KR20230100919A - Aminopyrimidinyl derivatives - Google Patents

Abstract

여기서 R¹ 및 R²는 각각 독립적으로 수소 또는 히드록시이고; 여기서 R¹ 및 R²는 둘 다 히드록시는 아니다.A pharmaceutical composition comprising a compound of formula (I) or a pharmaceutically acceptable salt thereof:

wherein R ¹ and R ² are each independently hydrogen or hydroxy; wherein R ¹ and R ² are not both hydroxy.

Description

Aminopyrimidinyl derivatives {AMINOPYRIMIDINYL DERIVATIVES}

The present invention provides pharmaceutically active aminopyrimidinyl ligands and analogs. Such compounds are useful for inhibiting Janus kinase (JAK). The present invention also relates to compositions comprising such compounds, methods of making such compounds, and methods of treating and preventing conditions mediated by JAKs, particularly TYK2/JAK1.

Protein kinases are a family of enzymes that catalyze the phosphorylation of specific residues in proteins, broadly classified as tyrosine and serine/threonine kinases. Mutation, over-expression or inappropriate regulation, dysregulation or deregulation of growth factors or cytokines, as well as inappropriate kinase activity resulting from over- or under-production, can lead to cancer, cardiovascular disease, allergies, asthma and other respiratory diseases, and autoimmune diseases. It has been implicated in many diseases including, but not limited to, inflammatory diseases, bone diseases, metabolic disorders, and nervous system and neurodegenerative disorders such as Alzheimer's disease. Inappropriate kinase activity triggers a variety of biological cell responses related to cell growth, cell differentiation, cell function, survival, apoptosis and cell motility implicated in the aforementioned diseases and related disorders.

Thus, protein kinases are emerging as an important class of enzymes as targets for therapeutic intervention. In particular, the JAK family of cellular protein tyrosine kinases (JAK1, JAK2, JAK3 and TYK2) play pivotal roles in cytokine signaling (Kisseleva et al., Gene, 2002, 285, 1; Yamaoka et al. Genome Biology 2004, 5, 253). Upon binding to its receptor, the cytokine activates JAK, which in turn phosphorylates the cytokine receptor, thereby docking to signaling molecules, particularly members of the signal transducer and activator of transcription (STAT) family that ultimately drive gene expression. area is created. A number of cytokines are known to activate the JAK family. These cytokines include the interferon (IFN) family (IFN-alpha, IFN-beta, IFN-omega, limitin, IFN-gamma, IL-10, IL-19, IL-20, IL-22), the gp130 family (IL- 6, IL-11, OSM, LIF, CNTF, NNT-1/BSF-3, G-CSF, CT-1, leptin, IL-12, IL-23), gamma C family (IL-2, IL-7 , TSLP, IL-9, IL-15, IL-21, IL-4, IL-13), IL-3 family (IL-3, IL-5, GM-CSF), single chain family (EPO, GH, PRL, TPO), receptor tyrosine kinases (EGF, PDGF, CSF-1, HGF), and G-protein coupled receptors (AT1).

There remains a need for new compounds that effectively and selectively inhibit specific JAK enzymes. JAK enzymes have been shown to be important for the differentiation and function of a number of cell types important in inflammatory and autoimmune diseases, including natural killer cells, B cells and T helper cell types. Aberrant JAK expression is associated with a number of autoimmune or inflammatory conditions. Modulation of immune activity through inhibition of JAK kinase activity may prove useful in the treatment of a variety of immune disorders (O'Shea JJ, Plenge R, Immunity, 36, 542-50 (2012); Murray, P.J., J. Immunol., 178, 2623-2629 (2007); Kisseleva, T., et al., Gene, 285, 1-24 (2002)).

JAK inhibitors known in the art often have properties that make them generally more suitable for oral administration and less suitable for topical administration. Thus, the present invention is not only a potent JAK inhibitor, particularly a TYK2/JAK1 inhibitor, but also exhibits high clearance in human hepatocytes, providing significantly improved systemic clearance, thereby reducing the risk of adverse side effects while maintaining efficacy in the skin upon topical administration. Provided are novel JAK inhibitors that reduce

The present invention provides a compound having the structure of Formula I, or a pharmaceutically acceptable salt thereof:

wherein R ¹ and R ² are each independently hydrogen or hydroxy; wherein R ¹ and R ² are not both hydroxy.

In another aspect, the present invention also provides a pharmaceutical composition comprising a pharmaceutically acceptable carrier and a compound of Formula I or a pharmaceutically acceptable salt thereof.

In another aspect, the invention also provides a method of treating a condition or disorder comprising:

arthritis such as rheumatoid arthritis, juvenile arthritis and psoriatic arthritis;

Autoimmune or inflammatory diseases or disorders such as Hashimoto's thyroiditis, autoimmune hemolytic anemia, autoimmune atrophic gastritis of pernicious anemia, autoimmune encephalomyelitis, autoimmune orchitis, Goodpasture's disease, autoimmune thrombocytopenia, sympathetic ophthalmitis, myasthenia gravis, Graves disease, primary biliary cirrhosis, autoimmune hepatitis, primary sclerosing cholangitis, chronic aggressive hepatitis, non-alcoholic fatty liver disease, non-alcoholic steatohepatitis ulcerative colitis and membranous glomerulopathy, systemic lupus erythematosus, rheumatoid arthritis, psoriatic Arthritis, Sjogren's syndrome, Reiter's syndrome, polymyositis, dermatomyositis, type I interferonopathy such as Aicardi-Goutieres syndrome and other Mendelian diseases of overexpression of type I systemic sclerosis, polyarteritis nodosa , multiple sclerosis, relapsing remitting multiple sclerosis, primary progressive multiple sclerosis, secondary progressive multiple sclerosis and bullous pemphigoid, and further autoimmune diseases such as those which may be O-cell (humoral) based or T-cell based , such as Cogan syndrome, ankylosing spondylitis, Wegener's granulomatosis, autoimmune alopecia, type I or juvenile onset diabetes, or thyroiditis;

Cancers or tumors such as digestive/gastrointestinal cancers, colon cancer, liver cancer, skin cancers such as mast cell tumors and squamous cell carcinomas, breast and mammary gland cancers, ovarian cancer, prostate cancer, lymphomas, leukemias such as acute myelogenous leukemia and chronic myelogenous leukemia, kidney cancer , lung cancer, muscle cancer, bone cancer, bladder cancer, brain cancer, melanoma such as oral and metastatic melanoma, Kaposi's sarcoma, myeloma such as multiple myeloma, myeloproliferative disorders, proliferative diabetic retinopathy, or angiogenesis-related disorders, eg solid tumors;

diabetes, such as type I diabetes or complications from diabetes;

Eye diseases, disorders or conditions, such as autoimmune diseases of the eye, keratoconjunctivitis, vernal conjunctivitis, uveitis such as uveitis associated with Behcet's disease and phaco-induced uveitis, keratitis, keratitis herpetiformis, keratitis, corneal epithelial dystrophy, corneal leukoplakia, ocular Pemphigus, Murren's ulcer, scleritis, Grave's ophthalmopathy, Vogt-Koyanagi-Harada syndrome, keratoconjunctivitis sicca (dry eye), flictenule, iridocyclitis, sarcoidosis, endocrine ophthalmopathy, sympathetic ophthalmia, allergic conjunctivitis, or ocular neovascularization;

intestinal inflammation such as Crohn's disease, ulcerative colitis, inflammatory bowel disease, celiac disease, proctitis, eosinophilic gastroenteritis, or mastocytosis;

neurodegenerative diseases such as motor neuron disease, Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, Huntington's disease, cerebral ischemia, or neurodegenerative diseases caused by traumatic injury, shock, glutamate neurotoxicity or hypoxia; ischemic/reperfusion injury in stroke, myocardial ischemia, renal ischemia, heart attack, cardiac hypertrophy, atherosclerosis and atherosclerosis, organ hypoxia or platelet aggregation;

Skin diseases, conditions or disorders such as atopic dermatitis, hand dermatitis, contact dermatitis, allergic contact dermatitis, irritant contact dermatitis, neurodermatitis, perioral dermatitis, retention dermatitis, hyperhidrotic eczema, dry dermatitis, nummularis Dermatitis, seborrheic dermatitis, eyelid dermatitis, diaper dermatitis, dermatomyositis, lichen planus, lichen sclerosus, alopecia areata, vitiligo, rosacea, epidermolysis bullosa, keratosis pilaris, pityriasis white, pemphigus, vulvovaginitis, acne, chronic spontaneous urticaria, Chronic idiopathic urticaria, chronic physical urticaria, Vogt-Koyanagi-Harada disease, Sutton nevus/nevi, post-inflammatory hypopigmentation, senile leukoplakia, chemical/drug-induced leukoplakia, cutaneous lupus erythematosus , discoid lupus, palmar plantar pustulosis, pemphigoid, sweet's syndrome, hidradenitis suppurativa, psoriasis, plaque psoriasis, pustular psoriasis, nail psoriasis, sinusoidal psoriasis, red psoriasis, psoriatic arthritis, erythematous psoriasis , or inverse psoriasis;

Allergic reactions such as allergic dermatitis in mammals (such as equine allergic diseases such as bite hypersensitivity), summer eczema, skin itching in horses, urticaria, inflammatory airway disease, recurrent airway obstruction, airway hyperresponsiveness, or chronic obstruction lung disease;

asthma and other obstructive airway diseases such as chronic or refractory asthma, late stage asthma, bronchitis, bronchial asthma, allergic asthma, intrinsic asthma, extrinsic asthma, or dust asthma; and

Transplant rejection, such as pancreatic islet transplant rejection, bone marrow transplant rejection, graft-versus-host disease, organ and cell transplant rejection, such as bone marrow, cartilage, cornea, heart, intervertebral disc, islet, kidney, limb, liver, lung, muscle, muscle Blast, nerve, pancreas, skin, small intestine, or organ, or xenograft.

The invention will be further understood from the following description given by way of example only. The present invention relates to a class of aminopyrimidinyl derivatives. In particular, the present invention relates to aminopyrimidinyl compounds useful as inhibitors of JAK. Although the present invention is not limited thereto, awareness of various aspects of the present invention will be obtained through the following discussion and examples.

The terms "isolated" and "in isolated form" for a compound are meant to refer to the physical state of the compound after the compound or salt thereof has been isolated from a synthetic process, eg, a reaction mixture. Accordingly, the terms “isolated” and “in isolated form” for a compound refer to a compound from a purification process or processes (eg, chromatography, recrystallization, etc.) described herein or well known to those skilled in the art, herein refers to the physical state of the compound after it has been obtained in sufficient purity to be characterized by standard analytical techniques well known to those skilled in the art. By way of example, the purification techniques disclosed herein (eg, LC-MS and LC-MS/MS techniques) result in isolated forms of compounds of interest. Such isolation and purification techniques are for products containing at least about 70%, at least about 80%, at least about 90%, at least about 95%, at least about 97%, or at least about 99% by weight of a compound or salt thereof. expected to lead to purity.

The term "subject" refers to a mammal, such as a human, livestock or companion animal. "Patient", "individual" or "subject", used interchangeably, is a mammal, more preferably a human.

The term "companion animal" or "companion animals" refers to an animal kept as a pet or household animal. Examples of companion animals include dogs, cats and rodents such as hamsters, guinea pigs, gerbils, etc., rabbits, ferrets and birds.

The term "livestock" refers to animals raised or raised in an agricultural environment for the manufacture of products such as food or fiber, or for their labor. In some embodiments, livestock is suitable for consumption by a mammal, such as a human. Examples of livestock animals include cows, goats, horses, pigs, sheep, such as rams, and rabbits, as well as birds, such as chickens, ducks, and turkeys.

Unless defined otherwise herein, scientific and technical terms used in connection with the present invention have the meanings commonly understood by one of ordinary skill in the art.

When substituents are described as "independently selected" from a group, each substituent is selected independently of the other. Thus, each substituent may be the same as or different from the other substituent(s).

As used herein, the term “treating”, unless indicated otherwise, means reversing, alleviating, inhibiting the progression of, or preventing the disorder or condition to which the term applies, or one or more symptoms of such disorder or condition. It means delaying its progression, delaying its onset, or preventing it. As used herein, the term "treatment", unless indicated otherwise, refers to the act of treating, such as "treating" as defined immediately above.

The term "selective", when used herein to describe a functionally defined receptor ligand or enzyme inhibitor, is selective for a defined receptor or enzyme subtype when compared to other receptors or enzyme subtypes in the same family. it means. For example, a selective TYK2/JAK1 inhibitor is a compound that inhibits a TYK2/JAK1 enzyme subtype more potently than any other JAK enzyme subtype. This selectivity is, in one embodiment, at least 2-fold (as measured using a conventional binding assay), or in another embodiment, at least 10-fold, or in a further embodiment, at least 100-fold.

The term "therapeutically effective" refers to the ability of an agent to prevent or reduce the severity of a disorder. The phrase “therapeutically effective” is to be understood as equivalent to the phrase “effective in treating, preventing or ameliorating” both—the severity and incidence of cancer, cardiovascular disease, or pain and inflammation relative to the treatment of each agent by itself. It is intended to quantify the amount of an agent that will achieve the purpose of mitigating.

"Pharmaceutically acceptable" means suitable for use in a subject.

1 is a crystalline ((S)-2,2-difluorocyclopropyl)((1R,5S)-3-(2-((1-propyl-1H-pyrazol-4-yl)amino)pyrimidine -4-yl)-3,8-diazabicyclo[3.2.1]octan-8-yl)methanone provides a powder X-ray diffraction pattern obtained for the diffraction peaks in Table 1 with 2-theta values. enumerate
Figure 2 provides the percent change in the CXCL10 biomarker following topical administration of 1% formulations of Example 1 and Compound 2 in a Th2 stimulated ex vivo human skin assay according to the disclosure described below.

The present invention relates to novel compounds that are JAK modulators useful for the treatment of diseases and conditions associated with dysregulation of JAKs, particularly TYK2/JAK1. The present invention further provides pharmaceutical compositions comprising such JAK enzyme modulators, as well as methods of treating and/or preventing such diseases and conditions.

According to a first aspect of the present invention there is provided a compound of formula (I) or a pharmaceutically acceptable salt thereof:

wherein R ¹ and R ² are each independently hydrogen or hydroxy; wherein R ¹ and R ² are not both hydroxy. A number of embodiments (E) of this first aspect (where E1 is the same for convenience) of the present invention are described below.

E1 A compound of formula I as defined above or a pharmaceutically acceptable salt thereof.

E2. The compound or a pharmaceutically acceptable salt thereof according to E1, wherein R ¹ is hydroxy and R ² is hydrogen.

E3. The compound according to E1, wherein R ¹ is hydrogen and R ² is hydroxy, or a pharmaceutically acceptable salt thereof.

E4. The compound according to E1 of Formula IA: or a pharmaceutically acceptable salt thereof:

.

.

E5. The compound according to E1 of Formula IB: or a pharmaceutically acceptable salt thereof:

.

.

E6. A compound according to E1 of Formula IC: or a pharmaceutically acceptable salt thereof:

.

.

E7. A compound according to E1 having formula ID: or a pharmaceutically acceptable salt thereof:

.

.

E8. The compound according to E1, or a pharmaceutically acceptable salt thereof, selected from the group consisting of:

((S)-2,2-difluorocyclopropyl)((1R,5S)-3-(2-((1-propyl-1H-pyrazol-4-yl)amino)pyrimidin-4-yl )-3,8-diazabicyclo[3.2.1]octan-8-yl)methanone;

((S)-2,2-difluorocyclopropyl)((1R,5S)-3-(2-((1-((R)-2-hydroxypropyl)-1H-pyrazole-4- yl)amino)pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octan-8-yl)methanone;

((S)-2,2-difluorocyclopropyl)((1R,5S)-3-(2-((1-((S)-2-hydroxypropyl)-1H-pyrazole-4- yl)amino)pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octan-8-yl)methanone; and

((S)-2,2-difluorocyclopropyl)((1R,5S)-3-(2-((1-(3-hydroxypropyl)-1H-pyrazol-4-yl)amino) pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octan-8-yl)methanone.

E9. ((S)-2,2-difluorocyclopropyl)((1R,5S)-3-(2-((1-propyl-1H-pyrazol-4-yl)amino)pyrimidin-4-yl )-3,8-diazabicyclo[3.2.1]octan-8-yl)methanone.

E10. ((S)-2,2-difluorocyclopropyl)((1R,5S)-3-(2-((1-((R)-2-hydroxypropyl)-1H-pyrazole-4- yl)amino)pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octan-8-yl)methanone.

E11. ((S)-2,2-difluorocyclopropyl)((1R,5S)-3-(2-((1-(3-hydroxypropyl)-1H-pyrazol-4-yl)amino) pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octan-8-yl)methanone.

E12. ((S)-2,2-difluorocyclopropyl)((1R,5S)-3-(2-((1-((S)-2-hydroxypropyl)-1H-pyrazole-4- yl)amino)pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octan-8-yl)methanone.

E13. The compound according to any one of E1 to E12 in isolated form or a pharmaceutically acceptable salt thereof.

E14. The compound according to any one of E1 to E13 in crystalline form or a pharmaceutically acceptable salt thereof.

E15. A pharmaceutical composition comprising a compound according to any one of E1 to E13 or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.

E16. TYK2/JAK1 inhibitor in a subject in need of treatment for a disease or condition that is an indication for a TYK2/JAK1 inhibitor, comprising administering to the subject a therapeutically effective amount of a compound according to any one of E1 to E14, or a pharmaceutically acceptable salt thereof. A method for treating or preventing a disease or condition that is an indication for a JAK1 inhibitor.

E17. Treating or preventing an inflammatory or autoimmune condition in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a compound according to any one of E1 to E14, or a pharmaceutically acceptable salt thereof. How to treat or prevent.

E18. Inflammation, autoimmune disease, neuroinflammation, arthritis, rheumatoid arthritis, spondyloarthropathy, systemic lupus erythematosus, lupus nephritis, osteoarthritis, gouty arthritis, pain, fever, pulmonary sarcoidosis, silicosis, cardiovascular disease, atherosclerosis, myocardial Infarction, thrombosis, congestive heart failure and cardiac reperfusion injury, cardiomyopathy, stroke, ischemia, reperfusion injury, brain oedema, brain trauma, neurodegeneration, liver disease, inflammatory bowel disease, Crohn's disease, ulcerative colitis, nephritis, retinitis, retina conditions, macular degeneration, glaucoma, diabetes (types 1 and 2), diabetic neuropathy, viral and bacterial infections, myalgia, endotoxin shock, toxic shock syndrome, osteoporosis, multiple sclerosis, endometriosis, menstrual cramps, vaginitis , candidiasis, cancer, fibrosis, obesity, muscular dystrophy, polymyositis, dermatomyositis, autoimmune hepatitis, primary biliary cirrhosis, primary sclerosing cholangitis, vitiligo, Alzheimer's disease, skin flushing, eczema, psoriasis, atopic dermatitis, sunburn, keloids, hypertrophic scars, rheumatic disease, urticaria, discoid lupus, cutaneous lupus, central nervous system lupus, psoriatic arthritis, asthma, allergic asthma, type I interferonopathy such as Ecardi Gutiérrez syndrome and type I interferon Other Mendelian diseases of overexpression, primary progressive multiple sclerosis, relapsing remitting multiple sclerosis, non-alcoholic fatty liver disease, non-alcoholic steatohepatitis, scleroderma, alopecia areata, scarring alopecia, pruritus, prurigo nodosa, CPUO, lichen disease , lichen planus, Steven's Johnson's syndrome, spondylosis, myositis, vasculitis, pemphigus, lupus, major depressive disorder, allergy, dry eye, transplant rejection, cancer, septic shock, cardiorespiratory dysfunction, acute respiratory disease, Ankylosing spondylitis, cachexia, chronic graft-versus-host disease, acute graft-versus-host disease, celiac sprue, idiopathic thrombocytopenic thrombotic purpura, thrombotic thrombocytopenic purpura, myasthenia gravis, Sjogren's syndrome, epidermal hyperplasia, cartilage Inflammation, osteolysis, juvenile arthritis, juvenile rheumatoid arthritis, juvenile rheumatoid arthritis polyarticular, juvenile rheumatoid arthritis polyarticular, systemic onset juvenile rheumatoid arthritis, juvenile ankylosing spondylitis, juvenile enteropathic arthritis, juvenile Reter syndrome, SEA syndrome, juvenile dermatomyositis, juvenile psoriatic arthritis, juvenile scleroderma, juvenile systemic lupus erythematosus, juvenile vasculitis, polyarticular rheumatoid arthritis, polyarticular rheumatoid arthritis, systemic onset rheumatoid arthritis, enteropathic arthritis, reactive arthritis, Letter's syndrome , myositis, polymyositis, dermatomyositis, polyarteritis nodosa, Wegener's granulomatosis, arteritis, polymyalgia rheumatica, sarcoidosis, sclerosis, primary biliary sclerosis, sclerosing cholangitis, dermatitis, Still's disease , chronic obstructive pulmonary disease, Guillain-Barre disease, Graves' disease, Addison's disease, Raynaud's phenomenon, psoriatic epidermal hyperplasia, plaque psoriasis, erythematous psoriasis, Treatment of a disease or condition selected from inverse psoriasis, pustular psoriasis, erythematous psoriasis, immune disorders associated with or resulting from the activation of pathogenic lymphocytes, non-infectious uveitis, Behcet's disease and Vogt-Koyanagi-Harada syndrome Or a method of treating or preventing said disease or condition in a subject in need thereof, comprising administering to a subject in need thereof a therapeutically effective amount of a compound according to any one of E1 to E14, or a pharmaceutically acceptable salt thereof.

E19. The method of any one of E16 to E18, wherein the compound is selected from the group consisting of:

((S)-2,2-difluorocyclopropyl)((1R,5S)-3-(2-((1-propyl-1H-pyrazol-4-yl)amino)pyrimidin-4-yl )-3,8-diazabicyclo[3.2.1]octan-8-yl)methanone;

((S)-2,2-difluorocyclopropyl)((1R,5S)-3-(2-((1-((R)-2-hydroxypropyl)-1H-pyrazole-4- yl)amino)pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octan-8-yl)methanone;

((S)-2,2-difluorocyclopropyl)((1R,5S)-3-(2-((1-((S)-2-hydroxypropyl)-1H-pyrazole-4- yl)amino)pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octan-8-yl)methanone; and

((S)-2,2-difluorocyclopropyl)((1R,5S)-3-(2-((1-(3-hydroxypropyl)-1H-pyrazol-4-yl)amino) pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octan-8-yl)methanone; or a pharmaceutically acceptable salt thereof.

E20. The compound according to any one of E16 to E18, wherein the compound is ((S)-2,2-difluorocyclopropyl)((1R,5S)-3-(2-((1-propyl-1H- pyrazol-4-yl)amino)pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octan-8-yl)methanone or a pharmaceutically acceptable salt thereof.

E21. The compound according to any one of E16 to E18, wherein the compound is in isolated form ((S)-2,2-difluorocyclopropyl)((1R,5S)-3-(2-((1-(3-hydroxy) Roxypropyl)-1H-pyrazol-4-yl)amino)pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octan-8-yl)methanone or a pharmaceutically acceptable salt thereof. .

E22. The method of any one of E16-E19, wherein the disease or condition is psoriasis.

E23. The method of any one of E16-E19, wherein the disease or condition is atopic dermatitis.

E24. The method of any one of E16-E19, wherein the disease or condition is hand eczema.

E25. The method of any one of E16-E19, wherein the disease or condition is pruritus.

E26. The method of any one of E16-E19, wherein the disease or condition is cutaneous lupus.

E27. The method of any one of E16 to E19, wherein the compound is administered topically.

E28. Use of a compound according to any one of E1 to E14, or a pharmaceutically acceptable salt thereof, for the manufacture of a medicament for the treatment of a disorder which is an indication for a TYK2/JAK1 inhibitor.

E29. A compound or a pharmaceutically acceptable salt thereof for use in the treatment of a disorder according to any one of E1 to E14, which is an indication for a TYK2/JAK1 inhibitor.

E30. A pharmaceutical combination comprising an isolated form of a compound of any one of E1 to E14, or a pharmaceutically acceptable salt thereof, or a pharmaceutically acceptable salt thereof, and at least one additional pharmacologically active compound.

E40. The crystalline form of the compound according to E4, having an X-ray powder diffraction pattern comprising diffraction peaks at 4.6, 9.2, 18.5, and 19.6 ± 0.2 degrees 2θ.

In certain embodiments, a therapeutically effective amount used according to the method is from 0.01 mg/kg body weight/day to 100 mg/kg body weight/day. In certain other embodiments, the therapeutically effective amount used in accordance with the methods is a therapeutically effective amount of from 0.1 mg/kg body weight/day to 10 mg/kg body weight/day.

In certain embodiments, the present invention provides a topical pharmaceutical formulation comprising an active agent present at a concentration of about 0.0001% to about 10.0% (w/w). In another embodiment, the active agent is present at a concentration of about 0.001% to about 3.0% (w/w). In another embodiment, the active agent is present at a concentration of about 0.01% to about 3.0% (w/w).

Compounds of the present invention that have the same molecular formula but differ in the nature or order of bonding of their atoms or the arrangement of their atoms in space are termed “isomers”. Isomers that differ in the arrangement of their atoms in space are termed “stereoisomers”. These stereoisomers are "R" or "S" depending on the configuration of the substituents around the chiral carbon atom. As used herein, the terms "R" and "S" refer to IUPAC 1974 Recommendations for Section E, Fundamental Stereochemistry, Pure Appl. Chem., 1976, 45: 13-30]. Enantiomers of the present invention designated by (R), (S) or * are substantially free of other enantiomers. “Substantially free” means having an enantiomeric excess greater than about 90%, preferably greater than about 95%, and more preferably greater than about 99%. With respect to enantiomeric excess, the term “about” means ±1.0%. The symbol * designates a chiral carbon atom as either (R) or (S) stereochemistry, depending on the configuration of the substituents around the chiral carbon atom. The present invention contemplates the various stereoisomers and mixtures thereof specifically included within the scope of the present invention. Stereoisomers include enantiomers and mixtures of enantiomers. Individual stereoisomers of the compounds of this invention can be prepared synthetically from commercially available starting materials containing asymmetric or chiral centers, or by preparation of racemic mixtures followed by resolutions well known to those skilled in the art. can do. These resolution methods include (1) attaching a chiral auxiliary to a mixture of enantiomers, separating the resulting mixture of diastereomers by recrystallization or chromatography, and liberating an optically pure product from the auxiliary, or (2) ) directly separating a mixture of optical enantiomers on a chiral chromatography column. Compounds of the present invention not designated as (R), (S), or * may occur as racemates (i.e., 50% (R) and 50% (S)) or as two species with an excess of one enantiomer. It can exist as a mixture of enantiomers. For example, an enantiomeric mixture is 51% (R) enantiomer and 49% (S) enantiomer or vice versa, or other than a racemic mixture of 50% (R) and 50% (S) ( R) and (S) in any combination.

All isomers (e.g., cis-, trans-, or diastereomers) of the compounds described herein, alone as well as in any mixture, are included within the scope of the described compounds. All these forms, including enantiomers, diastereomers, cis, trans, syn, anti, solvates (including hydrates), tautomers, and mixtures thereof, are included in the described compounds. Stereoisomeric mixtures, eg mixtures of diastereomers, can be separated into their corresponding isomers in a known manner by suitable separation methods. Diastereomeric mixtures can be separated into their individual diastereomers by, for example, fractional crystallization, chromatography, solvent partitioning, and similar procedures. This separation can be carried out at the level of one of the starting compounds or at the compound of formula I, IA, IB, IC or ID itself. Enantiomers may be separated via formation of diastereomeric salts, for example by salt formation with an enantiomerically pure chiral acid, or by chromatography using a chromatography substrate having a chiral ligand, for example HPLC. . The present invention relates to all pharmaceutically acceptable isotopically-labeled formulas I, IA, IB, in which one or more atoms are replaced by an atom having the same atomic number, but an atomic mass or mass number different from the atomic mass or mass number prevalent in nature. A compound of IC or ID or a pharmaceutically acceptable salt thereof.

Examples of isotopes suitable for inclusion in the compounds of the present invention include isotopes of hydrogen such as ² H and ³ H, carbon such as ¹¹ C, ¹³ C and ¹⁴ C, chlorine such as ³⁶ Cl, fluorine such as ¹⁸ F , iodine such as ¹²³ I and ¹²⁵ I, nitrogen such as ¹³ N and ¹⁵ N, oxygen such as ¹⁵ O, ¹⁷ O and ¹⁸ O, phosphorus such as ³² P, and sulfur such as ³⁵ S.

Certain isotopically-labeled compounds of Formulas I, IA, IB, IC or ID, or pharmaceutically acceptable salts thereof, for example compounds incorporating radioactive isotopes, or pharmaceutically acceptable salts thereof, are drugs and/or matrix tissue Useful for distribution studies. The radioactive isotopes tritium, ³ H, and carbon-14, ¹⁴ C, are particularly useful for this purpose in view of their ease of incorporation and convenient means of detection.

Substitution with heavier isotopes, such as deuterium, i.e. ² H, may provide certain therapeutic advantages due to greater metabolic stability, e.g. increased in vivo half-life or reduced dosage requirements, and is thus preferred in some circumstances. can do. Substitution with positron emitting isotopes such as ¹¹ C, ¹⁸ F, ¹⁵ O and ¹³ N may be useful in positron emission tomography (PET) studies to investigate substrate receptor occupancy. Isotopically-labeled compounds of formula I, IA, IB, IC or ID are generally known to those skilled in the art using an appropriate isotopically-labeled reagent in place of the previously used non-labeled reagent. It can be prepared by conventional techniques or by methods similar to those described in the accompanying Examples and Preparation Examples.

In therapeutic use to treat a disorder in a mammal, a compound of the present invention or a pharmaceutical composition thereof may be administered orally, parenterally, topically, rectally, transmucosally or enterally. Parenteral administration includes direct injection that produces a systemic effect or direct injection to the affected site. Topical administration includes treatment of the skin or organs readily accessible by topical application, such as the eye or ear. Transdermal delivery that produces a systemic effect is also included. Rectal administration includes the form of suppositories. Preferred routes of administration are oral and parenteral.

Pharmaceutically acceptable salts of a compound of formula I, IA, IB, IC or ID include acid addition and base salts thereof. Suitable acid addition salts are formed from acids which form non-toxic salts. Examples are acetate, adipate, aspartate, benzoate, besylate, bicarbonate/carbonate, bisulfate/sulfate, borate, camsylate, citrate, cyclamate, edisylate, esylate , formate, fumarate, gluceptate, gluconate, glucuronate, hexafluorophosphate, hybenzate, hydrochloride/chloride, hydrobromide/bromide, hydroiodide/iodide, isethionate , lactate, maleate, maleate, malonate, mesylate, methylsulfate, naphthylate, 2-naphsylate, nicotinate, nitrate, orotate, oxalate, palmitate, pamoate, phosphate /hydrogen phosphate/dihydrogen phosphate, pyroglutamate, saccharate, stearate, succinate, tannate, tartrate, tosylate, trifluoroacetate and xinophoate salts.

Suitable base salts are formed from bases that form non-toxic salts. Examples include aluminum, arginine, benzathine, calcium, choline, diethylamine, diolamine, glycine, lysine, magnesium, meglumine, olamine, potassium, sodium, tromethamine and zinc salts.

Hemi-salts of acids and bases may also be formed, for example hemisulphate and hemicalcium salts. For a review of suitable salts, see Handbook of Pharmaceutical Salts: Properties, Selection, and Use by Stahl and Wermuth (Wiley-VCH, 2002).

A pharmaceutically acceptable salt of a compound of formula I, IA, IB, IC or ID can be prepared by one or more of three methods, respectively: (i) by reacting a compound of formula I, IA, IB, IC or ID with the desired acid; (ii) by removing an acid- or base-labile protecting group from a suitable precursor of a compound of Formula I, IA, IB, IC or ID, or using a suitable acid or base, to prepare a suitable cyclic precursor, for example a lactone or lactam. by ring opening; or (iii) converting one salt of a compound of formula I, IA, IB, IC or ID into another by reaction with a suitable acid or base or by means of a suitable ion exchange column. All three reactions are typically carried out in solution. The resulting salt precipitates out and can be collected by filtration or can be recovered by evaporation of the solvent. The degree of ionization of the resulting salt may vary from fully ionized to nearly non-ionized.

The present invention also includes the following embodiments:

A compound of I, IA, IB, IC or ID, or a pharmaceutically acceptable salt thereof, as defined in any of the embodiments described herein, for use as a medicament;

Inflammation, autoimmune disease, neuroinflammation, arthritis, rheumatoid arthritis, spondyloarthropathy, systemic lupus erythematosus, lupus nephritis, osteoarthritis, gouty arthritis, pain, fever, pulmonary sarcoidosis, silicosis, cardiovascular disease, atherosclerosis, myocardial Infarction, thrombosis, congestive heart failure and cardiac reperfusion injury, cardiomyopathy, stroke, ischemia, reperfusion injury, brain oedema, brain trauma, neurodegeneration, liver disease, inflammatory bowel disease, Crohn's disease, ulcerative colitis, nephritis, retinitis, retina conditions, macular degeneration, glaucoma, diabetes (types 1 and 2), diabetic neuropathy, viral and bacterial infections, myalgia, endotoxin shock, toxic shock syndrome, osteoporosis, multiple sclerosis, endometriosis, menstrual cramps, vaginitis , candidiasis, cancer, fibrosis, obesity, muscular dystrophy, polymyositis, dermatomyositis, autoimmune hepatitis, primary biliary cirrhosis, primary sclerosing cholangitis, vitiligo, Alzheimer's disease, skin flushing, eczema, psoriasis, atopic dermatitis, sunburn, keloids, hypertrophic scars, rheumatic disease, urticaria, discoid lupus, cutaneous lupus, central nervous system lupus, psoriatic arthritis, asthma, allergic asthma, type I interferonopathy such as Ecardi Gutiérrez syndrome and type I interferon Overexpression of other Mendelian diseases, primary progressive multiple sclerosis, relapsing remitting multiple sclerosis, non-alcoholic fatty liver disease, non-alcoholic steatohepatitis, scleroderma, alopecia areata, spondylopathy, myositis, vasculitis, pemphigus, lupus, major depression Disorders, allergy, dry eye, transplant rejection, cancer, septic shock, cardiopulmonary dysfunction, acute respiratory disease, ankylosing spondylitis, cachexia, chronic graft-versus-host disease, acute graft-versus-host disease, celiac sprue, idiopathic Thrombocytopenic thrombotic purpura, myasthenia gravis, Sjogren's syndrome, epidermal hyperplasia, cartilage inflammation, bone decomposition, juvenile arthritis, juvenile rheumatoid arthritis, juvenile rheumatoid arthritis polyarticular, juvenile rheumatoid arthritis polyarticular, systemic onset juvenile rheumatoid arthritis, pediatric Ankylosing spondylitis, juvenile enteropathic arthritis, juvenile Lettert syndrome, SEA syndrome, juvenile dermatomyositis, juvenile psoriatic arthritis, juvenile scleroderma, juvenile systemic lupus erythematosus, juvenile vasculitis, oligoarticular rheumatoid arthritis, polyarticular rheumatoid arthritis, systemic Onset rheumatoid arthritis, enteropathic arthritis, reactive arthritis, Lettert syndrome, myositis, polymyositis, dermatomyositis, polyarteritis nodosa, Wegener's granulomatosis, arteritis, polymyalgia rheumatoid, sarcoidosis, sclerosis, primary biliary sclerosis, sclerosis Cholangitis, dermatitis, Still disease, chronic obstructive pulmonary disease, Guillain-Barre disease, Graves' disease, Addison's disease, Raynaud's phenomenon, psoriatic epidermoplasia, plaque psoriasis, red psoriasis, inverse psoriasis, pustular psoriasis, erythrodermic psoriasis, pathogenic lymphocytes I as defined in any one of the embodiments described herein for use in the treatment of an immune disorder associated with or arising from the activity of, non-infectious uveitis, Behcet's disease or Vogt-Koyanagi-Harada syndrome, a compound of IA, IB, IC or ID or a pharmaceutically acceptable salt thereof;

A therapeutically effective amount of a compound of Formula I, IA, IB, IC or ID, or a pharmaceutically acceptable salt thereof, as defined in any of the embodiments described herein, to a subject in need thereof for treatment of a disease that is an indication for an inhibitor of JAK. A method for treating a disease that is an indication for a JAK inhibitor in the subject, comprising administering;

A compound of Formula I, IA, IB, IC or ID, or a pharmaceutically acceptable compound thereof, as defined in any of the embodiments described herein, for the manufacture of a medicament for the treatment of a disease or condition that is an indication for an inhibitor of JAK. use of salt;

A compound of Formula I, IA, IB, IC or ID, or a pharmaceutically acceptable salt thereof, as defined in any of the embodiments described herein, for use in the treatment of a disease or condition that is an indication for an inhibitor of JAK;

A pharmaceutical for the treatment of a disease or condition that is an indication for an inhibitor of JAK comprising a compound of Formula I, IA, IB, IC or ID, or a pharmaceutically acceptable salt thereof, as defined in any of the embodiments described herein. composition.

The present invention also relates to the use of a compound of formula I, IA, IB, IC or ID, or a pharmaceutically acceptable salt thereof, in combination with another pharmacologically active compound, in particular one of the functionally defined classes or specific compounds listed below. any use, method or composition as defined above. These agents may be administered in the same or separate dosage forms, via the same or different routes of administration, and on the same or different dosing schedules according to standard pharmaceutical practice known to those skilled in the art.

Suitable agents for use in combination therapy with a compound of formula I, IA, IB, IC or ID, or a pharmaceutically acceptable salt thereof, are sulfasalazine, mesalazine, prednisone, azathioprine, infliximab, adali Mumab, belimumab, besertolizumab, natalizumab, vedolizumab, hydrocortisone, budesonide, cyclosporine, tacrolimus, fexofenadine, 6-mercaptopurine, methotrexate, ursodeoxycholic acid, obeticholic acid, antihistamine, Rifampin, prednisone, methotrexate, azathioprine, cyclophosphamide, hydroxychloroquine, mofetil, sodium mycophenolate, tacrolimus, leflunomide, chloroquine and quinacrine, thalidomide, rituxan, NSAIDs, solu medrol, depomedrol and dexamethasone.

Other suitable agents for use in combination therapy with a compound of formula I, IA, IB, IC or ID or a pharmaceutically acceptable salt thereof include: 5-lipoxygenase activating protein (FLAP) antagonists; leukotriene antagonists (LTRA), such as antagonists of LTB ₄ , LTC ₄ , LTD ₄ , LTE ₄ , CysLT ₁ or CysLT ₂ eg montelukast or zafirlukast; histamine receptor antagonists such as histamine type 1 receptor antagonists or histamine type 2 receptor antagonists such as loratidine, fexofenadine, desloratidine, levocetirizine, metapyrylene or cetirizine; α1-adrenoceptor agonists or α2-adrenoceptor agonists such as phenylephrine, methoxamine, oxymetazoline or methylnorephrine; muscarinic M3 receptor antagonists such as tiotropium or ipratropium; dual muscarinic M3 receptor antagonists/β2 agonists; PDE inhibitors such as PDE3 inhibitors, PDE4 inhibitors or PDE5 inhibitors, for example theophylline, sildenafil, vardenafil, tadalafil, ibudilast, cilomilast or roflumilast; sodium cromoglycate or sodium nedocromil; Cyclooxygenase (COX) inhibitors such as non-selective inhibitors (eg, aspirin or ibuprofen) or selective inhibitors (eg, celecoxib or valdecoxib); glucocorticosteroids such as fluticasone, mometasone, dexamethasone, prednisolone, budesonide, ciclesonide or beclamethasone; anti-inflammatory monoclonal antibodies such as infliximab, adalimumab, tanezumab, ranibizumab, bevacizumab or mepolizumab; β2 agonists such as salmeterol, albuterol, salbutamol, fenoterol or formoterol, especially long-acting β2 agonists; integrin antagonists such as natalizumab; adhesion molecule inhibitors such as VLA-4 antagonists; kinin B ₁ or B ₂ receptor antagonists; immunosuppressive agents such as inhibitors of the IgE pathway (eg omalizumab) or inhibitors of cyclosporine; matrix metalloprotease (MMP) inhibitors, such as inhibitors of MMP-9 or MMP-12; tachykinin NK ₁ , NK ₂ or NK ₃ receptor antagonists; protease inhibitors, such as inhibitors of elastase, chymase or cateopsin G; adenosine A _2a receptor agonists; adenosine A _2b receptor antagonists; urokinase inhibitors; dopamine receptor agonists (eg ropinirole), especially dopamine D2 receptor agonists (eg bromocriptine); modulators of the NFκB pathway, such as IKK inhibitors; additional modulators of cytokine signaling pathways such as inhibitors of JAK kinase, syk kinase, p38 kinase, SPHK-1 kinase, Rho kinase, EGF-R or MK-2; mucolytics, mucoactives or antitussives; Antibiotic; antiviral agents; vaccine; chemokines; epithelial sodium channel (ENaC) blockers or epithelial sodium channel (ENaC) inhibitors; nucleotide receptor agonists such as P2Y2 agonists; thromboxane inhibitors; niacin; 5-lipoxygenase (5-LO) inhibitors such as zileuton; attachment factors such as VLAM, ICAM or ELAM; CRTH2 receptor (DP ₂ ) antagonists; prostaglandin D ₂ receptor (DP ₁ ) antagonists; hematopoietic prostaglandin D2 synthase (HPGDS) inhibitors; interferon-β; soluble human TNF receptors such as etanercept; HDAC inhibitors; phosphoinositide 3-kinase gamma (PI3Kγ) inhibitors; phosphoinositide 3-kinase delta (PI3Kδ) inhibitors; CXCR-1 or CXCR-2 receptor antagonists; IRAK-4 inhibitors; and TLR-4 or TLR-9 inhibitors (including pharmaceutically acceptable salts of the specifically named compounds). An agent may be administered with another active agent, wherein the second active agent may be administered orally or topically.

Accordingly, the present invention provides treatment or prevention of a disease, condition or disorder associated with JAK in a subject, such as a human or non-human mammal, comprising administering to the subject an effective amount of one or more compounds described herein. Methods of treating or preventing a disease, condition or disorder associated with JAK are provided. Conditions for which selective targeting of the JAK pathway or modulation of the JAK kinase are considered therapeutically useful are, inter alia, arthritis, asthma, autoimmune diseases, cancers or tumors, diabetes, certain eye diseases, disorders or conditions, inflammation, intestinal inflammation, allergies or conditions. , neurodegenerative diseases, psoriasis and transplant rejection.

One method of carrying out the present invention is to administer a compound of Formula I, IA, IB, IC or ID in the form of a prodrug. Thus, certain derivatives of compounds of Formulas I, IA, IB, IC or ID, which themselves may have little or no pharmacological activity, when administered into or onto the body, for example as a It can be converted to compounds of Formulas I, IA, IB, IC or ID having the desired activity by catalytic cleavage, particularly hydrolytic cleavage catalyzed by esterase or peptidase enzymes. Such derivatives are referred to as “prodrugs”. Additional information on the use of prodrugs can be found in 'Pro-drugs as Novel Delivery Systems', Vol. 14, ACS Symposium Series (T. Higuchi and W. Stella) and 'Bioreversible Carriers in Drug Design', Pergamon Press, 1987 (Ed. E. B. Roche, American Pharmaceutical Association)]. See also Nature Reviews/Drug Discovery, 2008, 7, 355 and Current Opinion in Drug Discovery and Development, 2007, 10, 550.

The prodrugs according to the present invention are described, for example, in 'Design of Prodrugs' by H. Bundgaard (Elsevier, 1985), for example to those skilled in the art as 'pro-Moyer' It can be prepared by substituting the appropriate functional group present in the compound of formula I, IA, IB, IC or ID using a specific moiety known as t'.

Thus, the prodrugs according to the present invention are (a) ester or amide derivatives of a hydroxy group in a compound of formula I, IA, IB, IC or ID; (b) an ester, carbonate, carbamate, phosphate or ether derivative of a hydroxy group in a compound of formula I, IA, IB, IC or ID; (c) an amide, imine, carbamate or amine derivative of an amino group in a compound of formula I, IA, IB, IC or ID; (d) an oxime or imine derivative of a carbonyl group in a compound of formula I, IA, IB, IC or ID.

Some specific examples of prodrugs according to the present invention include:

(i) when the compound of formula I, IA, IB, IC or ID contains a hydroxyl function,

(ii) when the compound of formula I, IA, IB, IC or ID contains an alcohol function (—OH), the hydrogen of the alcohol function of its ester, such as the compound of formula I, IA, IB, IC or ID -CO(C ₁ -C ₈ alkyl) (eg, methylcarbonyl) or an alcohol is esterified with an amino acid;

(iii) when the compound of formula I, IA, IB, IC or ID contains an alcohol function (-OH), its ether, such as the hydrogen of the alcohol function of the compound of formula I, IA, IB, IC or ID (C ₁ -C ₈ alkyl)C(=O)OCH ₂ - or -CH ₂ OP(=O)(OH) ₂ ;

(iv) when the compound of formula I, IA, IB, IC or ID contains an alcohol function (-OH), its phosphate, such as the hydrogen of the alcohol function of the compound of formula I, IA, IB, IC or ID -P(=0)(OH) ₂ or -P(=0)(ONa) ₂ or -P(=0)(0 ^- ) ₂ compound replaced by Ca ²⁺ ;

(v) when the compound of formula I, IA, IB, IC or ID contains a secondary amino function (-NHR, where R ≠ H), its amide, for example, optionally, formulas I, IA , IB, IC or ID, wherein one or both of the hydrogens of the amino functional group are replaced by (C ₁ -C ₁₀ )alkanoyl, -COCH ₂ NH ₂ or the amino group is derivatized with an amino acid;

(vi) when the compound of formula I, IA, IB, IC or ID contains a secondary amino function (-NH ₂ or -NHR where R ≠ H), its amine, for example optionally , wherein one or both of the hydrogens of the amino functional group of the compound of formula I, IA, IB, IC or ID are replaced by -CH ₂ OP(=O)(OH) ₂ .

Reference to a compound of Formula I, IA, IB, IC or ID is intended to include the compound itself and prodrugs thereof. The present invention includes compounds of formula I, IA, IB, IC or ID, as well as pharmaceutically acceptable salts of such compounds.

Also included within the scope of the present invention are active metabolites of compounds of formula I, IA, IB, IC or ID, i.e. compounds formed in vivo, often by oxidation or dealkylation upon administration of the drug. Some examples of metabolites according to the present invention include:

(i) When the compound of formula I, IA, IB, IC or ID contains a methylene group, its hydroxymethylene derivative (-CH ₂ - -> -CHOH):

(ii) when the compound of formula I, IA, IB, IC or ID contains a tertiary amino group, its secondary amino derivative (-NRR' -> -NHR or -NHR'); and

(iii) when the compound of formula I, IA, IB, IC or ID contains a secondary amino group, its primary derivative (-NHR -> -NH ₂ ).

Pharmaceutical composition or formulation

In another embodiment, the invention provides a pharmaceutical composition or formulation comprising a therapeutically effective amount of a compound of the invention and a pharmaceutically acceptable diluent or carrier. The pharmaceutical compositions or formulations of the present invention may be administered to humans and other mammals topically, orally, parenterally, intracisternally, intravaginally, intraperitoneally, buccally, as an oral spray, as a nasal spray, rectally as a suppository, or as a liposome. It can be administered in the form of

A typical pharmaceutical composition or formulation is prepared by mixing a compound of the present invention and a carrier or diluent. Suitable carriers and diluents include materials such as carbohydrates, waxes, water soluble and/or swellable polymers, hydrophilic or hydrophobic materials, gelatin, oils, solvents, water and the like. The particular carrier or diluent employed will depend on the means and purpose for which the compounds of this invention are applied. Suitable aqueous solvents include water, ethanol, propylene glycol, polyethylene glycol (eg PEG400, PEG300) and the like and mixtures thereof. The formulation may also contain one or more buffers, stabilizers, surfactants, wetting agents, lubricants, emulsifiers, suspending agents, preservatives, antioxidants, opacifiers, glidants, processing aids, colorants, sweeteners, perfumes, flavoring agents, and other known The inclusion of excipients may provide an elegant formulation of a drug (ie, a compound of the present invention or a pharmaceutical composition thereof) or aid in the manufacture of a pharmaceutical product (ie, use in the manufacture of a medicament).

Formulations can be prepared using conventional dissolving and mixing procedures. For example, bulk drug substance (i.e., a compound of the present invention or a stabilized form of a compound (eg, a complex with a cyclodextrin derivative or other known complexing agent)) in the presence of one or more of the above-described suitable carriers. dissolve in solvent. The dissolution rate of poorly water-soluble compounds can be measured in spray-dried dispersions, such as Takeuchi, H., et al., in "Enhancement of the dissolution rate of a poorly water-soluble drug (tolbutamide) by a spray-drying solvent deposition method and disintegrants," J. Pharm. Pharmacol., 39, 769-773 (1987)]; and EP0901786 B1 (US2002/009494), incorporated herein by reference. The compounds of the present invention are typically formulated in pharmaceutical dosage forms to provide easily controllable dosages of the drug and to provide patients with an elegant and easily handled product.

Pharmaceutical compositions or preparations for application may be packaged in a variety of ways depending on the method used to administer the drug. Generally, an article for dispensing includes a container having placed therein a pharmaceutical formulation in an appropriate form. Suitable containers include materials such as bottles (plastic and glass), sachets, ampoules, plastic bags, metal cylinders, and the like. The container may also include a tamper-proof assembly to prevent indiscriminate access to the contents of the package. Additionally, a label describing the contents of the container is placed on the container. Such labels may also include appropriate warning statements.

The term “pharmaceutically acceptable carrier” refers to a carrier medium that provides adequate delivery of an effective amount of an active agent as defined herein, does not interfere with the effectiveness of the biological activity of the active agent, and is sufficiently non-toxic to the host or patient. Representative carriers include water, oils, both vegetable and mineral, cream bases, lotion bases, ointment bases, and the like. These bases include suspending agents, thickening agents, penetration enhancers, and the like. Additional information regarding carriers can be found in Remington: The Science and Practice of Pharmacy, 21st Ed., Lippincott, Williams & Wilkins (2005), incorporated herein by reference. Additional examples of materials that can serve as pharmaceutically acceptable carriers include sugars such as lactose, glucose and sucrose; starches such as corn starch and potato starch; cellulose and its derivatives, such as sodium carboxymethyl cellulose, ethyl cellulose and cellulose acetate; powdered tragacanth; malt; gelatin; talc; carriers such as cocoa butter and suppository wax; oils such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil and soybean oil; glycols such as propylene glycol; esters such as ethyl oleate and ethyl laurate; agar; buffers such as magnesium hydroxide and aluminum hydroxide; alginic acid; pyrogen-free water; isotonic saline; Ringer's solution; Ethyl alcohol and phosphate buffers, as well as other non-toxic compatible lubricants such as sodium lauryl sulfate and magnesium stearate, as well as colorants, release agents, coating agents, sweetening, flavoring and perfumeing agents, preservatives and antioxidants are also used by the manufacturer. may be present in the composition at the discretion of

The term "pharmaceutically acceptable topical carrier" refers to a pharmaceutically acceptable carrier as described herein above that is suitable for topical application. An inert liquid or cream vehicle capable of suspending or dissolving the active agent(s) and having the properties of being non-toxic and non-inflammatory when applied to the skin, fingernails, hair, claws or hooves is an example of a pharmaceutically acceptable topical carrier. . This term is also intended to encompass carrier materials specifically approved for use in topical cosmetics.

The term "topical administration" refers to the application of a pharmaceutical agent to the external surface of the skin, fingernails, hair, claws or hoofs such that the agent passes through the external surface of the skin, fingernails, hair, claws or hoofs and enters the underlying tissue. . Topical administration includes application of the composition to intact skin, fingernails, hair, toenails or hooves, or to broken, peeling or open wounds of the skin, fingernails, hair, toenails or hooves. Topical administration of pharmaceutical agents may result in limited distribution of the agent to the skin and surrounding tissues, or systemic distribution of the agent when the agent is removed from the treatment site by the bloodstream.

Dosage forms for topical or transdermal administration of a compound of this invention include ointments, pastes, creams, lotions, gels, powders, solutions, sprays, inhalants or patches. The active ingredient is admixed under sterile conditions with a pharmaceutically acceptable carrier and any necessary preservatives or buffers as may be required. Volatile compounds may require special formulations or mixtures with special packaging materials to ensure proper dosage delivery. In addition, compounds of the present invention with poor human skin permeability may require one or more permeation enhancers, while compounds that are rapidly absorbed through the skin may require absorption-retarding agents or formulations with barriers.

Ointments, pastes, creams, lotions, gels, powders, and solutions for topical administration may contain, in addition to the active compound of the present invention, a pharmaceutically acceptable carrier such as animal and vegetable fats, oils, waxes, paraffins, starches, tragacanth. , cellulose derivatives, polyethylene glycol, silicone, bentonite, silicic acid, talc, zinc oxide, preservatives, antioxidants, fragrances, emulsifiers, dyes, inert fillers, antiirritants, tackifiers, fragrances, opacifiers, antioxidants, gelling agents, stabilizers , surfactants, emollients, colorants, preservatives, buffers, permeation enhancers, or mixtures thereof. Topical carriers should not interfere with the effectiveness of the biological activity of the active agent and should not be detrimental to epithelial cells or their function.

The term "permeation enhancer" or "permeation enhancer" relates to an object intended to increase the permeability of the skin, fingernails, hair, claws or hooves to a drug, thereby increasing the rate at which a drug penetrates through the skin, fingernails, hair, claws or hooves. do. Enhanced permeation obtained through the use of such enhancers can be observed, for example, by measuring the rate of diffusion of the drug through animal or human skin, fingernails, hair, claws or hooves using a diffusion cell device. Diffusion cells are described in Merritt et al. Diffusion Apparatus for Skin Penetration, J of Controlled Release, 1 (1984) pp. 161-162]. The term “penetration enhancer” or “penetration enhancer” refers to an agent or mixture of agents that, alone or in combination, acts to increase the permeability of the skin, nails, hair or hooves to a drug.

The term “transdermal delivery” refers to the diffusion of an agent across the barriers of the skin, fingernails, hair, toenails or hooves resulting from topical administration or other application of the composition. The stratum corneum acts as a barrier, and few pharmaceutical agents are able to penetrate intact skin. In contrast, the epidermis and dermis are permeable to many solutes, so absorption of drugs that abrade or otherwise strip the stratum corneum to expose the epidermis occurs more readily through the skin, fingernails, hair, claws, or hooves. Transdermal delivery includes injection or other delivery through the skin, nails, hair, toenails or hooves or any part of the mucous membranes and absorption or permeation through the remainder. Absorption through intact skin, fingernails, hair, claws or hooves can be enhanced by incorporating the active agent into an appropriate pharmaceutically acceptable vehicle prior to application to the skin, fingernails, hair, claws or hooves. Passive topical administration may consist of direct application of the active agent to the treatment site in combination with an emollient or penetration enhancer. Transdermal delivery, as used herein, is intended to include delivery by permeation through or through the epidermis, ie the skin, fingernails, hair, claws or hoofs.

Powders and sprays may contain, in addition to the compound of the present invention, lactose, talc, silicic acid, aluminum hydroxide, calcium silicate and polyamide powder, or mixtures of these substances. Sprays may also contain customary propellants, such as chlorofluorohydrocarbons.

Solid dosage forms for oral administration include capsules, tablets, pills, powders and granules. In such solid dosage forms, the active compound may be added with one or more inert, pharmaceutically acceptable carriers such as sodium citrate or calcium phosphate, and/or a) fillers or extenders such as starch, lactose, sucrose, glucose, mannitol and salicylic acid; b) binders such as carboxymethylcellulose, alginates, gelatin, polyvinyl pyrrolidinone, sucrose and acacia; c) humectants such as glycerol; d) disintegrants such as agar-agar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates and sodium carbonate; e) dissolution retardants such as paraffin; f) absorption accelerators such as quaternary ammonium compounds; g) humectants such as cetyl alcohol and glycerol monostearate; h) absorbents such as kaolin and bentonite clay; and i) lubricants such as talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate, and mixtures thereof. In the case of capsules, tablets and pills, the dosage form may also include a buffering agent.

Solid compositions of a similar type may also be employed as fillers in soft and hard-filled gelatin capsules using lactose or milk sugar, as well as high molecular weight polyethylene glycols and the like.

Solid dosage forms of tablets, dragees, capsules, pills, and granules can be prepared with coatings and shells such as enteric coatings and other coatings well known in the pharmaceutical formulation art. They may optionally contain opacifying agents and may also be of a composition that they release the active ingredient(s) alone or preferentially in a specific part of the intestinal tract in a delayed manner. Examples of embedding compositions that can be used include polymeric materials and waxes.

Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, microemulsions, solutions, suspensions, syrups and elixirs. In addition to the active compound, liquid dosage forms may contain inert diluents commonly used in the art, such as for example water or other solvents, solubilizers and emulsifiers such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl Alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol, dimethylformamide, oils (especially cottonseed, peanut, corn, germ, olive, castor and sesame oils), glycerol, tetrahydrofurfuryl alcohol, polyethylene fatty acid esters of glycols and sorbitan, and mixtures thereof.

Besides inert diluents, the oral compositions may also contain adjuvants such as wetting, emulsifying and suspending agents, sweetening, flavoring and perfuming agents.

As used herein, the term "parenterally" refers to modes of administration including intravenous, intramuscular, intraperitoneal, intrasternal, subcutaneous, intraarticular injection and infusion. Pharmaceutical compositions of the present invention for parenteral injection include pharmaceutically acceptable sterile aqueous or non-aqueous solutions, dispersions, suspensions or emulsions, and sterile powders for reconstitution into sterile injectable solutions or dispersions. Examples of suitable aqueous and non-aqueous carriers, diluents, solvents or vehicles include water, ethanol, polyols (propylene glycol, polyethylene glycol, glycerol and the like), suitable mixtures thereof, vegetable oils (such as olive oil) and injectable organic esters such as ethyl Contains oleate. Adequate fluidity can be maintained, for example, by the use of coatings, such as lecithin, by maintenance of the required particle size in the case of dispersions, and also by the use of surfactants.

Injectable depot forms are prepared by forming microencapsulated matrices of the drug in biodegradable polymers such as polylactide-polyglycolide. Depending on the ratio of drug to polymer and the nature of the particular polymer used, the rate of drug release can be controlled. Examples of other biodegradable polymers include poly(orthoesters) and poly(anhydrides), and depot injectable formulations are also prepared by entrapping the drug in liposomes or microemulsions that are compatible with body tissues.

Injectable preparations can be sterilized, for example, by filtration through a bacteria-retaining filter, or by incorporating a sterilizing agent in the form of a sterile solid composition which can be dissolved or dispersed in sterile water or other sterile injectable medium immediately prior to use. there is.

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

Pharmaceutical compositions or preparations for rectal or vaginal administration are preferably suppositories, which may be prepared by mixing a compound of the present invention with a suitable non-irritating carrier such as cocoa butter, polyethylene glycol or suppository wax, which is solid at ambient temperature but It is liquid at body temperature and therefore melts in the rectum or vaginal cavity to release the active compound.

The compounds of the present invention may also be administered in the form of liposomes. Liposomes are generally derived from phospholipids or other lipid substances and are formed by mono- or multi-layered hydrated liquid crystals dispersed in an aqueous medium. Any non-toxic, physiologically acceptable, metabolizable lipid capable of forming liposomes may be used. The composition of the present invention in the form of a liposome may contain a stabilizer, a preservative, an excipient and the like in addition to the compound of the present invention. Preferred lipids are natural and synthetic phospholipids and phosphatidylcholine (lecithin), used individually or together. Methods for forming liposomes are known in the art. See, eg, Prescott, Ed., Methods in Cell Biology, Volume XIV, Academic Press, New York, N. Y. (1976), p 33 et seq.

The pharmaceutical compositions or preparations of the present invention may also contain adjuvants such as preservatives, wetting agents, emulsifying agents and dispersing agents. Prevention of the action of microorganisms can be ensured by various antibacterial and antifungal agents, such as parabens, chlorobutanol, phenol, sorbic acid and the like. It may also be desirable to include isotonic agents such as sugars, sodium chloride, and the like. Prolonged absorption of the injectable pharmaceutical form can be brought about using agents which delay absorption, for example, aluminum monostearate and gelatin.

A pharmaceutical composition or formulation of the present invention may be a suspension. Suspensions may contain, in addition to the active compounds, for example, as ethoxylated isostearyl alcohols, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum metahydroxide, bentonite, agar-agar and tragacanth, and mixtures thereof. May contain suspending agents.

Pharmaceutical compositions also include solvates and hydrates of the compounds of the present invention. The term “solvate” refers to a molecular complex of a compound represented by Formula I, IA, IB, IC or ID, including pharmaceutically acceptable salts thereof, and one or more solvent molecules. Such solvent molecules are those commonly used in the pharmaceutical arts known to be harmless to recipients, for example water, ethanol, ethylene glycol, (S)-propylene glycol, (R)-propylene glycol, and the like, and the term "hydrate" refers to Refers to a complex in which the solvent molecule is water. Solvates and/or hydrates are preferably in crystalline form. Other solvents may be used as intermediate solvates in the preparation of more preferred solvates. Intermediate solvents include, but are not limited to, methanol, methyl t-butyl ether, ethyl acetate, methyl acetate, 1,4-butyne-diol, and the like.

Actual dosage levels of active ingredients in pharmaceutical compositions of the present invention may vary in order to obtain an amount of active compound effective to achieve the desired therapeutic response for a particular patient, composition, and mode of administration. The dosage level selected will depend on the activity of the particular compound, the route of administration, the severity of the condition being treated, and the condition and medical history of the patient being treated. However, it is within the skill of the art to start with a dose of the compound at a level lower than that required to achieve the desired therapeutic effect and gradually increase the dosage until the desired effect is achieved.

The total daily dose of the compounds of the present invention administered to humans or lower animals may range from about 0.000001 to about 10 mg/kg/day. For oral administration, a more preferred dosage may range from about 0.001 to about 1 mg/kg/day. For topical administration, a more preferred dosage may range from 0.00001 mg/kg/day to about 5 mg/kg/day. If necessary, the effective daily dose can be divided into multiple doses, for example 2 to 4 individual doses per day, depending on the purpose of administration.

synthesis method

The following reaction schemes and written descriptions provide general details regarding the preparation of the compounds of this invention. The compounds of the present invention can be prepared by any method known in the art for the preparation of compounds of similar structure. In particular, the compounds of the present invention can be prepared by the procedures described with reference to the reaction schemes below, or by the specific methods described in the Examples, or by methods analogous to any of these.

The skilled person will understand that the experimental conditions set forth in the schemes below are illustrative of suitable conditions for carrying out the transformations presented, and that it may be necessary or desirable to vary the precise conditions used in the preparation of compounds of formula I, IA, IB, IC or ID. You will realize that you can.

In addition, those skilled in the art will recognize that it may be necessary or desirable to protect one or more sensitive groups at any stage of the synthesis of the compounds of the present invention, so as to avoid undesirable side reactions. In particular, it may be necessary or desirable to protect amino or carboxylic acid groups. The protecting groups used in the preparation of the compounds of the present invention can be used in a conventional manner. For example, Greene's Protective Groups in Organic Synthesis' by Theodora W Greene and Peter G M Wuts, third edition, (John Wiley and Sons, 1999), incorporated herein by reference, also describes methods for removing such groups. , especially chapters 7 ("Protection for the Amino Group") and 5 ("Protection for the Carboxyl Group").

All derivatives of formula I can be prepared by the procedures described in the general methods given below or by commercial variations thereof. The present invention also encompasses any one or more of these methods for preparing derivatives of formula I, IA, IB, IC or ID, as well as any novel intermediates used therein. A person skilled in the art will recognize that the reaction below can be heated either thermally or under microwave irradiation. It will also be appreciated that it may be necessary or desirable to perform the transformations in a different order than those depicted in the schemes, or to modify one or more of the transformations to provide the desired compounds of the present invention.

Those skilled in the art will also recognize that some compounds of the present invention are chiral and can therefore be prepared as racemic or non-racemic mixtures of enantiomers. Several methods for the separation of enantiomers are available and are well known to those skilled in the art. A preferred method for commercial separation of enantiomers is supercritical fluid chromatography using a chiral stationary phase.

Compounds of formula I, IA, IB, IC or ID are compounds A-1, A-2 and C-3 (A=n-propyl, 1-hydroxypropyl or 2-hydroxy as illustrated by Scheme A). propyl, or a protected form thereof). Compounds of formulas A-1, A-2 and C-3 are commercially available or can be synthesized by those skilled in the art according to the literature or preparation examples described herein. For this purpose, PG is a protecting group, typically tert-butoxycarbonyl. Compounds of formula A-3 can be prepared from compounds of formulas A-1 and A-2 according to process step (i), which is an aromatic substitution reaction in the presence of an organic base. Preferred conditions include triethylamine in methanol at 0° C. to room temperature. The compound of Formula A-5 undergoes process steps (ii) and (iii), under Buchwald-Hartwig cross-coupling conditions or by an acid and elevated temperature, to induce nucleophilicity with a compound of Formula C-3. It can be prepared from compounds of formula A-3 by a substitution reaction followed by a deprotection reaction mediated by an inorganic or organic acid. Typical Buchwald-Hartwig conditions include a suitable palladium catalyst, a suitable chelating phosphine ligand and an inorganic base in a suitable organic solvent at elevated temperature, either thermally or under microwave irradiation. Preferred conditions are a) sodium tert-butoxide with palladium(II) acetate and 2-dicyclohexylphosphino-2',4',6'-triisopropylbiphenyl or Xantphos ^™ , b) at 120-140°C. potassium phosphate or cesium carbonate in DMA under microwave irradiation, or c) BrettPhos ^™ Pd G3 at 40° C. with cesium carbonate as base and DMA or dioxane as solvent. Typical acidic conditions include a suitable inorganic acid in a suitable alcoholic solvent at elevated temperature, either thermally or under microwave irradiation. Preferred conditions include concentrated hydrochloric acid in iso-propanol at 140° C. under microwave irradiation. Alternatively, deprotection takes place in situ during process step (ii). A compound of formula A-6 (A=n-propyl, 1-hydroxypropyl or 2-hydroxypropyl, or a protected form thereof) is prepared in process step (iv) of formula BC(O)X, wherein X is chloro; hydroxy, a suitable leaving group, or an anhydride (which may be, for example, (S)-2,2-difluorocyclopropane-1-carboxylic acid), followed by an amide bond forming reaction with a compound of formula A- It can be prepared from the compound of 5. When the compound of formula BC(O)X is an acid chloride, preferred conditions include triethylamine in dichloromethane at room temperature. When the compound of formula BC(O)X is a carboxylic acid, activation of the carboxylic acid using a suitable organic base and a suitable coupling agent is employed. Preferred conditions include DIPEA or triethylamine with HATU in dichloromethane or DMF at room temperature.

Scheme A

Alternatively, compounds of Formulas I, IA, IB, IC or ID can be prepared from compounds A-3 and C-3 as illustrated by Scheme B. Compounds of Formula A-3 are prepared as described in Scheme A. Compounds of formula C-3 are commercially available or can be synthesized by those skilled in the art according to the literature or preparation examples described herein. Compounds of formula B-1 may be prepared from compounds of formula A-3 by process step (i) deprotection mediated by an inorganic or organic acid in a suitable organic solvent. Preferred conditions include hydrochloric acid or TFA in dioxane or DCM. Compounds of formula B-2 can be prepared from compounds of formula B-1 and BC(O)X according to the amide bond forming reaction as described in process step (ii), Scheme A. The compound of formula A-6 is subjected to a nucleophilic substitution reaction with the compound of formula C-3 mediated by an acid and high temperature or under Buchwald-Hartwig cross-coupling conditions as described in process step (iii), Scheme A. It can be prepared from the compound of formula B-2 according to.

Scheme B

The compounds of Formula C-3 used in Scheme A and Scheme B can be prepared from compounds of Formula C-1 as illustrated in Scheme C. Compounds of formula C-1 are commercially available or can be synthesized by those skilled in the art according to the literature or preparation examples described herein. A compound of formula C-2 is prepared by process steps (i) an alkylation reaction with an appropriately substituted alkyl halide of formula AX, wherein X is Cl, Br or I, in the presence of an inorganic or organic base and a solvent such as DMF; or from compounds of formula C-1 by addition reaction to an epoxide in the presence of an inorganic or organic base. A compound of formula C-3 may be prepared according to process step (ii) from a compound of formula C-2, wherein the reduction is typically carried out with a metal catalyst such as palladium or nickel, hydrogen gas at a pressure of 1 - 50 atm, and protic It is performed in the presence of a solvent such as methanol.

Scheme C

Preparation and Examples

The following non-limiting preparations and examples illustrate the preparation of compounds and salts of the present invention. In the examples and preparations given below, and in the reaction schemes mentioned above, the following abbreviations, definitions and analytical procedures may be mentioned. Other abbreviations common in the art may also be used. The compounds of the present invention are named using Chemdraw Professional ^™ version 18.0 (Perkin Elmer) or given names consistent with IUPAC nomenclature.

^{The 1} H nuclear magnetic resonance (NMR) spectra were consistent with the proposed structures in all cases. Characteristic chemical shifts (δ) are given in parts per million downfield from tetramethylsilane using the following common abbreviations for the names of the main peaks: eg s, singlet; d, doublet; t, triplet; q, quartet; m, multiplet; br, wide. For common NMR solvents the following abbreviations have been used: CD ₃ CN, deuteroacetonitrile; CDCl ₃ , deuterochloroform; DMSO-d ₆ , deuterodimethylsulfoxide; and CD ₃ OD, deuteromethanol. Where appropriate, tautomers can be reported within the NMR data; Some exchangeable protons may not be visible. Since the isolate is a mixture of two conformers, some resonances in the NMR spectrum appear as complex multiplets.

Mass spectra were recorded using electron impact ionization (EI), electrospray ionization (ESI) or atmospheric pressure chemical ionization (APCI). The observed ions are reported as MS m/z, and can be compound [M] ⁺ , compound + proton [MH] ⁺ , or compound + cation of sodium ion [MNa] ⁺ . In some cases, the only observed ion may be a fragment ion reported as [MH-(fragment loss)] ⁺ . Where appropriate, reported ions are assigned to isotopes of chlorine ( ³⁵ Cl and/or ³⁷ Cl), bromine ( ⁷⁹ Br and/or ⁸¹ Br) and tin ( ¹²⁰ Sn).

When TLC, chromatography or HPLC is used to purify a compound, a person skilled in the art can select any suitable solvent or combination of solvents to purify the target compound. Chromatographic separations (except HPLC) were performed using silica gel adsorbents unless otherwise indicated.

All reactions were performed using continuous stirring under an atmosphere of nitrogen or argon gas unless otherwise indicated. In some cases, the reaction was purged with nitrogen or argon gas before starting the reaction. In these cases, nitrogen or argon gas was bubbled through the liquid phase of the mixture for approximately the specified time. The solvents used were of commercial anhydrous grade. All starting materials were commercially available products. In some cases, starting materials were prepared according to reported literature procedures. As used herein, it will be clear to those skilled in the art that the term "concentrated" generally refers to the practice of solvent evaporation under reduced pressure, typically achieved by use of a rotary evaporator.

The following abbreviations are used herein:

ACN: acetonitrile;

ATM: atmospheric pressure

BrettPhos ^™ Pd G3: [(2-di-cyclohexylphosphino-3,6-dimethoxy-2',4',6'-triisopropyl-1,1'-biphenyl)-2-(2'-amino-1,1'-biphenyl)] palladium (II) methanesulfonate;

c: concentration

CDI: 1,1'-carbonyldiimidazole;

Cs ₂ CO ₃ : cesium carbonate;

DCM: dichloromethane

DIPEA: N,N-diisopropylethylamine;

DMA: N,N-dimethylacetamide;

DMF: N,N-dimethylformamide;

ESI: electrospray ionization;

EtOAc: ethyl acetate;

HATU: N-[(dimethylamino)-1H-1,2,3-triazolo-[4,5-b]pyridin-1-ylmethylene]-N-methylmethanaminium hexafluorophosphate N-oxide ;

HCl; Hydrochloric acid;

HPLC: high pressure liquid chromatography;

HRMS: high resolution mass spectrum;

H ₂ SO ₄ ; sulfuric acid;

kg: kilogram

KOH: potassium hydroxide;

MeOH: methanol;

MIBK: methyl isobutyl ketone;

mg: milligram;

mL: milliliters;

mmol: millimoles;

Mpa: megapascal;

MTBE: methyl tert-butyl ether;

NMT: hereinafter;

Pd/C: palladium on carbon;

TBAB: tetrabutylammonium bromide;

TFA: trifluoroacetic acid;

THF: tetrahydrofuran;

TLC: thin layer chromatography;

T3P: 2,4,6-tripropyl-1,3,5,2,4,6-trioxatriphosphorinane-2,4,6-trioxide;

Xantphos: 4,5-bis(diphenylphosphino)-9,9-dimethylxanthene.

Preparation Example 1. (S)-2,2-difluorocyclopropane-1-carboxylic acid 2,2',2"-nitrilotris(ethane-1-ol) salt.

To a 100 mL reactor was added ACN (50.0 mL) and triethanolamine (12.2 g, 1.0 eq). This solution was heated to 45° C. and (S)-2,2-difluorocyclopropane-1 prepared as described in preparation 68 of U.S. Pat. No. 9,663,526 in MTBE (50.0 mL, ~20% w/w). - A premixed solution of carboxylic acid (10.1 g, 1.0 eq) was added dropwise over 100 minutes. After addition, the reaction was held at 45° C. for 30 min and then cooled to 20° C. at a rate of 0.25° C./min. The mixture was granulated for 30 minutes, then filtered, washed with MTBE (40.0 mL) and dried under vacuum at 50°C. ¹ H NMR (400 MHz, DMSO-d ₆ ) δ 6.85 (s, 4H), 3.61 (t, J = 5.7 Hz, 6H), 2.97 (t, J = 5.7 Hz, 6H), 2.38 (ddd, J = 15.4, 10.8, 7.9 Hz, 1H), 1.84 - 1.62 (m, 2H) ¹³ C NMR (101 MHz, DMSO-d ₆ ) δ 169.0, 115.9, 113.1 (dd, J = 285.9, 281.2 Hz), 113.1, 110.3 , 57.4, 56.5, 27.7, 27.6 (dd, J = 12.0, 9.2 Hz), 27.6, 27.5, 16.2, 16.1 (t, J = 9.8 Hz), 16.0. mp: 82.4°C.

Preparation Example 2. 2,2-difluorocyclopropane-1-(S)-carboxylate (R)-N-benzyl-1-phenylethane-1-aminium salt

In a 250 mL vessel, MTBE (134 mL), (S)-2,2-difluorocyclopropane-1-carboxylic acid 2,2',2"-nitrilotris(ethane-1-ol) salt (20.0 g, 1.0 equiv.), and a premixed solution of sulfuric acid (4.3 mL, 1.1 equiv.) in water (86.0 mL) was added. The mixture was stirred until all solids dissolved, then allowed the layer to settle. Layer was separated, and the lower (aqueous) layer was back extracted with MTBE (58 mL) The combined organic layers were dried via azeotropic distillation to obtain ~15% (w/w) of (S)-2,2-di in MTBE To achieve final concentration of fluorocyclopropane-1-carboxylic acid, chiral amine (R)-(+)-N-benzyl-α-methylbenzylamine (13.0 g, 0.85 equiv) was added to the solution for ~1 hour. After addition of ~25% of the amine, the reaction was added dropwise over (R)-N-benzyl-1-phenylethan-1-amine (S)-2,2-difluorocyclopropane. -1-carboxylate (50 mg, 0.002 eq) was seeded in. After adding the amine, the slurry was allowed to granulate, then filtered and washed with MTBE (12.0 mL) pre-chilled to 10°C, The solid was dried under vacuum at 50° C. The crude solid (10.57 g) was returned to the same container, and ACN (35.0 mL) was added. The slurry was heated to 80° C. to completely dissolve the solid. The solution was 0.2° C./ml. min at 22° C. and granulated. The product was collected by filtration, washed with ACN (13.0 mL) and dried under vacuum at 50° C. ¹ H NMR (400 MHz, DMSO-d ₆ ) δ 9.25 (s, 2H), 7.46 (d, J = 6.9 Hz, 2H), 7.43 - 7.24 (m, 9H), 4.00 (q, J = 6.7 Hz, 1H), 3.74 (d, J = 13.3 Hz, 1H), 3.65 (s, 1H), 2.50 - 2.39 (m, 1H), 1.90 - 1.66 (m, 2H), 1.43 (d, J = 6.7 Hz, 3H) ¹³ C NMR (101 MHz, DMSO -d ₆ ) δ 168.5, 142.4, 137.1, 129.3, 129.0, 128.7, 128.0, 127.9, 127.6, 116.0, 113.2, 113.1 (dd, J = 286.1, 281.3 Hz), 110.3, 57 .2, 49.8, 27.6, 27.5, 27.5 (dd, J = 12.0, 9.3 Hz), 27.4, 22.5, 16.2, 16.1 (t, J = 9.8 Hz), 16.07. mp: 138.6°C.

Example 1

((S)-2,2-difluorocyclopropyl)((1R,5S)-3-(2-((1-propyl-1H-pyrazol-4-yl)amino)pyrimidin-4-yl )-3,8-diazabicyclo[3.2.1]octan-8-yl)methanone (IA).

Step 1A. 4-Nitro-1-propyl-1H-pyrazole. This step was performed in two parallel batches. Potassium carbonate (1.38 g, 9.99 mmol) was added in one portion to a 25° C. stirring mixture of 4-nitro-1H-pyrazole (360 g, 3.18 mol) in ACN (3 L) and the mixture was brought to about 50-60° C. heated with 1-Iodopropane (595.3 g, 3.5 mol) was added to the mixture over a period of about 20 minutes. The resulting reaction mixture was stirred at 50-60° C. for about 2 hours. The reaction mixture was cooled to about 25 °C. The two batches of the reaction mixture were combined, diluted with water (3 L), and concentrated under reduced pressure to remove most of the ACN. The resulting aqueous layer was extracted with EtOAc (4 L, 3 x 2 L). The combined organic extracts were dried over sodium sulfate and filtered. The filtrate was concentrated under reduced pressure to give 980.0 g (99%) of the title compound as an oil. ¹ H NMR (400 MHz, CDCl ₃ ) δ 8.15 (s, 1H), 7.99 (s, 1H), 4.08 (t, J = 7.2 Hz, 2H), 1.92-1.83 (m, 2H), 0.88 (t, J = 7.2 Hz, 2H).

Step 1B. 1-propyl-1H-pyrazol-4-amine hydrochloride. Pd/C (10% Pd, 71.06 g, 66.77 mmol) was added to a mixture of 4-nitro-1-propyl-1H-pyrazole (1.04 kg, 9.93 mol) in MeOH (5 L). The mixture was degassed and backfilled with argon gas (3x). The mixture was warmed to 35° C. and stirred for about 48 hours at about 2 Mpa under a hydrogen gas atmosphere. Hydrogen gas was refilled as needed to maintain the pressure at about 2 Mpa. The reaction mixture was filtered. The filtrate was concentrated to a solution (2.5 kg). The solution was added portionwise at approximately 5-10° C. to a solution of HCl in MeOH (5.99 mol, 2.35 L) and stirred for about 1 hour. The mixture was concentrated under reduced pressure to give a slurry (~1 L). EtOAc (500 mL) was added and the mixture was stirred for about 5 minutes. The mixture was filtered and the filter cake was washed with EtOAc (4 x 500 mL). The filter cake was dried under reduced pressure to give 550 g of the title compound as a solid. MS m/z 126.1 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO-d ₆ ) δ 10.35 (br s, 3H), 7.94 (s, 1H), 7.51 (s, 1H), 4.05 (t, J = 8.0 Hz, 2H), 1.78-1.69 (m, 2H), 0.79 (t, J = 8.0 Hz, 3H).

Step 1C. tert-Butyl (1R,5S)-3-(2-chloropyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]-octane-8-carboxylate. The title compound was purchased from STA Pharmaceutical Hong Kong Limited.

Step 1D. 4-((1R,5S)-3,8-diazabicyclo[3.2.1]octan-3-yl)-N-(1-propyl-1H-pyrazol-4-yl)pyrimidin-2-amine . In a 1 L OptiMax ^™ reaction vessel charged with 50 mL isopropanol and 375 mL water, tert-butyl (1R,5S)-3-(2-chloropyrimidin-4-yl)-3,8-diazabi Cyclo[3.2.1]octane-8-carboxylate (50.00 g, 153.9 mmol) and 1-propyl-1H-pyrazol-4-amine hydrochloride (49.76 g, 1.20 eq, 184.7 mmol) were added as a solid . The suspension was stirred at 65°C for about 2 hours and then cooled to 45°C. A 50% KOH aqueous solution (77 mL, 6.0 eq) was added to the mixture and stirred at 35° C. for about 20 hours. The suspension was then cooled to 15° C. and the solid was collected by filtration to yield 43.51 g (90%) of the title compound as an off-white solid. MS m/z 314 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO-d ₆ ) δ 8.76 (s, 1H), 7.86 (d, J = 5.9 Hz, 1H), 7.74 (s, 1H), 7.44 (s, 1H), 6.00 (d, J = 6.1 Hz, 1H), 3.98 (t, J = 6.9 Hz, 2H), 3.84 (s, 2H), 3.48 (d, J = 4.7 Hz, 2H), 2.93 (d, J = 11.8 Hz, 2H) , 2.39 (s, 1H), 1.83–1.44 (m, 6H), 0.82 (t, J = 7.4 Hz, 3H).

Step 1E. (S)-2,2-difluorocyclopropane-1-carboxylic acid. To a 2.0 L Optimax ^™ reaction vessel was charged 2,2-difluorocyclopropane-1-(S)-carboxylate (R)-N-benzyl-1-phenylethane-1-aminium in water (945 mL). Salt (prepared as described in Example 2) (150 g, 450 mmol), MTBE (1 L), and concentrated sulfuric acid (27.2 mL, 0.495 mol) were charged. The mixture was stirred at 25° C. for about 1 hour. The aqueous layer was separated and extracted with MTBE (630 mL). The combined organic layers were concentrated to give 56.7 g (95%) of the title compound as an oil.

Step 1F. ((S)-2,2-difluorocyclopropyl)((1R,5S)-3-(2-((1-propyl-1H-pyrazol-4-yl)amino)pyrimidin-4-yl )-3,8-diazabicyclo[3.2.1]octan-8-yl)methanone. To a 1.0 L Optimax reaction vessel charged with THF (595 mL) was added (S)-2,2-difluorocyclopropane-1-carboxylic acid (43.7 g, 1.25 equiv, 331 mmol). After cooling the solution to 0° C., CDI (59.7 g, 1.35 eq, 357 mmol) as a solid was added in portions. After stirring at 0° C. for about 90 min, water (17.0 g, 17.0 mL, 944 mmol) was added via addition funnel over 5 min. The mixture was stirred at 0 °C for about 20 minutes. 2-Hydroxypyridine N-oxide (1.47 g, 0.05 equiv, 13.2 mmol) and 4-((1R,5S)-3,8-diazabicyclo[3.2.1]octan-3-yl)-N- (1-Propyl-1H-pyrazol-4-yl)pyrimidin-2-amine (85.0 g, 264 mmol) was added as a solid to the reaction mixture. The reaction mixture was stirred at 0 °C for about 12 hours. The reaction mixture was diluted with 600 mL of MIBK and filtered through a thin layer of Celite ^™ to remove solid impurities. The filtrate was transferred to a separatory funnel and washed sequentially with 50% saturated sodium bicarbonate solution, 50% saturated ammonium chloride solution and 50% saturated brine. The organic phase was transferred to a 2 L Optimax ^™ reactor and THF was removed by vacuum distillation at 70°C. Heptane (600 mL) was slowly added to the solution at 70°C and it was stirred at 70°C for about 2 hours to give a white suspension. Another 600 mL of heptane was charged to the suspension over 1 hour using a dosing pump. The resulting suspension was stirred at 70° C. for about 2 hours and then cooled to 25° C. over about 3 hours. The mixture was stirred at 25 °C for an additional 8 hours. The suspension was filtered to yield 97.6 g (88%) of the title compound as a white solid. HRMS (LC-ESI) calcd for C ₂₀ H ₂₅ F ₂ N ₇ O [(M+H) ⁺ ] m/z 418.2162, found 418.2159. ¹ H NMR (400 MHz, DMSO-d ₆ ) δ 8.87 (s, 1H), 7.93 (d, J = 5.9 Hz, 1H),. 7.75 (s, 1H), 7.46 (d, J = 8.5 Hz, 1H), 6.11 (dd, J = 10.6, 6.0 Hz, 1H), 4.75–4.51 (m, 2H), 4.00 (t, J = 6.9 Hz) , 4H), 3.28–2.85 (m, 3H), 2.02–1.57 (m, 8H), 0.82 (t, J = 7.4 Hz, 3H).

Powder X-ray diffraction analysis

Powder X-ray diffraction analysis was performed using a Bruker AXS D8 Endeavor diffractometer equipped with a Cu radiation source. The diverging slits were set to 15 mm continuous illumination. Diffracted radiation was detected by a PSD-Lynx Eye detector, with the detector PSD aperture set at 4.129 degrees. The X-ray tube voltage and amperage were set to 40 kV and 40 mA, respectively. Data were collected with a theta-theta goniometer at Cu wavelengths from 3.0 to 40.0 degrees 2-theta using a step size of 0.00997 degrees and a step time of 1.0 seconds. The anti-scatter screen was set at a fixed distance of 1.5 mm. Samples were spun at 15/min during collection. Samples were prepared by placing them in a silicon low background sample holder and rotated during collection. Data were collected using Bruker DIFFRAC ^™ Plus software and analysis was performed by EVA DIFFRAC Plus software (v. 5.0.0.22). In general, a preliminary peak arrangement was made using a threshold value of 1 and a width value of 0.235. The automated batch result was visually confirmed, and the peak position was adjusted to the peak maximum. Peaks with a relative intensity of ≧3% were generally selected. Peaks that did not resolve or were consistent with noise were not selected. Typical errors associated with peak positions of crystalline materials from PXRD as stated in USP are up to +/- 0.2° 2-theta (USP-941).

Table 1. PXRD peak list for compound IA.

1 is a crystalline ((S)-2,2-difluorocyclopropyl)((1R,5S)-3-(2-((1-propyl-1H-pyrazol-4-yl)amino)pyrimidine Powder X-ray diffraction analysis of -4-yl)-3,8-diazabicyclo[3.2.1]octan-8-yl)methanone. Based on Table 1, crystalline Compound IA is characterized by an X-ray powder diffraction pattern with characteristic diffraction peaks at 4.6, 9.2, 18.5, and 19.6 ± 0.2 degrees 2 theta.

Example 2. ((S)-2,2-difluorocyclopropyl)((1R,5S)-3-(2-((1-((R)-2-hydroxypropyl)-1H-pyra zol-4-yl)amino)pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octan-8-yl)methanone (IB).

Step 2A. (R)-1-(4-nitro-1H-pyrazol-1-yl)propan-2-ol. To a flask charged with (R)-(+)-propylene oxide (12 mL) was added 4-nitro-1H-pyrazole (4.0 g, 40 mmol) followed by Cs ₂ CO ₃ (6.92 g, 21.2 mmol). did The resulting mixture was stirred at 15° C. for about 32 hours. Water (30 mL) was added and the mixture was extracted with EtOAc (3 x 40 ml). The combined organic layers were dried over sodium sulfate and filtered. The filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel chromatography eluting with a solvent mixture of EtOAc:petroleum ether (1:1) to yield 5.3 g (90%) of the title compound as a pale yellow oil. MS m/z 171.9 [M+H] ⁺ . ^1H NMR (400 MHz, CDCl ₃ ) δ 8.23 (d, J = 0.6 Hz, 1H), 8.09 (s, 1H), 4.34-4.26 (m, 1H), 4.25 (dd, J = 13.8, 2.6 Hz, 1H), 4.06 (dd, J = 13.7, 7.9 Hz, 1H), 2.47 (d, J = 3.8 Hz, 1H), 1.28 (d, J = 6.3 Hz, 3H).

Step 2B. (R)-1-(4-amino-1H-pyrazol-1-yl)propan-2-ol. Pd/C (10% Pd, 659 mg, 0.02 eq) was added. The resulting suspension was stirred at room temperature under 1 atm pressure of hydrogen gas for about 16 hours. The reaction mixture was filtered and the filtrate was concentrated under reduced pressure to give 4.3 g (98%) of the title compound as a dark oil. MS m/z 142.1 [M+H] ⁺ . ^1H NMR (400 MHz, CDCl ₃ ) δ 7.17 (d, J = 0.9 Hz, 1H), 7.03 (d, J = 0.9 Hz, 1H), 4.21-4.12 (m, 1H), 4.01 (dd, J = 13.8, 2.8 Hz, 1H), 3.85 (dd, J = 13.8, 8.0 Hz, 1H), 3.47 (s, 1H), 1.18 (d, J = 6.4 Hz, 3H). [α] ²⁰ _D = -10.862 (c = 0.2 g/100 mL, MeOH)

Step 2C. tert-Butyl (1R,5S)-3-(2-chloropyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octane-8-carboxylate. Triethylamine (7.9 ml, 56.5 mmol) was added. After about 10 minutes, 2,4-dichloropyrimidine (4.6 g, 31.1 mmol) was added to the mixture in about two equal portions. The resulting mixture was stirred at about 15° C. for about 5 hours. The mixture was concentrated under reduced pressure to obtain a slurry. The slurry was dissolved in dichloromethane (150 mL), washed with water (2 x 50 mL), dried over sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography eluting with 0-50% EtOAc in petroleum ether to yield 8.3 g (90%) of the title compound as a white solid. MS m/z 325.1 [M+H] ⁺ .

Step 2D. (1R,5S)-3-(2-chloropyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octane. A solution of HCl in MeOH (60 mL, 4.0 M) was dissolved in tert-butyl (1R,5S)-3-(2-chloropyrimidin-4-yl)-3,8-diazabicyclo[ 3.2.1]octane-8-carboxylate (10.3 g, 31.7 mmol) was added to a solution. After addition, the mixture was stirred at about 15° C. for about 3 hours. The reaction was concentrated under reduced pressure. Water (40 mL) was added to the resulting solid and cooled in an ice bath. Aqueous sodium hydroxide solution was slowly added to the stirred reaction mixture until a pH of about 10 was reached. The resulting slurry was extracted with dichloromethane (2 x 100 mL). The combined organic extracts were dried over sodium sulfate and filtered. The filtrate was concentrated under reduced pressure to yield 6.6 g (87%) of the title compound as a white solid. MS m/z 225.0 [M+H] ⁺ .

Step 2E. ((1R,5S)-3-(2-chloropyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octan-8-yl)((S)-2,2-difluoro rocyclopropyl)methanone. T3P (8.86 g, 13.9 mmol, 50% solution in EtOAc) was dissolved in ACN (4 mL) as (1R,5S)-3-(2-chloropyrimidin-4-yl)-3,8-diazabicyclo[3.2 .1] octane (1.56 g, 6.96 mmol) with (S)-2,2-difluorocyclopropane-1-carboxylic acid 2,2',2"-nitrile prepared as described in Preparation 1 In an ice bath cooled solution of tris(ethane-1-ol) salt (1.9 g, 6.96 mmol) and triethylamine (4.9 mL, 34.8 mmol) were added slowly The resulting reaction mixture was stirred in an ice bath for about 2 hours The aqueous solution of NaHCO ₃ (100 mL) was added to the ice-bath cooled mixture.Then the mixture was diluted with water (500 mL) and EtOAc (500 mL).The organic layer was isolated and with brine (2 x 500 mL) Washed, dried over sodium sulfate and filtered The filtrate was concentrated to give 2.3 g (100%) of the title compound as a yellow solid MS m/z 329.1 [M+H] ⁺ .[α] ²⁵ _D = + 38.010 (c=0.2 g/100 mL, MeOH).

Step 2F. ((S)-2,2-difluorocyclopropyl)((1R,5S)-3-(2-((1-((R)-2-hydroxypropyl)-1H-pyrazole-4- yl)amino)pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octan-8-yl)methanone. ((1R,5S)-3-(2-chloropyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octan-8-yl)((S)- in isopropanol (19.6 ml) (R)-1-(4-amino-1H-pyrazol-1-yl)propan-2-ol (258 mg , 1.83 mmol, 1.2 equiv) followed by concentrated hydrochloric acid (0.05 mL). The resulting solution was stirred at 76° C. for about 16 hours. The reaction mixture was concentrated under reduced pressure. The residue was taken up in water (20 mL), cooled in an ice-water bath, and neutralized to pH>7 by addition of ammonium hydroxide. The solution was extracted with dichloromethane (2 x 30 mL). The combined organic extracts were dried over sodium sulfate and filtered. The filtrate was concentrated under reduced pressure. The resulting crude product was analyzed by preparative HPLC (Welch Xtimate ^™ C18, 25 mm id x 150 mm; water (0.05% ammonia hydroxide v/v)-ACN; 13-43%, 7 min). to give 297.4 mg (45%) of the title compound as a white solid after lyophilization. MS m/z 434.2 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO-d ₆ ) δ 8.90 (s, 1H), 7.93 (d, J = 5.9 Hz, 1H), 7.79 (s, 1H), 7.44 (d, J = 6.4 Hz, 1H) , 6.14–6.10 (m, 1H), 4.88 (d, J = 3.9 Hz, 1H), 4.73–4.51 (m, 2H), 4.14 (br s, 1H), 3.95–3.90 (m, 3H), 3.33– 2.75 (m, 3H), 2.10–1.50 (m, 7H), 1.02 (d, J = 4.6 Hz, 3H). [α] ²⁰ _D = +52.807 (c = 0.2 g/100 mL, MeOH).

Example 3. ((S)-2,2-difluorocyclopropyl)((1R,5S)-3-(2-((1-((S)-2-hydroxypropyl)-1H-pyra zol-4-yl)amino)pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octan-8-yl)methanone (IC).

Step 3A. (S)-1-(4-nitro-1H-pyrazol-1-yl)propan-2-ol. To a flask charged with (S)-(-)-propylene oxide (6 mL) was added 4-nitro-1H-pyrazole (2.0 g, 20 mmol) followed by Cs ₂ CO ₃ (5.76 g, 17.7 mmol). did The resulting mixture was stirred at 15° C. for about 32 hours. Water (30 mL) was added and the mixture was extracted with EtOAc (3 x 40 ml). The combined organic layers were dried over sodium sulfate and filtered. The filtrate was concentrated under reduced pressure. The resulting residue was purified on silica gel chromatography, eluting with a solvent mixture of EtOAc:petroleum ether (1:1) to yield 2.2 g (70%) of the title compound as a pale yellow oil. MS m/z 171.9 [M+H] ⁺ . ¹ H NMR (400 MHz, CDCl ₃ ) δ 8.23 (s, 1H), 8.09 (s, 1H), 4.31-4.24 (m, 1H), 4.22 (dd, J = 13.8, 2.5 Hz, 1H), 4.06 ( dd, J = 13.7, 7.8 Hz, 1H), 2.49 (d, J = 3.8 Hz, 1H), 1.28 (d, J = 6.3 Hz, 3H).

Step 3B. (S)-1-(4-amino-1H-pyrazol-1-yl)propan-2-ol. Pd/C (10% Pd, 249 mg, 0.2 eq) was added. The resulting suspension was stirred at room temperature under 1 atm of hydrogen gas for about 16 hours. The reaction mixture was filtered and the filtrate was concentrated under reduced pressure to give 1.5 g (91%) of the title compound as a dark oil. MS m/z 142.1 [M+H] ⁺ . ^1H NMR (400 MHz, CDCl ₃ ) δ 7.19 (d, J = 0.9 Hz, 1H), 7.03 (d, J = 0.8 Hz, 1H), 4.18-4.10 (m, 1H), 4.02 (dd, J = 13.8, 2.8 Hz, 1H), 3.86 (dd, J = 13.8, 7.9 Hz, 1H), 3.47 (s, 1H), 1.21 (d, J = 6.4 Hz, 3H). [α] ²⁰ _D = +38.201 (c = 0.2 g/100 mL, MeOH).

Step 3C. ((S)-2,2-difluorocyclopropyl)((1R,5S)-3-(2-((1-((S)-2-hydroxypropyl)-1H-pyrazole-4- yl)amino)pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octan-8-yl)methanone. ((1R,5S)-3-(2-chloropyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octane-8 prepared as described in step 2E in isopropanol (21.7 mL) -yl)((S)-2,2-difluorocyclopropyl)methanone (500 mg, 1.52 mmol) in a solution of (S)-1-(4-amino-1H-pyrazol-1-yl) Propan-2-ol (258 mg, 1.83 mmol, 1.2 equiv) was added, followed by concentrated hydrochloric acid (1 mL). The resulting solution was stirred at 76° C. for about 16 hours. The reaction mixture was then concentrated under reduced pressure. The residue was taken up in water (20 mL), cooled with an ice-water bath, neutralized and adjusted to pH>7 by addition of ammonium hydroxide. The solution mixture was extracted with dichloromethane (2 x 30 ml). The combined organic extracts were dried over sodium sulfate and filtered. The filtrate was concentrated under reduced pressure. The resulting residue was purified by preparative HPLC (Welch Xtimate ^™ C18, 25 mm id x 150 mm; water (0.05% ammonia hydroxide v/v)-ACN; 14-44%, 7 min) 296.4 mg (45%) of the title compound was obtained as a white solid after lyophilization. MS m/z 434.1 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO-d ₆ ) δ 8.90 (s, 1H), 7.93 (d, J = 5.6 Hz, 1H), 7.80 (s, 1H), 7.44 (d, J = 5.6 Hz, 1H) , 6.13–6.09 (m, 1H), 4.88 (d, J = 3.9 Hz, 1H), 4.69–4.58 (m, 2H), 4.14 (br s, 1H), 3.95–3.92 (m, 3H), 3.32– 2.90 (m, 3H), 2.00–1.65 (m, 7H), 1.01 (d, J = 6.0 Hz, 3H). [α] ²⁰ _D = +56.761 (c = 0.2 g/100 mL, MeOH).

Example 4. ((S)-2,2-difluorocyclopropyl)((1R,5S)-3-(2-((1-(3-hydroxypropyl)-1H-(pyrazole-4 -yl)amino)pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octan-8-yl)methanone (ID).

Step 4A. 3-(4-nitro-1H-pyrazol-1-yl)propan-1-ol. To a mixture of 4-nitro-1H-pyrazole (1.13 g, 9.9 mmol) in dimethylformamide (20 mL) was added 3-bromopropanol (1.67 g, 12 mmol) and potassium carbonate (1.38 g, 9.99 mmol). did The mixture was heated to 60° C. for about 16 hours. The mixture was concentrated under reduced pressure. Water (100 mL) was added to the residue and the mixture was extracted with EtOAc (3 x 100 mL). The combined organic extracts were dried over sodium sulfate and filtered. The filtrate was concentrated under reduced pressure to give 1.7 g (99%) of the title compound. MS m/z 171.9 [M+H] ⁺ .

Step 4B. 3-(4-amino-1H-pyrazol-1-yl)propan-1-ol. To a mixture of 3-(4-nitro-1H-pyrazol-1-yl)propan-1-ol (1.70 g, 9.93 mmol) in MeOH (19.0 mL) was added Pd/C (10% Pd, 0.60 g, 0.06 mmol). ) was added. The mixture was stirred at room temperature under 1 atm of hydrogen gas for 16 hours. The mixture was filtered and the filtrate was concentrated to give 1.4 g (100%) of the title compound. ^1H NMR (400 MHz, CDCl ₃ ) δ 7.19-7.12 (m, 1H), 7.07-6.99 (m, 1H), 4.15 (t, J = 6.5 Hz, 2H), 3.60 (t, J = 5.9 Hz, 2H), 2.04–1.95 (m, 2H).

Step 4C. ((S)-2,2-difluorocyclopropyl)((1R,5S)-3-(2-((1-(3-hydroxypropyl)-1H-pyrazol-4-yl)amino) pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octan-8-yl)methanone. ((1R,5S)-3-(2-chloropyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octane-8 prepared as described in step 2E in isopropanol (24 mL) -yl)((S)-2,2-difluorocyclopropyl)methanone (600 mg, 1.8 mmol) in a mixture of 3-(4-amino-1H-pyrazol-1-yl)propan-1- Ol (515 mg, 3.6 mmol) and concentrated hydrochloric acid (1 mL) were added. The mixture was heated to 70° C. for about 16 hours. The mixture was concentrated under reduced pressure. The residue was dissolved in water (20 mL) and the pH was adjusted to >7 with ammonium hydroxide while cooling in an ice-water bath. The mixture was extracted with dichloromethane (2 x 30 mL). The organic extract was dried over sodium sulfate and filtered. The filtrate was concentrated. The residue was purified by preparative HPLC (Welch Xtimate ^™ C18, 25 mm id x 150 mm; water (0.05% ammonia hydroxide v/v)-ACN; 14-42%, 7 min) and lyophilized. Afterwards 432 mg (55%) of the title compound was obtained as a white solid. MS m/z 434.3 [M+H] ⁺ . ¹ H NMR (400 MHz, DMSO-d ₆ ) δ 8.90 (s, 1H), 7.93 (d, J = 5.6 Hz, 1H), 7.75 (s, 1H), 7.46 (d, J = 4.8 Hz, 1H) , 6.14–6.10 (m, 1H), 4.71–4.56 (m, 3H), 4.19 (br, 1H), 4.10 (t, J = 6.9 Hz, 2H), 3.38 (q, J = 5.9 Hz, 2H), 3.27-2.90 (m, 3H), 2.00-1.64 (m, 9H).

black protocol

JAK family caliper enzyme assay

Compounds of the invention were evaluated by in vitro methods to determine their respective ability to inhibit JAK kinases (TYK2, JAK1, JAK2, JAK3). Inhibitory activity was determined using a microfluidic assay (LabChip 3000 ^TM Mobility Change Technology, Caliper Life Science) to determine recombinant human 4 members of the JAK family, JAK1, JAK2, JAK3 and TYK2, respectively. Phosphorylation of synthetic peptides by the kinase domain was monitored. The reaction mixture contained 1 μM of fluorescently labeled synthetic peptide, a concentration below the apparent K _m , and 1 mM ATP.

Compounds in DMSO solution were added to 384-well plates. The reaction mixture contained 10 mM HEPES, pH 7.4, 10 mM MgCl ₂ , 0.01% BSA, 0.0005% Tween 20 ^™ , 1 mM ATP and 1 μM peptide substrate. The JAK1 and TYK2 assays contained 1 μM of IRStide peptide (5FAM-KKSRGDYMTMQID), and the JAK2 and JAK3 assays contained 1 μM of JAKtide peptide (FITC-KGGEEEEYFELVKK). Initiate the assay by adding 20 nM JAK1, 1 nM JAK2, 1 nM JAK or 1 nM TYK2 enzyme and incubate at room temperature for 3 h for JAK1, 60 min for JAK2, 75 min for JAK3 or 135 min for TYK2. Incubated. Enzyme concentration and incubation time were optimized for each new enzyme preparation and slightly modified over time to ensure 20% to 30% phosphorylation. The assay was stopped with 15 μL of 180 mM HEPES, pH 7.4, 20 mM EDTA, and 0.2% Coating Reagent 3. The assay plate was placed on a Caliper Life Sciences LC3000 instrument and each well was sampled using appropriate separation conditions to measure unphosphorylated and phosphorylated peptides.

data analysis

Data was collected using the HTS Well Analyzer software from Caliper Life Sciences. The data output for data analysis is the percent converted product calculated as the peak height (Equation 1).

Equation 1:

% converted product = 100*((product)/(product + substrate))

The percent effect at each compound concentration was calculated based on the positive and negative control wells contained within each assay plate (Equation 2). Positive control wells contained saturating concentrations of control compounds that produced phosphorylation levels comparable to background (i.e., fully inhibited JAK1, JAK2, JAK3 or TYK2). Negative control wells contained DMSO alone (at the same concentration as compound wells) used to establish baseline activity (ie, uninhibited JAK1, JAK2, JAK3 or TYK2) in the assay.

Equation 2:

% effect = 100*((sample well - negative control)/(positive control - negative control))

Percent effects were plotted against compound concentration compound. A 4-parameter logistic model was used to fit an unconstrained sigmoidal curve and determine the compound concentration required for 50% inhibition (IC50) (Equation 3).

Equation 3:

y = ((max-min)/(1+((x/IC50)^s)))+min

where max is the maximum asymptote (full inhibition), min is the minimum asymptote (no inhibition) and s is the slope factor. IC ₅₀ values are reported in nM units for each compound in Table 1.

Table 1.

HWB INF alpha induced STAT3 phosphorylation assay

Compounds of the present invention were evaluated for their ability to inhibit interferon alpha signaling in a human whole blood flow cytometry assay. Interferon alpha signals through TYK2 and JAK1. Test articles were prepared as 30 mM stock solutions in 100% DMSO and then diluted to 10 mM. An 11-point three dilution series was created in DMSO with a top concentration of 5 mM. Further dilution was performed by adding 4 μL of the test article solution to a top concentration of 400 μM to 96 μL of PBS. Human whole blood (HWB) was collected via venipuncture from healthy donors into Vacutainer ^™ collection tubes containing sodium heparin (catalog number 366480; Becton Dickinson, Franklin Lakes, NJ). Blood was warmed to 37° C. before use. To a 96-well polypropylene plate (VWR 10755-246), 90 μL of HWB was added per well, followed by the addition of 5 μL test article solution prepared above to give a top concentration of 20 μM. Plates were mixed and incubated at 37° C. for 60 minutes. To each well was added 5 μL of human IFN alpha (Universal Type I IFN, R&D Systems #11200-2; final concentration of 5000 U/ml) or D-PBS (unstimulated control); Mix and incubate at 37° C. for 15 minutes. Reactions were quenched by adding 700 μL/well of Lysis/Fixation Buffer (BD Phosflow 5x Lysis/Fixation Buffer (BD #558049)) to all wells and incubated at 37° C. for 20 minutes; After washing with FACS buffer [D-PBS containing 0.1% BSA and 0.1% sodium azide (Invitrogen cat#14190)], 400 μL ice-cold 90% methanol/water was added to each well and incubated at 4° C. for 30 minutes. Washed once more with FACS buffer, all samples were finally incubated in 150 μL/well of Alexa Fluor 647 conjugated anti-phospho-STAT3 (pY705) antibody (BD #557815) 1:150 in FACS buffer. Resuspended by dilution. Samples were incubated overnight at 4°C.

flow cytometry

Samples were transferred to 96-well U-bottom plates (Falcon #353077) and flow cytometric analysis was performed on an LSRFortessa (BD Biosciences) equipped with an HTS plate loader. Lymphocyte populations were gated for histogram analysis of pSTAT3. Background fluorescence was defined using unstimulated cells and the gate was placed below the peak to include ˜0.5% gated population. Histogram statistical analysis was performed using FACSDiva version 8.0 (BD Biosciences) software. The relative fluorescence unit (RFU), which measures the level of phospho STAT, was calculated by multiplying the percent positive population and its mean fluorescence. Data from 11 compound concentrations (at each concentration alone) were normalized as a percentage of control based on the formula:

% of control = 100 x (A-B)/(C-B)

where A is RFU from wells containing compound and cytokines, B is RFU (minimum fluorescence) from wells without cytokines and no compound, and C is RFU (maximum fluorescence) from wells containing only cytokines. Inhibition curves and IC50 values were determined using Prism version 7 software (GraphPad, La Jolla, Calif.).

Table 2.

Biomarker Tuning in a Th2 Stimulated In Vitro Human Skin Assay

Compounds of the present invention were evaluated for their ability to modulate the CXCL10 biomarker in Th2 stimulated human skin ex vivo after topical application.

The compound was dissolved in a formulation consisting of:

10% w/w white petrolatum,

5% w/w mineral oil,

10% w/w emulsifying wax,

2% w/w oleyl alcohol,

46% w/w water,

15% w/w diethylene glycol monoethyl ether,

10% w/w polyethylene glycol (PEG) 400, and

1% w/w 2-phenoxyethanol

Human skin freshly excised from the surgical specimen was cut to a thickness of 750 μm using a dermatome and tissue cultured such that the upper surface of the skin was exposed to air in a Franz cell donor chamber and the lower (dermis) exposed to keratinization medium. was mounted on a 7 mm static Franz cell. A total of two skin donors were used (n=6 per treatment per donor). Incubation was performed in a standard tissue culture incubator. Compounds were dissolved in the indicated formulations at a concentration of 1% w/w and added apically to the skin at 10 μL per sample (approximately 18 μL per cm ² ) in 4-6 repetitions per donor. After 16 hours, the medium was replaced with a stimulation cocktail containing anti-CD3 mAb, anti-CD28 mAb, IL-2, IL-4, IL-33 and TSLP. Incubation was continued for an additional 24 hours at 37°C. The skin was then harvested, treated with RNALater solution, and processed for RNA extraction and qPCR, which were run in duplicate. MMP12, CCL26, CXCL10 and Filaggrin can be measured. Results for each sample were normalized to its own GAPDH internal standard, which was not appreciably affected by the stimulus or compound. Fold changes were normalized to untreated samples. The results are summarized in Table 3 and FIG. 2 below.

Table 3.

In vitro human hepatocyte clearance assay

A high-throughput human hepatocyte matrix-depletion assay was performed in 384-well formatted as previously described [Di et al., Eur. J. Med. Chem., 2012, 57, 441]. Briefly, cryopreserved human hepatocytes were thawed and resuspended in Williams' E medium supplemented with HEPES and Na ₂ CO ₃ . Cells were counted using the trypan blue exclusion method. Test compounds were added to human hepatocytes suspended in Williams E medium buffer and incubated at 37° C. in a humidified CO ₂ incubator (75% relative humidity, 5% CO ₂ /air) for 4 hours. The final incubation contained 500,000 cells/mL and 1 μM test compound in a 15 μL total volume containing 0.1% DMSO. At various time points (0, 3, 10, 30, 60, 120 and 240 minutes), aliquots of samples were taken and quenched with cold acetonitrile containing internal standard. Samples were centrifuged at 3000 rpm for 10 minutes at 4°C and the supernatant was transferred to a new plate, which was sealed prior to LC-MS/MS analysis. The apparent metabolic intrinsic clearance (CL _{int, met, app} ) was determined based on the substrate-depletion half-life estimated from the ratio of the peak area response of each compound to the peak area response of the internal standard as previously described [Di et al., Eur. J. Med. Chem., 2012, 57, 441]. The results are summarized in Table 4 below.

Table 4.

Claims (11)

A pharmaceutical composition comprising a compound of formula (I) or a pharmaceutically acceptable salt thereof:

wherein R ¹ and R ² are each independently hydrogen or hydroxy; wherein R ¹ and R ² are not both hydroxy. 2. The pharmaceutical composition according to claim 1, wherein R ¹ is hydroxy and R ² is hydrogen. 2. The pharmaceutical composition according to claim 1, wherein R ¹ is hydrogen and R ² is hydroxy. A pharmaceutical composition comprising a compound of Formula IA: or a pharmaceutically acceptable salt thereof:

. A pharmaceutical composition comprising a compound of Formula IB: or a pharmaceutically acceptable salt thereof:

. A pharmaceutical composition comprising a compound of formula (IC) or a pharmaceutically acceptable salt thereof:

. A pharmaceutical composition comprising a compound of Formula ID: or a pharmaceutically acceptable salt thereof:

. The pharmaceutical composition according to claim 1 comprising a compound selected from the group consisting of or a pharmaceutically acceptable salt thereof:
((S)-2,2-difluorocyclopropyl)((1R,5S)-3-(2-((1-propyl-1H-pyrazol-4-yl)amino)pyrimidin-4-yl )-3,8-diazabicyclo[3.2.1]octan-8-yl)methanone;
((S)-2,2-difluorocyclopropyl)((1R,5S)-3-(2-((1-((R)-2-hydroxypropyl)-1H-pyrazole-4- yl)amino)pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octan-8-yl)methanone;
((S)-2,2-difluorocyclopropyl)((1R,5S)-3-(2-((1-((S)-2-hydroxypropyl)-1H-pyrazole-4- yl)amino)pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octan-8-yl)methanone; and
((S)-2,2-difluorocyclopropyl)((1R,5S)-3-(2-((1-(3-hydroxypropyl)-1H-pyrazol-4-yl)amino) pyrimidin-4-yl)-3,8-diazabicyclo[3.2.1]octan-8-yl)methanone. 9. A pharmaceutical composition according to any one of claims 1 to 8, wherein the compound is in isolated form. 9. A pharmaceutical composition according to any one of claims 1 to 8, wherein the compound is in crystalline form. According to any one of claims 1 to 8, inflammation, autoimmune disease, neuroinflammation, arthritis, rheumatoid arthritis, spondyloarthropathy, systemic lupus erythematosus, lupus nephritis, osteoarthritis, gouty arthritis, pain, fever, lung Sarcoidosis, silicosis, cardiovascular disease, atherosclerosis, myocardial infarction, thrombosis, congestive heart failure and cardiac reperfusion injury, cardiomyopathy, stroke, ischemia, reperfusion injury, brain edema, brain trauma, neurodegeneration, liver disease, inflammatory bowel Diseases, Crohn's disease, ulcerative colitis, nephritis, retinitis, retinopathy, macular degeneration, glaucoma, diabetes (types 1 and 2), diabetic neuropathy, viral and bacterial infections, myalgia, endotoxic shock, toxicity Shock syndrome, osteoporosis, multiple sclerosis, endometriosis, dysmenorrhea, vaginitis, candidiasis, cancer, fibrosis, obesity, muscular dystrophy, polymyositis, dermatomyositis, autoimmune hepatitis, primary biliary cirrhosis, primary sclerosing cholangitis, vitiligo, Alzheimer's disease, Skin flushing, eczema, psoriasis, atopic dermatitis, sunburn, keloids, hypertrophic scars, rheumatic disease, urticaria, discoid lupus, cutaneous lupus, central nervous system lupus, psoriatic arthritis, asthma, allergic asthma, type I interferon Conditions such as Aicardi-Goutieres syndrome and other Mendelian diseases of overexpression of type I interferon, primary progressive multiple sclerosis, relapsing remitting multiple sclerosis, non-alcoholic fatty liver disease, non-alcoholic steatohepatitis , scleroderma, alopecia areata, scarring alopecia, prurigo, prurigo nodosa, CPUO, lichen disease, lichen planus, Steven's Johnson's syndrome, spondylopathy, myositis, vasculitis, pemphigus, lupus, major depressive disorder, allergy, ocular xerosis, transplant rejection, cancer, septic shock, cardiopulmonary dysfunction, acute respiratory disease, ankylosing spondylitis, cachexia, chronic graft-versus-host disease, acute graft-versus-host disease, celiac sprue, idiopathic thrombocytopenic thrombotic Purpura, thrombotic thrombocytopenic purpura, myasthenia gravis, Sjogren's syndrome, epidermal hyperplasia, cartilage inflammation, bone decomposition, juvenile arthritis, juvenile rheumatoid arthritis, juvenile rheumatoid arthritis polyarticular, juvenile rheumatoid arthritis polyarticular, systemic onset juvenile rheumatoid arthritis , juvenile ankylosing spondylitis, juvenile enteropathic arthritis, juvenile Reter syndrome, SEA syndrome, juvenile dermatomyositis, juvenile psoriatic arthritis, juvenile scleroderma, juvenile systemic lupus erythematosus, juvenile vasculitis, hydroarticular rheumatoid arthritis, polyarticular Rheumatoid arthritis, systemic onset rheumatoid arthritis, enteropathic arthritis, reactive arthritis, Letter syndrome, myositis, polymyositis, dermatomyositis, polyarteritis nodosa, Wegener's granulomatosis, arteritis, polymyalgia rheumatica, sarcoidosis , cirrhosis, primary biliary sclerosis, sclerosing cholangitis, dermatitis, Still's disease, chronic obstructive pulmonary disease, Guillain-Barre disease, Graves' disease, Addison's disease , Raynaud's phenomenon, psoriatic epidermoplasia, plaque psoriasis, erythematous psoriasis, inverse psoriasis, pustular psoriasis, erythematous psoriasis, immune disorders associated with or arising from the activation of pathogenic lymphocytes, non-infectious uveitis, Behcet's disease ( A pharmaceutical composition for the treatment or prevention of a disease or condition selected from Behcet's disease) and Vogt-Koyanagi-Harada syndrome.

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