e.g., 16mg/day), about 24mg/day (e.g., 24mg/day) or about 32mg/day (e.g., 32mg/day).In certain embodiments, the amount of compound (I) or its pharmaceutically acceptable salt is about 8mg/day (e.g., 8mg/day), about 12mg/day (e.g., 12mg/day), about 16mg/day (e.g., 16mg/day) or about 24mg/day (e.g., 24mg/day).

In certain embodiments, the amount of compound (I) or its pharmaceutically acceptable salt administered in the method for treating hair loss disorder is 10.6mg/day of compound (I) phosphate (e.g., 5.3mg per day, twice).In certain embodiments, the amount of compound (I) or its pharmaceutically acceptable salt is 21.1mg/day of compound (I) phosphate (e.g., 10.5mg per day, twice).In certain embodiments, the amount of compound (I) or its pharmaceutically acceptable salt is 31.6mg/day of compound (I) phosphate (e.g., 15.8mg per day, twice).In certain embodiments, the amount of compound (I) or its pharmaceutically acceptable salt is 42.2mg/day of compound (I) phosphate (e.g., 21.1mg per day, twice).

In certain embodiments, the amount of compound (I) or its pharma- ceutically acceptable salt administered in the method for treating hair loss disorder is about 4 mg (e.g., 4 mg) twice per day. In a specific embodiment, compound (I) is administered as about 5.3 mg (e.g., 5.3 mg) of compound (I) phosphate twice per day. In certain embodiments, compound (I) or its pharma- ceutically acceptable salt administered in the method for treating hair loss disorder is about 8 mg (e.g., 8 mg) twice per day. In a specific embodiment, compound (I) is administered as about 10.5 mg (e.g., 10.5 mg) of compound (I) phosphate twice per day.

In certain embodiments, the amount of Compound (I) or its pharma- ceutically acceptable salt administered in the method for treating hair loss disorder is about 12 mg (e.g., 12 mg) twice per day. In a specific embodiment, Compound (I) is administered as about 15.8 mg (e.g., 15.8 mg) of Compound (I) phosphate twice per day. In certain embodiments, the amount of Compound (I) or its pharma- ceutically acceptable salt administered in the method for treating hair loss disorder is about 16 mg (e.g., 16 mg) twice per day. In a specific embodiment, Compound (I) is administered as about 21.1 mg (e.g., 21.1 mg) of Compound (I) phosphate twice per day. In certain embodiments, the hair loss disorder is alopecia areata. In certain embodiments, the subject is a human. In one embodiment, the subject is a human aged 6 years or older. Preferably, Compound (I) or its pharma- ceutically acceptable salt (e.g., phosphate) is orally administered in any of the dosages described herein. Preferably, Compound (I) or a pharma- ceutically acceptable salt thereof is administered orally in a pharmaceutical formulation that is a tablet, at any of the dosages described herein.

In another aspect, the present invention provides a method for inducing hair growth in a subject, the method comprising administering to a mammalian subject an effective amount of Compound (I) or a pharma- ceutically acceptable salt thereof (i.e., an equivalent amount of a pharma- ceutically acceptable salt, such as a phosphate salt), once or twice per day, wherein the amount of Compound (I) or a pharma- ceutically acceptable salt thereof is in the range of about 4 mg/day to about 50 mg/day (e.g., 4 mg/day to 50 mg/day), such as about 5 mg/day (e.g., 5 mg/day), about 10 mg/day (e.g., 10 mg/day), about 20 mg/day (e.g., 20 mg/day), about 30 mg/day (e.g., 30 mg/day), about 40 mg/day (e.g., 40 mg/day), or about 50 mg/day (e.g., 50 mg/day).

In certain embodiments, the amount of Compound (I) or a pharma- ceutically acceptable salt thereof administered in a method to induce hair growth is about 4 mg/day (e.g., 4 mg/day), about 8 mg/day (e.g., 8 mg/day), about 16 mg/day (e.g., 16 mg/day), about 32 mg/day (e.g., 32 mg/day), or about 48 mg/day (e.g., 48 mg/day).

In certain embodiments, the amount of Compound (I) or a pharma- ceutically acceptable salt thereof administered in a method for inducing hair growth is about 8 mg/day (e.g., 8 mg/day), about 16 mg/day (e.g., 16 mg/day), about 24 mg/day (e.g., 24 mg/day) or about 32 mg/day (e.g., 32 mg/day). In certain embodiments, the amount of Compound (I) or a pharma- ceutically acceptable salt thereof administered in a method for inducing hair growth is about 8 mg/day (e.g., 8 mg/day), about 12 mg/day (e.g., 12 mg/day), about 16 mg/day (e.g., 16 mg/day) or about 24 mg/day (e.g., 24 mg/day).

In certain embodiments, the amount of compound (I) or its pharma- ceutically acceptable salt administered in the method of inducing hair growth is 10.6 mg/day of compound (I) phosphate (e.g., 5.3 mg twice a day). In certain embodiments, the amount of compound (I) or its pharma- ceutically acceptable salt is 21.1 mg/day of compound (I) phosphate (e.g., 10.5 mg twice a day). In certain embodiments, the amount of compound (I) or its pharma- ceutically acceptable salt is 31.6 mg/day of compound (I) phosphate (e.g., 15.8 mg twice a day). In certain embodiments, the amount of compound (I) or its pharma- ceutically acceptable salt is 42.2 mg/day of compound (I) phosphate (e.g., 21.1 mg twice a day).

In certain embodiments, the amount of Compound (I) or a pharma- ceutically acceptable salt thereof administered in a method of inducing hair growth is about 4 mg (e.g., 4 mg) twice per day. In a specific embodiment, Compound (I) is administered as about 5.3 mg (e.g., 5.3 mg) of Compound (I) phosphate twice per day.

In certain embodiments, the amount of Compound (I) or a pharma- ceutically acceptable salt thereof administered in a method for inducing hair growth is about 8 mg (e.g., 8 mg) twice per day. In a specific embodiment, Compound (I) is administered as about 10.5 mg (e.g., 10.5 mg) of Compound (I) phosphate twice per day.

In certain embodiments, the amount of Compound (I) or a pharma- ceutically acceptable salt thereof administered in a method of inducing hair growth is about 12 mg (e.g., 12 mg) twice per day. In a specific embodiment, Compound (I) is administered as about 15.8 mg (e.g., 15.8 mg) of Compound (I) phosphate twice per day.

In certain embodiments, the amount of Compound (I) or a pharma- ceutically acceptable salt thereof administered in a method of inducing hair growth is about 16 mg (e.g., 16 mg) twice per day. In a specific embodiment, Compound (I) is administered as about 21.1 mg (e.g., 21.1 mg) of Compound (I) phosphate twice per day.

In certain embodiments, the subject suffers from hair loss disorder, and in further embodiments, the hair loss disorder is alopecia areata.In certain embodiments, the subject is a human.In one embodiment, the subject is a human aged 6 years or older.Preferably, compound (I) or its pharmaceutically acceptable salt (e.g., phosphate) is orally administered at any of the above dosages.Preferably, compound (I) or its pharmaceutically acceptable salt is orally administered at any of the above dosages in a pharmaceutical formulation that is a tablet.

Hair loss disorders include, but are not limited to, androgenetic alopecia, alopecia areata, telogen effluvium, alopecia totalis, and alopecia universalis.

Alopecia areata is an autoimmune disease that can affect up to 650,000 Americans at any one time, causing partial or complete loss of hair on the scalp and body. The most commonly affected area is the scalp, but any area that has hair can be affected alone or together with the scalp. Onset of the disease can occur throughout life, affecting both women and men. Alopecia areata can be associated with serious psychological consequences, such as anxiety and depression. Currently, there are no drugs approved by the US Food and Drug Administration (FDA) for the treatment of alopecia areata.

In specific embodiments, the condition is alopecia areata in a subject, such as a mammalian (e.g., human) patient in need thereof. In certain embodiments, the alopecia areata is moderate to severe alopecia areata (e.g., hair loss over at least 30% of the scalp, hair loss over at least 40% of the scalp, or hair loss over at least 50% of the scalp).

In one embodiment of any aspect, the compound is administered orally once per day. In another embodiment of any aspect, the compound is administered orally twice per day.

The effective dose will also vary, as recognized by those of skill in the art, depending on the disease being treated, the severity of that disease, the route of administration, the sex, age, and general health of the subject, the use of excipients, the possibility of co-use with other therapeutic treatments (e.g., the use of other drugs), and the judgment of the treating physician.

Administration of Compound (I) or a pharma- ceutically acceptable salt thereof (e.g., a phosphate salt) may be continued for as long as necessary to treat the alopecia, for example, for one week, for two weeks, for one month, for two months, for three months, for four months, for six months, for one year, for two years, for five years, for ten years, or longer.

The effectiveness of the treatment of hair loss disorders such as alopecia areata can be measured in various ways, some of which are known in the art.For example, the "severity of alopecia tool" (otherwise known as SALT) is a useful assessment scale for assessing the degree of hair loss, developed by the National Alopecia Areata Foundation Task Force.See, for example, Olsen EA, Hordinsky MK, Price VH, et al. Alopecia areata investigational assessment guidelines-Part II. J Am Acad Dermatol. 2004:51:440-447 (herein incorporated by reference). The patient's SALT score is calculated by measuring the percentage of hair loss in each of the four scalp regions and adding the totals to arrive at a composite score. Hair regrowth is reflected in a reduction in the SALT score. For example, no hair on the scalp will result in a SALT score of 100, and complete hair regrowth will result in a SALT score of 0. In certain embodiments, the methods of treatment described herein may result in at least a 10-point SALT score improvement after treatment (e.g., from a SALT score of 100 before treatment to a SALT score of 90 after treatment). In further embodiments, the methods of treatment described herein may result in at least a 20-point, 30-point, 40-point, 50-point, 60-point, 70-point, 80-point, 90-point, or 100-point SALT score improvement. In certain embodiments, the methods of treatment described herein may result in at least a 20% improvement from baseline in the patient's SALT score, or at least a 30% improvement from baseline in the patient's SALT score, or at least a 40% improvement from baseline in the patient's SALT score, or at least a 50% improvement from baseline in the patient's SALT score, or at least a 60% improvement from baseline in the patient's SALT score, or at least a 70% improvement from baseline in the patient's SALT score, following treatment.

In certain embodiments, treatment continues for a period of at least 4 weeks, or at least 8 weeks, or at least 12 weeks, or at least 16 weeks, or at least 20 weeks, or at least 24 weeks, or at least 28 weeks, or at least 32 weeks, or at least 36 weeks, or at least 40 weeks, or at least 44 weeks, or at least 48 weeks, or at least 52 weeks.

In certain embodiments, compound (I) or its pharmaceutically acceptable salt is administered in combination with a second therapeutic agent.Preferably, this second therapeutic agent is a drug useful for treating hair loss disorder or autoimmune condition, such as an inhibitor of JAK1, JAK2 or JAK3 and/or STAT1.Such inhibitors include ruxolitinib, tofacitinib, baricitinib, filgotinib and the like.Other orally administered second therapeutic agents include drugs used in the treatment of alopecia areata, such as oral corticosteroids.

In the case of pharmaceutical compositions containing a second therapeutic agent, the effective amount of the second therapeutic agent is about 20% to 100% of the dosage normally utilized in a monotherapy regimen using only that agent. Preferably, the effective amount is about 70% to 100% of the usual monotherapy dose. The usual monotherapy dosage of the second therapeutic agent is known in the art. See, for example, Wells et al., eds., Pharmacotherapy Handbook, 2nd Edition, Appleton and Lange, Stamford, Conn. (2000); PDR Pharmacopoeia, Tarascon Pocket Pharmacopoeia 2000, Deluxe Edition, Tarascon Publishing, Loma Linda, Calif. (2000); FDA approved labeling information for ruxolitinib and tofacitinib; and clinical trial information for baricitinib and filgotinib. Each of these references is incorporated herein by reference in its entirety.

It is expected that some of the second therapeutic agents mentioned above will act synergistically with the compounds of the present invention.If this synergy occurs, it will be possible to reduce the effective amount of the second therapeutic agent and/or compound (I) or its pharmaceutically acceptable salt from the amount required in monotherapy.This has the following advantages: minimizing the adverse side effects of either the second therapeutic agent or compound (I) or its pharmaceutically acceptable salt, synergistically improving efficacy, improving ease of administration or use, and/or reducing the overall cost of preparing or formulating the compound.

In another embodiment, any of the above-mentioned methods of treatment comprises the further step of co-administering one or more second therapeutic agents to the subject in need.The second therapeutic agent can be selected from any other therapeutic agent known to be useful in the treatment of hair loss disorders, such as alopecia areata.The selection of the second therapeutic agent also depends on the specific disease or condition to be treated.The examples of the second therapeutic agent that can be employed in the method of the present invention are those described above for use in the combination composition comprising compound (I) or its pharmaceutically acceptable salt and the second therapeutic agent.The additional therapeutic agent includes agents used in the treatment of alopecia areata, such as topical minoxidil, injected corticosteroids, and anthralin cream or ointment.

The term "co-administered" as used herein means that the second therapeutic agent may be administered together with compound (I) or a pharmaceutically acceptable salt thereof, either as part of a single dosage form (e.g., the composition of the present invention comprising a compound of the present invention and a second therapeutic agent as described above) or as separate multiple dosage forms. Alternatively, the additional agent may be administered prior to, consecutively with, or after administration of compound (I) or a pharmaceutically acceptable salt thereof. In such combination therapy treatment, both compound (I) or a pharmaceutically acceptable salt thereof and the second therapeutic agent are administered in a conventional manner. The administration of a composition of the present invention comprising both compound (I) or a pharmaceutically acceptable salt thereof and a second therapeutic agent to a subject does not exclude the separate administration of the same therapeutic agent (any other second therapeutic agent or compound (I) or a pharmaceutically acceptable salt thereof) to said subject at separate times during the course of treatment.

Effective amounts of these second therapeutic agents are known to those skilled in the art, and guidance for administration can be found in the patents and published patent applications referenced herein, as well as in Wells et al., eds., Pharmacotherapy Handbook, 2nd Edition, Appleton and Lange, Stamford, Conn. (2000); PDR Pharmacopoeia, Tarascon Pocket Pharmacopoeia 2000, Deluxe Edition, Tarascon Publishing, Loma Linda, Calif. (2000) and other medical texts. However, determining the optimal effective amount range of the second therapeutic agent is well within the purview of one skilled in the art.

In one embodiment of the present invention, when a second therapeutic agent is administered to a subject, the effective amount of compound (I) or a pharma- ceutically acceptable salt thereof is less than the effective amount when the second therapeutic agent is not administered. In another embodiment, the effective amount of the second therapeutic agent is less than the effective amount when the second therapeutic agent is not administered. In this way, undesirable side effects associated with high doses of either agent may be minimized. Other potential advantages, including but not limited to improved dosing regimens and/or reduced drug costs, will be apparent to those skilled in the art.

In yet another aspect, the present invention provides the use of compound (I) or a pharma- ceutically acceptable salt thereof (i.e., an equivalent amount of a pharma- ceutically acceptable salt (e.g., a phosphate salt) alone or together with one or more of the second therapeutic agents described above, in the manufacture of a medicament, either as a single composition or in separate dosage forms, for the treatment or prevention of a disease, disorder or condition of a subject as described above. Another aspect of the present invention is compound (I) or a pharma- ceutically acceptable salt thereof for use in the treatment or prevention of a disease, disorder or condition of a subject as detailed herein.

Another aspect of the present invention is a pharmaceutical composition comprising about 4 mg to about 50 mg (e.g., about 5 mg, about 10 mg, about 20 mg, about 30 mg, about 40 mg, or about 50 mg) of Compound (I) or an equivalent amount of a pharma- ceutically acceptable salt thereof, together with a pharma- ceutically acceptable carrier or diluent. In certain embodiments, the amount of Compound (I) or a pharma- ceutically acceptable salt thereof is about 4 mg, 8 mg, 16 mg, 24 mg, 32 mg, or 48 mg. In certain embodiments, the amount of Compound (I) or a pharma- ceutically acceptable salt thereof is 4 mg, 8 mg, 12 mg, or 16 mg. In certain embodiments, the amount of Compound (I) or a pharma- ceutically acceptable salt thereof is 5.3 mg of Compound (I) phosphate. In certain embodiments, the amount of Compound (I) or a pharma- ceutically acceptable salt thereof is 10.5 or 10.6 mg of Compound (I) phosphate. In certain embodiments, the amount of Compound (I) or its pharma- ceutically acceptable salt is 15.8 mg of Compound (I) phosphate. In certain embodiments, the amount of Compound (I) or its pharma- ceutically acceptable salt is 21.1 mg of Compound (I) phosphate. In certain embodiments, the pharmaceutical composition is a tablet.

Another embodiment of the present invention is a unit dose formulation comprising a range of about 4 mg to about 50 mg (e.g., about 5 mg, about 10 mg, about 20 mg, about 30 mg, about 40 mg, or about 50 mg) of Compound (I) or an equivalent amount of a pharma- ceutically acceptable salt thereof, together with a pharma- ceutically acceptable carrier or diluent. In certain embodiments, the amount of Compound (I) or a pharma- ceutically acceptable salt thereof is about 4 mg, 8 mg, 16 mg, 24 mg, 32 mg, or 48 mg. In certain embodiments, the amount of Compound (I) or a pharma- ceutically acceptable salt thereof is about 4 mg, 8 mg, 12 mg, or 16 mg. In certain embodiments, the amount of Compound (I) or a pharma- ceutically acceptable salt thereof is 5.3 mg of Compound (I) phosphate. In certain embodiments, the amount of Compound (I) or a pharma- ceutically acceptable salt thereof is 10.5 or 10.6 mg of Compound (I) phosphate. In certain embodiments, the amount of Compound (I) or a pharma- ceutically acceptable salt thereof is 15.8 mg of Compound (I) phosphate. In certain embodiments, the amount of Compound (I) or a pharma- ceutically acceptable salt thereof is 21.1 mg of Compound (I) phosphate. In certain embodiments, the unit dosage form is a tablet.

In one embodiment, every atom not designated as deuterium is present in Compound (I) or a pharma- ceutically acceptable salt thereof at the natural isotopic abundance of that atom.

The synthesis of compound (I) or a pharma- ceutically acceptable salt thereof (e.g., phosphate salt) can be easily achieved by suitable modification of the method described in U.S. Pat. No. 9,249,149, the teachings of which are incorporated herein by reference. For example, U.S. Pat. No. 9,249,149 describes the use of D9-intermediate 15 to produce D9-ruxolitinib product, and intermediate A in the method described in U.S. Pat. No. 9,249,149 can be used to prepare ruxolitinib product. <img file="JP7586868B2_D0008.tif" /> In addition, the use of intermediate B instead of intermediate 14 of U.S. Pat. No. 9,249,149 gives compound (I). <img file="JP7586868B2_D0009.tif" /> Compound (I) can be prepared using ruxolitinib, and removal of the amino protecting group can be achieved by basic cleavage (e.g., with sodium hydroxide). Phosphoric acid can be used to convert compound (I) (free base) to the phosphate salt of compound (I). A further method for preparing ruxolitinib (i.e., non-deuterated compound (I)) is disclosed in U.S. Pat. No. 9,000,161, and this method can be used with the use of a suitable deuteration reagent to prepare compound (I).

Such methods may be carried out utilizing corresponding deuterated and optionally isotopically containing reagents and/or intermediates to synthesize the compounds detailed herein, or with the aid of standard synthetic protocols known in the art for introducing isotopic atoms into a chemical structure.

The present invention also provides pharmaceutical compositions comprising an effective amount of Compound (I) or a pharma- ceutically acceptable salt thereof (i.e., an equivalent amount of a pharma- ceutically acceptable salt, such as a phosphate salt), and a pharma- ceutically acceptable carrier. The carrier is "acceptable" in the sense of being compatible with the other ingredients of the formulation and, in the case of a pharma- ceutically acceptable carrier, not deleterious to the recipient of the carrier in the amounts used in the drug. In certain embodiments, the pharmaceutical composition is provided as a unit dosage form.

The present invention relates to a pharmaceutical composition comprising a pharmaceutical carrier or diluent and 4 to 50 mg of a compound having the following structural formula: <img file="JP7586868B2_D0010.tif" /> or a pharma- ceutically acceptable salt thereof (i.e., an equivalent amount of a pharma- ceutically acceptable salt, such as a phosphate salt).

Pharmaceutically acceptable carriers, adjuvants and excipients that may be used in the pharmaceutical compositions of the present invention include, but are not limited to, ion exchangers, alumina, aluminum stearate, lecithin, serum proteins such as human serum albumin, buffer substances such as phosphates, glycine, sorbic acid, potassium sorbate, partial glyceride mixtures of saturated vegetable fatty acids, water, salts or electrolytes such as protamine sulfate, disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, zinc salts, colloidal silica, magnesium trisilicate, polyvinylpyrrolidone, cellulosic substances, polyethylene glycol, sodium carboxymethylcellulose, polyarylates, waxes, polyethylene-polyoxypropylene-block polymers, polyethylene glycol and wool fat.

If necessary, the solubility and bioavailability of the compounds of the present invention in pharmaceutical compositions can be enhanced by methods known in the art. One method includes the use of lipid additives in the formulation. See "Oral Lipid-Based Formulations:Enhancing the Bioavailability of Poorly Water-Soluble Drugs(Drugs and the Pharmaceutical Sciences)," David J.Hauss, ed.Informa Healthcare, 2007; and "Role of Lipid Excipients in Modifying Oral and Parenteral Drug Delivery:Basic Principles and Biological Examples," Kishor M.Wasan, ed.Wiley-Interscience, 2006.

Another known method of enhancing bioavailability is the use of amorphous forms of the compounds of the invention, optionally formulated with poloxamers (e.g., LUTROL and PLURONIC (BASF Corporation)) or block copolymers of ethylene oxide and propylene oxide. See U.S. Patent No. 7,014,866 and U.S. Patent Application Publication Nos. 20060094744 and 20060079502.

The pharmaceutical composition of the present invention includes those suitable for oral administration.Other formulations can be conveniently provided in unit dosage form (e.g., tablets, sustained release capsules, granules) and liposomes, and can be prepared by any method known in the field of pharmacy.See, for example, Remington: The Science and Practice of Pharmacy, Lippincott Williams &amp; Wilkins, Baltimore, MD (20th ed.2000).

Such preparatory methods include the step of bringing into association with the molecule to be administered ingredients such as the carrier which constitutes one or more accessory ingredients. In general, the compositions are prepared by uniformly and intimately bringing into association the active ingredients with liquid carriers, liposomes or finely divided solid carriers, or both, and then, if necessary, shaping the product.

In certain embodiments, the compound is administered orally.The composition of the present invention suitable for oral administration can be provided as separate units, such as capsules, sachets or tablets (each containing a predetermined amount of active ingredient); as powder or granules; as a solution or suspension in aqueous or non-aqueous liquid; as an oil-in-water liquid emulsion; as a water-in-oil liquid emulsion; filled in liposomes; or as a bolus, etc.Soft gelatin capsules can be useful for containing such suspensions, and can advantageously increase the absorption rate of the compound.In a specific embodiment, the compound is administered orally as a tablet.

For tablets for oral use, commonly used carriers include lactose and cornstarch. Lubricants (e.g. magnesium stearate) are also generally added. For oral administration in capsule form, useful diluents include lactose and dry cornstarch. For oral administration of aqueous suspension, active ingredient is combined with emulsifying and suspending agents. If necessary, certain sweeteners and/or flavorings and/or colorings can be added. In another embodiment, the composition is in the form of a tablet. In certain embodiments, the exemplary formulation for this tablet is disclosed in U.S. Pat. No. 8,754,224, the teachings of which are incorporated herein by reference.

In certain embodiments, the tablet formulation comprises about 4mg to about 50mg of Compound (I) or an equivalent amount of its pharma- ceutically acceptable salt (e.g., phosphate salt) and the following inactive ingredients: colloidal silicon dioxide, magnesium stearate, microcrystalline cellulose and povidone. Compression after wet granulation produces a tablet containing Compound (I) or its pharma- ceutically acceptable salt. For example, to prepare a 200mg tablet containing the equivalent amount of 16mg of Compound (I), 10.6wt% of Compound (I) phosphate salt and 64.44wt% of Avicel PH-101 microcrystalline cellulose are mixed in a high shear granulator, and 8.5wt% of Kollidon 30 aqueous solution (containing Kollidon 30, which is polyvinylpyrrolidone (povidone); 5wt% (based on the total weight of this formulation)) is added during mixing to form granules. The granules are tray dried in an oven at 60±10°C and milled using a Quadro Comil U5 mill. The granules retained on the screen are pushed through a #20 mesh screen using a stainless steel spatula. The resulting milled granules are mixed with Avicel PH-200 microcrystalline cellulose (18.5% by weight), Aerosil 200 colloidal silicon dioxide (0.5% by weight) and Hyqual magnesium stearate (1% by weight) in a Turbula mixer to form a final blend. This final blend is compressed into 200 mg tablets using a Riva Piccola rotary press equipped with a 0.451" x 0.229" D-type modified capsule shape tooling. Each tablet contains 21.1 mg of Compound (I) (equivalent to 16 mg of Compound (I) free base).

In certain embodiments, the tablet contains about 10.5 mg or about 10.6 mg of Compound (I) phosphate (equivalent to 8 mg of Compound (I) free base).

In a particular embodiment, the tablet comprises the following ingredients:

<tables><img file="JP7586868B2_D0011.tif" /></tables>

In another particular embodiment, the tablet comprises the following ingredients:

<tables><img file="JP7586868B2_D0012.tif" /></tables>

In an alternative specific embodiment, the tablet comprises the following ingredients:

<tables><img file="JP7586868B2_D0013.tif" /></tables>

In yet another particular embodiment, the tablet comprises the following ingredients:

<tables><img file="JP7586868B2_D0014.tif" /></tables>

In another embodiment, the composition of the invention further comprises a second therapeutic agent, which may be selected from any compound or therapeutic agent known to have or exhibit beneficial properties when administered together with a compound having the same mechanism of action as ruxolitinib.

Preferably, the second therapeutic agent is an agent useful in treating hair loss disorders or autoimmune conditions (e.g., an inhibitor of JAK1, JAK2 or JAK3 and/or STAT1). Such inhibitors include ruxolitinib, tofacitinib, baricitinib, filgotinib and the like. Other second therapeutic agents include oral corticosteroids.

In another embodiment, the present invention provides separate dosage forms of Compound (I) or a pharma- ceutically acceptable salt thereof and one or more of any of the second therapeutic agents described above, wherein Compound (I) or a pharma- ceutically acceptable salt thereof and the second therapeutic agent are associated with each other. The term "associated with each other" as used herein means that the separate dosage forms are packaged together or otherwise associated with each other such that it is readily apparent that the separate dosage forms are intended to be sold together and administered (within less than 24 hours of each other, either sequentially or simultaneously).

In the pharmaceutical composition of the present invention, compound (I) or its pharma- ceutically acceptable salt is present in an effective amount.As used herein, the term "effective amount" refers to an amount sufficient to treat the target disorder when administered in a suitable dosage regimen.

The interrelationship of dosages for animals and humans (based on milligrams per square meter of body surface) is described in Freireich et al., Cancer Chemother. Rep, 1966, 50:219. Body surface area can be approximately determined from the subject's height and weight. See, e.g., Scientific Tables, Geigy Pharmaceuticals, Ardsley, NY, 1970, 537.

In one embodiment, an effective amount of Compound (I) (either as the free base or an equivalent amount of a pharma- ceutically acceptable salt (e.g., a phosphate salt)) may range from about 4 mg to 50 mg per day (e.g., 4 mg to 50 mg per day), for example, about 5 mg/day (e.g., 5 mg/day), about 10 mg/day (e.g., 10 mg/day), about 20 mg/day (e.g., 20 mg/day), about 30 mg/day (e.g., 30 mg/day), about 40 mg/day (e.g., 40 mg/day), or about 50 mg/day (e.g., 50 mg/day). In certain embodiments, the amount may be about 4 mg/day (e.g., 4 mg/day), about 8 mg/day (e.g., 8 mg/day), about 16 mg/day (e.g., 16 mg/day), about 24 mg/day (e.g., 24 mg/day), about 32 mg/day (e.g., 32 mg/day), or about 48 mg/day (e.g., 48 mg/day). In one embodiment, a dose of about 4 mg/day (e.g., 4 mg/day), about 8 mg/day (e.g., 8 mg/day), about 16 mg/day (e.g., 16 mg/day), about 24 mg/day (e.g., 24 mg/day), about 32 mg/day (e.g., 32 mg/day) or about 48 mg/day (e.g., 48 mg/day) is administered once per day. In one embodiment, a dose of 16 mg/day is administered as two 8 mg tablets of Compound (I) (either as the free base or an equivalent amount of a pharma- ceutically acceptable salt (e.g., a phosphate salt)) administered together (i.e., as a single dose). In another embodiment, a dose of 16 mg/day is administered as one 16 mg tablet of Compound (I) (either as the free base or an equivalent amount of a pharma-ceutically acceptable salt (e.g., a phosphate salt). In another embodiment, a dose of 4 mg/day, 8 mg/day, 16 mg/day, 24 mg/day, 32 mg/day or 48 mg/day is administered in divided doses twice per day (e.g., a 48 mg/day dose is administered as 24 mg twice per day). In another embodiment, a dose of 8 mg/day, 16 mg/day, 24 mg/day or 32 mg/day is administered twice per day in divided doses (e.g., a dose of 32 mg/day is administered as 16 mg of Compound (I) (either as the free base or as an equivalent amount of a pharma- ceutically acceptable salt, e.g., a phosphate salt) twice per day (i.e., in separate doses). In a specific embodiment, a dose of 16 mg/day is administered as 8 mg of Compound (I) (either as the free base or as an equivalent amount of a pharma- ceutically acceptable salt, e.g., a phosphate salt) twice per day (i.e., in separate doses). References to the amount of Compound (I) or a pharma- ceutically acceptable salt thereof will be understood to include the amount of a pharma- ceutically acceptable salt, e.g., a phosphate salt, of Compound (I) that is equivalent to the stated value of Compound (I) as the free base (e.g., 10.5 mg of Compound (I) phosphate salt is equivalent to 8 mg of Compound (I) free base).

In certain embodiments, the effective amount of compound (I) or its pharma- ceutically acceptable salt is about 4 mg (e.g., 4 mg) twice per day. In a specific embodiment, the effective amount of compound (I) is administered as about 5.3 mg (e.g., 5.3 mg) of compound (I) phosphate twice per day. In certain embodiments, the effective amount of compound (I) or its pharma- ceutically acceptable salt is administered as about 8 mg (e.g., 8 mg) twice per day. In a specific embodiment, compound (I) is administered as about 10.5 mg (e.g., 10.5 mg) of compound (I) phosphate twice per day.

In certain embodiments, the effective amount of Compound (I) or a pharma- ceutically acceptable salt thereof is about 12 mg (e.g., 12 mg) twice per day. In a specific embodiment, the effective amount of Compound (I) is about 15.8 mg (e.g., 15.8 mg) of Compound (I) phosphate twice per day. In certain embodiments, the effective amount of Compound (I) or a pharma- ceutically acceptable salt thereof is about 16 mg (e.g., 16 mg) twice per day. In a specific embodiment, the effective amount of Compound (I) is about 21.1 mg (e.g., 21.1 mg) of Compound (I) phosphate twice per day.

<p>Example 1. Measurement of metabolic stability of D-ruxolitinib using CYP3A4 Supersomes Materials and Methods: Materials: CYP3A4 Supersomes were obtained from Corning Gentest. β-nicotinamide adenine dinucleotide phosphate, reduced form (NADPH), magnesium chloride (MgCl₂) and dimethylsulfoxide (DMSO) were purchased from Sigma-Aldrich. Deuterated test compounds were supplied by Concert Pharmaceuticals.</p><p>Determination of metabolic stability: 10 mM stock solutions of test compounds were prepared in DMSO. 7.5 mM stock solutions were diluted to 12.75 μM in acetonitrile (ACN). CYP3A4 supersomes were diluted in 3 mM MgCl₂The diluted supersomes were placed in triplicate into wells of a 96-well deep-well polypropylene plate. A 10 μL aliquot of 12.75 μM test compound was added to the supersomes and the mixture was preheated for 10 min. The reaction was started by the addition of preheated NADPH solution. The final reaction volume was 0.5 mL and contained 0.1 M potassium phosphate buffer (pH 7.4) and 3 mM MgCl.₂The mixture contained 50 pmol/mL CYP3A4 supersomes, 0.25 μM test compound, and 2 mM NADPH. The reaction mixture was incubated at 37° C. and 50 μL aliquots were removed at 0, 5, 10, 20, and 30 min and stopped by placing into a shallow-well 96-well plate containing 50 μL ice-cold ACN containing internal standard. The plate was stored at 4° C. for 20 min, after which the wells of the plate were filled with 100 μL water, followed by centrifugation to pellet precipitated proteins. The supernatant was transferred to another 96-well plate and analyzed for the amount of parent remaining by LC-MS/MS using an Applied Bio-systems mass spectrometer.</p><p>Data analysis: In vitro t_1/2was calculated from the slope of the linear regression of % parent remaining (ln) versus incubation time.</p><p>In vitro_1/2= 0.693/kk = - [slope of linear regression of % parent remaining (ln) vs incubation time]</p><p>Data analysis was performed using Microsoft Excel Software.</p><p>Compound (I) is a non-deuterated form of ruxolitinib._1/2Approximately 80% longer than_1/2The results show that formula (I) is substantially metabolically stable compared to ruxolitinib in the CYP3A4 supersome assay.</p><p>Example 2: Determination of metabolic stability of D-ruxolitinib using human liver microsomes Materials: Human liver microsomes (20 mg/mL) were obtained from Xenotech, LLC (Lenexa, KS). β-nicotinamide adenine dinucleotide phosphate, reduced form (NADPH), magnesium chloride (MgCl₂) and dimethylsulfoxide (DMSO) were purchased from Sigma-Aldrich. Deuterated test compounds were supplied by Concert Pharmaceuticals.</p><p>Determination of metabolic stability: A 7.5 mM stock solution of the test compound was prepared in DMSO. The 7.5 mM stock solution was diluted to 12.5 μM in acetonitrile (ACN). Human liver microsomes were diluted with 3 mM MgCl₂The diluted microsomes were placed in triplicate in wells of a 96-well deep-well polypropylene plate. A 10 μL aliquot of 12.5 μM test compound was added to the microsomes and the mixture was preheated for 10 min. The reaction was started by the addition of preheated NADPH solution. The final reaction volume was 0.5 mL and contained 0.1 M potassium phosphate buffer (pH 7.4) and 3 mM MgCl.₂The mixture contained 5 mg/mL human liver microsomes, 0.25 μM test compound, and 2 mM NADPH. The reaction mixture was incubated at 37° C. and 50 μL aliquots were removed at 0, 5, 10, 20, and 30 minutes and stopped by placing into a shallow-well 96-well plate containing 50 μL ice-cold ACN containing internal standard. The plate was stored at 4° C. for 20 minutes, after which the wells of the plate were filled with 100 μL water, followed by centrifugation to pellet precipitated proteins. The supernatant was transferred to another 96-well plate and analyzed for the amount of parent remaining by LC-MS/MS using an Applied Bio-systems mass spectrometer.</p><p>Data analysis: In vitro t_1/2was calculated from the slope of the linear regression of % parent remaining (ln) versus incubation time.</p><p>In vitro_1/2= 0.693/kk = - [slope of linear regression of % parent remaining (ln) vs incubation time]</p><p>Data analysis was performed using Microsoft Excel Software.</p><p>Compound (I) is a non-deuterated form of ruxolitinib._1/2Approximately 75% longer than_1/2The results show that compound (I) is substantially metabolically stable compared to ruxolitinib in the HLM assay.</p><p>Example 3: Human Study Single Ascending Dose (SAD) Study: Healthy volunteers were administered doses of Compound (I) or placebo under fasting conditions. The purpose of this study was to evaluate the pharmacokinetics of a single dose of 8 mg, 16 mg, 32 mg or 48 mg of Compound (I) as phosphate (e.g., 10.5 mg of Compound (I) phosphate is equivalent to 8 mg of Compound (I) free base). For each dose group, 6 subjects were administered Compound (I) and 2 subjects were administered placebo. The design of this study is shown in Figure 1A. Metabolites of Compound (I) were analyzed. Compound (I) doses were administered as Compound (I) phosphate as powder in capsules and administered with water. Preliminary results are shown in Figure 1B and Figure 1C.</p><p>Multiple ascending dose study: Healthy volunteers were administered doses of Compound (I) or placebo under fasting conditions. The purpose of this study was to evaluate the pharmacokinetics of daily administration of 8 mg, 16 mg, 24 mg, or 32 mg of Compound (I) as phosphate (e.g., 10.5 mg of Compound (I) phosphate is equivalent to 8 mg of Compound (I) free base). Compound (I) was administered once per day (QD) (8 mg, 24 mg, 32 mg doses) or twice per day (BID) (two 8 mg doses totaling 16 mg/day or two 16 mg doses totaling 32 mg/day) for seven consecutive days. For each dose group, eight subjects were administered Compound (I) and two subjects were administered placebo. The design of this study is shown in Figure 1A. Metabolites of Compound (I) were analyzed. Compound (I) doses were administered as 8 mg tablets with water. Preliminary results are shown in Figures 1B and 1C.</p><p>The final results are shown in the table below.</p><p><tables><img file="JP7586868B2_D0015.tif" /></tables></p><p>The preliminary results of the SAD study are shown in Figures 1B and 1C. The preliminary results of the MAD study are shown in Figures 2B and 2C. The final results of the MAD study are shown in the table below. In Figures 1 and 2, CTP-543 is compound (I) and is administered as the phosphate salt.</p><p><tables><img file="JP7586868B2_D0016.tif" /></tables></p><p><tables><img file="JP7586868B2_D0017.tif" /></tables></p><p>A total of 77 subjects were dosed in the SAD and MAD studies combined (60 received CTP-543 and 17 received placebo). CTP-543 was rapidly absorbed and did not accumulate with repeated dosing. No serious adverse events were reported, with headache being the most common adverse event reported. No dose interruptions or dose modifications related to CTP-543 occurred. Cases of mild neutropenia resolved or showed a tendency to resolve after completion of dosing. No severe neutropenia (grade 3 or 4) was observed.</p><p>In Phase 1 studies, Compound (I) was generally well tolerated. Based on these studies, doses of 8 mg BID (twice daily), 12 mg BID, 16 mg BID and 32 mg BID were selected.</p><p>Further findings from the Phase 1 study indicate that the mean systemic exposure of Compound (I) at 16 mg BID (the highest dose evaluated in the Phase 2a study described below) appears to be comparable to published findings regarding the reported mean exposure of 20 mg BID ruxolitinib, which has been shown to be effective in inducing hair regrowth in patients with moderate to severe alopecia (see, e.g., JCI Insight. 2016;1(15):e89790. doi:10.1172/jci.insight.89790.).</p><p>Pharmacodynamic analysis was also performed during the phase 1 MAD study to evaluate inhibition of IL-6-mediated JAK/STAT signaling and IFN-γ-mediated JAK/STAT signaling. Consistent with the established pharmacodynamic activity of CTP-543 (compound (I)), we observed a dose-dependent decrease in IL-6-stimulated phosphorylated STAT3 (pSTAT3). Overall, pSTAT3 inhibition in all treatment groups returned to near baseline values 24 hours after dosing. Similarly, IFN-γ-mediated STAT1 signaling, which is believed to play an important role in the pathogenesis of alopecia areata, was significantly inhibited in disease-relevant immune cell types at all doses evaluated.</p><p>The Phase 2a study is designed to evaluate the safety and efficacy of Compound (I) after 12 months of treatment with a primary efficacy analysis at 24 weeks. The Phase 2a study is a double-blind, randomized, placebo-controlled, parallel dose study to evaluate the safety and efficacy of Compound (I) in adult patients with moderate to severe alopecia areata. Approximately 100 patients will be randomized to receive one of four doses of Compound (I) as phosphate (e.g., 10.5 mg of Compound (I) phosphate is equivalent to 8 mg of Compound (I) free base). The four doses of Compound (I) are 4, 8 (i.e., about 10.5 mg of Compound (I) phosphate), 12 and 16 mg twice daily, with a placebo group. The primary prognostic indicator will utilize the Severity of Alopecia Tool (SALT) after 24 weeks of treatment. The study will include an additional 28 weeks of treatment with Compound (I) administered to all patients enrolled in the study.</p><p>Without further description, it is believed that one skilled in the art can use the above description and illustrative examples to make and use the compounds of the present invention and to practice the methods of the claims. It should be understood that the above discussion and examples merely provide a detailed description of certain preferred embodiments. It will be apparent to those skilled in the art that various modifications and equivalents are possible without departing from the spirit and scope of the present invention.</p>