WO2025109475A1 - Shp-1 agonists - Google Patents

Abstract

The present invention provides SHP-1 agonists of Formula I.

Description

  SHP-1 AGONISTS FIELD [0001] The present invention relates to SHP-1 agonists useful for the treatment of autoimmune disorders and certain cancers. BACKGROUND [0002] The Src homology region 2 domain-containing phosphatase 1 (SHP-1) is a tyrosine phosphatase and a key regulator of immune function known to exert a negative regulatory effect on cellular signaling in both the innate and adaptive immune systems. Mice that have a spontaneous mutation in the Ptpn6 gene resulting in loss of SHP-1 activity develop a spectrum of autoimmune and inflammatory symptoms (Claire L. Abram and Clifford A. Lowell, Shp1 function in myeloid cells, 102 JOURNAL OF LEUKOCYTE BIOLOGY 657 (2017)). Activation of SHP-1 is an attractive approach for direct treatment of certain carcinomas in view of its negative regulatory role in cell proliferation, migration, and invasion (Stephanie M. Stanford and Nunzio Bottini, Targeting protein phosphatases in cancer immunotherapy and autoimmune disorders, 22 NATURE REVIEWS DRUG DISCOVERY 273, 284 – 286 (April 2023)). SHP-1 agonists are needed to provide new treatments for autoimmune disorders and certain cancers. SUMMARY OF THE INVENTION [0003] An aspect of the invention is to provide SHP-1 agonist compounds of Formula: wherein:

R¹ is halo, amino, or C1 – C4 alkoxy; ;

R⁴ is C₂ – C₄ alkyl or C₂ – C₄ alkenyl; and   X is phenyl or pyridinyl optionally substituted with one substituent selected from halo, hydroxy, C1 – C4 alkyl, -NHR^a wherein R^a is H, C3 – C6 cycloalkyl, or phenyl, or C1 – C4 alkoxy optionally substituted with hydroxy or C₁ – C₄ alkoxy; or a pharmaceutically acceptable salt thereof. [0004] Another aspect of the invention is to provide SHP-1 agonist compounds of Formula I, or a pharmaceutically acceptable salt thereof, wherein R¹ is halo; R² is -X-NHR³; and R⁴ is C2 – C4 alkenyl. [0005] Another aspect of the invention is to provide SHP-1 agonist compounds of Formula I, or a pharmaceutically acceptable salt thereof, wherein R¹ is chloro; R² is -X-NHR³; and R⁴ is C2 – C4 alkenyl. [0006] Another aspect of the invention is to provide SHP-1 agonist compounds of Formula I, or a pharmaceutically acceptable salt thereof, wherein R¹ is chloro; R² is -X-NHR³; and R⁴ is ethenyl. [0007] Another aspect of the invention is to provide SHP-1 agonist compounds of Formula I, or a pharmaceutically acceptable salt thereof, wherein R¹ is halo; R² is -X-NHR³; X is phenyl or pyridinyl each optionally substituted with one substituent selected from the group consisting of halo and hydroxy; and R⁴ is C2 – C4 alkenyl. [0008] Another aspect of the invention is to provide SHP-1 agonist compounds of Formula I, or a pharmaceutically acceptable salt thereof, wherein R¹ is halo; R² is -X-NHR³; X is phenyl or pyridinyl; and R⁴ is C2 – C4 alkenyl. [0009] Another aspect of the invention is to provide SHP-1 agonist compounds of Formula I, or a pharmaceutically acceptable salt thereof, wherein R¹ is halo; R² is -X-NHR³; X is pyridinyl; and R⁴ is C2 – C4 alkenyl. [0010] Another aspect of the invention is to provide SHP-1 agonist compounds of Formula I, or a pharmaceutically acceptable salt thereof, wherein R¹ is halo; R² is -X-NHR³; X is pyridinyl optionally substituted with -NHR^a wherein R^a is C3 – C6 cycloalkyl or phenyl; and R⁴ is C2 – C4 alkenyl. [0011] An aspect of the invention is the compound of Formula: or a pharmaceutically acceptable

  [0012] An aspect of the invention is the compound of Formula: or a pharmaceutically acceptable

[0013] An aspect of the

or a pharmaceutically acceptable salt thereof. [0014] An aspect of the invention is the compound of Formula:

or a pharmaceutically acceptable salt thereof. [0015] Another aspect of the present invention provides a pharmaceutical composition comprising an agonist compound of Formula I or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier. [0016] A further aspect of the present invention provides a compound of Formula I or a pharmaceutically acceptable salt thereof for use in therapy.   [0017] Another aspect of the present invention provides a method for treating autoimmune disorders comprising administering to a patient in need thereof an effective amount of a compound of Formula I or a pharmaceutically acceptable salt thereof. [0018] Another aspect of the present invention provides a method for treating a SHP-1 susceptible cancer selected from bladder carcinoma, breast cancer, cervical squamous cell carcinoma, kidney renal papillary cell carcinoma, lung adenocarcinoma, pancreatic ductal adenocarcinoma, sarcoma, stomach adenocarcinoma, uterine corpus endometrial carcinoma, triple negative breast cancer, and colorectal cancer comprising administering to a patient in need thereof an effective amount of a compound of Formula I or a pharmaceutically acceptable salt thereof. [0019] A further aspect of the present invention provides the use of a compound of Formula I or a pharmaceutically acceptable salt thereof for the manufacture of a medicament for the treatment of autoimmune disorders. [0020] A further aspect of the present invention provides the use of a compound of Formula I or a pharmaceutically acceptable salt thereof for the manufacture of a medicament for the treatment of a SHP-1 susceptible cancer selected from bladder carcinoma, breast cancer, cervical squamous cell carcinoma, kidney renal papillary cell carcinoma, lung adenocarcinoma, pancreatic ductal adenocarcinoma, sarcoma, stomach adenocarcinoma, uterine corpus endometrial carcinoma, triple negative breast cancer, and colorectal cancer. DESCRIPTION OF THE DRAWINGS [0021] To assist those of ordinary skills in the relevant art in making and using the subject matter hereof, reference is made to the appended drawings, wherein: [0022] FIG. 1A is a graph of the recombinant SHP-1 phosphatase activity with 30 min (at room temperature) pre-incubation of NJH-04-021 at indicated concentrations performed at 37°C, expressed as rate of activity relative to DMSO control (in 4 replicates). [0023] FIG. 1B is a graph of the recombinant SHP-1 phosphatase activity with 30 min (at room temperature) pre-incubation of non-covalent control DGY-12-162 at indicated concentrations performed at 37°C, expressed as rate of activity relative to DMSO control (in 4 replicates). [0024] FIG. 1C is a graph of the recombinant SHP-1 phosphatase activity with 30 min (room temperature) pre-incubation of C-3 at indicated concentrations performed at 37°C, expressed as rate of activity relative to DMSO control (in 4 replicates).   [0025] FIG. 1D is a graph of the recombinant SHP-1 phosphatase activity with 30 min (room temperature) pre-incubation of non-covalent control MNN-03-006 at indicated concentrations performed at 37°C, expressed as rate of activity relative to DMSO control (in 4 replicates). [0026] FIG.2A is a graph of intact protein mass spectrometry of 2 µM recombinant wild type SHP- 1 incubated alone (48 h, 4°C). [0027] FIG.2B is a graph of intact protein mass spectrometry of 2 µM recombinant wild type SHP- 1 incubated with 5 times (10 µM) NJH-04-021 (48 h, 4°C). [0028] FIG.2C is a graph of intact protein mass spectrometry of 2 µM recombinant wild type SHP- 1 incubated with 10 times (20 µM) NJH-04-021 (48 h, 4°C). [0029] FIG.2D is a graph of intact protein mass spectrometry of 2 µM recombinant C102S mutant SHP-1 incubated alone (48 h, 4°C). [0030] FIG.2E is a graph of intact protein mass spectrometry of 2 µM recombinant C102S mutant SHP-1 incubated with 5 times (10 µM) NJH-04-021 (48 h, 4°C). [0031] FIG.2F is a graph of intact protein mass spectrometry of 2 µM recombinant C102S mutant SHP-1 incubated with 10 times (20 µM) NJH-04-021 (48 h, 4°C). [0032] FIG.3A is a western blot gel illustrating the NJH-04-021 and non-covalent control DGY- 12-162 induced IκBα degradation and NFκB p65 phosphorylation upon LPS stimulation (100 ng/ml, 15 min) at indicated concentrations (3 h pre-incubation) in immortalized bone marrow derived macrophages (iBMDMs). [0033] FIG. 3B is a western blot gel illustrating the C-3 and non-covalent control MNN-3-006 induced IκBα degradation and NFκB p65 phosphorylation upon LPS stimulation (100 ng/ml, 15 min) at indicated concentrations (3 h pre-incubation) in immortalized bone marrow derived macrophages (iBMDMs). [0034] FIG. 4 is a western blot gel illustrating the induction of IκBα degradation and NFκB p65 phosphorylation following LPS stimulation (100 ng/ml, 15 min) by 10 µM of select SHP-1 agonists (3 h pre-incubation). [0035] FIG. 5 is a western blot gel illustrating the induction of IκBα degradation and NFκB p65 phosphorylation following LPS stimulation (100 ng/ml, 15 min) by 50 µM of select SHP-1 agonists (3 h pre-incubation). DETAILED DESCRIPTION [0036] Terms used herein but not separately defined are taken to have their normal and customary meaning as understood by one of ordinary skill in the art.   [0037] The term “halo” is taken to mean fluoro, chloro, bromo, or iodo. [0038] The term “C1 – C4 alkoxy” is taken to mean a straight or branched alkyl chain of from 2 to 4 carbon atoms attached to an oxygen atom and includes methoxy, ethoxy, propoxy, isopropoxy, butoxy, isobutoxy, and the like. [0039] The term “C1 – C4 alkyl” is taken to mean a straight or branched alkyl chain of from 1 to 4 carbon atoms and includes methyl, ethyl, propyl, isopropyl, butyl, isobutyl and the like. Similarly, the term “C₂ – C₄ alkyl” is taken to mean a straight or branched alkyl chain of from 2 to 4 carbon atoms and includes ethyl, propyl, isopropyl, butyl, isobutyl and the like. [0040] The term “C₂ – C₄ alkenyl” is taken to mean a straight or branched alkenyl chain of from 2 to 4 carbon atoms and includes ethenyl, propenyl-2-yl, propen-3-yl, 2-methylpropen-3-yl, and the like. [0041] The term “C3 – C6 cycloalkyl” is taken to mean a carbocyclic ring of from 3 to 6 carbon atoms and includes cyclopropyl, cyclobutyl, cyclopentyl, and cyclohexyl. [0042] The term “patient” means mammal and “mammal” includes, but is not limited to, a human. [0043] “Therapeutically effective amount” means the dosage of a compound of Formula I, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition containing a compound of Formula I, or a pharmaceutically acceptable salt thereof, necessary to stimulate SHP-1 activity in a patient in need thereof. Anticipated dosages of a compound of Formula I, or a pharmaceutically acceptable salt thereof, are in the range of 1 mg/patient/day to 2000 mg/patient/day. The exact dosage required to treat a patient and the duration of treatment will be determined by a physician in view of the stage and severity of the disease as well as the specific needs and response of the individual patient. Dosage administration may be adjusted to provide an optimal therapeutic benefit to an individual patient and to manage or avoid drug-related toxicities. For example, in addition to single daily dosing, multiple smaller daily doses or administration on a staggered daily, weekly, or monthly schedule may be appropriate. [0044] The terms “treatment”, “treat”, and “treating” are meant to include the full spectrum of pharmaceutical intervention for a patient suffering from an autoimmune disorder, such as administration of a SHP-1 agonist of the present invention to alleviate, slow, or reverse one or more of a patient’s symptoms or to delay progression of the disorder even if the disorder is not actually eliminated. [0045] Autoimmune disorders are diseases that arise from an overactive immune response of the body against normally present substances and tissues. SHP-1 agonists are useful for the treatment of   autoimmune disorders due to SHP-1 regulatory activity on diverse cellular functions. For example, SHP-1 agonist treatment in a bleomycin-induced pulmonary fibrosis murine model suppressed inflammation and inhibited the transition of fibroblasts to myofibroblasts, suggesting that SHP-1 agonists would be useful for the treatment of pulmonary fibrosis (Shiao-Ya, et al., “Targeting pathogenic macrophages by the application of SHP-1 agonists reduces inflammation and alleviates pulmonary fibrosis”, 14 CELL DEATH AND DISEASE 352 (2023)). Further, stimulation of SHP-1 by administration of regorafenib to mice has been shown to prevent the development of severe arthritis in a human cartilage proteoglycan induced arthritis model or reduced disease severity when treatment was started after disease onset, suggesting the use of SHP-1 agonists in the prevention or treatment of rheumatoid arthritis (Adrien Markovics, et al., Regulation of autoimmune arthritis by the SHP-1 tyrosine phosphatase, 22 ARTHRITIS RESEARCH & THERAPY 160 (2020)). Also, activation of SHP-1 with boswellic acid has also been shown to inhibit the proliferation of hepatic stellate cells, suggesting the use of SHP-1 agonists for the treatment of hepatic fibrosis (Elena Tibaldi, et al., “The tyrosine phosphatase SHP-1 inhibits proliferation of activated hepatic stellate cells by impairing PDGF receptor signaling”, 1843(2) BIOCHIM BIOPHYS ACTA 288 (February 2014)). [0046] SHP-1 modulates the development and progression of certain cancers by attenuating or blocking signaling pathways that control cell proliferation, survival, migration, and invasion. Increased SHP-1 expression leads to better survival outcomes in patients suffering from bladder carcinoma, breast cancer, cervical squamous cell carcinoma, kidney renal papillary cell carcinoma, lung adenocarcinoma, pancreatic ductal adenocarcinoma, sarcoma, stomach adenocarcinoma, or uterine corpus endometrial carcinoma (Alessia Verone, et al., Shp1 in Solid Cancers and Their Therapy, 10 FRONTIERS IN ONCOLOGY 1 (June 2020)). SHP-1 agonists SC-43 and SC-78 have been shown to inactivate signal transducer and activator of transcription 3 (STAT3), suggesting their use in the treatment of colorectal cancer (Shin-Yi Chung, et al., Two novel SHP-1 agonists, SC-43 and SC-78, are more potent than regorafenib in suppressing the in vitro stemness of human colorectal cancer cells, 4 CELL DEATH DISCOVERY 82 (2018)). Additionally, SHP-1 agonist SP-60 has been taught to be useful in the treatment of triple-negative breast cancer with docetaxel (Chun-Yu Liu, et al., “Sorafenib analogue SC-60 induces apoptosis through the SHP-1/STAT3 pathway and enhances docetaxel cytotoxicity in triple-negative breast cancer cells”, 11 MOLECULAR ONCOLOGY 266 (2017)). [0047] A compound of Formula I or a pharmaceutically acceptable salt thereof is preferably formulated as a pharmaceutical composition using a pharmaceutically acceptable carrier and   administered by a variety of routes. Preferably, such compositions are for oral administration. Such pharmaceutical compositions and methods for preparing them are well known in the art. See, for example, REMINGTON: THE SCIENCE AND PRACTICE OF PHARMACY (L.V. Allen ed., Pharmaceutical Press, 22^nd Edition, 2012). [0048] A compound of Formula I or a pharmaceutically acceptable salt thereof may be administered either simultaneously with, or before, or after, one or more other therapeutic agents. The compound of Formula I or a pharmaceutically acceptable salt thereof, when administered with one or more therapeutic agents, may be administered separately, by the same or different route of administration, or together in the same pharmaceutical composition as the other therapeutic agent or agents. Where one or more additional therapeutic agents are administered, the administration of each therapeutic agent may be simultaneous, separate, or sequential. [0049] A compound of Formula I is capable of reacting with a number of inorganic and organic acids to form pharmaceutically acceptable acid addition salts. Such pharmaceutically acceptable salts and common methodology for preparing them are well known in the art. See, for example, P. Stahl, et al., HANDBOOK OF PHARMACEUTICAL SCIENCES, (VCHA/Wiley – VCH, 2002); S.M. Berge, et al., Pharmaceutical Salts, 66 JOURNAL OF PHARMACEUTICAL SCIENCES 1 (1977). [0050] Compounds of Formula I are named according to IUPAC, and may also be named according to CAS, and other naming conventions may be used to unambiguously identify a compound of Formula I or a pharmaceutically acceptable salt thereof. [0051] The compounds employed as initial starting materials in the synthesis of compounds of Formula I are well known and, to the extent not commercially available, are readily synthesized using specific references provided, by standard procedures commonly employed by those of ordinary skill in the art or are found in general reference texts. Examples of known procedures and methods include those in general reference texts such as: COMPREHENSIVE ORGANIC TRANSFORMATIONS (VCH Publishers Inc., 1989); COMPENDIUM OF ORGANIC SYNTHETIC METHODS (Wiley Interscience, Volumes 1 – 10, 1974 – 2002); Michael B. Smith and Jerry March, ADVANCED ORGANIC CHEMISTRY, REACTIONS, MECHANISMS, AND STRUCTURE (Wiley Interscience, 5^th ed.2001); Francis A. Carey and Richard J. Sundberg, ADVANCED ORGANIC CHEMISTRY, PART B, REACTIONS AND SYNTHESIS (Kluwer Academic/Plenum Publishers, 4^th ed.2000), and references cited therein. [0052] Certain intermediates described in the following preparations may contain one or more nitrogen protecting groups. It is understood that protecting groups may be varied as appreciated by one of skill in the art depending on the actual reaction conditions and the particular transformations   to be performed. The protection and deprotection conditions are well known to the skilled artisan and are described in the literature (See for example "Greene's Protective Groups in Organic Synthesis", Fifth Edition, by Peter G.M. Wuts and Theodora W. Greene, John Wiley and Sons, Inc.2014). [0053] The compounds of Formula I or pharmaceutically acceptable salts thereof may be prepared by a variety of procedures known in the art, some of which are illustrated in the Schemes, Preparations, and Examples below. The specific steps and methodology for each of the synthetic routes described may be combined in different ways, or in conjunction with steps from different schemes, to prepare compounds of Formula I or pharmaceutically acceptable salts thereof. The products of each step in the schemes below may be isolated by conventional methods well known in the art, including extraction, evaporation, precipitation, chromatography, filtration, trituration, and crystallization. In the schemes below, all substituents are as previously defined unless otherwise indicated. The reagents and starting materials are readily available to one of ordinary skill in the art. [0054] Certain abbreviations are defined as follows: “AcOH” means acetic acid; “BINAP” means 2,2′-bis(diphenylphosphino)-1,1′-binaphthyl; “Boc2O” means di-tert-butyl decarbonate; “DCM” means dichloromethane; “DIEA” or “DiPEA” means N, N-diisopropylethylamine; “DMAP” means dimethylaminopyridine; “DMF” means dimethylformamide; “DMSO” means dimethyl sulfoxide; “DPPA” means diphenylphosphoryl azide; “EtOAc” means ethyl acetate; “EtOH” means ethanol; “HATU” means hexafluorophosphate azabenzotriazole tetramethyl uronium; “IPA” means isopropanol; “MeOH” means methanol; “NaOtBu” means sodium tert-butoxide; “NMP” means N- methyl-2-pyrrolidinone; “PG” means protecting group; “PMB” means para-methoxybenzyl; “PMBNH2” means para-methoxybenzylamine; “TBSCl” means tert-butyldimethylsilyl chloride; “tBuOH” means tert-butanol; “TEA” means triethylamine; “THF” means tetrahydrofuran.  

  Scheme 1

[0055]

with a suitable pyridine 1, 2 or 7. In Step 1, the pyridine 1 is reacted with an appropriate amine, such as   PMBNH2, under palladium-catalyzed amination conditions well known to the skilled artisan to give 3. Suitable palladium-catalyzed amination conditions include reacting a compound bearing an appropriate halogen on an aromatic moiety, such as bromine or chlorine, with a suitable nucleophilic reactant such as an amine. The reaction occurs in the presence of a suitable palladium catalyst, such as tBuXPhos Pd G3, and an appropriate organic base, such as NaOtBu, in a suitable organic solvent, such as dioxane, at 100°C at least. The PG moiety on the pyridyl amine 3 is a standard amine protecting group standard to the art, including PMB protecting groups. The removal of the PG moiety on the protected pyridyl amine 3 is achieved under acidic conditions, such as TFA in DCM. The pyridine 2 is treated under nucleophilic aromatic substitution conditions standard to the skilled person in the art. Suitable nucleophilic aromatic substitution conditions include reacting a compound bearing an appropriate halogen on an aromatic moiety, such as fluorine or chlorine, with a suitable nucleophilic reactant, such as NaOCH3, in an appropriate organic solvent, such as DMSO, at 120°C at least and for at least 12 hours. In Step 4, the pyridine 4 is reacted with a suitable PG precursor, such as (Boc)2O, in presence of an appropriate organic base, such as DMAP and/or TEA, in a suitable organic solvent, such as DCM. The protected intermediate is reacted with an appropriate amine, such as PMBNH₂, under palladium-catalyzed amination conditions similar to Step 1. The intermediate 6 is afforded by hydrogenation conditions including Pd/C and hydrogen gas (50 psi) in an appropriate solvent, such as MeOH, at 50°C at least. The pyridyl amines 5, 6 and 7 are reacted with a suitable heterobiaryl species under nucleophilic aromatic substitution conditions, in presence of an appropriate base, such as NaH, in a suitable organic solvent, such as DMF, THF or NMP. The nitro group of intermediates Steps 5 and 7 are treated under reductive conditions standard to the skilled person with suitable reductive agents, such as iron powder and ammonium chloride, in an appropriate organic solvent, such as EtOH. The PG moiety on the intermediate of Step 6 is a standard amine protecting group, including a carbamate protecting group, and it is removed under acidic conditions similar to Step 3. The resulting amine 8, where Z is NH2 or OCH3, is reacted with a suitable acyl chloride, such as acryloyl chloride or propionyl chloride, in presence of an appropriate base, such as TEA or DiPEA, in a suitable organic solvent, such as DCM or THF.   Scheme 2 [0056]

beginning with a suitable arylamine 10 where Y is N or optionally substituted C and G is either nitro or amino. In Step 1, the arylamine 10 is reacted with an appropriate heterobiaryl halide 11 under nucleophilic aromatic substitution conditions to give 12. The reaction occurs in presence of a suitable base, such as DiPEA or Cs2CO3, in an appropriate organic solvent, such as IPA or DMF at a reaction temperature of 80°C at least. Alternatively, the reaction occurs in presence of a suitable organic solvent, such as EtOH under reflux conditions. In Step 2, it is not always necessary to protect the amine of intermediate 12 but if chosen, it can be reacted with an appropriate PG precursor, such as (Boc)₂O, in presence of a suitable organic base, such as DMAP, in an appropriate organic solvent, such as DCM. The intermediate in Step 2 can then be treated under nucleophilic aromatic substitution conditions with a suitable nucleophile, such as PMBNH₂ or MeOH activated by a standard base, such as NaH or DiPEA, in an appropriate organic solvent, such as MeOH or tBuOH, to prepare 13. The nitro group of the intermediate (G = nitro) is treated under reductive conditions known to the skilled person with suitable reductive agents, such as iron powder and ammonium chloride, or iron powder and AcOH, or Pd/C and hydrogen gas, in an appropriate organic solvent, such as THF or EtOH. If necessary, the PG moiety can be removed under suitable acidic conditions, such as TFA in DCM. The product 14 is obtained by reacting the resulting amines 12 or 13 with an appropriate acyl chloride, such as acryloyl   chloride or propionyl chloride, in the presence of an appropriate base such as NaHCO3, TEA, or DiPEA in a suitable organic solvent such as THF, DCM, or DMF. Alternatively, the amine 12 can be reacted with a suitable acid, such as acrylic acid, under amide coupling conditions well known to the skilled person and include using an appropriate coupling reagent, such as HATU, in the presence of a suitable organic base, such as TEA, in a suitable solvent, such as DCM, for at least 12 hours. Scheme 3 [0057] Scheme 3

beginning with a suitable amine 15. In Step 1, the amine 15 is reacted with heteroaryl halide 16 where R⁵ is F, C1 – C4 alkoxy optionally substituted with OPg or C₁ – C₄ alkoxy under nucleophilic aromatic substitution conditions well known to the skilled artisan to give 17. The reaction occurs in the presence of a   suitable base, such as NaH, in an appropriate organic solvent, such as NMP or DMF at a reaction temperature of 90°C at least. Optionally, the intermediate 17 is reacted with a suitable nucleophilic species under nucleophilic aromatic substitution conditions in the presence of an appropriate base, such as NaH, in an appropriate organic solvent, such as DMF at a reaction temperature of 90°C at least to give 18 in Step 3. The carboxylic acid intermediates 17 and 18 are treated under Curtius rearrangement conditions with a suitable azide donor, such as DPPA, and an appropriate nucleophile, such as tBuOH, in presence of a suitable organic base, such as TEA, to give the protected aryl amine 19. The removal of the PG moiety on the amine of the intermediate 19 is achieved under acidic conditions standard to the art. The amine 20 is obtained by reacting a suitable acid, such as HCl, to a stirring solution of 19 in an appropriate solvent, such as MeOH or EtOAc, at ambient temperature. One skilled in the art would understand that the acidic conditions employed in Step 5 are susceptible to remove a multitude of protecting groups. As such, if the R-group on 19 bears a moiety for which the protecting group is removed in Step 5, then the intermediate can be reacted with a suitable PG precursor, such as TBSCl, in presence of appropriate organic bases, such as DMAP and TEA, in a suitable organic solvent, such as DCM. The amine 20 is reacted with an appropriate acyl chloride, such as acryloyl chloride, in presence of an appropriate base, such as Na₂CO₃ or TEA, in a suitable organic solvent, such as THF or DCM. It is not always necessary to remove the PG on the R⁵-group of 21 but if chosen, it can be treated under acidic conditions standard to the art, with a suitable acid, such as TFA, in an appropriate solvent, such as DCM, at ambient temperature for at least 1 hour.

  Scheme 4 Br Cl R⁶ N 3 Cl

S_{tep 4}

6

R _R ⁶ Cl [0058] Scheme 4

with a suitable pyridine 22. In Step 1, the pyridine 22 is reacted with an appropriate amine, such as a C₃ – C6 cycloalkylamine or aniline, under palladium-catalyzed amination or nucleophilic aromatic substitution conditions, including a suitable palladium catalyst, such as RuPhos Pd G3, in the presence of an appropriate base, such as Cs₂CO₃ or DiPEA, in a suitable solvent, such as dioxane or DMF, at 100°C at least to provide the substituted pyridine 23 where R^a’ is a C₃ – C₆ cycloalkyl or phenyl. The heterobiarylamine 15 is prepared by reacting the suitable heterobiaryl species 11 with ammonia hydrate at 85°C at least, for 5 hours at least. In Step 3, the heterobiarylamine 15 is reacted with one of the pyridines 23 or 24 under palladium-catalyzed amination conditions. The reaction occurs in the   presence of a suitable palladium catalyst, such as tBuXPhos Pd G3, and an appropriate organic base, such as NaOtBu, in a suitable organic solvent, such as dioxane, at 100°C at least to provide a compound of formula 25 where R⁶ is either C₁ – C₄ alkoxy or -NHR^a’ where R^a’ is a C₃ – C₆ cycloalkyl or phenyl. When R⁶ is methoxy, the methyl group may be cleaved at any convenient point in the synthesis to provide the corresponding hydroxy derivative under standard conditions well known to the skilled artisan, such reaction with a Lewis acid, such as AlCl₃, in an appropriate solvent, such as 1,2-dichloroethane, at 80 °C at least to provide the corresponding hydroxy derivative. In Step 4, the nitro group of 25 is reduced under standard conditions known to the skilled person, such as reaction with iron powder and either ammonium chloride or AcOH, in an appropriate organic solvent such as EtOH and H₂O. The resulting amine 26 is reacted with a suitable acid, such as acrylic acid, under amide coupling conditions well known to the skilled person and include using an appropriate coupling reagent, such as HATU, in the presence of a suitable organic base, such as DiPEA or TEA, in an appropriate solvent, such as DMF or THF, for at least 4 hours to provide compounds of formula 27.  

  Scheme 5 [0059]

with a suitable amine 28. In Step 1, the amine 28 is reacted with an appropriate PG precursor such as (Boc)₂O in presence of a suitable organic base, such as TEA, in an appropriate organic solvent, such as DCM. The resulting intermediate 29 is then treated under standard palladium-catalyzed amination conditions to prepare the amide 30 in Step 2. Suitable palladium-catalyzed amination conditions are well known to the skilled person and include reacting a solution of 29, an appropriate organopalladium catalyst such as Pd2(dba)3, a suitable organophosphorous ligand, such as BINAP, and an appropriate amine donor, such as diphenylmethanimine, in a suitable solvent, such as toluene, in the presence of a suitable organic base, such as NaOtBu, under N2 atmosphere at 80°C at least. The removal of the PG on the newly formed amine of 30 is treated under basic conditions standard to the art, with a suitable base, such as NH₂OH, in an appropriate solvent, such as MeOH, at ambient temperature for at least 2   hours. The resulting aryl amine 31 is reacted with a heterobiaryl species 11 under nucleophilic aromatic substitution conditions well known to the skilled artisan to give 24. The reaction occurs in the presence of a suitable base, such as NaH, in an appropriate organic solvent, such as DMF. The PG moiety on the amine of the intermediate 32 is a standard amine protecting group well known to the skilled artisan, including carbamate and amide protecting groups. The intermediate amine 33 is obtained by reacting a suitable acid such as HCl to a stirring solution of 32 in an appropriate solvent, such as EtOAc, at ambient temperature for at least 2 hours. The resulting amine 33 is reacted with a suitable acyl chloride, such as acryloyl chloride, in presence of an appropriate base, such as Na2CO3, in a suitable organic solvent, such as THF. EXAMPLES Preparation 1 5-Chloropyrazolo[1,5-a]pyrimidin-7-amine [0060] A mixture of 5,7- (10 g, 53.19 mmol) in NH3.H2

O (25 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 85°C for 5 hours under N₂ atmosphere. The reaction mixture was filtered and the filter cake was concentrated under reduced pressure to give a residue. The residue was used as is in the next step. The title compound (7 g, crude) was obtained as a yellow solid. MS (m/z): 169 (M+H). Preparation 2 2-Chloro-6-methoxypyridin-4-amine [0061] A mixture of 2,6-

36.81 mmol), NaOMe (2.98 g, 55.21 mmol) in DMSO (30 mL) was degassed and purged with N₂ for 3 times, and then the mixture was stirred at 120°C for 12 hours under N₂ atmosphere. The reaction mixture was diluted with H₂O (30 mL) and extracted with EtOAc (90 mL). The combined organic layers were dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by silica gel chromatography (1-5% EtOAc/Petroleum ether). The title compound was obtained as a purple solid (2.2 g, 13.87 mmol, 37.69% yield). MS (m/z): 159 (M+H).   Preparation 3 tert-Butyl 6-chloro-1,3-dihydro-2H-pyrrolo[3,4-c]pyridine-2-carboxylate [0062] To a solution of 6-chloro-2,3-dihydro-1H-pyrrolo[3,4-c]pyridine (1.00 g, 5.23 mmol, HCl) in DCM (10 mL) was added TEA (635 mg, 6.28 mmol, 874 μL) and Boc2O (1.26 g, 5.76 mmol, 1.32 mL). The mixture was stirred at 25°C for 3 hours. The reaction mixture was diluted with H₂O (20 mL) and extracted with EtOAc (30 mL). The combined organic layers were washed with brine (20 mL), dried over Na2SO4, filtered and concentrated under reduced pressure to give the title compound as a gray solid (1.30 g, 5.10 mmol, 97.5% yield). MS (m/z): 255 (M+H). [0063] The following compounds were prepared by the method of Preparation 3: 1H NMR Prep. Chemical name Structu (400 No. re MHz, CDCl3) O δ 6.75 (s, tert-Butyl (tert- O N 1H), 6.45 4 butoxycarbonyl)(2-chloro- 6-meth O N Cl (s, 1H), oxypyridin-4- 3.95 (s, yl)carbamate O O 3H), 1.47 (s, 18H) Preparation 5 tert-Butyl (tert-butoxycarbonyl)(2-methoxy-6-((4-methoxybenzyl)amino)pyridin-4-yl)carbamate [0064] A mixture of tert-butyl (tert-butoxycarbonyl)(2-chloro-6-methoxypyridin-4-yl)carbamate (4.5 g, 12.54 mmol), PMBNH₂ (1.89 g, 13.80 mmol, 1.79 mL), tBuXPhos Pd G3 (996.24 mg, 1.25 mmol) and NaOtBu (2.41 g, 25.08 mmol) in dioxane (25 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 100°C for 12 hours under N₂ atmosphere. The reaction   mixture was diluted with H2O (100 mL) and extracted with EtOAc (90 mL). The combined organic layers were dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by silica gel chromatography (1-25% EtOAc/Petroleum ether). The title compound was obtained as a yellow solid (3.2 g, 6.96 mmol, 55.53% yield). MS (m/z): 460 (M+H). [0065] The following compounds were prepared by the method of Preparation 4: Prep. Chemical name Struc MS (m/z) No. ture (M+H) N²,N⁶-Bis(4- 6 methoxybenzyl)-4- 395 nitropyridine-2,6-diamine 7 6-Bromo-N-cyclopropyl-4- nitropyridin-2-amine 258 N²-(5-Chloropyrazolo[1,5- 8 a]pyrimidin-7-yl)-N⁶- cyclopropyl-4- 346 nitropyridine-2,6-diamine Preparation 9 tert-Butyl (2-amino-6-methoxypyridin-4-yl)carbamate [0066] To a solution of Pd/C (1 g, 1.67 mmol, 10% purity) in MeOH (20 mL) was added tert-butyl (tert-butoxycarbonyl)(2-methoxy-6-((4-methoxybenzyl)amino)pyridin-4-yl)carbamate (2.40 g, 5.56 mmol). The mixture was stirred at 50°C for 12 hours under H2 (50 Psi). The reaction mixture was filtered and the filtrate was concentrated under reduced pressure to remove solvent. The residue was purified by silica gel chromatography (1-100% EtOAc/Petroleum ether). The title compound was obtained as a white solid (1 g, 3.07 mmol, 55.2% yield). MS (m/z): 240 (M+H).   Preparation 10 tert-Butyl 6-((diphenylmethylene)amino)-1,3-dihydro-2H-pyrrolo[3,4-c]pyridine-2-carboxylate [0067] To a solution of tert- pyrrolo[3,4-c]pyridine-2-carboxylate

(1.11 g, 6.12 mmol, 1.03 mL), g, 6.12 mmol, 1.03 mL), BINAP (635 mg, 1.02 mmol) and tBuONa (735 mg, 7.66 mmol) in toluene (10 mL) was added Pd₂(dba)₃ (233 mg, 255 μmol) under N₂. The mixture was stirred at 80°C for 1 hour. The solution was concentrated in vacuum and the residue was diluted with H2O (100 mL) and extracted with EtOAc (90 mL). Organic layer was washed with brine (100 mL), dried over Na₂SO₄, filtered and filtrate was concentrated in vacuum. The residue was purified by silica gel chromatography (1-100% EtOAc/Petroleum ether) to obtain the title compound as a light yellow solid (954 mg, 1.91 mmol, 37.4% yield, 79.9% purity). MS (m/z): 400 (M+H). Preparation 11 tert-Butyl 6-amino-1,3-dihydro-2H-pyrrolo[3,4-c]pyridine-2-carboxylate [0068] To a solution of tert-

amino)-1,3-dihydro-2H-pyrrolo[3,4- c]pyridine-2-carboxylate (954 mg, 2.39 mmol) in MeOH (9 mL) was added NH2OH (315 mg, 4.78 mmol, 50.0% purity). The mixture was stirred at 20°C for 2 hours. The reaction mixture was concentrated in vacuum and the residue was purified by silica gel chromatography (5-10% EtOAc/Petroleum ether) to give the title compound as a light yellow solid (488 mg, 1.67 mmol, 69.7% yield, 80.3% purity). MS (m/z): 236 (M+H). Preparation 12 N¹-(5-Chloropyrazolo[1,5-a]pyrimidin-7-yl)benzene-1,3-diamine   [0069] To a solution of 5,7-dichloropyrazolo[1,5-a]pyrimidine (188.0 mg, 1.00 mmol) and 5- benzene-1,3-diamine (216 mg, 2.00 mmol) in IPA (5 mL) was added DiPEA (0.348 mL, 2.00 mmol). The reaction was allowed to stir at 85˚C for 2 hours. The reaction was concentrated in vacuo and carried onto the next step as a crude of the title compound. MS (m/z): 260 (M+H). [0070] The following compounds were prepared by the method of Preparation 12: Prep. Chemical nam MS (m/z) No. e Structure (M+H) 5-Bromo-N¹-(5- 13 chloropyrazolo[1,5- a]pyrimidin-7-yl)benzene- 340 1,3-diamine N¹-(5-Chloropyrazolo[1,5- 14 a]pyrimidin-7-yl)-5- methoxybenzene-1,3- 291 diamine tert-Butyl 6-((5- chloropyrazolo[1,5- 15 a]pyrimidin-7-yl)amino)- 1,3-dihydro-2H- 387 pyrrolo[3,4-c]pyridine-2- c^arboxylate 5-Chloro-N-(3- 16 nitrophenyl)pyrazolo[1,5- 290 a]pyrimidin-7-amine 5-Chloro-N-(3-methyl-5- 17 nitrophenyl)pyrazolo[1,5- 304 a]pyrimidin-7-amine 5-Chloro-N-(4-nitropyridin- 2-yl)pyrazolo[1,5- 291 a]pyrimidin-7-amine 5-Chloro-N-(5-nitropyridin- 3-yl)pyrazolo[1,5- 291 a]pyrimidin-7-amine tert-Butyl (2-((5- chloropyrazolo[1,5- a]pyrimidin-7-yl)amino)-6- 391 methoxypyridin-4- yl)carbamate 1H NMR (400 MHz, d6- DMSO) δ NH₂ 10.78 (s, N 1H), 8.25 N²-(5-Chloropyrazolo[1,5- (d, 1H), a]pyrimidin-7-yl)-4- O₂N NH 8.02 (s, nitropyridine-2,6-diamine N 1H), 7.52 N (s, 1H), N Cl 7.04 (s. 2H), 6.83 (d, 1H), 6.58 (s, 1H). 2-((5-Chloropyrazolo[1,5- a]pyrimidin-7-yl)amino)-6- (2- 364 methoxyethoxy)isonicotinic acid   2-((5-Chloropyrazolo[1,5- a]pyrimidin-7-yl)amino)-6- 23 (2-((tetrahydro-2H-pyran- 2- 434 yl)oxy)ethoxy)isonicotinic acid 24 6-Bromo-4-nitro-N- phenylpyridin-2-amine - Preparation 25 2-((5-Chloropyrazolo[1,5-a]pyrimidin-7-yl)amino)-6-fluoroisonicotinic acid [0071] To a solution of 5-

7-amine (349 mg, 2.07 mmol) in DMF (3 mL) was added NaH (226 mg, 5.66 mmol, 60.0% purity) at 0°C. After stirring the reaction mixture for 30 min, 2,6-difluoroisonicotinic acid (300 mg, 1.89 mmol) was added, the mixture was stirred at 90°C for 12 hours. The reaction mixture was quenched by addition H₂O (4 mL) at 0°C, and then adjusted pH = 2 with 2M HCl. The crude was filtered and the filter cake was obtained as a yellow solid. The residue was used directly for next step as the title compound (60.0 mg, 195 μmol, 10.3% yield). MS (m/z): 308 (M+H).     Preparation 26 5-Chloro-N-(5-methoxy-4-nitropyridin-2-yl)pyrazolo[1,5-a]pyrimidin-7-amine [0072] To a solution of 5- 7-amine (25.0 mg, 0.150 mmol) and 2-

bromo-5-methoxy-4-nitropyridine mg, in 1,4-dioxane (1.5 mL) was added NaOtBu (29.0 mg, 0.300 mmol) and tBuXPhos Pd G3 (24.0 mg, 0.030 mmol). The solution was flushed with N₂ for 15 minutes, and then allowed to heat at 110˚C for 12 hours. The reaction was concentrated with silica and purified through normal phase chromatography using a solid load method (0-100% EtOAc/hexane) to afford the title compound (27.0 mg, 57%, 0.084 mmol). MS (m/z): 331 (M+H). [0073] The following compounds were prepared by the method of Preparation 26: Prep. Chemical name Struct MS (m/z) No. ure (M+H) N²-(5-Chloropyrazolo[1,5- 27 a]pyrimidin-7-yl)-4-nitro- N⁶-phenylpyridine-2,6- - diamine Preparation 28 tert-Butyl (5-chloropyrazolo[1,5-a]pyrimidin-7-yl)(3-nitrophenyl)carbamate [0074] The title compound was

in Preparation 3 and was obtained as a white solid (312 mg, yield 80%). MS (m/z): 334 (M-55).     Preparation 29 5-Methoxy-N-(3-nitrophenyl)pyrazolo[1,5-a]pyrimidin-7-amine [0075] To a solution of 60% NaH MeOH (15 mL) was added tert-butyl (5- chloropyrazolo[1,5-a]pyrimidin-7-yl)

(310 mg, 0.80 mmol). The mixture was stirred at room temperature overnight. The mixture was quenched with H₂O (50 mL) and extracted with EtOAc (150 mL), the combined organic was washed with brine (100 mL), dried over anhydrous Na2SO4, filtered and concentrated to give the title compound as a white solid (200 mg, yield 88%). MS (m/z): 286 (M+H). Preparation 30 N⁵-(4-Methoxybenzyl)-N⁷-(3-nitrophenyl)pyrazolo[1,5-a]pyrimidine-5,7-diamine [0076] To a solution of 5-

a]pyrimidin-7-amine (500 mg, 1.73 mmol) and (4-methoxyphenyl)methanamine (356 mg, 2.60 mmol) in tBuOH (30 mL) was added DiPEA (670 mg, 5.19 mmol). The mixture was stirred at reflux for 48 h. After removal of the solvent, the residue was purified with prep-HPLC to give the title compound as a colorless oil (320 mg, yield 47%). MS (m/z): 391(M+H). Preparation 31 tert-Butyl (2-((5-chloropyrazolo[1,5- amino)-6-fluoropyridin-4-yl)carbamate

  [0077] To a solution of 2-((5-chloropyrazolo[1,5-a]pyrimidin-7-yl)amino)-6-fluoroisonicotinic acid (150 mg, 487 μmol) in tBuOH (3 mL) was added DPPA (147 mg, 536 μmol, 115 μL) and TEA (54.2 mg, 536 μmol, 74.6 μL) at 20°C. The mixture was stirred at 20°C for 1 hour. Then the mixture was stirred at 80°C for 11 hours. The reaction mixture was concentrated under reduced pressure to remove the solvent. The residue was diluted with NaHCO3 (aq, 5 mL) and extracted with EtOAc (15 mL) dried over Na₂SO₄, filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep-TLC (SiO₂, DCM) to obtain the title compound as a yellow solid (110 mg, 290 μmol, 59.5% yield). MS (m/z): 379 (M+H). [0078] The following compounds were prepared by the method of Preparation 31: Prep. Chemical nam MS (m/z) No. e Structure (M+H) tert-Butyl (2-((5- chloropyrazolo[1,5- 32 a]pyrimidin-7- yl)amino)-6-(2- 435 methoxyethoxy)pyridin- 4-yl)carbamate tert-Butyl (2-((5- chloropyrazolo[1,5- a]pyrimidin-7- 33 yl)amino)-6-(2- ((tetrahydro-2H-pyran- 505 2- yl)oxy)ethoxy)pyridin- 4-yl)carbamate Preparation 34 6-((5-Chloropyrazolo[1,5-a]pyrimidin-7-yl)amino)-4-nitropyridin-3-ol

  [0079] To a solution of 5-chloro-N-(5-methoxy-4-nitropyridin-2-yl)pyrazolo[1,5-a]pyrimidin-7- amine (27.0 mg, 0.0840 mmol) in 1,2-dichloroethane (0.780 mL) was added aluminum trichloride (170 mg, 1.20 mmol) and the reaction was heated at 80˚C for 30 minutes. The reaction was quenched with 5 mL of NaHCO3 and the product was subsequently extracted with EtOAc (15 mL). The organic layer was washed with brine, dried over Na2SO4, filtered, and concentrated in vacuo. The residue was purified using normal phase purification (0-20% MeOH/DCM) to afford the title compound (37.0 mg, 39%, 0.0600 mmol) as an oil. MS (m/z): 307 (M+H). Preparation 35 N²-(5-Chloropyrazolo[1,5-a]pyrimidin-7-yl)pyridine-2,4-diamine [0080] To a solution of 5-chloro-N-(4-nitropyridin-2-yl)pyrazolo[1,5-a]pyrimidin-7-amine (526 mg, 1.81 mmol) in EtOH (30 mL) and H2O (1 mL) was added Fe powder (484 mg, 18.1 mmol) and NH₄Cl (484 mg, 9.05 mmol). The mixture was stirred at reflux overnight. After cooled down to room temperature, the mixture was filtered through Celite, the cake was washed with EtOH (20 mL). The combined filtrate and washing were evaporated to leave the crude product, which was purified by prep-HPLC to obtain the title compound as a white solid (245 mg, yield 52%). MS (m/z): 261 (M+H). [0081] The following compounds were prepared by the method of Preparation 35: Prep. Che MS (m/z) No. mical name Structure (M+H) N¹-(5- 36 Methoxypyrazolo[1,5- a]pyrimidin-7- 256 yl)benzene-1,3-diamine N¹-(5- Chloropyrazolo[1,5- 37 a]pyrimidin-7-yl)-5- 274 methylbenzene-1,3- diamine 4-Amino-6-((5- chloropyrazolo[1,5- a]pyrimidin-7- 277 yl)amino)pyridin-3-ol N³-(5- Chloropyrazolo[1,5- a]pyrimidin-7- 261 yl)pyridine-3,5- diamine NH₂ N²-(5- N Chloropyrazolo[1,5- a]pyrimidin-7- H₂N NH 276 yl)pyridine-2,4,6- N triamine N N Cl N²-(5- Chloropyrazolo[1,5- a]pyrimidin-7-yl)-N⁶- 316 cyclopropylpyridine- 2,4,6-triamine N²-(5- Chloropyrazolo[1,5- a]pyrimidin-7-yl)-N⁶- - phenylpyridine-2,4,6- triamine Preparation 43 N⁷-(3-Aminophenyl)-N⁵-(4-methoxybenzyl)pyrazolo[1,5-a]pyrimidine-5,7-diamine   [0082] To a solution of N⁵-(4-methoxybenzyl)-N⁷-(3-nitrophenyl)pyrazolo[1,5-a]pyrimidine-5,7- diamine (320 mg, 0.82 mmol) in MeOH (20 mL) was added 10 % Pd/C (200 mg). The mixture was stirred at room temperature under hydrogen (1 atm) overnight. The mixture was filtered and concentrated to leave crude of the title compound as a yellow solid (226 mg, yield 77%). MS (m/z): 361 (M+H). Preparation 44 N⁷-(3-Aminophenyl)pyrazolo[1,5-a]pyrimidine-5,7-diamine [0083] To a solution of N⁷-(3-

pyrazolo[1,5-a]pyrimidine-5,7- diamine (226 mg, 0.63 mmol) in THF (10 mL) was added TFA (5.0 mL). The mixture was stirred at room temperature overnight. After removal of the solvent, the residue was purified with prep-HPLC to afford the title compound as a brown solid (40 mg, yield 27%). MS (m/z): 241 (M+H) [0084] The following compounds were prepared by the method of Preparation 44: Prep. Chemical nam MS (m/z) No. e Structure (M+H) F N²-(5- N Chloropyrazolo[1,5- 45 a]pyrimidin-7-yl)-6- H₂N NH 279 fluoropyridine-2,4- N diamine N N Cl 5-Chloro-N-(2,3- dihydro-1H-pyrrolo[3,4- 46 c]pyridin-6- 287 yl)pyrazolo[1,5- a]pyrimidin-7-amine N²-(5- Chloropyrazolo[1,5- a]pyrimidin-7-yl)-6-(2- 335 methoxyethoxy)pyridine- 2,4-diamine N²-(5- Chloropyrazolo[1,5- a]pyrimidin-7-yl)-6- 290 methoxypyridine-2,4- diamine 1H NMR (400 MHz, d6- DMSO) δ 7.34 (d, N²-(4-Methoxybenzyl)- 1H), 7.18 4-nitropyridine-2,6- (dd, 2H), diamine 6.82 (dd, 2H), 6.27 (s, 1H), 4.38 (d, 2H), 3.71 (s, 3H) Preparation 50 6-(2-((tert-Butyldimethylsilyl)oxy)ethoxy)-N²-(5-chloropyrazolo[1,5-a]pyrimidin-7- yl)pyridine-2,4-diamine

  [0085] To a solution of tert-butyl (2-((5-chloropyrazolo[1,5-a]pyrimidin-7-yl)amino)-6-(2- ((tetrahydro-2H-pyran-2-yl)oxy)ethoxy)pyridin-4-yl)carbamate (1.82 g, 3.98 mmol) in HCl/MeOH (4.00 M, 995 μL). The mixture was stirred at 25°C for 1 hour. The mixture was concentrated under reduced pressure, then dissolved DCM (15 mL) and added TEA (1.21 g, 11.9 mmol, 1.66 mL), DMAP (97.2 mg, 796 μmol), TBSCl (660 mg, 4.38 mmol, 539 μL). The mixture was stirred at 25°C for 1 hour. The reaction mixture was quenched by addition H₂O (20 mL) at 25°C, and then extracted with DCM (30 mL). The combined organic layers were washed with brine (20 mL), dried over Na₂SO₄, filtered and concentrated under reduced pressure to give the title product as a green solid (2.56 g, crude) was used into the next step without further purification. MS (m/z): 435 (M + H). Example 1 N-(3-((5-Chloropyrazolo[1,5-a]pyrimidin-7-yl)amino)phenyl)acrylamide (NJH-04-21) Cl [0086] To a crude solution of

a]pyrimidin-7-yl)benzene-1,3-diamine (259.7 mg, 1.00 mmol) in THF (2 mL) was added an aqueous solution of saturated sodium bicarbonate (2 mL). The mixture was cooled to 0˚C and acryloyl chloride (0.08 mL, 1.00 mmol) was added dropwise and the reaction was allowed to stir for 15 minutes. The reaction was concentrated in vacuo and purified by prep-HPLC (85% to 10% H₂O (0.05% TFA) to MeOH (0.05% TFA)) to afford the title compound (226.3 mg, 72%, 0.072 mmol) as a white solid. MS (m/z): 314 (M+H). [0087] The following compounds were prepared by the method of Example 1: Example MS No. Chemical name Structure (m/z) (M+H) O N-(3-((5-Chloropyrazolo[1,5- a]pyri N NH 2 midin-7- H yl)amino)phenyl)propionamide 316 N N (DGY-12-162) N Cl N-(3-((5-Chloropyrazolo[1,5- a]pyrimidin-7-yl)amino)-5- methylphenyl)acrylamide 328 (C-5) N-(3-Bromo-5-((5- chloropyrazolo[1,5-a]pyrimidin- 7-yl)amino)phenyl)acrylamide 394 (DGY-13-019) N-(3-((5-Chloropyrazolo[1,5- a]pyrimidin-7-yl)amino)-5- methoxyphenyl)acrylamide 344 (DGY-13-034) O N N-(2-((5-Chloropyrazolo[1,5- a]pyrimidin-7-yl)amino)pyridin- N NH H 4-yl)propionamide 317 N N (MNN-03-006) N Cl N-(2-((5-Chloropyrazolo[1,5- a]pyrimidin-7-yl)amino)-6- fluoropyridin-4-yl)acrylamide 333 (E5) 1-(6-((5-Chloropyrazolo[1,5- a]pyrimidin-7-yl)amino)-1,3- dihydro-2H-pyrrolo[3,4- 341 c]pyridin-2-yl)prop-2-en-1-one (E6) N-(2-((5-Chloropyrazolo[1,5- a]pyrimidin-7-yl)amino)-6-(2- methoxyethoxy)pyridin-4- 389 yl)acrylamide (E9) N-(2-((5-Chloropyrazolo[1,5- a]pyrimidin-7-yl)amino)pyridin- 4-yl)acrylamide 315 (C3) N-(3-((5-Methoxypyrazolo[1,5- a]pyrimidin-7- yl)amino)phenyl)acrylamide 310 (C7) N-(3-((5-Aminopyrazolo[1,5- a]pyrimidin-7- yl)amino)phenyl)acrylamide 295 (C8) N-(2-((5-Chloropyrazolo[1,5- a]pyrimidin-7-yl)amino)-6- methoxypyridin-4-yl)acrylamide 345 (T3) N-(2-Amino-6-((5- chloropyrazolo[1,5-a]pyrimidin- 7-yl)amino)pyridin-4- 330 yl)acrylamide (T4)   Example 15 N-(2-((5-Chloropyrazolo[1,5-a]pyrimidin-7-yl)amino)-6-(2-hydroxyethoxy)pyridin-4-yl)acrylamide (E11) [0088] To a solution of 6-(2-

oxy)ethoxy)-N²-(5-chloropyrazolo[1,5- a]pyrimidin-7-yl)pyridine-2,4-diamine (1.50 g, 3.45 mmol), Na2CO3 (1.10 g, 10.3 mmol) in THF (15 mL) acryloyl chloride was added (234 mg, 2.59 mmol, 210 μL) at 0°C. The mixture was stirred at 25°C for 1 hour. Then the reaction mixture was concentrated under reduced pressure to give a residue, added DCM (1 mL) and TFA (0.330 mL) at 25°C for 1 hour. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (FA condition) to obtain the title compound as a white solid (39.0 mg, 79.3 μmol, 2.30% yield, 99.4% purity, TFA). MS (m/z): 375 (M+H). Example 16 N-(2-((5-chloropyrazolo[1,5-a]pyrimidin-7-yl)amino)-5-hydroxypyridin-4-yl)acrylamide (MNN-02-143) [0089] To a solution of acrylic

mmol) in DMF (1.11 mL) was added DiPEA (0.0289 mL, 0.0166 mmol) and HATU (33.6 mg, 0.0884 mmol) at 0˚C and the reaction was allowed to stir for 15 minutes. Subsequently 4-amino-6-((5-chloropyrazolo[1,5-a]pyrimidin-7- yl)amino)pyridin-3-ol (15.3 mg, 0.0553 mmol) was added and the reaction was let to stir for 12 hours. The reaction was purified by prep-HPLC (85% to 10% H2O (0.05% TFA) to MeOH (0.05% TFA)) to afford the title compound as a white solid (0.420 mg, 2%, 0.0130 mmol). MS (m/z): 331 (M+H). [0090] The following compounds were prepared by the method of Example 16:   Example MS No. Chemical name Structure (m/z) (M+H) N-(5-((5-Chloropyrazolo[1,5- 17 a]pyrimidin-7-yl)amino)pyridin- 3-yl)acrylamide 315 (T1) NH N-(2-((5-Chloropyrazolo[1,5- a]pyrimidin-7-yl)amino)-6- O N 18 (cyclopropylamino)pyridin-4- 370 yl)acrylamide N NH H (T15) N N N Cl N-(2-((5-Chloropyrazolo[1,5- a]pyrimidin-7-yl)amino)-6- 19 (phenylamino)pyridin-4- 406 yl)acrylamide (T17) Protein expression and purification [0091] C-terminal 6x His-tag constructs of human wild type (WT) or C102S full-length (residues 1-595) and residues 1-524 SHP-1 were expressed using pET 21b(+) vector in Rosetta 2 and purified via affinity and size-exclusion chromatography. Briefly, transformed cells were cultured at 37^C in Terrific Broth (TB) medium for 6h, and auto-induced at 20^C for 18 h. Cell pellets were lysed by high pressure microfluidizer in lysis buffer containing 50 mM HEPES, 500 mM NaCl, 1 mM TCEP, 20 mM imidazole, pH 7.5, 0.1% IGEPAL, 10% glycerol, 1 mM phenylmethylsulfonyl fluoride (PMSF). Following lysis, the supernatant was applied to Ni-NTA for affinity binding at 4^C for 1 h with rocking, and the resin was washed by fast protein liquid chromatography (FPLC) with wash buffer containing 25 mM HEPES, 1.5 M NaCl, 1 mM TCEP, 20 mM imidazole, pH 7.5, and eluted with elution buffer containing 25 mM HEPES, 500 mM NaCl, 1 mM TCEP, 400 mM imidazole, 10% glycerol, pH 7.5. The final protein was purified by size exclusion using a HiLoad 16/600 Superdex 200 pg column (Cytiva) in buffer containing 20 mM HEPES, 200 mM NaCl, 1 mM TCEP, pH 7.5. The resultant SHP-1 containing fractions were concentrated to 10 mg/ml and stored at -80^C.   Intact protein mass spectrometry [0092] WT or C102S SHP-1 recombinant protein (final concentration 2 ^M) was incubated with 5 equivalents (10 ^M) or 10 equivalents (20 ^M) of the indicated SHP-1 agonists at 4^C for 24 h in reaction buffer (25 mM HEPES, pH 7.4, 50 mM NaCl, 2.5 mM EDTA, 100 ^M TCEP). The samples were then diluted 1 in 2 with Buffer A (0.1% formic acid, 5% acetonitrile, 95% H20), and 3.5 ^g per sample was subjected to LC-MS analysis using a PLRP-S 100A, 2.1 x 50 mM, 5 ^M column (Agilent) on a Q-Exactive HF-X. Separation buffers A (0.1% formic acid, 5% acetonitrile, 95% H20) and B (0.1% formic acid in 95% acetonitrile, 5% H₂0) were used for the runs at 60^C, 0.3 ml min^-1. The gradient used was 15% Buffer B, 0.5 min, ramped up to 95% over 4.5 min. Positive ion modes were collected with full scan analysis over 900 – 2600 m/z at 7500 resolution, 1 x 10⁵ AGC, 25 ms maximum ion accumulation time and 60 eV in-source collision-induced dissociation. Data processing was performed using Thermo Biopharma Finder software version 4.1 (Thermo Fisher Scientific). All exemplified compounds were tested essentially as described above and the data from these experiments are as displayed in the following table. Data for the compound of Example 1 (NJH-04- 021) are shown in Figures 2A – 2F). E_{XAMPLE # COMPOUND #} ^{10 ^M (5 eq) Incubation} 20 ^M (10 eq) Incubation (% binding) (% binding) 1 NJH-04-021 15.2% 46.3% 2 DGY-12-162 - - 3 C5 10.8% 29.6% 4 DGY-13-019 100% 100% 5 DGY-13-034 17.4% 37.1% 6 MNN-03-006 - - 7 E5 100% 100% 8 E6 12.5% 12% 9 E9 21.1% 25% 10 C3 88.7% 100% 11 C7 16.7% 31.9% 12 C8 - 10.1% 13 T3 39.3% 47.5% 14 T4 42.2% 70.7%   15 E11 24.8% 24% 16 MNN-02-143 100% 100% 17 T1 - 20.9% 18 T15 3.6% 13.8% 19 T17 30.6% 54.1% SHP-1 Phosphatase Activity Assay (Figures 1A – 1D) [0093] SHP-1 phosphatase activity was assessed by monitoring dephosphorylation of the synthetic substrate DiFMUP (6,8-Difluoro-4-Methylumbelliferyl Phosphate) (Thermo Fisher Scientific, #D6567) to the fluorogenic product DiFMU (6,8-Difluoro-7-Hydroxy-4-Methylcoumarin) (Thermo Fisher Scientific, #D6566). Briefly, 20 nM recombinant SHP-1 was incubated with NJH-04-021, DGY-12-162, C-3 or MNN-3-006 at the indicated concentrations for 30 min at room temperature in activity buffer (25 mM HEPES, pH 7.4, 50 mM NaCl, 2.5 mM EDTA, 100 ^M TCEP). Following incubation, the reaction was initiated by addition of 20 ^l enzyme to 80 ^l DiFMUP substrate (final concentration 50 ^M) in a 96-well plate (Corning, #3603) at 37^C. Fluorescence at excitation/emission wavelengths of 355/460 nm was continuously measured in a fluorescence-based SpectraMax M5 microplate reader (Molecular Devices) and converted to phosphate equivalents released using a DiFMU reference standard curve. The turnover to one picomole of DiFMU is equivalent to release of one picomole of phosphate. Rate of SHP-1 activity in the presence of each agonist was calculated as picomole phosphate release per min in fold change relative to DMSO control, and the rates fitted on a non-linear regression model on GraphPad Prism 9, with calculated EC50 and Vmax of the reactions as indicated. [0094] Compounds of the invention were tested essentially as described above and representative data are presented in the following table: EXAMPLE COMPOUND # FIGURE EC₅₀ (µM) V_max (pmol/min) 1 NJH-04-021 1A 28.2 14.2 2 DGY-12-162 1B - 1.8 6 MNN-03-006 1D - 4.5 10 C-3 1C 12.2 12.2 Cellular NF^B activation assay [0095] iBMDMs seeded in 24-well plates at 2x10⁶/ml were allowed to adhere overnight. The following day, cells were pre-treated with the indicated concentrations of compounds for 3 h,   followed by lipopolysaccharide (LPS) (InvivoGen, LPS-EK Ultrapure, #tlrl-peklps) induction at 100 ng/ml for 15 min. Cells were washed with ice-cold phosphate-buffered saline (PBS) and lysed in 80 μl/well of Triton-lysis buffer (25 mM Tris-HCl, pH 7.5, 100 mM NaCl, 2.5 mM EDTA, 2.5 mM EGTA, 20 mM NaF, 1 mM Na3VO4, 20 mM sodium β-glycerophosphate, 10 mM sodium pyrophosphate, 0.5% Triton X-100, Roche EDTA-free protease inhibitor cocktail (Roche, #11836170001) and 0.1% β-mercaptoethanol). Lysates were pre-cleared by centrifugation at 21,000 g, 10 min, 4°C, reduced with 4x NuPAGE LDS sample buffer (Invitrogen, #NP0007) containing 5% β-mercaptoethanol, and boiled at 95°C for 10 min. Equal amounts of proteins were resolved using NuPAGE 4-12% Bis-Tris 15-well or 26-well gels (Invitrogen, #NP0336BOX or #WG1403BOX), transferred onto PVDF membranes (Thermo Fisher Scientific, iBlot 2 Gel Transfer Device, #IB21001), and immunoblotted with anti-IRAK1 (Cell Signaling, #4504, rabbit mAb; 1:1000 dilution), anti-phospho-I^B^ (Ser32/36) (Cell Signaling, #9246; mouse mAb; 1:1000 dilution), anti- I^B^ (Cell Signaling, #9242, rabbit pAb; 1:1000 dilution), anti-phospho-NF^B p65 (Ser536) (Cell Signaling, #3033, rabbit mAb; 1:1000 dilution), anti-NF^B p65 (Cell Signaling, #8242, rabbit mAb; 1:1000 dilution), and anti-β-actin (Cell Signaling, #3700, mouse mAb; 1:3000 dilution) antibodies, diluted in TBS-T (Boston BioProducts, Inc., #IBB-180X) containing 5% BSA, followed by incubation with horseradish peroxidase-conjugated anti-rabbit or anti-mouse secondary antibodies (Promega, #W4011 or #W4021), diluted 1:10000 in TBS-T containing 5% milk, and visualized using ECL western blotting substrates (Pierce ECL Western Blotting Substrate, #32106). Representative compounds of the invention were tested essentially as described above and data from these tests are illustrated in Figures 3A, 3B, 4, and 6. Cytokine production quantification via ELISA assays [0096] iBMDMs seeded in 96-well plates at 2x10⁶/ml were allowed to adhere overnight. The following day, cells were pre-treated with the compound of Example 1 (NJH-04-021) or Example 10 (C-3) at concentrations from 10¹ to 10⁶ nM for 3 h, followed by LPS induction at 100 ng/ml for 6 h. IL-6 and TNF^ cytokine levels in cell culture supernatants were quantified using mouse IL-6 or TNF^ DuoSet ELISA kits (R&D Systems, #DY406 or #DY410) according to manufacturer’s protocols. Absorbance values at 450 and 540 nm were determined with a microplate reader (BMG LABTECH FLUOstar Omega microplate reader), and wavelength corrected absorbances were used to plot standard curves from which cytokine levels from supernatant samples were calculated by intrapolation using a 4-parameter logistic (4PL) regression on GraphPad Prism 9. For IL-1^   measurement using mouse IL-1^/IL-1F2 DuoSet ELISA kit (R&D Systems, #DY401), cells were pre-treated with 100 ng/ml LPS for 3 h, medium refreshed and treated with NJH-04-021 and C-3 at a range of concentrations for 45 min, followed by treatment with 5 mM ATP for 45 min. IL-1^ levels were then assessed from cell culture supernatants. IC50 values for each compound are recorded in the following tables. Inhibition of IL-1β Production EXAMPLE IC50 (µM) 1 153.3 10 6.7 Inhibition of IL-6 Production EXAMPLE IC50 (µM) 1 10.8 10 7.4 Inhibition of TNFα Production EXAMPLE IC50 (µM) 1 11.7 10 12.7

Claims

  CLAIMS We claim: 1. A compound of Formula I: wherein:

R¹ is halo, amino, or C1 – C4 alkoxy; ;

X is phenyl or pyridinyl optionally substituted with one substituent selected from halo, hydroxy, C1 – C4 alkyl, -NHR^a wherein R^a is H, C3 – C6 cycloalkyl, or phenyl, or C1 – C4 alkoxy optionally substituted with hydroxy or C₁ – C₄ alkoxy; or a pharmaceutically acceptable salt thereof. 2. A compound of Claim 1 or a pharmaceutically acceptable salt thereof wherein R¹ is halo; R² is -X-NHR³; and R⁴ is C₂ – C₄ alkenyl. 3. A compound of any of Claims 1 – 2 or a pharmaceutically acceptable salt thereof wherein R¹ is chloro. 4. A compound of any of Claims 1 – 3 or a pharmaceutically acceptable salt thereof wherein R⁴ is ethenyl. 5. A compound of any of Claims 1 – 4 or a pharmaceutically acceptable salt thereof wherein X is phenyl or pyridinyl each optionally substituted with one substituent selected from the group consisting of halo and hydroxy.   6. A compound of any of Claims 1 – 5 or a pharmaceutically acceptable salt thereof wherein X is phenyl or pyridinyl. 7. A compound of any of Claims 1 – 4 or a pharmaceutically acceptable salt thereof wherein X is pyridinyl optionally substituted with -NHR^a wherein R^a is C₃ – C₆ cycloalkyl or phenyl. 8. A pharmaceutical composition comprising a compound of any of Claims 1 – 7 or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier. 9. A compound of any of Claims 1 – 7 or a pharmaceutically acceptable salt thereof for use in therapy. 10. The use of a compound of any of Claims 1 – 7 or a pharmaceutically acceptable salt thereof for the manufacture of a medicament for the treatment of autoimmune disorders. 11. The use of a compound of any of Claims 1 – 7 or a pharmaceutically acceptable salt thereof for the manufacture of a medicament for the treatment of a SHP-1 susceptible cancer selected from bladder carcinoma, breast cancer, cervical squamous cell carcinoma, kidney renal papillary cell carcinoma, lung adenocarcinoma, pancreatic ductal adenocarcinoma, sarcoma, stomach adenocarcinoma, uterine corpus endometrial carcinoma, triple negative breast cancer, and colorectal cancer. 12. A method for treating autoimmune disorders comprising administering to a patient in need thereof an effective amount of a compound of any of Claims 1 -7 or a pharmaceutically acceptable salt thereof. 13. A method for treating a SHP-1 susceptible cancer selected from bladder carcinoma, breast cancer, cervical squamous cell carcinoma, kidney renal papillary cell carcinoma, lung adenocarcinoma, pancreatic ductal adenocarcinoma, sarcoma, stomach adenocarcinoma, uterine corpus endometrial carcinoma, triple negative breast cancer, and colorectal cancer comprising   administering to a patient in need thereof an effective amount of a compound of any of Claims 1 – 7 or a pharmaceutically acceptable salt thereof.