T-type calcium channel antagonists and uses thereof

Abstract

A method of reducing Miro1 levels in a cell is disclosed, comprising contacting the cell with an effective amount of a T-type calcium channel antagonist.

Description

[Technical Field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims priority to U.S. Provisional Application No. 63 / 156,806, filed March 4, 2021, and U.S. Provisional Application No. 63 / 228,505, filed August 2, 2021, each of which is incorporated herein in its entirety for all purposes. [Background technology]

[0002] Neurons are metabolically active cells with high energy demands far from the cell body. Consequently, these cells are particularly dependent on mitochondrial function, as reflected by the finding that mitochondrial dysfunction disorders often have a neurodegenerative component. Recent discoveries highlight the neuron's particular dependence on the dynamic properties of mitochondria. Mitochondria are dynamic organelles by several criteria. They participate in repeated cycles of fusion and fission, which serve to mix the contents of the mitochondrial mass with lipids. In addition, mitochondria are actively recruited to subcellular sites, such as neuronal axons and dendrites. Finally, the quality of the mitochondrial mass is maintained through mitophagy, a form of autophagy in which defective mitochondria are selectively degraded (mitochondrial quality control). Defects in key features of mitochondrial dynamics, such as mitochondrial fusion, fission, transport, and mitophagy, have been associated with neurodegenerative disorders. Several major neurodegenerative disorders, including Parkinson's disease, Alzheimer's disease, and Huntington's disease, involve disruptions in mitochondrial dynamics.

[0003] Mitochondrial movement is tightly regulated to maintain energy homeostasis and prevent oxidative stress. Mitochondrial movement ceases before the initiation of mitophagy, a key cellular mechanism by which depolarized mitochondria are degraded and digested via autophagosomes and lysosomes. Cessation of movement sequesters damaged mitochondria, immobilizing them and preventing the reintroduction of damage to other healthy mitochondria.

[0004] Miro is a mitochondrial outer membrane (OMM) protein that anchors the microtubule motors kinesin and dynein to mitochondria (Glater et al., “Axonal transport of mitochondria requires milton to recruit kinesin heavy chain and is light chain independent,” The Journal of Cell Biology (2006) 173:545-557; and Koutsopoulos et al., “Human Miltons associate with mitochondria and induce microtubule-dependent remodeling of mitochondrial networks,” Biochimica et biophysica acta (2010) 1803:564-574). This depolarization-induced mitochondrial arrest is achieved by removing Miro from the surface of damaged mitochondria (Wang et al., “PINK1 and Parkin target Miro for phosphorylation and degradation to arrest mitochondrial motility,” Cell (2011) 147:893-906). Miro is then degraded by the proteasome ( Wang et al., 2011 ).Evidence suggests that two PD-associated proteins, PINK1 (PTEN-induced putative kinase 1) and Parkin, cooperate to target Miro for degradation (Ashrafi et al., “Mitophagy of damaged mitochondria occurs locally in distal neuronal axons and requires PINK1 and Parkin,” The Journal of cell biology (2014) 206:655-670; Liu et al., “Parkinson's disease-associated kinase PINK1 regulates Miro protein level and axonal transport of mitochondria,” PLoS genetics (2012) 8:e1002537; and Wang et al., 2011). Mutations in PINK1 or Parkin are associated with rare forms of recessive, early-onset PD.

[0005] Alterations in mitochondrial transport function are one of the pathogenic alterations in major adult-onset neurodegenerative diseases (Sheng ZH, Cai Q, “Mitochondrial transport in neurons: impact on synaptic homeostasis and neurodegeneration,” Nat. Rev. Neurosci., (2012) Jan. 5;13(2):77-93). In mutant LRRK 2GS2019 cells, the mitochondrial outer membrane protein Miro is stabilized and remains on damaged mitochondria longer than normal, prolonging active transport and inhibiting mitochondrial degradation (Hsieh et al., “Functional Impairment in Miro Degradation and Mitophagy Is a Shared Feature in Familial and Sporadic Parkinson's Disease,” Cell Stem Cell (2016) Dec. 1;19(6):709-724). Miro degradation and mitochondrial motor function are also impaired in sporadic PD patients (Hsieh et al., 2016). Persistent retention of Miro and its subsequent downstream consequences may constitute a central element in PD pathogenesis.

[0006] T-type voltage-dependent calcium channels, or T-type calcium channels, are low-voltage-activated calcium channels that are inactivated during hyperpolarization of the cell membrane, but subsequently open upon depolarization. There are three known subtypes of T-type calcium channels: Ca V lc3.1, Ca V lc3.2, and Ca V lc3.3 is present. Calcium influx into various cells results in a number of different associated physiological responses. Activation of voltage-dependent calcium channels in cardiac and smooth muscle cells directly initiates contraction by increasing cytoplasmic calcium concentration. T-type calcium channels are present not only in cardiac and smooth muscle cells, but also in many neuronal cells within the central nervous system.

[0007] Novel compounds and methods for reducing intracellular Miro1 levels are needed to treat neurodegenerative diseases, such as Parkinson's disease. Summary of the Invention

[0008] In one aspect, the present disclosure provides a compound of formula (II): [ka] or a pharmaceutically acceptable salt thereof, wherein Ring B is C6 to C 10 aryl or 5-10 membered heteroaryl; R 1 , R 2 and R 3 are each independently H, halogen, CN, OR 11 , N.R. 12a R 12b , C1-C6 alkyl, C2-C6 alkoxyalkyl, C1-C6 haloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, phenyl, 5-6 membered heteroaryl, C3-C7 cycloalkyl, -(CH2) n -(C3-C7 cycloalkyl), 4- to 7-membered heterocyclyl, or -(CH2) n -(4- to 7-membered heterocyclyl), wherein said phenyl, heteroaryl, cycloalkyl or heterocyclyl optionally contains one, two or three halogen atoms, CN, OR 11 , N.R. 12a R 12b , optionally substituted by C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylamino, or C1-C6 haloalkyl; R 4a and R 4b are each independently H or C1-C6 alkyl, or R 4a and R 4b and the carbon atoms to which they are attached together form a C3-C5 cycloalkyl; R 6 is H, C1-C6 alkyl, or C1-C6 haloalkyl; X1 and X 5 are each independently N or CR 8 where X 1 and X 5 At least one of the following is CR 8 and; X 2 , X 3 and X 4 are each independently N, NR 9 , O, S or CR 9 where X 2 , X 3 and X 4 At least one of the following is N or NR 9 , O, or S; R 8 is H, halogen, CN, OR 11 , C1-C6 alkyl, C2-C6 alkoxyalkyl, or C1-C6 haloalkyl; R 9 is H, C1-C6 alkyl, C2-C6 alkoxyalkyl, C1-C6 haloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, or 4-7 membered heterocyclyl, wherein said cycloalkyl or heterocyclyl may be optionally substituted by 1, 2, or 3 halogen, CN, OH, NH2, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylamino, or C1-C6 haloalkyl; Each R 11 , R 12a and R 12b are each independently H, C1-C6 alkyl, C2-C6 alkoxyalkyl, C1-C6 haloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, or 4-7 membered heterocyclyl, wherein said cycloalkyl or heterocyclyl is optionally substituted by 1, 2, or 3 halogen or CN; and n is 1, 2 or 3; provided that the compound has the following structure: [ka] [ka] or [ka] or a pharmaceutically acceptable salt thereof provided that the compound does not have

[0009] The compound of formula (II) is X 1 is CR 8 For example, R 8 can be H. Compounds of formula (II) each independently represent N, NR 9 , or CR 9 is X 2 , X 3 , and X 4 may contain, but X 2 , X 3 , and X 4 At least one of the following is N or NR 9 This is subject to the condition that:

[0010] The present disclosure also provides a compound of formula (III): [ka] (In the formula, R 1 , R 2 , R 3 , R 4a , R 4b , R 6 , R 8 , and R 9 is as defined elsewhere herein) or a pharmaceutically acceptable salt thereof.

[0011] The compounds of formula (II) or (III), or pharmaceutical salts thereof, each independently represent R which is H, halogen, CN, OH, NH, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylamino, C2-C6 alkoxyalkyl, C1-C6 haloalkyl, or C3-C7 cycloalkyl.1 , R 2 and R 3 wherein said cycloalkyl is optionally substituted by 1, 2, or 3 halogen, CN, OH, NH, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylamino, or C1-C6 haloalkyl.

[0012] The compounds of formula (II) or (III) or pharmaceutical salts thereof may be R 1 and R 2 are each independently H, halogen, C1-C6 alkyl, C1-C6 alkoxy, C2-C6 alkoxyalkyl, C1-C6 haloalkyl, or C3-C7 cycloalkyl, where the cycloalkyl is optionally substituted with 1, 2, or 3 halogen, CN, OH, NH2, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylamino, or C1-C6 haloalkyl. For example, R 1 and R 2 can each independently be H, halogen, C1-C3 alkyl, C1-C3 alkoxy, C2-C3 alkoxyalkyl, C1-C3 haloalkyl, or C3-C5 cycloalkyl, wherein the cycloalkyl is optionally substituted with 1, 2, or 3 halogen, CN, C1-C3 alkyl, C1-C3 alkoxy, C1-C3 alkylamino, or C1-C3 haloalkyl.

[0013] The compounds of formula (II) or (III) or pharmaceutical salts thereof each independently have R which is H or CH 4a and R 4b It can have:

[0014] The compounds of formula (II) or (III) or pharmaceutical salts thereof may comprise R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , R 8 , R 9 , R 10 , R 11 , R 12 , R 13 , R 14 , R 15 , R 16 , R 17 , R 18 , R 19 , R 20 6 For example, R 6 can be CH3.

[0015] The present disclosure also provides a compound of formula (IV): [ka] (In the formula, R 1 , R 2 , R 4a , R 6 and R 9 is as defined anywhere herein) or a pharmaceutically acceptable salt thereof.

[0016] The compounds of formula (II), (III) or (IV) or pharmaceutical salts thereof may comprise R 1 -C 6 alkyl, C 2 -C 6 alkoxyalkyl, C 1 -C 6 haloalkyl, C 2 -C 6 alkenyl, C 2 -C 6 alkynyl, C 3 -C 7 cycloalkyl, or 4-7 membered heterocyclyl. 9 wherein the cycloalkyl or heterocyclyl is optionally substituted with 1, 2, or 3 halogen, CN, OH, NH, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylamino, or C1-C6 haloalkyl. For example, R 9 can be C1-C6 alkyl, C2-C6 alkoxyalkyl, C1-C6 haloalkyl, C3-C7 cycloalkyl, or 4-7 membered heterocyclyl, wherein said cycloalkyl or heterocyclyl is optionally substituted by 1, 2, or 3 halogen, CN, OH, NH2, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylamino, or C1-C6 haloalkyl.

[0017] The compounds of formula (II), (III), or (IV) or pharmaceutical salts thereof can have ring B that is phenyl or 5-6 membered heteroaryl. For example, ring B can be phenyl or pyridyl.

[0018] The present disclosure also provides a compound of formula (V): [ka] or a pharmaceutically acceptable salt thereof, wherein R 1 , R 2 , R6 , and R 9 is as defined elsewhere herein.

[0019] Exemplary T-type calcium channel antagonists (antagonists) of the present disclosure can have the structure of any one of the compounds in Table 1. Additional structures of compounds of the present disclosure can be found in Table 2.

[0020] The present disclosure includes pharmaceutical compositions of the compounds described herein or pharmaceutically acceptable salts thereof.

[0021] Further provided herein is a method of treating a neurodegenerative disease in a subject (also referred to as a patient) comprising administering a compound described herein or a pharmaceutically acceptable salt thereof.

[0022] In another embodiment, a method for reducing Miro1 levels in a cell is described, comprising contacting the cell with an effective amount of a T-type calcium channel antagonist. The T-type calcium channel antagonist can have at least about 1.2-fold or greater selectivity for T-type calcium channels over one or more of L-type, N-type, P-type, and / or R-type calcium channels. The method includes contacting the cell with a T-type calcium channel antagonist having the structure of formula (I): [ka] or a pharmaceutically acceptable salt thereof, Ring A and ring B each independently represent a C6 to C 10 aryl or 5-10 membered heteroaryl; R 1 , R 2 and R 3 are each independently H, halogen, CN, OR 11 , N.R. 12a R 12b, C1-C6 alkyl, C2-C6 alkoxyalkyl, C1-C6 haloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, phenyl, 5-6 membered heteroaryl, C3-C7 cycloalkyl, or 4-7 membered heterocyclyl, wherein said phenyl, heteroaryl, cycloalkyl, or heterocyclyl optionally contains one, two, or three halogen atoms, CN, OR 11 , N.R. 12a R 12b , optionally substituted by C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylamino, or C1-C6 haloalkyl; R 4a and R 4b are each independently H or C1-C6 alkyl; R 5 is H or C1-C6 alkyl; R 6a and R 6b are each independently H or C1-C6 alkyl; R 7 is H, halogen, CN, OR 11 , N.R. 12a R 12b , C1-C6 alkyl, C2-C6 alkoxyalkyl, C1-C6 haloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, phenyl, 5-6 membered heteroaryl, C3-C7 cycloalkyl, or 4-7 membered heterocyclyl, wherein said phenyl, heteroaryl, cycloalkyl, or heterocyclyl optionally contains 1, 2, or 3 halogen atoms, CN, OR 11 , N.R. 12a R 12b , optionally substituted by C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylamino, or C1-C6 haloalkyl; R 8is H, C1-C6 alkyl, C2-C6 alkoxyalkyl, C1-C6 haloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, or 4-7 membered heterocyclyl, wherein said cycloalkyl or heterocyclyl is optionally substituted by 1, 2, or 3 halogen, CN, OH, NH2, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylamino, or C1-C6 haloalkyl; R 9 is H, C1-C6 alkyl, C2-C6 alkoxyalkyl, C1-C6 haloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, or 4-7 membered heterocyclyl, wherein said cycloalkyl or heterocyclyl is optionally substituted by 1, 2, or 3 halogen, CN, OH, NH2, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylamino, or C1-C6 haloalkyl; and Each R 11 , R 12a and R 12b are each independently H, C1-C6 alkyl, C2-C6 alkoxyalkyl, C1-C6 haloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, or 4-7 membered heterocyclyl, wherein said cycloalkyl or heterocyclyl is optionally substituted with 1, 2, or 3 halogen or CN. Ring A can be a 9- to 10-membered heteroaryl. Ring B can be phenyl or a 5- to 6-membered heteroaryl. For example, the T-type calcium channel antagonist can have the structure of Formula (II), (III), (IV), or (V), or a pharmaceutically acceptable salt thereof.

[0023] The method for reducing Miro1 levels described herein can be carried out in any suitable cell. For example, the cell can be a muscle cell. In another example, the cell can be a neuron. The reduced Miro1 levels can be in vitro or ex vivo. Alternatively, the reduced Miro1 levels can be in vivo.

[0024] In another aspect, provided herein is a method for identifying a subject (subject or patient) at risk for developing a Miro1-associated disorder, the method comprising: a) detecting whether the Miro1 level in a biological sample obtained from the subject and treated with a mitochondrial stress response factor (stressor) is equal to or higher than the control Miro1 level in a control biological sample obtained from the subject and untreated; and b) identifying the subject as at risk for developing a Miro1-associated disorder if the Miro1 level is equal to or higher than the control Miro1 level in the control biological sample, wherein the biological sample and the control biological sample comprise iPSCs or cells differentiated from iPSCs. The mitochondrial stress response factor can be carbonyl cyanide 3-chlorophenylhydrazone (CCCP). The method can further include treating the subject at risk for developing a Miro1-associated disorder by administering a therapeutically effective amount of a compound described herein or a pharmaceutically acceptable salt thereof.

[0025] In another embodiment, a method for identifying a subject at risk of developing a Miro1-associated disorder is provided, the method comprising: (a) detecting whether the Miro1 level in a biological sample obtained from the subject and treated with a mitochondrial stress response factor is equal to or higher than the control Miro1 level in an untreated biological sample obtained from the subject; and (b) identifying the subject as at risk of developing a Miro1-associated disorder if the Miro1 level in the biological sample is equal to or higher than the Miro1 level in the control biological sample, where the mitochondrial stress response factor can be carbonyl cyanide 3-chlorophenylhydrazone (CCCP); and (c) treating the subject at risk of developing a Miro1-associated disorder by administering a therapeutically effective amount of a compound described herein or a pharmaceutically acceptable salt thereof. The biological sample and the control biological sample can comprise fibroblasts.

[0026] The present disclosure further describes a method for treating a neurodegenerative disorder in a subject in need of treatment, the method comprising: (a) detecting whether the Miro1 level in a biological sample obtained from the subject and treated with a mitochondrial stress response factor is equal to or higher than the control Miro1 level in a control biological sample; (b) identifying the subject as a candidate for treatment if the Miro1 level in the biological sample is equal to or higher than the control Miro1 level; and (c) administering to the subject a compound described herein or a pharmaceutically acceptable salt thereof. The mitochondrial stress response factor can be carbonyl cyanide 3-chlorophenylhydrazone (CCCP). The biological sample and the control biological sample can comprise fibroblasts. Neurodegenerative disorders include drug-induced parkinsonism, progressive supranuclear palsy, vascular parkinsonism, dementia with Lewy bodies, diffuse Lewy body disease, corticobasal degeneration, multiple system degeneration (Shy-Drager syndrome), Parkinson's disease, Alzheimer's disease, Pick's disease, frontotemporal dementia, multiple system atrophy, vascular dementia, and progressive supranuclear palsy (Steel-Richardson syndrome). The subject can be asymptomatic for the neurodegenerative disorder.

[0027] The Miro1 level relative to the control Miro1 level can be determined by any method in the art, such as the methods further described herein. For example, the ratio of the Miro1 level to the control Miro1 level can be about 0.5 to about 10. In another example, the ratio of the Miro1 level to the control Miro1 level can be about 0.7 to about 4. [Brief explanation of the drawings]

[0028] [Figure 1] FIG. 1 shows the effect of fed benidipine on the decline in locomotor activity in a Drosophila fly model of Parkinson's disease. [Figure 2-1]Figure 2 shows Miro1 response to carbonyl cyanide 3-chlorophenylhydrazone (CCCP) in induced pluripotent stem cells (iPSCs). (A) Schematic of the readout. (B) Example readouts using Healthy-2, PD-29, and Risk-39. Cell lysates were blotted as indicated. [Figure 2-2] Figure 2(C) Summary of Western blotting results. Normalized Miro1 intensities between "DMSO-treated" and "CCCP-treated" subjects within the same subject were compared by Student's t-test. The number of subjects with P > 0.05 combined with subjects who showed significant Miro1 upregulation after CCCP or P < 0.05 for Miro1 reduction after CCCP is shown in the "No. (Miro1 DMSO vs. CCCP, P > 0.05 or < 0.05)" column. Fisher's exact test was used to determine P values ​​for specific subject groups compared to "healthy." [Figure 2-3] Figure 2(D) shows the relative ratio of the average normalized Miro1 intensity (CCCP-treated vs. DMSO-treated) of the same subject measured by Western blotting. n = 3–55. [Figure 2-4] Figure 2 (E-G) Validation of the ELISA for Miro1. (E) A representative standard curve is shown. LLOD = 0.112 ng / mL. Dynamic range = 0.625–40 ng / mL. (F) Plate-to-plate variability was demonstrated by measuring the same sample (healthy-1) on four different plates. [Figure 2-5] Figure 2(G) demonstrates intraplate variability by measuring the same sample (healthy-1) four times within the same plate. (H) Heatmap shows the relative ratio of mean Miro1 values ​​(CCCP-treated divided by DMSO-treated) from the same subject measured by ELISA. n = 3–4. *P < 0.05; **P < 0.01; ***P < 0.001. [Figure 3-1]Figure 3 shows correlation analysis of Miro1 ratio in iPSCs. (A) One-way ANOVA was used to determine significant differences across all subject groups. (B) Two-way ANOVA was used to determine the interaction between sex and genetic background, with Miro1 ratio as the response variable. [Figure 3-2] Figure 3 (C, D). Multivariate regression was used to determine the interaction between age and genetic background (C) or age and sex (D) with the Miro1 ratio as the response variable. All P values ​​were calculated by linear fit. [Figure 4-1] Figure 4 shows correlation analysis of Miro1 ratio in fibroblasts. (A) One-way ANOVA was used. (B) Two-way ANOVA was used to determine the interaction between sex and genetic background, with Miro1 ratio as the response variable. [Figure 4-2] Figure 4 (C, D). Multivariate regression was used to determine the interaction between age and genetic background (C) or age and sex (D) with the Miro1 ratio as the response variable. All P values ​​were calculated by linear fit. [Figure 5-1] Figure 5 shows interactions between demographic and clinical variables. (A) Representative partial regression plots show the effect of a single variable, Mini-Mental State Examination (MMSE), on the Miro1 ratio. Figure 5 (B-D) Representative partial regression plots show interactions between two variables affecting the Miro1 ratio. (B) Hoehn & Yahr Scale (HYS) and age at onset. (C) MMSE and HYS. (D) Age at onset and MMSE. [Figure 5-2] Figure 5 shows interactions between demographic and clinical variables. (A) Representative partial regression plots show the effect of a single variable, Mini-Mental State Examination (MMSE), on the Miro1 ratio. Figure 5 (B-D) Representative partial regression plots show interactions between two variables affecting the Miro1 ratio. (B) Hoehn & Yahr Scale (HYS) and age at onset. (C) MMSE and HYS. (D) Age at onset and MMSE. DETAILED DESCRIPTION OF THE INVENTION

[0029] The present disclosure provides, inter alia, methods and compositions in which selective T-type calcium channel antagonists (also referred to as antagonists), but not selective L-type or N-type calcium channel antagonists, or mixed L-type / N-type or L-type / N-type / P-type calcium channel antagonists, can reduce intracellular Miro1 levels. Thus, selective T-type voltage-dependent calcium channel antagonists, or mixed selective calcium channel antagonists with T-type antagonist activity, may be useful for reducing Miro1 levels, for example, in diseases or conditions that would benefit from reduced Miro1 levels in neuronal cells. Such diseases or conditions include neurodegenerative diseases such as Parkinson's disease.

[0030] Also disclosed are amide compounds of formula (II). See Examples 1-55. Such compounds exhibited T-type calcium channel antagonist activity (Example 60). Thus, compounds of formula (II) can reduce intracellular Miro1 levels and may be useful for treating neurodegenerative diseases such as Parkinson's disease.

[0031] Furthermore, we provide a method for identifying subjects at risk for developing a Miro1-associated disorder by measuring Miro1 levels in a biological sample from a subject treated with a mitochondrial stress response factor and comparing them with control Miro1 levels measured in a corresponding untreated biological sample. Miro1-associated disorders include any of the neurodegenerative diseases described herein, such as Parkinson's disease. As shown in Example 64, in normal subjects, Miro1 levels in cells derived from the subject's biological sample, such as induced pluripotent stem cells (iPSCs) or skin fibroblasts treated with the mitochondrial stress response factor CCCP, were lower than control Miro1 levels in untreated cells. However, in Parkinson's disease patients and subjects at risk for developing Parkinson's disease, Miro1 levels were equal to or higher than control Miro1 levels (Figures 2-4). Therefore, Miro1 levels can be used to identify subjects at risk for developing a Miro1-associated disorder, such as symptomatic and asymptomatic individuals with Parkinson's disease. Furthermore, higher Miro1 levels, which correlate with cognitive decline as measured by MMSE score (Figure 5), suggest that they can be used in conjunction with cognitive impairment to monitor the progression of Parkinson's disease. These results suggest that Miro1 levels can be used to identify subjects with pre-symptomatic Miro1-related disorders, including Parkinson's disease, and treat these subjects with compounds that can reduce Miro1 levels before symptoms appear. I. Definition

[0032] An "alkyl" is a straight-chain or branched saturated monovalent hydrocarbon. For example, an alkyl group can have 1 to 18 carbon atoms (i.e., C 1-18 alkyl) or 1 to 8 carbon atoms (i.e., C 1-8 alkyl) or 1 to 6 carbon atoms (i.e., C 1-6 alkyl) or 1 to 4 carbon atoms (i.e., C 1-4Examples of alkyl groups include, but are not limited to, methyl (Me, -CH), ethyl (Et, -CHCH), 1-propyl (n-Pr, n-propyl, -CHCHCH), 2-propyl (i-Pr, i-propyl, -CH(CH)), 1-butyl (n-Bu, n-butyl, -CHCHCHCHCH), 2-methyl-1-propyl (i-Bu, i-butyl, -CHCH(CH)), 2-butyl (s-B u, s-butyl, -CH(CH3)CH2CH3), 2-methyl-2-propyl (t-Bu, t-butyl, -C(CH3)3), 1-pentyl (n-pentyl, -CH2CH2CH2CH2CH3), 2-pentyl (-CH(CH3)CH2CH2CH3), 3-pentyl (-CH(CH2CH3)2), 2-methyl-2-butyl (-C(CH3)2CH2CH3), 3-methyl-2-butyl (-CH(CH3)CH(CH3)2), 3-Methyl-1-butyl (-CH2CH2CH(CH3)2), 2-methyl-1-butyl (-CH2CH(CH3)CH2CH3), 1-hexyl (-CH2CH2CH2CH2CH2CH2CH3), 2-hexyl (-CH(CH3)CH2CH2CH2CH2CH3), 3-hexyl (-CH(CH2CH3)(CH2CH2CH3)), 2-methyl-2-pentyl (-C(CH3)2CH2CH2CH3), 3-methyl-2-pentyl (-

[0023] Other alkyl groups include heptyl, octyl, nonyl, decyl, undecyl, dodecyl, pentadecyl, hexadecyl, heptadecyl, and octadecyl.

[0033] "Alkenyl" is a straight or branched chain hydrocarbon containing at least two carbon atoms and at least one double bond. Alkenyl includes C, C 2-3 , C 2-4, C 2-5 , C 2-6 , C 2-7 , C 2-8 , C 2-9 , C 2-10 , C3, C 3-4 , C 3-5 , C 3-6 , C4, C 4-5 , C 4-6 , C5, C 5-6 and C6. Alkenyl groups can have any suitable number of double bonds, including, but not limited to, 1, 2, 3, 4, 5, or more. Examples of alkenyl groups include, but are not limited to, vinyl (ethenyl), propenyl, isopropenyl, 1-butenyl, 2-butenyl, isobutenyl, butadienyl, Alkenyl groups include 1-pentenyl, 2-pentenyl, isopentenyl, 1,3-pentadienyl, 1,4-pentadienyl, 1-hexenyl, 2-hexenyl, 3-hexenyl, 1,3-hexadienyl, 1,4-hexadienyl, 1,5-hexadienyl, 2,4-hexadienyl, and 1,3,5-hexatrienyl. Alkenyl groups can be substituted or unsubstituted.

[0034] "Alkynyl" means either a straight or branched chain hydrocarbon having at least two carbon atoms and at least one triple bond. Alkynyl includes C, C 2-3 , C 2-4 , C 2-5 , C 2-6 , C 2-7 , C 2-8 , C 2-9 , C 2-10 , C3, C 3-4 , C 3-5 , C 3-6 , C4, C 4-5 , C 4-6 , C5, C 5-6and C6. Examples of alkynyl groups include, but are not limited to, acetylenyl, propynyl, 1-butynyl, 2-butynyl, butadiynyl, 1-pentynyl, 2-pentynyl, isopentynyl, 1,3-pentadiynyl, 1,4-pentadiynyl, 1-hexynyl, 2-hexynyl, 3-hexynyl, 1,3-hexadiynyl, 1,4-hexadiynyl, 1,5-hexadiynyl, 2,4-hexadiynyl, or 1,3,5-hexatriynyl. Alkynyl groups can be substituted or unsubstituted.

[0035] "Alkoxy" refers to an alkyl group having an oxygen atom connecting the alkyl group to the point of attachment, i.e., alkyl-O-. As with alkyl groups, alkoxy groups are C 1-6 Alkoxy groups can have any suitable number of carbon atoms, such as , ...

[0036] "Alkoxyalkyl" refers to an alkoxy group linked to an alkyl group that is linked to the rest of the compound, such that the alkyl group is divalent. Alkoxyalkyl is an alkyl group having 2 to 6 carbon atoms (C 2-6 Alkoxyalkyl), 2-5(C 2-5 Alkoxyalkyl), 2-4(C 2-4 alkoxyalkyl), or 2-3(C 2-3The number of carbons herein refers to the total number of carbons contained in the alkoxy group and the alkyl group. For example, C alkoxyalkyl refers to ethoxy (C alkoxy) linked to butyl (C alkyl), and n-propoxy (C alkoxy) linked to isopropyl (C alkyl). The alkoxy and alkyl herein are as defined above, and when alkyl is divalent, include, but are not limited to, methoxymethyl (CHOCH-), methoxyethyl (CHOCHCH-), and the like.

[0037] "Halo" or "halogen" as used herein refers to fluoro (-F), chloro (-Cl), bromo (-Br), and iodo (-I).

[0038] As used herein, "haloalkyl" refers to an alkyl, as defined herein, in which one or more hydrogen atoms of the alkyl are independently replaced by halo substituents, which may be the same or different. For example, C 1-4 Haloalkyl is C 1-4 C in which one or more of the alkyl hydrogen atoms have been replaced by halo substituents 1-4 Examples of haloalkyl groups include, but are not limited to, fluoromethyl, fluorochloromethyl, difluoromethyl, difluorochloromethyl, trifluoromethyl, 1,1,1-trifluoroethyl, and pentafluoroethyl.

[0039] "Cycloalkyl" refers to a monocyclic saturated or partially unsaturated all-carbocyclic ring (i.e., C 3-20Cycloalkyl) refers to a monocyclic saturated or partially unsaturated all-carbocyclic ring, e.g., having 3 to 12 ring atoms, e.g., 3 to 10 ring atoms, or 3 to 8 ring atoms, or 3 to 6 ring atoms, or 3 to 5 ring atoms, or 3 to 4 ring atoms. The term "cycloalkyl" also includes polycondensed saturated and partially unsaturated all-carbocyclic ring systems (e.g., ring systems containing 2, 3, or 4 carbocyclic rings). Thus, cycloalkyl includes polycyclic carbocycles, e.g., bicyclic carbocycles (e.g., bicyclic carbocycles having 6 to 12 ring carbon atoms such as bicyclo[3.1.0]hexane and bicyclo[2.1.1]hexane), and polycyclic carbocycles (e.g., tricyclic and tetracyclic carbocycles having 20 or fewer ring carbon atoms). The rings of a fused polycyclic ring system can be connected to each other through fused, spiro, and bridged bonds, if permitted by valence requirements. Non-limiting examples of monocyclic cycloalkyls include cyclopropyl, cyclobutyl, cyclopentyl, 1-cyclopent-1-enyl, 1-cyclopent-2-enyl, 1-cyclopent-3-enyl, cyclohexyl, 1-cyclohex-1-enyl, 1-cyclohex-2-enyl, and 1-cyclohex-3-enyl.

[0040] As used herein, "heterocyclyl" or "heterocycle" or "heterocycloalkyl" refers to a saturated or partially unsaturated non-aromatic monocyclic or polycyclic ring system having at least one heteroatom (i.e., at least one ring heteroatom selected from oxygen, nitrogen, and sulfur) in the ring, wherein the polycyclic ring system includes at least one non-aromatic ring containing at least one heteroatom. The polycyclic ring system may also include other aromatic and non-aromatic rings. Unless otherwise specified, a heterocyclyl group contains 3 to 20 ring atoms, e.g., 3 to 12 ring atoms, e.g., 3 to 10 ring atoms, or 3 to 8 ring atoms, or 3 to 6 ring atoms, or 3 to 5 ring atoms, or 4 to 6 ring atoms, or 4 to 5 ring atoms. Thus, the term includes saturated or partially unsaturated monocyclic rings (e.g., 3-, 4-, 5-, 6-, or 7-membered rings) having 1 to 6 ring carbon atoms and 1 to 3 ring heteroatoms selected from the group consisting of oxygen, nitrogen, and sulfur in the ring. The heteroatom is optionally selected from -N(-OH)-, ═N(-O - )-, -S(=O)-, or -S(=O)2-. The rings of fused polycyclic (e.g., bicyclic heterocyclyl) systems can be connected to each other via fused, spiro, and bridged bonds, for example, where permitted by valence requirements. Heterocycles include, but are not limited to, azetidine, aziridine, imidazolidine, morpholine, oxirane (epoxide), oxetane, thietane, piperazine, piperidine, pyrazolidine, piperidine, pyrrolidine, pyrrolidinone, tetrahydrofuran, tetrahydrothiophene, dihydropyridine, tetrahydropyridine, quinuclidine, 2-oxa-6-azaspiro[3.3]heptan-6-yl, 6-oxa-1-azaspiro[ 3.3]heptan-1-yl, 2-thia-6-azaspiro[3.3]heptan-6-yl, 2,6-diazaspiro[3.3]heptan-2-yl, 2-azabicyclo[3.1.0]hexan-2-yl, 3-azabicyclo[3.1.0]hexanyl, 2-azabicyclo[2.1.1]hexanyl, 2-azabicyclo[2.2.1]heptan-2-yl, 4-azaspiro[2.4]heptanyl, 5-azaspiro[2.4]heptanyl, and the like.

[0041] As used herein, "aryl" refers to an all-carbon aromatic monocyclic ring system or an all-carbon fused polycyclic ring system in which at least one ring is aromatic. For example, in some embodiments, aryl groups have 6 to 20 carbon atoms, 6 to 14 carbon atoms, or 6 to 12 carbon atoms. Aryl includes phenyl radicals. Aryl also includes fused polycyclic ring systems (e.g., ring systems containing 2, 3, or 4 rings) having 9 to 20 carbon atoms, e.g., 9 to 16 carbon atoms, in which at least one ring is aromatic and the other rings may or may not be aromatic (i.e., carbocyclic). Such fused polycyclic ring systems may be optionally substituted with one or more (e.g., 1, 2, or 3) oxo groups on any carbocyclic portion of the fused polycyclic ring system. The rings of a fused polycyclic ring system may be connected to each other through fused, spiro, and bridged bonds, where permitted by valence requirements. It should also be understood to refer to aryls composed of members within a specific range of atom numbers, such as 6- to 10-membered aryls, where the atom number range refers to the total number of ring atoms in the aryl. For example, 6-membered aryls include phenyl, and 10-membered aryls include naphthyl and 1,2,3,4-tetrahydronaphthyl. Non-limiting examples of aryl groups include phenyl, indenyl, naphthyl, 1,2,3,4-tetrahydronaphthyl, anthracenyl, etc.

[0042] As used herein, "heteroaryl" refers to an aromatic monocyclic ring having at least one atom other than carbon in the ring, where the atom is selected from the group consisting of oxygen, nitrogen, and sulfur; "heteroaryl" also includes fused polycyclic ring systems having at least one such aromatic ring, which fused polycyclic ring systems are further described below. Thus, "heteroaryl" includes an aromatic monocyclic ring having 1 to 6 carbon atoms and 1 to 4 heteroatoms selected from the group consisting of oxygen, nitrogen, and sulfur. The sulfur and nitrogen atoms may be present in oxidized form, provided the ring is aromatic. Exemplary heteroaryl ring systems include, but are not limited to, pyridyl, pyrimidinyl, oxazolyl, or furyl. "Heteroaryl" also encompasses fused polycyclic ring systems (e.g., ring systems containing 2, 3, or 4 rings), where a heteroaryl group as defined above is fused with one or more rings selected from heteroaryl (e.g., to form 1,8-naphthyridinyl), heterocycle (e.g., to form 1,2,3,4-tetrahydro-1,8-naphthyridinyl), carbocycle (e.g., to form 5,6,7,8-tetrahydroquinolyl), and aryl (e.g., to form indazolyl) to form a fused polycyclic ring system. Thus, a heteroaryl (aromatic monocyclic or fused polycyclic ring system) has 1 to 20 carbon atoms and 1 to 6 heteroatoms in the heteroaryl ring. Such fused polycyclic ring systems may optionally be substituted on the carbocyclic or heterocyclic moieties with one or more (e.g., 1, 2, 3, or 4) oxo groups. The rings of a fused polycyclic ring system can be joined to each other through fused, spiro, and bridged bonds, where permitted by valence requirements. It should be understood that the individual rings of the fused polycyclic ring system can be joined to each other in any order. It should be understood that the point of attachment of a heteroaryl or heteroaryl fused polycyclic ring system can be at any suitable atom of the heteroaryl or heteroaryl fused polycyclic ring system, including carbon atoms and heteroatoms (e.g., nitrogen). It should also be understood that when reference is made to a specific range of atoms that are members of a heteroaryl (e.g., a 5- to 10-membered heteroaryl), the atom range is in terms of the total ring atoms of the heteroaryl, and includes carbon atoms and heteroatoms.For example, a 5-membered heteroaryl would include thiazolyl and a 10-membered heteroaryl would include quinolinyl. Exemplary heteroaryls include, but are not limited to, pyridyl, pyrrolyl, pyrazinyl, pyrimidinyl, pyridazinyl, pyrazolyl, thienyl, indolyl, imidazolyl, oxazolyl, isoxazolyl, thiazolyl, furyl, oxadiazolyl, thiadiazolyl, quinolyl, isoquinolyl, benzothiazolyl, benzoxazolyl, indazolyl, quinoxalyl, quinazolyl, 5,6,7,8-tetrahydroisoquinolinyl, benzofuranyl, benzimidazolyl, thianaphthenyl, pyrrolo[2,3-b]pyridinyl, quinazolinyl-4(3H)-one, and triazolyl.

[0043] Also provided are compounds described herein, or pharmaceutically acceptable salts, isotopes, or mixtures thereof, in which 1 to n hydrogen atoms bonded to a single carbon atom are replaced by deuterium or D, where n is the number of hydrogen atoms in the molecule. As is known in the art, deuterium atoms are non-radioactive isotopes of hydrogen atoms. Such compounds can enhance resistance to metabolism and therefore may be useful for increasing the half-life of the compounds described herein, or pharmaceutically acceptable salts, isotopes, or mixtures thereof, when administered to a mammal. See, for example, Foster, "Deuterium Isotope Effects in Studies of Drug Metabolism," Trends Pharmacol. Sci., 5(12):524-527 (1984). Such compounds can be synthesized by means well known in the art, for example, by using starting materials in which one or more hydrogen atoms have been replaced with deuterium.

[0044] Examples of isotopes that can be incorporated into the disclosed compounds include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus, fluorine, chlorine, and iodine, e.g., 2 H, 3 H, 11 C. 13 C. 14 C. 13 N,15 N, 15 O. 17 O. 18 O. 31 P, 32 P, 35 S, 18 F, 36 Cl, 123 I, 125 I. Substitution with positron generating isotopes, e.g., 11 C. 18 F, 15 O and 13 N may be useful in positron emission topography (PET) studies to examine substrate receptor occupancy. In general, isotopically labeled compounds of formula (I) can be prepared by conventional techniques known to those skilled in the art, or by processes similar to those described in the examples provided below, using an appropriate isotopically labeled reagent in place of the conventionally used non-labeled reagent.

[0045] Compounds of the disclosure include compounds described herein, for example, compounds of the disclosure include compounds of formula (I), (II), (III), (IV), and (V), including compounds of the Examples.

[0046] As used herein, "composition" is intended to encompass a product containing the specified ingredients in the specified amounts, as well as any product resulting directly or indirectly from combining the specified ingredients in the specified amounts. "Pharmaceutically acceptable" means that the carrier, diluent, or excipient must be compatible with the other ingredients of the formulation and not deleterious to the recipient thereof.

[0047] "Pharmaceutically effective amount" refers to the amount of a compound of the present disclosure in a formulation or combination thereof that provides the desired therapeutic or pharmacological result.

[0048] A "pharmaceutically acceptable excipient" includes, but is not limited to, any adjuvant, carrier, filler, glazing agent, sweetener, diluent, preservative, dye / colorant, flavor enhancer, surfactant, wetting agent, dispersing agent, suspending agent, stabilizer, isotonic agent, solvent, or emulsifier approved by the U.S. Food and Drug Administration (FDA) as acceptable for use in humans or domestic animals.

[0049] As used herein, "treatment" or "treating" or "treating" refers to an approach for obtaining beneficial or desired results. For purposes of this disclosure, beneficial or desired results include, but are not limited to, alleviating symptoms and / or reducing the severity and / or preventing worsening of symptoms associated with a disease or condition. In some embodiments, "treatment" or "treating" includes one or more of the following: a) inhibiting a disease or condition (e.g., reducing one or more symptoms resulting from a disease or condition and / or diminishing the severity of the disease or condition); b) slowing or halting the development of one or more symptoms associated with a disease or condition (e.g., stabilizing a disease or condition, slowing worsening or progression of a disease or condition); and c) palliating a disease or condition, e.g., regressing clinical symptoms, improving a disease state, alleviating pathology, slowing disease progression, improving quality of life, and / or prolonging survival.

[0050] As used herein, a "therapeutically effective amount" or "effective amount" refers to an amount effective to elicit a desired biological or medical response, including an amount of a compound that, when administered to a subject to treat a disease, is sufficient to effect treatment of such disease. The effective amount may vary depending on the compound, the disease, and its severity, as well as age, weight, and the like. An effective amount may include a range of amounts. As understood in the art, an effective amount may be a single dose or multiple doses, and the single dose or multiple doses may be the amount needed to achieve a desired therapeutic endpoint. An effective amount may be considered in the context of administering one or more therapeutic agents, and may be considered to have been administered in an effective amount if, in conjunction with one or more other agents, a desired or beneficial result can be achieved or is achieved. The appropriate dose of any co-administered compound may optionally be reduced due to the combined effects (e.g., additive or synergistic) of the compounds.

[0051] "Administering" refers to oral administration, administration as a suppository, topical contact, parenteral, intravenous, intraperitoneal, intramuscular, intralesional, intranasal or subcutaneous administration, intrathecal administration, or implantation of a sustained-release device, e.g., a mini-osmotic pump, into the subject, etc. Administration can be carried out according to a schedule that dictates frequency of administration, dosage, and other factors.

[0052] As used herein, "co-administration" refers to administration of a unit dose of a compound disclosed herein before or after administration of a unit dose of one or more additional therapeutic agents, e.g., administration of a compound disclosed herein within seconds, minutes, or hours of administration of one or more additional therapeutic agents. In some embodiments, a unit dose of a compound disclosed herein is administered first, followed within seconds or minutes by a unit dose of one or more additional therapeutic agents. Alternatively, in other embodiments, a unit dose of one or more additional therapeutic agents is administered first, followed within seconds or minutes by a unit dose of a compound disclosed herein. In some embodiments, a unit dose of a compound disclosed herein is administered first, followed several hours (e.g., 1-12 hours) later by a unit dose of one or more additional therapeutic agents. In other embodiments, a unit dose of one or more additional therapeutic agents is administered first, followed several hours (e.g., 1-12 hours) later by a unit dose of a compound disclosed herein. Co-administration of the compounds disclosed herein generally refers to the simultaneous or sequential administration of a compound disclosed herein with one or more additional therapeutic agents such that a therapeutically effective amount of each agent is present in the patient's body.

[0053] "Subject (or subject or patient)" refers to an animal, such as a mammal, including, but not limited to, a primate (e.g., a human), cow, sheep, goat, horse, dog, cat, rabbit, rat, mouse, etc. In some embodiments, the subject is a human.

[0054] "Disease" or "condition" refers to a condition or state of a patient or subject that can be treated by a compound, pharmaceutical composition, or method provided herein.

[0055] II. Compounds The present disclosure provides compounds useful as T-type voltage-dependent calcium channel antagonists.

[0056] In some embodiments, the compound has the structure of Formula (I): [ka] or a pharmaceutically acceptable salt thereof, wherein: Ring A and ring B each independently represent a C6 to C 10 aryl or 5-10 membered heteroaryl; R 1 , R 2 and R 3 are each independently H, halogen, CN, OR 11 , N.R. 12a R 12b , C1-C6 alkyl, C2-C6 alkoxyalkyl, C1-C6 haloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, phenyl, 5-6 membered heteroaryl, C3-C7 cycloalkyl, or 4-7 membered heterocyclyl, wherein said phenyl, heteroaryl, cycloalkyl, or heterocyclyl optionally contains one, two, or three halogen atoms, CN, OR 11 , N.R. 12a R 12b , optionally substituted by C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylamino, or C1-C6 haloalkyl; R 4a and R 4b are each independently H or C1-C6 alkyl; R 5 is H or C1-C6 alkyl; R 6a and R 6b are each independently H or C1-C6 alkyl; R 7 is H, halogen, CN, OR 11 , N.R. 12a R 12b , C1-C6 alkyl, C2-C6 alkoxyalkyl, C1-C6 haloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, phenyl, 5-6 membered heteroaryl, C3-C7 cycloalkyl, or 4-7 membered heterocyclyl, wherein said phenyl, heteroaryl, cycloalkyl, or heterocyclyl optionally contains one, two, or three halogen atoms, CN, OR 11 , N.R. 12a R12b , optionally substituted by C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylamino, or C1-C6 haloalkyl; R 8 is H, C1-C6 alkyl, C2-C6 alkoxyalkyl, C1-C6 haloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, wherein said cycloalkyl or heterocyclyl is optionally substituted by 1, 2, or 3 halogen, CN, OH, NH2, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylamino, or C1-C6 haloalkyl; R 9 is H, C1-C6 alkyl, C2-C6 alkoxyalkyl, C1-C6 haloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, or 4-7 membered heterocyclyl, wherein said cycloalkyl or heterocyclyl is optionally substituted by 1, 2, or 3 halogen, CN, OH, NH2, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylamino, or C1-C6 haloalkyl; and Each R 11 , R 12a and R 12b are each independently H, C1-C6 alkyl, C2-C6 alkoxyalkyl, C1-C6 haloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, or 4-7 membered heterocyclyl, wherein said cycloalkyl or heterocyclyl is optionally substituted with 1, 2, or 3 halogen or CN.

[0057] In some embodiments, ring A is a 9-10 membered heteroaryl.

[0058] In some embodiments, ring A is pyrrolopyridine, isoxazolopyridine, pyrazolopyridine, pyrazolopyridazine, or triazolopyridine. In some embodiments, ring A is pyrazolo[3,4-c]pyridin-5-yl, pyrazolo[4,3-c]pyridin-6-yl, isoxazolo[4,5-c]pyridin-6-yl, isoxazolo[5,4-c]pyridin-5-yl, pyrazolo[3,4-b]pyridin-5-yl, pyrazolo[3,4-c]pyridazin-5-yl, [1,2,3]triazolo[4,5-c]pyridin-6-yl, or pyrrolo[2,3-c]pyridin-5-yl. In some embodiments, ring A is pyrazolopyridine.

[0059] In some embodiments, the compound has the structure of Formula (II): [ka] or a pharmaceutically acceptable salt thereof, Ring B is C6 to C 10 aryl or 5-10 membered heteroaryl; R 1 , R 2 and R 3 are each independently H, halogen, CN, OR 11 , N.R. 12a R 12b , C1-C6 alkyl, C2-C6 alkoxyalkyl, C1-C6 haloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, phenyl, 5-6 membered heteroaryl, C3-C7 cycloalkyl, -(CH2) n -(C3-C7 cycloalkyl), 4- to 7-membered heterocyclyl, or -(CH2) n -(4- to 7-membered heterocyclyl), wherein said phenyl, heteroaryl, cycloalkyl or heterocyclyl optionally contains one, two or three halogen atoms, CN, OR 11 , N.R. 12a R 12b , optionally substituted by C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylamino, or C1-C6 haloalkyl; R 4a and R 4b are each independently H or C1-C6 alkyl, or R 4a and R 4b and the carbon atom to which they are attached together form a C3-C5 cycloalkyl; R 6 is H, C1-C6 alkyl, or C1-C6 haloalkyl; X 1 and X 5 are each independently N or CR 8 where X 1 and X 5 At least one of the following is CR 8 and; X 2 , X 3 and X 4 are each independently N, NR 9 , O, S or CR 9 where X 2 , X 3 and X 4 At least one of the following is N or NR 9 , O, or S; R 8 is H, halogen, CN, OR 11 , C1-C6 alkyl, C2-C6 alkoxyalkyl, or C1-C6 haloalkyl; R 9 is H, C1-C6 alkyl, C2-C6 alkoxyalkyl, C1-C6 haloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, or 4-7 membered heterocyclyl, wherein said cycloalkyl or heterocyclyl may be optionally substituted by 1, 2, or 3 halogen, CN, OH, NH2, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylamino, or C1-C6 haloalkyl; Each R 11 , R 12a and R 12bare each independently H, C1-C6 alkyl, C2-C6 alkoxyalkyl, C1-C6 haloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, or 4-7 membered heterocyclyl, wherein said cycloalkyl or heterocyclyl is optionally substituted by 1, 2, or 3 halogen or CN; and n is 1, 2, or 3.

[0060] In some embodiments, X 1 is CR 8 is.

[0061] In some embodiments, R 8 is H.

[0062] In some embodiments, X 2 , X 3 , and X 4 are independently N, NR 9 or CR 9 where X 2 , X 3 and X 4 At least one of is N or NR 9 This is subject to the condition that:

[0063] In some embodiments, X 1 is N or CR 8 and X 2 and X 3 are each independently N or NR 9 and X 4 is N, NR 9 , or CR 9 and X 5 is CR 8 For example, X 1 is CR 8 and X 2 and X 3 are each independently N or NR 9 X can be 4 is N, NR 9 , or CR 9 X can be 5is CR 8 In some embodiments, X 1 is N or CR 8 and X 2 is N or NR 9 and X 3 is N, NR 9 or CR 9 and X 4 is N or CR 9 and X 5 is CR 9 In some embodiments, X 1 is CR 8 and X 2 is N or NR 9 and X 3 is N, NR 9 , or CR 9 and X 4 is N or CR 9 and X 5 is CR 9 is.

[0064] In some embodiments of the compounds of Formula (I) and / or (II), the compounds have the following structure: [ka] or a pharmaceutically acceptable salt thereof.

[0065] In some embodiments of the compounds of Formula (I) and / or (II), the compounds have the following structure: [ka] or a pharmaceutically acceptable salt thereof.

[0066] In some embodiments of the compound of Formula (II), the compound has the following structure: [ka] or a pharmaceutically acceptable salt thereof, wherein: Ring B is C6 to C 10aryl or 5-10 membered heteroaryl; R 1 , R 2 and R 3 are each independently H, halogen, CN, OR 11 , N.R. 12a R 12b , C1-C6 alkyl, C2-C6 alkoxyalkyl, C1-C6 haloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, phenyl, 5-6 membered heteroaryl, C3-C7 cycloalkyl, -(CH2) n -(C3-C7 cycloalkyl), 4- to 7-membered heterocyclyl, or -(CH2) n -(4- to 7-membered heterocyclyl), wherein said phenyl, heteroaryl, cycloalkyl or heterocyclyl optionally contains one, two or three halogen atoms, CN, OR 11 , N.R. 12a R 12b , optionally substituted by C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylamino, or C1-C6 haloalkyl; R 4a and R 4b are each independently H or C1-C6 alkyl, or R 4a and R 4b and the carbon atoms to which they are attached together form a C3-C5 cycloalkyl; R 6 is H, C1-C6 alkyl, or C1-C6 haloalkyl; X 1 and X 5 are each independently N or CR 8 where X 1 and X 5 At least one of the following is CR 8 and; X 2 , X 3 and X 4 are each independently N, NR 9 , O, S or CR 9 where X 2 , X 3 and X 4At least one of the following is N or NR 9 , O, or S; R 8 is H, halogen, CN, OR 11 , C1-C6 alkyl, C2-C6 alkoxyalkyl, or C1-C6 haloalkyl; R 9 is H, C1-C6 alkyl, C2-C6 alkoxyalkyl, C1-C6 haloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, or 4-7 membered heterocyclyl, wherein said cycloalkyl or heterocyclyl may be optionally substituted by 1, 2, or 3 halogen, CN, OH, NH2, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylamino, or C1-C6 haloalkyl; Each R 11 , R 12a and R 12b is independently H, C1-C6 alkyl, C2-C6 alkoxyalkyl, C1-C6 haloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, or 4-7 membered heterocyclyl, wherein said cycloalkyl or heterocyclyl may be optionally substituted by 1, 2, or 3 halogen or CN; and n is 1, 2 or 3; However, the compound has the following structure: [ka] [ka] or [ka] or a pharmaceutically acceptable salt thereof The condition is that the applicant does not have

[0067] In some embodiments, the compound has the structure of Formula (III): [ka] or a pharmaceutically acceptable salt thereof.

[0068] In some embodiments of compounds of Formula (I), (II), and / or (III), R 1 , R 2 and R 3 are each independently H, halogen, CN, OH, NH2, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylamino, C2-C6 alkoxyalkyl, C1-C6 haloalkyl, or C3-C7 cycloalkyl, wherein the cycloalkyl is optionally substituted with 1, 2, or 3 halogen, CN, OH, NH2, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylamino, or C1-C6 haloalkyl.

[0069] In some embodiments of compounds of Formula (I), (II) and / or (III), R 3 is H, halogen, or CN. In some embodiments, R 3 is H or halogen. In some embodiments, R 3 is H.

[0070] In some embodiments of compounds of Formula (I), (II), and / or (III), R 4b is H or CH3.

[0071] In some embodiments of compounds of Formula (I), (II), and / or (III), R 4a and R 4b are each independently H or CH3.

[0072] In some embodiments of compounds of Formula (I), (II), and / or (III), Ring B is phenyl, 1H-benzo[d]imidazol-5-yl, 1H-indazol-5-yl, benzo[d][1,3]dioxol-5-yl, 2H-indazol-6-yl, thiophen-2-yl, pyridin-2-yl, or pyridin-3-yl; R 1is F, Me, iPr, CF3, CF2CH3, cyclopropyl, 1-fluorocyclopropyl, 1-cyanocyclopropyl, 2,2-difluorocyclopropyl, 1-trifluoromethylcyclopropyl, 1,1-difluoromethylcyclopropyl; R 2 is H, F, Cl, CN, or CH3; R 3 is H or CH3.

[0073] In some embodiments, the compound has the structure of Formula (IV): [ka] or a pharmaceutically acceptable salt thereof.

[0074] In some embodiments of compounds of Formula (I), (II), (III), and / or (IV), Ring B is phenyl or 5-6 membered heteroaryl. In some embodiments, Ring B is phenyl or pyridyl.

[0075] In some embodiments of compounds of Formula (I), (II), and / or (III), R 4a is H or CH3.

[0076] In some embodiments, the compound has the structure of Formula (V): [ka] or a pharmaceutically acceptable salt thereof.

[0077] In some embodiments of compounds of Formula (I), (II), (III), (IV) and / or (V), R 1 is H, halogen, C1-C6 alkyl, C1-C6 alkoxy, or C2-C6 alkoxyalkyl. In some embodiments, R 1 is H, halogen, C1-C3 alkyl, or C1-C3 haloalkyl. In some embodiments, R 1 is H or halogen. In some embodiments, R1 is C1-C3 haloalkyl. In some embodiments, R 1 is CF3.

[0078] In some embodiments of compounds of Formula (I), (II), (III), (IV), and / or (V), R 2 is H, halogen, C1-C6 alkyl, C1-C6 alkoxy, or C2-C6 alkoxyalkyl. In some embodiments, R 2 is H, halogen, C1-C3 alkyl, or C1-C3 haloalkyl. In some embodiments, R 2 is H or halogen. In some embodiments, R 2 is H.

[0079] In some embodiments of compounds of Formula (I), (II), (III), (IV), and / or (V), R 1 and R 2 are each independently H, halogen, C1-C6 alkyl, C1-C6 alkoxy, C2-C6 alkoxyalkyl, C1-C6 haloalkyl, or C3-C7 cycloalkyl, wherein the cycloalkyl is optionally substituted with 1, 2, or 3 halogen, CN, OH, NH2, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylamino, or C1-C6 haloalkyl. In some embodiments, R 1 and R 2 are each independently H, halogen, C1-C3 alkyl, C1-C3 alkoxy, C2-C3 alkoxyalkyl, C1-C3 haloalkyl, or C3-C5 cycloalkyl, wherein the cycloalkyl is optionally substituted with 1, 2, or 3 halogen, CN, C1-C3 alkyl, C1-C3 alkoxy, C1-C3 alkylamino, or C1-C3 haloalkyl. In some embodiments, R 1 and R 2 are each independently H, halogen, C1-C3 alkyl, or C1-C3 haloalkyl. 1 and R2 are each independently H or halogen. In some embodiments, R 1 and R 2 are each independently H, C1-C3 alkyl, or C1-C3 haloalkyl. 1 and R 2 are each independently H or C1-C3 haloalkyl.

[0080] In some embodiments of compounds of Formula (I), (II), (III), (IV), and / or (V), R 1 is H or C1-C3 haloalkyl, and R 2 is H or halogen. In some embodiments, R 1 is C1-C3 haloalkyl, and R 2 is H. In some embodiments, R 1 is CF3 and R 2 is H.

[0081] In some embodiments of compounds of Formula (I), (II), (III), (IV), and / or (V), R 6 is H or C1-C3 alkyl. In some embodiments, R 6 is CH3.

[0082] In some embodiments of compounds of Formula (I), (II), (III), (IV), and / or (V), R 9 is C1-C6 alkyl, C2-C6 alkoxyalkyl, C1-C6 haloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C7 cycloalkyl, or 4-7 membered heterocyclyl, wherein said cycloalkyl or heterocyclyl is optionally substituted with 1, 2, or 3 halogen, CN, OH, NH2, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylamino, or C1-C6 haloalkyl. 9is C1-C6 alkyl, C2-C6 alkoxyalkyl, C1-C6 haloalkyl, C3-C7 cycloalkyl, or 4-7 membered heterocyclyl, wherein said cycloalkyl or heterocyclyl is optionally substituted with 0, 1, 2, or 3 halogen, CN, OH, NH2, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 alkylamino, or C1-C6 haloalkyl. 9 is C1-C3 alkyl, C2-C3 alkoxyalkyl, or C1-C3 haloalkyl, wherein the cycloalkyl or heterocyclyl is optionally substituted with 0, 1, 2, or 3 F, Cl, CN, OH, NH2, C1-C3 alkyl, C1-C3 alkoxy, C1-C3 alkylamino, or C1-C3 haloalkyl. 9 is C1-C3 alkyl, C2-C3 alkoxyalkyl, or C1-C3 haloalkyl. 9 is C1-C3 haloalkyl. In some embodiments, R 9 is CH2CF3.

[0083] In some embodiments of the compounds of Formula (I), (II), (III), (IV), and / or (V), the compound has the following structure: [ka] or a pharmaceutically acceptable salt thereof.

[0084] Depending on the specific substitution pattern, the compounds of formula (I), (II), (III), (IV), and / or (V), or pharmaceutically acceptable salts thereof, may have one or more stereocenters. In certain instances, it may be advantageous to separate and evaluate individual enantiomers and / or diastereomers to facilitate compound evaluation. Methods for producing individual diastereomers and / or enantiomers are known in the art, including, but not limited to, chiral chromatography, such as supercritical fluid chromatography on a chiral amylose column, and diastereoselective and / or enantioselective synthesis using chiral auxiliaries, such as organometallic addition to chiral sulfinimines. See, for example, the procedure described in Example 8. Thus, in some embodiments, the compound is enantiomerically enriched and present in about 90% to about 99.999% enantiomeric excess (ee), e.g., about 90% to about 99.99% enantiomeric excess, about 93% to 99.99%, about 95% to about 99.99%, about 95% to about 99.9%, about 97% to about 99.9%, about 98% to about 99.9%, or about 99% to about 99.99% ee. Unless otherwise indicated, the predominant isomer is that of the stereochemistry shown in the compound structures herein. In some embodiments, the compound is predominantly the R isomer. In some embodiments, the compound is predominantly the S isomer.

[0085] In some embodiments of a compound of Formula (I), (II), (III), (IV) and / or (V), the compound has the structure of any one of the compounds in Table 1 and Table 2, or a pharmaceutically acceptable salt thereof.

[0086] In some embodiments of a compound of Formula (I), (II), (III), (IV) and / or (V), the compound has the structure of any one of the compounds in Table 1, or a pharmaceutically acceptable salt thereof. [Table 1-1] [Table 1-2] [Table 1-3] [Table 1-4] [Table 1-5]

[0087] For example, in some embodiments, compounds of the present disclosure have the following structure: [ka] or a pharmaceutically acceptable salt thereof.

[0088] In some embodiments of the compounds of Formula (I), (II), (III), (IV) and / or (V), the compound has the structure of any one of the compounds set forth in Table 2, or a pharmaceutically acceptable salt thereof. [Table 2-1] [Table 2-2] [Table 2-3]

[0089] In some embodiments of the compound of Formula (I), the compound has the structure of any of the compounds in Table 3, or a pharmaceutically acceptable salt thereof. [Table 3]

[0090] The compounds described herein are T-type calcium channel antagonists or T-type voltage-dependent calcium channel antagonists. Thus, the compounds of the present disclosure can exhibit T-type calcium channel antagonist activity in one or more assays known in the art or described herein. [Compound activity]

[0091] The compounds of the present disclosure generally have activity in one or more T-type calcium channel antagonist assays described herein or known in the art. In some embodiments, the compounds of the present disclosure have T-type calcium channel antagonist activity in a patch clamp assay. In some embodiments, the compounds have IC 50 In some embodiments, the compound has an ICICM in a patch clamp assay. 50 In some embodiments, the compound has an ICI <10 μM in a patch clamp assay. 50 In some embodiments, the compound has an ICIM in a patch clamp assay. 50 In some embodiments, the compound has an ICIOnM in a patch clamp assay. 50 In some embodiments, the compound has an IC ... 50 In some embodiments, the compound has an IC<0.05<0.1 nM in a patch clamp assay. 50 In some embodiments, the compound has an IC 50 <0.01 nM.

[0092] In some embodiments, the compounds have an IC in the range of about 0.01 nM to about 20 μM in a patch clamp assay. 50 In some embodiments, the compounds have an IC in the range of about 0.01 nM to about 10 μM in a patch clamp assay. 50In some embodiments, the compounds have an IC in the range of about 0.01 nM to about 1 μM in a patch clamp assay. 50 In some embodiments, the compounds have an IC in the range of about 0.01 nM to about 100 nM in a patch clamp assay. 50 In some embodiments, the compounds have an IC in the range of about 0.01 nM to about 10 nM in a patch clamp assay. 50 In some embodiments, the compounds have an IC in the range of about 0.1 nM to about 100 nM in a patch clamp assay. 50 In some embodiments, the compounds have an IC in the range of about 0.1 nM to about 10 nM in a patch clamp assay. 50 It has. [Higher compound selectivity than other ion channels]

[0093] In general, compound selectivity is preferred to produce a desired pharmacological effect while reducing the impact of undesired off-target effects. Thus, in some embodiments, compounds of the present disclosure are selective for T-type calcium channels over other types of T-type calcium channels, including L-type, N-type, P-type, and / or R-type calcium channels. In some embodiments, compounds of the present disclosure are more selective for T-type calcium channels than L-type calcium channels. In some embodiments, compounds of the present disclosure are more selective for T-type calcium channels than N-type calcium channels. In some embodiments, compounds of the present disclosure are more selective for T-type calcium channels than P-type calcium channels. In some embodiments, compounds of the present disclosure are more selective for T-type calcium channels than R-type calcium channels.

[0094] The standard (comparator) ion channel for selectivity determination is Ca V 1.2 (L type), Ca V 1.3 (L type), Ca V 2.1 (P type), Ca V 2.2 (N type), Ca V2.3 (L type), K V 1.5, K V 4.3, KChIP2, K V 7.1, KCNE1(minK), K V 7.2, K V 7.3, K V 11.1(hERG), HCN4, Kir2.1, Kir3.1, Kir3.4, Na V 1.1, Na V 1.2, Na V 1.5, and Na V 1.6 is an example.

[0095] Selectivity can be determined by comparing the antagonist activity of a given compound at least two different ion channel types in a comparative assay format. Any suitable assay capable of measuring calcium channel antagonist activity can be used to determine selectivity. In some embodiments, selectivity is measured using the IC 50 or by comparing % inhibition values. Exemplary assays that can be used to determine activity and selectivity are further described below. For example, the antagonist IC of a compound obtained for each calcium channel by Patz clamp assay can be used to compare the IC of a compound obtained for each calcium channel by Patz clamp assay. 50 By comparing the values, R-type calcium channel Ca V T-type calcium channel Ca higher than 2 V 3.1 can be measured. V 3.1 About IC 50 = 100 nM and Ca V 2.3 About IC 50 A given compound has Ca = 1000 nM V Ca than 2.3 V 3.1, and therefore has 10-fold greater selectivity for T-type calcium channels over R-type calcium channels.

[0096] In some embodiments, compounds of the present disclosure have at least about 1.2, 1.5, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 30, 40, 50, 60, 70, 80, 90, 100, 200, 300, 400, 500, 1000, 2000, 3000, 4000, 5000, 10000, 100,000 or more fold selectivity for T-type calcium channels over one or more (e.g., 2, 3 or more) other calcium channel types, including L-type, N-type, P-type, and / or R-type calcium channels. Selectivity for T-type calcium channels over other calcium channel types, including L-type, N-type, P-type, and / or R-type calcium channels, can be determined by any assay known in the art, for example, by patch clamp assay. In some embodiments, compounds of the present disclosure have selectivity for T-type calcium channels over one or more, e.g., two, three, or more, other calcium channel types, including L-type, N-type, P-type, and / or R-type calcium channels, that is at least about 1.2-fold or greater. In some embodiments, compounds of the present disclosure have selectivity for T-type calcium channels over one or more, e.g., two, three, or more, other calcium channel types, including L-type, N-type, P-type, and / or R-type calcium channels, that is at least about 1.5-fold or greater. In some embodiments, compounds of the present disclosure have selectivity for T-type calcium channels over one or more, e.g., two, three, or more, other calcium channel types, including L-type, N-type, P-type, and / or R-type calcium channels, that is at least about 10-fold or greater. In some embodiments, compounds of the present disclosure have selectivity for T-type calcium channels over one or more, e.g., two, three, or more, other calcium channel types, including L-type, N-type, P-type, and / or R-type calcium channels, that is at least about 100-fold or greater.In some embodiments, compounds of the present disclosure have selectivity for T-type calcium channels over one or more, e.g., two, three, or more, other calcium channel types, including L-type, N-type, P-type, and / or R-type calcium channels, by at least about 1,000-fold or more. In some embodiments, compounds of the present disclosure have selectivity for T-type calcium channels over one or more, e.g., two, three, or more, other calcium channel types, including L-type, N-type, P-type, and / or R-type calcium channels, by at least about 10,000-fold or more. In some embodiments, compounds of the present disclosure have selectivity for T-type calcium channels over one or more, e.g., two, three, or more, other calcium channel types, including L-type, N-type, P-type, and / or R-type calcium channels, by at least about 100,000-fold or more.

[0097] In some embodiments, compounds of the present disclosure have selectivity for T-type calcium channels over one or more, e.g., two, three, or more, additional calcium channel types, including L-type, N-type, P-type, and / or R-type calcium channels, in the range of about 1.2-fold to about 100,000-fold. Selectivity for T-type calcium channels over additional calcium channels, including L-type, N-type, P-type, and / or R-type calcium channels, can be determined by any assay known in the art, e.g., by patch clamp assay. In some embodiments, compounds of the present disclosure have selectivity for T-type calcium channels over one or more, e.g., two, three, or more, additional calcium channels, including L-type, N-type, P-type, and / or R-type calcium channels, in the range of about 1.2-fold to about 10,000-fold. In some embodiments, compounds of the present disclosure have selectivity for T-type calcium channels over one or more, e.g., two, three, or more, additional calcium channels, including one or more, e.g., two, three, or more, L-type, N-type, P-type, and / or R-type calcium channels, ranging from about 1.2-fold to about 100-fold. In some embodiments, compounds of the present disclosure have selectivity for T-type calcium channels over one or more, e.g., two, three, or more, additional calcium channels, including one or more, e.g., two, three, or more, L-type, N-type, P-type, and / or R-type calcium channels, ranging from about 1.2-fold to about 100-fold. In some embodiments, compounds of the present disclosure have selectivity for T-type calcium channels over one or more, e.g., two, three, or more, additional calcium channels, including one or more, e.g., two, three, or more, L-type, N-type, P-type, and / or R-type calcium channels, ranging from about 10-fold to about 100,000-fold. In some embodiments, compounds of the present disclosure have selectivity for T-type calcium channels in the range of about 100-fold to about 100,000-fold over one or more, e.g., two, three, or more, additional calcium channels, including L-type, N-type, P-type, and / or R-type calcium channels. III. Composition

[0098] The present disclosure provides, inter alia, compositions of one or more compounds that are T-type calcium channel antagonists, as disclosed herein, that can act through T-type calcium channels to reduce intracellular Miro1 levels.

[0099] In some embodiments, the composition comprises a compound of the present disclosure, or a salt thereof. In some embodiments, the composition further comprises a carrier or excipient.

[0100] In some embodiments, the present disclosure provides pharmaceutical compositions or formulations comprising a pharmaceutically effective amount of a compound of the present disclosure, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier or excipient. The pharmaceutical composition can deliver an amount of the compound of the present disclosure sufficient to produce a therapeutically effective treatment, as further described below. Also provided are pharmaceutical formulations comprising a pharmaceutically effective amount of a compound of Formula (I), (II), (III), (IV), and / or (V), or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier or excipient.

[0101] The compounds herein are formulated with conventional carriers and excipients selected in accordance with ordinary practice. Tablets contain excipients, glazes, fillers, binders, etc. Aqueous formulations are prepared in sterile form and, if intended for delivery other than oral administration, are generally isotonic. All formulations will optionally contain excipients such as those listed in the "Handbook of Pharmaceutical Excipients" (1986). Excipients include ascorbic acid and other antioxidants, chelating agents such as EDTA, and carbohydrates such as dextran, hydroxyalkylcellulose, hydroxyalkylmethylcellulose, and stearic acid. The pH of the formulations ranges from about 3 to about 11, but is usually about 7 to 10.

[0102] While it is possible for the active ingredients to be administered alone, it may be more preferable to present them as pharmaceutical formulations. Both veterinary and human formulations contain at least one active ingredient, as defined above, together with one or more acceptable carriers and any other therapeutic ingredients, particularly additional therapeutic ingredients as discussed herein. The carrier must be "acceptable" in the sense of being physiologically compatible with the other ingredients of the formulation and with the recipient.

[0103] Formulations include those suitable for the above-mentioned routes of administration. Formulations can be conveniently presented in unit dosage form and can be prepared by any of the methods well known in the pharmaceutical arts. In general, formulation techniques and preparations can be found in Remington's Pharmaceutical Sciences (Mack Publishing Co, Easton, PA). Such methods include the step of combining the active ingredient with the carrier, which constitutes one or more accessory ingredients. In general, formulations are prepared by uniformly and intimately contacting the active ingredient with liquid carriers or finely divided solid carriers, or both, and then, if necessary, shaping the product.

[0104] Formulations suitable for oral administration may be presented as discrete units such as capsules, sachets, or tablets, each containing a predetermined amount of the active ingredient; as a powder or granules; as a solution or suspension in an aqueous or non-aqueous liquid; or as an oil-in-water liquid or water-in-oil emulsion. The active ingredient may also be administered as a bolus, electuary, or paste.

[0105] Tablets are optionally prepared by compression or molding, with one or more accessory ingredients. Compressed tablets can be prepared by compressing the active ingredient in a free-flowing form such as powder or granules in a suitable machine. Optionally, they are mixed with binders, lubricants, inert diluents, preservatives, surfactants or dispersing agents, and diluents. Molded tablets can be prepared by molding a mixture of powdered active ingredient moistened with an inert liquid in a suitable machine. Tablets can optionally be provided with a film coating or scoring, and can optionally be formulated to provide slow or controlled release of the active ingredient therefrom.

[0106] The pharmaceutical preparations herein comprise a combination of one or more pharmaceutically acceptable carriers or excipients, optionally with other therapeutic agents. Pharmaceutical preparations containing the active ingredient may be in any form suitable for the intended administration method. For example, for oral use, tablets, troches, lozenges, aqueous or oily suspensions, dispersible powders or granules, emulsions, hard or soft capsules, liquids, syrups, or elixirs can be prepared. Compositions for oral use can be prepared according to any method known in the art for the manufacture of pharmaceutical compositions, and such compositions can contain one or more agents, such as sweeteners, flavorings, coloring agents, and preservatives, to provide a palatable formulation. Tablets containing the active ingredient in a mixture with non-toxic pharmaceutically acceptable excipients suitable for the manufacture of tablets are preferred. These excipients include, for example, inert diluents such as calcium or sodium carbonate, lactose, calcium or sodium phosphate; granulating and disintegrating agents such as corn starch or alginic acid; binders such as starch, gelatin, or acacia; and lubricants such as magnesium stearate, stearic acid, or talc. Tablets may be uncoated or may be coated by known techniques, including microencapsulation, to delay disintegration and adsorption in the gastrointestinal tract, thereby providing a sustained action over a longer period. For example, a time-delay material such as glyceryl monostearate or glyceryl distearate, alone or with a wax, may be employed.

[0107] Formulations suitable for topical administration to the eye also include eye drops in which the active ingredient is dissolved or suspended in a suitable carrier, particularly an aqueous solvent. The active ingredient may be present in a concentration of about 0.5 to about 20%, for example, about 0.5 to about 10%, e.g., about 1.5% (w / w). IV. Method (A) Assay

[0108] Calcium channel antagonist activity, including T-type calcium channel antagonist activity, can be evaluated using patch clamp assay or fluorescence imaging plate reader (FLIPR) assay known in the art.For example, see Leech, CA and Holz, GG, IV.Methods Cell Biol.(1994)40:135-151;Bell, DC and Dallas, ML, Br.J.Pharmacology(2018)175:2312-2321;Bezencon et al., J.Med.Chem.(2017)60:9769;Uebele et al., Cell Biochem Biophys(2009)55:81;Xiang et al., ACS Chemical Neuroscience(2011)2:730;and U.S. Patent No. 10,562,857.

[0109] A T-type calcium channel antagonist capable of reducing Miro1 levels and / or activity can be demonstrated, for example, by any convenient method in the art for detecting Miro1 levels and / or activity in the presence and absence of a T-type calcium channel antagonist. For example, the level and / or phosphorylation state of Miro protein (Ser156, Thr298, or Thr299 of Miro1 or Miro2; see, e.g., Wang et al., Cell (2011) 147(4):893-906) can be detected, for example, by immunoprecipitation with a mitochondrial transport protein-specific antibody followed by Western blotting with a phospho-specific or conventional antibody; a decrease in Miro protein phosphorylation and / or total Miro protein levels, or a decrease in Khc phosphorylation after contact with the agent, can indicate that the agent will treat Parkinson's disease. As an example, the level and / or ubiquitination of Miro protein can be detected, for example, by immunoprecipitation with a mitochondrial transport protein-specific antibody followed by immunoblotting with a ubiquitin-specific antibody, in which case an increase in ubiquitination after contact with a candidate agent may indicate that the agent will treat Parkinson's disease. As another example, the ability of a target mitochondrial protein to transport mitochondria within a cell can be assessed, for example, by treating cultured cells (e.g., neurons) with a T-type calcium channel antagonist and then observing intracellular mitochondrial transport compared to cells not treated with the T-type calcium channel antagonist, for example, using live cell imaging techniques (see, e.g., Brickley and Stephenson, J. Biol. Chem. 286(20):8079-92 (2011); Misko et al., J. Neurosci. 30(19):4232-40 (2010); Russo, GJ et al., J. Neurosci. 29(17):5443-55 (2009)).As another example, because the formation of a complex between Miro (e.g., Miro1 and Miro2), TRAK (e.g., TRAK1 and TRAK2), and Khc is essential for mitochondrial transport within neurons (see, e.g., Brickley & Stephenson, J. Biol. Chem. 286(20):18079-92(2011)), the effect of a T-type calcium channel antagonist on Miro function can be assessed by assessing the ability of Miro, TRAK, and Khc to form a complex in the presence of the T-type calcium channel antagonist. Such assessment can be performed using any technique for measuring protein-protein interactions, including, but not limited to, co-immunoprecipitation and affinity purification techniques. Such ability is assessed in cells with familial PD mutations, such as PINK1 or LRRK2 mutations.

[0110] Affinity assays, often immunoassays, are assays or analytical procedures that rely on the binding of a target molecule, i.e., Miro1, to a receptor, antibody, or another macromolecule. Detection methods are used to determine the presence and extent of the binding complex formed. Many formats for such assays are known in the art and are suitable for detecting Miro1 degradation associated with mitochondrial uncoupling or depolarization. In some embodiments, the assay format is suitable for high-throughput analysis.

[0111] Suitable assay formats include immunoassays that utilize antibodies specific for Miro1. Antibodies suitable for this purpose are commercially available, e.g., as polyclonal or monoclonal compositions, and include monoclonals CL1095 and CL1083 from Invitrogen, clone 4H4 from Sigma-Aldrich, and anti-RhoT1 antibody (A-8) from Santa Cruz Biotechnology.

[0112] Assays of interest include, for example, Western blot, immunohistochemistry, immunoprecipitation, and particularly immunoassays such as enzyme-linked immunosorbent assay (ELISA), radioimmunoassay (RIA), and enzyme-linked immunosorbent assay (EIA).

[0113] Enzyme-linked immunosorbent assays (ELISAs) are used to qualitatively and quantitatively analyze the presence or concentration of specific soluble antigens, such as Miro1, in liquid samples such as cell lysates. These assays generally utilize the ability of multiwell plates to bind antibodies that capture the cognate antigen. Colorimetric endpoint methods are typically used, which can be detected by absorbance wavelength and quantified from a known standard curve of antigen or antibody dilutions. Detection antibodies are often labeled with enzymes such as horseradish peroxidase or alkaline phosphatase, fluorescent tags, electrochemiluminescent labels, or intermediate labels such as biotin.

[0114] Common ELISA formats include sandwich ELISA, so named because the analyte is sandwiched between two different antibodies. The capture substrate in this format is a capture antibody, often a monoclonal antibody, which increases assay specificity and reduces background noise. The analyte is bound to the capture antibody and then detected by binding to the detection antibody. A variation of the monoclonal ELISA assay, called a single molecule assay (Simoa), uses beads coated with a capture antibody; each bead binds to one or zero target molecules, and each bead is detected using a separate antibody (detection antibody) and a labeling enzyme.

[0115] Other ELISA formats include indirect ELISA, in which the capture substrate is the specific antigen to be tested for and the detection process is mediated by a primary antibody and an enzyme-conjugated secondary antibody reactive with it. Thus, the primary antibody that recognizes the antigen is not labeled. In direct ELISA, the capture substrate is the specific antigen to be tested for. An enzyme that catalyzes a color-changing reaction is conjugated (bound) to the antigen-detecting antibody.

[0116] Immuno-PCR (I-PCR) is a technique that combines the sensitivity of nucleic acid amplification by PCR with the specificity of antibody-based assays to enhance detection sensitivity.

[0117] An exemplary method for measuring the reduction of Miro1 following compound administration is described in Hsieh, CH, Li, L, Vanhauwaert, R, Nguyen, KT, Davis, MD, Bu, G et al., Miro1 Marks Parkinson's Disease Subset and Miro1 Reducer Rescues Neuron Loss in Parkinson's Models. Cell Metab. (2019) 1131-1140.

[0118] B. How to Reduce Miro1 Provided herein are methods for reducing intracellular Miro1 levels, comprising contacting a cell with an effective amount of a T-type calcium channel antagonist. In some embodiments, the T-type calcium channel antagonist has the structure of one or more compounds of formula (I), (II), (III), (IV), and / or (V) described herein. For example, the method for reducing intracellular Miro1 levels can be carried out using any one of the compounds in Tables 1, 2, and 3, or a pharmaceutically acceptable salt thereof. The present disclosure further demonstrates that calcium channel antagonists having at least some activity against T-type calcium channels can result in a dose-dependent reduction in Miro1 levels in Western blot and fibroblast assays (Examples 61 and 62). Compounds with T-type calcium channel antagonist activity and the ability to decrease Miro1 include benidipine (1,4-dihydro-2,6-dimethyl-4-(3-nitrophenyl)-3,5-pyridinedicarboxylic acid methyl 1-(phenylmethyl)-3-piperidinyl ester); MK-8998 ((R)-2-(4-isopropylphenyl)-N-(1-(5-(2,2,2-trifluoroethoxy)pyridin-2-yl)ethyl)acetamide); ABT-639 (5-[(8aR)-3,4,6,7,8,8a-hexahydro-1H-pyrrolo[1,2-a]pyrazine-2-yl]ethyl)acetamide); carbonyl]-4-chloro-2-fluoro-N-(2-fluorophenyl)benzenesulfonamide; ACT-709478 (N-(1-((5-cyanopyridin-2-yl)methyl)-1H-pyrazol-3-yl)-2-(4-(1-(trifluoromethyl)cyclopropyl)phenyl)acetamide); and zonisamide (1,2-benzisoxazole-3-methanesulfonamide). One exemplary compound, benidipine, shows in vivo activity in rescuing locomotor defects in a Drosophila model of Parkinson's disease (Example 63).

[0119] T-type calcium channel antagonists are described in U.S. Patent No. 8,377,968; U.S. Patent No. 9,403,798; U.S. Patent No. 10,562,857; U.S. Patent Application Publication No. 20120264804; U.S. Patent No. 20150329533; U.S. Patent No. 20160340322; European Patent No. EP3572403; PCT Publication No. WO2011022315; WO2012094615; WO2013148640; WO2018200850; WO2019175395; WO2020072773; and WO021007487, each of which is incorporated herein by reference in its entirety.Any one of the T-type calcium channel antagonists in the publications described above can be used in the methods described herein.

[0120] In the methods described herein, a reduction in Miro1 levels refers to a reduction in the amount and / or activity of Miro1 protein. In some embodiments, a reduction in Miro1 levels refers to a reduction in the amount of Miro1 protein, resulting in a reduction in Miro1 activity, as determined by any assay, including assays known in the art and described in this disclosure. In Examples 37-39 described herein, T-type calcium channel antagonists did not induce the Miro1 levels observed in naive healthy fibroblasts or control iPSC DA neuron cells in disease-derived fibroblasts or iPSC DA neuron cells. Furthermore, T-type calcium channel antagonists did not reduce Miro1 levels in disease-derived fibroblasts or iPSC DA neuron cells to levels lower than the control levels observed in naive healthy fibroblasts or control iPSC DA neuron cells.

[0121] Any suitable cells can be used in the methods of reducing or downregulating Miro1 levels described herein. Cultured cells can be derived from a patient or control sample and can be modified to produce genetically modified cells, cells differentiated in vitro, cells exposed to a candidate therapeutic agent, etc. In some embodiments, the cells are muscle cells. For example, the muscle cells can be cardiac cells, i.e., cardiomyocytes. In some embodiments, the cells are kidney cells. In some embodiments, the cells are hepatocytes. In some embodiments, the cells are neuronal cells. The method can be performed on cells in vitro, ex vivo, or in vivo. In some embodiments, the reduced Miro1 levels are in vitro or ex vivo. In some embodiments, the reduced Miro1 levels are in vivo.

[0122] The methods described herein can be used with any suitable biological sample containing cells. The methods can be performed using biological samples obtained from subjects, including but not limited to fibroblasts. Examples include skin fibroblasts, peripheral blood lymphocytes, iPSCs, etc.

[0123] Miro1 levels measured using the methods described herein can be compared to control Miro1 levels using any method known in the art, see, for example, the ELISA assay described in Hsieh, CH et al., Cell Meta (2019) 1131-1140.

[0124] In some embodiments, the T-type calcium channel antagonist reduces Miro1 levels to the normal range. For example, in some embodiments, the Miro1 normal range can be the range observed between untreated or naive healthy fibroblasts or iPSC DA neuron cells (at the high end of the normal range) and healthy fibroblasts or iPSC DA neuron cells challenged with a mitochondrial stress response factor (at the low end of the range).

[0125] In some embodiments, the T-type calcium channel antagonist reduces or down-regulates the level of Miro1 to within about 50%, about 40%, about 30%, or about 20% of the control level of Miro1. In some embodiments, the control level of Miro1 is measured in control cells from a control subject who does not have or is not suspected of having a disease or disorder mediated by abnormal Miro1 levels. For example, the T-type calcium channel antagonist can down-regulate the Miro1 level in neuronal cells from a Parkinson's disease patient to within about 50% of the control level of Miro1 in control neuronal cells from an age-matched patient who does not have or is not suspected of having Parkinson's disease.

[0126] The Miro1 level in a cell after contact with a T-type calcium channel antagonist may be higher or lower than the control level of Miro1 in a control cell. In some embodiments, the Miro1 level in a cell after contact with a T-type calcium channel antagonist is about 20%, about 30%, about 40%, or about 50% higher than the control Miro1 level in a control cell. In some embodiments, the Miro1 level in a cell after contact with a T-type calcium channel antagonist is about 20% to about 50% higher than the control Miro1 level in a control cell. In some embodiments, the Miro1 level in a cell after contact with a T-type calcium channel antagonist is about 20%, about 30%, about 40%, or about 50% lower than the control Miro1 level in a control cell. In some embodiments, the Miro1 level in a cell after contact with a T-type calcium channel antagonist is about 20% to about 50% lower than the control Miro1 level in a control cell.

[0127] Any suitable concentration of intracellular T-type calcium channel antagonist can be used to reduce intracellular Miro1 levels to a desired level. In some embodiments, the intracellular concentration of the T-type calcium channel antagonist can be about 1 nM to about 100 μM, e.g., about 1 nM to about 10 μM, about 1 nM to about 1 μM, about 10 nM to about 100 μM, about 10 nM to about 1 μM, about 100 nM to about 100 μM, about 100 nM to about 1 μM, about 1 μM to about 100 μM, or about 1 μM to about 10 μM.

[0128] In some embodiments, the T-type calcium channel antagonist reduces the level or biological activity of Miro1 by about 20% or more, e.g., about 30% or more, about 40% or more, or about 50% or more, about 60% or more, about 70% or more, or about 80% or more, e.g., about 90%, about 95%, or about 100%, relative to an untreated control not contacted with the T-type calcium channel antagonist. In some embodiments, the T-type calcium channel antagonist reduces the level or biological activity of Miro1 by about 20% relative to an untreated control not contacted with the T-type calcium channel antagonist. In some embodiments, the T-type calcium channel antagonist reduces the level or biological activity of Miro1 by about 25% relative to an untreated control not contacted with the T-type calcium channel antagonist. In some embodiments, the T-type calcium channel antagonist reduces the level or biological activity of Miro1 by about 30% relative to an untreated control not contacted with the T-type calcium channel antagonist. In some embodiments, the T-type calcium channel antagonist reduces the level or biological activity of Miro1 by about 35% relative to an untreated control not contacted with the T-type calcium channel antagonist. In some embodiments, the T-type calcium channel antagonist reduces the level or biological activity of Miro1 by about 40% relative to an untreated control not contacted with the T-type calcium channel antagonist. In some embodiments, the T-type calcium channel antagonist reduces the level or biological activity of Miro1 by about 45% relative to an untreated control not contacted with the T-type calcium channel antagonist. In some embodiments, the T-type calcium channel antagonist reduces the level or biological activity of Miro1 by about 50% relative to an untreated control not contacted with the T-type calcium channel antagonist. In some embodiments, the T-type calcium channel antagonist reduces the level or biological activity of Miro1 by about 55% relative to an untreated control not contacted with the T-type calcium channel antagonist.In some embodiments, the T-type calcium channel antagonist reduces the level or biological activity of Miro1 by about 60% relative to an untreated control not contacted with the T-type calcium channel antagonist. In some embodiments, the T-type calcium channel antagonist reduces the level or biological activity of Miro1 by about 65% relative to an untreated control not contacted with the T-type calcium channel antagonist. In some embodiments, the T-type calcium channel antagonist reduces the level or biological activity of Miro1 by about 70% relative to an untreated control not contacted with the T-type calcium channel antagonist. In some embodiments, the T-type calcium channel antagonist reduces the level or biological activity of Miro1 by about 75% relative to an untreated control not contacted with the T-type calcium channel antagonist. In some embodiments, the T-type calcium channel antagonist reduces the level or biological activity of Miro1 by about 80% relative to an untreated control not contacted with the T-type calcium channel antagonist. In some embodiments, the T-type calcium channel antagonist reduces the level or biological activity of Miro1 by about 85% relative to an untreated control not contacted with the T-type calcium channel antagonist. In some embodiments, the T-type calcium channel antagonist reduces the level or biological activity of Miro1 by about 90% relative to an untreated control not contacted with the T-type calcium channel antagonist. In some embodiments, the T-type calcium channel antagonist reduces the level or biological activity of Miro1 by about 95% relative to an untreated control not contacted with the T-type calcium channel antagonist.

[0129] In some embodiments, the T-type calcium channel antagonist reduces the level or biological activity of Miro1 by about 20% to about 100% relative to an untreated control not contacted with the T-type calcium channel antagonist. In some embodiments, the T-type calcium channel antagonist reduces the level or biological activity of Miro1 by about 20% to about 90% relative to an untreated control not contacted with the T-type calcium channel antagonist. In some embodiments, the T-type calcium channel antagonist reduces the level or biological activity of Miro1 by about 20% to about 80% relative to an untreated control not contacted with the T-type calcium channel antagonist. In some embodiments, the T-type calcium channel antagonist reduces the level or biological activity of Miro1 by about 20% to about 70% relative to an untreated control not contacted with the T-type calcium channel antagonist. In some embodiments, the T-type calcium channel antagonist reduces Miro1 levels or biological activity by about 20% to about 60% relative to an untreated control not contacted with the T-type calcium channel antagonist. In embodiments, the T-type calcium channel antagonist reduces Miro1 levels or biological activity by about 20% to about 50% relative to an untreated control not contacted with the T-type calcium channel antagonist. In some embodiments, the T-type calcium channel antagonist reduces Miro1 levels or biological activity by about 20% to about 40% relative to an untreated control not contacted with the T-type calcium channel antagonist. In some embodiments, the T-type calcium channel antagonist reduces Miro1 levels or biological activity by about 30% to about 50% relative to an untreated control not contacted with the T-type calcium channel antagonist. In some embodiments, the T-type calcium channel antagonist reduces Miro1 levels or biological activity by about 40% to about 60% relative to an untreated control not contacted with the T-type calcium channel antagonist.

[0130] C. Methods for identifying subjects at risk for developing a Miro1-associated disorder A defect in the ability to degrade or clear (exclude) Miro1 from cells is believed to correlate with the onset of a Miro1-associated disorder, e.g., a neurodegenerative disorder such as Parkinson's disease, e.g., before the subject (patient) exhibits overt symptoms of the neurodegenerative disorder, e.g., one or more of the symptoms described herein. Thus, in some embodiments, the subject is asymptomatic for the neurodegenerative disorder. Example 64 presents experimental data supporting the correlation of a defect in the ability to excrete Miro1 from cells derived from Parkinson's disease patients or from individuals at risk for developing Parkinson's disease. See also Figures 2-5, which show the correlation of high Miro1 ratios with Parkinson's disease in patient-derived skin fibroblasts and iPSC cells. Thus, Miro1 can be used as a predictive biomarker for neurodegenerative disease in subjects at risk for developing such disorders, e.g., as an early step in treating neurodegenerative disease before symptoms appear. At-risk subjects may have a family history of developing a neurodegenerative disorder, may exhibit, for example, genetic markers associated with an increased risk of developing a neurodegenerative disorder, such as the LRRK2 G2019S mutation associated with Parkinson's disease, or may have no known risk of developing a neurodegenerative disease.

[0131] As described in Hsieh, CH et al., Cell Meta (2019) 1131-1140 and PCT Publication WO 2021 / 046368, Miro1 levels in cells treated with mitochondrial stress response factors are expected to be lower than control Miro1 levels in untreated controls because the mitophagy process is induced by mitochondrial stress response factors. In biological sample cells derived from a subject, Miro1 levels equal to or higher than control Miro1 levels in cells treated with mitochondrial stress response factors can indicate a neurodegenerative disorder correlated with defective mitophagy.

[0132] In some embodiments, a method for identifying a subject at risk of developing a Miro1-associated disorder includes: (a) detecting whether the Miro1 level in a biological sample obtained from the subject treated with a mitochondrial stress response factor is equal to or greater than the control Miro1 level in a control biological sample; and (b) identifying the subject as at risk of developing a Miro1-associated disorder if the Miro1 level in the biological sample is equal to or greater than the control Miro1 level in the biological sample, wherein the biological sample and the control biological sample contain iPSCs or cells differentiated from iPSCs. In some embodiments, the method further includes treating the subject at risk of developing a Miro1-associated disorder by administering a therapeutically effective amount of a compound described herein or a pharmaceutically acceptable salt thereof. In certain embodiments, the compound is a compound described in Table 1 or a pharmaceutically acceptable salt thereof.

[0133] In some embodiments, a method for identifying a subject at risk of developing a Miro1-associated disorder includes: (a) detecting whether the Miro1 level in a biological sample obtained from the subject treated with a mitochondrial stress response factor is equal to or greater than the control Miro1 level in a control biological sample obtained from the same subject but untreated; (b) identifying the subject as at risk of developing a Miro1-associated disorder if the Miro1 level in the biological sample is equal to or greater than the control Miro1 level in the control biological sample; and (c) treating the subject at risk of developing a Miro1-associated disorder by administering a therapeutically effective amount of a compound described herein or a pharmaceutically acceptable salt thereof. In some embodiments, the compound is a compound of Formula (I), (II), (III), (IV), and / or (V), or a pharmaceutically acceptable salt thereof. In certain embodiments, the compound is a compound described in Table 1, or a pharmaceutically acceptable salt thereof.

[0134] A Miro-1-associated disorder is any disease or disorder correlated with abnormal degradation and / or clearance of Miro1 protein, including any one of the neurodegenerative diseases described herein. In some embodiments, the Miro1-associated disorder is drug-induced parkinsonism, progressive supranuclear palsy, vascular parkinsonism, dementia with Lewy bodies, diffuse Lewy body disease, corticobasal degeneration, multiple system degeneration (Shy-Drager syndrome), Parkinson's disease, Alzheimer's disease, Pick's disease, frontotemporal dementia, multiple system atrophy, vascular dementia, or progressive supranuclear palsy (Steel-Richardson syndrome). In some embodiments, the Miro-1-associated disorder is Parkinson's disease. In some embodiments, the Miro1-associated disorder is Alzheimer's disease. In some embodiments, the Miro1-associated disorder is Pick's disease. In some embodiments, the Miro1-associated disorder is frontotemporal dementia. In some embodiments, the Miro1-associated disorder is multiple system atrophy.

[0135] Also provided is a method for treating a neurodegenerative disease in a subject in need of treatment, the method comprising: a) detecting whether the Miro1 level in a biological sample obtained from the subject and treated with a mycotondrial stress response factor is equal to or higher than the control Miro1 level in a control biological sample obtained from the subject and untreated; b) determining whether the Miro1 level in the biological sample is equal to or higher than the control Miro1 level in a control biological sample obtained from the subject and untreated; and c) administering to the subject a therapeutically effective amount of a compound as described herein, or a pharmaceutically acceptable salt thereof. In some embodiments, the neurodegenerative disease is drug-induced parkinsonism, progressive supranuclear palsy, vascular parkinsonism, dementia with Lewy bodies, diffuse Lewy body disease, corticobasal degeneration, multiple system degeneration (Shy-Drager syndrome), Parkinson's disease, Alzheimer's disease, Pick's disease, frontotemporal dementia, multiple system atrophy, vascular dementia, or progressive supranuclear palsy (Steel-Richardson syndrome). In some embodiments, the compound is a compound of Formula (I), (II), (III), (IV), and / or (V), or a pharmaceutically acceptable salt thereof. In some embodiments, the compound is a compound set forth in Table 1, or a pharmaceutically acceptable salt thereof. Example 65 illustrates one compound of the present disclosure, Example 15, demonstrating dose-dependent rescue of Miro1 defects in fibroblasts from TauP301L donors at risk for developing frontotemporal dementia (FID).

[0136] Any suitable biological sample described herein can be used in this method. In some embodiments, the biological sample and the control biological sample comprise fibroblasts. For example, skin fibroblasts can be obtained directly from a subject. In some embodiments, the biological sample and the control biological sample comprise iPSCs or cells differentiated from iPSCs.

[0137] iPSCs can be obtained directly from a subject or cultured from other cell types obtained from a subject according to any method known in the art. See Shi, Y et al., AL Nature Review, Vol. 16, pp. 115-130 (2017), and references cited therein. For example, iPSCs can be dedifferentiated from fibroblasts obtained directly from a subject. Furthermore, iPSCs can be redifferentiated into a variety of different cell types, including neurons, skin cells, blood cells, and liver cells.

[0138] Such cells differentiated from a subject's iPSCs can be used to conveniently determine personalized treatments without obtaining target cell types directly from the subject. In an exemplary embodiment, skin fibroblasts can be obtained from a subject at risk for developing Parkinson's disease. The skin fibroblasts can be dedifferentiated into iPSCs, which can then be redifferentiated into motor neurons. Motor neurons differentiated from iPSCs can be tested in the assays described herein for Miro1 deficiency, with or without treatment with a mitochondrial stress response factor, to identify whether the subject will respond to a Miro1-reducing therapy, such as one containing a compound of the present disclosure.

[0139] Methods for comparing Miro1 levels with control Miro1 levels are known in the art. When determining whether Miro1 levels in a biological sample obtained from a subject and treated with a mitochondrial stress response factor are equal to or greater than the control Miro1 level in an untreated control biological sample obtained from the same subject, the ratio of the Miro1 level to the control Miro1 level can be compared. In some embodiments, the ratio of the Miro1 level to the control Miro1 level is about 0.5 to about 10, e.g., about 0.5 to about 5, about 0.6 to about 6, about 0.7 to about 4, about 0.7 to about 3, about 0.8 to about 3, or about 0.9 to about 2. For example, the ratio of the Miro1 level to the control Miro1 level can be about 0.5 to about 10; in another example, the ratio of the Miro1 level to the control Miro1 level can be about 0.7 to about 4.

[0140] In the method described herein, any mitochondrial stress response factor known in the art can be used.Suitable mitochondrial stress response factors include mitochondrial depolarizing agents, such as carbonyl cyanide 4-(trifluorooxy)phenylhydrazone (FCCP) and carbonyl cyanide 3-chlorophenylhydrazone (CCCP); mitochondrial electron transport chain inhibitors, such as complex I inhibitors, such as rotenone, piericidin A, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and paraquat, complex III inhibitors, such as antimycin A, complex V inhibitors, such as oligomycin A, and nitrochondria membrane potassium ionophores, such as valinomycin; metabolic modulators, such as insulin signaling modulators, such as metformin, and inhibitors of the mTOR main signaling pathway required for cell growth and metabolism, such as rapamycin.

[0141] D. Treatment method Provided herein are methods for treating a neurodegenerative disorder, comprising administering to a subject a therapeutically effective amount of a T-type calcium channel antagonist described herein, or a pharmaceutical composition thereof. In some embodiments, the T-type calcium channel antagonist is a compound of Formula (I), (II), (III), (IV), and / or (V), or a pharmaceutically acceptable salt thereof. In some embodiments, the method delivers a therapeutically effective amount of a compound of the present disclosure, or a pharmaceutical composition thereof, sufficient to treat one or more symptoms of the conditions described below.

[0142] Neurodegenerative disorders included in the methods of the present disclosure include, but are not limited to, neurological disorders that share similar symptoms with Parkinson's disease-related disorders. In some cases, neurological disorders may exhibit symptoms similar to Parkinson's disease, atypical Parkinson's disease, or Parkinson's plus disease. Examples include, but are not limited to, drug-induced parkinsonism, progressive supranuclear palsy, vascular parkinsonism, dementia with Lewy bodies, diffuse Lewy body disease, corticobasal degeneration, multiple system degeneration (Shy-Drager syndrome), Alzheimer's disease, Pick's disease, frontotemporal dementia, multiple system atrophy, vascular dementia, and progressive supranuclear palsy (Steel-Richardson syndrome). Other diseases included in the method of the present invention include age-related dementia, vascular dementia, diffuse white matter disease (Binswanger's disease), dementia of endocrine or metabolic origin, dementia due to head trauma, and diffuse brain injury, dementia pugilistica, and frontal lobe dementia. In some cases, neurological diseases do not respond well to dopaminergic treatment and can be caused by various vascular, drug-related, infectious, toxic, structural, and other known secondary causes. Drug-induced parkinsonism can be caused by drugs that block high-affinity postsynaptic dopamine D2 receptors, such as antipsychotics and antiemetics, as well as sodium valproate, antidepressants, reserpine, tetrabenazine, etc.

[0143] Various subjects (patients) are suitable for treatment with the T-type calcium channel antagonists of the present disclosure.Suitable subjects include those with Parkinson's disease symptoms, such as bradykinesia, repetitive movements, tremors, limb stiffness, gait and balance problems, loss of eye muscle strength, loss of ability to aim, weakness, sensory loss, non-motor symptoms such as REM sleep behavior disorder, mood disorders and neuropsychiatric symptoms including cognitive changes, anxiety, apathy, changes in thinking ability, attention or alertness level, as well as visual hallucinations, intellectual and functional deterioration, forgetfulness, personality changes, autonomic dysfunction affecting the cardiovascular system, respiratory system, genitourinary system, gastrointestinal tract and sweating function, difficulty breathing and swallowing, inability to sweat, orthostatic hypotension, pain, constipation, and a decreased sense of smell, such as anosmia.In some embodiments, patients (subjects) may experience significant speech or language disorders, prominent frontal presentation (PSP-F) and freezing of gait.

[0144] In some embodiments, the subject may not exhibit any overt symptoms of Parkinson's disease. In some cases, the subject may exhibit infections, low body temperature, brittle bones, stiff joints, arthritis, hunched posture, slow movements, low overall energy, constipation, urinary incontinence, memory loss, slower thinking, impaired reflexes, poor balance, decreased vision, reduced peripheral vision, hearing loss, wrinkled skin, greasy hair, weight loss, or loss of muscle tissue.

[0145] In some embodiments, the subject is selected from those diagnosed with Alzheimer's disease; subjects who have experienced one or more strokes; subjects who have suffered a traumatic head injury; individuals with high serum cholesterol levels; subjects with proteinaceous lesions, including deposits in brain tissue; subjects who have had one or more cardiac events; subjects undergoing cardiac surgery; and subjects with multiple sclerosis.

[0146] In some embodiments, the subject has a neurological disorder including amyotrophic lateral sclerosis, degenerative ataxia, motor neuron diseases such as corticobasal degeneration, ALS-Parkinsonism dementia complex (ALS-PDC) disease in Guam, subacute sclerosing panencephalitis nephroscleritis, Huntington's disease, Parkinson's disease, synucleinopathy, primary progressive aphasia, striatonigral degeneration, Machado-Joseph disease / Spinocerebellar ataxia type 3 and olivopontine camphoratic degeneration, Gilles De La Tourette's disease, Symptoms may be associated with neurological disorders including motor neuron diseases such as Tourette's syndrome, bulbar and pseudobulbar palsy, spinal and spinal muscular atrophy (Kennedy's disease), primary lateral sclerosis, familial spastic paraplegia, Werdnich-Hoffmann disease, Kugelberg-Welander disease, Tay-Sachs disease, Sandohoff disease, familial spastic disorders, Wohlfart-Kugelberg-Welander disease, spastic paraplegia, progressive multifocal leukoencephalopathy, and prion diseases (including Creutzeldt-Jakob disease, Gestmann-Sträussler-Scheinker disease, Kuru, and fatal familial insomnia). It may also refer to other neurodegenerative disorders resulting from cerebral ischemia or cerebral infarction, including embolic and thrombotic occlusion, and any type of intracranial hemorrhage (including, but not limited to, epidural, subdural, subarachnoid and intracerebral hemorrhage), and intracranial and intraspinal lesions (including, but not limited to, contusion, penetrating, shearing, compression, and laceration). [Example]

[0147] V. Working Examples JPEG2024508903000031.jpg219170

[0148] Example 1 (R)-2-(4-isopropylphenyl)-N-(1-(1-methyl-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide Step 1: Synthesis of 5-bromo-1H-pyrazolo[3,4-c]pyridine [ka]

[0149] To a solution of 6-bromo-4-methylpyridin-3-amine (1.0 g) in acetic acid (60 mL) was added sodium nitrite (0.37 mL) while cooling in an ice bath. The reaction was stirred at room temperature overnight. The reaction mixture was then concentrated and treated with saturated aqueous NaHCO3. The aqueous mixture was extracted with EA. The organic layer was separated, washed with brine, dried over Na2SO4, filtered, and concentrated in vacuo. The residue was purified by silica gel column chromatography to give 5-bromo-1H-pyrazolo[3,4-c]pyridine (442 mg). LC / MS ESI (m / z): 198 [M+H] + .

[0150] Step 2: Synthesis of 1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde [ka]

[0151] To a solution of 5-bromo-1H-pyrazolo[3,4-c]pyridine (3.3 g) in anhydrous THF (100 mL) was added n-BuLi (20.0 mL, 2N in THF) at -78 °C. The mixture was stirred at -78 °C for 1 h. Then, DMF (3.87 mL) was added to the mixture at -78 °C and stirred at -78 °C for 1 h. The reaction mixture was warmed to room temperature and quenched with NH4Cl solution. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over Na2SO4, filtered, and concentrated to give a residue that was purified by column chromatography on silica gel (DCM:MeOH = 1:0 to 20:1) to give 1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde (1.0 g) as a white solid. LC / MS ESI (m / z): 148 [M+H] + .

[0152] Step 3: Synthesis of 1-methyl-1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde [ka]

[0153] To a solution of 1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde (1 g) in DMF (15 mL) was added CsCO (4.4 g) and iodomethane (1.1 g). The mixture was stirred at 25 °C for 12 h. The mixture was then diluted with water and extracted with EtOAc. The combined extracts were dried, filtered, and concentrated to give 1-methyl-1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde (400 mg) [ 1 H-NMR (400 MHz, DMSO-d6) δ 10.12 (s, 1H); 9.35 (s, 1H); 8.43 (s, 2H); 4.25 (s, 3H)] and 2-methyl-2H-pyrazolo[3,4-c]pyridine-5-carbaldehyde (200 mg) [ 1 H-NMR (400 MHz, DMSO-d6) δ 10.05 (s, 1H); 9.27 (s, 1H); 8.80 (s, 1H); 8.43 (d, J = 1.2 Hz, 1H); 4.31 (s, 3H)] was obtained as a yellow solid. LC / MS ESI (m / z): 162 [M+H] + .

[0154] Step 4: Synthesis of (R,E)-2-methyl-N-((1-methyl-1H-pyrazolo[3,4-c]pyridin-5-yl)methylene)propane-2-sulfinamide [ka]

[0155] To a solution of 1-methyl-1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde (235 mg) in DCM (15 mL) was added (R)-2-methylpropane-2-sulfinamide (230 mg) and CuSO (349 mg). The reaction mixture was stirred at room temperature overnight. The solution was filtered through a pad of Celite, and the filtrate was concentrated. The residue was purified by silica gel column to give (R,E)-2-methyl-N-((1-methyl-1H-pyrazolo[3,4-c]pyridin-5-yl)methylene)propane-2-sulfinamide (340 mg). 1 H-NMR (400MHz, CDCl3) δ9.06(s,1H);8.83(s,1H);8.34(s,1H);8.17(s,1H);4.25(s,3H);1.31(s,9H)ppm. LC / MS ESI(m / z):265[M+H] + .

[0156] Step 5: Synthesis of (R)-2-methyl-N-((R)-1-(1-methyl-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)propane-2-sulfinamide [ka] To a solution of (R,E)-2-methyl-N-((1-methyl-1H-pyrazolo[3,4-c]pyridin-5-yl)methylene)propane-2-sulfinamide (347 mg) in anhydrous THF (20 mL) was added dropwise CHMgBr (3.0 M in ether, 2.2 mL). The reaction was stirred at −60° C. for 2 hours. The reaction was then quenched with saturated NHCl and extracted with EtOAc. The organic layer was separated, washed with additional saturated NaCl, dried over NaSO, filtered, and concentrated in vacuo. The residue was then purified by silica gel column chromatography to give (R)-2-methyl-N-((R)-1-(1-methyl-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)propane-2-sulfinamide (314 mg) as a pale yellow oil. LC / MS ESI(m / z):281[M+H] + .

[0157] Step 6: Synthesis of (R)-1-(1-methyl-1H-pyrazolo[3,4-c]pyridin-5-yl)ethan-1-amine hydrochloride [ka] To a solution of (R)-2-methyl-N-((R)-1-(1-methyl-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)propane-2-sulfinamide (57 mg) in dioxane (2.0 mL), HCl-dioxane (4 M in dioxane, 1.0 mL) was added, and the reaction mixture was stirred at room temperature for 30 minutes. The mixture was then concentrated to give crude (R)-1-(1-methyl-1H-pyrazolo[3,4-c]pyridin-5-yl)ethan-1-amine hydrochloride, which was used in the next step. LC / MS ESI (m / z): 177 [M+H] + .

[0158] Step 7: Synthesis of (R)-2-(4-isopropylphenyl)-N-(1-(1-methyl-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide [ka]

[0159] A solution of crude (R)-1-(1-methyl-1H-pyrazolo[3,4-c]pyridin-5-yl)ethan-1-amine hydrochloride (70 mg), 2-[4-(propan-2-yl)phenyl]acetic acid (77.88 mg), EDCI (91.38 mg), HOBt (64.41 mg), and DIEA (0.08 mL) in DMF (5 mL) was stirred at 25° C. for 3 hours. The mixture was diluted with water and extracted with EtOAc. The combined extracts were dried, filtered, and concentrated to give a residue. The residue was purified by preparative HPLC [column: YMC-Actus Triart C18, 250 × 21 mm; mobile phase: 20%–95% MeCN / HO (0.1% NH₃·HO); flow rate: 15 mL / min; wavelength: 220 nm / 254 nm] and SFC [column: ChiralPak® IA, 250 × 21.3 mm internal diameter (ID), 5 μm; mobile phase: A CO₂, B MEOH + 0.1% NH₃·HO, gradient: 35% B, wavelength: 220 nm] to give (R)-2-(4-isopropylphenyl)-N-(1-(1-methyl-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide (17.1 mg) as a white solid. LC / MS ESI (m / z): 337 (M+H). + . 1 H-NMR(400MHz,CDCl3)δ8.83(s,1H);7.98(s,1H);7.50(s,1H);7.20(s,4H);6.62(d,J=7.6Hz,1H);5.30-5.2 3(m,1H);4.18(s,3H);3.60-3.52(m,2H);2.95-2.88(m,1H);1.46(d,J=6.8Hz,3H);1.26(d,J=8.0Hz,6H)ppm. Example 2 (R)-2-(4-isopropylphenyl)-N-(1-(2-methyl-2H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide

[0160] Step 1: Synthesis of (R,E)-2-methyl-N-((2-methyl-2H-pyrazolo[3,4-c]pyridin-5-yl)methylene)propane-2-sulfinamide [ka]

[0161] To a solution of 2-methyl-2H-pyrazolo[3,4-c]pyridine-5-carbaldehyde (210 mg) in anhydrous dichloromethane (10 mL), (R)-2-methylpropane-2-sulfinamide (190 mg) and CuSO4 (520 mg) were added, and the reaction mixture was stirred at room temperature overnight.The mixture was then filtered, and the filtrate was concentrated under reduced pressure.The residue was purified by silica gel column to obtain (R,E)-2-methyl-N-((2-methyl-2H-pyrazolo[3,4-c]pyridin-5-yl)methylene)propane-2-sulfinamide (421 mg, crude) as a white solid.LC / MS ESI: 265 (M+H) + . 1 H-NMR (400MHz, DMSO) δ9.25(s,1H);8.71(s,1H);8.59(s,1H);8.49(s,1H);4.31(s,3H);1.21(s,9H).

[0162] Step 2: Synthesis of (R)-2-methyl-N-((R)-1-(2-methyl-2H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)propane-2-sulfinamide [ka]

[0163] To a solution of (R,E)-2-methyl-N-((2-methyl-2H-pyrazolo[3,4-c]pyridin-5-yl)methylene)propane-2-sulfinamide (274 mg) in THF (8.0 mL) was added methylmagnesium bromide (1.38 mL, 3 M in ether) at −78° C., and the mixture was stirred at −78° C. for 4 h. The mixture was then quenched with NH4Cl solution and extracted with EtOAc. The combined extracts were dried, filtered, and concentrated to give (R)-2-methyl-N-((R)-1-(2-methyl-2H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)propane-2-sulfinamide (285 mg). LC / MS ESI: 281 (M+H). + . 1H-NMR(400MHz,CDCl3)δ9.20(s,1H);7.90(s,1H);7.49(d,J=0.8Hz,1H);4.65-4.62(m ,1H);4.43(d,J=6.0Hz,1H);4.28(d,J=3.0Hz,3H);1.57(d,J=6.7Hz,3H);1.25(s,9H).

[0164] Step 3: Synthesis of (R)-1-(2-methyl-2H-pyrazolo[3,4-c]pyridin-5-yl)ethan-1-amine hydrochloride [ka]

[0165] To a solution of (R)-2-methyl-N-((R)-1-(2-methyl-2H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)propane-2-sulfinamide (215 mg) in dioxane (6 mL) was added 4N HCl-dioxane (2 mL) and the reaction was stirred at room temperature for 1 hour. The reaction was then concentrated and used directly in the next step. LC / MS ESI: 177 (M+H). + .

[0166] Step 4: Synthesis of (R)-2-(4-isopropylphenyl)-N-(1-(2-methyl-2H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide [ka]

[0167] (R)-1-(2-Methyl-2H-pyrazolo[3,4-c]pyridin-5-yl)ethan-1-amine hydrochloride (135 mg), EDCI (177 mg), HOBt (125 mg), and 2-(4-isopropylphenyl)acetic acid (151 mg) were dissolved in anhydrous N,N-dimethylformamide (5.0 mL), and then DIEA (0.51 mL, 3.064 ml) was added to the solution. The reaction mixture was stirred at room temperature for 1 hour and extracted from water with ethyl acetate. The organic layer was separated, washed with saturated aqueous NaCl, and concentrated in vacuo. The residue was purified by preparative HPLC [column: YMC-Actus Triart C18, 250 × 21 mm; mobile phase: 20%–95% MeCN / HO (0.1% NH₃·HO); flow rate: 15 mL / min; wavelength: 220 nm / 254 nm] and SFC [column: ChiralPak® IA, 250 × 21.3 mm internal diameter (ID), 5 μm; mobile phase: CO₂ for phase A, MEOH + 0.1% NH₃·HO for phase B; gradient: B 40%; flow rate: 50 mL / min; backpressure: 100 bar; column temperature: 35 °C; wavelength: 220 nm] to give (R)-2-(4-isopropylphenyl)-N-(1-(2-methyl-2H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide (46.5 mg). LC / MS ESI: 337 (M+H) + . 1 H-NMR(400MHz,CDCl3)δ9.11(s,1H);7.89(s,1H);7.38(s,1H);7.18(s,4H);6.64(d,J=6.8Hz,1H);5.24-5. 16(m,1H);4.27(s,3H);3.54-3.52(m,2H);2.93-2.86(m,1H);1.45(d,J=6.8Hz,3H);1.24(d,J=6.9Hz,6H). Example 3 (R)-2-(4-isopropylphenyl)-N-(1-(1-methyl-1H-pyrazolo[4,3-c]pyridin-6-yl)ethyl)acetamide

[0168] Step 1: Synthesis of methyl 1-methyl-1H-pyrazolo[4,3-c]pyridine-6-carboxylate and methyl 2-methyl-2H-pyrazolo[4,3-c]pyridine-6-carboxylate [ka]

[0169] To a solution of methyl 1H-pyrazolo[4,3-c]pyridine-6-carboxylate (2.0 g) in anhydrous DMF (20 mL) was added CsCO (5.5 g) and iodomethane (1.8 g), and the reaction mixture was stirred at room temperature for 18 hours. Water and EtOAc were then added to the reaction mixture. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over NaSO, filtered, and concentrated to give a residue that was purified by silica gel column chromatography to give methyl 1-methyl-1H-pyrazolo[4,3-c]pyridine-6-carboxylate (1.2 g) and methyl 2-methyl-2H-pyrazolo[4,3-c]pyridine-6-carboxylate (0.54 g).

[0170] Methyl 1-methyl-1H-pyrazolo[4,3-c]pyridine-6-carboxylate: LC / MS ESI (m / z) = 192 [M+H] + . 1 H-NMR (400MHz, DMSO-d6) δ9.17(d,J=1.2Hz,1H);8.44(t,J=1.0Hz,1H);8.41(d,J=0.9Hz,1H);4.17(s,3H);3.92(s,3H).

[0171] Methyl 2-methyl-2H-pyrazolo[4,3-c]pyridine-6-carboxylate: LC / MS ESI (m / z) = 192 [M+H] + . 1 H-NMR (400MHz, DMSO-d6) δ9.23(d,J=1.2Hz,1H);8.78(s,1H);8.28(t,J=1.0Hz,1H);4.28(s,3H);3.89(s,3H).

[0172] Step 2: Synthesis of (1-methyl-1H-pyrazolo[4,3-c]pyridin-6-yl)methanol [ka]

[0173] To a solution of methyl 1-methyl-1H-pyrazolo[4,3-c]pyridine-6-carboxylate (350 mg) in THF (5 mL) was added lithium aluminum hydride (208.42 mg) at 0° C., and the mixture was stirred at 25° C. for 2 hours. The mixture was then quenched with aqueous NaOH (2 M) and extracted with DCM. The combined extracts were dried, filtered, and concentrated to give (1-methyl-1H-pyrazolo[4,3-c]pyridin-6-yl)methanol (160 mg) as a yellow oil, which was used directly in the next step. LC / MS ESI (m / z): 164 [M+H] + .

[0174] Step 3: Synthesis of 1-methyl-1H-pyrazolo[4,3-c]pyridine-6-carbaldehyde [ka]

[0175] To a solution of (1-methyl-1H-pyrazolo[4,3-c]pyridin-6-yl)methanol (160 mg) in DCM (5 mL) was added manganese dioxide (426.3 mg). The mixture was stirred at 25° C. for 3 hours. The mixture was then filtered, and the filtrate was concentrated in vacuo to give a residue that was used directly in the next reaction. LC / MS ESI (m / z): 265 [M+H] + .

[0176] Step 4: Synthesis of methyl (R,E)-2-methyl-N-((1-methyl-1H-pyrazolo[4,3-c]pyridin-6-yl)methylene)propane-2-sulfinamide [ka]

[0177] To a solution of (1-methyl-1H-pyrazolo[4,3-c]pyridin-6-yl)methanol (140 mg) in CHCl (10 mL) was added (R)-2-methylpropane-2-sulfinamide (105.3 mg) and copper(II) sulfate (693.3 mg) at room temperature, and the mixture was stirred at room temperature overnight. LC / MS showed that the reaction was complete. The reaction mixture was quenched with ice water and then extracted twice with EtOAc. The combined extracts were concentrated, and the residue was purified by silica gel column chromatography (PE:EA=2:1) ​​to give (R,E)-2-methyl-N-((1-methyl-1H-pyrazolo[4,3-c]pyridin-6-yl)methylene)propane-2-sulfinamide (110 mg) as a yellow solid. LC / MS ESI (m / z): 265 [M+H] + .

[0178] Step 5: Synthesis of (R)-2-methyl-N-((R)-1-(1-methyl-1H-pyrazolo[4,3-c]pyridin-6-yl)ethyl)propane-2-sulfinamide [ka]

[0179] To a solution of (R,E)-2-methyl-N-((1-methyl-1H-pyrazolo[4,3-c]pyridin-6-yl)methylene)propane-2-sulfinamide (110 mg) in THF (5 mL) at −78° C. under N was added CHMgBr (0.41 mL, 3 M in ether), and the mixture was stirred at −78° C. for 1 h. LC / MS showed the reaction was complete. The reaction mixture was then quenched with ice water and extracted twice with EtOAc. The combined extracts were dried over NaSO, filtered, and concentrated. The residue was purified by silica gel column chromatography (DCM / MeOH=10:1) to give (R)-2-methyl-N-((R)-1-(1-methyl-1H-pyrazolo[4,3-c]pyridin-6-yl)ethyl)propane-2-sulfinamide (80 mg) as a pale yellow oil. LC / MS ESI(m / z):281[M+H] + .

[0180] Step 6: Synthesis of (R)-1-(1-methyl-1H-pyrazolo[4,3-c]pyridin-6-yl)ethan-1-amine hydrochloride [ka]

[0181] To a solution of (R)-2-methyl-N-((R)-1-(1-methyl-1H-pyrazolo[4,3-c]pyridin-6-yl)ethyl)propane-2-sulfinamide (85 mg) in dioxane (3.0 mL) was added 4N HCl solution in dioxane (253.9 mg) at room temperature, and the mixture was stirred at room temperature for 2 hours. LC / MS showed the reaction was complete. The solvent was removed to give (R)-1-(1-methyl-1H-pyrazolo[4,3-c]pyridin-6-yl)ethan-1-amine hydrochloride (85 mg). LC / MS ESI (m / z): 177 [M+H] + .

[0182] Step 7: Synthesis of (R)-2-(4-isopropylphenyl)-N-(1-(1-methyl-1H-pyrazolo[4,3-c]pyridin-6-yl)ethyl)acetamide [ka]

[0183] To a solution of (R)-1-(1-methyl-1H-pyrazolo[4,3-c]pyridin-6-yl)ethan-1-amine hydrochloride (91 mg) in CHCl (5 mL) was added 2-(4-isopropylphenyl)acetic acid (92 mg), HATU (392 mg), and triethylamine (156 mg), and the mixture was stirred at room temperature for 2 hours. LC / MS showed the reaction was complete. The reaction mixture was then quenched with ice water and extracted twice with EtOAc. The combined extracts were concentrated, and the residue was purified by silica gel column chromatography (PE:EA = 3:1) followed by preparative HPLC [column: YMC-Actus Triart C18, 250 × 21 mm; mobile phase: 20% to 95% MeCN / HO (0.1% FA); flow rate: 15 mL / min; wavelength: 220 nm / 254 nm] and SFC (mobile phase: CO in phase A and MeOH + 0.1% NH₃·HO in phase B; gradient: B 40%) to give (R)-2-(4-isopropylphenyl)-N-(1-(1-methyl-1H-pyrazolo[4,3-c]pyridin-6-yl)ethyl)acetamide (28 mg) as a white solid. LC / MS ESI (m / z): 337 [M+H]. + . 1 H-NMR(400MHz,CDCl3)δ8.82(s,1H);8.14(s,1H);7.36(s,1H);7.20-7.11(m,4H);6.87(d,J=7.5Hz,1H);5. 28-5.24(m,1H);4.06(s,3H);3.57(s,2H);2.89-2.85(m,1H);1.50(t,J=8.0Hz,3H);1.23(t,J=8.0Hz,6H). Example 4 (R)-2-(4-isopropylphenyl)-N-(1-(2-methyl-2H-pyrazolo[4,3-c]pyridin-6-yl)ethyl)acetamide

[0184] Step 1: (2-methyl-2H-pyrazolo[4,3-c]pyridin-6-yl)methanol [ka]

[0185] To a solution of methyl 2-methyl-2H-pyrazolo[4,3-c]pyridine-6-carboxylate (530 mg) in THF (10 mL) was added lithium aluminum hydride (315.6 mg) at 0° C., and the mixture was stirred at 25° C. for 3 hours. The mixture was then quenched with aqueous NaOH (2M) and extracted with DCM. The combined extracts were dried, filtered, and concentrated to give (2-methyl-2H-pyrazolo[4,3-c]pyridin-6-yl)methanol (280 mg, colorless oil), which was used directly in the next step. LC / MS ESI (m / z): 164 [M+H] + .

[0186] Step 2: 2-methyl-2H-pyrazolo[4,3-c]pyridine-6-carbaldehyde [ka]

[0187] To a solution of (2-methyl-2H-pyrazolo[4,3-c]pyridin-6-yl)methanol (280 mg) in DCM (5 mL) was added manganese dioxide (0.12 mL), and the mixture was stirred at 25° C. for 3 h. The mixture was then filtered, and the filtrate was concentrated in vacuo to give crude 2-methyl-2H-pyrazolo[4,3-c]pyridine-6-carbaldehyde, which was used directly in the next step. LC / MS ESI (m / z): 162 [M+H] + .

[0188] Step 3: (R,E)-2-methyl-N-((2-methyl-2H-pyrazolo[4,3-c]pyridin-6-yl)methylene)propane-2-sulfinamide [ka]

[0189] A solution of 2-methyl-2H-pyrazolo[4,3-c]pyridine-6-carbaldehyde (280 mg), (R)-2-methylpropane-2-sulfinamide (316 mg), and copper sulfate pentahydrate (465 mg) in DCM (5.0 mL) was stirred at 25 °C for 3 hours. The mixture was then filtered and concentrated to give a yellow oil, which was purified by column chromatography on silica gel (DCM:MeOH = 50:1 to 20:1) to give (R,E)-2-methyl-N-((2-methyl-2H-pyrazolo[4,3-c]pyridin-6-yl)methylene)propane-2-sulfinamide (400 mg) as a yellow solid. LC / MS ESI (m / z): 265 [M+H] + .

[0190] Step 4: (R)-2-methyl-N-((R)-1-(2-methyl-2H-pyrazolo[4,3-c]pyridin-6-yl)ethyl)propane-2-sulfinamide [ka]

[0191] To a solution of (R,E)-2-methyl-N-((2-methyl-2H-pyrazolo[4,3-c]pyridin-6-yl)methylene)propane-2-sulfinamide (180 mg) in THF (5 mL) was added CHMgBr (3 M in ether, 2.0 mL) at −70° C., and the mixture was stirred at −70° C. for 2 hours. The reaction mixture was then quenched with saturated aqueous ammonium chloride solution (5 mL) and extracted with EtOAc (10 mL). The organic layer was dried, filtered, and concentrated to give a yellow oil, which was purified by column chromatography on silica gel (DCM:MeOH=50:1→20:1) to give (R)-2-methyl-N-((R)-1-(2-methyl-2H-pyrazolo[4,3-c]pyridin-6-yl)ethyl)propane-2-sulfinamide (40 mg) as a white solid. LC / MS ESI(m / z):281[M+H] + .

[0192] Step 5: (R)-1-(2-methyl-2H-pyrazolo[4,3-c]pyridin-6-yl)ethan-1-amine hydrochloride [ka]

[0193] To a solution of 2-methyl-N-[(1R)-1-{2-methyl-2H-pyrazolo[4,3-c]pyridin-6-yl}ethyl]propane-2-sulfinamide (40 mg) in dioxane (2 mL) was added HCl / dioxane (4N in dioxane, 1 mL), and the mixture was stirred at 25° C. for 30 minutes. The mixture was then concentrated to give (R)-1-(2-methyl-2H-pyrazolo[4,3-c]pyridin-6-yl)ethan-1-amine hydrochloride (30 mg, crude) as a white solid. LC / MS ESI (m / z): 177 [M+H] + .

[0194] Step 6: (R)-2-(4-isopropylphenyl)-N-(1-(2-methyl-2H-pyrazolo[4,3-c]pyridin-6-yl)ethyl)acetamide [ka]

[0195] A solution of (R)-1-(2-methyl-2H-pyrazolo[4,3-c]pyridin-6-yl)ethan-1-amine hydrochloride (30 mg), 2-[4-(propan-2-yl)phenyl]acetic acid (33.4 mg), HATU (97 mg), and TEA (0.07 mL) in DCM (2.0 mL) was stirred for 12 hours at 25° C. The mixture was diluted with water and extracted with EtOAc. The combined extracts were dried, filtered, and concentrated to give a residue, which was purified by preparative HPLC [Column: Shim-pack GIST C18, 250 × 21 mm; Mobile phase: 10% to 85% MeCN / HO (0.1% NH HO); Flow rate: 15 mL / min; Wavelength: 220 nm / 254 nm] to give (R)-2-(4-isopropylphenyl)-N-(1-(2-methyl-2H-pyrazolo[4,3-c]pyridin-6-yl)ethyl)acetamide (14 mg) as a colorless oil. LC / MS ESI (m / z): 337 (M+H). + .

[0196] 1 H-NMR(400MHz,CDCl3)δ9.02(s,1H);7.40(s,1H);7.20(s,4H);6.67(d,J=7.6Hz,1H);5.26-6.19(m, 1H);4.25(s,3H);3.57-3.56(m,2H);2.94-2.87(m,1H);1.46(d,J=6.8Hz,3H);1.25(d,J=8.0Hz,6H). Example 5 (R)-2-(4-isopropylphenyl)-N-(1-(3-methylisoxazolo[4,5-c]pyridin-6-yl)ethyl)acetamide

[0197] Synthesis of 4,6-dichloro-N-methoxy-N-methylnicotinamide [ka]

[0198] To a solution of 4,6-dichloronicotinic acid (5.0 g) in DCM (70 mL) was added HATU (10.9 g, 28.6) at room temperature, and the reaction mixture was stirred for 10 minutes. N-Methoxymethanamine HCl (3.0 g) and TEA (10.8 mL) were then added at room temperature, and the mixture was stirred at room temperature for 2 hours. The mixture was then quenched with water and extracted with EA. The combined extracts were washed with brine, dried over Na2SO4, and concentrated in vacuo to give 4,6-dichloro-N-methoxy-N-methylnicotinamide (6.0 g) as a yellow oil. LC / MS ESI: 234 (M+H) + . 1 H-NMR (400MHz, DMSO-d6) δ 8.58 (s, 1H); 7.96 (s, 1H); 3.49 (s, 3H); 3.32 (s, 3H).

[0199] Synthesis of 1-(4,6-dichloropyridin-3-yl)ethan-1-one [ka]

[0200] 4,6-Dichloro-N-methoxy-N-methylnicotinamide (6.0 g) was dissolved in anhydrous THF (86 mL) and stirred at 0-5°C for 10 minutes. Then, MeMgBr (14 mL, 3 M in ether) was slowly added to the solution. After 2 hours at 0-5°C, the starting material was consumed, and the reaction was quenched with saturated aqueous ammonium chloride and extracted with ethyl acetate. The combined extracts were washed with brine, dried over Na2SO4, and evaporated to give an oil. This oil was purified by silica gel column chromatography to give 1-(4,6-dichloropyridin-3-yl)ethan-1-one (4.3 g). LC / MS ESI: 190 (M+H). + . 1 H-NMR (400MHz, CDCl3) δ8.62(s,1H); 7.46(s,1H); 2.68(s,3H).

[0201] Step 3: Synthesis of (E)-1-(4-chloro-6-styrylpyridin-3-yl)ethan-1-one and (E)-1-(6-chloro-4-styrylpyridin-3-yl)ethan-1-one [ka]

[0202] To a mixture of 1-(4,6-dichloropyridin-3-yl)ethan-1-one (228 mg) and (E)-styrylboronic acid (266 mg) in DMF (6.0 mL) was added Pd(PPh3)4 (100 mg) and 2N Cs2CO3 (1.8 mL). The reaction mixture was filled with N2 three times and stirred at 80 °C for 6 h. 50 mL of EA was added to the reaction mixture. The organic phase was washed with brine, dried over Na2SO4, and filtered. The filtrate was concentrated in vacuo, and the residue was purified by silica gel column chromatography (10% EA in PE) to give (E)-1-(4-chloro-6-styrylpyridin-3-yl)ethan-1-one and (E)-1-(6-chloro-4-styrylpyridin-3-yl)ethan-1-one as a mixture of isomers. LC / MS ESI(m / z):258(M+H) + .

[0203] Synthesis of 1-(4-chloro-6-((E)-styryl)pyridin-3-yl)ethan-1-one oxime (N201201-118) [ka] To a solution of (E)-1-(4-chloro-6-styrylpyridin-3-yl)ethan-1-one and (E)-1-(6-chloro-4-styrylpyridin-3-yl)ethan-1-one in EtOH (25 mL) and HO (5 mL) was added hydroxylamine hydrochloride (0.80 g) and NaCO (1.2 g). The reaction mixture was stirred at 65 °C overnight. The reaction mixture was diluted with EA and water. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over NaSO, filtered, and concentrated to give a residue that was purified by silica gel column chromatography to give 1-(4-chloro-6-((E)-styryl)pyridin-3-yl)ethan-1-one oxime (307 mg). LC / MS ESI: 273 (M+H). + .

[0204] Synthesis of (E)-3-methyl-6-styrylisoxazolo[4,5-c]pyridine (N201201-122) [ka]

[0205] To a solution of 1-(4-chloro-6-((E)-styryl)pyridin-3-yl)ethan-1-one oxime (307 mg) in DMSO (15 mL) was added K2CO3 (466.741 mg) and the reaction was stirred at 100 °C for 2 h. After cooling to room temperature, water and EtOAc were added. The two phases were separated and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over Na2SO4, filtered, and concentrated to give a residue which was purified by silica gel column chromatography to give (E)-3-methyl-6-styrylisoxazolo[4,5-c]pyridine (235 mg). LC / MS ESI: 237 (M+H) + . 1 H-NMR(400MHz,DMSO-d6)δ9.15(d,J=0.7Hz,1H);7.87-7.83(m,2H);7.70(d,J=7.3Hz,2H);7.46-7.42(m,3H);7.36(d,J=7.3Hz,1H);2.64(s,3H).

[0206] Synthesis of 3-methylisoxazolo[4,5-c]pyridine-6-carbaldehyde [ka]

[0207] Ozone was bubbled through a solution of (E)-3-methyl-6-styrylisoxazolo[4,5-c]pyridine (486 mg) in anhydrous DCM (20 mL) at −78° C. for 10 minutes. The ozone gas was then purged with nitrogen for 10 minutes, and PPh3 (644 mg) was added at room temperature. The reaction mixture was stirred for an additional 40 minutes. The solvent was filtered, and the filtrate was concentrated in vacuo to give a residue that was purified by silica gel column chromatography to give 3-methylisoxazolo[4,5-c]pyridine-6-carbaldehyde (150 mg). LC / MS ESI: 163 (M+H). + . 1 H-NMR (400MHz, CDCl3) δ10.27(s,1H);9.15(d,J=0.4Hz,1H);8.14(d,J=0.8Hz,1H);2.74(s,3H).

[0208] Synthesis of (R,E)-2-methyl-N-((3-methylisoxazolo[4,5-c]pyridin-6-yl)methylene)propane-2-sulfinamide [ka]

[0209] To a solution of 3-methylisoxazolo[4,5-c]pyridine-6-carbaldehyde (150 mg) in anhydrous DCM (15 mL) was added (R)-2-methylpropane-2-sulfinamide (145.8 mg) and CuSO (738 mg), and the reaction mixture was stirred at room temperature overnight. The reaction mixture was then filtered, and the filtrate was concentrated in vacuo. The residue was purified by silica gel column chromatography to give (R,E)-2-methyl-N-((3-methylisoxazolo[4,5-c]pyridin-6-yl)methylene)propane-2-sulfinamide (197 mg) as a white solid. LC / MS ESI: 266 (M+H). + . 1 H-NMR (400MHz, CDCl3) δ9.10(d,J=0.9Hz,1H);8.86(s,1H);8.19(d,J=0.9Hz,1H);2.72(s,3H);1.32(s,9H)ppm.

[0210] Synthesis of (R)-2-methyl-N-((R)-1-(3-methylisoxazolo[4,5-c]pyridin-6-yl)ethyl)propane-2-sulfinamide (N201201-131) [ka]

[0211] To a solution of (R,E)-2-methyl-N-((3-methylisoxazolo[4,5-c]pyridin-6-yl)methylene)propane-2-sulfinamide (197 mg) in anhydrous THF (10 mL) was added CHMgBr (1.2 mL, 3.0 M in ether) dropwise at −78 °C under N. The Grignard reagent was added at a rate such that the internal reaction temperature did not exceed −60 °C. After the addition, the reaction mixture was stirred at −78 °C for 2 h, then warmed to room temperature and quenched with saturated aqueous ammonium chloride solution. The organic layer was separated, and the aqueous layer was extracted once with ethyl acetate. The combined organic layers were dried over Na2SO4, filtered, and concentrated to give a residue which was purified by silica gel column chromatography to give (R)-2-methyl-N-((R)-1-(3-methylisoxazolo[4,5-c]pyridin-6-yl)ethyl)propane-2-sulfinamide (187 mg). LC / MS ESI: 282 (M+H). + . 1 H-NMR (400MHz, CDCl3) δ8.93(d,J=1.0Hz,1H);7.47(d,J=1.2Hz,1H);4.72-4.63(m,2H);2.64(s,3H);1.57(d,J=6.5Hz,3H);1.26(s,9H).

[0212] Synthesis of (R)-1-(3-methylisoxazolo[4,5-c]pyridin-6-yl)ethan-1-amine hydrochloride [ka]

[0213] To a solution of (R)-2-methyl-N-((R)-1-(3-methylisoxazolo[4,5-c]pyridin-6-yl)ethyl)propane-2-sulfinamide (172 mg) in dioxane (6.0 mL) was added 4N HCl-dioxane (2.0 mL). The reaction mixture was stirred at room temperature for 20 minutes, and the mixture was concentrated to give (R)-1-(3-methylisoxazolo[4,5-c]pyridin-6-yl)ethan-1-amine hydrochloride as a residue. The residue was used directly in the next step. LC / MS ESI: 178 (M+H). + .

[0214] Synthesis of (R)-2-(4-isopropylphenyl)-N-(1-(3-methylisoxazolo[4,5-c]pyridin-6-yl)ethyl)acetamide [ka]

[0215] (R)-1-(3-methylisoxazolo[4,5-c]pyridin-6-yl)ethan-1-amine hydrochloride (108 mg) was dissolved in anhydrous DCM (10 mL), followed by the addition of 2-(4-isopropylphenyl)acetic acid (119.5 mg), HATU (347.6 mg), and DIPEA (0.2 mL). The reaction was stirred at room temperature for 1 hour, and the mixture was diluted with water. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL x 3). The combined organic layers were dried over NaSO, filtered, and concentrated to give a residue that was purified by preparative HPLC [Column: Shim-pack GIST C18, 250 × 21 mm; Mobile phase: 10%-95% MeCN / HO (0.1% NH HO); Flow rate: 15 mL / min; Wavelength: 220 nm / 254 nm] to give (R)-2-(4-isopropylphenyl)-N-(1-(3-methylisoxazolo[4,5-c]pyridin-6-yl)ethyl)acetamide (93.7 mg) as a white solid. LC / MS ESI: 338 (M+H). + . 1 H-NMR(400MHz,CDCl3)δ8.83(s,1H);7.34(s,1H);7.23-7.18(m,4H);6.60(d,J=7.3Hz,1H);5.29-5. 22(m,1H);3.57(s,2H);2.95-2.90(m,1H);2.63(s,3H);1.46(d,J=6.9Hz,3H);1.26(d,J=6.9Hz,6H). Example 6 (R)-2-(4-isopropylphenyl)-N-(1-(3-methylisoxazolo[5,4-c]pyridin-5-yl)ethyl)acetamide

[0216] Synthesis of 2,5-dichloro-N-methoxy-N-methylisonicotinamide. [ka]

[0217] To a solution of 2,5-dichloroisonicotinic acid (5.0 g) in DCM (70 mL) was added HATU (10.9 g) at room temperature, and the reaction mixture was stirred at room temperature for 10 minutes. Then, N-methoxymethanamine hydrochloride (3.0 g) and TEA (10.8 mL) were added to the reaction mixture, and the mixture was stirred at room temperature for 2 hours. The reaction was quenched with water and extracted with EA. The organic layer was separated, washed with brine, dried over Na2SO4, and concentrated in vacuo. The residue was purified by silica gel column chromatography to give 2,5-dichloro-N-methoxy-N-methylisonicotinamide (6.0 g) as a white solid.

[0218] Synthesis of 1-(2,5-dichloropyridin-4-yl)ethan-1-one (N201201-101). [ka]

[0219] 2,5-Dichloro-N-methoxy-N-methylisonicotinamide (5.0 g) was added to anhydrous tetrahydrofuran (71 mL) under N2 and stirred at 0-5 °C for 10 min. Next, CH3MgBr (11 mL, 3.0 M in ether) was slowly added to the solution, and the reaction was stirred at 0-5 °C for 2 h. The reaction mixture was quenched with saturated aqueous ammonium chloride solution and extracted with ethyl acetate. The combined extracts were washed with brine, dried over Na2SO4, filtered, and evaporated to give an oil, which was purified by silica gel column chromatography to give 1-(2,5-dichloropyridin-4-yl)ethan-1-one (3.4 g). LC / MS ESI (m / z): 189.9 [M+H] + .

[0220] Synthesis of (E)-1-(5-chloro-2-styrylpyridin-4-yl)ethan-1-one [ka]

[0221] To a solution of 1-(2,5-dichloropyridin-4-yl)ethan-1-one (1.1 g), (E)-styrylboronic acid (1.3 g) in DMF, 2.0 N CsCO (6.0 mL) and Pd(PPh) (420 mg) were added. The reaction mixture was stirred at 80 °C for 5 h. EA (50 mL) and HO (40 mL) were added. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over NaSO, filtered, and concentrated to give a residue, which was purified by silica gel column chromatography (eluted with 10% EA in PE) to give (E)-1-(5-chloro-2-styrylpyridin-4-yl)ethan-1-one as a yellow solid (900 mg).

[0222] Synthesis of 1-(5-chloro-2-((E)-styryl)pyridin-4-yl)ethan-1-one oxime [ka]

[0223] To a solution of 1-(5-chloro-2-styrylpyridin-4-yl)ethan-1-one (1.48 g) in EtOH (25 mL) and HO (5 mL) was added NHOH·HCl (0.8 g) and NaCO (1.21 g). The reaction was stirred at 65 °C overnight. The reaction was then diluted with EA and water. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over NaSO, filtered, and concentrated to give a residue that was purified by silica gel column chromatography to give 1-(5-chloro-2-((E)-styryl)pyridin-4-yl)ethan-1-one oxime (294 mg). LC / MS ESI (m / z): 273 [M+H] + . 1 H-NMR(400MHz,DMSO-d6)δ11.02(s,1H);8.71(s,1H);7.76-7.70(m,3H);7.57(s,1H);7.49(t,J=7.4Hz,2H);7.40(t,J=11.7Hz,2H);2.18(s,3H)ppm.

[0224] Synthesis of (E)-3-methyl-5-styrylisoxazolo[5,4-c]pyridine [ka]

[0225] To a solution of 1-(5-chloro-2-((E)-styryl)pyridin-4-yl)ethan-1-one oxime (294 mg) in DMSO (10 mL) was added K2CO3 (448.2 mg). The reaction was stirred at 100 °C for 2 h. After cooling to room temperature, water and EtOAc were added to the reaction mixture. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over Na2SO4, filtered, and concentrated to give a residue that was purified by silica gel column chromatography to give (E)-3-methyl-5-styrylisoxazolo[5,4-c]pyridine (231 mg) as a yellow solid. LC / MS ESI (m / z): 237 [M+H] + . 1 H-NMR (400MHz, DMSO-d6) δ9.17(s,1H);8.06(s,1H);7.69(t,J=11.2Hz,3H);7.50-7.39(m,3H);7.33(t,J=7.3Hz,1H);2.63(s,3H).

[0226] Synthesis of 3-methylisoxazolo[5,4-c]pyridine-5-carbaldehyde (N201201-109 / 115) [ka]

[0227] Ozone was bubbled through a solution of (£)-3-methyl-5-styrylisoxazolo[5,4-c]pyridine (483 mg) in anhydrous DCM (10 mL) at -78 °C for 10 min. The excess ozone gas was then purged with nitrogen, followed by the addition of triphenylphosphine (644 mg). The mixture was stirred for an additional 40 min and then concentrated in vacuo to give a residue that was purified by silica gel column chromatography to give 3-methylisoxazolo[5,4-c]pyridine-5-carbaldehyde (200 mg). LC / MS ESI (m / z): 163 [M+H] + .

[0228] 1 H-NMR (400MHz, CDCl3) δ10.23(s,1H);9.16(s,1H);8.35(d,J=0.9Hz,1H);2.70(s,3H).

[0229] Synthesis of (R,E)-2-methyl-N-((3-methylisoxazolo[5,4-c]pyridin-5-yl)methylene)propane-2-sulfinamide [ka]

[0230] To a solution of 3-methylisoxazolo[5,4-c]pyridine-5-carbaldehyde (0.23 g) in anhydrous dichloromethane (10 mL) was added (R)-2-methylpropane-2-sulfinamide (0.26 g) and CuSO (1.15 g), and the reaction was stirred at room temperature overnight. The reaction solution was then filtered and concentrated in vacuo. The residue was purified by silica gel column chromatography to give (R,E)-2-methyl-N-((3-methylisoxazolo[5,4-c]pyridin-5-yl)methylene)propane-2-sulfinamide (309 mg) as a white solid. LC / MS ESI (m / z): 266 [M+H] + . 1 H-NMR (400MHz, DMSO-d6) δ9.33(d,J=1.1Hz,1H);8.69-8.62(m,2H);2.69(s,3H);1.24(s,9H)ppm.

[0231] Synthesis of (R)-2-methyl-N-((R)-1-(3-methylisoxazolo[5,4-c]pyridin-5-yl)ethyl)propane-2-sulfinamide [ka]

[0232] To a solution of (R,E)-2-methyl-N-((3-methylisoxazolo[5,4-c]pyridin-5-yl)methylene)propane-2-sulfinamide (337 mg) in anhydrous tetrahydrofuran (5.0 mL) was added CHMgBr (1.7 mL, 3.0 M in EtO) dropwise at −78 °C. The Grignard reagent was added at a rate such that the internal reaction temperature did not exceed −60 °C. After the addition, the mixture was stirred at −78 °C for 3 h and then allowed to warm to room temperature. The reaction was quenched with saturated aqueous ammonium chloride solution. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over Na2SO4, filtered, and concentrated to give a residue that was purified by silica gel column chromatography to give (R)-2-methyl-N-((R)-1-(3-methylisoxazolo[5,4-c]pyridin-5-yl)ethyl)propane-2-sulfinamide (226 mg). LC / MS ESI(m / z): 282 [M+H] + .

[0233] Synthesis of (R)-1-(3-methylisoxazolo[5,4-c]pyridin-5-yl)ethan-1-amine hydrochloride [ka]

[0234] To a solution of (R)-2-methyl-N-((R)-1-(3-methylisoxazolo[5,4-c]pyridin-5-yl)ethyl)propane-2-sulfinamide (219 mg) in dioxane (3 mL) was added 4N HCl-dioxane (1 mL), and the reaction was stirred at room temperature for 30 minutes. The mixture was then concentrated in vacuo to give crude (R)-1-(3-methylisoxazolo[5,4-c]pyridin-5-yl)ethan-1-amine hydrochloride, which was used directly in the next step. LC / MS ESI (m / z): 178 [M+H] + .

[0235] Synthesis of (R)-2-(4-isopropylphenyl)-N-(1-(3-methylisoxazolo[5,4-c]pyridin-5-yl)ethyl)acetamide [ka]

[0236] (R)-1-(3-methylisoxazolo[5,4-c]pyridin-5-yl)ethan-1-amine hydrochloride (137 mg) was dissolved in anhydrous DCM (15 mL), and then 2-[4-(propan-2-yl)phenyl]acetic acid (151.6 mg), HATU (587.9 mg), and TEA (0.32 mL) were added to the solution. The reaction was stirred at room temperature for 1 hour and then diluted with water. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over NaSO, filtered, and concentrated to give a residue that was purified by preparative HPLC [Column: YMC-Actus Triart C18, 250 × 21 mm; Mobile phase: 20%-95% MeCN / HO (0.1% NH HO); Flow rate: 15 mL / min; Wavelength: 220 nm / 254 nm] to give (R)-2-(4-isopropylphenyl)-N-(1-(3-methylisoxazolo[5,4-c]pyridin-5-yl)ethyl)acetamide (146.8 mg) as a colorless oil. LC / MS ESI (m / z): 338 [M+H] + . 1H-NMR(400MHz,CDCl3)δ8.88(s,1H);7.48(s,1H);7.20-7.16(m,4H);6.55(d,J=7.6Hz,1H);5.32-5. 25(m,1H);3.55(s,2H);2.92-2.89(m,1H);2.60(s,3H);1.47(d,J=6.8Hz,3H);1.25(d,J=6.9Hz,6H). Example 7 (R)-2-(4-isopropylphenyl)-N-(1-(1-propyl-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide

[0237] Synthesis of methyl 1-propyl-1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde [ka]

[0238] A solution of 1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde (480 mg), 1-iodopropane (0.35 mL), and CsCO (1.6 g) in DMF (1.0 mL) was stirred at 25 °C for 12 h. The mixture was diluted with water and extracted with EtOAc. The combined extracts were dried, filtered, and concentrated to give a residue that was purified by column chromatography on silica gel (DCM:MeOH = 1:0 to 20:1) to give 1-propyl-1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde (185 mg) as a white solid. 1 H-NMR (400 MHz, CDCl) δ 10.22 (s, 1H); 9.06 (s, 1H); 8.39 (d, J = 1.2 Hz, 1H); 8.24 (s, 1H); 4.51 (t, J = 7.2 Hz, 2H); 2.06-2.04 (m, 2H); 0.96 (t, J = 7.2 Hz, 3H). 2-Propyl-2H-pyrazolo[3,4-c]pyridine-5-carbaldehyde (125 mg) was obtained as a white solid. 1 H-NMR(400MHz,CDCl3):δ10.18(s,1H);9.32(s,1H);8.32(d,J=1.2Hz,1H);8 .20(s,1H);4.49(t,J=7.2Hz,2H);2.12-2.08(m,2H);0.98(t,J=7.2Hz,3H).

[0239] Synthesis of (R,E)-2-methyl-N-((1-propyl-1H-pyrazolo[3,4-c]pyridin-5-yl)methylene)propane-2-sulfinamide (N201201-161) [ka]

[0240] To a solution of 1-propyl-1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde (185 mg) in anhydrous DCM (11 mL) was added (R)-2-methylpropane-2-sulfinamide (154.0 mg) and CuSO4 (234.1 mg). The reaction mixture was stirred at room temperature overnight. The solid was filtered through Celite. The filtrate was then concentrated, and the resulting residue was purified by silica gel column chromatography to give (R,E)-2-methyl-N-((1-propyl-1H-pyrazolo[3,4-c]pyridin-5-yl)methylene)propane-2-sulfinamide (233 mg). LC / MS ESI (m / z): 293 [M+H] + . 1 H-NMR(400MHz,DMSO-d6)δ9.36(s,1H);8.64(s,1H);8.54(s,1H);8.40(s,1H) ;4.57(t,J=6.9Hz,2H);1.94-1.89(m,2H);1.22(s,9H);0.84(t,J=7.4Hz,3H).

[0241] Synthesis of (R)-2-methyl-N-((R)-1-(1-propyl-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)propane-2-sulfinamide (N201201-166) [ka]

[0242] To a solution of (R,E)-2-methyl-N-((1-propyl-1H-pyrazolo[3,4-c]pyridin-5-yl)methylene)propane-2-sulfinamide (233 mg) in anhydrous THF (10 mL) was added 3 M CHMgBr (1.328 mL, 3 M in ether) dropwise under N, and the reaction mixture was stirred at −60° C. for 2 h. The reaction mixture was washed with aqueous NHCl and extracted with EA. The combined extracts were washed with saturated NaCl, dried over NaSO, filtered, and concentrated in vacuo. The residue was then purified by silica gel column chromatography to give (R)-2-methyl-N-((R)-1-(1-propyl-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)propane-2-sulfinamide (146 mg). LC / MS ESI (m / z): 309 [M+H] + . 1 H-NMR(400MHz,DMSO-d6)δ9.14(s,1H);8.19(d,J=0.6Hz,1H);7.80(d,J=0.6Hz,1H);5.63(d,J=7.5Hz,1 H);4.62-4.44(m,2H);1.93-1.84(m,2H);1.48(d,J=6.8Hz,3H);1.14(s,9H);0.83(t,J=7.4Hz,3H)ppm.

[0243] Synthesis of (R)-1-(1-propyl-1H-pyrazolo[3,4-c]pyridin-5-yl)ethanamine hydrochloride [ka]

[0244] To a solution of (R)-2-methyl-N-((R)-1-(1-propyl-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)propane-2-sulfinamide (95 mg) in dioxane (3 mL) was added 4N HCl-dioxane (1.0 mL). The reaction was stirred at room temperature for 30 minutes. The mixture was then concentrated to give (R)-1-(1-propyl-1H-pyrazolo[3,4-c]pyridin-5-yl)ethanamine hydrochloride, which was used directly in the next step. LC / MS ESI (m / z): 205 [M+H] + .

[0245] Synthesis of (R)-2-(4-isopropylphenyl)-N-(1-(1-propyl-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide [ka]

[0246] To a solution of (R)-1-(1-propyl-1H-pyrazolo[3,4-c]pyridin-5-yl)ethanamine hydrochloride (82 mg) in anhydrous DMF, DIEA (0.12 mL) was added (Solution A). To a solution of 2-(4-isopropylphenyl)acetic acid (78.7 mg) in anhydrous DMF, HATU (167.9 mg) was added and stirred at room temperature for 3 minutes (Solution B). Solution A was added to Solution B and stirred at room temperature for 1 hour. The mixture was diluted with water, and the aqueous phase was extracted with DCM (10 mL × 3). The combined extracts were dried over Na2SO4, filtered, and concentrated to give a residue that was purified by preparative HPLC [Column: YMC Triart C18, 250 x 20 mm ID, 5 μm; Mobile phase: 10% to 95% MeCN / HO (0.1% FA); Flow rate: 14 mL / min; Wavelength: 220 nm / 254 nm] to give (R)-2-(4-isopropylphenyl)-N-(1-(1-propyl-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide. LC / MS ESI (m / z): 365 [M+H] + . 1 H-NMR(400MHz,CDCl3)δ8.82(s,1H);7.98(s,1H);7.49(s,1H);7.19(s,4H);6.62(d,J=7.8Hz,1H);5.28-5.21(m,1H);4.42(t,J=7.0 Hz,2H);3.56-3.55(m,2H);2.92-2.88(m,1H);2.00-1.96(m,2H);1.46(d,J=6.8Hz,3H);1.25(d,J=6.9Hz,6H);0.93(t,J=7.4Hz,3H). Example 8 (R)—N-(1-(1-ethyl-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)-2-(4-isopropylphenyl)acetamide

[0247] Synthesis of 5-bromo-1H-pyrazolo[3,4-c]pyridine [ka]

[0248] To a solution of 6-bromo-4-methylpyridin-3-amine (1.0 g) in acetic acid (60 mL) was added sodium nitrite (0.37 mL) in an ice bath. The reaction was stirred at room temperature overnight. The reaction was then concentrated and treated with saturated aqueous NaHCO3. The aqueous mixture was extracted with EA. The combined extracts were washed with brine, dried over Na2SO4, filtered, and concentrated in vacuo. The residue was purified by silica gel column chromatography to give 5-bromo-1H-pyrazolo[3,4-c]pyridine (442 mg). LC / MS ESI (m / z): 198 [M+H] + .

[0249] Synthesis of 1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde [ka]

[0250] To a solution of 5-bromo-1H-pyrazolo[3,4-c]pyridine (3.3 g) in anhydrous THF (100 mL) was added n-BuLi (20.0 mL, 2N in THF) at −78° C. The mixture was stirred at −78° C. for 1 hour. Then, DMF (3.87 mL) was added to the mixture at −78° C., and the mixture was stirred at −78° C. for 1 hour. The reaction was warmed to room temperature and quenched with NH4Cl solution. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over Na2SO4, filtered, and concentrated to give a residue, which was purified by column chromatography on silica gel (DCM:MeOH = 1:0 to 20:1) to give HCl. 1 H-Pyrazolo[3,4-c]pyridine-5-carbaldehyde (1.0 g) was obtained as a white solid. LC / MS ESI (m / z): 148 [M+H] + .

[0251] Synthesis of 1-ethyl-1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde [ka]

[0252] A solution of 1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde (480 mg), ethyl iodide (0.35 mL), and CsCO (1.6 g) in DMF (1.0 mL) was stirred at 25 °C for 12 h. The mixture was then diluted with water and extracted with EtOAc. The combined extracts were dried, filtered, and concentrated to give a residue that was purified by column chromatography on silica gel (DCM:MeOH = 1:0 to 20:1) to give 1-ethyl-1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde (185 mg) as a white solid. 1 H-NMR (400 MHz, DMSO-d6) δ 10.11 (s, 1H); 9.39 (s, 1H); 8.53-8.39 (m, 2H); 4.65 (q, J = 7.2 Hz, 2H); 1.47 (t, J = 7.2 Hz, 3H). 2-Propyl-2H-pyrazolo[3,4-c]pyridine-5-carbaldehyde (125 mg) was obtained as a white solid. 1 H-NMR (400MHz, DMSO):10.05(s,1H);9.36-9.23(m,1H);8.85(s,1H);8.43(d,J=1.2Hz,1H);4.60(q,J=7.2Hz,2H);1.55(t,J=7.2Hz,3H).

[0253] Synthesis of (R,E)-N-((1-ethyl-1H-pyrazolo[3,4-c]pyridin-5-yl)methylene)-2-methylpropane-2-sulfinamide [ka]

[0254] To a solution of 1-ethyl-1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde (186 mg) in DCM (10 mL) was added 2-methylpropane-2-sulfinamide (157.6 mg) and CuSO (239.4 mg). The reaction mixture was stirred at room temperature overnight. The mixture was then filtered, and the filtrate was concentrated in vacuo to give a residue that was purified by silica gel to give (R,E)-N-((1-ethyl-1H-pyrazolo[3,4-c]pyridin)-5-yl)methylene)-2-methylpropane-2-sulfinamide (207 mg) as a white solid. LC / MS ESI (m / z): 279 [M+H] + . 1 H-NMR(400MHz,DMSO-d6)δ9.35(s,1H);8.64(s,1H);8.54(d,J=1.2Hz,1H);8. 39(d,J=0.6Hz,1H);4.66-4.61(m,2H);1.47(t,J=7.2Hz,3H);1.22(s,9H)ppm.

[0255] Synthesis of (R)-N-((R)-1-(1-ethyl-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)-2-methylpropane-2-sulfinamide [ka]

[0256] To a solution of (R,E)-N-((1-ethyl-1H-pyrazolo[3,4-c]pyridin-5-yl)methylene)-2-methylpropane-2-sulfinamide (207 mg) in THF (15 mL) was added CHMgBr (1.24 mL, 3.0 M in ether) dropwise. The reaction mixture was stirred at -60 °C for 2 h. The reaction mixture was then quenched with saturated NHCl solution. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over Na2SO4, filtered, and concentrated to give a residue which was purified by silica gel column chromatography to give (R)-N-((R)-1-(1-ethyl-1H)-pyrazolo[3,4-c]pyridin-5-yl)ethyl)-2-methylpropane-2-sulfinamide (103 mg) as a yellow oil: LC / MS ESI(m / z): 295 [M+H] + .

[0257] Synthesis of (R)-1-(1-ethyl-1H-pyrazolo[3,4-c]pyridin-5-yl)ethan-1-amine hydrochloride [ka]

[0258] To a solution of (R)-N-((R)-1-(1-ethyl-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)-2-methylpropane-2-sulfinamide (142 mg) in dioxane (3 mL) was added 4N HCl-dioxane (1 mL), and the reaction mixture was stirred at room temperature for 30 minutes. The mixture was then concentrated to give (R)-1-(1-ethyl-1H-pyrazolo[3,4-c]pyridin-5-yl)ethan-1-amine hydrochloride, which was used directly in the next step. LC / MS ESI (m / z): 191 [M+H] + .

[0259] Synthesis of (R)-N-(1-(1-ethyl-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)-2-(4-isopropylphenyl)acetamide [ka]

[0260] To a solution of (R)-1-(1-ethyl-1H-pyrazolo[3,4-c]pyridin-5-yl)ethanamine hydrochloride (134 mg) in anhydrous DMF, DIEA (0.5 mL) was added to give solution A. To a solution of 2-(4-isopropylphenyl)acetic acid (95.9 mg) in anhydrous DMF, HATU (225 mg) was added and stirred at room temperature for 3 minutes to give solution B. Solution A was added to solution B and stirred at room temperature for 1 hour. The reaction mixture was diluted with EA and water. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over Na2SO4, filtered, and concentrated to give a residue that was purified by preparative HPLC [column: Xbudge prep C18, 250 × 19 mm, 5 μm OBD; mobile phase: 10%–95% MeCN / HO (0.1% FA); flow rate: 20 mL / min; wavelength: 205 nm / 254 nm] and SFC (column: ChiralPak® IA, 250 × 21.3 mm ID, 5 μm, mobile phase: CO2 in phase A and MEOH + 0.1% NH3·HO in phase B, gradient: B 40%, wavelength: 220 nm) to give (R)—N-(1-(1-ethyl-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)-2-(4-isopropylphenyl)acetamide (44.8 mg) as a white solid. LC / MS ESI(m / z):351[M+H] + . 1 H-NMR(400MHz,CDCl3)δ8.83(s,1H);7.97(s,1H);7.49(s,1H);7.19(s,4H);6.62(d,J=7.2Hz,1H);5.28-5.21(m,1H);4. 52(q,J=7.3Hz,2H);3.55(s,2H);2.92-2.87(m,1H);1.56(t,J=7.3Hz,3H);1.46(d,J=6.8Hz,3H);1.25(d,J=6.9Hz,6H). Example 9 (R)-N-(1-(1-ethyl-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)-2-(4-isopropylphenyl)acetamide

[0261] Step 1: Synthesis of 1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde [ka]

[0262] A solution of 1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde (760 mg), 1,1,1-trifluoro-2-iodoethane (0.51 mL), and CsCO (2.5 g) in DMF (10 mL) was stirred at 100 °C for 12 h. The mixture was diluted with water and extracted with EtOAc. The combined extracts were dried, filtered, and concentrated to give a residue. The residue was purified by silica gel column chromatography (DCM:MeOH = 1:0 to 20:1) to give 1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde (500 mg) as a yellow solid. LC / MS ESI (m / z): 230 [M+H] + .

[0263] Step 2: Synthesis of (R,E)-2-methyl-N-((1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)methylene)propane-2-sulfinamide [ka]

[0264] A solution of 1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde (500 mg), (R)-2-methylpropane-2-sulfinamide (396.7 mg), and copper sulfate pentahydrate (1.0 g) in DCM (15 mL) was stirred at 25 °C for 12 hours. The mixture was filtered and concentrated to give a residue. The residue was purified by silica gel column chromatography (DCM:MeOH = 1:0 to 20:1) to give (R,E)-2-methyl-N-((1-(2,2,2-trifluoroethyl))-1H-pyrazolo[3,4-c]pyridin-5-yl)methylene)propane-2-sulfinamide (450 mg) as a yellow solid. LC / MS ESI (m / z): 333 [M+H] + .

[0265] Step 3: Synthesis of (R)-2-methyl-N-((R)-1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)propane-2-sulfinamide [ka]

[0266] To a solution of (R,E)-2-methyl-N-((1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)methylene)propane-2-sulfinamide (450 mg) in THF (5 mL) was added CHMgBr (3.0 M in ether, 1.35 mL) at −78° C. The mixture was stirred at −78° C. for 2 hours. The mixture was quenched with NHCl solution and extracted with EtOAc. The combined extracts were dried, filtered, and concentrated to give a residue. The residue was purified by column chromatography on silica gel (DCM:MeOH=1:0 to 20:1) to give (R)-2-methyl-N-((R)-1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)propane-2-sulfinamide (350 mg) as a yellow solid. LC / MS ESI(m / z):349[M+H] + .

[0267] Step 4: Synthesis of (R)-1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethan-1-amine hydrochloride [ka]

[0268] To a solution of (R)-2-methyl-N-((R)-1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)propane-2-sulfinamide (300 mg) in dioxane (2.0 mL) was added HCl / dioxane (4N in dioxane, 2.0 mL). The mixture was stirred at 25° C. for 30 minutes. The mixture was concentrated to give (R)-1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethan-1-amine hydrochloride (300 mg, crude) as a white solid. LC / MS ESI (m / z): 245 [M+H] + .

[0269] Step 5: Synthesis of (R)-2-(4-isopropylphenyl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide [ka]

[0270] A solution of (R)-1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethan-1-amine hydrochloride (300 mg), 2-[4-(propan-2-yl)phenyl]acetic acid (240.8 mg), HATU (560.49 mg) and DIEA (0.41 mL) in DCM (10 mL) was stirred at 25° C. for 12 hours. The mixture was washed with water. The organic layer was dried, filtered and concentrated to give a residue. The residue was purified by preparative HPLC [column: YMC-Actus Triart C18, 250 × 21 mm; mobile phase: 50% to 95% MeCN / HO (0.1% FA); flow rate: 20 mL / min; wavelength: 220 nm / 254 nm] and SFC (column: ChiralPak® IA, 250 × 21.3 mm ID, 5 μm; mobile phase: CO in phase A and MEOH + 0.1% NH₃·HO in phase B; gradient: B 40%, wavelength: 220 nm) to give (R)-2-(4-isopropylphenyl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide (128.6 mg) as a white solid. LC / MS ESI (m / z): 405 (M+H). + . 1 H-NMR(400MHz,CDCl3)δ8.85(s,1H);8.08(s,1H);7.54(s,1H);7.24-7.12(m,4H);6.58(d,J=7.6Hz,1H);5.27-5.2 4(m,1H);5.03(q,J=8.4Hz,2H);3.59-3.51(m,2H);2.95-2.85(m,1H);1.46(d,J=6.8Hz,3H);1.25(d,J=6.6Hz,3H). 19 F-NMR (377MHz, CDCl3)δ -70.8ppm. Example 10 (R)—N-(1-(1-methyl-1H-pyrazolo[3,4-b]pyridin-5-yl)ethyl)-2-(4-(1-(trifluoromethyl)cyclopropyl)phenyl)acetamide

[0271] Step 1: Synthesis of 1-methyl-1H-pyrazolo[3,4-b]pyridine-5-carbaldehyde [ka]

[0272] To a solution of 1H-pyrazolo[3,4-b]pyridine-5-carbaldehyde (594 mg) in anhydrous DMF (10 mL) was added Cs2CO3 (1.97 g) and iodomethane (630.3 mg), and the reaction mixture was stirred at room temperature overnight. The reaction mixture was diluted with EA and water. The organic layer was separated, washed with saturated NaCl solution, and concentrated in vacuo. The residue was purified by silica gel column chromatography to give 1-methyl-1H-pyrazolo[3,4-b]pyridine-5-carbaldehyde (285 mg). 1-Methyl-1H-pyrazolo[3,4-b]pyridine-5-carbaldehyde: LC / MS ESI (m / z): 162 [M+H] + . 1 H-NMR (400MHz, DMSO-d6) δ10.15(d,J=1.3Hz,1H);9.06(s,1H);8.81(s,1H);8.41(d,J=2.1Hz,1H);4.12(d,J=1.2Hz,3H).

[0273] 2-Methyl-2H-pyrazolo[3,4-b]pyridine-5-carbaldehyde: LC / MS ESI(m / z): 162[M+H] + . 1 H-NMR (400MHz, DMSO) δ10.07(s,1H);9.01(d,J=2.1Hz,1H);8.86(d,J=2.1Hz,1H);8.75(s,1H);4.25(s,3H).

[0274] Step 2: Synthesis of (R,E)-2-methyl-N-((1-methyl-1H-pyrazolo[3,4-b]pyridin-5-yl)methylene)propane-2-sulfinamide [ka]

[0275] To a solution of 1-methyl-1H-pyrazolo[3,4-b]pyridine-5-carbaldehyde (285 mg) in anhydrous DCM (15 mL) was added (R)-2-methylpropane-2-sulfinamide (278.6 mg) and CuSO (423.4 mg), and the reaction was stirred at room temperature overnight. The solution was filtered through a pad of Celite, and the filtrate was concentrated. The residue was purified by silica gel column to give (R,E)-2-methyl-N-((1-methyl-1H-pyrazolo[3,4-b]pyridin-5-yl)methylene)propane-2-sulfinamide (158 mg). LC / MS ESI (m / z): 265 [M+H] + . 1 H-NMR (400MHz, CDCl3) δ9.06(d,J=1.9Hz,1H);8.74(s,1H);8.51(d,J=1.9Hz,1H);8.13(s,1H);4.20(s,3H);1.29(s,9H)ppm.

[0276] Step 3: Synthesis of (R)-2-methyl-N-((R)-1-(1-methyl-1H-pyrazolo[3,4-b]pyridin-5-yl)ethyl)propane-2-sulfinamide [ka]

[0277] To a solution of (R,E)-2-methyl-N-((1-methyl-1H-pyrazolo[3,4-b]pyridin-5-yl)methylene)propane-2-sulfinamide (407 mg) in anhydrous THF (30 mL) was added CHMgBr (9.0 mL, 3.0 M in ether) dropwise, and the reaction mixture was stirred at −78° C. for 6 h. The reaction mixture was quenched with saturated aqueous NHCl and extracted with EtOAc. The combined extracts were separated, washed with saturated NaCl solution, dried over NaSO, filtered, and concentrated in vacuo. The residue was purified by silica gel column chromatography to give (R)-2-methyl-N-((R)-1-(1-methyl-1H-pyrazolo[3,4-b]pyridin-5-yl)ethyl)propane-2-sulfinamide (236 mg). LC / MS ESI(m / z): 281 [M+H] + .1 H-NMR(400MHz,CDCl3)δ8.56(d,J=2.0Hz,1H);8.04-7.95(m,2H);4.79-4.73( m,1H);4.16(s,3H);3.43(d,J=2.7Hz,1H);1.63(d,J=6.7Hz,3H);1.21(s,9H).

[0278] Step 4: Synthesis of (R)-1-(1-methyl-1H-pyrazolo[3,4-b]pyridin-5-yl)ethan-1-amine hydrochloride [ka]

[0279] To a solution of (R)-2-methyl-N-((R)-1-(1-methyl-1H-pyrazolo[3,4-b]pyridin-5-yl)ethyl)propane-2-sulfinamide (77 mg) in dioxane (3 mL) was added 4N HCl-dioxane (1.0 mL), and the reaction was stirred at room temperature for 30 minutes. The reaction mixture was then concentrated to give (R)-1-(1-methyl-1H-pyrazolo[3,4-b]pyridin-5-yl)ethan-1-amine hydrochloride, which was used directly in the next step. LC / MS ESI (m / z): 177 [M+H] + .

[0280] Step 5: Synthesis of (R)—N-(1-(1-methyl-1H-pyrazolo[3,4-b]pyridin-5-yl)ethyl)-2-(4-(1-(trifluoromethyl)cyclopropyl)phenyl)acetamide [ka]

[0281] To a solution of (R)-1-(1-methyl-1H-pyrazolo[3,4-b]pyridin-5-yl)ethan-1-amine hydrochloride (107 mg) in anhydrous DMF (4.0 mL) was added DIEA (0.5 mL) to give solution A. To a solution of 2-(4-(1-(trifluoromethyl)cyclopropyl)phenyl)acetic acid (134.8 mg) in anhydrous DMF (4.0 mL) was added HATU (230.9 mg) and stirred at room temperature for 10 minutes to give solution B. Solution B was then added to solution A and stirred at room temperature for 1 hour. The reaction was then diluted with EA and water. The organic layer was separated, washed with saturated NaCl solution, dried over Na2SO4, and concentrated in vacuo. The residue was then purified by preparative HPLC (column: YMC-Triart C18, 250 x 20 mm ID, 5 μm; mobile phase: 20% to 95% MeCN / HO (0.1% FA); flow rate: 15 mL / min; wavelength: 220 nm / 254 nm) to give (R)-N-(1-(1-methyl-1H-pyrazolo[3,4-b]pyridin-5-yl)ethyl)-2-(4-(1-(trifluoromethyl)cyclopropyl)phenyl)acetamide (67.3 mg). LC / MS ESI (m / z): 403 [M+H] + . 1 H-NMR(400MHz,CDCl3)δ8.48(s,1H);7.94(s,1H);7.88(s,1H);7.43(d,J=8.0Hz,2H);7.22(d,J=8.2Hz,2H);5.71(d,J =7.4Hz,1H);5.30-5.23(m,1H);4.14(s,3H);3.56(s,2H);1.52(d,J=7.0Hz,3H);1.37-1.34(m,2H);1.02-1.01(m,2H). 19 F-NMR (377MHz, CDCl3)δ -70.08(s). Example 11 (R)-2-(4-isopropylphenyl)-N-(1-(2-propyl-2H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide

[0282] Synthesis of (R,E)-2-methyl-N-((2-propyl-2H-pyrazolo[3,4-c]pyridin-5-yl)methylene)propane-2-sulfinamide [ka]

[0283] To a solution of 2-propyl-2H-pyrazolo[3,4-c]pyridine-5-carbaldehyde (34 mg) in CHCl (2 mL) was added (R)-2-methylpropane-2-sulfinamide (28.3 mg) and copper sulfate pentahydrate (129.5 mg) at room temperature. The reaction mixture was stirred at room temperature overnight, and LC / MS showed the reaction was complete. The reaction mixture was quenched with ice water and then extracted with EtOAc. The combined extracts were concentrated, and the residue was purified by silica gel column chromatography (PE:EA=3:1) to give (R,E)-2-methyl-N-((2-propyl-2H-pyrazolo[3,4-c]pyridin-5-yl)methylene)propane-2-sulfinamide (25 mg, yellow solid). LC / MS ESI (m / z): 293 [M+H] + .

[0284] Synthesis of (R)-2-methyl-N-((R)-1-(2-propyl-2H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)propane-2-sulfinamide [ka]

[0285] To a solution of (R,E)-2-methyl-N-((2-propyl-2H-pyrazolo[3,4-c]pyridin-5-yl)methylene)propane-2-sulfinamide (100 mg) in THF (3.0 mL) was added CHMgBr (0.45 mL; 3 M in ether) under nitrogen at −78° C. The mixture was stirred at −78° C. for 2 hours, and LC / MS indicated the reaction was complete. The reaction mixture was quenched with ice water and then extracted twice with EtOAc. The combined extracts were concentrated, and the residue was purified by silica gel column chromatography (PE:EA=3:1) to give (R)-2-methyl-N-((R)-1-(2-propyl-2H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)propane-2-sulfinamide (80 mg) as a yellow solid. LC / MS ESI(m / z): 309 [M+H] + .

[0286] Synthesis of (R)-1-(2-propyl-2H-pyrazolo[3,4-c]pyridin-5-yl)ethan-1-amine hydrochloride [ka]

[0287] To a solution of (R)-2-methyl-N-((R)-1-(2-propyl-2H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)propane-2-sulfinamide (80 mg) in dioxane (3 mL) was added HCl-dioxane (0.18 mL, 4N in dioxane) at room temperature. The mixture was stirred at room temperature for 2 hours, and LC / MS showed the reaction was complete. The solvent was concentrated to give (R)-1-(2-propyl-2H-pyrazolo[3,4-c]pyridin-5-yl)ethan-1-amine hydrochloride (120 mg, yellow solid), which was used directly in the next step. LC / MS ESI (m / z): 204 [M+H] + .

[0288] Step 4: Synthesis of (R)-2-(4-isopropylphenyl)-N-(1-(2-propyl-2H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide [ka]

[0289] To a solution of 2-(4-isopropylphenyl)acetic acid (50 mg) in DMF (3.0 mL) was added (R)-1-(2-propyl-2H-pyrazolo[3,4-c]pyridin)-5-yl)ethan-1-amine hydrochloride (57.0 mg), HATU (160 mg), and DIEA (108.6 mg). The mixture was stirred at room temperature for 2 hours, and LC / MS showed the reaction was complete. The reaction mixture was quenched with ice water and then extracted twice with EtOAc. The combined extracts were concentrated, and the residue was purified by column chromatography on silica gel (PE:EA = 1:1) and further purified by preparative HPLC [column: YMC-Triart C18, 250 × 20.0 mm; mobile phase: 80% to 5% HO (0.1% FA), 20% to 95% MeCN; flow rate: 20 mL / min; wavelength: 220 nm / 254 nm] and SFC (column: ChiralPak® IA, 250 × 21.3 mm internal diameter (ID), 5 μm; mobile phase: CO2 in phase A, MEOH + 0.1% NH3·HO in phase B, gradient: B 40%, wavelength: 220 nm) to give (R)-2-(4-isopropylphenyl)-N-(1-(2-propyl-2H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide (6.7 mg) as a colorless oil. LC / MS ESI(m / z):365[M+H] + .

[0290] 1 H-NMR(400MHz,CDCl3)δ9.12(s,1H);7.89(s,1H);7.39(s,1H);7.17(s,4H);6.66(d,J=7.7Hz,1H);5.23-5.15(m,1H);4.42(t,J=7.1H) z,2H);3.53-3.52(m,2H);2.90-2.85(m,1H);2.08-2.02(m,2H));1.44(t,J=6.9Hz,3H);1.24(d,J=6.9Hz,6H);0.95(t,J=7.4Hz,3H). Example 12 (R)—N-(1-(1-methyl-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)-2-(4-(1-(trifluoromethyl)cyclopropyl)phenyl)acetamide

[0291] Synthesis of (R)-N-(1-(1-methyl-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)-2-(4-(1-(trifluoromethyl)cyclopropyl)phenyl)acetamide [ka]

[0292] To a solution of (R)-1-(1-methyl-1H-pyrazolo[3,4-c]pyridin-5-yl)ethan-1-amine hydrochloride (41.2 mg) in anhydrous DMF (5.0 mL) was added DIEA (0.21 mL) to give solution A. To a solution of 2-(4-(1-(trifluoromethyl)cyclopropyl)phenyl)acetic acid (51.9 mg) in anhydrous DMF (5.0 mL) was added HATU (88.9 mg) and stirred at room temperature for 5 minutes to give solution B. Solution A was added to solution B and stirred at room temperature for 1 hour. The reaction was then diluted with EA and water. The organic layer was separated, washed with saturated NaCl, dried over Na2SO4, and concentrated in vacuo to give a residue that was purified. Purification by SFC (column: YMC-Actus Triart C18, 250 x 21 mm; mobile phase: 20%-95% MeCN / HO (0.1% FA); flow rate: 14 mL / min; wavelength: 220 nm / 254 nm) and SFC (column: ChiralPak® IA, 250 x 21.3 mm ID, 5 μm; mobile phase: CO2 in phase A, MEOH + 0.1% NH3·HO in phase B, gradient: B 40%, wavelength: 220 nm) gave (R)—N-(1-(1-methyl-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)-2-(4-(1-(trifluoromethyl)cyclopropyl)phenyl)acetamide (28.5 mg). LC / MS ESI (m / z): 403 [M+H] + . 1H-NMR(400MHz,CDCl3)δ8.80(s,1H);7.97(s,1H);7.49(s,1H);7.42(d,J=8.0Hz,2H);7.24(d,J=8.1Hz,2H);6.74(d,J =7.6Hz,1H);5.27-5.20(m,1H);4.17(s,3H);3.57(s,2H);1.47(d,J=6.8Hz,3H);1.36-1.34(m,2H);1.04-1.00(m,2H). 19 F-NMR (377MHz, CDCl3)δ -70.07(s). Example 13 (R)-2-(4-isopropylphenyl)-N-(1-(1-methyl-1H-pyrazolo[3,4-c]pyridazin-5-yl)ethyl)acetamide

[0293] Synthesis of 5-chloro-1-methyl-1H-pyrazolo[3,4-c]pyridazin-3-ol [ka]

[0294] A solution of methyl 3,6-dichloropyridazine-4-carboxylate (500 mg), methylhydrazine sulfate (417 g), and DIEA (1.1 g) in MeOH (20 mL) was stirred at 70° C. for 12 hours. The mixture was concentrated to give a residue, which was purified by column chromatography on silica gel (DCM:MeOH=50:1 to 10:1) to give 5-chloro-1-methyl-1H-pyrazolo[3,4-c]pyridazin-3-ol (400 mg) as an orange solid. LC / MS ESI (m / z): 185 (M+H). + . 1 H-NMR (400MHz, CDCl3) δ11.90(s,1H);8.23(s,1H);3.96(s,3H).

[0295] Synthesis of 5-chloro-1-methyl-1H-pyrazolo[3,4-c]pyridazin-3-yl trifluoromethanesulfonate [ka]

[0296] To a solution of 5-chloro-1-methyl-1H-pyrazolo[3,4-c]pyridazin-3-ol (2.0 g) in DCM (30 mL) was added N-bis(trifluoromethanesulfonyl)aniline (7.7 g), TEA (4.5 mL), and DMAP (264.7 mg). The mixture was stirred at 25 °C for 12 h and then washed with water. The organic layer was dried, filtered, and concentrated to give a residue, which was purified by silica gel chromatography (PE:DCM = 50:1 to 0:1) to give 5-chloro-1-methyl-1H-pyrazolo[3,4-c]pyridazin-3-yl trifluoromethanesulfonate (600 mg) as a white solid. LC / MS ESI (m / z): 317 [M+H] + .

[0297] Synthesis of 5-chloro-1-methyl-1H-pyrazolo[3,4-c]pyridazine [ka]

[0298] A solution of 5-chloro-1-methyl-1H-pyrazolo[3,4-c]pyridazin-3-yl trifluoromethanesulfonate (1.1 g), Pd(PPh3)4 (0.20 g), and triethylsilane (1.12 mL) in DMF (2 mL) was stirred at 60 °C for 12 h. The mixture was diluted with water and DCM, the two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over Na2SO4, filtered, and concentrated to give a residue that was purified by silica gel chromatography (PE:DCM = 50:1 to 0:1) to give 5-chloro-1-methyl-1H-pyrazolo[3,4-c]pyridazine (400 mg) as a white solid. LC / MS ESI (m / z): 169 [M+H] + . 1 H-NMR (400MHz, CDCl3) δ8.00 (s, 1H); 7.79 (s, 1H); 4.28 (s, 3H).

[0299] Synthesis of 1-methyl-5-vinyl-1H-pyrazolo[3,4-c]pyridazine [ka]

[0300] A solution of 5-chloro-1-methyl-1H-pyrazolo[3,4-c]pyridazine (400 mg), vinyl pinacol borate (0.80 mL), KPO (1.2 g), and dichloro[1,1′-bis(dicyclohexylphosphino)ferrocene]palladium(II) (179.3 mg) in dioxane (20 mL) and HO (4 mL) was sparged with N and stirred at 100 °C under N for 12 h. The mixture was diluted with water and extracted with EtOAc. The combined extracts were dried, filtered, and concentrated to give a residue that was purified by chromatography (PE:DCM = 50:1 to 0:1) to give 1-methyl-5-vinyl-1H-pyrazolo[3,4-c]pyridazine (400 mg, crude) as a yellow solid, which was used in the next step. LC / MS ESI (m / z): 161 [M+H] + .

[0301] Synthesis of 1-methyl-1H-pyrazolo[3,4-c]pyridazine-5-carbaldehyde [ka]

[0302] Ozone was bubbled through a solution of 1-methyl-5-vinyl-1H-pyrazolo[3,4-c]pyridazine (382 mg) in anhydrous DCM (10 mL) at −78° C. for 10 min. The excess ozone was purged with nitrogen for 10 min, after which PPh3 (751 mg) was added and stirred at room temperature for 40 min. The reaction mixture was concentrated in vacuo to give the crude product, which was used in the next step. LC / MS ESI (m / z): 163 [M+H] + .

[0303] Synthesis of (R,E)-2-methyl-N-((1-methyl-1H-pyrazolo[3,4-c]pyridazin-5-yl)methylene)propane-2-sulfinamide [ka]

[0304] To a solution of 1-methyl-1H-pyrazolo[3,4-c]pyridazine-5-carbaldehyde (400 mg) in DCM (10 mL) was added CuSO (1.18 g) and (R)-2-methylpropane-2-sulfinamide (358.8 mg), and the reaction mixture was stirred at room temperature overnight. The solution was filtered through a pad of Celite, and the filtrate was concentrated. The residue was purified by silica gel column to give (R,E)-2-methyl-N-((1-methyl-1H-pyrazolo[3,4-c]pyridazin-5-yl)methylene)propane-2-sulfinamide (255 mg). LC / MS ESI (m / z): 266 [M+H] + . 1 H-NMR (400MHz, CDCl3) δ9.26 (s, 1H); 8.59 (s, 1H); 8.24 (s, 1H); 4.42 (s, 3H); 1.32 (s, 9H).

[0305] Synthesis of (R)-2-methyl-N-((R)-1-(1-methyl-1H-pyrazolo[3,4-c]pyridazin-5-yl)ethyl)propane-2-sulfinamide [ka]

[0306] To a solution of (R,E)-2-methyl-N-((1-methyl-1H-pyrazolo[3,4-c]pyridazin-5-yl)methylene)propane-2-sulfinamide (205 mg) in anhydrous THF (15 mL) was added 3 M CHMgBr (1.29 mL, 3.0 M in ether) at −78 °C under nitrogen. The reaction mixture was stirred at −78 °C for 2 h. Then, the reaction mixture was quenched with saturated NHCl solution and extracted with EA. The combined extracts were washed with brine, dried over NaSO, and concentrated in vacuo. The residue was purified by silica gel column chromatography to give (R)-2-methyl-N-((R)-1-(1-methyl-1H-pyrazolo[3,4-c]pyridazin-5-yl)ethylpropane-2-sulfinamide (50 mg). LC / MS ESI (m / z): 282 [M+H] + .1 H-NMR (400MHz, CDCl3) δ8.08(s,1H);7.87(s,1H);4.95(t,J=6.8Hz,1H);4.35(s,3H);1.71(d,J=6.8Hz,3H);1.26(s,9H)ppm.

[0307] Synthesis of (R)-1-(1-methyl-1H-pyrazolo[3,4-c]pyridazin-5-yl)ethan-1-amine hydrochloride [ka]

[0308] To a solution of (R)-2-methyl-N-((R)-1-(1-methyl-1H-pyrazolo[3,4-c]pyridazin-5-yl)ethyl)propane-2-sulfinamide (55 mg) in dioxane (1.5 mL) was added 4M HCl-dioxane (1 mL), and the reaction mixture was stirred at room temperature for 30 minutes. The mixture was then concentrated to give (R)-1-(1-methyl-1H-pyrazolo[3,4-c]pyridazin-5-yl)ethan-1-amine hydrochloride as a residue, which was used in the next step. LC / MS ESI (m / z): 178 [M+H] + .

[0309] Synthesis of (R)-2-(4-isopropylphenyl)-N-(1-(1-methyl-1H-pyrazolo[3,4-c]pyridazin-5-yl)ethyl)acetamide (N201201-266) [ka]

[0310] To a solution of (R)-1-(1-methyl-1H-pyrazolo[3,4-c]pyridazin-5-yl)ethan-1-amine hydrochloride (61 mg) in DMF (1 mL) was added DIEA (242.6 mg, 0.31 mL) to give solution A. To a solution of 2-(4-isopropylphenyl)acetic acid (55.8 mg) in DMF (1 mL) was added HATU (13.1 mg) and stirred at room temperature for 5 minutes to give solution B. Solution B was added to solution A and stirred at room temperature for 1 hour. The reaction was then diluted with EA and water. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over Na2SO4, filtered, and concentrated to give a residue. Purification by preparative HPLC (column: YMC-Actus Triart C18, 250 × 21 mm; mobile phase: 20% to 95% MeCN / HO (0.1% TFA); flow rate: 15 mL / min; wavelength: 220 nm / 254 nm) gave (R)-2-(4-isopropylphenyl)-N-(1-(1-methyl-1H-pyrazolo[3,4-c]pyridazin-5-yl)ethyl)acetamide (14.6 mg). LC / MS ESI (m / z): 338 [M+H] + . 1 H-NMR(400MHz,CDCl3)δ8.06(s,1H);7.75(s,1H);7.19(s,4H);6.68(d,J=7.6Hz,1H);5.48-5.41(m,1H );4.34(s,3H);3.55-3.54(m,2H);2.93-2.86(m,1H);1.60(d,J=6.9Hz,3H);1.24(d,J=6.9Hz,6H)ppm. Example 14 (R)—N-(1-(1-(2,2-difluoropropyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)-2-(4-isopropylphenyl)acetamide

[0311] Synthesis of 2,2-difluoropropyl 4-methylbenzenesulfonate [ka]

[0312] To a solution of 2,2-difluoropropan-1-ol (2.0 g) in anhydrous DCM (40 mL) was added TEA (8.7 mL). TsCl (11.9 g) was added to the reaction solution at 0 °C, and the reaction mixture was stirred at room temperature overnight. The reaction mixture was then diluted with water, the two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over Na2SO4, filtered, and concentrated to give a residue, which was purified by silica gel column chromatography to give 2,2-difluoropropyl 4-methylbenzenesulfonate (5 g). LC / MS ESI (m / z): 251 [M+H] + . 1 H-NMR (400MHz, CDCl3) δ7.81(d,J=8.3Hz,2H);7,37(d,J=8.1Hz,2H);4.09(t,J=11.1Hz,2H);2.47(s,3H);1.64(t,J=18.7Hz,3H)ppm. 19 F-NMR (377MHz, CDCl3)δ -98.43(s).

[0313] Synthesis of (2,2-difluoropropyl)-1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde [ka]

[0314] To a solution of 1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde (498 mg) in anhydrous DMF (30 mL) was added CsCO (1.3 g) and 2,2-difluoropropyl 4-methylbenzenesulfonate (1.0 g). The reaction mixture was stirred at 100 °C overnight, and then the reaction mixture was diluted with EA and water. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over NaSO, filtered, and concentrated to give a residue, which was purified by silica gel column chromatography to give 1-(2,2-difluoropropyl)-1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde (181 mg). LC / MS ESI (m / z): 226 [M+H] + . 1H-NMR (400MHz, CDCl3) δ10.23(s,1H);9.13(s,1H);8.41(d,J=1.2Hz,1H);8.29(s,1H);4.88(t,J=12.1Hz,2H);1.67(t,J=18.7Hz,3H)ppm. 19 F-NMR (377MHz, CDCl3)δ -93.83(s).

[0315] Synthesis of (R,E)-N-((1-(2,2-difluoropropyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)methylene)-2-methylpropane-2-sulfinamide [ka]

[0316] To a solution of 1-(2,2-difluoropropyl)-1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde (209 mg) in anhydrous DCM (14 mL) was added CuSO (444.4 mg) and (R)-2-methylpropane-2-sulfinamide (146.2 mg). The reaction mixture was stirred overnight at room temperature, then the solution was filtered through a pad of Celite, and the filtrate was concentrated. The residue was purified by silica gel column to give (R,E)-N-((1-(2,2-difluoropropyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)methylene)-2-methylpropane-2-sulfinamide (229 mg). LC / MS ESI (m / z): 329 [M+H] + . 1 H-NMR(400MHz,CDCl3)δ9.12(s,1H);8.84(s,1H);8.36(d,J=1.1Hz,1H);8.23(d ,J=0.6Hz,1H);4.86(t,J=12.1Hz,2H);1.66(t,J=18.4Hz,3H);1.31(s,9H)ppm. 19 F-NMR (377MHz, CDCl3)δ -93.83(s).

[0317] Synthesis of (R)-N-((R)-1-(1-(2,2-difluoropropyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)-2-methylpropane-2-sulfinamide [ka]

[0318] To a solution of (R,E)-N-((1-(2,2-difluoropropyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)methylene)-2-methylpropane-2-sulfinamide (229 mg) in anhydrous THF (9.0 mL) at −78° C. under nitrogen, CHMgBr (1.2 mL, 3.0 M in ether) was added dropwise. The reaction was stirred at −78° C. for 2 h. The reaction was then quenched with aqueous NHCl and extracted with EA. The organic layer was separated, washed with brine, dried, and concentrated in vacuo. The residue was purified by silica gel column chromatography to give (R)-N-((R)-1-(1-(2,2-difluoropropyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)-2-methylpropane-2-sulfinamide (193 mg) as a yellow oil. LC / MS ESI(m / z):345[M+H] + . 1 H-NMR(400MHz,CDCl3)δ8.97(s,1H);8.06(d,J=0.5Hz,1H);7.62(d,J=0.9Hz,1H);4.79(t,J =12.0Hz,2H);4.73-4.66(m,1H);4.49(d,J=6.2Hz,1H);1.68-1.56(m,6H);1.25(s,9H)ppm. 19 F-NMR(377MHz, CDCl3)δ -93.56(d,J=5.5Hz).

[0319] Synthesis of (R)-1-(1-(2,2-difluoropropyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethan-1-amine hydrochloride [ka]

[0320] To a solution of (R)-N-((R)-1-(1-(2,2-difluoropropyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)-2-methylpropane-2-sulfinamide (153 mg) in dioxane (3.0 mL) was added 4N HCl-dioxane (1.5 mL). The reaction mixture was stirred at room temperature for 30 minutes, and then the mixture was concentrated to give crude (R)-1-(1-(2,2-difluoropropyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethan-1-amine hydrochloride, which was used directly in the next step. LC / MS ESI (m / z): 241 [M+H] + .

[0321] Synthesis of (R)-N-(1-(1-(2,2-difluoropropyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)-2-(4-isopropylphenyl)acetamide [ka]

[0322] To a solution of (R)-1-(1-(2,2-difluoropropyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethan-1-amine hydrochloride (123 mg) in anhydrous DMF (3 mL) was added DIEA to give solution A. To a solution of 2-(4-isopropylphenyl)acetic acid (80.75 mg) in anhydrous DMF, HATU (156 mg) was added and stirred at room temperature for 15 minutes to give solution B. Solution B was added to solution A and stirred at room temperature for 1 hour. The reaction mixture was then diluted with EA and water. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over Na2SO4, filtered, and concentrated to give a residue that was purified by [column: YMC-Actus Triart C18, 250 × 21 mm; mobile phase: 20%–95% MeCN / HO (0.1% FA); flow rate: 15 mL / min; wavelength: 220 nm / 254 nm] and SFC (column: ChiralPak® IA, 250 × 21.3 mm ID, 5 μm; mobile phase: CO2 for phase A, MeOH + 0.1% NH3·HO for phase B; gradient: B 40%; wavelength: 220 nm) to give (R)—N-(1-(1-(2,2-difluoropropyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)-2-(4-isopropylphenyl)acetamide (98.6 mg). LC / MS ESI(m / z):401[M+H] + . 1 H-NMR(400MHz,CDCl3)δ8.88(s,1H);8.04(s,1H);7.51(s,1H);7.19(s,4H);6.61(d,J=7.4Hz,1H);5.29-5.22(m,1H);4.7 8(t,J=12.0Hz,2H);3.55-3.54(m,2H);2.94-2.87(m,1H);1.66-1.56(m,3H);1.46(d,J=6.8Hz,3H);1.25(d,J=6.9Hz,6H). 19 F-NMR (377MHz, CDCl3)δ -93.56(s). Example 15 (R)—N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)-2-(4-(trifluoromethyl)phenyl)acetamide

[0323] Synthesis of (R)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)-2-(4-(trifluoromethyl)phenyl)acetamide [ka]

[0324] A solution of 2-(4-(trifluoromethyl)phenyl)acetic acid (96.4 mg) and HATU (269.5 mg) in 3 mL of DMF was stirred at room temperature for 15 minutes (Solution A). DIEA (0.48 mL) was added to (R)-1-[1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl]ethan-1-amine hydrochloride (see Example 9, 150 mg) in 2 mL of DMF until the pH of the solution was higher than 7 using moistened pH paper (Solution B). Solution B was added to Solution A, and the reaction was stirred for 1 hour, at which point LCMS indicated the reaction was complete. The reaction was diluted with EA and water. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 2). The combined organic layers were dried over Na2SO4, filtered, and concentrated in vacuo. The residue was purified by silica gel column chromatography, eluting with methanol (0–7%) in DCM to give a crude product, which was purified by preparative HPLC (column: YMC-Actus Triart C18, 250 × 20 mm; mobile phase: 28%–95% MeCN / HO (0.1% FA); flow rate: 15 mL / min; wavelength: 220 nm / 254 nm) and SFC (column: ChiralPak TM Purification by HPLC (IA, 250 × 21.2 mm ID, 5 μm; mobile phase: CO₂ in phase A, MEOH + 0.1% NH₃·H₂O in phase B; gradient: B 40%; flow rate: 50 mL / min; wavelength: 210 nm) gave N-[(1R)-1-[1-(2,2,2-trifluoroethyl)])-1H-pyrazolo[3,4-c]pyridin-5-yl]ethyl]-2-[4-(trifluoromethyl)phenyl]acetamide (92.1 mg) as a white solid. LC / MS ESI (m / z): 431 [M+H] + . 1H-NMR(400MHz,CDCl3):δ8.87(s,1H);8.10(s,1H);7.63-7.51(m,3H);7.40(d,J=8.0Hz,2H);6.7 2(d,J=6.9Hz,1H);5.26-5.24(m,1H);5.04(q,J=8.3Hz,2H);3.63(s,2H);1.48(d,J=6.8Hz,3H). 19 F-NMR (377MHz, CDCl3): δ -62.54, -70.81. Example 16 (R)-2-(6-(1,1-difluoroethyl)pyridin-3-yl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide

[0325] Synthesis of 2-(1,1-difluoroethyl)-5-methylpyridine [ka]

[0326] To a solution of 1-(5-methylpyridin-2-yl)ethan-1-one (2.0 g) in DCM (30 mL) was added DAST (19.6 mL) at 0 °C. The mixture was stirred at 25 °C for 48 h. The mixture was quenched with NaHCO solution. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over NaSO, filtered, and concentrated to give a residue, which was purified by silica gel chromatography (PE:EA = 1:0 to 20:1) to give 2-(1,1-difluoroethyl)-5-methylpyridine (1.1 g) as a colorless oil. LC / MS ESI (m / z): 158 [M+H] + .

[0327] Synthesis of 5-(bromomethyl)-2-(1,1-difluoroethyl)pyridine [ka]

[0328] To a solution of 2-(1,1-difluoroethyl)-5-methylpyridine (1.02 g) in CCl4 (30 mL) was added NBS (1.28 g) and AIBN (0.048 mL). The reaction mixture was stirred at 80 °C overnight. The reaction mixture was filtered, and the filtrate was concentrated in vacuo to give a residue that was purified by silica gel column chromatography (PE:EA = 1:0 to 10:1) to give 5-(bromomethyl)-2-(1,1-difluoroethyl)pyridine (780 mg). 1 H-NMR (400MHz, CDCl3) δ8.65(d,J=1.6Hz,1H);7.85-7.83(m,1H);7.66-7.64(m,1H);4.49(s,2H);2.07-1.97(m,3H). LC / MS ESI(m / z):237[M+H] + .

[0329] Synthesis of 2-(6-(1,1-difluoroethyl)pyridin-3-yl)acetonitrile [ka]

[0330] To a solution of 5-(bromomethyl)-2-(1,1-difluoroethyl)pyridine (780 mg) in DMF (15 mL) was added TMSCN (1.6 mL) and K2CO3 (685 mg). The reaction was stirred at 50 °C overnight. The reaction was diluted with DCM and water. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over Na2SO4, filtered, and concentrated to give a residue that was purified by silica gel column chromatography (PE:EA = 10:1) to give 2-[6-(1,1-difluoroethyl)pyridin-3-yl]acetonitrile (194 mg). LC / MS ESI (m / z): 183 [M+H] + .

[0331] Synthesis of 2-(6-(1,1-difluoroethyl)pyridin-3-yl)acetic acid [ka]

[0332] A solution of 2-[6-(1,1-difluoroethyl)pyridin-3-yl]acetonitrile (194 mg) in concentrated aqueous HCl (3.0 mL) was stirred at 100° C. for 2 hours. The mixture was added to water and extracted with EA. The combined extracts were dried, filtered, and concentrated to give 2-(6-(1,1-difluoroethyl)pyridin-3-yl)acetic acid (110 mg, crude) as a white solid. LC / MS ESI (m / z): 200 [MH] - .

[0333] Synthesis of (R)-2-(6-(1,1-difluoroethyl)pyridin-3-yl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide [ka]

[0334] A solution of 2-[6-(1,1-difluoroethyl)pyridin-3-yl]acetic acid (110 mg) and HATU (312 mg) in DMF (2.0 mL) was stirred at room temperature for 15 minutes. Next, (R)-1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethan-1-amine hydrochloride (173.6 mg) and DIEA (0.542 mL, 3.281 ml) were added. The mixture was stirred at room temperature for 1 hour, and LC / MS showed the reaction was complete. The reaction solution was diluted with EA and water, the two layers were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over Na2SO4, filtered, and concentrated to give a residue, which was purified by preparative HPLC [column: YMC-Actus Triart C18, 250 × 20 mm; mobile phase: 15%-95% MeCN / HO (1% FA); flow rate: 15 mL / min; wavelength: 220 nm / 254 nm] and SFC (column: ChiralPak IA, 250 × 21.3 mm Purification by HPLC (ID, 5 μm, mobile phase: CO₂ in phase A, IPA + 0.1% NH₃·H₂O in phase B, gradient: B 40%, flow rate: 50 mL / min; wavelength: 210 nm) gave (R)-2-(6-(1,1-difluoroethyl)pyridin-3-yl)-N-(1-(1-(2,2,2-trifluoroethyl))-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide (121 mg) as a white solid. LC / MS ESI (m / z): 428 [M+H] + . 1 H-NMR(400MHz,CDCl3)δ8.93(s,1H);8.54(s,1H);8.13(s,1H);7.78(dd,J=8.1Hz,2.1Hz,1H);7.60(dd,J=4.2Hz,3.2Hz,2H );6.96(t,J=14.2Hz,1H);5.28-5.24(m,1H);5.07(q,J=8.3Hz,2H);3.62(s,2H);2.05-1.95(m,3H);1.52(t,J=6.3Hz,3H). 19 F-NMR (377MHz, CDCl3)δ -70.76, -90.68. Example 17 (R)—N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)-2-(5-(1-(trifluoromethyl)cyclopropyl)thiophen-2-yl)acetamide

[0335] Step 1: Synthesis of (1-(5-bromothiophen-2-yl)-2,2,2-trifluoroethoxy)trimethylsilane [ka]

[0336] To a solution of 5-bromothiophene-2-carbaldehyde (8.4 mL, 70.7 mmol) in 1,2-dimethoxyethane (30 mL) was added (trifluoromethyl)trimethylsilane (13.7 mL, 91.9 mmol) and cesium fluoride (0.26 mL, 7.01 mmol) at 0 °C. The mixture was stirred at room temperature for 3 hours, and TLC showed that the reaction was complete. The reaction mixture was quenched with ice water and then extracted twice with EtOAc. The combined extracts were concentrated, and the residue was purified by silica gel column chromatography (PE:EA = 3:1) to give (1-(5-bromothiophen-2-yl)-2,2,2-trifluoroethoxy)trimethylsilane (19.6 g) as a colorless oil.

[0337] Step 2: Synthesis of 1-(5-bromothiophen-2-yl)-2,2,2-trifluoroethan-1-ol [ka]

[0338] To a solution of (1-(5-bromothiophen-2-yl)-2,2,2-trifluoroethoxy)trimethylsilane (19.4 g, 58.2 mmol) in MeOH (20 mL) was added HCl (9.7 mL, 116.4 mmol, 12 mol / L) at 0° C. The mixture was stirred at room temperature for 3 hours, and TLC showed that the reaction was complete. The reaction mixture was quenched with ice water and then extracted twice with EtOAc. The combined extracts were concentrated, and the residue was purified by silica gel column chromatography (PE:EA=5:1) to give 1-(5-bromothiophen-2-yl)-2,2,2-trifluoroethan-1-ol (14.8 g) as a colorless oil.

[0339] Step 3: Synthesis of 1-(5-bromothiophen-2-yl)-2,2,2-trifluoroethan-1-one [ka]

[0340] To a solution of 1-(5-bromothiophen-2-yl)-2,2,2-trifluoroethan-1-ol (17.3 g, 66.3 mmol) in CHCl (30 mL) was added Dess-Martin periodinane (30.9 mL, 99.4 mmol) at 0 °C. The mixture was warmed to room temperature and stirred for 3 h. TLC showed the reaction was complete. The reaction mixture was quenched with ice water and then extracted twice with EtOAc. The combined extracts were concentrated, and the residue was purified by silica gel column chromatography (100% PE) to give 1-(5-bromothiophen-2-yl)-2,2,2-trifluoroethan-1-one (10.4 g) as a pale yellow oil. 1 H-NMR (400MHz, CDCl3): δ 7.64 (dd, J=2.5, 1.3Hz, 1H); 7.17-7.12 (m, 1H). 19 F-NMR (377MHz, CDCl3): δ -72.2.

[0341] Step 4: Synthesis of 2-bromo-5-(3,3,3-trifluoroprop-1-en-2-yl)thiophene [ka]

[0342] Methyltriphenylphosphonium iodide (1.8 g, 4.6 mmol) was dissolved in THF (25 mL) under an argon atmosphere. The suspension was cooled to 0 °C, followed by the dropwise addition of n-BuLi (2.5 mL, 2.5 N). The mixture was stirred at 0 °C for 10 minutes, and then the reaction was cooled to -78 °C. 1-(5-bromothiophen-2-yl)-2,2,2-trifluoroethanone (1.0 g, 3.9 mmol) in THF (1.0 mL) was added. The mixture was stirred at -78 °C for 30 minutes, then stirred at room temperature overnight. The reaction was quenched with saturated NaHCO3, washed with NH4Cl, and extracted with petroleum ether (10 mL). The organic layer was washed with saturated NaCl, dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated, and the residue was purified by silica gel column chromatography (PE:EA=10:1) to give 2-bromo-5-(3,3,3-trifluoroprop-1-en-2-yl)thiophene as a colorless oil (435 mg). 1 H-NMR (400MHz, CDCl3): δ7.00-6.98 (m, 1H); 6.96-6.93 (m, 1H); 5.82 (s, 1H); 5.75 (s, 1H). 19 F-NMR (377MHz, CDCl3): δ -65.9.

[0343] Step 5: Synthesis of 2-bromo-5-(1-(trifluoromethyl)cyclopropyl)thiophene [ka]

[0344] To an oven-dried 20 mL vial containing 2-bromo-5-(3,3,3-trifluoroprop-1-en-2-yl)thiophene (200 mg, 0.78 mmol) and methyldiphenylsulfonium tetrafluoroborate (291 mg, 1.0 mmol) in anhydrous tetrahydrofuran (5 mL) was added sodium bis(trimethylsilyl)amide (1 M in THF, 1.2 mL) under nitrogen at 0 °C. The reaction mixture was stirred at 0 °C for 10 min and then at room temperature for 1 h. The reaction was quenched with 250 μL of methanol, and the crude mixture was concentrated in vacuo to give a residue that was purified by automated flash column chromatography (eluent: 100% PE → 50% PE in EA) to give 2-bromo-5-(1-(trifluoromethyl)cyclopropyl)thiophene as a colorless oil (70 mg). 1 H-NMR (400MHz, CDCl3): δ6.90 (d, J=3.6Hz, 1H); 6.86 (d, J=4.0Hz, 1H); 1.42-1.39 (m, 2H); 1.15-1.11 (m, 2H). 19 F-NMR (377MHz, CDCl3): δ -70.4.

[0345] Step 6: Synthesis of ethyl 2-(5-(1-(trifluoromethyl)cyclopropyl)thiophen-2-yl)acetate [ka]

[0346] A Schlenk tube equipped with a stir bar was evacuated and refilled with nitrogen. Pd2(allyl)2Cl2 (3.65 mg, 0.01 mmol), BINAP (18.7 mg, 0.03 mmol), DMAP (6.1 mg, 0.05 mmol), and potassium ethyl malonate (128 mg, 0.75 mmol) were added. 2-Bromo-5-(1-(trifluoromethyl)cyclopropyl)thiophene (136 mg, 0.5 mmol) and mesitylene (2 mL) were then added. The reaction mixture was stirred at room temperature under N2 in a sealed tube for 10 minutes and then heated at 120 °C overnight. Upon completion, the reaction was cooled to room temperature and diluted with EA (30 mL). The resulting mixture was washed with saturated NaCl and concentrated. The residue was purified by flash column chromatography (eluted with 5% to 10% PE in EA) to give ethyl 2-(5-(1-(trifluoromethyl)cyclopropyl)thiophen-2-yl)acetate as a yellow liquid (50 mg). 1 H-NMR(400MHz, CDCl3):δ 6.95(d,J=3.6Hz,1H);6.78(d,J=4.0Hz,1H);4.21-4.16(m,2H);3.76(s,2H);1.40-1.37(m,2H);1.28(t,J=6.8Hz,3H);1.16-1.12(m,2H)ppm. 19 F-NMR (377MHz, CDCl3): δ -70.34ppm.

[0347] Step 7: Synthesis of 2-(5-(1-(trifluoromethyl)cyclopropyl)thiophen-2-yl)acetic acid [ka]

[0348] To a mixture of ethyl 2-((5-(1-(trifluoromethyl)cyclopropyl)thiophen-2-yl)acetate (50 mg, 0.18 mmol) in ethanol (10 mL) was added NaOH (7.2 mg, 0.18 mmol), and the mixture was stirred under reflux for 16 h. After cooling to room temperature, the pH of the reaction mixture was adjusted to 2 by adding 1 N hydrogen chloride. The reaction mixture was then extracted with ethyl acetate (2 × 20 mL), and the combined extracts were washed with saturated NaCl. The organic phase was concentrated to give 2-(5-(1-(trifluoromethyl)cyclopropyl)thiophen-2-yl)acetic acid as a brown solid (40 mg). 1 H-NMR (400MHz, CDCl3): 6.96 (d, J = 3.6 Hz, 1H); 6.80 (d, J = 3.6 Hz, 1H); 3.82 (s, 2H); 1.41-1.38 (m, 2H); 1.16-1.14 (m, 2H). 19 F-NMR (377MHz, CDCl3): δ -70.4.

[0349] Step 8: Synthesis of (R)—N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)-2-(5-(1-(trifluoromethyl)cyclopropyl)thiophen-2-yl)acetamide [ka]

[0350] A solution of 2-(5-(1-(trifluoromethyl)cyclopropyl)thiophen-2-yl)acetic acid (158 mg) and HATU (360 mg) in DMF (2.5 mL) was stirred at room temperature for 15 minutes. Then, (R)-1-[1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl]ethan-1-amine hydrochloride (200 mg) and DIEA (0.63 mL) were added, and the mixture was stirred for 1 hour. LC / MS showed that the reaction was complete. The reaction mixture was diluted with EA and water. The organic layer was separated, washed with brine, dried over Na2SO4, filtered, and concentrated in vacuo. The residue was purified by preparative HPLC [column: YMC-Actus Triart C18, 250 × 21 mm; mobile phase: 30%–95% MeCN with HO (0.1% FA); flow rate: 15 mL / min; wavelength: 220 nm / 254 nm] and SFC [column: ChiralPak® IA, 250 × 21.3 mm ID, 5 μm; mobile phase: CO in phase A, MEOH + 0.1% NH₃·HO in phase B, gradient: B 40%; flow rate: 50 mL / min; wavelength: 210 nm] to give (R)—N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)-2-(5-(1-(trifluoromethyl)cyclopropyl)thiophen-2-yl)acetamide (80.9 mg) as a white solid. LC / MS ESI(m / z):477[M+H] + . 1 H-NMR(400MHz,CDCl3)δ8.90(s,1H);8.12(s,1H);7.60(s,1H);6.97(d, J=3.5Hz,2H);6.79(d,J)=3.5Hz,1H);5.28- 5.24(m,1H);5.05(q,J=8.3Hz,2H);3.78-3.68(m,2H);1.51(d,J=6.8Hz,3H);1.39-1.35(m,2H);1.13-1.11(m,2H). 19 F-NMR (377MHz, CDCl3)δ -70.32, -70.81. Example 18 (R)—N-(1-(1-((1-fluorocyclopropyl)methyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)-2-(4-isopropylphenyl)acetamide

[0351] Step 1: Synthesis of 1-(1-fluorocyclopropyl)methyl)-1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde [ka]

[0352] To a solution of 1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde (546 mg) in anhydrous DMF (1.5 mL) was added CsCO (1.45 g) and (1-fluorocyclopropyl)methyl 4-methylbenzenesulfonate (1.09 g), and the reaction was stirred at room temperature for 4 hours. The reaction was diluted with EA and water. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over NaSO, filtered, and concentrated to give a residue that was purified on a silica gel column to give 1-((1-fluorocyclopropyl)methyl)-1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde (192 mg). LC / MS ESI (m / z): 230 [M+H] + . 1 H-NMR(400MHz,CDCl3)δ10.23(s,1H);9.18(d,J=0.7Hz,1H);8.41(d,J=1.1Hz, 1H);8.27(s,1H);4.88(d,J=21.2Hz,2H);1.29-1.10(m,2H);1.05-0.91(m,2H).

[0353] Synthesis of (R,E)-N-((1-((1-fluorocyclopropyl)methyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)methylene)-2-methylpropane-2-sulfinamide [ka]

[0354] To a solution of 1-((1-fluorocyclopropyl)methyl)-1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde (192 mg) in anhydrous DCM (8.0 mL) was added CuSO4 (419.4 mg) and (R)-2-methylpropane-2-sulfinamide (138.0 mg). The reaction mixture was stirred at room temperature overnight. The solution was then filtered through a pad of Celite, and the filtrate was concentrated. The residue was purified on a silica gel column to give (R,E)-N-((1-((1-fluorocyclopropyl)methyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)methylene)-2-methylpropane-2-sulfinamide (294 mg). LC / MS ESI(m / z): 323 [M+H] + . 1 H-NMR(400MHz,CDCl3)δ9.17(d,J=0.8Hz,1H);8.83(s,1H);8.35(d,J=1.1Hz,1H);8.20(d,J =0.5Hz,1H);4.86(d,J=21.0Hz,2H);1.31(s,9H);1.22-1.13(m,2H);1.01-0.93(m,2H)ppm.

[0355] Synthesis of (R)-N-((R)-1-(1-((1-fluorocyclopropyl)methyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)-2-methylpropane-2-sulfinamide [ka]

[0356] To a solution of (R,E)-N-((1-((1-fluorocyclopropyl)methyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)methylene)-2-methylpropane-2-sulfinamide (290 mg) in anhydrous THF (12 mL) at −78° C. under nitrogen was added CHMgBr (1.499 mL, 3.0 M in ether) dropwise and the reaction was stirred at −78° C. for 2 h. The reaction was then quenched with aqueous NHCl. The two phases were separated and the aqueous phase was extracted with EA. The combined organic layers were dried over Na2SO4, filtered, and concentrated in vacuo to give a residue which was purified by silica gel column chromatography to give (R)-N-((R)-1-(1-((1-fluorocyclopropyl)methyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)-2-methylpropane-2-sulfinamide (228 mg). LC / MS ESI(m / z): 339 [M+H] + .

[0357] Synthesis of (R)-1-(1-((1-fluorocyclopropyl)methyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethan-1-amine hydrochloride [ka]

[0358] To a solution of (R)-N-((R)-1-(1-((1-fluorocyclopropyl)methyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)-2-methylpropane-2-sulfinamide (212 mg) in dioxane (5 mL) was added 4N HCl-dioxane (2.5 mL). The reaction was stirred at room temperature for 30 minutes. The mixture was concentrated and used directly in the next step. LC / MS ESI (m / z): 235 [M+H] + .

[0359] Synthesis of (R)-N-(1-(1-((1-fluorocyclopropyl)methyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)-2-(4-isopropylphenyl)acetamide [ka]

[0360] To a solution of (R)-1-(1-((1-fluorocyclopropyl)methyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethan-1-amine hydrochloride (170 mg) in anhydrous DMF, DIEA (487 mg) was added to obtain solution A. To a solution of 2-(4-isopropylphenyl)acetic acid (107.8 mg) in anhydrous DMF, HATU (252.9 mg) was added and stirred at room temperature for 15 minutes to obtain solution B. Solution B was added to solution A, and the resulting mixture was stirred at room temperature for 1 hour. The reaction mixture was then diluted with EA and water. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over Na2SO4, filtered, and concentrated to give a residue that was purified by preparative HPLC (column: YMC-Actus Triart C18, 250 × 21 mm; mobile phase: 20% to 95% MeCN / HO (1% NH3 HO); flow rate: 15 mL / min; wavelength: 220 nm / 254 nm) to give (R)—N-(1-(1-((1-fluorocyclopropyl)methyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)-2-(4-isopropylphenyl)acetamide (162.2 mg). LC / MS ESI (m / z): 395 [M+H] + . 1 H-NMR(400MHz,CDCl3)δ8.95(s,1H);8.03(s,1H);7.54(s,1H);7.19(s,4H);6.74(d,J=6.9Hz,1H);5.30-5.23(m,1H);4.80(d,J=2 1.1Hz,2H);3.55(s,2H);2.93-2.86(m,1H);1.49(d,J=6.8Hz,3H);1.24(d,J=6.9Hz,6H);1.18-1.13(m,2H), 0.95-0.93(m,2H)ppm; 19 F-NMR (377MHz, CDCl3)δ -183.10(s). Example 19 (R)-2-(4-(1,1-difluoroethyl)phenyl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide

[0361] Step 1: 2-(4-(1,1-difluoroethyl)phenyl)ethyl acetate [ka]

[0362] To a solution of 1-bromo-4-(1,1-difluoroethyl)benzene (0.54 mL), potassium ethyl malonate (924.0 mg), allylpalladium chloride dimer (26.5 mg), BINAP (135.2 mg), and DMAP (442.2 mg) in 1,3,5-trimethylbenzene (10 mL), N was added and stirred at 140° C. for 1 hour. The mixture was then stirred at 120° C. for an additional 12 hours. The reaction was diluted with water and extracted with EtOAc. The combined extracts were dried, filtered, and concentrated to give a residue that was purified by column chromatography on silica gel (PE:EA=20:1) to give ethyl 2-[4-(1,1-difluoroethyl)phenyl]acetate (180 mg) as a colorless oil. 1 H-NMR (400MHz, CDCl3) δ7.47(d,J=8.4Hz,2H);7.34(d,J=8.0Hz,2H);4.20-4.12(m,2H);3.64(s,2H);1.91(t,J=18.4Hz,3H);1.26(t,J=7.2Hz,3H).

[0363] Step 2: 2-(4-(1,1-difluoroethyl)phenyl)acetic acid [ka] To a solution of ethyl 2-[4-(1,1-difluoroethyl)phenyl]acetate (150 mg) in MeOH (0.5 mL) was added NaOH (2 M in HO, 2.0 mL), and the mixture was stirred at room temperature. 12 h at 25 °C. The mixture was adjusted to pH = 3 and extracted with EtOAc. The combined extracts were dried, filtered, and concentrated to give 2-[4-(1,1-difluoroethyl)phenyl]acetic acid (110 mg) as a white solid. LC / MS ESI (m / z): 199 [MH] - .

[0364] Step 3: (R)-2-(4-(1,1-difluoroethyl)phenyl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide [ka]

[0365] A solution of (1R)-1-[1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl]ethan-1-amine hydrochloride (140 mg), 2-[4-(1,1-difluoroethyl)phenyl]acetic acid (114.8 mg), HATU (239.8 mg), and DIEA (0.28 mL) in DMF (5.0 mL) was stirred at 25 °C for 1 hour. The mixture was diluted with water and extracted with DCM. The combined extracts were dried, filtered, and concentrated to give a residue. The residue was purified by preparative HPLC [column: YMC-Actus Triart C18, 250 × 20 mm; mobile phase: 25% to 95% MeCN / HO (0.1% FA); flow rate: 15 mL / min; wavelength: 220 nm / 254 nm] to give (R)-2-(4-(1,1-difluoroethyl)phenyl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo)[3,4-c]pyridin-5-yl)ethyl)acetamide (109.9 mg) as a white solid. LC / MS ESI (m / z): 427 (M+H). + . 1 H-NMR(400MHz,CDCl3)δ8.91(s,1H);8.12(s,1H);7.59(s,1H);7.46(d,J=8.0Hz,2H);7.33(d,J=8.0Hz,2H);6.77 (s,1H);5.28-5.24(m,1H);5.05(q,J=8.4Hz,2H);3.61(s,2H);1.91(t,J=18.0Hz,3H);1.50(d,J=6.8Hz,3H)ppm. 19 F-NMR(377MHz, CDCl3)δ -70.78;-87.35ppm. Example 20 (R)-2-(6-Isopropylpyridin-3-yl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide

[0366] Synthesis of methyl 6-(prop-1-en-2-yl)nicotinate [ka]

[0367] A solution of methyl 6-bromonicotinate (7.0 g) in dioxane / water (20 mL, 9:1) was added with 4,4,5,5-tetramethyl-2-(prop-1-en-2-yl)-1,3,2-dioxaborolane (6.1 mL), Pd(PPh3)4 (0.75 g), PPh3 (1.70 g), and K2CO3 (13.44 g). The mixture was purged with N2 and stirred under nitrogen at 80 °C overnight, at which point LC / MS indicated the reaction was complete. The reaction mixture was quenched with ice water and then extracted twice with EtOAc. The combined extracts were concentrated, and the residue was purified by silica gel column chromatography (PE:EA = 10:1) to give methyl 6-(prop-1-en-2-yl)nicotinate (4.9 g) as a white solid. 1 H-NMR(400MHz,CDCl3)δ9.36-8.82(m,1H);8.24(dd,J=8.3, 2.2Hz,1H);7.54(d,J=8.3H z,1H);6.00(d,J=0.5Hz,1H);5.53-5.02(m,1H);3.94(s,3H);2.23(d,J=0.5Hz,3H)ppm.

[0368] Synthesis of methyl 6-isopropylnicotinate [ka]

[0369] To a solution of methyl 6-(prop-1-en-2-yl)nicotinate (2.5 g) in MeOH (20 mL) was added 10% Pd / C (1.46 mL). The mixture was stirred under hydrogen at room temperature for 2 hours, and LC / MS showed the reaction was complete. The reaction was filtered, and the solvent was concentrated to give a residue that was purified by silica gel column chromatography (PE:EA=5:1) to give methyl 6-isopropylnicotinate (2.0 g) as a colorless oil. 1H-NMR (400MHz, CD3OD) δ9.01(d,J=2.1Hz,1H);8.29(dd,J=8.2Hz,2.1Hz,1H);7.44(d,J=8.2Hz,1H);3.93(s,3H);3.13(m,1H);1.32(d,J=6.9Hz,6H)ppm.

[0370] Synthesis of (6-isopropylpyridin-3-yl)methanol [ka]

[0371] To a solution of methyl 6-isopropylnicotinate (1.5 g) in THF (10 mL) was added LiAlH4 (0.95 g), and the mixture was stirred at room temperature for 4 hours. LC / MS showed that the reaction was complete. The reaction mixture was then quenched with ice water and extracted twice with EtOAc. The combined extracts were concentrated, and the residue was purified by silica gel column chromatography (PE:EA=1:1) to give (6-isopropylpyridin-3-yl)methanol (1.0 g) as a colorless oil. 1 H-NMR(400MHz,DMSO-d6)δ8.41(d,J=1.7Hz,1H);7.63(dd,J=8.0Hz,2.3Hz,1H);7.23(d,J=8.0 Hz,1H);5.24(t,J=5.7Hz,1H);4.48(d,J=5.6Hz,2H);3.05-2.95(m,1H);1.22(d,J=6.9Hz,6H). LC / MS ESI(m / z):152[M+H] + .

[0372] Step 4: Synthesis of 5-(bromomethyl)-2-isopropylpyridine [ka]

[0373] To a solution of (6-isopropylpyridin-3-yl)methanol (1.0 g) in CHCl (10 mL) was added phosphorus tribromide (1.24 mL). The mixture was stirred at room temperature overnight, at which point LC / MS indicated the reaction was complete. The reaction mixture was quenched with ice water and then extracted twice with EtOAc. The combined extracts were concentrated, and the residue was purified by silica gel column chromatography (PE:EA=5:1) to give 5-(bromomethyl)-2-isopropylpyridine (1.1 g) as a colorless oil. 1 H-NMR(400MHz,DMSO-d6)δ8.61-8.47(m,1H);7.86-7.69(m,1H);7.30(t,J=11.2Hz,1H);4.72(s,2H);3.12-2.90(m,1H);1.22(d,J=6.9Hz,6H). LC / MS ESI(m / z):214[M+H] + .

[0374] Step 5: Synthesis of 2-(6-isopropylpyridin-3-yl)acetonitrile [ka]

[0375] To a solution of 5-(bromomethyl)-2-isopropylpyridine (1.1 g) in DMF (15 mL) was added KCN (1.67 g). The mixture was stirred at 50° C. for 3 hours. LC / MS and TLC showed the reaction was complete. The reaction mixture was quenched with ice water and then extracted twice with EtOAc. The combined extracts were concentrated, and the residue was purified by silica gel column chromatography (PE:EA=5:1) to give 2-(6-isopropylpyridin-3-yl)acetonitrile (120 mg) as a colorless oil. 1 H-NMR (400MHz, CDCl3) δ8.49(d,J=1.7Hz,1H);7.65(d,J=8.1Hz,1H);7.23(d,J=8.1Hz,1H);3.74(s,2H);3.21-2.91(m,1H);1.31(d,J=6.9Hz,6H)ppm. LC / MS ESI(m / z):161[M+H] + .

[0376] Step 6: Synthesis of 2-(6-isopropylpyridin-3-yl)acetic acid hydrochloride [ka]

[0377] To a solution of 2-(6-isopropylpyridin-3-yl)acetonitrile (120 mg) in water (3.0 mL) was added 12N HCl (1.4 g). The mixture was stirred at 100° C. for 2 hours, and LC / MS showed the reaction was complete. The reaction mixture was quenched with ice water and then extracted twice with EtOAc. The aqueous phase was evaporated to dryness to give 2-(6-isopropylpyridin-3-yl)acetic acid hydrochloride (130 mg) as a brown solid, which was used in the next step. LC / MS ESI (m / z): 180 [M+H] + .

[0378] Step 7: Synthesis of (R)-2-(6-isopropylpyridin-3-yl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide [ka]

[0379] A solution of (1R)-1-[1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl]ethan-1-amine hydrochloride (140 mg), 2-(6-isopropylpyridin-3-yl)acetic acid hydrochloride (102.0 mg), HATU (239.8 mg), and DIEA (0.28 mL) in DMF (4.0 mL) was stirred at 25° C. for 1 hour. The mixture was diluted with water and DCM. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over Na2SO4, filtered, and concentrated to give a residue that was purified by preparative HPLC [column: YMC-Actus Triart C18, 250 × 21 mm; mobile phase: 10%–85% MeCN / HO (0.1% NH3·HO); flow rate: 15 mL / min; wavelength: 220 nm / 254 nm] and SFC (column: ChiralPak® IA, 250 × 21.3 mm ID, 5 μm, mobile phase: CO2 in phase A, MEOH+0.1% NH3·HO in phase B, gradient: B 40%, wavelength: 220 nm) to give (R)-2-(6-isopropylpyridin-3-yl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide (67.9 mg) as a white solid. LC / MS ESI(m / z):406(M+H) + . 1 H-NMR(400MHz,CDCl3)δ8.87(s,1H);8.50(s,1H);8.09(s,1H);7.81-7.74(m,1H);7.56(s,1H);7.27(s,1H),6.98-6.90 (m,1H);5.29-5.22(m,1H);5.11-5.02(m,2H);3.60(s,2H);3.27-3.17(m,1H);1.50(d,J=6.8Hz,3H);1.34-1.32(m,6H). 19 F-NMR (377MHz, CDCl3)δ -70.79. Example 21 (R)-2-(6-(1-(difluoromethyl)cyclopropyl)pyridin-3-yl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide

[0380] Step 1: 1-(5-bromopyridin-2-yl)cyclopropane-1-carbonitrile [ka]

[0381] A solution of 2-(5-bromopyridin-2-yl)acetonitrile (5.0 g), 1,2-dibromoethane (2.2 mL), tetrabutylammonium bromide (7.9 mL), and CHCN (80 mL) was added to NaOH (10.1 g) in HO (20 mL). The mixture was stirred at 25 °C for 12 hours. The mixture was diluted with water and extracted with EtOAc. The combined extracts were dried, filtered, and concentrated to give a residue that was purified by silica gel chromatography. (PE:EA = 1:0 to 50:1) to give 1-(5-bromopyridin-2-yl)cyclopropane-1-carbonitrile (4.5 g) as a white solid. 1 H-NMR (400MHz, DMSO-d6) δ8.66-8.62(m,1H), 8.08(dd, J=8.4, 2.4Hz,1H), 7.51(dd, J=8.4, 0.6Hz,1H), 1.91-1.78(m,2H), 1.73-1.61(m,2H).

[0382] Step 2: 1-(5-bromopyridin-2-yl)cyclopropane-1-carbaldehyde [ka]

[0383] To a solution of 1-(5-bromopyridin-2-yl)cyclopropane-1-carbonitrile (4.5 g) in THF (80 mL) was added diisobutylaluminum hydride (1.0 M in THF, 40.3 mL) at 0° C., and the reaction mixture was stirred at 0° C. for 2 hours. The reaction was quenched with MeOH (20 mL) and 1N HCl (30 mL) and extracted with EtOAc. The combined extracts were dried, filtered, and concentrated to give 1-(5-bromopyridin-2-yl)cyclopropane-1-carbaldehyde (1.8). g) as a yellow oil. LC / MS ESI (m / z): 246 [M+H] + .

[0384] Step 3: 5-bromo-2-(1-(difluoromethyl)cyclopropyl)pyridine [ka]

[0385] To a solution of 1-(5-bromopyridin-2-yl)cyclopropane-1-carbaldehyde (1.8 g) in DCM (30 mL) was added DAST (3.2 mL). The mixture was stirred at 25 °C for 12 h. The reaction was then quenched with NaHCO3 solution at 0 °C and washed with water. The organic layer was dried, filtered, and concentrated to give a residue, which was purified by silica gel chromatography (PE: EtOAc = 1:0 to 50:1) to give 5-bromo-2-[1-(difluoromethyl)cyclopropyl]pyridine (700 mg) as a colorless oil. LC / MS ESI (m / z): 248 [M+H] + .

[0386] Step 4: 2-(6-(1-(difluoromethyl)cyclopropyl)pyridin-3-yl)ethyl acetate [ka]

[0387] A solution of [1-(difluoromethyl)cyclopropyl]pyridine (700 mg), potassium 1-ethyl-3-propanedioate (720 mg), allylpalladium chloride dimer (103.2 mg), BINAP (527.1 mg), and DMAP (344.7 mg) in mesitylene (10 mL) was filled with N three times and stirred at 140 °C for 1 h. The mixture was then stirred at 120 °C for an additional 12 h. The mixture was concentrated to give a residue, which was subjected to silica gel chromatography (PE: EtOAc = 1:0 to 20:1) to give ethyl 2-{6-[1-(difluoromethyl)cyclopropyl]pyridin-3-yl}acetate (430 mg) as a yellow oil. LC / MS ESI (m / z): 256 [M+H] + .

[0388] Step 5: 2-(6-(1-(difluoromethyl)cyclopropyl)pyridin-3-yl)acetic acid [ka]

[0389] To a solution of ethyl 2-{6-[1-(difluoromethyl)cyclopropyl]pyridin-3-yl}acetate (430 mg) in MeOH (1.0 mL) was added NaOH (1 M in HO, 4.0 mL) and stirred at 25° C. for 30 minutes. The mixture was then concentrated, diluted with water, and extracted with EtOAc. The aqueous phase was adjusted to pH 4 with 1N aqueous HCl, followed by lyophilization to give 2-(6-(1-(difluoromethyl)cyclopropyl)pyridin-3-yl)acetic acid (230 mg) as a white solid. LC / MS ESI (m / z): 226 [MH] - .

[0390] Step 6: ((R)-2-(6-(1-(difluoromethyl)cyclopropyl)pyridin-3-yl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H)-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide [ka]

[0391] A solution of (methyl)cyclopropyl)pyridin-3-yl)acetic acid (130 mg), (R)-1-[1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl]ethan-1-amine hydrochloride (153.7 mg), HATU (239.3 mg), and DIEA (0.284 mL) in DMF (5 mL) was stirred for 12 hours at 25° C. The mixture was diluted with water and extracted with DCM. The combined extracts were dried, filtered, and concentrated to give a residue, which was purified by preparative HPLC [column: Shimpak GISTC18250*20 mm; mobile phase: 10%-95% MeCN and HO (0.1% NHHO); flow rate: 15 mL / min; wavelength: 220 nm / 254 nm] to give (R)-2-(6-(1-(difluoromethyl)cyclopropyl)pyridin-3-yl)-N-(1-(1-(2,2,2-trifluoroethyl))-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide (89.4 mg) as a white solid. LC / MS ESI(m / z): 454 (M+H)+; 1 H-NMR(400MHz,CDCl3)δ8.90(s,1H), 8.43(s,1H), 8.11(s,1H), 7.62(dd, J=8.2, 2.3Hz,1H), 7.58(s,1H), 7.32(d, J=8.0Hz,1H), 6.82(d , J=7.4Hz,1H), 6.18(t,J=57.1Hz,1H), 5.27-5.24(m,1H), 5.06(q, J=8.3Hz,2H), 3.55(s,2H), 1.51(d, J=6.8Hz,3H), 1.40~1.11(m,4H); 19 F-NMR (377MHz, CDCl3) δ -70.80, -119.76. Example 22 (R)-2-(4-(1-(difluoromethyl)cyclopropyl)phenyl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide

[0392] Synthesis of 1-(4-bromophenyl)cyclopropane-1-carbonitrile [ka]

[0393] To a 250 mL round-bottom flask containing 1,2-dibromoethane (6.17 mL) and a toluene solution (40 mL) of 2-(4-bromophenyl)acetonitrile (10 g), 50% aqueous NaOH (40 mL) and tetrabutylammonium bromide (3.17 mL) were added at room temperature. The reaction mixture was vigorously stirred at room temperature for 24 hours. The reaction mixture was then poured into 450 mL of ice-water solution, and this solution was extracted with EA (130 mL × 3). The combined extracts were washed with water (150 mL × 2), brine (150 mL), and finally dried over anhydrous Na2SO4. The solvent was removed under vacuum to give the crude product, which was purified by silica gel flash chromatography to give 1-(4-bromophenyl)cyclopropane-1-carbonitrile as a yellow oil (7.75 g). 1 H-NMR (400MHz, CDCl3) δ7.51-7.44(m,2H);7.21-7.11(m,2H);1.81-1.67(m,2H);1.45-1.31(m,2H)ppm.

[0394] Synthesis of 1-(4-bromophenyl)cyclopropane-1-carbaldehyde [ka]

[0395] A mixture of 1-(4-bromophenyl)cyclopropane-1-carbonitrile (4.0 g) and diisobutylaluminum hydride (36.02 mL) in THF (4.0 mL) was stirred at 0° C. for 1 h, and the reaction mixture was allowed to warm to room temperature. Upon completion of the reaction, the reaction mixture was cooled to 0° C. and quenched with MeOH. The reaction was allowed to warm slowly to room temperature over 15 min, and then 1 M HCl was added. The reaction was extracted with EA and washed with NaCl solution. The organic phase was concentrated to give 1-(4-bromophenyl)cyclopropane-1-carbaldehyde as a yellow oil (3.79 g, crude). 1 H-NMR (400MHz, CDCl3) δ9.16(s,1H);7.51-7.46(m,2H);7.20-7.15(m,2H);1.60-1.56(m,2H);1.41-1.37(m,2H)ppm.

[0396] Synthesis of 1-bromo-4-(1-(difluoromethyl)cyclopropyl)benzene [ka]

[0397] To a solution of 1-(4-bromophenyl)cyclopropane-1-carbaldehyde (3.79 g) in DCM (40 mL) was added DAST (22 g), and the reaction was stirred at room temperature overnight. The reaction was diluted with DCM and water. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over Na2SO4, filtered, and concentrated to give a residue that was purified by silica gel column chromatography (PE:EA = 10:1) to give 1-bromo-4-[1-(difluoromethyl)cyclopropyl]benzene as a yellow oil (2.2 g). 1 H-NMR (400MHz, CDCl3) δ7.56-7.35(m,2H);7.29-7.26(m,2H);5.58(t,J=56.0Hz,1H);1.21-1.07(m,2H);0.99-0.86(m,2H)ppm. 19 F-NMR (377MHz, CDCl3)δ -117.06ppm.

[0398] Synthesis of ethyl 2-(4-(1-(difluoromethyl)cyclopropyl)phenyl)acetate [ka]

[0399] To a solution of 1-bromo-4-[1-(difluoromethyl)cyclopropyl]benzene (636 mg) in mesitylene (10 mL) was added diallylpalladium dichloride (18.8 mg), BINAP (96.2 mg), DMAP (31.4 mg), and potassium ethyl malonate (657.2 mg). The reaction mixture was filled with N2 three times and stirred at 120 °C overnight. The reaction mixture was diluted with EA and water. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over Na2SO4, filtered, and concentrated to give a residue that was purified by silica gel column chromatography (PE:EA = 10:1) to give ethyl 2-(4-(1-(difluoromethyl)cyclopropyl)phenyl)acetate (267 mg) as a yellow oil. 1 H-NMR(400MHz,CDCl3)δ7.36(d,J=8.0Hz,2H);7.26-7.20(m,2H);5.64(t,J=56.0Hz,1H);4.2 0-4.10(m,2H);3.60(s,2H);1.26(t,J=4.0Hz,3H);1.16-1.12(m,2H);0.98-0.92(m,2H)ppm. 19 F-NMR (377MHz, CDCl3)δ -117.51ppm.

[0400] Synthesis of 2-(4-(1-(difluoromethyl)cyclopropyl)phenyl)acetic acid [ka]

[0401] In a 25 mL round-bottom flask, ethyl 2-(4-(1-(difluoromethyl)cyclopropyl)phenyl)acetate (66 mg) was mixed with 1 M aqueous NaOH (2.0 mL) and MeOH (2.0 mL) to give a colorless solution. The reaction mixture was stirred at room temperature for 1 h. Then, water (20 mL) was added, and the aqueous layer was washed with EA. The aqueous layer was acidified to pH < 3 with 1 M aqueous HCl, and the mixture was extracted with EA. The combined EtOAc extracts from the acidic aqueous layer were washed with saturated NaCl, dried over Na2SO4, filtered, and concentrated to give 2-(4-(1-(difluoromethyl)cyclopropyl)phenyl)acetic acid as a white solid (244 mg, crude). 1 H-NMR (400MHz, CDCl3) δ7.37(d, J=8.0Hz,2H);7.25(d,J=8.0Hz,2H);5.63(t,J=56.0Hz,1H);3.64(s,2H);1.16-1.11(m,2H);0.97-0.92(m,2H)ppm. 19 F-NMR (377MHz, CDCl3)δ -117.43ppm.

[0402] Synthesis of (R)-2-(4-(1-(difluoromethyl)cyclopropyl)phenyl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-indazol-5-ylethyl)acetamide [ka]

[0403] A solution of 2-(4-(1-difluoromethyl)cyclopropyl)phenyl)acetic acid (164 mg) and HATU (413.5 mg) in DMF (5.0 mL) was stirred at room temperature for 15 minutes. Then, (R)-1-(1-(2,2,2-trifluoroethyl)-1H-indazol-5-yl)ethan-1-amine hydrochloride (230.2 mg) and DIEA (0.7 × 2 mL) were added. The reaction mixture was stirred for 1 hour, and LC / MS showed the reaction was complete. The reaction was diluted with EA and water. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic phases were dried over Na2SO4, filtered, and concentrated to give a residue, which was purified by preparative HPLC. Purification by YMC-Actus Triart C18, 250 x 20 mm; mobile phase: 25% to 95% MeCN / HO (0.1% FA); flow rate: 15 mL / min; wavelength: 220 nm / 254 nm gave (R)-2-(4-(1-(difluoromethyl)cyclopropyl)phenyl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-indazol-5-yl)ethyl)acetamide (160 mg) as a tan solid. LC / MS ESI (m / z): 453 (M+H). + ;1H-NMR(400MHz,CDCl3):δ8.88(s,1H);8.11(s,1H);7.57(s,1H);7.36(d, J=8.1Hz,2H);7.23(d, J=8.2Hz,2H);6.75(s,1H);5.63(t,J=57. 3Hz,1H);5.26-5.23(m,1H);5.04(q,J=8.3Hz,2H);3.57(s,2H);1.49(d,J=6.8Hz,3H);1.14(dd,J=6.3Hz,4.7Hz,2H);0.95(d,J=2.1Hz,2H). 19 F-NMR (377MHz, CDCl3)δ -70.81, -117.15. Example 23 (R)-2-(6-cyclopropylpyridin-3-yl)-N-(1-(1-(4-fluorophenyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide

[0404] Step 1: Synthesis of 5-bromo-1-(4-fluorophenyl)-1H-pyrazolo[3,4-c]pyridine [ka]

[0405] A suspension of 5-bromo-1H-pyrazolo[3,4-c]pyridine (4.0 g), 4-fluoro-1-iodobenzene (6.95 g), 2-isobutyrylcyclohexanone (1.69 mL), CuI (0.77 g), and CsCO (12.5 g) in 30 mL was degassed with nitrogen for 30 minutes. The reaction mixture was then heated to 100 °C and stirred overnight. The reaction mixture was then cooled and diluted with DCM. The organic phase was washed with 1N aqueous NaOH and water, dried over NaSO, and concentrated to dryness. The solid residue was suspended in ether, stirred for 30 minutes, and filtered to collect the desired product, 5-bromo-1-(4-fluorophenyl)-1H-pyrazolo[3,4-c]pyridine, as a white solid (1.66 g). LC / MS ESI (m / z): 292 (M+H). + . 1 H-NMR (400MHz, CDCl3) δ8.93(s,1H);8.24-8.14(m,1H);7.91(d,J=4.0Hz,1H);7.76-7.63(m,2H);7.33-7.27(m,2H)ppm. 19 F-NMR(377MHz, CDCl3)δ -113.02.

[0406] Step 2: Synthesis of 1-(4-fluorophenyl)-5-vinyl-1H-pyrazolo[3,4-c]pyridine [ka]

[0407] To a solution of 5-bromo-1-(4-fluorophenyl)-1H-pyrazolo[3,4-c]pyridine (2.24 g) in 5:1 dioxane / water (100 mL) was added 2-ethenyl-4,4,5,5-tetramethyl-1,3,2-dioxaborolane (1.95 mL), KPO (4.07 g), and Pd(dppf)Cl (0.56 g). The reaction mixture was purged with N (three times) and stirred at 100 °C overnight. The reaction mixture was diluted with EA and water. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined extracts were dried over Na2SO4, filtered, and concentrated to give a residue which was purified using silica gel column chromatography eluting with 10% ethyl acetate in petroleum ether to give 1-(4-fluorophenyl)-5-vinyl-1H-pyrazolo[3,4-c]pyridine (1.21 g). LC / MS ESI(m / z): 240(M+H)+. 1 H-NMR:(400MHz,CDCl3)δ9.17(s,1H);8.26(d,J=10.0Hz,1H);7.76-7.65(m,3H);7.3 2-7.27(m,2H);7.03-6.88(m,1H);6.31(d,J=16.0Hz,1H);5.50(d,J=12.0Hz,1H)ppm.

[0408] Step 3: Synthesis of 1-(4-fluorophenyl)-1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde [ka]

[0409] To a solution of 1-(4-fluorophenyl)-5-vinyl-1H-pyrazolo[3,4-c]pyridine (1.2 g) in THF (80 mL) and HO (10 mL) was added sodium periodate (6.4 g) and potassium osmate dihydrate (0.18 g). The reaction mixture was stirred at room temperature for 0.5 h and then diluted with EA and water. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over NaSO, filtered, and concentrated to give a residue that was purified using silica gel column chromatography eluting with 14% ethyl acetate in petroleum ether to give 1-(4-fluorophenyl)-1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde (701 mg). LC / MS ESI (m / z): 242 (M+H). + .

[0410] Step 4: Synthesis of (R,E)—N-((1-(4-fluorophenyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)methylene)-2-methylpropane-2-sulfinamide [ka]

[0411] A solution of 1-(4-fluorophenyl)-1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde (700 mg), (R)-(+)-2-methyl-2-propanesulfinamide (457 mg), and CuSO (1.4 g) in DCM (20 mL) was stirred at room temperature overnight. LC / MS showed the reaction was complete. The reaction mixture was diluted with DCM and water. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over NaSO, filtered, and concentrated to give a residue. Purification by silica gel chromatography (PE:EA = 5:1) gave (R,E)-N-((1-(4-fluorophenyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)methylene)-2-methylpropane-2-sulfinamide (811 mg). LC / MS ESI(m / z):345(M+H) + .

[0412] Step 5: (R)—N—((R)-1-(1-(4-fluorophenyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)-2-methylpropane-2-sulfinamide [ka]

[0413] To (R,E)-N-((1-(4-fluorophenyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)methylene)-2-methylpropane-2-sulfinamide (300 mg) in THF (10 mL) was added CHMgBr (4.1 mL, 3.0 M in EtO). The reaction mixture was stirred at -78 °C for 2 hours. The reaction mixture was diluted with DCM and water. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over NaSO, filtered, and concentrated to give a residue, which was purified by silica gel chromatography (DCM:MeOH = 15:1) to give (R)-N-((R)-1-(1-(4-fluorophenyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)-2-methylpropane-2-sulfinamide as a yellow solid (602 mg). LC / MS ESI(m / z):361(M+H) + .

[0414] Step 6: Synthesis of (R)-1-(1-(4-fluorophenyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethan-1-amine hydrochloride [ka]

[0415] To (R)-N-((R)-1-(1-(4-fluorophenyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)-2-methylpropane-2-sulfinamide (602 mg) in dioxane (10 mL) was added 4N HCl-dioxane (5.0 mL), and the reaction mixture was stirred at room temperature for 30 minutes. The reaction mixture was evaporated in vacuo. The resulting solid was filtered, washed with EtOAc, and then dried under vacuum to give (R)-1-(1-(4-fluorophenyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethan-1-amine hydrochloride as a white solid (545 mg, crude). LC / MS ESI (m / z): 257 (M+H). + .

[0416] Step 7: Synthesis of (R)-2-(6-cyclopropylpyridin-3-yl)-N-(1-(1-(4-fluorophenyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide [ka]

[0417] A solution of 2-(6-cyclopropylpyridin-3-yl)acetic acid (250 mg, crude) and HATU (321.9 mg) in DMF (5.0 mL) was stirred at room temperature for 15 minutes. Then, (R)-1-(1-(4-fluorophenyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethan-1-amine hydrochloride (205.4 mg, crude) and DIEA (0.56 mL) were added until the pH of the solution was higher than 7, as measured by pH paper. After stirring for 1 hour, LC / MS showed the reaction was complete. The reaction was diluted with EA and water, and the two phases were separated. The aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over NaSO, filtered, and concentrated to give a residue. The residue was precipitated in CHCN and washed with CHCN to give 149.8 mg of the product as a white solid. LC / MS ESI(m / z):416(M+H) + . 1H-NMR(400MHz,CDCl3)δ9.07(s,1H);8.34(s,1H);8.20(s,1H);7.74-7.67(m,2H);7.58-7.50(m,2H);7.33-7.26(m,2H);7.10(d, J=8.0Hz,1H);6.74(d,J=7.0Hz,1H);5.35-5.17(m,1H);3.53(s,2H);2.11-2.02(m,1H);1.49(d,J=6.8Hz,3H);1.04-0.97(m,4H). 19 F-NMR(377MHz, CDCl3)δ -113.66. Example 24 (R)-2-(4-cyclopropylphenyl)-N-(1-(3-(2,2,2-trifluoroethyl)-3H-[1,2,3]triazolo[4,5-c]pyridin-6-yl)ethyl)acetamide

[0418] Synthesis of 2-chloro-5-fluoropyridine 1-oxide [ka]

[0419] To a solution of 2-chloro-5-fluoropyridine (19.28 mL) in 30% H2O2 (96 mL) was added TFA (175 mL), and the reaction mixture was stirred at 70 °C overnight. The reaction mixture was concentrated in vacuo and diluted with toluene. The reaction solution was then concentrated in vacuo again and diluted with DCM and water. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL x 3). The combined organic layers were dried over Na2SO4, filtered, and concentrated to give 2-chloro-5-fluoropyridine 1-oxide (26.2 g) as a crude product. LC / MS ESI (m / z): 148 [M+H] + . 1 H-NMR (400MHz, CDCl3) δ8.35-8.28(m,1H); 7.52-7.43(m,1H); 7.11-7.01(m,1H)ppm.

[0420] Synthesis of 2-chloro-5-fluoro-4-nitropyridine 1-oxide [ka]

[0421] To a solution of 2-chloro-5-fluoropyridine 1-oxide (10 g) in H2SO4 (102 mL) was added KNO3 (27.4 g) slowly at room temperature. The reaction mixture was stirred under nitrogen at 110 °C for 16 hours. The reaction mixture was poured onto ice at 0 °C and neutralized by the dropwise addition of 30% ammonium hydroxide, using an ice bath to maintain the temperature below 15 °C. The precipitated pale yellow crystals were collected by filtration. The precipitate was then purified by flash chromatography on silica gel to give 2-chloro-5-fluoro-4-nitropyridine 1-oxide (1.6 g). LC / MS ESI (m / z): 193 [M+H] + . 1 H-NMR (400MHz, CDCl3) δ8.41(d,J=5.7Hz,1H);8.31(d,J=8.4Hz,1H).

[0422] Synthesis of 2-chloro-4-nitro-5-((2,2,2-trifluoroethyl)amino)pyridine 1-oxide [ka]

[0423] To a solution of 2-chloro-5-fluoro-4-nitropyridine 1-oxide (1.89 g) in THF (24 mL) was added 2,2,2-trifluoroethan-1-amine (1.568 mL), and the reaction mixture was stirred at room temperature overnight. The reaction mixture was then diluted with DCM and water. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over Na2SO4, filtered, and concentrated to give a residue that was further concentrated under high vacuum to give 2-chloro-4-nitro-5-((2,2,2-trifluoroethyl)amino)pyridine 1-oxide (2.1 g). LC / MS ESI (m / z): 272 [M+H] + . 1 H-NMR (400MHz, CDCl3): δ8.33 (s, 1H); 8.16 (s, 1H); 7.98 (brs, 1H); 3.98-3.87 (m, 2H).

[0424] 6-Chloro-N 3Synthesis of -(2,2,2-trifluoroethyl)pyridine-3,4-diamine [ka]

[0425] To a solution of 2-chloro-4-nitro-5-((2,2,2-trifluoroethyl)amino)pyridine 1-oxide (2.0 g) in glacial acetic acid (21 mL) was added Fe (0.209 mL) slowly at 0 °C. The reaction mixture was stirred at room temperature for 1 h and diluted with EA and water. The two phases were separated and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over Na2SO4, filtered and concentrated to give a residue which was concentrated in vacuo to give 6-chloro-N 3 -(2,2,2-trifluoroethyl)pyridine-3,4-diamine (2.28 g, crude) was obtained. LC / MS ESI (m / z): 226 [M+H] + .

[0426] Synthesis of 6-chloro-3-(2,2,2-trifluoroethyl)-3H-[1,2,3]triazolo[4,5-c]pyridine [ka]

[0427] 6-Chloro-N in 30 mL of water 3 To a solution of -(2,2,2-trifluoroethyl)pyridine-3,4-diamine (2.67 g) and HSO (1.8 mL) under nitrogen at 0 °C, NaNO (1.63 g) was slowly added. The reaction mixture was stirred at 0 °C for 4 hours, and then the reaction mixture was neutralized to pH 8 with saturated NaHCO. The resulting solid was collected and purified by silica gel column to give 6-chloro-3-(2,2,2-trifluoroethyl)-3H-[1,2,3]triazolo[4,5-c]pyridine (872 mg). LC / MS ESI (m / z): 237 [M+H] + . 1 H-NMR (400MHz, CDCl3) δ8.96 (s, 1H); 8.06 (s, 1H); 5.38-5.32 (m, 2H). 19F-NMR (377MHz, CDCl3)δ -70.30(s).

[0428] Synthesis of 3-(2,2,2-trifluoroethyl)-6-vinyl-3H-[1,2,3]triazolo[4,5-c]pyridine [ka]

[0429] To a solution of 6-chloro-3-(2,2,2-trifluoroethyl)-3H-[1,2,3]triazolo[4,5-c]pyridine (872 mg) in dioxane-water (25 mL) was added vinylboronic acid pinacol ester (149.55 mg), KPO (1.9 g), and Pd(dcpf)Cl (276.38 mg). The reaction mixture was flushed with N and stirred overnight at 100 °C under nitrogen. The reaction was diluted with EA and water and filtered through Celite. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over NaSO, filtered, and concentrated to give a residue that was purified by silica gel column chromatography to give 3-(2,2,2-trifluoroethyl)-6-vinyl-3H-[1,2,3]triazolo[4,5-c]pyridine (807 mg). LC / MS ESI(m / z):229[M+H] + . 1 H-NMR (400MHz, CDCl3) δ9.12(s,1H);7.93(d,J=0.9Hz,1H);7.01-6.94(m,1H);6.38-6.33(m,1H);5.57-5.54(s,1H);5.38-5.32(s,2H).

[0430] Synthesis of 3-(2,2,2-trifluoroethyl)-3H-[1,2,3]triazolo[4,5-c]pyridine-6-carbaldehyde [ka]

[0431] To a solution of 3-(2,2,2-trifluoroethyl)-6-vinyl-3H-[1,2,3]triazolo[4,5-c]pyridine (802 mg) in THF-water (8:1, 30 mL) was added NaIO (4.5 g) and potassium osmate (1.3 g), and the reaction mixture was stirred at room temperature for 1 hour. The reaction mixture was filtered through Celite and diluted with DCM and water. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over NaSO, filtered, and concentrated to give a residue that was purified by silica gel column chromatography to give 3-(2,2,2-trifluoroethyl)-3H-[1,2,3]triazolo[4,5-c]pyridine-6-carbaldehyde (584 mg). LC / MS ESI (m / z): 231 [M+H] + . 1 H-NMR (400MHz, CDCl3) δ10.29(s,1H);9.29(s,1H);8.72(s,1H);5.46-5.40(m,2H).

[0432] Synthesis of (R,E)-2-methyl-N-((3-(2,2,2-trifluoroethyl)-3H-[1,2,3]triazolo[4,5-c]pyridin-6-ylmethylene)propane-2-sulfinamide [ka]

[0433] To a solution of 3-(2,2,2-trifluoroethyl)-3H-[1,2,3]triazolo[4,5-c]pyridine-6-carbaldehyde (584 mg) in anhydrous DCM (15 mL) was added CuSO4 (1.2 g) and (R)-2-methylpropane-2-sulfinamide (399.8 mg). The reaction was stirred at room temperature overnight. The solution was filtered through a pad of Celite, and the filtrate was concentrated. The residue was purified on a silica gel column to give (R,E)-2-methyl-N-((3-(2,2,2-trifluoroethyl)-3H-[1,2,3]triazolo[4,5]-c]pyridin-6-yl)methylene)propane-2-sulfinamide (840 mg). LC / MS ESI (m / z): 334 [M+H] + .1 H-NMR (400MHz, CDCl3) δ9.26(s,1H);8.90(s,1H);8.74(d,J=0.8Hz,1H);5.45-5.36(m,2H);1.32(s,9H).

[0434] Synthesis of (R)-2-methyl-N-((R)-1-(3-(2,2,2-trifluoroethyl)-3H-[1,2,3]triazolo[4,5-c]pyridin-6-yl)ethyl)propane-2-sulfinamide [ka]

[0435] To a solution of (R,E)-2-methyl-N-((3-(2,2,2-trifluoroethyl)-3H-[1,2,3]triazolo[4,5-c]pyridin-6-yl)methylene)propane-2-sulfinamide in anhydrous THF (26 mL). MeMgBr (4.20 mL, 3.0 M in ether) was added dropwise at −78° C. under nitrogen. The reaction mixture was stirred at −78° C. for 2 h. The reaction solution was then quenched with aqueous NH4Cl and extracted with EA. The combined extracts were separated, washed with brine, and concentrated in vacuo. The residue was then purified by silica gel column chromatography to give (R)-2-methyl-N-((R)-1-(3-(2,2,2-trifluoroethyl)-3H-[1,2,3]triazolo[4,5-c]pyridin-6-yl)ethyl)propane-2-sulfinamide (690 mg). LC / MS ESI (m / z): 350 [M+H] + . 1 H-NMR(400MHz,CDCl3)δ9.12(s,1H);7.98(d,J=0.7Hz,1H);5.43-5.31(m,2H) ;4.80-4.72(m,1H);4.51(d,J=6.7Hz,1H);1.61(d,J=6.7Hz,3H);1.25(s,9H).

[0436] Synthesis of (R)-1-(3-(2,2,2-trifluoroethyl)-3H-[1,2,3]triazolo[4,5-c]pyridin-6-yl)ethan-1-amine hydrochloride [ka]

[0437] To a solution of (R)-2-methyl-N-((R)-1-(3-(2,2,2-trifluoroethyl)-3H-[1,2,3]triazolo[4,5-c]pyridin-6-yl)ethyl)propane-2-sulfinamide (680 mg) in dioxane (6.0 mL) was added 4N HCl-dioxane (2.0 mL), and the reaction mixture was stirred at room temperature for 1 hour. The mixture was then concentrated in vacuo and used directly in the next step. LC / MS ESI (m / z): 246 [M+H] + .

[0438] Synthesis of (R)-2-(4-cyclopropylphenyl)-N-(1-(3-(2,2,2-trifluoroethyl)-3H-[1,2,3]triazolo[4,5-c]pyridin-6-yl)ethyl)acetamide (N201201-361) [ka]

[0439] To a solution of (R)-1-[3-(2,2,2-trifluoroethyl)-3H-[1,2,3]triazolo[4,5-c]pyridin-6-yl]ethan-1-amine hydrochloride (150 mg) in anhydrous DMF, DIEA (0.607 mL) was added to give solution A. To a solution of 2-(4-cyclopropylphenyl)acetic acid (129.3 mg) in DMF (1 mL), HATU (255.85 mg) was added and stirred at room temperature for 15 minutes to give solution B. Solution B was added to solution A, and the mixture was stirred at room temperature for 1 hour. The reaction mixture was diluted with EA and water. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over Na2SO4, filtered, and concentrated to give a residue, which was purified by preparative HPLC [column: YMC-Actus Triart C18, 250 × 21 mm; mobile phase: 20%–95% MeCN / H2O (1% NH3·H2O); flow rate: 15 mL / min; wavelength: 220 nm / 254 nm] and SFC (column: ChiralPakTM Purification with ®IA, 250 x 21.3 mm ID, 5 μm; mobile phase: CO A and MEOH + 0.1% NH H O B; gradient: B 35%; wavelength: 220 nm) gave (R)-2-(4-cyclopropylphenyl)-N-(1-(3-(2,2,2-trifluoroethyl)-3H-[1,2,3]triazolo[4,5-c]]pyridin-6-yl)ethyl)acetamide (89.3 mg). LC / MS ESI (m / z): 404 [M+H] + ; 1 H-NMR(400MHz,CDCl3):δ9.06(s,1H);7.90(s,1H);7.16(d,J=8.1Hz,2H);7.05(d,J=8.1Hz,2H);6.48(d,J=7.6Hz,1H );5.37-5.31(m,3H);3.56(s,2H);1.92-1.86(m,1H);1.48(d,J=6.9Hz,3H);0.98-0.96(m,2H);0.70-0.68(m,2H)ppm. 19 F-NMR (377MHz, CDCl3)δ -70.30(s). Example 25 (R)-2-(6-cyclopropylpyridin-3-yl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide

[0440] Synthesis of ethyl 2-(6-cyclopropylpyridin-3-yl)acetate [ka]

[0441] To a solution of ethyl 2-(6-chloropyridin-3-yl)acetate (1.5 g), cyclopropylboronic acid (1.94 g), and potassium carbonate (1.04 g) in dioxane (40 mL) was added Pd(PPh3)4 (0.87 g), and the mixture was filled with N2 three times and stirred at 110 °C overnight. After cooling to room temperature, the reaction was diluted with EA (30 mL) and water (40 mL), and the two layers were separated. The aqueous layer was then extracted with EA (30 mL). The combined organic layers were washed with water and brine (40 mL each), dried over Na2SO4, filtered, and concentrated. The crude product was purified by silica gel chromatography (5-18% EA / PE) to give ethyl 2-(6-cyclopropylpyridin-3-yl)acetate (1.21 g) as a colorless oil. LC / MS ESI (m / z): 192 (M+H). + . 1 H-NMR(400MHz,CDCl3)δ8.33(d,J=4.0Hz,1H);7.51-7.48(m,1H);7.09(d,J= 8.0Hz,1H);3.70(s,3H);3.57(s,2H);2.06-2.01(m,1H);1.05-0.92(m,4H).

[0442] Synthesis of 2-(6-cyclopropylpyridin-3-yl)acetic acid [ka]

[0443] To a 250 mL round-bottom flask was added methyl 2-(6-cyclopropylpyridin-3-yl)acetate (1.2 g) and 50 mL of MeOH. Aqueous NaOH (1 N, 50 mL) was added to the mixture to give a colorless solution. The reaction mixture was stirred at room temperature for 1 hour, and the aqueous layer was extracted with EA. The aqueous layer was acidified with 1 N aqueous HCl to pH < 3. The aqueous phase was then extracted with EA. The EtOAc extract from the acidified aqueous phase was washed with saturated NaCl solution, filtered, and concentrated to give 2-(6-cyclopropylpyridin-3-yl)acetic acid as a white solid (2.31 g, crude).

[0444] Synthesis of (R)-2-(6-cyclopropylpyridin-3-yl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide [ka]

[0445] 2-(6-Cyclopropylpyridin-3-yl)acetic acid (250 mg, crude) and HATU (321.9 mg) in DMF (5.0 mL) were stirred at room temperature for 15 minutes. Next, (R)-1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethan-1-amine hydrochloride (179.2 mg) and DIEA (0.56 mL) were added. The mixture was stirred for 1 hour, and LC / MS showed the reaction was complete. The reaction was diluted with EA and water. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 2). The combined organic layers were dried over NaSO, filtered, and concentrated in vacuo to give a residue that was purified by preparative HPLC [Column: YMC-Actus Triart C18, 250 × 20 mm; Mobile phase: 10%-95% MeCN / HO (1% NH HO); Flow rate: 15 mL / min; Wavelength: 220 nm / 254 nm] to give (R)-2-(6-cyclopropylpyridin-3-yl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide (141.8 mg) as a white solid. LC / MS ESI (m / z): 404 (M+H). + . 1 H-NMR(400MHz,CDCl3):δ8.86(s,1H);8.38(s,1H);8.08(s,1H);7.64(s,1H);7.54(d,J=4.0Hz,1H);7.10(d,J=8.0Hz,1H);6 .84(s,1H);5.32-5.20(m,1H);5.14-4.95(m,2H);3.65-3.47(m,2H);2.17(s,1H);1.49(d,J=8.0Hz,3H);1.11-1.01(m,4H). 19 F-NMR(377MHz, CDCl3)δ -70.78. Example 26 (R)—N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)-2-(6-(1-(trifluoromethyl)cyclopropyl)pyridin-3-yl)acetamide

[0446] Synthesis of methyl 6-(3,3,3-trifluoroprop-1-en-2-yl)nicotinate [ka]

[0447] A mixture of methyl 6-bromonicotinate (5.47 g), 4,4,6-trimethyl-2-(3,3,3-trifluoroprop-1-en-2-yl)-1,3,2-dioxaborionane (7.87 g), PdCl(dppf) (1.85 g), and KCO (2 M aqueous solution, 26.8 mL) in acetonitrile (104 mL) was backfilled with N three times and heated at 80 °C for 90 min. LC / MS indicated the formation of the desired product. The reaction mixture was cooled to room temperature and partitioned between water and ethyl acetate. The two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over NaSO, filtered, and concentrated to give a residue, which was purified by silica gel column chromatography (0–20% ethyl acetate / PE). Methyl 6-(3,3,3-trifluoroprop-1-en-2-yl)nicotinate was obtained as a colorless oil (3.6 g). LC / MS ESI (m / z): 232 [M+H] + . 1 H-NMR (400MHz, CDCl3): δ9.22(d,J=2.4Hz,1H);8.75-8.74(m,1H);8.34-8.31(m,1H);8.25(d,J=1.2Hz,1H);7.58(d,J=11.6Hz,1H);3.97(s,3H). 19 F-NMR (376.48MHz, CDCl3δ -63.98ppm.

[0448] Synthesis of methyl 6-(1-(trifluoromethyl)cyclopropyl)nicotinate [ka]

[0449] To a suspension of methyl 6-(3,3,3-trifluoroprop-1-en-2-yl)nicotinate (2.47 g) and methyl diphenylsulfonium tetrafluoroborate (4.0 g) in anhydrous tetrahydrofuran (30 mL) at 0° C. under nitrogen, sodium bis(trimethylsilyl)amide (2 M in THF) was added. The reaction mixture was stirred at 0° C. for 10 minutes and then at room temperature for 1 hour. Methanol (250 μL) was added to quench the reaction. The crude mixture was concentrated in vacuo to give a residue that was purified by silica gel column chromatography (10% PE / EA) to give methyl 6-(1-(trifluoromethyl)cyclopropyl)nicotinate (1.3 mg) as a white solid. LC / MS ESI (m / z): 246 [M+H] + . 1 H-NMR (400MHz, CDCl3): δ9.09(d,J=2.4Hz,1H);8.26-8.23(m,1H);7.64(d,J=8.4Hz,1H);3.95(s,3H);1.54-1.53(m,2H);1.50-1.49(m,2H)ppm. 19 F-NMR (376.48MHz, CDCl3): δ -67ppm.

[0450] Synthesis of 6-(1-(trifluoromethyl)cyclopropyl)nicotinic acid [ka]

[0451] To a solution of methyl 6-(1-(trifluoromethyl)cyclopropyl)nicotinate (345 mg) in MeOH (20 mL) was added 4.0 mL of NaOH (2.0 M). The mixture was stirred at 65 °C for 2 h, and TLC showed the reaction was complete. The solvent was concentrated in vacuo, and the reaction mixture was adjusted to pH 2-3 with 1 N HCl. The reaction mixture was extracted with EA (50 mL), washed with brine, dried over anhydrous Na2SO4, filtered, and concentrated to give 6-(1-(trifluoromethyl)cyclopropyl)nicotinic acid as a white solid (300 mg). LC / MS ESI (m / z): 232 [M+H] + .

[0452] Synthesis of 2-diazo-1-(6-(1-(trifluoromethyl)cyclopropyl)pyridin-3-yl)ethanone [ka]

[0453] 6-(1-(trifluoromethyl)cyclopropyl)nicotinic acid (300 mg) in DCM (10 mL) was cooled to 0 °C. Oxalyl chloride (1.1 mL) and DMF (2 drops) were added, and the resulting solution was stirred at room temperature for 2 h. The volatiles were then removed under vacuum. The residue was redissolved in DCM (10 mL) and cooled to 0 °C. TMSCHN2 (1.28 mL, 2 M solution in hexanes) and TEA (0.33 mL) were slowly added, and the resulting solution was maintained at 5 °C for 12 h. The reaction mixture was then filtered, and the filtrate was concentrated under reduced pressure to give 2-diazo-1-(6-(1-(trifluoromethyl)cyclopropyl)pyridin-3-yl)ethenone as a brown residue, which was used directly in the next step. LC / MS ESI (m / z): 256 [M+H] + .

[0454] Step 5: Synthesis of methyl 2-((6-(1-(trifluoromethyl)cyclopropyl)pyridin-3-yl)acetate [ka]

[0455] To a mixture of 2-diazo-1-(6-(1-(trifluoromethyl)cyclopropyl)pyridin-3-yl)ethanone (600 mg, crude) in methanol (20 mL) was added AgO (175 mg). The reaction mixture was stirred at 65 °C for 2 h. TLC showed that a trace amount of product had formed, so an additional 142 mg of AgO was added. The mixture was stirred at 65 °C for another 2 h. The reaction mixture was concentrated in vacuo, and the residue was purified by silica gel column chromatography (10% EA to 20% EA in PE) to give methyl 2-(6-(1-(trifluoromethyl)cyclopropyl)pyridin-3-yl)acetate as a yellow oil (100 mg). LC / MS ESI (m / z): 260 [M+H] + .

[0456] Step 6: Synthesis of 2-(6-(1-(trifluoromethyl)cyclopropyl)pyridin-3-yl)acetic acid [ka]

[0457] To a solution of methyl 2-((6-(1-(trifluoromethyl)cyclopropyl)pyridin-3-yl)acetate (100 mg) in MeOH (10 mL) was added 2.0 mL of NaOH (2.0 M). The mixture was stirred at 65 °C for 2 h. After completion, the reaction mixture was removed under reduced pressure and the pH of the reaction mixture was adjusted to 2-3 with 1 N HCl. The reaction mixture was extracted with EA (50 mL × 3). The combined extracts were washed with brine, dried over anhydrous Na2SO4, filtered, and concentrated to give 2-(6-(1-(trifluoromethyl)cyclopropyl)pyridin-3-yl)acetic acid as a white solid (70 mg). LC / MS ESI (m / z): 246 [M+H] + . 1 H-NMR (400MHz, CDCl3): δ8.43(d, J=1.6Hz,1H); 7.78-7.75(m,1H); 7.59-7.56(m,1H); 3.69(s,2H); 1.43-1.40(m,2H); 1.32-1.29(m,2H)ppm. 19F-NMR (CDC13, -376.48): δ -69.78ppm.

[0458] Step 7: Synthesis of (R)—N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)-2-(6-(1-(trifluoromethyl)cyclopropyl)pyridin-3-yl)acetamide [ka]

[0459] To a mixture of 2-(6-(1-(trifluoromethyl)cyclopropyl)pyridin-3-yl)acetic acid (125.0 mg) in DMF (1.0 mL), HATU (213 mg) was added, and the mixture was stirred at room temperature for 5 minutes to give solution A. To another round-bottom flask containing (R)-1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethanamine hydrochloride (186 mg), DMF (1.0 mL) and DIEA (197 mg) were added, and the mixture was stirred for 1 minute (solution B). Solution B was added to solution A, and the reaction mixture was stirred at room temperature for 2 hours. EA (50 mL) and water (40 mL) were added, and the organic layer was washed with water and brine, dried over Na2SO4, filtered, and concentrated. The residue was purified by preparative HPLC [column: YMC-Actus Triart C18, 250 x 21 mm; mobile phase: 10%-85% MeCN / HO (0.1% FA); flow rate: 15 mL / min; wavelength: 220 nm / 254 nm] to give a white solid (106.9 mg). LC / MS ESI (m / z): 472 (M+H). + ; 1 H-NMR(400MHz,CDCl3):δ8.90(s,1H);8.43(d,J=2.0Hz,1H);8.11(s,1H);7.64-7.62(m,1H);7.58(s,1H);7.52-7.49(m, 1H);6.84(d,J=7.6Hz,1H);5.30-5.23(m,1H);5.08-5.02(m,2H);3.56(s,2H);1.51(d,J=6.4Hz,3H);1.41-1.37(m,4H). 19F-NMR (377MHz, CDCl3): δ -67.70; -70.79ppm. Example 27 (R)-2-(4-cyclopropylphenyl)-N-(1-(1-(4-fluorophenyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide

[0460] Step 1: Synthesis of ethyl 2-(4-cyclopropylphenyl)acetate [ka]

[0461] To a solution of ethyl 4-bromophenylacetate (2.0 g), cyclopropylboronic acid (0.92 g), tricyclohexylphosphine (0.23 g), and KPO (6.11 g) in toluene / water (40 mL, 20:1) was added palladium(II) acetate (0.09 g). The reaction mixture was purged with N and stirred at 100 °C overnight. After cooling to room temperature, the reaction was diluted with EA and water, and the two layers were separated. The aqueous layer was extracted with EA (20 mL). The combined organic layers were washed with water and brine (40 mL each), dried over NaSO, filtered, and concentrated. The residue was purified by silica gel chromatography (5–18% EA / PE) to give ethyl 2-(4-cyclopropylphenyl)acetate (1.368 g) as a pale yellow oil. 1 H-NMR(400MHz,CDCl3)δ7.16(d,J=8.0Hz,1H);7.02(d,J=8.0Hz,1H);4.15(q,J=8.0Hz,2H); 3.56(s,2H);1.90-1.84(m,1H);1.24(t,J=8.0Hz,3H);0.95-0.92(m,2H);0.7-0.63(m,2H).

[0462] Step 2: Synthesis of 2-(4-cyclopropylphenyl)acetic acid [ka]

[0463] To a solution of ethyl 2-(4-cyclopropylphenyl)acetate in THF (20.0 mL) and MeOH (8.0 mL) was added 2.5 M aqueous LiOH (12 mL), and the resulting solution was stirred at room temperature for 2 h. After completion, the mixture was acidified with 1N aqueous HCl, and the aqueous phase was extracted with DCM. The combined organic layers were dried over Na2SO4, filtered, and concentrated to give 2-(4-cyclopropylphenyl)acetic acid (1.04 g) as a yellow solid, which was used in the next step.

[0464] Step 3: Synthesis of (R)-2-(4-cyclopropylphenyl)-N-(1-(1-(4-fluorophenyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide [ka]

[0465] A solution of 2-(4-cyclopropylphenyl)acetic acid (100 mg) and HATU (323.7 mg) in DMF (5.0 mL) was stirred at room temperature for 15 minutes. Then, (R)-1-(1-(4-fluorophenyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethan-1-amine hydrochloride (206.6 mg) and DIEA (0.56 mL) were added. The reaction mixture was stirred at room temperature for 1 hour, after which LC / MS showed that the reaction was complete. The reaction mixture was diluted with EA and water. The two phases were then separated. The aqueous phase was extracted with EA (10 mL × 3). The combined organic layers were dried over Na2SO4, filtered, and concentrated in vacuo. The residue was purified by preparative HPLC [column: YMC-Actus Triart C18, 250 × 20 mm; mobile phase: 20% to 80% MeCN / HO (0.1% FA); flow rate: 15 mL / min; wavelength: 220 nm / 254 nm] to obtain the compound (R)-2-(4-cyclopropylphenyl)-N-(1-(1-(4-fluorophenyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide (140 mg) as a white solid. LC / MS ESI (m / z): 415 (M+H). + . 1H-NMR(400MHz,CDCl3)δ9.06(s,1H);8.20(s,1H);7.76-7.65(m,2H);7.60(s,1H);7.32-7.26(m,2H);7.15(d,J=8.1Hz,2H);7.07-7.01(m ,2H);6.61(s,1H);5.29-5.24(m,1H);3.64-3.44(m,2H);1.93-1.82(m,1H);1.48(d,J=6.8Hz,3H);1.02-0.88(m,2H);0.75-0.61(m,2H). 19 F-NMR (377MHz, CDCl3)δ -113.57. Example 28 (R)-2-(4-cyclopropylphenyl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide

[0466] Synthesis of (R)-2-(4-cyclopropylphenyl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide [ka]

[0467] To a solution of (R)-1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethan-1-amine hydrochloride (151 mg) in anhydrous DMF (1 mL) was added DIEA (0.61 mL, 3.71 mmol) to give solution A. To a solution of 2-(4-cyclopropylphenyl)acetic acid (130.8 mg) in DMF, HATU (258.6 mg) was added and stirred at room temperature for 5 minutes to give solution B. Solution B was added to solution A and stirred at room temperature for 1 hour. The reaction was diluted with EA and water, the two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic layers were dried over Na2SO4, filtered, and concentrated to a residue that was purified by preparative HPLC [column: YMC-Actus Triart C18, 250 × 20 mm; Na2SO4; mobile phase: 20%–95% MeCN / HO (0.1% FA); flow rate: 15 mL / min; wavelength: 220 nm / 254 nm] and SFC (column: ChiralPak® IA, 250 × 21.3 mm ID, 5 μm; mobile phase: CO2 in phase A and MeOH + 0.1% NH3·HO in phase B; gradient: B 40%; wavelength: 220 nm) to give (R)-2-(4-cyclopropylphenyl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide as a white solid (97 mg). LC / MS ESI(m / z):403[M+H] + . 1 H-NMR(400MHz,CDCl3)δ8.86(s,1H);8.08(s,1H);7.53(s,1H);7.16-7.03(m,4H);6.53(d,J=7.6Hz,1H);5.30-5.23(m,1 H);5.06-5.00(m,2H);3.54-3.53(m,2H);1.93-1.86(m,1H);1.45(d,J=6.8Hz,3H);1.03-0.90(m,2H);0.69-0.68(m,2H). 19 F-NMR (377MHz, CDCl3)δ -70.81(s). Example 29 (R)-2-(4-(1-fluorocyclopropyl)phenyl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide

[0468] Synthesis of 1-(2-bromo-1-fluoroethyl)-4-methylbenzene [ka]

[0469] To an ice-cold solution of 1-ethenyl-4-methylbenzene (9.45 g) in DCM (80.0 mL) were added triethylamine trihydrofluoride (40 mL) and N-bromosuccinimide (17.0 g). After stirring at 0 °C for 30 minutes, the reaction mixture was warmed to room temperature and stirred overnight. The reaction mixture was poured into ice water (500 mL) and neutralized with aqueous NH4OH. The two phases were separated, and the aqueous phase was extracted with DCM (600 mL). The combined organic layers were washed with 0.1 M HCl (600 mL), 5% NaHCO3, and water (200 mL) and dried over Na2SO4. The organic layer was then filtered and concentrated in vacuo. The residue was purified on a silica gel column (5% EA in PE) to give 1-(2-bromo-1-fluoroethyl)-4-methylbenzene (13.69 g) as a colorless oil. 1 H-NMR (400MHz, CDCl3) δ7.26(d,J=8.0,2H);7.16(d,J=8.0Hz,2H);5.66-5.63(m,1H);5.54-5.51(m,1H));3.70-3.61(m,3H);2.35-2.32(m,1H). 19 F-NMR (377MHz, CDCl3)δ -172.35(s).

[0470] Synthesis of 1-(1-fluorovinyl)-4-methylbenzene [ka]

[0471] 1-(2-Bromo-1-fluoroethyl)-4-methylbenzene (2.0 g) was dissolved in pentane (60 mL) and tBuOK (2.09 g) was added slowly at 0 °C. The reaction mixture was stirred under reflux for 1 hour. After cooling to room temperature, the mixture was poured into ice water (50 mL). After phase separation, the aqueous phase was extracted with pentane (300 mL). The combined organic layers were washed with 5% NaHCO (50 mL), 0.05 M HCl (25 mL), and water (50 mL). The organic phase was dried (Na SO ) and filtered. The filtrate was concentrated in vacuo to give a residue, which was purified on a silica gel column (5% EA in PE) to give 1-(1-fluorovinyl)-4-methylbenzene (711 mg) as a colorless liquid. 1 H-NMR:(400MHz,DMSO-d6)δ7.51(d,J=8.0Hz,2H);7.26(d,J=8.0Hz,2H);5.36-5.22(m,1H);4.90-4.85(m,1H);2.33(s,3H). 19 F-NMR (377MHz, DMSO) δ -107.97.

[0472] Synthesis of 1-(2,2-dichloro-1-fluorocyclopropyl)-4-methylbenzene [ka]

[0473] A mixture of 1-(1-fluorovinyl)-4-methylbenzene (500 mg), CHCl (10 mL), NaOH (40%, 10 mL), and benzyltriethylammonium chloride (30 mg) was stirred at room temperature overnight. The mixture was poured into HO (50 mL) and then extracted with CHCl (3 × 20 mL). The CHCl layer was washed with HO (3 × 20 mL), dried (MgSO), and filtered. The filtrate was concentrated under reduced pressure to give crude 1-(2,2-dichloro-1-fluorocyclopropyl)-4-methylbenzene (500 mg) as a colorless liquid, which was used directly in the next reaction. 1H-NMR:(400MHz,DMSO-d6)δ7.40(t,J=12.7Hz,2H);7.28(d,J=8.0Hz,2H);2.86-2.63(m,1H);2.39-2.35(m,1H);2.33(s,3H).

[0474] Synthesis of 1-(1-fluorocyclopropyl)-4-methylbenzene [ka]

[0475] To a mixture of 1-(2,2-dichloro-1-fluorocyclopropyl)-4-methylbenzene (500 mg) in THF (10 mL) was added lithium aluminum tetrahydride (436 mg) at 0 °C, and the resulting mixture was stirred at room temperature overnight. The mixture was poured into HO (50 mL) and then extracted with EA (3 × 20 mL). The combined EA layers were washed with HO (3 × 20 mL), dried (MgSO), and filtered. The filtrate was concentrated under reduced pressure to give crude 1-(1-fluorocyclopropyl)-4-methylbenzene (250 mg) as a colorless liquid, which was used directly in the next step. 1 H-NMR:(400MHz,DMSO-d6)δ7.25-7.04(m,4H);2.38-2.25(m,3H);1.51-1.33(m,2H);1.10-1.01(m,2H).

[0476] Step 5: Synthesis of 1-(bromomethyl)-4-(1-fluorocyclopropyl)benzene [ka]

[0477] To a solution of 1-(1-fluorocyclopropyl)-4-methylbenzene (200 mg) in CCl4 (4 mL) was added AIBN (0.02 mL) and NBS (355.5 mg). The reaction was stirred at 80 °C for 16 h. The reaction was diluted with DCM and water. The organic layer was separated and concentrated in vacuo to give an oil, which was purified by column chromatography to give 1-(bromomethyl)-4-(1-fluorocyclopropyl)benzene (170 mg) as a colorless oil. 1 H-NMR (400MHz, CD3OD) δ7.40(d,J=8.2Hz,2H);7.23(d,J=8.2Hz,2H);4.55(s,2H);1.52-1.37(m,2H);1.14-1.04(m,2H).

[0478] Step 6: Synthesis of 2-(4-(1-fluorocyclopropyl)phenyl)acetonitrile [ka]

[0479] To a solution of 1-(bromomethyl)-4-(1-fluorocyclopropyl)benzene (100.0 mg, crude) in DMSO (5.0 mL) was added NaCN (107.0 mg), and the reaction mixture was stirred at 40° C. overnight. The reaction mixture was then poured into water (10 mL) and extracted with EA (20 mL). The organic layer was separated, dried (NaSO), filtered, and concentrated in vacuo to give a residue that was purified on a silica gel column (5% EA in PE) to give 2-(4-(1-fluorocyclopropyl)phenyl)acetonitrile (30 mg) as a colorless oil. 1 H-NMR (400MHz, CDCl3) δ7.30 (dd, J=22.6Hz, 8.2Hz, 4H); 3.75 (s, 2H); 1.55-1.43 (m, 2H); 1.11-1.00 (m, 2H).

[0480] Step 7: Synthesis of 2-(4-(1-fluorocyclopropyl)phenyl)acetic acid [ka]

[0481] To a solution of 2-[4-(1-fluorocyclopropyl)phenyl]acetonitrile (30 mg) in HO (3 mL) was added sodium peroxide (40 mg). The mixture was stirred at 50° C. for 24 hours. Then, the mixture was stirred at 60° C. for 24 hours. The pH of the mixture was adjusted to 4 with 1N HCl solution and extracted with EtOAc. The combined organic layers were dried, filtered, and concentrated to give 2-[4-(1-fluorocyclopropyl)phenyl]acetic acid (30 mg) as a white solid. LC / MS ESI (m / z): 193 [MH] - . 1 H-NMR (400MHz, CDCl3) δ7.29(d,J=8.2Hz,2H);7.24(d,J=8.2Hz,2H);3.66(s,1H);1.53-1.48(m,1H);1.48-1.42(m,2H);1.10-1.01(m,2H).

[0482] Step 8: Synthesis of (R)-2-(4-(1-fluorocyclopropyl)phenyl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide [ka]

[0483] A solution of 2-[4-(1-fluorocyclopropyl)phenyl]acetic acid (30 mg), (R)-1-[1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl]ethan-1-amine hydrochloride (49.0 mg), HATU (64.6 mg), and DIEA (0.077 mL) in DMF (5 mL) was stirred at 25° C. for 12 hours. The mixture was diluted with water and extracted with DCM. The combined organic extracts were dried, filtered, and concentrated to give a residue. The residue was purified by preparative HPLC (column: YMC-Actus Triart C18, 250 × 21 mm; mobile phase: 25% to 95% MeCN / HO (0.1% FA); flow rate: 15 mL / min; wavelength: 220 nm / 254 nm) to give (R)-2-(4-(1-fluorocyclopropyl)phenyl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-[c]pyridin-5-yl)ethyl)acetamide (26.9 mg) as a white solid. LCMS ESI (m / z): 421 (M+H). + . 1 H-NMR(400MHz,CDCl3)δ8.90(s,1H);8.12(s,1H);7.59(s,1H);7.31-7.27(m,2H);7.24(d,J=8.4Hz,2H) ;6.71(s,1H);5.35-5.20(m,1H);5.18-4.93(m,2H);3.59(s,2H);1.52-1.46(m,5H);1.10-1.02(s,2H). 19 F-NMR(377MHz, CDCl3)δ -70.78;-179.26. Example 30 Diastereomer A of 2-(4-(2,2-difluorocyclopropyl)phenyl)-N-((R)-1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide

[0484] Step 1: 1-Bromo-4-(2,2-difluorocyclopropyl)benzene [ka] A solution of 1-bromo-4-ethenylbenzene (2.50 mL), trimethyl(bromodifluoromethyl)silane (4.46 mL), and tetrabutylammonium bromide (0.178 mL) in toluene (50 mL) was stirred at 110° C. for 12 hours. The mixture was concentrated to a residue, and the residue was purified by silica gel chromatography (PE:EA=1:0 to 50:1) to give racemic 1-bromo-4-(2,2-difluorocyclopropyl)benzene (4.2 g) as a colorless oil. 1 H-NMR (400MHz, CDCl3) δ7.50-7.43(d,J=8.5Hz,2H);7.10(d,J=8.5Hz,2H);2.78-2.61(m,1H);1.92-1.74(m,1H);1.66-1.56(m,1H).

[0485] Step 2: Synthesis of ethyl 2-(4-(2,2-difluorocyclopropyl)phenyl)acetate [ka] A solution of 1-bromo-4-(2,2-difluorocyclopropyl)benzene (4 g), potassium 1-ethyl-3-propanedioate (4.38 g), allylpalladium chloride dimer (0.13 g), BINAP (0.64 g), and DMAP (2.10 g) in mesitylene (10 mL) was purged with N. The mixture was stirred at 140 °C for 1 h and then at 120 °C for 12 h. The mixture was concentrated to give a residue, which was purified by silica gel chromatography (PE: EtOAc = 1:0 to 50:1) to give ethyl 2-[4-(2,2-difluorocyclopropyl)phenyl]acetate (1.54 g) as a white solid. LCMS ESI (m / z): 241 (M+H). + .

[0486] Step 3: 2-(4-(2,2-difluorocyclopropyl)phenyl)acetic acid [ka]

[0487] To a solution of racemic 2-[4-(2,2-difluorocyclopropyl)phenyl]ethyl acetate (1.3 g) in MeOH (4.0 mL) was added NaOH (1 M in HO, 30 mL). The mixture was stirred at 25 °C for 12 h. The reaction mixture was then concentrated, diluted with HO, and washed with EtOAc. The aqueous phase was adjusted to pH = 3 with 1 N HCl and extracted with EtOAc. The combined organic extracts were dried, filtered, and concentrated to give racemic 2-(4-(2,2-difluorocyclopropyl)phenyl)acetic acid as a white solid. [LC / MS ESI (m / z): 211 [MH] - . 1 H-NMR (400 MHz, CDCl) δ 7.27-7.23 (d, J = 8.4 Hz, 2H); 7.20 (d, J = 8.4 Hz, 2H); 3.64 (s, 2H); 2.81-2.64 (m, 1H); 1.90-1.71 (m, 1H); 1.65-1.60 (m, 1H). Racemic 2-(4-(2,2-difluorocyclopropyl)phenyl)acetic acid was separated by chiral preparative SFC (column: ChiralPak TM (Trademark) AD, 250 × 21.2 mm ID, 5 μm, mobile phase: A is CO, B is MeOH, gradient: B 15%; wavelength: 220 nm) to give 2-[4-(2,2-difluorocyclopropyl)phenyl]acetic acid enantiomer P1 (200 mg) and 2-[4-(2,2-difluorocyclopropyl)phenyl]acetic acid enantiomer P2 (240 mg) as white solids.

[0488] Step 4A: 2-(4-((R or S)-2,2-difluorocyclopropyl)phenyl)-N-((R)-1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide [ka]

[0489] A solution of 2-[4-(2,2-difluorocyclopropyl)phenyl]acetic acid enantiomer P2 (120 mg), (R)-1-[1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl]ethan-1-amine hydrochloride (165.7 mg), HATU (236.5 mg), and DIEA (0.28 mL) in DMF (5 mL) was stirred at 25° C. for 12 hours. The mixture was diluted with water and extracted with DCM. The combined organic extracts were dried, filtered, and concentrated to give a residue. The residue was purified by preparative HPLC (column: GEMINI TM Purification with HPLC™ C18, 250 x 21.2 mm x 5 μm; mobile phase: 25% to 95% MeCN / HO (0.1% FA); flow rate: 15 mL / min; wavelength: 220 nm / 254 nm gave 2-(4-((R or S)-2,2-difluorocyclopropyl)phenyl)-N-((R)-1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide (79.6 mg) as a white solid. LCMS ESI (m / z): 439 (M+H). + . 1 H-NMR(400MHz,CDCl3)δ8.88(s,1H);8.12(s,1H);7.59(s,1H);7.25(d,J=8.4Hz,2H);7.19(d,J=8.4Hz,2H);6.70(d,J=7.2Hz,1H); 5.33-5.16(m,1H);5.15-4.93(m,2H);3.58(s,2H);2.88-2.60(m,1H);1.92-1.75(m,1H);1.70-1.58(m,1H);1.50(d,J=6.8Hz,3H). 19 F-NMR(377MHz, CDCl3)δ -70.79;-125.90;-142.41. Example 31 2-(4-(2,2-difluorocyclopropyl)phenyl)-N-((R)-1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide diastereomer B

[0490] Step 4B: 2-(4-((R or S)-2,2-difluorocyclopropyl)phenyl)-N-((R)-1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide [ka]

[0491] A solution of 4-(2,2-difluorocyclopropyl)phenyl]acetic acid enantiomer P1 (100 mg), (R)-1-[1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl]ethan-1-amine hydrochloride (138.1 mg), HATU (197.1 mg), and DIEA (0.23 mL) in DMF (5 mL) was stirred for 12 h at 25° C. The mixture was diluted with water and extracted with DCM. The combined organic extracts were dried, filtered, and concentrated to give a residue that was purified by preparative HPLC (column: YMC-Actus Triart C18, 250 × 21 mm; mobile phase: 55% to 95% MeCN / HO (0.1% FA); flow rate: 20 mL / min; wavelength: 220 nm / 254 nm) to give 2-(4-((R or S)-2,2-difluorocyclopropyl)phenyl)-N-((R)-1-(1-(2,2,2-)trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide (50.7 mg, white solid). LCMS ESI (m / z): 439 (M+H). + . 1 H-NMR(400MHz,CDCl3)δ8.88(s,1H);8.12(s,1H);7.58(s,1H);7.25(d,J=8.4Hz,2H);7.20(d,J=8.4Hz,2H);6.72(s,1H);5.38 -5.20(m,1H);5.14-4.95(m,2H);3.58(s,2H);2.86-2.62(m,1H);1.95-1.76(m,1H);1.70-1.58(m,1H);1.49(d, J=6.8Hz,3H). 19 F-NMR(377MHz, CDCl3)δ -70.80;-125.92;-142.30. Example 32 (R)-2-(4-isopropylphenyl)-N-(1-(3-methyl-1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide

[0492] Synthesis of 2-bromo-5-fluoro-N-methoxy-N-methylisonicotinamide [ka]

[0493] To a solution of 2-bromo-5-fluoroisonicotinic acid (10 g) in DCM (400 mL) was added HATU (22.47 g), TEA (18.9 mL), and N,O-dimethylhydroxylamine hydrochloride (4.8 g). The reaction mixture was stirred under nitrogen at 20° C. for 16 hours. The solvent was evaporated under reduced pressure to give a residue. The residue was dissolved in EA (500 mL) and washed twice with water (500 mL), then with brine (500 mL). The organic layer was separated, dried over Na2SO4, filtered, and concentrated to give a residue that was purified by Biotage column (0% to 15% EA with PE) to give 2-bromo-5-fluoro-N-methoxy-N-methylisonicotinamide (10.3 g) as a yellow oil: LCMS ESI (m / z): 262.9 (M+H). + .

[0494] Synthesis of 1-(2-bromo-5-fluoropyridin-4-yl)ethan-1-one [ka]

[0495] 2-Bromo-5-fluoro-N-methoxy-N-methylisonicotinamide (10.3 g) was suspended in THF (200 mL) and stirred at 0° C. for 5 minutes. Methylmagnesium bromide (19.58 mL, 3.0 M in ether) was slowly added to the reaction mixture under a nitrogen atmosphere, and the reaction was stirred at 20° C. for 16 hours. The reaction mixture was quenched with saturated ammonium chloride. The two phases were separated, and the aqueous phase was extracted with ethyl acetate. The combined organic phases were washed with brine, dried over Na2SO4, filtered, and concentrated to give a residue that was purified on a Biotage® column (0% to 10% EA / PE) to give 1-(2-bromo-5-fluoropyridin-4-yl)ethan-1-one (8.3 g) as a colorless oil. LCMS ESI (m / z): 218.0 (M+H). + .

[0496] Synthesis of 5-bromo-3-methyl-1H-pyrazolo[3,4-c]pyridine [ka]

[0497] To a solution of 1-(2-bromo-5-fluoropyridin-4-yl)ethan-1-one (8.3 g) in ethylene glycol (50 mL) was added hydrazine hydrate (4.793 mL), and the reaction mixture was stirred at 120 °C under a nitrogen atmosphere for 72 h. Upon completion, the reaction mixture was diluted with EA (200 mL), and the resulting mixture was washed twice with water (2 L) and brine (500 mL). The organic layer was separated, dried over Na2SO4, filtered, and evaporated to give a residue that was purified by Biotage® column (0% to 20% EA / PE) to give 5-bromo-3-methyl-1H-pyrazolo[3,4-c]pyridine (2.7 g) as a white solid. LCMS ESI (m / z): 212 (M+H). + . 1 H-NMR (400MHz, CDCl3): δ8.80(d,J=0.9Hz,1H);7.80(d,J=0.9Hz,1H);2.60(s,3H).

[0498] Synthesis of 5-bromo-3-methyl-1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridine [ka]

[0499] To a solution of 5-bromo-3-methyl-1H-pyrazolo[3,4-c]pyridine (5.30 g) in 26 mL of anhydrous DMF was added 2,2,2-trifluoroethyl trifluoromethanesulfonate (4.3 mL) and CsCO (9.82 g). The reaction mixture was stirred under a nitrogen atmosphere at 25 °C for 16 h. Upon completion, the reaction mixture was poured into H2O and extracted three times with EA. The combined organic layers were washed with brine, dried over Na2SO4, filtered, and evaporated in vacuo. The residue was purified by Biotage® column (0%-10% EA / PE; 80 g cartridge column) to give 5-bromo-3-methyl-1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridine (5.72 g) as a white solid. LCMS ESI (m / z): 294 (M+H). + . 1 H-NMR (400MHz, CDCl3) δ8.69(s,1H);7.79(d,J=0.9Hz,1H);5.02-4.87(m,2H);2.57(s,3H). 19 F-NMR (377MHz, CDCl3)δ -70.91(s).

[0500] Synthesis of 3-methyl-1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde [ka]

[0501] To a solution of 5-bromo-3-methyl-1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridine (2 g) in anhydrous DMF (60 mL) was added Na2CO3 (1.082 g), Pd(dppf)2Cl2 (498 mg), and Et3SiH (2.2 mL). The reaction mixture was filled with N2 and stirred at 100 °C under a CO2 (7 bar) atmosphere for 16 h. After completion, water was added to the reaction mixture and extracted with ethyl acetate. The combined organic extracts were washed with brine, dried over Na2SO4, filtered, and concentrated to a black oil, which was purified by Biotage® column (0%-20% EA with PE; 40 g cartridge column). The fractions were collected and evaporated to give 3-methyl-1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde (1.2 g) as a yellow solid. LCMS ESI (m / z): 244 (M+H). + . 1 H-NMR (400MHz, CDCl3): δ10.24(s,1H);9.04(s,1H);8.37(d,J=0.6Hz,1H);5.10-4.98(m,2H);2.66(s,3H). 19 F-NMR (377MHz, CDCl3)δ -70.80(s).

[0502] Synthesis of (R,E)-2-methyl-N-((3-methyl-1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)methylene)propane-2-sulfinamide [ka]

[0503] To a solution of 3-methyl-1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridine-5-carbaldehyde (1.2 g) in anhydrous DCM (15 mL) was added CuSO4 (2.36 g) and (R)-2-methylpropane-2-sulfinamide (0.78 g). The reaction mixture was stirred at room temperature overnight. The solution was filtered through a pad of Celite, and the filtrate was concentrated. The residue was purified on a silica gel column to give (R,E)-2-methyl-N-((3-methyl-1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)methylene)propane-2-sulfinamide (1.5 g). LCMS ESI (m / z): 347 (M+H). + . 1 H-NMR (400MHz, CDCl3) δ9.02(s,1H);8.85(s,1H);8.31(d,J=1.1Hz,1H);5.07-4.95(m,2H);2.66(s,3H);1.31(s,9H).

[0504] Synthesis of (R)-2-methyl-N-((R)-1-(3-methyl-1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)propane-2-sulfinamide [ka]

[0505] To a solution of (R,E)-2-methyl-N-((3-methyl-1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)methylene)propane-2-sulfinamide (1.6 g, 4.62 mmol) in anhydrous THF (17 mL) was added methylmagnesium bromide (7.699 mL, 3.0 M in ether) dropwise at −78° C. under nitrogen. The reaction mixture was stirred at −78° C. for 2 h. The reaction solution was then quenched with aqueous NH4Cl. The two phases were separated and the aqueous phase was extracted with EA. The combined organic layers were washed with brine, dried over Na2SO4, filtered and concentrated in vacuo. The residue was purified by silica gel column chromatography to give (R)-2-methyl-N-((R)-1-(3-methyl-1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)propane-2-sulfinamide (1.5 g, 4.139 mmol) as a white solid. LCMS ESI(m / z): 363(M+H) + .

[0506] Synthesis of (R)-1-(3-methyl-1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethan-1-amine hydrochloride [ka]

[0507] To a solution of (R)-2-methyl-N-[(R)-1-[3-methyl-1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl]ethyl]propane-2-sulfinamide (1.0 g, 2.76 mmol) in dioxane (4.0 mL), 4N HCl-dioxane (2.0 mL) was added, and the reaction mixture was stirred at room temperature for 1 hour. The mixture was then concentrated to give a residue that was used directly in the next step. LCMS ESI (m / z): 259 (M+H). + .

[0508] Synthesis of (R)-2-(4-isopropylphenyl)-N-(1-(3-methyl-1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide [ka]

[0509] To (R)-1-(3-methyl-1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethan-1-amine hydrochloride (238 mg) in anhydrous DMF (1.5 mL) was added DIEA (0.6 mL) to obtain solution A. To a solution of 2-(4-isopropylphenyl)acetic acid (126.4 mg) in DMF (1 mL) was added HATU (296.5 mg) and stirred at room temperature for 15 minutes to obtain solution B. Solution A was added to the solution and stirred at room temperature for 1 hour. The reaction mixture was diluted with EA and water. The two phases were separated, and the organic layer was washed with brine, dried over Na2SO4, filtered, and concentrated in vacuo. The residue was purified by preparative HPLC (column: AZZOTA C18, 30 x 250 mm x 10 μm; mobile phase: 25%-95% MeCN / HO (1% FA); flow rate: 15 mL / min; wavelength: 220 nm / 254 nm) and SFC (column: ChiralPak® IG, 250 x 21.2 mm ID, 5 μm; mobile phase: CO2 in phase A, MEOH + 0.1% NH3·HO in phase B; gradient: B 40%; wavelength: 220 nm) to give (R)-2-(4-isopropylphenyl)-N-(1-(3-methyl-1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide as a white solid (100.2 mg). LCMS ESI (m / z): 419.5 (M+H). + . 1H-NMR(400MHz,CDCl3)δ8.78(s,1H);7.47(s,1H);7.19(s,4H);6.58(s,1H);5.34-5.20(m,1H);5.01-4.87( m,2H);3.55(d,J=1.8Hz,2H);2.95-2.85(m,1H);2.56(s,3H);1.48(d,J=6.8Hz,3H);1.25(d,J=6.9Hz,6H). 19 F-NMR(377MHz, CDCl3)δ -70.92. Example 33 (R)-2-(4-cyclopropylphenyl)-N-(1-(3-methyl-1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide

[0510] (R)-2-(4-cyclopropylphenyl)-N-(1-(3-methyl-1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide [ka]

[0511] To a solution of (R)-1-[3-methyl-1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl]ethan-1-amine hydrochloride (242 mg) in anhydrous DMF (1.5 mL), DIEA (0.6 mL) was added to give solution A. To a solution of 2-(4-cyclopropylphenyl)acetic acid (111.58 mg) in DMF (1.0 mL), HATU (264.83 mg) was added and stirred at room temperature for 15 minutes to give solution B. Solution A was added to solution B and stirred at room temperature for 1 hour. The reaction mixture was diluted with EA and water. The organic layer was washed with aqueous NaCl, dried over Na2SO4, filtered, and concentrated in vacuo. The residue was purified by preparative HPLC (column: YMC-Actus Triart C18, 250 × 21 mm; mobile phase: 40%–90% MeCN / HO (0.1% NH₃·HO); flow rate: 15 mL / min; wavelength: 220 nm / 254 nm) and SFC (column: ChiralPak® IG, 250 × 21.2 mm ID, 5 μm; mobile phase: CO in phase A, MEOH + 0.1% NH₃·HO in phase B; gradient: B 45%; flow rate: 50 mL / min; wavelength: 220 nm) to give (R)-2-(4-cyclopropylphenyl)-N-(1-(3-methyl-1-(2,2,2-trifluoroethyl))-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide as a white solid (86.7 mg). LCMS ESI(m / z):417.5(M+H) + . 1 H-NMR(400MHz,CDCl3)δ8.78(s,1H);7.44(d,J=0.9Hz,1H);7.15(d,J=8.1Hz,2H);7.04(d,J=8.1Hz,2H);6.52(d,J=7.4Hz,1H);5.31-5.22(m ,1H);5.00-4.87(m,2H);3.54(d,J=2.4Hz,2H);2.55(s,3H);1.93-1.8 4(m,1H);1.46(d,J=6.8Hz,3H);1.00-0.92(m,2H);0.72-0.64(m,2H). 19 F-NMR (377MHz, CDCl3)δ -70.92(s). Example 34 (R)-2-(5-cyclopropylpyridin-2-yl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide

[0512] Synthesis of methyl 2-(5-cyclopropylpyridin-2-yl)acetate [ka]

[0513] To a solution of methyl 2-(5-bromopyridin-2-yl)acetate (1 g) in toluene-water (10:1, 16.5 mL) was added potassium cyclopropyltrifluoroborate (1.29 g), CsCO (4.25 g), Pd(OAc) (0.20 g), and di(adamantan-1-yl)(butyl)phosphine (0.31 g). The reaction mixture was filled with N and stirred at 110 °C overnight. The reaction mixture was diluted with EA and water. The two phases were separated, and the organic layer was washed with saturated NaCl, dried over NaSO, filtered, and concentrated in vacuo. The resulting residue was purified by silica gel column chromatography to give methyl 2-(5-cyclopropylpyridin-2-yl)acetate (582 mg) as an oil. LCMS ESI (m / z): 192 (M+H). + .

[0514] Synthesis of 2-(5-cyclopropylpyridin-2-yl)acetic acid [ka]

[0515] To a solution of methyl 2-(5-cyclopropylpyridin-2-yl)acetate (582 mg) in MeOH-HO (2:1, 9.5 mL) was added NaOH (183 mg). The reaction mixture was stirred at room temperature for 3 hours. Then, MeOH was removed under reduced pressure, and the pH of the aqueous layer was adjusted to pH = 5 with 2 M HCl while cooling in an ice bath. Ethyl acetate was then added to the acidic aqueous layer, and the resulting precipitate was collected by filtration. The aqueous layer was then extracted with DCM-MeOH (10:1), and the combined organic layers were concentrated in vacuo. Both the residue and the precipitate were combined to give 2-(5-cyclopropylpyridin-2-yl)acetic acid (354 mg) as a white solid. LCMS ESI (m / z): 178 (M+H). + .1 H-NMR(400MHz,DMSO-d6)δ8.64(d,J=2.0Hz,1H);8.05-8.00(m,1H);7.73(d,J=8 .3Hz,1H);4.06(s,2H);2.18-2.08(s,1H);1.15-1.08(m,2H);0.93-0.85(m,2H).

[0516] Synthesis of (R)-2-(5-cyclopropylpyridin-2-yl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide [ka]

[0517] To a solution of (R)-1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethan-1-amine hydrochloride (308 mg) in anhydrous DMF (1.5 mL) was added DIEA (1 mL) to give solution A. To a solution of 2-(5-cyclopropylpyridin-2-yl)acetic acid (150 mg) in DMF (1 mL) was added HATU (405.76 mg) and stirred at room temperature for 5 minutes to give solution B. Solution A was added to solution B, and the reaction mixture was stirred at room temperature for 1 hour. The reaction mixture was then diluted with EA and water. The organic layer was washed with aqueous NaCl, dried over Na2SO4, filtered, and concentrated in vacuo. The residue was purified by preparative HPLC (column: YMC-Actus Triart C18, 250 x 21 mm; mobile phase: 10% to 90% MeCN / HO (0.1% FA); flow rate: 15 mL / min; wavelength: 220 nm / 254 nm) to give (R)-2-(5-cyclopropylpyridin-2-yl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide as a white solid (135.4 mg). LCMS ESI (m / z): 404.4 (M+H). + . 1H-NMR(400MHz,CDCl3)δ8.94(s,1H);8.36(d,J=1.9Hz,1H);8.09(s,1H);7.82(d,J=7.2Hz,1H);7.60(s,1H);7.52(d,J=7.7Hz,1H);7.40(d,J=8.2H) z,1H);5.32-5.18(m,1H);5.11-4.98(m,2H);3.87(d,J=2.3Hz,2H);2.00- 1.90(m,1H);1.53(d,J=6.9Hz,3H);1.17-1.06(m,2H);0.81-0.72(m,2H). 19 F-NMR (377MHz, CDCl3)δ -70.81(s). Example 35 (R)-2-(5-chloro-6-(1,1-difluoroethyl)pyridin-3-yl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide

[0518] Step 1: Synthesis of 5-bromo-3-chloro-N-methoxy-N-methylpicolinamide [ka]

[0519] To a solution of 5-bromo-3-chloropyridine-2-carboxylic acid (4.76 g) in DCM (10 mL) was added HATU (9.94 g), methoxy(methyl)amine (3.396 mL), and TEA (8.4 mL). The reaction mixture was stirred at room temperature for 2 hours. The reaction mixture was then diluted with DCM and water, and the two phases were separated. The organic phase was washed with saturated NaCl, dried over Na2SO4, filtered, and concentrated in vacuo. The residue was purified using silica gel column chromatography eluting with ethyl acetate (15%) in petroleum ether to give 5-bromo-3-chloro-N-methoxy-N-methylpicolinamide (5.11 g) as a white solid. LCMS ESI (m / z): 281 (M+H) + .

[0520] Step 2: Synthesis of 1-(5-bromo-3-chloropyridin-2-yl)ethan-1-one [ka]

[0521] To a solution of 5-bromo-3-chloro-N-methoxy-N-methylpicolinamide (5.48 g) in THF (65 mL) was added methylmagnesium bromide (32.6 mL, 3.0 M in EtO) under nitrogen at −78° C. The reaction mixture was stirred at −78° C. for 2 hours. Upon completion, the reaction mixture was diluted with DCM and water. The two phases were separated, and the aqueous phase was extracted with DCM (30 mL×3). The combined organic phases were dried over NaSO, filtered, and concentrated to give a residue. The residue was purified by silica gel column chromatography eluting with ethyl acetate (6%) in petroleum ether to give 1-(5-bromo-3-chloropyridin-2-yl)ethan-1-one as a white solid (602 mg). LCMS ESI (m / z): 234 (M+H). + . 1 H-NMR (400MHz, CDCl3) δ8.59(d,J=1.6Hz,1H);7.98(d,J=1.9Hz,1H);2.67(s,3H).

[0522] Step 3: Synthesis of 5-bromo-3-chloro-2-(1,1-difluoroethyl)pyridine [ka]

[0523] To a solution of 1-(5-bromo-3-chloropyridin-2-yl)ethan-1-one (2 g) in 1,2-dichloroethane (40 mL) was added DAST (11.27 mL). The mixture was stirred in a sealed tube at 80° C. for 12 hours. The mixture was quenched with saturated sodium bicarbonate and extracted with DCM. The combined organic layers were dried, filtered, and concentrated to give a residue that was purified by silica gel chromatography (PE:EA=20:1) to give 5-bromo-3-chloro-2-(1,1-difluoroethyl)pyridine (1.0 g) as a yellow oil.

[0524] 1H-NMR (400MHz, CDCl3) δ8.55(d,J=2.0Hz,1H);7.98(d,J=2.0Hz,1H);2.06(t,J=18.8Hz,3H).

[0525] Step 4: Synthesis of ethyl 2-(5-chloro-6-(1,1-difluoroethylpyridin-3-yl)acetate [ka]

[0526] A solution of 5-bromo-3-chloro-2-(1,1-difluoroethyl)pyridine (200 mg), potassium ethyl malonate (199.1 mg), allylpalladium chloride dimer (34.24 mg), BINAP (97.11 mg), and DMAP (95.27 mg) in mesitylene (10 mL) was filled with N2 and the mixture was stirred at 140 °C for 1 hour. Then, the mixture was stirred at 120 °C for 12 hours. The mixture was concentrated to give a residue, which was purified by silica gel chromatography (PE:EA = 10:1) to give ethyl 2-[5-chloro-6-(1,1-difluoroethyl)pyridin-3-yl]acetate (50 mg) as a yellow oil. LCMS ESI (m / z): 264 (M+H) + .

[0527] Step 5: Synthesis of 2-(5-chloro-6-(1,1-difluoroethyl)pyridin-3-yl)acetic acid [ka]

[0528] To a solution of 2-[5-chloro-6-(1,1-difluoroethyl)pyridin-3-yl]ethyl acetate (50 mg) in MeOH (1.0 mL) was added NaOH (1 M in HO, 1.0 mL). The mixture was stirred at 25 °C for 12 hours. The mixture was then concentrated, diluted with water, and extracted with EtOAc. The aqueous layer was adjusted to pH = 3 with 1N HCl and extracted with DCM. The combined organic layers were dried, filtered, and concentrated to give 2-[5-chloro-6-(1,1-difluoroethyl)pyridin-3-yl]acetic acid (25 mg) as a white solid. LCMS ESI (m / z): 234 (MH) - .

[0529] Step 6: Synthesis of (R)-2-(5-chloro-6-(1,1-difluoroethyl)pyridin-3-yl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide [ka]

[0530] A solution of 2-[5-chloro-6-(1,1-difluoroethyl)pyridin-3-yl]acetic acid (80 mg), (1R)-1-[1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl]ethan-1-amine hydrochloride (107.80 mg), HATU (142.0 mg), and DIEA (0.168 mL) in DMF (1.0 mL) was stirred at 25° C. for 2 hours. The reaction mixture was diluted with water and extracted with DCM. The combined organic layers were dried, filtered, and concentrated to give a residue. The residue was purified by preparative HPLC (column: YMC-Actus Triart, C18, 250 × 20 mm; mobile phase: 30% to 95% MeCN / HO (0.1% FA); flow rate: 15 mL / min; wavelength: 220 nm / 254 nm) to give (R)-2-(5-chloro-6-(1,1-difluoroethyl)pyridin-3-yl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide (82.3 mg) as a white solid. LCMS ESI (m / z): 462 (M+H)+ . 1 H-NMR(400MHz,CDCl3)δ8.97(s,1H);8.40(s,1H);8.17(s,1H);7.80(d,J=1.6Hz,1H);7.67(s,1H);7.02(d,J= 14.4Hz,1H);5.38-5.24(m,1H);5.19-5.00(m,2H);3.61(s,2H);2.07(t,J=18.8Hz,3H);1.57(d,J=6.8Hz,3H). 19 F-NMR(377MHz, CDCl3)δ -70.73;-89.61. Example 36 (R)-2-(5-chloro-6-cyclopropylpyridin-3-yl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide [ka]

[0531] Step 1: Synthesis of 3-chloro-2-cyclopropyl-5-methylpyridine. A solution of 2-bromo-3-chloro-5-methylpyridine (4.2 g), cyclopropylboronic acid (1.92 g), Pd(PPh) (2.35 g), and KPO (12.95 g) in toluene (80 mL) / HO (8 mL) was stirred at 110 °C under N for 12 hours. The reaction completion was monitored by LCMS. The mixture was concentrated in vacuo to give a residue, which was purified by silica gel column chromatography (PE:EA = 10:1) to give 3-chloro-2-cyclopropyl-5-methylpyridine (1.4 g) as a colorless oil. LCMS ESI (m / z): 168 (M+H). + .

[0532] Step 2: Synthesis of 5-(bromomethyl)-3-chloro-2-cyclopropylpyridine. A solution of 3-chloro-2-cyclopropyl-5-methylpyridine (960 mg), NBS (1.12 g), and AIBN (0.042 mL) in carbon tetrachloride (15 mL) was stirred at 80° C. under N for 12 hours. The reaction mixture was filtered and concentrated to give a residue. The residue was purified by silica gel column chromatography (PE:EA=30:1) to give 5-(bromomethyl)-3-chloro-2-cyclopropylpyridine (700 mg) as a colorless oil. LCMS ESI (m / z): 246 (M+H). + . [ka]

[0533] Step 3: Synthesis of 2-(5-chloro-6-cyclopropylpyridin-3-yl)acetonitrile. A solution of 5-(bromomethyl)-3-chloro-2-cyclopropylpyridine (600 mg, 2.43 mmol) and NaCN (239 mg) in DMF (5 mL) was stirred at 25° C. for 12 hours. The reaction completion was monitored by LCMS. The mixture was diluted with water and extracted with DCM. The mixture was dried, filtered, and concentrated to give 2-(5-chloro-6-cyclopropylpyridin-3-yl)acetonitrile (380 mg) as a yellow oil. LCMS ESI (m / z): 193 (M+H). + .

[0534] Step 4: Synthesis of 2-(5-chloro-6-cyclopropylpyridin-3-yl)acetic acid. To a solution of 2-(5-chloro-6-cyclopropylpyridin-3-yl)acetonitrile (380 mg) in MeOH (2 mL) was added NaOH (10 mL, 20.0 mmol, 2N). The mixture was stirred at 25° C. for 12 hours, and the reaction was complete by LCMS. The reaction was then extracted with EtOAc. The aqueous layer was adjusted to pH=3 with 1N HCl solution and lyophilized to give a white solid. The white solid was dissolved in MeOH / DCM (10:1), filtered, and concentrated to give 2-(5-chloro-6-cyclopropylpyridin-3-yl)acetic acid (150 mg) as a white solid. LCMS ESI (m / z): 210 (MH). - . [ka]

[0535] Step 5: (R)-2-(5-chloro-6-cyclopropylpyridin-3-yl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide. A solution of 2-(5-chloro-6-cyclopropylpyridin-3-yl)acetic acid (150 mg), (R)-1-[1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl]ethan-1-amine hydrochloride (258.89 mg), HATU (296 mg) and DIEA (0.35 mL) in DMF (5 mL) was stirred at 25° C. for 12 hours. The mixture was diluted with water and extracted with DCM. The organic layer was dried, filtered and concentrated to give a residue. The residue was purified by preparative HPLC (column: YMC-Actus Triart C18, 250 × 21 mm; mobile phase: 20% to 95% MeCN / HO (0.1% FA); flow rate: 15 mL / min; wavelength: 220 nm / 254 nm) to give (R)-2-(5-chloro-6-cyclopropylpyridin-3-yl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide (203 mg) as a white solid. LCMS ESI (m / z): 438 (M+H). + .1 H-NMR(400MHz,CDCl3)δ8.97(s,1H);8.22(d,J=2.0Hz,1H);8.15(s,1H);7.64(s,1H);7.61(d,J=2.0Hz,1H);7.03-6.89(m,1H);5 .36-5.21(m,1H);5.16-5.01(m,2H);3.51(s,2H);2.56-2.42(m,1H);1.54(d,J=6.8Hz,3H);1.12-1.07(m,2H);1.06-1.01(m,2H). 19 F-NMR(377MHz, CDCl3)δ -70.73. Example 37 (R)-2-(3-chloro-4-(1,1-difluoroethyl)phenyl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide [ka]

[0536] Step 1: 4-Bromo-2-chloro-N-methoxy-N-methylbenzamide. To a solution of 4-bromo-2-chlorobenzoic acid (2.0 g) in DCM (25 mL) was added HATU (4.20 g), methoxy(methyl)amine hydrochloride (4.54 g), and TEA (3.5 mL). The reaction was then stirred at room temperature overnight and was monitored by LCMS for completion. The reaction was diluted with DCM and water, and the organic layer was separated, washed with saturated NaCl solution, and concentrated in vacuo. The residue was purified by silica gel column chromatography eluting with 25% ethyl acetate in petroleum ether to give 4-bromo-2-chloro-N-methoxy-N-methylbenzamide (2.209 g) as a pale yellow oil. LCMS ESI (m / z): 279 [M+H] + .

[0537] Step 2: 1-(4-Bromo-2-chlorophenyl)ethan-1-one. To a solution of 4-bromo-2-chloro-N-methoxy-N-methylbenzamide (1.07 g) in THF (10 mL) was added MeMgBr (6.4 mL, 3.0 M in EtO). The reaction was stirred at −78° C. for 2 hours and then at room temperature overnight. The reaction was diluted with DCM and water, and the two phases were separated. The aqueous phase was further extracted with DCM (30 mL × 3). The combined organic phase was dried over anhydrous NaSO, filtered, and concentrated to give a residue that was purified by silica gel column chromatography (PE:EA=10:1) to give 1-(4-bromo-2-chlorophenyl)ethan-1-one as a pale yellow solid (820 mg). 1 H-NMR (400MHz, CDCl3) δ7.61 (s, 1H); 7.50-7.44 (m, 2H); 2.64 (s, 3H). 19 F-NMR(377MHz, CDCl3)δ -87.65. [ka]

[0538] Step 3: 4-Bromo-2-chloro-1-(1,1-difluoroethyl)benzene. To a solution of 1-(4-bromo-2-chlorophenyl)ethan-1-one (2.2 g) in BAST (2.6 mL) was added two drops of methanol. Upon completion of the addition, the reaction mixture was placed under a nitrogen atmosphere and heated to 70°C overnight. The reaction mixture was cooled to room temperature and quenched with 50 mL of water. The resulting mixture was extracted twice with 50 mL of ether. The combined ether phase was washed twice with water, twice with aqueous sodium bicarbonate, once with 10% aqueous citric acid, and once with brine, and then dried over magnesium sulfate. The solvent was removed under reduced pressure, and purification by column chromatography on silica gel (2% EA in PE) afforded 4-bromo-2-chloro-1-(1,1-difluoroethyl)benzene (1.71 g) as a colorless oil. 1 H-NMR (400MHz, CDCl3) δ7.62 (d, J = 4.0 Hz, 1H); 7.50-7.43 (m, 2H); 2.02 (t, J = 18.4Hz, 3H). 19F-NMR(377MHz, CDCl3)δ -87.65.

[0539] Step 4: Ethyl 2-(3-chloro-4-(1,1-difluoroethyl)phenyl)acetate. To a solution of 4-bromo-2-chloro-1-(1,1-difluoroethyl)benzene (1.5 g) in mesitylene (2 mL) was added diallylpalladium dichloride (1.56 mg), BINAP (0.22 g), DMAP (0.72 g), and potassium ethyl malonate (1.50 g). The reaction mixture was filled with N and stirred at 120 °C overnight. The reaction was diluted with EA and water, the two phases were separated, and the aqueous phase was extracted with DCM (30 mL × 3). The combined organic phase was dried over Na2SO4, filtered, and concentrated to give a residue, which was purified by silica gel column chromatography (PE:EA=20:1) to give ethyl 2-(3-chloro-4-(1,1-difluoroethyl)phenyl)acetate (590 mg, 2.246 mmol) as a pale yellow oil. 1 H-NMR(400MHz,CDCl3)δ7.56(d,J=8.0Hz,1H);7.38(s,1H);7.24(d,J=8.0Hz,1H);4 .17(q,J=20.0Hz,2H);3.61(s,2H);2.03(t,J=36.0Hz,3H);1.27(t,J=16.0Hz,3H). 19 F-NMR(377MHz, CDCl3)δ -87.45. [ka]

[0540] Step 5: 2-(3-chloro-4-(1,1-difluoroethyl)phenyl)acetic acid. A 25 mL round-bottom flask was charged with ethyl 2-(3-chloro-4-(1,1-difluoroethyl)phenyl)acetate (590 mg), 1 M aqueous NaOH solution (10 mL), and MeOH (10 mL). The reaction mixture was stirred at room temperature for 1 hour. Completion of the reaction was monitored by LCMS. Then, water (50 mL) was added, and the aqueous layer was washed with EA. The aqueous layer was acidified with 1 M aqueous HCl solution to pH < 3, and the mixture was extracted with EA. The combined organic phases were washed with saturated NaCl, dried over Na2SO4, filtered, and concentrated to give 2-(3-chloro-4-(1,1-difluoroethyl)phenyl)acetic acid (465 mg) as a yellow-green solid. 1 H-NMR (400MHz, CDCl3) δ7.58(d,J=8.0Hz,1H);7.39(s,1H);7.23(d,J=8.0Hz,1H);3.67(s,2H);2.03(t,J=36.0Hz,3H). 19 F-NMR(377MHz, CDCl3)δ -87.53. LCMS ESI(m / z):233(MH) + . [ka]

[0541] Step 6: (R)-2-(3-chloro-4-(1,1-difluoroethyl)phenyl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo)[3,4-c]pyridin-5-yl)ethyl)acetamide. A solution of 2-(3-chloro-4-(1,1-difluoroethyl)phenyl)acetic acid (120 mg) and HATU (292 mg) in 2 mL of DMF was stirred at room temperature for 15 minutes (Solution A). In a separate container, (R)-1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethan-1-amine hydrochloride (186 mg) and DMF (2.0 mL) were added. DIEA (0.566 mL) was added until the pH of the solution was greater than 7 using moist pH paper (Solution B). Solution B was added to solution A, and the reaction was stirred for 1 h, at which point LCMS indicated the reaction was complete. The reaction was diluted with EA and water, the two phases were separated, and the aqueous phase was extracted with EA (10 mL × 2). The combined organic phases were dried over anhydrous Na2SO4, filtered, and concentrated in vacuo. The residue was purified by preparative HPLC (column: AZZOTA C18, 30 × 250 mm × 10 μm; mobile phase: 35% to 95% MeCN + HO (0.1% FA); flow rate: 20 mL / min; wavelength: 220 nm / 254 nm) to give (R)-2-(3-chloro-4-(1,1-difluoroethyl)phenyl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide (130 mg) as a white solid. 1 H-NMR(400MHz,CDCl3)δ8.93(s,1H);8.14(s,1H);7.62(s,1H);7.55(d,J=8.0Hz,1H);7.36(s,1H);7.24(d,J=8.0Hz,1H) ;6.85(d,J=8.0Hz,1H);5.32-5.25(m,1H);5.10-5.03(m,2H);3.57(s,2H);2.02(t,J=16.0Hz,3H);1.53(d,J=8.0Hz,3H). 19 F-NMR(377MHz, CDCl3)δ -70.76;-87.39. LC / MS ESI(m / z):461[M+H] + . Example 38 (R)-2-(4-(1,1-difluoroethyl)-3-fluorophenyl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide [ka]

[0542] 4-Bromo-2-fluoro-N-methoxy-N-methylbenzamide. To a solution of 4-bromo-2-fluorobenzoic acid (1.0 g) in DCM (15 mL) was added HATU (2.26 g), methoxy(methyl)amine hydrochloride (4.54 g), and TEA (1.9 mL). The reaction mixture was stirred at room temperature overnight and then diluted with DCM and water. The organic layer was separated, washed with saturated NaCl solution, and concentrated in vacuo. The residue was purified by silica gel column chromatography eluting with 17% ethyl acetate in petroleum ether to give the title compound 4-bromo-2-fluoro-N-methoxy-N-methylbenzamide (1.166 g) as a yellow oil. 1 H-NMR (400MHz, CDCl3) δ7.37-7.29(m,3H);3.55(s,3H);3.34(s,3H). 19 F-NMR (377MHz, CDCl3)δ -110.95. LCMS ESI(m / z):262(M+H) + .

[0543] 1-(4-Bromo-2-fluorophenyl)ethan-1-one. To a solution of 4-bromo-2-fluoro-N-methoxy-N-methylbenzamide (1.116 g) in THF (10 mL) was added MeMgBr (7.1 mL, 3.0 M in EtO). The reaction mixture was stirred at -78 °C for 2 hours and then at room temperature overnight. The reaction was complete as monitored by LCMS. The reaction was diluted with DCM and water, the two phases were separated, and the aqueous phase was extracted with DCM (30 mL × 3). The combined organic phases were dried over NaSO, filtered, and concentrated to give a residue that was purified by silica gel column chromatography (PE:EA = 10:1) to give 1-(4-bromo-2-fluorophenyl)ethan-1-one (667 mg) as a pale yellow solid. 1H-NMR (400MHz, CDCl3) δ7.77 (t, J = 8.0 Hz, 1H); 7.40-7.34 (m, 2H); 2.63 (d, J = 4.0Hz, 3H). 19 F-NMR (377MHz, CDCl3)δ -107.01(s). [ka]

[0544] Synthesis of 4-bromo-1-(1,1-difluoroethyl)-2-fluorobenzene. To a solution of 1-(4-bromo-2-fluorophenyl)ethan-1-one (667 mg) in BAST (0.8 mL) was added two drops of methanol. Upon completion of the addition, the reaction mixture was placed under a nitrogen atmosphere and heated to 70 °C overnight. The reaction mixture was cooled to room temperature and quenched with 10 mL of water. The resulting mixture was extracted twice with 20 mL of ether. The combined ether phase was washed twice with water, twice with aqueous sodium bicarbonate, once with 10% aqueous citric acid, and once with brine, and then dried over magnesium sulfate. The solvent was removed under reduced pressure, and the mixture was purified on a silica gel column (3% EA in PE) to give 4-bromo-1-(1,1-difluoroethyl)-2-fluorobenzene (294 mg) as a colorless oil. 1 H-NMR (400MHz, CDCl3) δ7.44-7.40(m,1H); 7.36-7.31(m,2H); 2.02-1.93(m,3H). 19 F-NMR(377MHz, CDCl3)δ -87.13;-112.27.

[0545] Synthesis of ethyl 2-(4-(1,1-difluoroethyl)-3-fluorophenyl)acetate. To a solution of 4-bromo-1-(1,1-difluoroethyl)-2-fluorobenzene (290 mg) in mesitylene (5 mL) was added BINAP (45 mg), potassium 1-ethyl-3-propanedioate (310 mg), DMAP (148 mg, 1.21 mmol), and diallylpalladium dichloride (1.6 mg). The reaction mixture was purged with N and stirred at 120 °C overnight. The reaction was complete as monitored by LCMS. The reaction mixture was diluted with EA and water, the two phases were separated, and the aqueous phase was extracted with DCM (10 mL × 3). The combined organic phase was dried over Na2SO4, filtered, and concentrated to give a residue, which was purified by silica gel column chromatography (4% ethyl acetate in petroleum ether) to give ethyl 2-(4-(1,1-difluoroethyl)-3-fluorophenyl)acetate (121 mg) as a pale yellow oil. 1 H-NMR (400MHz, CDCl3) δ7.451-7.47(m,1H);7.11-1.07(m,2H);4.17(q,J=8.0Hz,2H);3.63(s,2H);2.03-1.94(m,3H);1.27(t,J=8.0Hz,3H). 19 F-NMR(377MHz, CDCl3)δ -86.88;-114.99. [ka]

[0546] 2-(4-(1,1-Difluoroethyl)-3-fluorophenyl)acetic acid. To a 25 mL round-bottom flask was added ethyl 2-(4-(1,1-difluoroethyl)-3-fluorophenyl)acetate (121 mg), 1 M aqueous NaOH (3 mL), and MeOH (3 mL). The reaction mixture was stirred at room temperature for 1 h. Then, water (10 mL) was added, and the aqueous layer was washed with EA. The aqueous layer was acidified with 1 M aqueous HCl to pH < 3, and the mixture was extracted with EA. The combined organic phases were washed with saturated NaCl, dried over Na2SO4, filtered, and concentrated to give the product 2-(4-(1,1-difluoroethyl)-3-fluorophenyl)acetic acid (100 mg) as a yellow solid. LCMS ESI (m / z): 217 (MH) + . [ka]

[0547] (R)-2-(4-(1,1-difluoroethyl)-3-fluorophenyl)-N-(1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethyl)acetamide. A solution of 2-(4-(1,1-difluoroethyl)-3-fluorophenyl)acetic acid (100 mg) and HATU (261 mg) in 2 mL of DMF was stirred at room temperature for 15 minutes (Solution A). DIEA (0.46 mL) was added to a solution of (R)-1-(1-(2,2,2-trifluoroethyl)-1H-pyrazolo[3,4-c]pyridin-5-yl)ethan-1-amine hydrochloride (167 mg) in 2 mL of DMF, and the pH of the solution was greater than 7 as determined by moistened pH paper (Solution B). Solution B was added to solution A, and the reaction mixture was stirred at room temperature for 1 hour, at which point LCMS indicated the reaction was complete. The reaction was diluted with EA and water. The two phases were separated, and the aqueous phase was extracted with EA (10 mL × 2). The combined organic phases were dried over Na2SO4, filtered, and concentrated in vacuo. The residue was purified by preparative HPLC (column: AZZOTA C18, 30 × 250 mm × 10 μm; mobile phase: 30% to 95% MeOH + HO (0.1% FA); flow rate: 20 mL / min; wavelen...

Claims

1. Compound of formula (II): 【Chemistry 1】 or a pharma- ceutically acceptable salt thereof, Ring B is C ６ ~C １０ aryl or 5-10 membered heteroaryl; R １ , R ２ and R ３ are each independently H, halogen, CN, OR １１ , N.R. １２ａ R １２ｂ , C １ ~C ６ Alkyl, C ２ ~C ６ Alkoxyalkyl, C １ ~C ６ Haloalkyl, C ２ ~C ６ Alkenyl, C ２ ~C ６ Alkynyl, phenyl, 5-6 membered heteroaryl, C ３ ~C ７ Cycloalkyl, -(CH ２ ) ｎ - (C ３ ~C ７ cycloalkyl), 4- to 7-membered heterocyclyl, or -(CH ２ ) ｎ -(4- to 7-membered heterocyclyl), wherein said phenyl, heteroaryl, cycloalkyl or heterocyclyl optionally comprises 1, 2 or 3 halogen, CN, OR １１ , N.R. １２ａ R １２ｂ , C １ ~C ６ Alkyl, C １ ~C ６ Alkoxy, C １ ~C ６ Alkylamino, or C １ ~C ６ optionally substituted by haloalkyl; R ４ａ and R ４ｂ are each independently H or C １ ~C ６ alkyl or R ４ａ and R ４ｂ and the carbon atom to which they are bonded are C ３ ~C ５ Forming a cycloalkyl; R ６ is H, C １ ~C ６ Alkyl, or C １ ~C ６ haloalkyl; X １ and X ５ are each independently N or CR ８ where X １ and X ５ At least one of the following is CR ８ and X ２ , X ３ and X ４ are each independently N, NR ９ , O, S or CR ９ where X ２ , X ３ and X ４ At least one of the following is N or NR ９ , O, or S; R ８ is H, halogen, CN, OR １１ , C １ ~C ６ Alkyl, C ２ ~C ６ Alkoxyalkyl, or C １ ~C ６ haloalkyl; R ９ is H, C １ ~C ６ Alkyl, C ２ ~C ６ Alkoxyalkyl, C １ ~C ６ Haloalkyl, C ２ ~C ６ Alkenyl, C ２ ~C ６ Alkynyl, C ３ ~C ７ cycloalkyl, or 4- to 7-membered heterocyclyl, said cycloalkyl or heterocyclyl optionally containing 1, 2 or 3 halogens, CN, OH, NH ２ , C １ ~C ６ Alkyl, C １ ~C ６ Alkoxy, C １ ~C ６ Alkylamino, or C １ ~C ６ may be substituted by haloalkyl; Each R １１ , R １２ａ and R １２ｂ are each independently H, C １ ~C ６ Alkyl, C ２ ~C ６ Alkoxyalkyl, C １ ~C ６ Haloalkyl, C ２ ~C ６ Alkenyl, C ２ ~C ６ Alkynyl, C ３ ~C ７ cycloalkyl, or 4- to 7-membered heterocyclyl, said cycloalkyl or heterocyclyl being optionally substituted by 1, 2 or 3 halogen or CN; and n is 1, 2 or 3; provided that the compound has the following structure: 【Chemistry 2】 【Chemistry 3】 or 【Chemistry 4】 or a pharma- ceutically acceptable salt thereof The compound is provided that it does not have the formula:

2. X １ is CR ８ 2. The compound of claim 1 ,

3. R ８ The compound according to claim 1 or 2, wherein is H.

4. X ２ , X ３ , and X ４ are each independently N, NR ９ , or C.R. ９ where X ２ , X ３ , and X ４ At least one of the following is N or NR: ９ A compound according to any one of claims 1 to 3, provided that

5. Structure of formula (III): 【Chemistry 5】 5. The compound according to any one of claims 1 to 4, having the formula:

6. R １ , R ２ and R ３ are each independently H, halogen, CN, OH, or NH ２ , C １ ~C ６ Alkyl, C １ ~C ６ Alkoxy, C １ ~C ６ Alkylamino, C ２ ~C ６ Alkoxyalkyl, C １ ~C ６ Haloalkyl, or C ３ ~C ７ cycloalkyl, wherein said cycloalkyl optionally contains one, two or three halogens, CN, OH, NH ２ , C １ ~C ６ Alkyl, C １ ~C ６ Alkoxy, C １ ~C ６ Alkylamino, or C １ ~C ６ The compound according to any one of claims 1 to 5, which is optionally substituted by haloalkyl.

7. R １ and R ２ are each independently H, halogen, or C １ ~C ６ Alkyl, C １ ~C ６ Alkoxy, C ２ ~C ６ Alkoxyalkyl, C １ ~C ６ Haloalkyl, or C ３ ~C ７ Cycloalkyl, said cycloalkyl optionally containing 1, 2 or 3 halogens, CN, OH, NH ２ , C １ ~C ６ Alkyl, C １ ~C ６ Alkoxy, C １ ~C ６ Alkylamino, or C １ ~C ６ A compound according to any one of claims 1 to 6, optionally substituted by haloalkyl.

8. R １ and R ２ are each independently H, halogen, or C １ ~C ３ Alkyl, C １ ~C ３ Alkoxy, C ２ ~C ３ Alkoxyalkyl, C １ ~C ３ Haloalkyl, or C ３ ~C ５ cycloalkyl, said cycloalkyl optionally containing 1, 2 or 3 halogens, CN, C １ ~C ３ Alkyl, C １ ~C ３ Alkoxy, C １ ~C ３ Alkylamino, or C １ ~C ３ A compound according to any one of claims 1 to 7, optionally substituted by haloalkyl.

9. R ４ａ and R ４ｂ are each independently H or CH ３ The compound according to any one of claims 1 to 8,

10. R ６ is H or C １ ~C ３ The compound according to any one of claims 1 to 9, which is alkyl.

11. R ６ CH ３ The compound according to any one of claims 1 to 10,

12. Structure of formula (IV): 【Chemistry 6】 12. The compound of any one of claims 1 to 11, having the formula: or a pharma- ceutically acceptable salt thereof.

13. R ９ But, C １ ~C ６ Alkyl, C ２ ~C ６ Alkoxyalkyl, C １ ~C ６ Haloalkyl, C ２ ~C ６ Alkenyl, C ２ ~C ６ Alkynyl, C ３ ~C ７ cycloalkyl, or 4- to 7-membered heterocyclyl, said cycloalkyl or heterocyclyl optionally containing 1, 2 or 3 halogens, CN, OH, NH ２ , C １ ~C ６ Alkyl, C １ ~C ６ Alkoxy, C １ ~C ６ Alkylamino, or C １ ~C ６ A compound according to any one of claims 1 to 12, optionally substituted by haloalkyl.

14. R ９ But, C １ ~C ６ Alkyl, C ２ ~C ６ Alkoxyalkyl, C １ ~C ６ Haloalkyl, C ３ ~C ７ cycloalkyl, or 4- to 7-membered heterocyclyl, said cycloalkyl or heterocyclyl optionally containing 1, 2, or 3 halogens, CN, OH, NH ２ , C １ ~C ６ Alkyl, C １ ~C ６ Alkoxy, C １ ~C ６ Alkylamino, or C １ ~C ６ A compound according to any one of claims 1 to 13, optionally substituted by haloalkyl.

15. The compound of any one of claims 1 to 14, wherein Ring B is phenyl or 5-6 membered heteroaryl.

16. The compound of any one of claims 1 to 15, wherein Ring B is phenyl or pyridyl.

17. Structure of formula (V): 【Chemistry 7】 17. The compound of any one of claims 1 to 16, having the formula: or a pharma- ceutically acceptable salt thereof.

18. The compound of any one of claims 1 to 17 having the structure of any one of the compounds in Table 1 or Table 2.

19. A pharmaceutical composition comprising a compound according to any one of claims 1 to 18, or a pharma- ceutically acceptable salt thereof, and a pharma- ceutically acceptable carrier.

20. 20. A pharmaceutical composition for treating a neurodegenerative disease in a subject, comprising a compound according to any one of claims 1 to 18 or a pharma- ceutically acceptable salt thereof.

21. A method for reducing Miro1 levels in a cell, comprising contacting the cell with an effective amount of a T-type calcium channel antagonist.

22. The T-type calcium channel antagonist has the structure of formula (I): 【Chemistry 8】 or a pharma- ceutically acceptable salt thereof, Ring A and ring B each independently represent C ６ ~C １０ aryl or 5-10 membered heteroaryl; R １ , R ２ and R ３ are each independently H, halogen, CN, OR １１ , N.R. １２ａ R １２ｂ , C １ ~C ６ Alkyl, C ２ ~C ６ Alkoxyalkyl, C １ ~C ６ Haloalkyl, C ２ ~C ６ Alkenyl, C ２ ~C ６ alkynyl, phenyl, 5-6 membered heteroaryl, C ３ ~C ７ cycloalkyl, or 4- to 7-membered heterocyclyl, wherein said phenyl, heteroaryl, cycloalkyl, or heterocyclyl optionally contains 1, 2, or 3 halogen, CN, OR １１ , N.R. １２ａ R １２ｂ , C １ ~C ６ Alkyl, C １ ~C ６ Alkoxy, C １ ~C ６ Alkylamino, or C １ ~C ６ optionally substituted by haloalkyl; R ４ａ and R ４ｂ are each independently H or C １ ~C ６ is alkyl; R ５ is H or C １ ~C ６ is alkyl; R ６ａ and R ６ｂ are each independently H or C １ ~C ６ is alkyl; R ７ is H, halogen, CN, OR １１ , N.R. １２ａ R １２ｂ , C １ ~C ６ Alkyl, C ２ ~C ６ Alkoxyalkyl, C １ ~C ６ Haloalkyl, C ２ ~C ６ Alkenyl, C ２ ~C ６ Alkynyl, phenyl, 5-6 membered heteroaryl, C ３ ~C ７ cycloalkyl, or 4- to 7-membered heterocyclyl, wherein said phenyl, heteroaryl, cycloalkyl, or heterocyclyl optionally contains 1, 2, or 3 halogen, CN, OR １１ , N.R. １２ａ R １２ｂ , C １ ~C ６ Alkyl, C １ ~C ６ Alkoxy, C １ ~C ６ Alkylamino, or C １ ~C ６ optionally substituted by haloalkyl; R ８ is H, C １ ~C ６ Alkyl, C ２ ~C ６ Alkoxyalkyl, C １ ~C ６ Haloalkyl, C ２ ~C ６ Alkenyl, C ２ ~C ６ Alkynyl, C ３ ~C ７ cycloalkyl, or 4- to 7-membered heterocyclyl, said cycloalkyl or heterocyclyl optionally containing 1, 2, or 3 halogens, CN, OH, NH ２ , C １ ~C ６ Alkyl, C １ ~C ６ Alkoxy, C １ ~C ６ Alkylamino, or C １ ~C ６ optionally substituted by haloalkyl; R ９ is H, C １ ~C ６ Alkyl, C ２ ~C ６ Alkoxyalkyl, C １ ~C ６ Haloalkyl, C ２ ~C ６ Alkenyl, C ２ ~C ６ Alkynyl, C ３ ~C ７ cycloalkyl, or 4- to 7-membered heterocyclyl, said cycloalkyl or heterocyclyl optionally containing 1, 2, or 3 halogens, CN, OH, NH ２ , C １ ~C ６ Alkyl, C １ ~C ６ Alkoxy, C １ ~C ６ Alkylamino, or C １ ~C ６ optionally substituted by haloalkyl; and Each R １１ , R １２ａ and R １２ｂ are each independently H, C １ ~C ６ Alkyl, C ２ ~C ６ Alkoxyalkyl, C １ ~C ６ Haloalkyl, C ２ ~C ６ Alkenyl, C ２ ~C ６ Alkynyl, C ３ ~C ７ 22. The method of claim 21, wherein said cycloalkyl or heterocyclyl is optionally substituted with 1, 2 or 3 halogen or CN.

23. The T-type calcium channel antagonist having the structure of formula (II): 【Chemistry 9】 or a pharma- ceutically acceptable salt thereof, Ring B is C ６ ~C １０ aryl or 5-10 membered heteroaryl; R １ , R ２ and R ３ are each independently H, halogen, CN, OR １１ , N.R. １２ａ R １２ｂ , C １ ~C ６ Alkyl, C ２ ~C ６ Alkoxyalkyl, C １ ~C ６ Haloalkyl, C ２ ~C ６ Alkenyl, C ２ ~C ６ Alkynyl, phenyl, 5-6 membered heteroaryl, C ３ ~C ７ Cycloalkyl, -(CH ２ ) ｎ - (C ３ ~C ７ cycloalkyl), 4- to 7-membered heterocyclyl, or -(CH ２ ) ｎ -(4- to 7-membered heterocyclyl), wherein said phenyl, heteroaryl, cycloalkyl or heterocyclyl optionally comprises 1, 2 or 3 halogen, CN, OR １１ , N.R. １２ａ R １２ｂ , C １ ~C ６ Alkyl, C １ ~C ６ Alkoxy, C １ ~C ６ Alkylamino, or C １ ~C ６ optionally substituted by haloalkyl; R ４ａ and R ４ｂ are each independently H or C １ ~C ６ alkyl or R ４ａ and R ４ｂ and the carbon atom to which they are attached are C ３ ~C ５ Forming a cycloalkyl; R ６ is H, C １ ~C ６ Alkyl, or C １ ~C ６ haloalkyl; X １ and X ５ are each independently N or CR ８ where X １ and X ５ At least one of the following is CR ８ and X ２ , X ３ and X ４ are each independently N, NR ９ , O, S or CR ９ where X ２ , X ３ and X ４ At least one of the following is N or NR ９ , O, or S; R ８ is H, halogen, CN, OR １１ , C １ ~C ６ Alkyl, C ２ ~C ６ Alkoxyalkyl, or C １ ~C ６ haloalkyl; R ９ is H, C １ ~C ６ Alkyl, C ２ ~C ６ Alkoxyalkyl, C １ ~C ６ Haloalkyl, C ２ ~C ６ Alkenyl, C ２ ~C ６ Alkynyl, C ３ ~C ７ cycloalkyl, or 4- to 7-membered heterocyclyl, said cycloalkyl or heterocyclyl optionally containing 1, 2 or 3 halogens, CN, OH, NH ２ , C １ ~C ６ Alkyl, C １ ~C ６ Alkoxy, C １ ~C ６ Alkylamino, or C １ ~C ６ may be substituted by haloalkyl; Each R １１ , R １２ａ and R １２ｂ are each independently H, C １ ~C ６ Alkyl, C ２ ~C ６ Alkoxyalkyl, C １ ~C ６ Haloalkyl, C ２ ~C ６ Alkenyl, C ２ ~C ６ Alkynyl, C ３ ~C ７ cycloalkyl, or 4- to 7-membered heterocyclyl, said cycloalkyl or heterocyclyl being optionally substituted by 1, 2 or 3 halogen or CN; and 22. The method of claim 21 , wherein n is 1, 2 or 3.

24. The T-type calcium channel antagonist has the structure of formula (III): 【Chemistry 10】 or a pharma- ceutically acceptable salt thereof.

25. The T-type calcium channel antagonist, comprising: 【Chemistry 11】 or a pharma- ceutically acceptable salt thereof.

26. The T-type calcium channel antagonist having the structure of formula (V): 【Chemistry 12】 or a pharma- ceutically acceptable salt thereof.

27. The method of any one of claims 21 to 26, wherein the cell is a muscle cell.

28. The method of any one of claims 21 to 27, wherein the cell is a neuronal cell.

29. 1. A method for identifying a subject at risk for developing a Miro1-associated disorder, comprising: a) detecting whether the Miro1 level in a biological sample obtained from a subject and treated with a mitochondrial stress response factor is equal to or greater than a control Miro1 level in a control biological sample obtained from the subject and untreated; and b) identifying a subject as at risk for developing a Miro1-associated disorder if the Miro1 level in said biological sample is equal to or greater than the control Miro1 level in said control biological sample; Including, The method, wherein the biological sample and the control biological sample contain iPSCs or cells differentiated from iPSCs.

30. 1. A method for identifying a subject at risk for developing a Miro1-associated disorder, comprising: a) detecting whether the Miro1 level in a biological sample obtained from a subject and treated with a mitochondrial stress response factor is equal to or greater than a control Miro1 level in a control biological sample obtained from the subject and untreated; b) identifying the subject as at risk for developing a Miro1-associated disorder if the Miro1 level in the biological sample is equal to or greater than the control Miro1 level in the control biological sample; and c) treating a subject at risk of developing said Miro1-associated disorder by administering a therapeutically effective amount of a compound according to any one of claims 1 to 18 or a pharma- ceutically acceptable salt thereof; A method comprising:

31. A pharmaceutical composition for use in a method for treating a neurodegenerative disease in a subject in need thereof, comprising a therapeutically effective amount of a compound according to any one of claims 1 to 18, or a pharma- ceutical acceptable salt thereof, The method further comprising: a) detecting whether the level of Miro1 in a biological sample obtained from a subject and treated with a mitochondrial stress response factor is equal to or greater than a control Miro1 level in a control biological sample obtained from the subject and untreated; b) identifying the subject as at risk for developing a Miro1-associated disorder if the Miro1 level in the biological sample is equal to or greater than the control Miro1 level in the control biological sample; and c) administering said pharmaceutical composition to said subject. A pharmaceutical composition comprising:

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