Heterocyclic compounds for imaging aggregated tau in tauopathy

Abstract

Provided are heterocyclic compounds of formula I or formula II, or their pharmaceutically acceptable salts, solvates, hydrates, tautomers, or stereoisomers, for imaging tau aggregates. Compounds of formula I or formula II can be used to detect tau aggregates in the diagnosis or monitoring of progression of diseases or disorders such as Alzheimer's disease, corticobasal degeneration, and progressive supranuclear palsy.

Description

[Technical Field]

[0001] Cross-reference with related applications This application claims priority to U.S. Provisional Patent Application No. 63 / 465,817, filed on 11 May 2023. The contents of this application are incorporated herein by reference in their entirety. Statement on federally funded research

[0002] This invention was made with government support under U19NS110456, granted by the National Institutes of Health. The government has certain rights to this invention. [Background technology]

[0003] To gain a deeper understanding of the pathophysiology of tauopathy, such as Alzheimer's disease, progressive supranuclear palsy (PSP), and corticobasal degeneration (CBD), the development of biomarkers to evaluate the accumulation of tau protein in the human brain is essential. Significant progress has been made in the development of positron emission tomography (PET) biomarkers for AD over the past 20 years. While three FDA-approved amyloid-beta plaque PET radiopharmaceuticals have become available in the last decade for broad imaging of AD amyloid-beta plaques, first and second-generation AD tau PET tracers are still under development. While AD aggregates are composed of a mixture of 3R-tau and 4R-tau, PSP and CBD tau aggregates are primarily composed of 4R-tau. Therefore, PET imaging agents developed for imaging tau deposits in AD (such as AV-1451, PI-2620, MK-6240, and PM-PBB3) may not bind to tau deposits in PSP and CBD patients with high affinity. Consequently, the development of high-affinity 4R-tau selective PET imaging agents is one of the biggest challenges in PSP and CBD research.

[0004] There is a need in the art for a 4R-tau selective binder, and this disclosure satisfies that need.

Summary of the Invention

[0005] According to one aspect of the present disclosure, compounds having the structure of Formula I or Formula II, or pharmaceutically acceptable salts, solvates, hydrates, tautomers or stereoisomers thereof are provided.

Chemical Formula

Chemical Formula

[0006] In some embodiments, X1 is N, R1 is H or a halogen, R2 is H, R3 is H or a halogen, R4 is an alkoxy, R5 is a halogen, and R6 is OH. In some embodiments, the halogen is F, Cl, I, or Br. In some embodiments, the halogen is F.

[0007] In some embodiments, the compound has the following structure. [ka] [ka]

[0008] In some embodiments, the compound has the following structure. [ka]

[0009] In some embodiments, any of the compounds described herein selectively bind to 4R tau.

[0010] In some embodiments, any of the compounds described herein include one or more detection labels.

[0011] In some embodiments, the detection label is selected from the group consisting of radionuclides, positron-emitting nuclides, gamma-ray-emitting nuclides, and fluorescent labels.

[0012] In some embodiments, the detection marker is 2 H, 3 H, 18 F, 123 I, 124 I, 125 I, 131 I, 11 C, 13 C, 14 C, 13 N,15 N, 15 O, 75 Br, 76 Br and 77 Selected from the group consisting of Br.

[0013] In some embodiments, the detection marker is 18 It is F.

[0014] In some embodiments, any of the compounds described herein are intended for use in the detection of disorders or diseases associated with tau aggregates.

[0015] In some embodiments, the disease or disorder is a disease or disorder associated with 4R tauopathy or 4R tau aggregates.

[0016] In some embodiments, the disease or disorder is a neurodegenerative disease or disorder.

[0017] In some embodiments, the disorder or disease is selected from the group consisting of Alzheimer's disease, progressive supranuclear palsy, Pick's disease, frontotemporal dementia with parkinsonism-17 (FTDP-17), primary age-related tauopathy (PART) dementia, argyrophilic grain disease, glial globular tauopathy, chronic traumatic encephalopathy (CTE), corticobasal syndrome (CBS), and corticobasal degeneration.

[0018] In some embodiments, detection is performed by one or more of the following: positron emission tomography (PET), single-photon emission computed tomography (SPECT), magnetic resonance imaging, or autoradiography.

[0019] In some embodiments, detection is performed by positron emission tomography (PET).

[0020] In one embodiment, a method is provided for diagnosing a disease or disorder in a subject, or for monitoring the progression of a disease or disorder in a subject. The method comprises: administering a detectable amount of an isotope-labeled compound of formula I or formula II, or a pharmaceutically acceptable salt, solvate, hydrate, tautomer, or stereoisomer thereof, to a subject.

[0021] In some embodiments, the disease or disorder is associated with tau aggregates.

[0022] In some embodiments, the disease or disorder is a disease or disorder associated with 4R tauopathy or 4R tau aggregates.

[0023] In some embodiments, the disease or disorder is a neurodegenerative disease or disorder.

[0024] In some embodiments, the disease or disorder is selected from the group consisting of Alzheimer's disease, progressive supranuclear palsy, Pick's disease, frontotemporal dementia (FTLDs), frontotemporal dementia with parkinsonism-17 (FTDP-17), primary age-related tauopathy (PART) dementia, argyrophilic grain disease, glial globular tauopathy, chronic traumatic encephalopathy (CTE), corticobasal syndrome (CBS), and corticobasal degeneration.

[0025] In some embodiments, the compound is 2 H, 3 H, 18 F, 123 I, 124 I, 125 I, 131 I, 11 C, 13 C, 14 C, 13 N, 15 N, 15 O, 75 Br, 76 Br or 77 It is isotope-labeled with Br.

[0026] In some embodiments, detection is performed by one or more of the following: positron emission tomography (PET), single-photon emission computed tomography (SPECT), magnetic resonance imaging, or autoradiography.

[0027] In all methods described herein, the subject may be human.

[0028] Both the summary above and the following descriptions and detailed descriptions of the drawings are illustrative and explanatory. They are intended to provide further details of the disclosure but are not intended to be restrictive. Other purposes, advantages, and novel features will be readily apparent to those skilled in the art from the following detailed description of the disclosure. [Brief explanation of the drawing]

[0029] [Figure 1] Figure 1 shows the X-ray crystal structure of compound 38.

[0030] [Figure 2] Figure 2 shows the X-ray crystal structure of compound 65.

[0031] [Figure 3] Figure 3 shows the X-ray crystal structure of compound 66.

[0032] [Figure 4] Figure 4 shows the X-ray crystal structure of compound 69 (JSS20-183A).

[0033] [Figure 5] Figure 5 shows brain PET images and time-activity curves of [18F]68 in a male rhesus macaque (12 kg).

[0034] [Figure 6] Figure 6 shows brain PET images and time-activity curves of [18F]69 ([18F]JSS20-183A) in a male rhesus macaque (13 kg).

[0035] [Figure 7] Figure 7 shows postmortem medial frontal cortical sections from PSP, CBD, and AD subjects, and cingulate gyrus sections from a 60-year-old control subject, to which 5 nM [3H]69 ([3H]JSS20-183A) was added, and adjacent sections were autoblocked by adding 1 mM unlabeled 69 (JSS20-183A). [Modes for carrying out the invention]

[0036] Detailed description I. Overview This invention aims to discover novel compounds that selectively bind to tau, particularly 4R-tau. These compounds are useful, for example, in methods for imaging aggregated tau in tauopathy.

[0037] Tau is a protein that stabilizes the internal skeleton of brain nerve cells (neurons). This internal skeleton is tubular in shape, through which nutrients and other essential substances move to various parts of the neuron. Tauopathy is an age-related neurodegenerative disease characterized by the presence of abnormally phosphorylated tau aggregates. In these neurodegenerative diseases, tau detaches from microtubules and forms insoluble aggregates, leading to tauopathy.

[0038] Six different isoforms of the microtubule-associated protein tau exist in the human adult brain: 0N3R, 0N4R, 1N3R, 1N4R, 2N3R, and 2N4R. These isoforms differ in the number of N-terminal insertions (0N, 1N, 2N) and the number of C-terminal repeat domains (3R or 4R), and their expression varies depending on the brain region and developmental stage. In human adults, 3R and 4R tau isoforms are present in nearly equal proportions, with 1N3R and 1N4R being the most abundant.

[0039] As used herein, "tau" refers to a highly soluble microtubule-binding protein primarily found in nerve cells, and includes six major isoforms, as well as truncated or cleaved forms, and other modified forms resulting from phosphorylation, glycosylation, glycation, prolyl isomerization, nitration, acetylation, polyamination, ubiquitination, sumoylation, and oxidation.

[0040] While all tau isoforms can aggregate to form neurofibrillary tangles, some tauopathies, such as Pick's disease and progressive supranuclear palsy, are characterized by the accumulation of specific tau isoforms. Aggregates in AD consist of a mixture of 3R-tau and 4R-tau, while tau aggregates in PSP and CBD consist primarily of 4R-tau (Buee L; Delacourte A, “Comparative biochemistry of tau in progressive supranuclear palsy, corticobasal degeneration, FTDP-17 and Pick's disease,” Brain Pathology 1999, 9(4), 681-693). Expression of the 3R isoform causes deeper axonal transport and motor impairments than that of the 4R isoform, ultimately leading to a shorter lifespan. In contrast, the 4R isoform causes greater neurodegeneration and learning and memory impairments. Tau isoform expression is directly related to brain development. During neurogenesis, only the shortest TAU ​​isoform, 0N3R, is expressed; however, in the adult brain, all six isoforms are present, with 3R and 4R being expressed in nearly equal amounts.

[0041] Diagnosing 4R-tau-specific neurological disorders offers numerous advantages. Developing biomarkers to assess tau protein accumulation in living human brains is crucial for a better understanding of the pathophysiology of tauopathies, not only in Alzheimer's disease (AD) but also in non-Alzheimer's diseases (NAD) such as progressive supranuclear palsy (PSP) and corticobasal degeneration (CBD). 4R-tau biomarkers will not only help clinicians make more definitive disease diagnoses but also assist in confirming target engagement and determining administration regimens for new anti-tau therapies currently under development.

[0042] Over the past 20 years, there has been much progress in the development of positron emission tomography (PET) biomarkers for Alzheimer's disease (AD). While several FDA-approved amyloid-beta-black PET radiopharmaceuticals (AV-1451, PI-2620, MK-6240, PM-PBB3) have become available in the last decade for extensive imaging of amyloid-beta blacks in AD, first and second-generation AD tau PET tracers are still under development. AD aggregates contain a mixture of 3R-tau and 4R-tau, while tau aggregates in PSP and CBD primarily contain 4R-tau. Therefore, PET imaging agents developed to image tau deposits in AD (AV-1451, PI-2620, MK-6240, PM-PBB3) may not bind to tau deposits in PSP and CBD patients with high affinity. Therefore, the development of high-affinity 4R-tau selective PET imaging agents is one of the biggest challenges in PSP and CBD research (Betthauser TJ., “In vitro evidence for a nonselective 4R tau PET tracer,” Mol Psychiatry. 2023 Jan 19. doi:10.1038 / s41380-023-01950-2. Epub ahead of print). For example, [ 18 F]PI-2620 and [ 18PET imaging studies using 18F-APN-1607 in patients with PSP and CBD are being conducted. (Li et al., “Progressive Supranuclear Palsy Neuroimage Initiative (PSPNI). Clinical Utility of 18F-APN-1607 Tau PET Imaging in Patients with Progressive Supranuclear Palsy,” Mov Disord. 2021 Oct;36(10):2314-2323; Tagai et al., “An optimized reference tissue method for quantification of tau protein depositions in diverse neurodegenerative disorders by PET with 18F-PM-PBB3 (18F-APN-1607),” Neuroimage. 2022 Dec 1;264:119763).

[0043] Previously reported tau ligands include the following: [ka] 18 F-flow taucipyr (AV-1451) is a tau PET ligand that binds to paired helical filaments of tau in aging and AD, but its usefulness in detecting aggregates in frontotemporal dementia is unclear (Tsai et al., " 18F-flortaucipir (AV-1451) tau PET in frontotemporal dementia syndromes, "Alzheimer's Research and Therapy, 11(13)(2019)." PI-2620 and MK-6240 are second-generation tau PET tracers (Malarte et al., "Discriminative binding of tau PET tracer PI2620, MK6240, and RO948 in Alzheimer's Disease, corticobasal degeneration and progressive supranuclear palsy brains," Molecular Psychiatry, 28:1272-1283(2023)), as are PM-PBB3 and CBD-2115 (Zhou et al., "Dissecting the Binding Profile of PET Tracer to Corticobasal Degeneration Tau Fibrils," ACS). Chem. Neurosci., 12:3487-3496(2021)).

[0044] However, the in vivo specific signal from existing 4R-tau tracers is quite low, and it is highly likely that improvements in the binding affinity of 4R-tau PET tracers are necessary to improve the imaging signal of 4R-tau biomarkers in PSP and CBD patients. This invention surprisingly and unexpectedly solves this problem that existed in the prior art.

[0045] In one embodiment of this disclosure, the compounds described in formulas I and II selectively bind to 4R tau, as defined by whether the compound binds to other protein binding sites (e.g., aggregated amyloid beta and aggregated α-synuclein). The binding affinity (K) of the compound is defined as follows: d or K i ) was measured at other protein binding sites, and K is approximately 50 nm larger at other binding sites. d and / or Ki The value indicates that the compound does not show a significant specific imaging signal in vivo.

[0046] Non-limiting examples of disorders or diseases associated with tau aggregates, including 4R-tau aggregates, include Alzheimer's disease, progressive supranuclear palsy, Pick's disease, frontotemporal dementia (FTLDs), frontotemporal dementia with parkinsonism-17 (FTDP-17), primary age-related tauopathy (PART) dementia, argyrophilic grain disease, glial spheroid tauopathy, chronic traumatic encephalopathy (CTE), corticobasal syndrome (CBS), and corticobasal degeneration.

[0047] II. Experimental Results In an exemplary embodiment, Enamine Z3777013540 (Z-3540 or Compound 1) was identified by molecular similarity fingerprint search of an Enamine compound library based on CBD-2115. Z-3540 is an exemplary compound according to the present invention. 3 [H]Z-3540 was synthesized and evaluated in binding assays in human CBD, PSP, and AD tissues.

[0048] III.Compound This specification provides heterocyclic compounds for imaging aggregated tau in tauopathy. To date, no high-affinity 4R-tau selective PET tracer suitable for neuroimaging exists.

[0049] The heterocyclic compounds described herein have a structure according to formula I or formula II, or a pharmaceutically acceptable salt, solvate, hydrate, tautomer, or stereoisomer thereof. [ka] [ka] Here, X1 is N, O, or S. X2, X3, X4, and X5 are each independently either N or CH. X6 is CH or O, R1 stands for H, Halogen, CN, Me3N + X7 - , Et3N + X7 - , Pr3N + X7 - Bu3N + X7 - or other suitable ammonium salts, OH (wherein R1 is OH, R1 is not substituted at position 5 of the indole ring), OMs, OTs, OTf, O-nosilates, O-brosilates, or other suitable sulfonate species, SnMe3, SnEt3, SnPr3, SnBu3, or other suitable trialkyltin species, I(O-acyl)2, ArI + X7 - IR7, S(O)Ar, SO2Ar, BF3K, B(OH)2, B(OMe)2, B(OEt)2, B(Opr)2, B(OiPr)2, B(OiPr)3Li, β-pinacol, β-neopentyl glycol, or other suitable boron species. Here, (1) X7 - (2)ArI is a suitable counterion. + X7 - (3) IR7 is a suitable iodonium species known in the field, R2 is H, alkyl, tert-butoxycarbonyl, tosyl, benzyl, p-methoxybenzyl, CBz, Fmoc, or other suitable nitrogen protecting group. R3 stands for H, Halogen, NO2, Me3N + X7 - , Et3N + X7 - , Pr3N + X7 - Bu3N + X7 -or other suitable ammonium salts, OH, OMs, OTs, OTf, O-nosylate, O-brosylate, or other suitable sulfonate species, SnMe3, SnEt3, SnPr3, SnBu3, or other suitable trialkyltin species, I(O-acyl)2, ArI + X7 - , IR7, S(O)Ar, SO2Ar, BF3K, B(OH)2, B(OMe)2, B(OEt)2, B(Opr)2, B(OiPr)2, B(OiPr)3Li, B-pinacol, B-neopentyl glycol, or other suitable boron species, where (1) X7 - is a suitable counterion, (2) ArI + X7 - is a suitable iodonium species known in the art, and (3) IR7 is a suitable iodonium ylide known in the art, R4 is H, halogen, OH, alkoxy, carbonyl, or OEtOEtF, R5 is H, halogen, OH, OMs, OTs, OTf, O-nosylate, O-brosylate, or other suitable sulfonate species, R6 is H, halogen, OH, O-acyl, O-alkyl, OBoc, or other suitable oxygen protecting species, or R5 and R6 are an epoxide: [Chemical formula] form m and n are each independently 0 or 1, and o is 0, 1, or 2.

[0050] In some embodiments, X1 is N, R1 is H or halogen, R2 is H, R3 is H or halogen, R4 is alkoxy, R5 is halogen, and R6 is OH. In some embodiments, the halogen is F, Cl, I, or Br. In some embodiments, the halogen is F.

[0051] The compound of formula I may have one of the following structures. [ka] [ka]

[0052] The compound of formula II may have one of the following structures. [ka]

[0053] In some embodiments, the compound of formula I has one of the following structures. [ka]

[0054] In some embodiments, the compound of formula II has the following structure. [ka]

[0055] Salts of the compounds of formula I or formula II may be pharmaceutically acceptable salts. Other salts may be useful in the preparation of the compounds according to this disclosure or their pharmaceutically acceptable salts.

[0056] If the compounds of this disclosure are acidic, a suitable “pharmaceutically acceptable salt” means a salt prepared in the form of a pharmaceutically acceptable non-toxic base, including inorganic and organic bases. Salts derived from inorganic bases include aluminum salts, ammonium salts, calcium salts, copper salts, ferric salts, ferrous salts, lithium salts, magnesium salts, manganese(III) salts, manganese(II) salts, potassium salts, sodium salts, and zinc salts. Particularly preferred are ammonium salts, calcium salts, magnesium salts, potassium salts, and sodium salts. Salts derived from pharmaceutically acceptable organic non-toxic bases include primary, secondary, and tertiary amines, substituted amines including naturally occurring substituted amines, cyclic amines, and salts of basic ion exchange resins, such as arginine, betaine, caffeine, choline, N-dibenzylethylenediamine, diethylamine, 2-diethylaminoethanol, 2-dimethylaminoethanol, ethanolamine, ethylenediamine, N-ethylmorpholine, N-ethylpiperidine, glucamine, glucosamine, histidine, hydravamin, isopropylamine, lysine, methylglucamine, morpholine, piperazine, piperidine, polyamine resins, procaine, purines, theobromine, triethylamine, trimethylamine, tripropylamine, and tromethamine.

[0057] When the compound of the present invention is basic, the salt can be prepared from pharmaceutically acceptable non-toxic acids, including inorganic and organic acids. Such acids include acetic acid, benzenesulfonic acid, benzoic acid, camphorsulfonic acid, citric acid, ethanesulfonic acid, fumaric acid, gluconic acid, glutamic acid, hydrobromic acid, hydrochloric acid, isethionic acid, lactic acid, maleic acid, malic acid, mandelic acid, methanesulfonic acid, mucinic acid, nitric acid, pamoic acid, pantothenic acid, phosphoric acid, succinic acid, sulfuric acid, tartaric acid, and p-toluenesulfonic acid. Particularly preferred are citric acid, hydrobromic acid, hydrochloric acid, maleic acid, phosphoric acid, sulfuric acid, and tartaric acid.

[0058] An "amine protecting group" is any protecting group suitable for protecting an amine group during a planned chemical reaction. Examples of suitable protecting groups are well known to those skilled in the art. These include, but are not limited to, carbamates, amides, imides, N-alkylamines, N-arylamines, imines, and enamines. Suitable protecting groups are described, for example, in the textbook Greene and Wuts, Protecting groups in Organic Synthesis, third edition, pages 494-653, which is included herein by reference. Specific preferred examples of amine protecting groups include tert-butyloxycarbonyl (Boc), trimethylsilylethoxycarbonyl (Teoc), carbobenzyloxy (Cbz), p-methoxybenzylcarbonyl (Moz or MeOZ), 9-fluorenylmethyloxycarbonyl (Fmoc), acetyl (Ac), or trifluoroacetyl.

[0059] Compounds of formula I or II may be labeled. “Label” (which may be a radioactive label or other detectable label, or a tag, marker, detectable marker, tracer, radiotracer, or equivalent) means any atom or group suitable for in vivo or in vitro imaging and / or assay (e.g., identification, imaging, diagnosis, evaluation, detection and / or quantification), and particularly suitable for imaging and diagnosis. Suitable labels include, for example, radioisotopes (which may also be called “radioactive labeled atoms”), radionuclides, isotopes, positron-emitting nuclides, gamma-ray-emitting nuclides, fluorescent groups, luminescent groups, chromogenic groups, biotin (in combination with streptavidin complexing), or photoaffinity groups. The type of label chosen depends on the desired detection method. The location on which the label is incorporated into or bound to the compound of the present invention is not particularly limited.

[0060] Examples of isotopes (such as radioisotopes, radionuclides, positron-emitting nuclides, and gamma-ray-emitting nuclides) that may be used to label the compounds of the present invention include: 2 H,3 H, 18 F, 123 I, 124 I, 125 I, 131 I, 11 C, 13 C, 14 C, 13 N, 15 N, 15 O, 75 Br, 76 Br and 77 Br are included, but not limited to these. In some embodiments, the detection label is 18 F.

[0061] In some embodiments, any of the compounds disclosed herein includes one or more detection labels. In some embodiments, the R1 and R3 substituents are detectably labeled. In some embodiments, in any one of the compounds described herein, one or more H, one or more C, one or more N, one or more O, one or more F, one or more Br, and / or one or more I are 3 H; 11 C, 13 C or 14 C; 13 N; 15 O; 18 F; 75 Br or 76 Br; or 123 I, 124 I, 125 I, 131 I may be substituted. In some embodiments, the detection label is 18 F.

[0062] In some embodiments, the compounds of the Disclosure selectively bind to 4R-tau. In some embodiments, the compounds of the Disclosure can bind to 4R-tau aggregates both in vivo and in vitro. In some embodiments, the compounds of the Disclosure may be used as diagnostic agents (for in vivo and / or in vitro diagnosis) in the diagnosis of diseases or disorders associated with 4R-tau aggregates (i.e., tauopathies). In some embodiments, the compounds of the Disclosure are for use in the detection of diseases or disorders associated with tau aggregates. In some embodiments, the disease or disorder is 4R-tauopathy or a disease or disorder associated with 4R-tau aggregates. In some embodiments, the disease or disorder is a neurodegenerative disease or disorder.

[0063] IV. Method Furthermore, this specification provides methods for diagnosing a disease or disorder in a subject, or for monitoring the progression of a disease or disorder in a subject. In one embodiment, the disease or disorder is a neurological disorder characterized by the accumulation of 4R tau aggregates in the subject.

[0064] In another embodiment, the method includes diagnosing subjects having non-AD tauopathy characterized by the accumulation of 4R tau aggregates in the subject.

[0065] In yet another embodiment, this includes methods for diagnosing subjects who have a disease or disorder other than AD, such as methods for ruling out a diagnosis of AD.

[0066] In one embodiment, the method includes diagnosing a subject having a neurological disorder characterized by the accumulation of 4R tau aggregates in the subject, and then treating the disorder or disability. The disorder or disability may be, for example, progressive supranuclear palsy, Pick's disease, frontotemporal dementia (FTLDs), frontotemporal dementia with parkinsonism-17 (FTDP-17), primary age-related tauopathy (PART) dementia, argyurophilic grain disease, glial spheroid tauopathy, chronic traumatic encephalopathy (CTE), corticobasal syndrome (CBS), or corticobasal degeneration.

[0067] The methods of the present disclosure may include (a) administering a detectable amount of an isotope-labeled compound of formula I or formula II, or a pharmaceutically acceptable salt, solvate, hydrate, tautomer, or stereoisomer thereof, to a subject; and (b) detecting the binding of the compound to tau aggregates in the patient, particularly to 4R-tau aggregates present in the patient.

[0068] In some embodiments, detection is performed by one or more of the following: positron emission tomography (PET), single-photon emission computed tomography (SPECT), magnetic resonance imaging, or autoradiography.

[0069] V. Definition As used herein, the term "alkyl" refers to both linear and branched saturated hydrocarbon groups. Examples of alkyl groups include methyl, ethyl, n-propyl, iso-propyl, n-butyl, t-butyl, i-butyl, sec-butyl, pentyl, and hexyl groups. Examples of unbranched alkyl groups include, but are not limited to, methyl, ethyl, n-propyl, and n-butyl groups. Examples of branched alkyl groups include, but are not limited to, isopropyl, tert-butyl, isobutyl, 1-ethylpropyl, and 1-ethylbutyl groups.

[0070] As used herein, the terms "halogen" or "halo" refer to fluorine (F), chlorine (Cl), bromine (Br), or iodine (I).

[0071] As used herein, the term "alkoxy" refers to an alkyl group consisting of the indicated number of carbon atoms linked via an oxygen bridge.

[0072] As used herein, the terms “heterocyclyl,” “heterocycle,” or “heterocyclic formula” refer to stable 5- to 7-membered monocyclic or stable 8- to 11-membered bicyclic heterocyclic rings, which may be either saturated or unsaturated, and consist of a carbon atom and one to four heteroatoms selected from the group consisting of N, O, and S, and include any bicyclic group formed by the condensation of any of the above-defined heterocyclic formulas with a benzene ring. Heterocyclic rings may be bonded via any heteroatoms or carbon atoms, insofar as they form a stable structure. The terms heterocyclyl, heterocycle, or heterocyclic formula include heteroaryl groups.

[0073] As used herein, “pharmaceutically acceptable” means a compound, material, composition, and / or dosage form that is suitable for use in contact with human and animal tissues within the bounds of sound medical judgment, without excessive toxicity, irritation, allergic reactions, or other problems or complications commensurate with a reasonable benefit / risk ratio.

[0074] Where used herein, “approximately” is to be understood by those skilled in the art and varies to some extent depending on the context in which it is used. In cases where there is a use of a term that is unclear to those skilled in the art, considering the context in which the term is used, “approximately” means within plus or minus 10% of the given term.

[0075] The use of “a,” “an,” “the,” and similar reference terms in the context of describing elements (particularly in the context of the claims below) shall be construed to cover both singular and plural forms unless otherwise indicated herein or unless the context clearly contradicts it. Unless otherwise indicated herein, the descriptions of ranges of values ​​are intended to function simply as abbreviations for each distinct value that falls within the range, and each distinct value is incorporated herein as if it were written separately herein. All methods described herein may be performed in any suitable order unless otherwise indicated herein or unless the context clearly contradicts it. Any examples or use of exemplary language (e.g., “like”) provided herein are intended simply to make embodiments clearer and, unless otherwise stated, do not limit the claims. Nothing in this specification should be construed as indicating that any unclaimed component is essential.

[0076] Various embodiments are described below. Note that no particular embodiment is intended to be exhaustive or limiting to the broader aspects described herein. One aspect described in relation to a particular embodiment is not necessarily limited to that embodiment and may be implemented in conjunction with any other embodiment(s).

[0077] Therefore, although the present invention is described in general terms, it will be more readily understood by referring to the following examples. These examples are provided for illustrative purposes only and are not intended to limit the present invention. [Examples]

[0078] Example 1. Identification of compounds with high affinity for aggregated 4R-tau.

[0079] The objective of this example was to identify compounds that have a high affinity for aggregated 4R-tau.

[0080] Three commercially available 6-fluoroindole-based compounds 1-3 (listed below) were found to have high affinity for aggregated 4R-tau in human brain tissue samples from patients with AD, PSP, and CBD (Table 1). [ka] [Table 1]

[0081] These compounds were identified by combining ultra-high-throughput in silico screening with in vitro binding assays for "hit confirmation." Subsequent chemosynthesis and structure-activity relationship studies identified numerous 4R-tau ligands (listed below) that exhibited high affinity in human brain tissue samples from patients with AD, PSP, and CBD (Table 2 below). [ka]

[0082] Example 2. Synthesis of 4R-tauligand

[0083] The objective of this example was to synthesize various 4R-tau ligands.

[0084] NMR spectra were obtained at specified frequencies using a Bruker Avance III spectrometer. Spectra in CDCl3 are referenced to internal TMS, while spectra in other deuterated solvents are referenced to solvent residual protons.

[0085] Compounds 4 and 5 (Scheme 1) were prepared according to the previously described procedure (Kumar, A., et al.; Synthesis 2008, 707-710). Compound 6, on the other hand, is commercially available. Compounds 4-6 were coupled with 5-bromo-2-chloropyrimidine under Pd catalysis to obtain compounds 7-11, and all N-Boc protected (7, 9, and 11) and N-deprotected (8 and 10) compounds were isolated. Compound 7 was reacted with either piperidine, 4-fluoropiperidine hydrochloride, or 4,4-difluoropiperidine hydrochloride to obtain compounds 12, 13, and 14, respectively, and then deprotected with TFA to obtain compounds 15, 16, and 17, respectively. Compound 10, and Boc-protected compounds 7 and 11 were reacted with trans / racemic-4-fluoro-3-hydroxypyrrolidine hydrochloride to obtain compounds 18, 19, and 20, respectively. The N-Boc protecting groups of 19 and 20 were then removed with TFA to obtain compounds 21 and 22, respectively. 5-bromo-2-chloropyrimidine was reacted with various cyclic amines (Scheme 2) to obtain compounds 23, 24, (R)-25, and 26. Compound 26 was oxidized with Dess-Martin periodinane to obtain compound 27. 2,5-dibromopyridine was reacted with piperidine or morpholine (Scheme 3) to give compounds 28 and 29, respectively. 2,5-dibromopyrazine was reacted with morpholine (Scheme 3) to give compound 30. Pd-catalyzed coupling (Scheme 4) of compound 4 with compounds 23, (R)-25, 27, 28, and 29 yielded N-Boc-protected compounds 31, (R)-32, 33, 34, and 35, and N-deprotected compound 36, respectively. Next, compounds 31-35 were deprotected with TFA to give compounds 36, (R)-37, 38, 39, and 40, respectively (the structure of 38 was confirmed by X-ray crystallography, Figure 1). Commercially available compounds 41, 6, and 42 (Scheme 5) were coupled with compound 26 to give compounds 43, 44, and 45, respectively. Next, compounds 44 and 45 were deprotected with TFA to give compounds 46 and 47, respectively.

[0086] [ka]

[0087] [ka]

[0088] [ka]

[0089] [ka]

[0090] [ka]

[0091] 2,6-difluoropyridine and 2-chloro-6-nitropyridine were reacted with piperidine or morpholine, respectively (Scheme 6) to obtain compounds 48, 49, 50, and 51, which were then brominated with NBS to obtain compounds 52, 53, 54, and 55 (Smith, AC, et al, Organic Process Research & Development 2018, 22, 681-696). 2-bromo-4-fluoropyridine was reacted with morpholine (Scheme 7) to obtain compound 56, which was then brominated to obtain compound 57. 2,5-dibromo-3-fluoropyridine, 5-bromo-2-chloro-3-fluoropyridine, and 5-bromo-2-chloro-4-fluoropyridine (Scheme 8) were coupled with piperidine or morpholine under Pd catalysis, respectively (Ji, J., et al, Organic Letters 2003, 5, 4611-4614; Stroup, BW, et al, Organic Letters 2007, 9, 2039-2042) to obtain compounds 58, 59, 60, 61, and 62.

[0092] [ka]

[0093] [ka]

[0094] [ka]

[0095] Compounds 52-55 were coupled with compound 6 under Pd catalysis (Scheme 9) to obtain N-Boc-protected compounds 63, 64, 65, and 66, along with N-deprotected compound 67, respectively. Next, compounds 63 and 64 were deprotected with TFA to obtain compounds 68 and 69 (JSS20-183A), respectively. X-ray crystallography confirmed the structures of compound 65 (Figure 2), compound 66 (Figure 3), and compound 69 (Figure 4).

[0096] [ka]

[0097] Compound 42 was coupled with compounds 24 and 29 (Scheme 10) to obtain compounds 70 and 71, respectively, which were then deprotected with TFA to obtain compounds 72 and 73, respectively. Compound 4 was coupled with 52 (Scheme 11) to obtain compound 74, which was then deprotected with TFA to obtain compound 75. Compound 76 was coupled with compounds 30 and 57-59 (Scheme 11) to obtain compounds 77, 78, 79, and 80, respectively, which were then deprotected with TFA to obtain compounds 81, 82, 83, and 84, respectively. Compound 6 was coupled with compounds 57-59 (Scheme 11) to obtain compounds 85, 86, and 87, respectively, which were then deprotected with TFA to obtain compounds 88, 89, and 90, respectively.

[0098] [ka]

[0099] [ka]

[0100] Compound 6 was coupled with compound 60 (Scheme 12) to obtain compound 91, which was deprotected with TFA to obtain compound 92. Compounds 6 and 76 were coupled with compounds 61 and 62, respectively (Scheme 12) to obtain compounds 93, 94, 95, and 96, respectively, which were deprotected with TFA to obtain compounds 97, 98, 99, and 100, respectively.

[0101] [ka]

[0102] Compound 44 was deprotonated with NaH (Scheme 13), then O-alkylated with 2-(2-fluoroethoxy)ethyl 4-methylbenzenesulfonate to obtain compound 101, which was deprotected with TFA to obtain compound 102. Compound 2 was deprotonated with NaH (Scheme 14), then N-methylated with methyl tosylate to obtain compound 103.

[0103] [ka]

[0104] [ka]

[0105] method

[0106] General matters

[0107] The solvents and reagents used were those provided as they were. NMR spectra were obtained at the specified frequencies using a Bruker Avance III spectrometer. 1The 1H NMR spectra are referenced to internal TMS, while the spectra in DMSO-d6, CD3OD, or acetone-d6 are referenced to residual protons in the solvent. 13 13C NMR spectra are based on the resonance of the solvent. 19 The 1F NMR spectrum is not referenced. Radial chromatography was performed using a Harrison Research Chromatotron (trademark) with a Miles Scientific silica rotor. The silica gel used was Silicycle SiliaFlash (registered trademark) P60 40-63 μm (230-400 mesh).

[0108] chemistry

[0109] tert-butyl 6-fluoro-2-(trimethylstanyl)-1H-indole-1-carboxylate (compound 4) Compound 104 (2.23 g, 9.48 mmol) and Me3SnCl (2.09 g, 10.49 mmol, 1.1 equiv.) were flushed with N2 (g) for 40 minutes, then dissolved in freshly distilled THF (100 mL), and cooled in a CH3CN / dry ice bath. LDA solution (2.0 M THF / heptane / ethylbenzene, 6 mL, 12 mmol, 1.3 equiv.) was added dropwise over 3 minutes, and the reaction mixture was stirred at CH3CN / dry ice temperature for 5 minutes, then warmed to ambient temperature and stirred for 4 hours. H2O (0.5 mL) was added, the mixture was stirred for 5 minutes, and then concentrated to obtain a dark brown oil. CH2Cl2 and hexane were added and removed to obtain a dark green / brown syrup / residue, which was dissolved in CH2Cl2 and purified by vacuum flash chromatography on silica (15 cm h × 4 cm id). A colorless syrup (3.64 g) was slowly solidified with CH2Cl2 / hexane at 25% (200 mL) and 50% (100 mL). The crude product was purified twice by radial chromatography (4 mm silica) and hexane (100 mL) to obtain compound 4 (3.52 g, 93%) as a white crystalline solid. 1H NMR (500MHz, CDCl3) δ7.64(dd, 1H, J=10.5Hz, J=2.0Hz), 7.41(dd, 1H, J=8.5Hz, J=5.5Hz), 6.94(td, 1H, J=9.0Hz, J=2.0Hz), 6.68(t, 1H, 3 J SnH =9.0Hz), 1.70(s, 9H), 0.30(t, 9H, 2 J SnH (=28.0Hz).

[0110] tert-butyl 2-(2-chloropyrimidine-5-yl)-6-fluoro-1H-indole-1-carboxylate (compound 7) and 2-(2-chloropyrimidine-5-yl)-6-fluoro-1H-indole (compound 8) Compound 4 (0.38g, 9.55×10 -4 mol), 5-bromo-2-chloropyrimidine (0.25g, 1.29 mmol, 1.4 equiv.), Pd(PPh3)4 (80mg, 6.92 × 10) -5 The mixture was stirred under reflux with N2 (g) for 15 hours, then cooled to ambient temperature and stirred for 3 hours. The reaction mixture was poured onto dry silica (55 mm h × 45 mm id) and eluted under vacuum. Orange / brown residues (0.33 g) were obtained from hexane (50 mL), CH2Cl2 (100 mL), %MeOH / CH2Cl2-1% (100 mL), 2.5% (200 mL), 5% (100 mL), and 10% (50 mL). Purification by radial chromatography (2 mm silica) and hexane / Â / NEt3v / v / v 90:8:2 (100 mL), 75:20:5 (100 mL), and 50:45:5 (50 mL) yielded compound 7 (57 mg, 17%) as an off-white solid and compound 8 (30 mg, 13%) as a pale yellowish-brown solid.

[0111] compound 7 1H NMR (300MHz, CDCl3) δ8.68(s, 2H), 7.94(dd, 1H, J=10.5Hz, J=2.4Hz), 7.53(dd, 1H, J=8 .4Hz, J=5.4Hz), 7.06(td, 1H, J=8.7Hz, J=2.4Hz), 6.68(d, 1H, J=0.6Hz), 1.48(s, 9H). HRMS(ESI)[M+H] + In C 17 H 16 Calculated value of ClFN3O2: 348.0910, measured value: 348.0902.

[0112] compound 8 1 H NMR (300MHz, acetone-d6) δ11.10(br s, 1H), 9.15(s, 2H), 7.64(dd, 1H, J=8.7Hz, J=5.4Hz), 7.19(m-duplicate resonance, 2H), 6.91(ddd, 1H, J=9.9Hz, J=8.7Hz, J=2.1Hz). HRMS(ESI)[M+H] + In C 12 Calculated value for H8ClFN3: 248.0385, measured value: 248.0382.

[0113] tert-butyl 2-(2-chloropyrimidine-5-yl)-6-cyano-1H-indole-1-carboxylate (compound 9) and 2-(2-chloropyrimidine-5-yl)-1H-indole-6-carbonitrile (compound 10) 1,4-Dioxane was purged with N2 (g) for 40 minutes. Compound 5 (0.46 g, 1.14 mmol), 5-Bromo-2-chloropyrimidine (0.23 g, 1.19 mmol), Pd(dppf)Cl2 (78 mg, 1.07 × 10⁻¹⁴) -4After flushing with N2(g) for 15 minutes with mol (0.09 equiv.) and Na2CO3 (0.14g, 1.32 mmol, 1.2 equiv.), 1,4-dioxane (20 mL) was added. The reaction mixture was stirred under reflux under N2(g) for 4 hours, cooled, and the solvent was removed to obtain a brown residue, which was dissolved in CH2Cl2 and poured onto dry silica (55 mm h × 45 mm id) and eluted under vacuum. Orange / brown residue (0.16 g, crude compound 9) and orange / brown solid (62 mg, crude compound 10) were obtained from CH2Cl2 (100 mL), %MeOH / CH2Cl2-1% (100 mL), 2% (100 mL), 3% (200 mL), and 4% (100 mL).

[0114] compound 9 The residue was dissolved in CH2Cl2 / MeOH and poured onto dry silica (33 mm h × 33 mm id), and eluted under vacuum. A dark orange residue was obtained from 1% (100 mL) and 2% (100 mL) of %MeOH / CH2Cl2, which was dried briefly under vacuum. The residue was dissolved / suspended in CHCl3 (1 mL), filtered, and the precipitate was washed with CHCl3 (1 mL × 2). The filtrate was purified by radial chromatography (1 mm silica) and 75:20:5 v / v / v hexane / Â1 / NEt3 (100 mL) to obtain compound 9 (38 mg, 9%) as a white solid. 1 H NMR (300MHz, CDCl3) δ8.71(s, 2H), 8.58(d, 1H, J=0.6Hz), 7.69(d, 1H, J=8.1Hz), 7.56(dd, 1H, J=8.1Hz, J=1.5Hz), 6.77(s, 1H), 1.49(s, 9H). HRMS(ESI)[M+H] + In C 18 H 16 Calculated value of O2N4Cl: 355.0956, measured value: 355.0958.

[0115] compound 10 The solid was dissolved / suspended in CHCl3 (2.5 mL), filtered, the precipitate was washed with CHCl3 (2.5 mL x 3), and dried under vacuum to obtain compound 10 (29 mg, 10%) as a yellow / orange solid. 1H NMR (400MHz, DMSO-d6) δ12.44(s, 1H), 9.31(s, 2H), 7.96(s, 1H), 7.79(d, 1H, J=8.0Hz), 7.40(d, 1H, J=8.0Hz), 7.36(s, 1H).

[0116] tert-butyl 2-(2-chloropyrimidine-5-yl)-1H-indole-1-carboxylate (compound 11) 1-Boc-Indole-2-boronic acid (compound 6) (0.25g, 9.58 x 10⁻⁶) -4 mol), 5-bromo-2-chloropyrimidine (0.18 g, 9.31 × 10⁻¹⁰ -4 mol), and Pd(dppf)Cl2(52 mg, 7.11 × 10⁻¹⁰) -5 The mixture (mol, 0.08 equiv.) was flushed with N2(g) for 10 minutes, then 1,4-dioxane (10 mL) was added, followed by K2CO3 / H2O solution (1.5 mL, 2 M, 3 mmol, 3.2 equiv.). The reaction mixture was stirred under reflux under N2(g) for 7 hours, cooled to ambient temperature, stirred overnight, filtered through Celite, and the Celite was washed with 1,4-dioxane. The filtrate was concentrated to obtain a brown oil, which was removed by adding CHCl3 and hexane to obtain a brown syrup. This was dissolved in CH2Cl2, poured onto dry silica (45 mm h × 45 mm id), and eluted under vacuum. A yellowish-brown solid (0.19 g) was obtained from CH2Cl2 (200 mL), %MeOH / CH2Cl2-1% (100 mL), 2% (50 mL), and 3% (50 mL). Purification by radial chromatography (2 mm silica) with CH2Cl2 (100 mL) yielded an off-white solid (0.11 g), which was then purified again by radial chromatography (2 mm silica) with hexane / Â / NEt3v / v / v 90:8:2 (35 mL), 85:12:3 (100 mL), and 75:20:5 (50 mL) to obtain compound 11 (82 mg, 27%) as a white solid. 1H NMR (400MHz, CDCl3) δ8.69(s, 2H), 8.20(d, 1H, J=8.4Hz), 7.60(d, 1H, J=7.6Hz), 7.40(ddd, 1H, J= 8.4Hz, J=7.2Hz, J=1.2Hz), 7.30(ddd, 1H, J=7.8Hz, J=7.2Hz, J=0.8Hz), 6.71(s, 1H), 1.49(s, 9H).

[0117] tert-butyl 6-fluoro-2-(2-(piperidine-1-yl)pyrimidine-5-yl)-1H-indole-1-carboxylate (compound 12) Compound 7 (42 mg, 1.21 × 10⁻⁶) -4 (mol), piperidine (0.05 mL, 5.1 × 10) -4 mol, 4.2equiv.), K2CO3 (24mg, 1.74×10 -4 1.4 mol (equiv.) and CH3CN (10 mL) were stirred under reflux with N2 (g) for 75 minutes and then cooled to ambient temperature. CH3CN was removed to obtain a residue, which was dissolved in CH2Cl2 and poured onto dry silica (33 mm h × 33 mm id) and eluted under vacuum. Compound 12 (47 mg, 98%) was obtained as a pale yellowish-brown solid from CH2Cl2 (25 mL), %MeOH / CH2Cl2-1% (50 mL), 2% (75 mL), and 3% (50 mL). 1 H NMR (500MHz, CDCl3) δ8.33(s, 2H), 7.93(dd, 1H, J=10.5Hz, J=2.5Hz), 7.45(dd, 1H, J=8.5Hz, J=5.5Hz), 7. 01(td, 1H, J=8.5Hz, J=2.5Hz), 6.48(s, 1H), 3.84(t, 4H, J=5.0Hz), 1.71(m, 2H), 1.62(m, 4H), 1.48(s, 9H).

[0118] tert-butyl 6-fluoro-2-(2-(4-fluoropiperidine-1-yl)pyrimidine-5-yl)-1H-indole-1-carboxylate (compound 13) Compound 7 (60mg, 1.73×10 -4 mol), 4-fluoropiperidine·HCl (34 mg, 2.44 × 10⁻⁶)-4 mol, 1.4equiv.), K2CO3 (88mg, 6.37×10 -4 (mol, 3.7 equiv.) and CH3CN (5 mL) were stirred under reflux with N2 (g) for 5 hours and then cooled. The solvent was removed to obtain the residue, which was dissolved in CH2Cl2 and poured onto dry silica (33 mm h × 33 mm id). Elution was performed under vacuum with CH2Cl2 (125 mL) to obtain compound 13 (64 mg, 89%) as a white foam. 1 H NMR (300MHz, CDCl3) δ8.35(s, 2H), 7.93(dd, 1H, J=10.8Hz, J=2.4Hz), 7.46(dd, 1H, J=8.7Hz, J=5.7Hz), 7.01(td , 1H, J=8.7Hz, J=2.4Hz), 6.50(s, 1H), 4.99(tt, 0.5H, J=6.3Hz, J=3.3Hz) and 4.83(dt, 0.5H, J=9.3Hz, J=4.8Hz)( 2 J HF =48.3Hz), 3.97(t, 4H, J=5.7Hz), 1.93(m, 4H), 1.48(s, 9H). HRMS(ESI)[M+H] + In C 22 H 25 Calculated value of F2N4O2: 415.1940, measured value: 415.1944.

[0119] tert-butyl 2-(2-(4,4-difluoropiperidine-1-yl)pyrimidine-5-yl)-6-fluoro-1H-indole-1-carboxylate (compound 14) Compound 7 (94.5mg, 2.72×10 -4 mol), 4,4-difluoropiperidine·HCl (60 mg, 3.81 × 10⁻¹⁰ -4(mol, 1.4 equiv.), K2CO3 (186 mg, 1.35 mmol, 5 equiv.), and CH3CN (20 mL) were stirred under reflux with N2 (g) for 21 hours, then cooled to ambient temperature, filtered, and the precipitate was washed with CH3CN. After removing the CH3CN, an orange residue was obtained, which was then removed by adding CH2Cl2 and hexane, and the mixture was dried under vacuum to obtain a pale orange solid. The solid was dissolved in CH2Cl2, poured onto dry silica (45 mm h × 45 mm id), and eluted under vacuum with CH2Cl2 (350 mL) to obtain a colorless residue. Purification by radial chromatography (2 mm silica) with %CH2Cl2 / hexane-25% (50 mL) and 50% (125 mL) yielded a white solid (46 mg). This was further purified by radial chromatography (1 mm silica) with %CH2Cl2 / hexane-25% (100 mL), 35% (100 mL), and 50% (50 mL) to obtain compound 14 (32 mg, 27%) as a white solid. 1 H NMR (400MHz, CDCl3) δ8.37(s, 2H), 7.93(dd, 1H, J=10.8Hz, J=2.0Hz), 7.46(dd, 1H, J=8.4Hz, J=5.6Hz), 7.0 2(td, 1H, J=8.8Hz, J=2.0Hz), 6.51(s, 1H), 4.03(t, 4H, J=6.0Hz), 2.02(septet, 4H, J=6.0Hz), 1.48(s, 9H). HRMS(ESI)[M+H] + In C 22 H 24 Calculated value for O2N4F3: 433.1846, measured value: 433.1836.

[0120] 6-Fluoro-2-(2-(piperidine-1-yl)pyrimidine-5-yl)-1H-indole (compound 15) Compound 12 (41 mg, 1.03 × 10⁻⁶) -4The compound (mol) was dissolved in TFA (1 mL, 13 mmol, 126 equiv.), stirred at ambient temperature for 15 minutes, and poured into a mixture of NaHCO3 (1.23 g, 14.64 mmol, 1.1 equiv. TFA), H2O (30 mL), and CH2Cl2 (30 mL). The mixture was stirred, the layers were separated, and the aqueous layer was extracted with CH2Cl2 (10 mL). The combined CH2Cl2 layers were washed with brine (15 mL) and dried on MgSO4. The solution was concentrated and poured onto dry silica (33 mm h × 33 mm id), and eluted under vacuum. Compound 15 (27 mg, 88%) was obtained as a pale yellow solid from hexane / siRNA / NET3 v / v / v 90:8:2 (50 mL), 75:20:5 (50 mL), and 50:45:5 (75 mL). 1 H NMR (300MHz, CDCl3) δ8.56(s, 2H), 8.20(br s, 1H), 7.49(dd, 1H, J=8.7Hz, J=5.4Hz), 7.08(dd, 1H, J=9.6Hz, J=2.1Hz), 6.88(ddd, 1H, J=9. 6Hz, J=8.7Hz, J=2.4Hz), 6.62(dd, 1H, J=2.1Hz, J=0.9Hz), 3.84(t, 4H, J=5.4Hz), 1.68(m, 6H), 13 C NMR (125MHz, DMSO-d6) δ160.08, 158.71(d, 1 J FC =234.4Hz), 154.49, 136.68(d, J FC =13.0Hz), 134.17(d, J FC =3.4Hz), 125.44, 120.46(d, J FC =10.2Hz), 114.48, 107.73(d, J FC =24.1Hz), 97.05(d, J FC =25.7Hz), 96.91, 44.34, 25.25, 24.27, 19 F NMR (470.6MHz, DMSO-d6) δ-121.61(m). HRMS(ESI)[M+H] + In C 17 H 18 FN4 calculated value: 297.1510, measured value: 297.1515.

[0121] 6-Fluoro-2-(2-(4-Fluoropiperidine-1-yl)pyrimidine-5-yl)-1H-indole (Compound 16). Compound 13 (59mg, 1.42×10 -4 The compound (mol) was dissolved in TFA (1.5 mL, 19.5 mmol, 137 equiv.), stirred at ambient temperature for 20 minutes, and then poured into a mixture of NaHCO3 (1.82 g, 21.67 mmol, 1.1 equiv. TFA), H2O (45 mL), and CH2Cl2 (45 mL). After stirring the mixture until the color disappeared, the layers were separated, and the aqueous layer was extracted with CH2Cl2 (10 mL). The combined CH2Cl2 layers were washed with brine (20 mL) and dried on MgSO4. The solution was concentrated and poured onto dry silica (33 mm h × 33 mm id), and eluted under vacuum. Compound 16 (44 mg, 99%) was obtained as a pale yellowish-brown solid from hexane / siRNA / NET3 v / v / v 90:8:2 (50 mL), 75:20:5 (50 mL), and 50:45:5 (100 mL). 1 H NMR (300MHz, CDCl3) δ8.58(s, 2H), 8.21(br s, 1H), 7.50(dd, 1H, J=8.7Hz, J=5.4Hz), 7.08(dd, 1H, J=9.6Hz, J=2.1Hz), 6.89(ddd, 1H, J=9.6Hz, J=8.7Hz, J=2. 1Hz), 6.64(dd, 1H, J=2.1Hz, J=0.9Hz), 5.00(tt, 0.5H, J=6.0Hz, J=3.3Hz) and 4.83(dt, 0.5H, J=9.6Hz, J=4.8Hz)( 2 J HF =48.3Hz), 3.97(m, 4H), 1.96(m, 4H), 1 H NMR (300MHz, DMSO-d6) δ11.58(s, 1H), 8.83(s, 2H), 7.49(dd, 1H, J=8.7Hz, J=5.4Hz), 7.13(dd, 1H, J =9.9Hz, J=2.1Hz), 6.85(ddd, 1H, J=9.9Hz, J=8.7Hz, J=2.1Hz), 6.82(d, 1H, J=1.8Hz), 4.93(dtt, 1H, 2 J HF=48.6Hz, J=7.2Hz, J=3.6Hz), 3.97(m, 2H), 3.78(m, 2H), 1.93(m, 2H), 1.72(m, 2H). HRMS(ESI)[M+H] + In C 17 H 17 Calculated value for F2N4: 315.1416, measured value: 315.1420.

[0122] 2-(2-(4,4-difluoropiperidine-1-yl)pyrimidine-5-yl)-6-fluoro-1H-indole (compound 17) Compound 14 (27mg, 6.24×10 -5 The compound (mol) was dissolved in TFA (0.6 mL, 7.8 mmol, 125 equiv.), stirred at ambient temperature for 20 minutes, and poured over a mixture of NaHCO3 (0.856 g, 10.19 mmol, 1.3 equiv. TFA), H2O (20 mL), and CH2Cl2 (20 mL). After stirring the mixture for 15 minutes, the layers were separated, and the aqueous layer was extracted with CH2Cl2 (5 mL × 2). The combined CH2Cl2 layers were washed with brine (15 mL) and dried on MgSO4. The solution was poured over dry silica (33 mm h × 33 mm id) and eluted under vacuum. Compound 17 (18 mg, 87%) was obtained as an off-white solid from CH2Cl2 (50 mL) %MeOH / CH2Cl2-1% (50 mL), 2% (25 mL), and 4% (50 mL). 1 H NMR (300MHz, CDCl3) δ8.60(s, 2H), 8.23(br s, 1H), 7.51(dd, 1H, J=8.7Hz, J=5.4Hz), 7.08(dd, 1H, J=9.6Hz, J=2.1Hz), 6.90(ddd, 1H, J=9.6Hz, J=8.7Hz, J=2.1Hz), 6.66(d, 1H, J=1.5Hz), 4.03(t, 4H, J=5.7Hz), 2.04(tt, 4H, 3 J HF =13.5Hz, J=5.7Hz). HRMS(ESI)[M+H] + In C 17 H 16 Calculated value for F3N4: 333.1322, measured value: 333.1314.

[0123] 2-(2-(3-fluoro-4-hydroxypyrrolidine-1-yl)pyrimidine-5-yl)-1H-indole-6-carbonitrile (compound 18) Compound 10 (29mg, 1.14×10 -4 (mol), trans / racemi-4-fluoro-3-hydroxypyrrolidine·HCl (33 mg, 2.33 × 10⁻⁶) -4 mol, 2equiv.), K2CO3 (120mg, 8.68×10 -4 7.6 mol (equiv.) and 10 mL of CH3CN were stirred under reflux with N2 (g) for 3 hours, cooled, and the CH3CN was removed to obtain a dark orange residue. The residue was dissolved / suspended in CH2Cl2 / MeOH, filtered, and the precipitate was washed with CH2Cl2. The solvent was removed from the filtrate to obtain an orange residue, which was dried briefly under vacuum (56 mg). The residue was dissolved in CH2Cl2 / MeOH, poured onto dry silica (33 mm h × 33 mm id), and eluted under vacuum. Compound 18 (9 mg, 24%) was obtained as a pale yellowish-brown solid from 1% (100 mL), 2.5% (100 mL), and 5% (100 mL) of %MeOH / CH2Cl2. 1 H NMR (300MHz, DMSO-d6) δ12.08(s, 1H), 8.92(s, 2H), 7.82(s, 1H), 7.67(d, 1H, J=8.1Hz), 7. 33(dd, 1H, J=8.1Hz, J=1.5Hz), 6.98(d, 1H, J=1.5Hz), 5.61(d, 1H, J=3.9Hz), 5.10(dm, 1H, 2 J HF =49.8Hz), 4.37(m, 1H), 3.88(s, 1H), 3.77(m, 1H), 3.69(s, 2H). HRMS(ESI)[M+H] + In C 17 H 13 FN5O calculated value: 322.1099, measured value: 322.1093.

[0124] tert-butyl 6-fluoro-2-(2-(3-fluoro-4-hydroxypyrrolidine-1-yl)pyrimidine-5-yl)-1H-indole-1-carboxylate (compound 19) Compound 7 (56mg, 1.61×10 -4 mol), trans / racemi-4-fluoro-3-hydroxypyrrolidine·HCl (52 mg, 3.67 × 10⁻⁶) -4 (mol, 2.3 equiv.), K2CO3 (168 mg, 1.22 mmol, 7.5 equiv.), and CH3CN (15 mL) were stirred under reflux with N2 (g) for 3 hours and then cooled to ambient temperature. CH3CN was removed to obtain an off-white solid, which was dissolved / suspended in CH2Cl2, filtered, and the precipitate was washed with CH2Cl2. CH2Cl2 was removed from the filtrate to obtain a yellow syrup, which was dried under vacuum (89 mg), then dissolved in CH2Cl2 / MeOH, poured onto dry silica (45 mm h × 45 mm id), and eluted under vacuum. %MeOH / CH2Cl2 - 1% (50 mL), 2% (75 mL), 3% (100 mL), 4% (50 mL). The desired fractions were combined and purified by radial chromatography (1 mm silica) using %MeOH / CH2Cl2-1% (50 mL), 2% (75 mL), and 3% (25 mL) to obtain compound 19 (41 mg, 61%) as an off-white solid. 1 H NMR (300MHz, CDCl3) δ8.39(s, 2H), 7.91(dd, 1H, J=10.8Hz, J=2.4Hz), 7.46(dd, 1H, J=8.7Hz, J=5.7Hz), 7.01(td, 1H, J=8.7Hz, J=2.4Hz), 5.11(dm, 1H, 2 J HF =51.0Hz), 4.59(dm, 1H, J=6.9Hz), 4.03(m, 1H), 3.90(m, 3H), 1.92(br d, 1H, J=2.7Hz), 1.50(s, 9H).

[0125] tert-butyl 2-(2-(3-fluoro-4-hydroxypyrrolidine-1-yl)pyrimidine-5-yl)-1H-indole-1-carboxylate (compound 20) Compound 11 (79 mg, 2.40 × 10⁻⁶) -4 mol), trans / racemi-4-fluoro-3-hydroxypyrrolidine·HCl (97 mg, 6.85 × 10) -4Compound 20 (85 mg, 89%) was obtained as an off-white foam from CH2Cl2 (25 mL), %MeOH / CH2Cl2-100 (15 mL), K2CO3 (158 mg, 1.14 mmol, 4.7 equiv.), and CH3CN (15 mL) under reflux with N2 (g) for 5 hours. The mixture was then cooled to ambient temperature. The CH3CN was removed to obtain a residue, which was briefly dried under vacuum. This residue was then dissolved in CH2Cl2, poured onto dry silica (33 mm h × 33 mm id), and eluted under vacuum. Compound 20 (85 mg, 89%) was obtained from CH2Cl2 (25 mL), %MeOH / CH2Cl2-1% (50 mL), 2% (75 mL), and 3% (50 mL). 1 ¹H NMR (300MHz, CDCl3) δ 8.41 (s, 2H), 8.17 (d, 1H, J=8.4Hz), 7.55 (d, 1H, J=7.5Hz), 7.33 (partially separated ddd, 1H, J=7.8Hz, J=1.2Hz), 7.25 (m, 1H, partially obscured by CHCl3 resonance), 6.55 (s, 1H), 5.10 (dm, 1H, 2 J HF =50.7Hz), 4.59(m, 1H), 4.03(m, 1H), 3.91(m, 3H), 1.89(d, 1H, J=3.9Hz), 1.50(s, 9H).

[0126] 4-Fluoro-1-(5-(6-Fluoro-1H-Indole-2-yl)pyrimidine-2-yl)pyrrolidine-3-ol (Compound 21) Compound 19 (27mg, 6.48×10 -5 The compound (mol) was dissolved in TFA (0.75 mL, 9.7 mmol, 150 equiv.), stirred at ambient temperature for 15 minutes, and then poured into a mixture of NaHCO3 (0.91 g, 10.83 mmol, 1.1 equiv. TFA), H2O (25 mL), and CH2Cl2 (25 mL). After stirring the mixture for 5 minutes, the layers were separated, and the aqueous layer was extracted with CH2Cl2 (5 mL × 2). The combined CH2Cl2 layer was dried on MgSO4, the solvent was removed to obtain an off-white solid, which was dried under vacuum to obtain compound 21 (17 mg, 83%). 1H NMR (300MHz, acetone-d6) δ10.71(br s, 1H), 8.80(s, 2H), 7.52(dd, 1H, J=8.7Hz, J=5.4Hz), 7.11(dd, 1H, J=9.9Hz, J= 2.1Hz), 6.84(ddd, 1H, J=9.9Hz, J=8.7Hz, J=2.4Hz), 6.79(m, 1H), 5.12(dm, 1H, 2 J HF =51.0Hz), 4.67(d, 1H, J=3.9Hz), 4.51(m, 1H), 3.96(d, 1H, J=2.1Hz), 3.85(m, 1H), 3.79(m, 2H), 13 C NMR (125MHz, DMSO-d6) δ158.74(d, 1 J FC =234.6Hz), 159.10, 154,57, 136.75(d, J FC =12.6Hz), 134.13(d, J FC =3.5Hz), 125.43, 120.51, 115.14, 107.76(d, J FC =24.3Hz), 97.09(d, J FC =25.5Hz), 97.07, 95.16(d, J FC =177.0Hz), 71.40(d, J FC =27.2Hz), 52.55, 50.71(d, J FC (=21.9Hz), 19 F NMR (470.6MHz, DMSO-d6) δ -121.54(m), -181.85(m). HRMS(ESI)[M+H] + In C 16 H 13 Calculated value of F2N4O: 315.1052, measured value: 315.1048.

[0127] 1-(5-(1H-indole-2-yl)pyrimidine-2-yl)-4-fluoropyrrolidine-3-ol (compound 22) Compound 20 (79 mg, 1.98 × 10⁻⁶) -4Dissolve compound 20 (mol) in CH2Cl2 (5 mL), add HCl / 1,4-dioxane (4 M, 1 mL, 4 mmol), stir the mixture for 5 hours, filter, and wash the precipitate with EtOEt (2 mL x 3). Remove the solvent from the filtrate to obtain a pale yellow solid, which was dried briefly under vacuum and dissolved in CH2Cl2 (10 mL). Add H2O (10 mL), followed by concentrated NH4OH (aq) (5 drops), and stir the mixture for 5 minutes (pH 11-12). Add brine (5 mL), mix and separate the layers, and dry the CH2Cl2 layer on MgSO4. Analysis by TLC (3% MeOH / CH2Cl2) revealed that N-Boc protected compound 20 remained. Remove the solvent to obtain a white foamy substance, which was dried under vacuum (50 mg). The foamy substance was dissolved in TFA (2 mL, 26 mmol) and stirred for 15 minutes. This mixture was then poured over a mixture of CH2Cl2 (40 mL), H2O (40 mL), and NaHCO3 (2.42 g, 28.81 mmol, 1.1 equiv. TFA). After stirring the mixture for 15 minutes, the layers were separated, and the H2O layer was extracted with CH2Cl2 (10 mL). The combined CH2Cl2 layer was washed with brine (25 mL), dried on MgSO4, and the solvent was removed to obtain an off-white solid (24 mg). Purification by flash column chromatography on silica (3% MeOH / CH2Cl2) yielded compound 22 (21 mg, 36%) as an off-white solid. 1 ¹H NMR (400MHz, DMSO-d6) δ 11.47 (s, 1H), 8.86 (s, 2H), 7.50 (d, 1H, J=8.0Hz), 7.38 (d, 1H, J=8.0Hz), 7.08 (partially separated ddd, 1H, J=7.2Hz, J=1.2Hz), 6.99 (partially separated ddd, 1H, J=7.2Hz, J=1.2Hz), 6.81 (d, 1H, J=1.2Hz), 5.60 (d, 1H, J=3.6Hz), 5.09 (dm, 1H, 2 J HF =50.8Hz), 4.37(m, 1H), 3.86(s, 1H), 3.79(m, 1H), 3.67(s, 2H). HRMS(ESI)[M+H] + In C 16 H 14 FN4O calculated value: 297.1152, measured value: 297.1145.

[0128] 5-Bromo-2-(4-methoxypiperidine-1-yl)pyrimidine (Compound 23) 5-Bromo-2-chloropyrimidine (0.88 g, 4.55 mmol), 4-methoxypiperidine (0.65 mL, 5.25 mmol, 1.2 equiv.), K2CO3 (0.89 g, 6.44 mmol, 1.4 equiv.), and CH3CN (25 mL) were stirred under reflux for 1 hour under N2 (g), then cooled to ambient temperature, filtered, and the precipitate was washed with CH3CN. After removing CH3CN, a pale yellow residue was obtained, which was dissolved in CH2Cl2, poured onto dry silica (55 mm h × 45 mm id), and eluted under vacuum. Compound 23 (1.21 g, 98%) was obtained as a white solid from CH2Cl2 (50 mL), %MeOH / CH2Cl2-1% (100 mL), 2% (100 mL), 3% (100 mL), and 4% (100 mL). 1 H NMR (300MHz, CDCl3) δ8.27(s, 2H), 4.19(m, 2H), 3.44(m, 3H), 3.39(s, 3H), 1.92(m, 2H), 1.57(m, 2H).

[0129] 4-(5-bromopyrimidine-2-yl)morpholine (compound 24) 5-Bromo-2-chloropyrimidine (1.48 g, 7.65 mmol), morpholine (1 mL, 11.6 mmol, 1.5 equiv.), K2CO3 (1.30 g, 9.41 mmol, 1.2 equiv.), and CH3CN (55 mL) were stirred under reflux with N2 (g) for 3 hours, then cooled to ambient temperature, filtered, and the precipitate was washed with CH3CN. The solvent was removed from the filtrate to obtain a white solid, which was dissolved in CH2Cl2 and poured onto dry silica (55 mm h × 45 mm id) and eluted under vacuum. Compound 24 (1.74 g, 93%) was obtained as a white solid from CH2Cl2 (100 mL), %MeOH / CH2Cl2-1% (100 mL), 2% (100 mL), 3% (100 mL), and 4% (100 mL). 1 H NMR (300MHz, CDCl3) δ8.31(s, 2H), 3.76(s, 8H), 13C NMR (125MHz, CDCl3) δ160.15, 158.09, 106.39, 66.85, 44.52. HRMS(ESI)[M+H] + In C8H 11 Calculated value of BrN3O: 244.0080, measured value: 244.0084.

[0130] (R)-5-bromo-2-(3-fluoropyrrolidine-1-yl)pyrimidine ((R)-compound 25) 5-Bromo-2-chloropyrimidine (0.51 g, 2.64 mmol), (R)-3-fluoropyrrolidine·HCl (0.49 g, 3.90 mmol, 1.5 equiv.), K2CO3 (1.13 g, 8.18 mmol, 3.1 equiv.), and CH3CN (30 mL) were stirred under reflux with N2 (g) for 90 minutes, then cooled to ambient temperature, filtered, and the precipitate was washed three times with phenylethylamine. The solvent was removed from the filtrate to obtain a white solid, which was dissolved in CH2Cl2 and poured onto dry silica (55 mm h × 45 mm id) and eluted under vacuum. From %CH2Cl2 / hexane-50% (50 mL), 75% (100 mL), CH2Cl2 (200 mL), %MeOH / CH2Cl2-1% (100 mL), 2% (100 mL), 3% (100 mL), 5% (100 mL), and 10% (50 mL), (R)-compound 25 (0.50 g, 77%) was obtained as an off-white solid. 1 H NMR (300MHz, CDCl3) δ8.32(s, 2H), 5.36(dt, 1H, 2 J HF =52.8, J=3.3Hz), 3.99-3.57(m, 4H), 2.45-1.99(m, 2H).

[0131] 1-(5-bromopyrimidine-2-yl)piperidine-4-ol (compound 26) 5-Bromo-2-chloropyrimidine (0.97 g, 5.01 mmol), 4-hydroxypiperidine (0.56 g, 5.54 mmol, 1.1 equiv.), K2CO3 (0.92 g, 6.66 mmol, 1.3 equiv.), and CH3CN (30 mL) were stirred under reflux for 1 hour under N2 (g), then cooled to ambient temperature, filtered, and the precipitate was washed with CH3CN. CH3CN was removed from the filtrate to obtain a solid, which was dissolved / suspended in CH2Cl2, poured onto dry silica (55 mm h × 45 mm id), and eluted under vacuum. Compound 26 (1.27 g, 98%) was obtained as a white solid from %MeOH / CH2Cl2-1% (100 mL), 2% (100 mL), 3% (200 mL), and 5% (150 mL). 1 H NMR (300MHz, CDCl3) δ8.27(s, 2H), 4.33(dtd, 2H, J=13.8Hz, J=4.8Hz, J=0.9Hz), 3.96(apparent o ctet, 1H, J=4.2Hz), 3.33(ddd, 2H, J=13.5Hz, J=9.6Hz, J=3.3Hz), 1.94(m, 2H), 1.53(m, 3H).

[0132] 1-(5-bromopyrimidine-2-yl)piperidine-4-one (compound 27) Compound 26 (0.54 g, 2.09 mmol) was dissolved in CH2Cl2 (25 mL), then H2O (0.05 mL) was added, followed by Dess-Martin periodinane (1.03 g, 2.43 mmol, 1.2 equiv.). The reaction mixture was stirred under reflux for 7 hours, then cooled to ambient temperature and stirred for 17 hours. The mixture was then poured into a mixture of CH2Cl2 (25 mL), H2O (50 mL), Na2S2O3·5H2O (2.17 g, 8.74 mmol), and NaHCO3 (2.25 g, 26.78 mmol). After stirring the mixture for 2 hours, the layers were separated, and the H2O layer was extracted with CH2Cl2 (10 mL × 2). The combined CH2Cl2 layers were washed with brine (25 mL) and dried on MgSO4. The solution was concentrated and poured onto dry silica (55 mm h × 45 mm id), and eluted under vacuum. Off-white solids (0.55 g) were obtained from CH2Cl2 (200 mL), %MeOH / CH2Cl2-1% (100 mL), 2.5% (100 mL), and 5% (200 mL). By purification using radial chromatography (4 mm silica) with %MeOH / CH2Cl2-1% (100 mL), 2% (100 mL), 4% (50 mL), and 5% (100 mL), recovered compound 26 (0.11 g, 20%) and compound 27 (0.38 g, 71%) were recovered as white solids. 1 H NMR (300MHz, CDCl3) δ8.35 (s, 2H), 4.10 (t, 4H, J=6.3Hz), 2.50 (t, 4H, J=6.3Hz).

[0133] 5-Bromo-2-(piperidine-1-yl)pyridine (compound 28) 2,5-Dibromopyridine (0.87 g, 3.67 mmol) and piperidine (5 mL, 50.6 mmol, 13.8 equiv.) were stirred in a heated sand bath (105 °C) under N2 (g) for 15 hours, and then cooled to ambient temperature. Depositphotos (5 mL) was added, the mixture was filtered, and the precipitate was washed with Depositphotos (5 mL x 2). The filtrate was concentrated to obtain the oil / residue, then dissolved in CH2Cl2, poured onto dry silica (55 mm h x 45 mm id), and eluted under vacuum. Compound 28 (0.86 g, 97%) was obtained as a colorless oil from hexane (50 mL), hexane / Depositphotos / Net3 v / v / v 90:8:2 (100 mL), and 75:20:5 (100 mL). 1 H NMR (300MHz, CDCl3) δ8.16 (d, 1H, J=2.4Hz), 7.48 (dd, 1H, J=9.3Hz, J=2.4Hz), 6.54 (d, 1H, J=9.3Hz), 3.50 (m, 4H), 1.63 (m, 6H).

[0134] 4-(5-bromopyridine-2-yl)morpholine (compound 29) 2,5-Dibromopyridine (0.87 g, 3.67 mmol) and morpholine (5 mL, 58 mmol, 16 equiv.) were stirred in a heated sand bath (~110°C) under N2 (g) for 15 hours, and then cooled to ambient temperature. Depositphotos (5 mL) was added, the mixture was filtered, and the precipitate was washed with Depositphotos (5 mL × 2). The filtrate was concentrated, diluted with hexane, and poured onto dry silica (55 mm h × 45 mm id), and eluted under vacuum. Compound 29 (0.86 g, 96%) was obtained as an off-white solid from hexane / Depositphotos / Net3v / v / v 90:8:2 (100 mL) and 75:20:5 (200 mL). 1 H NMR (300MHz, CDCl3) δ8.21(d, 1H, J=2.4Hz), 7.56(dd, 1H, J=9.0Hz, J=2.4Hz), 6.53(d, 1H, J=9.0Hz), 3.81(t, 4H, J=4.8Hz), 3.47(t, 4H, J=4.8Hz).

[0135] 4-(5-bromopyrazine-2-yl)morpholine (compound 30) Morpholine (0.3 mL, 3.5 mmol) was added to a suspension of NaH (87 mg, 90%, 3.26 mmol) in THF (6 mL) under N2 (g) at 0°C, and the mixture was stirred for 15 minutes. Then, 2,5-dibromopyrazine (0.81 g, 3.41 mmol) in THF (6 mL) was added, and the reaction mixture was stirred overnight under reflux. The solvent was evaporated, the residue was dissolved in toluene (20 mL), washed with H2O (10 mL), and dried on MgSO4. The solvent was removed, and the residue was purified by flash column chromatography (4:1 v / v hexane / toluene) to obtain compound 30 as a white solid (0.86 g, quantitative). 1 ¹H NMR (CDCl3, 500MHz): δ 8.16 (d, 1H, J=2.0Hz), 7.86 (d, 1H, J=2.5Hz), 3.83 (apparent t, 4H, J=8.0Hz), 3.53 (apparent t, 4H, J=8.0Hz). HRMS (ESI) [M+H] + In C8H 11 ON3Br calculated value: 244.0080, measured value: 244.0076.

[0136] tert-butyl 6-fluoro-2-(2-(4-methoxypiperidine-1-yl)pyrimidine-5-yl)-1H-indole-1-carboxylate (compound 31). Compound 4 (0.65g, 1.63mmol), Compound 23 (0.49g, 1.80mmol, 1.1equiv.), Pd(dppf)Cl2 (0.11g, 1.50×10 -4After flushing with N2(g) for 10 minutes with mol(0.09 equiv.) and Na2CO3 (0.56g, 5.28 mmol, 3.2 equiv.), 1,4-dioxane (45mL) was added. The reaction mixture was stirred under reflux under N2(g) for 15 hours, cooled to ambient temperature, filtered through Celite, and washed with 1,4-dioxane. The filtrate was concentrated to form a brown syrup, dried under vacuum, dissolved in CH2Cl2, poured onto dry silica (55mm h × 45mm id), and eluted under vacuum. Brown residues (0.65g) were obtained from CH2Cl2 (100mL), %MeOH / CH2Cl2-1% (100mL), 2.5% (100mL), and 5% (150mL). The residues were dissolved in CH2Cl2, poured onto dry silica (55mm h × 45mm id), and eluted under vacuum. A dark orange solid (0.52 g) was obtained from hexane (25 mL), hexane / Â / NEt3v / v / v 75:20:5 (100 mL), 50:45:5 (100 mL), and 20:75:5 (150 mL). Purification by radial chromatography (2 mm silica), CH2Cl2 (100 mL), and 1% MeOH / CH2Cl2 (50 mL) yielded crude compound 31 (0.30 g, light brown syrup) and crude compound 36 (91 mg, light brown residue - further purification below).

[0137] Crude compound 31 was purified by radial chromatography (2 mm silica) with hexane / Â / NEt3v / v / v 95:4:1 (100 mL) and 90:8:2 (25 mL) to obtain a viscous, slightly yellowish residue. This residue was further purified by radial chromatography (2 mm silica) with CHCl3 (50 mL), %EtOH / CHCl3-1% (50 mL), and 2% (25 mL) to obtain compound 31 (0.26 g, 37%) as a slightly yellowish, viscous syrup that slowly solidified. 1H NMR (300MHz, CDCl3) δ8.34(s, 2H), 7.93(dd, 1H, J=10.8Hz, J=2.4Hz), 7.45(dd, 1H, J=8.7Hz, J=5.4Hz), 7.01(td, 1 H, J=8.7Hz, J=2.4Hz), 6.49(s, 1H), 4.34(m, 2H), 3.48(m, 3H), 3.41(s, 3H), 1.96(m, 2H), 1.59(m, 2H), 1.47(s, 9H). HRMS(ESI)[M+H] + In C 23 H 28 Calculated value for FN4O3: 427.2140, measured value: 427.2131.

[0138] tert-butyl(R)-6-fluoro-2-(2-(3-fluoropyrrolidine-1-yl)pyrimidine-5-yl)-1H-indole-1-carboxylate((R)-compound 32) Compound 4 (0.40g, 1.00mmol), (R)-Compound 25 (0.30g, 1.22mmol, 1.2equiv.), Pd(dppf)Cl2 (76mg, 1.04×10 -41,4-Dioxane (25 mL) was stirred under reflux with N2 (g) for 6 hours and then cooled to ambient temperature. The reaction mixture was filtered through Celite, washed with EtOAC, and the filtrate was concentrated to obtain a brown syrup. CH2Cl2 and hexane were added to remove the residue, and the brown residue was dried under vacuum. The residue was dissolved in CH2Cl2 (+ a few drops of MeOH), poured onto dry silica (55 mm h × 45 mm id), and eluted under vacuum. Dark orange / brown residue (0.37 g) was obtained from CH2Cl2 (100 mL), %MeOH / CH2Cl2-1% (100 mL), 2.5% (100 mL), and 5% (200 mL). The residue was dissolved in CH2Cl2 / MeOH and purified by radial chromatography (2mm silica) with hexane / Âi / NEt3v / v / v 90:8:2 (100mL) to obtain a pale yellow solid (0.17g). This was further purified by radial chromatography (2mm silica) with hexane / Âi / NEt3v / v / v 90:8:2 (100mL) and 75:20:5 (100mL) to obtain a pale yellow solid (0.14g). A third purification by radial chromatography (2mm silica) with CHCl3 (150mL) and 1% EtOH / CHCl3 (50mL) yielded (R)-compound 32 (0.13g, 32%) as an off-white solid. 1 H NMR (400MHz, CDCl3) δ8.39(s, 2H), 7.92(dd, 1H, J=10.6Hz, J=2.4Hz), 7.46(dd, 1H , J=8.8Hz, J=5.4Hz), 7.01(td, 1H, J=8.8Hz, J=2.4Hz), 6.51(s, 1H), 5.40(dt, 1H, 2 J HF =52.8Hz, J=3.2Hz), 4.03(ddd, 1H, 3 J HF=25.2Hz, J=13.6Hz, J=1.6Hz), 3.93(t, 1H, J=10.0Hz), 3.81(dd, 0.5H, J=13.6Hz, J=3 .6Hz), 3.76-3.69(m-overlapping resonance, 1.5H), 2.46-2.37(m, 1H), 2.26-2.06(m, 1H), 1.49(s, 9H). HRMS(ESI)[M+H] + In C 21 H 23 Calculated value for O2N4F2: 401.1784, measured value: 401.1774.

[0139] tert-butyl 6-fluoro-2-(2-(4-oxopiperidine-1-yl)pyrimidine-5-yl)-1H-indole-1-carboxylate (compound 33) 1,4-Dioxane was bubbling with N2(g) for 30 minutes. Compound 4 (0.24g, 6.03 × 10⁻⁶) -4 mol), compound 27 (0.18g, 7.03×10 -4 mol, 1.2equiv.), Pd(dppf)Cl2 (46.5mg, 6.36×10 -5Flushing of mol (0.1 equiv.) and Na2CO3 (0.14 g, 1.32 mmol, 2.2 equiv.) with N2(g) for 10 minutes, then adding 1,4-dioxane (25 mL), stirring the mixture under reflux with N2(g) for 5 hours, and then cooling to ambient temperature. The mixture was filtered through Celite, and the Celite was washed with siRNA. The filtrate was concentrated to obtain a dark orange oil, which was removed by adding CH2Cl2 and hexane to obtain a dark orange residue. This residue was dissolved in CH2Cl2, poured onto dry silica (55 mm h × 45 mm id), and eluted under vacuum. A pale orange solid (0.134 g) was obtained from hexane / siRNA / NEt3 v / v / v 90:8:2 (100 mL), 75:20:5 (100 mL), 50:45:5 (200 mL), and 20:75:5 (100 mL). Purification by radial chromatography (2 mm silica) and hexane / Â / NEt3v / v / v 90:8:2 (100 mL) and 75:20:5 (100 mL) yielded an off-white solid (79 mg). This was further purified by radial chromatography (2 mm silica) and %MeOH / CH2Cl2-1% (50 mL) and 2% (50 mL) to obtain compound 33 (70 mg, 28%) as an off-white solid. 1 H NMR (300MHz, CDCl3) δ8.42(s, 2H), 7.93(dd, 1H, J=10.8Hz, J=2.4Hz), 7.47(dd, 1H, J=8.7Hz, J=5.4Hz), 7 .02(td, 1H, J=8.7Hz, J=2.4Hz), 6.53(s, 1H), 4.20(t, 4H, J=6.3Hz), 2.54(t, 4H, J=6.3Hz), 1.49(s, 9H). HRMS(ESI)[M+H] + In C 22 H 24 Calculated value for O3N4F: 411.1827, measured value: 411.1815.

[0140] tert-butyl 6-fluoro-2-(6-(piperidine-1-yl)pyridine-3-yl)-1H-indole-1-carboxylate (compound 34) 1,4-dioxane (30 mL) was purged with N2 (g) for 30 minutes. Compound 4 (0.44 g, 1.11 mmol), Compound 28 (0.29 g, 1.20 mmol, 1.1 equiv.), Pd(dppf)Cl2 (0.10 g, 0.14 mmol, 0.1 equiv.), and Na2CO3 (0.54 g, 5.09 mmol, 4.6 equiv.) were flushed with N2 (g) for 10 minutes, and then 1,4-dioxane was added. The reaction mixture was stirred under reflux under N2 (g) for 5 hours, then cooled to ambient temperature, filtered through Celite, and washed with Celite using SiO2. The solvent was removed from the filtrate to obtain a brown oil, which was then removed by adding CH2Cl2 and hexane to obtain a brown residue, which was dried under vacuum. The residue was dissolved in CH2Cl2, poured onto dry silica (55 mm h × 4 mm id), and eluted under vacuum. Using %CH2Cl2 / hexane-50% (100 mL), 75% (100 mL), CH2Cl2 (200 mL), and %MeOH / CH2Cl2-1% (100 mL), 2.5% (100 mL), and 5% (100 mL), compound 4 (crude 0.22 g, 94 mg after subsequent purification, recovery rate 21%) and brown residue (0.31 g) were obtained. The brown residue was dissolved in CH2Cl2, poured onto dry silica (55 mm h × 45 mm id), and eluted under vacuum. Brown residue (0.27 g) was obtained from CH2Cl2 (100 mL), %MeOH / CH2Cl2-1% (100 mL), 2% (100 mL), 3% (200 mL), and 5% (200 mL). This was purified by radial chromatography (2 mm silica) and CHCl3 (100 mL) to obtain a light brown solid (0.12 g). Purification by radial chromatography (2 mm silica) and hexane / siRNA / NET3v / v / v 90:8:2 (100 mL) and 75:20:5 (25 mL) yielded compound 34 (89 mg, 20%) as an off-white solid. 1H NMR (300MHz, CDCl3) δ8.22(d, 1H, J=2.4Hz), 7.92(dd, 1H, J=10.8Hz, J=2.4Hz), 7.45(m, 2H), 6.99(t d, 1H, J=8.7Hz, 2.4Hz), 6.67(d, 1H, J=8.7Hz), 6.46(s, 1H), 3.59(m, 4H), 1.67(m, 6H), 1.43(s, 9H). HRMS(ESI)[M+H] + In C 23 H 27 Calculated value of FN3O2: 396.2082, measured value: 396.2081.

[0141] tert-butyl 6-fluoro-2-(6-morpholinopyridine-3-yl)-1H-indole-1-carboxylate (compound 35) Compound 4 (0.41 g, 1.03 mmol), Compound 29 (0.26 g, 1.07 mmol, 1 equiv.), Pd(dppf)Cl2 (81 mg, 0.11 mmol, 0.1 equiv.), Na2CO3 (0.32 g, 3.02 mmol, 2.9 equiv.), and 1,4-dioxane (25 mL) were stirred under reflux with N2 (g) for 6 hours and then cooled to ambient temperature. The reaction mixture was filtered through Celite, washed with EtOAC, and the filtrate was concentrated to obtain a brown oil. CH2Cl2 and hexane were then added to remove the oil and obtain a brown residue, which was dried under vacuum. The residue was dissolved in CH2Cl2, poured onto dry silica (55 mm h × 45 mm id), and eluted under vacuum. A yellow syrup (0.14 g) was obtained from hexane / siRNA / NEt3v / v / v 90:8:2 (125 mL), 75:20:5 (100 mL), 50:45 (150 mL), and 20:75:5 (200 mL). By purification using radial chromatography (2 mm silica) and hexane / siRNA / siRNA / NEt3v / v / v 90:8:2 (125 mL) and 75:20:5 (100 mL), compound 35 (66 mg, 16%) was obtained as a pale yellow foam. 1H NMR (400MHz, CDCl3) δ8.26(d, 1H, J=2.0Hz), 7.91(dd, 1H, J=10.6Hz, J=2.0Hz), 7.54(dd, 1H, J=8.8Hz, J=2.4Hz), 7.45(dd, 1H, J=8.8Hz, J =5.4Hz), 7.00(td, 1H, J=8.8Hz, J=2.4Hz), 6.67(d, 1H, J=8.8Hz), 6.48(s, 1H), 3.85(t, 4H, J=4.8Hz), 3.56(t, 4H, J=4.8Hz), 1.44(s, 9H). HRMS(ESI)[M+H] + In C 22 H 25 Calculated value of FN3O3: 398.1875, measured value: 398.1866.

[0142] 6-Fluoro-2-(2-(4-methoxypiperidine-1-yl)pyrimidine-5-yl)-1H-indole (compound 36) Crude compound 36 (91 mg - obtained from the coupling reaction of compound 31 above) was purified by radial chromatography (1 mm silica), CH2Cl2 (50 mL), %MeOH / CH2Cl2 - 1% (50 mL), 2% (75 mL) to obtain compound 36 (46 mg, 9%) as a yellowish-brown solid. 1 H NMR (300MHz, CDCl3) δ8.57(s, 2H), 8.23(br s, 1H), 7.50(dd, 1H, J=8.7Hz, J=5.4Hz), 7.08(dd, 1H, J=9.6Hz, J=2.1Hz), 6.89(ddd, 1H, J=9.7Hz, J=8. 7Hz, J=2.4Hz), 6.63(d, 1H, J=1.5Hz), 4.31(m, 2H), 3.50(m, 3H), 3.41(s, 3H), 1.97(m, 2H), 1.62(m, 2H), 13 C NMR (125MHz, DMSO-d6) δ160.04, 158.74(d, 1 J FC =234.4Hz), 154.52, 136.70(d, J FC =12.7Hz), 134.06(d, J FC =3.5Hz), 125.42, 120.51(d, J FC =10.2Hz), 114.81, 107.76(d, JFC =24.4Hz), 97.08(d, J FC =25.7Hz), 97.05, 75.49, 54.97, 40.97, 30.20, 19 F NMR (470.6MHz, DMSO-d6) δ-121.53(m).

[0143] Compound 36 from N-Boc deprotection of compound 31 Compound 31 (64.5mg, 1.51×10 -4 A mixture of mol) and TFA (1.5 mL, 19.5 mmol, 129 equiv.) was stirred at ambient temperature for 15 minutes, then poured over a mixture of NaHCO3 (1.84 g, 21.90 mmol, 1.1 equiv. TFA), H2O (45 mL), and CH2Cl2 (45 mL). The mixture was stirred, the layers were separated, and the H2O layer was extracted with CH2Cl2 (10 mL). The combined CH2Cl2 layers were washed with brine (20 mL) and dried on MgSO4. The solution was concentrated and poured onto dry silica (33 mm h × 33 mm id) and eluted under vacuum. Compound 36 (30 mg, 61%) was obtained as a pale yellow solid from hexane / siRNA / NET3 v / v / v 90:8:2 (50 mL), 75:20:5 (50 mL), and 50:45:5 (100 mL). 1 H NMR (300MHz, CDCl3) δ8.57(s, 2H), 8.24(br s, 1H), 7.49(dd, 1H, J=8.7Hz, J=5.4Hz), 7.07(dd, 1H, J=9.3Hz, J=2.1Hz), 6.89(ddd, 1H, J=9.6Hz, J=8.7Hz, J=2.4 Hz), 6.62 (partially isolated dd, 1H, J=2.1Hz, J=0.6Hz), 4.31(m, 2H), 3.50(m, 3H), 3.41(s, 3H), 1.97(m, 2H), 1.62(m, 2H). HRMS(ESI)[M+H] + In C 18 H 20 FN4O calculated value: 327.1616, measured value: 327.1620.

[0144] (R)-6-fluoro-2-(2-(3-fluoropyrrolidine-1-yl)pyrimidine-5-yl)-1H-indole((R)-compound 37) (R)-Compound 32 (0.12g, 3.00×10 -4 The solution was dissolved in TFA (3 mL, 38.9 mmol, 130 equiv.) and stirred at ambient temperature for 20 minutes. Then, it was poured into a mixture of NaHCO3 (3.95 g, 47.02 mmol, 1.2 equiv. TFA), H2O (75 mL), and CH2Cl2 (75 mL). After stirring the mixture for 20 minutes, the layers were separated, and the aqueous layer was extracted with CH2Cl2 (25 mL × 2). The combined CH2Cl2 layers were washed with brine (25 mL), dried on MgSO4, and concentrated (a precipitate was formed). The precipitate was redissolved by adding MeOH, and then the solution was poured onto dry silica (55 mm h × 45 mm id) and eluted under vacuum. Yellow solids (85 mg) were obtained from %MeOH / CH2Cl2-1% (100 mL), 2.5% (100 mL), 5% (200 mL), and 10% (50 mL). This solid was placed in a medium-mesh frit filter, washed with 1:1 v / v CH2Cl2 / hexane (1 mL x 7), and dried under vacuum to obtain (R)-compound 37 (67 mg, 74%) as a pale yellow solid. 1 ¹H NMR (300MHz, acetone-d6) δ 10.71 (br s, 1H), 8.80 (s, 2H), 7.52 (dd, 1H, J=8.4Hz, J=5.4Hz), 7.10 (dd, 1H, J=9.9Hz, J=2.1Hz), 6.84 (ddd, 1H, J=9.9Hz, J=8.7Hz, J=2.4Hz), 6.79 (s, 1H), 5.45 (dm, 1H, 2 J HF =52.8Hz), 4.01-3.58(m, 4H), 2.37-2.12(m, 2H). HRMS(ESI)[M+H] + In C 16 H 15 N4F2 calculated value: 301.1259, measured value: 301.1253.

[0145] 1-(5-(6-fluoro-1H-indole-2-yl)pyrimidine-2-yl)piperidine-4-one (compound 38) Compound 33 (65mg, 1.58×10 -4 The solution was dissolved in TFA (1.6 mL, 20.8 mmol, 131 equiv.) and stirred at ambient temperature for 20 minutes. This mixture was then poured into a mixture of NaHCO3 (2.13 g, 25.36 mmol, 1.2 equiv. TFA), H2O (40 mL), and CH2Cl2 (40 mL). After stirring the mixture for 10 minutes, the layers were separated, and the aqueous layer was extracted with CH2Cl2 (15 mL × 2). The combined CH2Cl2 layers were washed with brine (25 mL), dried on MgSO4, concentrated, poured onto dry silica (55 mm h × 45 mm id), and eluted under vacuum. Off-white solids (43 mg) were obtained from %MeOH / CH2Cl2-1% (100 mL), 2.5% (100 mL), 5% (200 mL), and 10% (50 mL). Compound 38 (38 mg, 77%) was obtained as a pale yellowish-brown solid by radial chromatography (1 mm silica), using 1% (50 mL) and 2.5% (25 mL) concentrations of %MeOH / CH2Cl2. 1 H NMR (300MHz, CDCl3) δ8.65(s, 2H), 8.27(br s, 1H), 7.52(dd, 1H, J=8.7Hz, J=5.4Hz), 7.10(dd, 1H, J=9.3Hz, J=2.1Hz), 6.91(ddd, 1H, J=9.6Hz, J=8.7Hz, J=2.1Hz), 6.68(dd, 1H, J=2.1Hz, J=0.6Hz), 4.20(t, 4H, J=6.0Hz), 2.55(t, 4H, J=6.0Hz). HRMS(ESI)[M+H] + In C 17 H 16 Calculated value of FN4O: 311.1303, measured value: 311.1296. X-ray quality crystals were grown by slow evaporation of acetone.

[0146] 6-fluoro-2-(6-(piperidine-1-yl)pyridine-3-yl)-1H-indole (compound 39) Compound 34 (82 mg, 2.07 × 10⁻⁶) -4The solution was dissolved in TFA (2 mL, 26 mmol, 126 equiv.) and stirred at ambient temperature for 20 minutes. The mixture was then poured into a mixture of Na2CO3 (2.43 g, 28.93 mmol, 1.1 equiv. TFA), H2O (60 mL), and CH2Cl2 (60 mL). After stirring the mixture for 10 minutes, the layers were separated, and the aqueous layer was extracted with CH2Cl2 (10 mL × 3). The combined CH2Cl2 layers were washed with brine (30 mL), dried on MgSO4, and then poured onto dry silica (33 mm h × 33 mm id) and eluted under vacuum. Off-white solids (58 mg) were obtained from %MeOH / CH2Cl2-1% (50 mL), 2% (75 mL), and 3% (50 mL). Compound 39 (50 mg, 82%) was obtained as an off-white solid by purification using radial chromatography (1 mm silica), hexane / Â1 / NEt3v / v / v 90:8:2 (40 mL), and 75:20:5 (50 mL). 1 H NMR (300MHz, CDCl3) δ8.46(d, 1H, J=2.4Hz), 8.28(br s, 1H), 7.68(dd, 1H, J=9.0Hz, J=2.4Hz), 7.48(dd, 1H, J=8.7Hz, J=5.4Hz), 7.06(dd, 1H, J=9.6Hz, J=1.8Hz), 6.8 6(ddd, 1H, J=9.6Hz, J=8.7Hz, J=2.1Hz), 6.70(d, 1H, J=8.7Hz), 6.61(d, 1H, J=1.5Hz), 3.59(m, 4H), 1.67(m, 6H), 13 C NMR (125MHz, DMSO-d6) δ158.57(d, 1 J FC =233.9Hz), 157.98, 144.42, 136.86(d, J FC (=3.5Hz), 136.66(d, J) FC =12.6Hz), 134.13, 125.61, 120.25(d, J FC =9.9Hz), 116.64, 107.49(d, J FC =24.0Hz), 106.98, 96.95(d, J FC =25.5Hz), 96.51, 45.50, 24.99, 24.29, 19F NMR (470.6MHz, DMSO-d6) δ-122.04(m). HRMS(ESI)[M+H] + In C 18 H 19 FN3 calculated value: 296.1558, measured value: 296.1568.

[0147] 4-(5-(6-fluoro-1H-indole-2-yl)pyridine-2-yl)morpholine (compound 40) Compound 35 (61mg, 1.53×10 -4 The compound (mol) was dissolved in TFA (1.5 mL, 19.5 mmol, 127 equiv.), stirred at ambient temperature for 20 minutes, and then poured into a mixture of Na2CO3 (1.82 g, 21.67 mmol, 1.1 equiv. TFA), H2O (40 mL), and CH2Cl2 (40 mL). After stirring the mixture for 15 minutes, the layers were separated, and the aqueous layer was extracted with CH2Cl2 (15 mL × 2). The combined CH2Cl2 layers were washed with brine (25 mL), dried on MgSO4, and then poured onto dry silica (33 mm h × 33 mm id) and eluted under vacuum. Compound 40 (42 mg, 92%) was obtained as a pale yellowish-brown solid from %MeOH / CH2Cl2-1% (50 mL), 2% (75 mL), and 3% (50 mL). 1 H NMR (300MHz, CDCl3) δ8.50 (d, 1H, J=2.1Hz), 8.28 (br s, 1H), 7.75(dd, 1H, J=8.7Hz, J=2.4Hz), 7.49(dd, 1H, J=8.7Hz, J=5.4Hz), 7.07(dd, 1H, J=9.3Hz, J=2.1Hz), 6.88(ddd, 1H, J=9. 6Hz, J=8.7Hz, J=2.1Hz), 6.71(d, 1H, J=8.7Hz), 6.65(dd, 1H, J=2.1Hz, J=0.6Hz), 3.85(t, 4H, J=4.8Hz), 3.57(t, 4H, J=4.8Hz), 13 C NMR (125MHz, DMSO-d6) δ158.66(d, 1 J FC =234.3Hz), 158.19, 144.28, 136.72(d, J FC =12.7Hz), 136.54(d, J FC=3.4Hz), 134.19, 125.55, 120.41(d, J FC =10.1Hz), 117.95, 107.60(d, J FC =24.3Hz), 106.92, 97.00(d, J FC =25.5Hz), 96.99, 65.91, 45.04, 19 F NMR (470.6MHz, DMSO-d6) δ-121.79. HRMS(ESI)[M+H] + In C 17 H 17 FN3O calculated value: 298.1350, measured value: 298.1345.

[0148] 1-(5-(5-fluorobenzofuran-2-yl)pyrimidine-2-yl)piperidine-4-ol (compound 43) 1,4-Dioxane (30 mL) and H2O (6 mL) were combined and purged with N2 (g) for 45 minutes. 5-Fluorobenzofuran-2-boronic acid (compound 41) (0.25 g, 1.39 mmol), compound 26 (0.42 g, 1.63 mmol, 1.2 equiv.), Pd(dppf)Cl2·CH2Cl2 (97 mg, 1.19 × 10⁻¹⁴) -4After flushing with N2(g) for 10 minutes with mol, 0.09 equiv., and Na2CO3 (0.71g, 6.70 mmol, 4.8 equiv.), H2O / 1,4-dioxane mixture (17 mL) was added. The reaction mixture was stirred under N2(g) in a heated sand bath (~95°C) for 14 hours, then cooled to ambient temperature, filtered through Celite, and washed with ethyl acetate. The filtrate was concentrated, ethyl acetate was added to remove it, and a brown residue was obtained. This was dissolved in CH2Cl2, poured onto dry silica (55 mm h × 45 mm id), and eluted under vacuum. Hexane / ethyl acetate / Net3 v / v / v 75:20:5 (100 mL), 50:45:5 (150 mL), 20:75:5 (250 mL), ethyl acetate (50 mL). The desired fractions were combined and purified by radial chromatography (2 mm silica) using %EtOH / CHCl3-1% (50 mL), 2% (75 mL), and 3% (100 mL) to obtain a yellowish-brown solid (77 mg). This solid was suspended in CHCl3 (2 mL), filtered, washed with CHCl3 (1.5 mL x 2), and dried under vacuum to obtain compound 43 (47 mg, 11%) as a white solid. 1 ¹H NMR (300MHz, DMSO-d6) δ 8.85 (s, 2H), 7.60 (dd, 1H, J=9.0Hz, J=4.2Hz), 7.43 (dd, 1H, J=9.0Hz, J=2.4Hz), 7.26 (s, 1H), 7.10 (td, 1H, J=9.0Hz, J=2.4Hz), 4.77 (d, 1H, J=4.2Hz), 4.30 (partially separated dddd, 2H, J=13.5Hz, J=4.5Hz), 3.77 (apparent octet, 1H, J=4.2Hz), 3.39 (partially obscured by ddd-H2O resonance, 2H, J=9.6Hz, J=3.3Hz), 1.80 (m, 2H), 1.36 (m, 2H). HRMS(ESI)[M+H] + In C 17 H 17 Calculated value of FN3O2: 314.1299, measured value: 314.1296.

[0149] tert-butyl 2-(2-(4-hydroxypiperidine-1-yl)pyrimidine-5-yl)-1H-indole-1-carboxylate (compound 44) 1,4-Dioxane (10 mL) and H2O (2 mL) were combined and purged with N2 (g) for 10 minutes. Compound 26 (0.24 g, 9.30 × 10) -4 (mol), Boc-indole-2-boronic acid (compound 6) (0.26g, 9.96 × 10⁻⁶) -4 mol, 1.1equiv.), Pd(dppf)Cl2 (94mg, 1.15×10 -4 After flushing with N2(g) for 15 minutes with mol, 0.1 equiv., and Na2CO3 (0.45g, 4.25 mmol, 4.6 equiv.), an H2O / 1,4-dioxane mixture was added. The reaction mixture was stirred in a heated sand bath (85°C) under N2(g) for 16 hours, then cooled to ambient temperature, filtered through Celite, and washed with SiO2. The solvent was removed to obtain a black syrup, which was dissolved in CH2Cl2 and poured onto dry silica (45mm h × 45mm id) and eluted under vacuum. CH2Cl2 (100mL), %MeOH / CH2Cl2 - 1% (100mL), 2.5% (100mL), 5% (100mL). The desired fractions were combined, concentrated, poured onto dry silica (45mm h × 45mm id), and eluted under vacuum. From hexane / Ã / NEt3v / v / v 90:8:2 (50 mL), 75:20:5 (100 mL), 50:45:5 (100 mL), and 20:75:5 (200 mL), an orange / yellowish-brown foamy substance (0.25 g) was obtained. By purification using radial chromatography (2 mm silica), CHCl3 (50 mL), and %EtOH / CHCl3-1% (50 mL), 2% (75 mL), and 3% (50 mL), compound 44 (0.23 g, 63%) was obtained as a pale yellow foamy substance. 1¹H NMR (400MHz, CDCl3) δ 8.36 (s, 2H), 8.19 (d, 1H, J=8.4Hz), 7.54 (d, 1H, J=6.0Hz), 7.32 (apparent t, 1H, J=7.6Hz), 7.25 (apparent t - partially hidden by CHCl3 resonance, 1H, J=7.6Hz), 6.53 (s, 1H), 4.46 (partially separated dddd, 2H, J=13.6Hz, J=4.4Hz), 3.99 (br s, 1H), 3.40 (partially separated ddd, 2H, J=11.4Hz, J=2.8Hz), 1.98 (m, 2H), 1.57 (m, 2H), 1.48 (s, 10H).

[0150] tert-butyl 5-fluoro-2-(2-(4-hydroxypiperidine-1-yl)pyrimidine-5-yl)-1H-indole-1-carboxylate (compound 45) 1,4-Dioxane (30 mL) and H2O (6 mL) were combined and purged with N2 (g) for 45 minutes. N-Boc-5-Fluorindole-2-boronic acid (compound 42) (0.25 g, 8.96 × 10⁻⁶) -4 mol), compound 26 (0.25g, 9.69×10 -4 mol, 1.1equiv.), Pd(dppf)Cl2·CH2Cl2 (79mg, 9.67×10 -5After flushing with N2(g) for 10 minutes with mol (0.1 equiv.) and Na2CO3 (0.47g, 4.43 mmol, 5 equiv.), a H2O / 1,4-dioxane mixture (16 mL) was added. The reaction mixture was stirred under N2(g) in a heated sand bath (~95°C) for 14 hours, then cooled to ambient temperature, filtered through Celite, and washed with ethyl acetate. The filtrate was concentrated to obtain an oil, then ethyl acetate was added and removed to obtain a brown residue, which was dissolved in CH2Cl2 and poured onto dry silica (55 mm h × 45 mm id) and eluted under vacuum. A yellowish-brown foamy substance (0.23 g) was obtained from hexane / ethyl acetate / Net3 v / v / v 75:20:5 (100 mL), 50:45:5 (150 mL), and 20:75:5 (200 mL). By purification using radial chromatography (2 mm silica), %EtOH / CHCl 3-1% (50 mL), and 2% (75 mL), compound 45 (0.17 g, 46%) was obtained as an off-white foam. 1 ¹H NMR (400MHz, CDCl3) δ 8.35 (s, 2H), 8.14 (dd, 1H, J=8.8Hz, J=4.8Hz), 7.19 (dd, 1H, J=8.8Hz, J=2.4Hz), 7.04 (td, 1H, J=9.2Hz, J=2.4Hz), 6.49 (s, 1H), 4.46 (partially separated dddd, 2H, J=13.6Hz, J=4.4Hz), 4.00 (apparent octet, 1H, J=4.4Hz), 3.40 (ddd, 1H, J=10.0Hz, J=3.2Hz), 1.98 (m, 2H), 1.56 (m, 3H), 1.47 (s, 9H).

[0151] 1-(5-(1H-indole-2-yl)pyrimidine-2-yl)piperidine-4-ol (compound 46). Compound 44 (83mg, 2.10×10 -4The solution was dissolved in TFA (2 mL, 26 mmol, 124 equiv.) and stirred at ambient temperature for 20 minutes. Then it was poured into a mixture of NaHCO3 (2.40 g, 28.57 mmol, 1.1 equiv. TFA), H2O (60 mL), and CH2Cl2 (60 mL). The mixture was stirred until gas generation ceased, then the layers were separated, and the aqueous layer was extracted with CH2Cl2 (10 mL). The combined CH2Cl2 layers were washed with brine (25 mL), dried on MgSO4, and concentrated until a precipitate was formed. MeOH was added (a few drops) until the precipitate dissolved, and the solution was poured onto dry silica (55 mm h × 45 mm id) and eluted under vacuum. Off-white solids were obtained from %MeOH / CH2Cl2 concentrations of 1% (100 mL), 2.5% (100 mL), 5% (150 mL), and 10% (50 mL). This solid was dissolved / suspended in CHCl3 (~4 mL), and then MeOH (~0.5 mL) was added to dissolve the remaining solid. The solution was purified by radial chromatography (2 mm silica) using CHCl3 (25 mL), %EtOH / CHCl3-1% (50 mL), 2% (75 mL), 3% (50 mL), 4% (50 mL), and 10% (60 mL) to obtain compound 46 (58 mg, 94%), yielding an off-white solid. 1 H NMR (400MHz, acetone-d6) δ10.59(br s, 1H), 8.80(s, 2H), 7.54(d, 1H, J=7.6Hz), 7.37(d, 1H, J=8.4Hz), 7.08(td, 1H, J= 7.6Hz, J=1.2Hz), 7.01(td, 1H, J=7.6Hz, J=1.2), 6.78(d, 1H, J=1.2Hz), 4.42(partially Decomposed dddd, 2H, J=13.6Hz, J=4.4Hz), 3.91 (apparent octet, 1H, J=4.4Hz), 3.84 (d, 1H, J=4.4Hz), 3.42 (ddd, 2H, J=13.4Hz, J=9.8Hz, J=3.2Hz), 1.90 (m, 2H), 1.48 (m, 2H). HRMS(ESI)[M+H] + In C 17 H 19 Calculated value of N4O: 295.1553, measured value: 295.1553.

[0152] 1-(5-(5-fluoro-1H-indole-2-yl)pyrimidine-2-yl)piperidine-4-ol (compound 47) Compound 45 (90mg, 2.18×10 -4 Dissolve (mol) in TFA (2 mL, 26 mmol, 119 equiv.), stir at ambient temperature for 20 minutes, and pour into a mixture of NaHCO3 (2.45 g, 29.16 mmol, 1.1 equiv. TFA), H2O (60 mL), and CH2Cl2 (60 mL). Stir the mixture for 20 minutes, separate the layers, and extract the aqueous layer with CH2Cl2 (10 mL). Wash the combined CH2Cl2 layer with brine (25 mL) and dry on MgSO4. The solution was concentrated and poured onto dry silica (55 mm h × 45 mm id) and eluted under vacuum. From CH2Cl2 (50 mL), %MeOH / CH2Cl2-1% (100 mL), 2.5% (100 mL), 5% (100 mL), and 10% (100 mL), a slightly yellowish solid (54 mg) was obtained. By purification using radial chromatography (2 mm silica) and hexane / siRNA / NET3v / v / v 75:20:5 (100 mL), 50:45:5 (100 mL), and 20:75:5 (250 mL), compound 47 (47 mg, 69%) was obtained as a pale yellowish-brown solid. 1 H NMR (300MHz, acetone-d6) δ10.67(br s, 1H), 8.79(s, 2H), 7.36(dd, 1H, J=8.7Hz, J=5.4Hz), 7.23(dd, 1H, J=9.6Hz, J=2.7Hz), 6.87(ddd, 1H, J=9.6Hz, J=8.7Hz, J=2.7Hz), 6.77(m, 1H), 4.41(partially isolated dddd, 2H, J=13 .5Hz, J=4.5Hz), 3.92 (apparent octet, 1H, J=4.2Hz), 3.83 (d, 1H, J=4.2Hz), 3.43 (ddd, 2H, J= 13.2Hz, J=9.6Hz, J=3.3Hz), 1.90(m, 2H), 1.48(dddd, 2H, J=13.2Hz, J=9.0Hz, J=4.2Hz). HRMS(ESI)[M+H] + In C 17 H 18 FN4O calculated value: 313.1459, measured value: 313.1456.

[0153] 2-Fluoro-6-(piperidine-1-yl)pyridine (compound 48) 2,6-Difluoropyridine (1.34 g, 11.64 mmol), piperidine (1.2 mL, 12.1 mmol, 1 equiv.), i-Pr2NEt (3.2 mL, 18.4 mmol, 1.6 equiv.), and 1,4-Dioxane (10 mL) were stirred under reflux with N2 (g) for 90 minutes and then cooled to ambient temperature. The reaction mixture was concentrated to a yellow oil, CH2Cl2 and hexane were added, and the solution was poured onto dry silica (55 mm h × 45 mm id) and eluted under vacuum. Compound 48 (1.50 g, 71%) was obtained as a colorless oil from hexane (100 mL), %CH2Cl2 / hexane-10% (100 mL), 25% (200 mL), 50% (100 mL), 75% (100 mL), and CH2Cl2 (100 mL). 1 H NMR (300MHz, CDCl3) δ7.48 (dd, 1H, J HF =16.7Hz, J=8.1Hz), 6.40(dd, 1H, J=8.1Hz, J=2.7Hz), 6.09(dd, 1H, J=7.8Hz, J=3.0Hz), 3.51(m, 4H), 1.64(m, 6H), 19 F NMR (282.5MHz, CDCl3)δ-68.60(d, J=6.5Hz), 13 C NMR (125MHz, CDCl3) δ162.96(d, 1 J CF =233.0Hz), 158.58(d, 3 J CF =15.9Hz), 141.73(d, 3 J CF =8.4Hz), 102.69(d, 4 J CF =4.0Hz), 94.86(d, 2 J CF =37.6Hz), 46.12, 25.46, 24.70). HRMS(ESI)[M+H] + In C 10 H 14 FN2 calculated value: 181.1136, measured value: 181.1133.

[0154] 4-(6-fluoropyridine-2-yl)morpholine (compound 49) 2,6-Difluoropyridine (1.34 g, 11.64 mmol), morpholine (1 mL, 11.6 mmol, 1 equiv.), i-Pr2NEt (3 mL, 17.2 mmol, 1.5 equiv.), and 1,4-Dioxane (10 mL) were stirred under reflux with N2 (g) for 90 minutes, cooled to ambient temperature, concentrated to an oil, and dried under vacuum to obtain an off-white solid. The solid was dissolved in CH2Cl2, poured onto dry silica (55 mm h × 45 mm id), and eluted under vacuum. Compound 49 (0.72g, 34%) was obtained as a colorless oil from hexane (50mL), %CH2Cl2 / hexane-25% (50mL), 50% (75mL), 75% (100mL), CH2Cl2 (500mL), %MeOH / CH2Cl2-2% (75mL), 5% (50mL), and 10% (100mL), and gradually turned into a white solid. 1 H NMR (300MHz, CDCl3)δ7.55(dd, 1H, J CF =16.5Hz, J=8.1Hz), 6.41(dd, 1H, J=8.1Hz, J=2.7Hz), 6.21(dd, 1H, J=7.8Hz, J=2.7Hz), 3.80(t, 4H, J=4.8Hz), 3.50(t, 4H, J=4.8Hz), 19 F NMR (282.5MHz, CDCl3)δ-68.47(d, J=7.1Hz), 13 C NMR (125MHz, CDCl3) δ162.87(d, 1 J CF =234.6Hz), 158.64(d, 3 J CF =15.5Hz), 142.07(d, 3 J CF =8.3Hz), 102.74(d, 4 J CF =4.0Hz), 96.74(d, 2 J CF =37.1Hz), 66.65, 45.40. HRMS(ESI)[M+H] + In C9H 12Calculated value of FN2O: 183.0928, measured value: 183.0926.

[0155] 2-Nitro-6-(piperidine-1-yl)pyridine (compound 50) 2-Chloro-6-nitropyridine (0.28 g, 1.77 mmol), piperidine (0.2 mL, 2.0 mmol, 1.1 equiv.), i-Pr2NEt (0.5 mL, 2.9 mmol, 1.6 equiv.), and 1,4-dioxane (5 mL) were stirred under reflux with N2 (g) for 2 hours and then cooled to ambient temperature. The reaction mixture was poured onto dry silica (55 mm h × 45 mm id) and eluted under vacuum. A yellow / orange syrup (0.27 g) was obtained from hexane (50 mL), hexane / siRNA / siRNA 3v / v / v 90:8:2 (100 mL), and 75:20:5 (150 mL). By purification using radial chromatography (2 mm silica), hexane / siRNA / NET3v / v / v 90:8:2 (100 mL), and 75:20:5 (25 mL), compound 50 (0.16 g, 44%) was obtained as an orange syrup. 1 H NMR (300MHz, CDCl3) δ7.64 (dd, 1H, J=8.4Hz, J=7.5Hz), 7.37 (d, 1H, J=7.5Hz), 6.89 (d, 1H, J=8.4Hz), 3.63 (m, 4H), 1.67 (m, 6H).

[0156] 4-(6-nitropyridine-2-yl)morpholine (compound 51) 2-chloro-6-nitropyridine (0.28 g, 1.77 mmol), morpholine (0.2 mL, 2.3 mmol, 1.3 equiv.), i-Pr2NEt (0.5 mL, 2.9 mmol, 1.6 equiv.), and 1,4-dioxane (5 mL) were stirred under reflux under N2 (g) for 4 hours, then cooled to ambient temperature, filtered, and the precipitate was washed with ethyl acetate. The solvent was removed from the filtrate to obtain an orange residue, which was dissolved in CH2Cl2 and poured onto dry silica (55 mm h × 45 mm id) and eluted under vacuum. Yellow / orange solids were obtained from CH2Cl2 (200 mL), %MeOH / CH2Cl2-1% (100 mL), 2.5% (100 mL), and 5% (150 mL). By purification using radial chromatography (2 mm silica), hexane / siRNA / NET3v / v / v 90:8:2 (100 mL), and 75:20:5 (75 mL), compound 51 (0.14 g, 38%) was obtained as a yellow / orange solid. 1 H NMR (300MHz, CDCl3) δ7.73(dd, 1H, J=8.4Hz, J=7.5Hz), 7.49(d, 1H, J=7.5Hz), 6.90(d, 1H, J=8.4Hz), 3.83(t, 4H, J=5.1Hz), 3.63(t, 4H, J=5.1Hz), 13 C NMR (125MHz, CDCl3) δ158.17, 156.06, 140.49, 111.79, 106.21, 66.67, 45.27. HRMS(ESI)[M+H] + In C9H 12 Calculated value of N3O3: 210.0873, measured value: 210.0871.

[0157] 3-Bromo-2-fluoro-6-(piperidine-1-yl)pyridine (compound 52) Compound 48 (0.77 g, 4.27 mmol) was dissolved in CH3CN (25 mL) under N2 (g) and cooled to 0°C. NBS (0.86 g, 4.83 mmol, 1.1 equiv.) was added, and the reaction mixture was stirred at 0°C for 5 minutes, then warmed to ambient temperature and stirred for 6 hours. CH3CN was removed to obtain a pale yellowish-brown residue, which was dissolved in CH2Cl2 and poured onto dry silica (55 mm h × 45 mm id). Elution was performed under vacuum with CH2Cl2 (150 mL) to obtain a colorless oil (1.26 g). Purification by radial chromatography (2 mm silica) and %CH2Cl2 / hexane-10% (100 mL) and 25% (50 mL) yielded compound 52 (0.78 g, 70%) as a colorless oil. 1 ¹H NMR (300MHz, CDCl3) δ 7.57 (apparent t, 1H, J=8.7Hz), 6.33 (dd, 1H, J=8.7Hz, J=1.5Hz), 3.49 (m, 4H), 1.62 (m, 6H), 19 F NMR (470.6MHz, CDCl3)δ-66.52(d, J=8.5Hz), 13 C NMR (125MHz, CD3-OD) δ159.43(d, 1 J CF =229.3Hz), 158.53(d, 3 J CF =14.9Hz), 145.54(d, 3 J CF (=2.4Hz), 106.08(d, 4J) CF =4.4Hz), 87.26(d, 2 J CF =38.4Hz), 47.17, 26.48, 25.66. HRMS(ESI)[M+H] + In C 10 H 13 79 Calculated value of BrFN2: 259.0241, measured value: 259.0238.

[0158] 4-(5-bromo-6-fluoropyridine-2-yl)morpholine (compound 53) Compound 49 (0.36 g, 1.98 mmol) was dissolved in CH3CN (12 mL) under N2 (g) and cooled to 0°C. NBS (0.39 g, 2.19 mmol, 1.1 equiv.) was added, and the reaction mixture was stirred at 0°C for 5 minutes, then warmed to ambient temperature and stirred for 6 hours. CH3CN was removed to obtain a red / brown syrup, which was dissolved in CH2Cl2 and poured onto dry silica (45 mm h × 45 mm id) and eluted under vacuum. A white solid (0.54 g) was obtained by dissolving CH2Cl2 (300 mL), %MeOH / CH2Cl2-1% (100 mL), and 2% (100 mL). By purification using radial chromatography (2 mm silica) and %CH2Cl2 / hexane-25% (100 mL), 50% (100 mL), and 75% (100 mL), compound 53 (0.28 g, 54%) was obtained as a white solid. 1 ¹H NMR (300MHz, CDCl3) δ 7.65 (apparent t, 1H, J=8.7Hz), 6.34 (dd, 1H, J=8.7Hz, J=1.5Hz), 3.79 (t, 4H, J=4.8Hz), 3.48 (t, 4H, J=4.8Hz), 19 F NMR (470.6MHz, CDCl3)δ-66.22(d, J=8.5Hz), 13 C NMR (125MHz, CDCl3) δ158.35(d, 1 J CF =241.5Hz), 157.37(d, 3 J CF =23.8Hz), 144.64(d, 3 J CF =2.6Hz), 104.63(d, 4 J CF =4.5Hz), 89.09(d, 2 J CF =38.5Hz), 66.60, 45.44. HRMS(ESI)[M+H] + In C9H 11 79 Calculated value of BrFN2O: 261.0033, measured value: 261.0063.

[0159] 3-Bromo-2-nitro-6-(piperidine-1-yl)pyridine (compound 54) Compound 50 (0.16g, 7.72×10 -4 The mixture (mol) was flushed with N2 (g), then dissolved in CH3CN (10 mL), and cooled to 0°C. NBS (0.14 g, 7.87 × 10⁻⁶) -4 A mole of 54 (0.15 g, 68%) was added, the reaction mixture was heated to ambient temperature, and stirred for 5.5 hours under N2(g). CH3CN was removed to obtain an orange syrup, then CH2Cl2 and hexane were added and removed to obtain an orange residue, which was dissolved in CH2Cl2 and poured onto dry silica (45 mm h × 45 mm id) and eluted under vacuum. A deep yellow syrup was obtained from hexane (50 mL), %CH2Cl2 / hexane-25% (100 mL), 50% (100 mL), and CH2Cl2 (150 mL). By purification by radial chromatography (2 mm silica), %CH2Cl2 / hexane-25% (100 mL), and 50% (50 mL), compound 54 (0.15 g, 68%) was obtained as a yellow syrup. 1 H NMR (500MHz, CDCl3) δ7.64(d, 1H, J=9.0Hz), 6.66(d, 1H, J=9.0Hz), 3.54(t, 4H, J=5.5Hz), 1.66(m, 2H), 1.62(m, 4H), 13 C NMR (125MHz, CDCl3) δ156.45, 156.14, 144.12, 111.10, 92.85, 46.17, 25.49, 24.52. HRMS(ESI)[M+H] + In C 10 H 13 79 Calculated value of BrN3O2: 286.0186, measured value: 286.0172.

[0160] 4-(5-bromo-6-nitropyridine-2-yl)morpholine (compound 55) Compound 51 (0.12g, 5.74×10 -4 Dissolve (mol) in CH3CN (10 mL), cool to 0°C, and add NBS (0.15 g, 8.43 × 10⁻⁶). -4(mol, 1.5 equiv.) was added. The reaction mixture was stirred in a capped flask at ambient temperature for 6 hours and concentrated to obtain an orange oil. This was then repeated twice, adding and removing CH2Cl2 and hexane to obtain an orange solid, which was dried under vacuum. The solid was dissolved in CH2Cl2 and poured onto dry silica (45 mm h × 45 mm id), and eluted under vacuum. A yellow solid (0.13 g) was obtained from CH2Cl2 (400 mL) and 1% MeOH / CH2Cl2 (100 mL). Purification by radial chromatography (2 mm silica) and CHCl3 (100 mL) yielded compound 55 (82 mg, 50%) as a yellow solid. 1 H NMR (400MHz, CDCl3) δ7.76 (d, 1H, J=9.2Hz), 6.68 (d, 1H, J=9.2Hz), 3.80 (t, 4H, J=4.8Hz), 3.54 (t, 4H, J=4.8Hz), 13 C NMR (125MHz, CDCl3) δ156.65, 156.07, 144.61, 110.99, 94.88, 66.51, 45.21. HRMS(ESI)[M+H] + In C9H 11 79 Calculated value of BrN3O3: 287.9978, measured value: 287.9981.

[0161] 4-(4-fluoropyridine-2-yl)morpholine (compound 56) 2-Bromo-4-fluoropyridine (1.0g, 5.68 mmol), Morpholine (0.50g, 5.74 mmol), Sodium tert-butoxide (0.58g, 6.04 mmol), Toluene (50 mL), Pd2(dba)3 (73 mg, 7.97 × 10⁻¹⁵) -5 (mol) and XantPhos (0.14g, 2.42 × 10) -4 The mixture was flushed with N2(g) for 10 minutes and then heated overnight at 100°C. The reaction mixture was cooled to ambient temperature, filtered through a short silica gel pad, and washed with Â. The filtrate was evaporated to dryness, and the residue was purified by radial chromatography (hexane / Â v / v 95:5 to 80:20) to obtain compound 56 (0.88 g, 85%) as a white solid.1 ¹H NMR (CDCl3, 300MHz): δ 8.13 (dd, 1H, J=9.4Hz, J=5.4Hz), 6.41 (ddd, 1H, J=8.1Hz, J=5.4Hz, J=2.1Hz), 6.28 (dd, 1H, J=12.3Hz, J=2.1Hz), 3.81 (apparent t, 4H, J=4.8Hz), 3.49 (apparent t, 4H, J=4.8Hz). HRMS (ESI) [M+H] + In C9H 12 ON2F calculated value: 183.0928, measured value: 183.0931.

[0162] 4-(5-bromo-4-fluoropyridine-2-yl)morpholine (compound 57) Compound 56 (0.87 g, 4.78 mmol) was dissolved in CH3CN (50 mL), cooled to 0°C, and then NBS (0.85 g, 4.78 mmol) was added. The reaction mixture was stirred at 0°C for 90 minutes, the solvent was removed, and the residue was purified by flash column chromatography (4:1 v / v hexane / siRNA) to obtain compound 57 (1.20 g, 96%) as an off-white solid. 1 ¹H NMR (300 MHz, CDCl3): δ 8.21 (d, 1H, J=9.9 Hz), 6.36 (d, 1H, J=11.5 Hz), 3.80 (apparent t, 4H, J=4.8 Hz), 3.48 (apparent t, 4H, J=4.8 Hz). HRMS (ESI) [M+H] + In C9H 11 Calculated ON2BrF value: 261.0033, measured value: 261.0029.

[0163] 5-Bromo-3-fluoro-2-(piperidine-1-yl)pyridine (compound 58) 2,5-Dibromo-3-fluoropyridine (0.27g, 1.06 mmol), piperidine (85mg, 1.00 mmol), sodium tert-butoxide (144mg, 1.50 mmol), toluene (20mL), Pd2(dba)3 (18mg, 1.97×10) -5 mol), and XantPhos (35 mg, 6.05 × 10) -5The mixture was flushed with N2(g) for 5 minutes and then heated overnight at 100°C. The reaction mixture was cooled to ambient temperature, filtered through a short silica gel pad, and washed with Â. The filtrate was evaporated to dryness, and the residue was purified by radial chromatography (7:3 v / v hexane / Â) to obtain compound 58 (0.15 g, 58%) as a white solid. 1 H NMR (300MHz, CDCl3) δ8.02 (dd, 1H, J=2.1Hz, J=0.9Hz), 7.33 (dd, 1H, J=12.0Hz, J=2.1Hz), 3.41 (m, 4H), 1.65 (m, 6H). HRMS(ESI)[M+H] + In C 10 H 13 Calculated N2BrF value: 259.0241, measured value: 259.0237.

[0164] 4-(5-bromo-3-fluoropyridine-2-yl)morpholine (compound 59) 2,5-Dibromo-3-fluoropyridine (0.27g, 1.06 mmol), Morpholine (87mg, 1.00 mmol), Sodium tert-butoxide (144mg, 1.50 mmol), Toluene (20mL), Pd2(dba)3 (18mg, 1.97×10) -5 mol), and XantPhos (35 mg, 6.05 × 10) -5 The mixture was flushed with N2(g) for 5 minutes and then heated overnight at 100°C. The reaction mixture was cooled to ambient temperature, filtered through a short silica gel pad, and washed with ÃO (4 × 10 mL). The filtrate was evaporated to dryness, and the residue was purified by radial chromatography (hexane / ÃO v / v 95:5 to 80:20) to obtain compound 59 (0.16 g, 61%) as a white solid. 1 H NMR (300MHz, CDCl3) δ8.05 (dd, 1H, J=1.8Hz, J=0.9Hz), 7.39 (dd, 1H, J=12.0Hz, J=1.8Hz), 3.82 (apparent t, 4H, J=4.8Hz), 3.46 (apparent t, 4H, J=4.8Hz). HRMS(ESI)[M+H] + In C9H 11Calculated ON2BrF value: 261.0033, measured value: 261.0044.

[0165] 2-Chloro-3-fluoro-5-(piperidine-1-yl)pyridine (compound 60) 5-Bromo-2-chloro-3-fluoropyridine (0.27g, 1.28mmol), piperidine (88mg, 1.03mmol), sodium tert-butoxide (144mg, 1.50mmol), toluene (20mL), Pd2(dba)3 (18mg, 1.97×10) -5 mol), and XantPhos (35 mg, 6.05 × 10) -5 The mixture was flushed with N2(g) for 5 minutes and then heated overnight at 100°C. The reaction mixture was cooled to ambient temperature, filtered through a short silica gel pad, and washed with ethyl acetate. The filtrate was evaporated to dryness, and the residue was purified by radial chromatography (7:3 v / v hexane / ethyl acetate) to obtain compound 60 (0.22 g, 99%) as a white solid. 1 H NMR (300MHz, CDCl3) δ7.84 (d, 1H, J=2.7Hz), 6.95 (dd, 1H, J=11.4Hz, J=2.7Hz), 3.20 (m, 4H), 1.67 (m, 6H). HRMS(ESI)[M+H] + In C 10 H 13 Calculated value of N2ClF: 215.0746, measured value: 215.0743.

[0166] 4-(6-chloro-5-fluoropyridine-3-yl)morpholine (compound 61) 5-Bromo-2-chloro-3-fluoropyridine (0.27g, 1.28mmol), Morpholine (0.11g, 1.26mmol), Sodium tert-butoxide (144mg, 1.50mmol), Toluene (20mL), Pd2(dba)3 (18mg, 1.97×10) -5 mol), and XantPhos (35 mg, 6.05 × 10) -5The mixture was flushed with N2(g) for 5 minutes and then heated overnight at 100°C. The reaction mixture was cooled to ambient temperature, filtered through a short silica gel pad, and washed with ethyl acetate. The filtrate was evaporated to dryness, and the residue was purified by radial chromatography (7:3 v / v hexane / ethyl acetate) to obtain compound 61 (0.25 g, 92%) as a white solid. 1 H NMR (300MHz, CDCl3) δ7.85 (d, 1H, J=2.7Hz), 6.97 (dd, 1H, J=10.8Hz, J=2.7Hz), 3.87 (m-AA'XX', 4H, J=4.8Hz, J=1.8Hz), 3.19 (m-AA'XX', 4H, J=4.8Hz, J=1.8Hz). HRMS(ESI)[M+H] + In C9H 11 Calculated value of ON2ClF: 217.0539, measured value: 217.0536.

[0167] 4-(6-chloro-4-fluoropyridine-3-yl)morpholine (compound 62) 2-Chloro-4-fluoro-5-bromopyridine (0.27g, 1.28 mmol), morpholine (87mg, 1.00 mmol), sodium tert-butoxide (144mg, 1.50 mmol), toluene (20 mL), Pd2(dba)3 (18mg, 1.97 × 10⁻¹⁰) -5 mol), and XantPhos (35 mg, 6.05 × 10) -5 The mixture was flushed with N2(g) for 5 minutes and then heated overnight at 100°C. The reaction mixture was cooled to ambient temperature, filtered through a short silica gel pad, and washed with Â. The filtrate was evaporated to dryness, and the residue was purified by radial chromatography (hexane / Â v / v 95:5 to 80:20) to obtain compound 62 (97 mg, 45%) as a white solid. 1 H NMR (300MHz, CDCl3) δ7.98 (d, 1H, J=10.5Hz), 7.04 (d, 1H, J=11.1Hz), 3.86 (m-AA'XX', 4H, J=4.8Hz, J=1.8Hz), 3.12 (m-AA'XX', 4H, J=4.8Hz, J=1.8Hz). HRMS(ESI)[M+H] +In C9H 11 Calculated value of ON2ClF: 217.0539, measured value: 217.0536.

[0168] tert-butyl 2-(2-fluoro-6-(piperidine-1-yl)pyridine-3-yl)-1H-indole-1-carboxylate (compound 63) 1,4-Dioxane (30 mL) and H2O (6 mL) were combined and purged with N2 (g) for 30 minutes. N-Boc-Indole-2-boronic acid (compound 6) (0.25 g, 9.58 × 10⁻⁶) -4 mol), compound 52 (0.31g, 1.20mmol, 1.2equiv.), Pd(dppf)Cl2 (95mg, 1.30×10 -4 The mixture was flushed with N2(g) for 10 minutes with mol, 0.14 equiv., and Na2CO3 (0.47g, 4.43 mmol, 4.6 equiv.), then 15 mL of H2O / 1,4-dioxane mixture was added. The reaction mixture was stirred under reflux under N2(g) for 4 hours, then cooled to ambient temperature, filtered through Celite, and washed with 1,4-dioxane and then siRNA. The solvent was removed from the filtrate to obtain a brown oil, which was dried under vacuum to obtain a brown residue. The residue was dissolved in CH2Cl2, poured onto dry silica (55 mm h × 45 mm id), and eluted under vacuum with CH2Cl2 (250 mL) to obtain a very pale yellowish-brown syrup (0.19 g). By purification using radial chromatography (2 mm silica) and 90:8:2 v / v / v hexane / siRNA / NEt3 (100 mL), compound 63 (87 mg, 23%) was obtained as a white foam. 1 H NMR (300MHz, CDCl3) δ8.20(d, 1H, J=8.4Hz), 7.55(m, 2H), 7.31(ddd, 1H, J=8.4Hz, J=7.2Hz, J=1.2Hz), 7.22(td , 1H, J=7.2Hz, J=0.9Hz), 6.53(s, 1H), 6.49(dd, 1H, J=8.4Hz, J=1.8Hz), 3.58(m, 4H), 1.66(m, 6H), 1.45(s, 9H), 19F NMR (282.5MHz, CDCl3) δ-67.96 (d, J=9.6Hz). HRMS(ESI)[M+H] + In C 23 H 27 Calculated value of FN3O2: 396.2082, measured value: 396.2074.

[0169] tert-butyl 2-(2-fluoro-6-morpholinopyridine-3-yl)-1H-indole-1-carboxylate (compound 64) 1,4-Dioxane (30 mL) and H2O (6 mL) were combined and purged with N2 (g) for 30 minutes. N-Boc-Indole-2-boronic acid (6) (0.24 g, 9.19 × 10⁻⁶) -4 mol), compound 53 (0.25g, 9.58×10 -4 mol), Pd(dppf)Cl2(0.11g, 1.50×10 -4 After flushing with N2(g) for 15 minutes with mol (0.16 equiv.) and Na2CO3 (0.30 g, 2.83 mmol, 3.1 equiv.), a H2O / 1,4-dioxane mixture (15 mL) was added. The reaction mixture was stirred under reflux under N2(g) for 4 hours, then cooled to ambient temperature, filtered through Celite, and washed with 1,4-dioxane and then siRNA. The solvent was removed from the filtrate to obtain a brown oil, which was dried under vacuum to obtain a brown residue. The residue was dissolved in CH2Cl2, poured onto dry silica (55 mm h × 45 mm id), and eluted under vacuum. CH2Cl2 (100 mL), %MeOH / CH2Cl2 - 1% (100 mL), 2.5% (100 mL), 5% (150 mL). The target fractions were combined, concentrated, and poured onto dry silica (55 mm h × 45 mm id), and eluted under vacuum. A brown residue (0.23 g) was obtained from hexane / siRNA / NEt3v / v / v 75:20:5 (300 mL) and 50:45:5 (100 mL). Purification by radial chromatography (2 mm silica) and hexane / siRNA / siRNA / NEt3v / v / v 90:8:2 (100 mL) and 75:20:5 (100 mL) yielded compound 64 (0.20 g, 55%) as a white foam. 1H NMR (300MHz, CDCl3) δ8.20(d, 1H, J=8.1Hz), 7.62(dd, 1H, J=9.6Hz, J=8.1Hz), 7.54(d, 1H, J=7.2Hz), 7.32(ddd, 1H, J=8.4Hz, J=7.2Hz, J=1.2H z), 7.23(td, 1H, J=7.5Hz, J=0.9Hz), 6.55(s, 1H), 6.48(dd, 1H, J=8.1Hz, J=1.8Hz), 3.82(t, 4H, J=4.8Hz), 3.54(t, 4H, J=4.8Hz), 1.46(s, 9H), 13 C NMR (125MHz, CDCl3) δ159.39(d, 1 J FC =237.7Hz), 158.10(d, J FC =15.6Hz), 150.26, 142.03(d, J FC =4.8Hz), 137.27, 133.70(d, J FC =4.3Hz), 129.22, 124.62, 123.01, 120.57, 115.69, 110.61, 105.72(d, J FC =31.2Hz), 102.33(d, J FC =3.8Hz), 83.81, 66.73, 45.64, 28.01, 19 F NMR (282.5MHz, CDCl3) δ-67.85 (d, J=9.9Hz). HRMS(ESI)[M+H] + In C 22 H 25 Calculated value of FN3O3: 398.1875, measured value: 398.1867.

[0170] tert-butyl 2-(2-nitro-6-(piperidine-1-yl)pyridine-3-yl)-1H-indole-1-carboxylate (compound 65) 1,4-Dioxane (15 mL) and H2O (3 mL) were combined and purged with N2 (g) for 30 minutes. N-Boc-Indole-2-boronic acid (6) (92 mg, 3.52 × 10⁻⁶) -4 mol), compound 54 (110mg, 3.84×10 -4 mol), Pd(dppf)Cl2(32mg, 4.37×10 -5The mixture was flushed with N2(g) for 10 minutes with mol, 0.1 equiv., and Na2CO3 (196 mg, 1.85 mmol, 5.2 equiv.), then 10 mL of H2O / 1,4-dioxane mixture was added. The reaction mixture was stirred under reflux under N2(g) for 5 hours, then cooled to ambient temperature, filtered through Celite, and washed with ethyl acetate. The solvent was removed from the filtrate to obtain a dark green / black residue, which was dissolved in CH2Cl2 and poured onto dry silica (45 mm h × 45 mm id) and eluted under vacuum. A dark orange syrup (0.14 g) was obtained by eluting with 75:20:5 v / v / v hexane / ethyl acetate / NET3 (200 mL). Purification by radial chromatography (2 mm silica) with hexane / Â / NEt3v / v / v 90:8:2 (100 mL) and 75:20:5 (100 mL) yielded a yellow solid (65 mg). This was further purified by radial chromatography (2 mm silica) using CHCl3 (50 mL) to obtain an orange foamy substance (56 mg). Purification by radial chromatography (1 mm silica) with %CH2Cl2 / hexane-25% (25 mL), 50% (50 mL), and CH2Cl2 (25 mL) yielded compound 65 (48 mg, 32%) as a yellow / orange foamy substance. 1 ¹H NMR (400MHz, CDCl3) δ 8.20 (d, 1H, J=8.4Hz), 7.58 (d, 1H, J=8.8Hz), 7.53 (d, 1H, J=8.0Hz), 7.33 (apparent t, 1H, J=7.6Hz), 7.24 (apparent t - partially obscured by CHCl3 resonance, 1H, J=7.2Hz), 6.86 (d, 1H, J=8.8Hz), 6.50 (s, 1H), 3.66 (m, 4H), 1.67 (m, 6H), 1.41 (s, 9H), 13 C NMR (125MHz, CDCl3) δ157.10, 154.90, 150.07, 142.88, 136.97, 134.21, 129.21, 124.83 , 123.08, 120.72, 116.06, 111.29, 110.65, 109.53, 83.90, 46.24, 27.97, 25.60, 24.74. HRMS(ESI)[M+H] + In C 23 H 27Calculated value of N4O4: 423.2027, measured value: 423.2030. X-ray quality crystals were grown by slow evaporation of CH2Cl2 / hexane.

[0171] tert-butyl 2-(6-morpholino-2-nitropyridine-3-yl)-1H-indole-1-carboxylate (compound 66) 1,4-Dioxane (15 mL) and H2O (3 mL) were combined and purged with N2 (g) for 30 minutes. N-Boc-Indole-2-boronic acid (6) (60 mg, 2.30 × 10⁻⁶) -4 mol), compound 55 (66mg, 2.30×10 -4 mol), Pd(dppf)Cl2(19mg, 2.60×10 -5 The mixture was flushed with N2(g) and Na2CO3 (117 mg, 1.10 mmol, 4.8 equiv.), then H2O / 1,4-dioxane mixture (10 mL) was added. The reaction mixture was stirred under reflux under N2(g) for 4 hours, then cooled to ambient temperature, filtered through Celite, and washed with SiO2. The filtrate was concentrated, then SiO2 was added and removed to obtain a dark green / brown oil. CH2Cl2 and hexane were added and removed to obtain a residue, which was dissolved in CH2Cl2 and poured onto dry silica (45 mm h × 45 mm id) and eluted under vacuum. From CH2Cl2 (200 mL), %MeOH / CH2Cl2-1% (100 mL), and 2% (150 mL), a dark yellow / brown residue (69 mg) was obtained. By purification using radial chromatography (1 mm silica) and CH2Cl2 (100 mL), compound 66 (36 mg, 37%) was obtained as a yellow / orange foamy substance. 1 H NMR(400 MHz, CDCl3)δ8.18(d, 1H, J=8.1Hz), 7.67(d, 1H, J=8.1Hz), 7.54(d, 1H, J=7.6Hz), 7.34(ddd, 1H, J=8.4Hz, J=7.2Hz, J=1.2Hz), 7.25(m-partially obscured by CHCl3 resonance, 1H), 6.88(d, 1H, J=8.4Hz), 6.52(s, 1H), 3.84(t, 4H, J=4.8Hz), 3.65(t, 4H, J=4.8Hz), 1.43(s, 9H),13 C NMR (100MHz, CDCl3) δ157.25, 154.70, 150.06, 143.25, 136.89, 133.83, 129.17, 125 .00, 123.17, 120.83, 116.09, 113.03, 110.87, 109.56, 84.12, 66.67, 45.35, 28.01. HRMS(ESI)[M+H] + In C 22 H 25 Calculated value of N4O5: 425.1820, measured value: 425.1815. X-ray quality crystals were grown by slow evaporation of CH2Cl2 / hexane.

[0172] 4-(5-(1H-indole-2-yl)-6-nitropyridine-2-yl)morpholine (compound 67) It was isolated as a byproduct from the reaction of compound 66. 1 H NMR (400MHz, acetone-d6) δ10.53(br s, 1H), 8.09(d, 1H, J=8.8Hz), 7.56(d, 1H, J=7.6Hz), 7.41(d, 1H, J=8.4Hz), 7.20(d, 1H, J=8.8Hz), 7. 13(t, 1H, J=7.6Hz), 7.04(t, 1H, J=7.6Hz), 6.49(s, 1H), 3.77(t, 4H, J=4.8Hz), 3.62(t, 4H, J=4.8Hz), 13 C NMR (125MHz, DMSO-d6) δ156.31, 154.79, 140.95, 136.77, 130.23, 128.21, 121.94, 120.14, 119.51, 111.30, 110.32, 107.21, 100.09, 65.66, 44.65. HRMS(ESI)[M+H] + In C 17 H 17 Calculated value of N4O3: 325.1295, measured value: 325.1295.

[0173] 2-(2-fluoro-6-(piperidine-1-yl)pyridine-3-yl)-1H-indole (compound 68) Compound 63 (71mg, 1.80×10 -4After stirring mol) and TFA (1.8 mL, 23.4 mmol, 130 equiv.) for 20 minutes, the mixture was poured over a mixture of NaHCO3 (2.29 g, 27.26 mmol, 1.2 equiv. TFA), H2O (45 mL), and CH2Cl2 (45 mL). The mixture was stirred for 10 minutes, the layers were separated, and the aqueous layer was extracted with CH2Cl2 (20 mL). The combined CH2Cl2 layer was washed with brine (25 mL), dried on MgSO4, concentrated, poured onto dry silica (33 mm h × 33 mm id), and eluted under vacuum. Compound 68 (25 mg, 47%) was obtained as an off-white solid from hexane / siRNA / NET3 v / v / v 90:8:2 (25 mL), 75:20:5 (200 mL), and 50:45:5 (50 mL). 1 H NMR (400 MHz, CDCl3)δ8.77(br s, 1H), 7.92(dd, 1H, J=10.6Hz, J=8.4Hz), 7.59(d, 1H, J=8.0Hz), 7.39(d, 1H, J=7.6Hz), 7.17(td, 1H, J=7.6Hz, J=0.8Hz ), 7.10(td, 1H, J=7.6Hz, J=0.8Hz), 6.72(d, 1H, J=1.6Hz), 6.54(dd, 1H, J=8.4Hz, J=2.4Hz), 3.59(m, 4H), 1.67(m, 6H), 13 C NMR (125MHz, DMSO-d6) δ158.00(d, 1 J FC =236.7Hz), 156.04(d, J FC =16.5Hz), 139.16(d, J FC =4.4Hz), 136.42, 131.47(d, J FC =7.3Hz), 128.57, 121.11, 119.60, 119.21, 111.00, 103.91(d, J FC (=3.5Hz), 100.65(d, J) FC =28.0Hz), 99.81(d, J FC =8.3Hz), 45.44, 24.93, 24.09, 19 F NMR (470.6MHz, DMSO-d6)δ-67.52, 19 F NMR (376.5MHz, CDCl3)δ-70.32. HRMS(ESI)[M+H]+ In C 18 H 19 FN3 calculated value: 296.1558, measured value: 296.1604.

[0174] 4-(6-fluoro-5-(1H-indole-2-yl)pyridine-2-yl)morpholine (Compound 69, JSS20-183A) Compound 64 (76mg, 1.91×10 -4 After stirring mol) and TFA (1.8 mL, 23.4 mmol, 123 equiv.) for 20 minutes, the mixture was poured over a mixture of NaHCO3 (2.22 g, 26.43 mmol, 1.1 equiv. TFA), H2O (45 mL), and CH2Cl2 (45 mL). The mixture was stirred for 10 minutes, the layers were separated, and the aqueous layer was extracted with CH2Cl2 (20 mL). The combined CH2Cl2 layer was washed with brine (25 mL), dried on MgSO4, concentrated, poured onto dry silica (33 mm h × 33 mm id), and eluted under vacuum. Compound 69 (54 mg, 95%) was obtained as a pale yellowish-brown solid from hexane / siRNA / Net3 v / v / v 75:20:5 (100 mL), 50:45:5 (200 mL), and 20:75:5 (75 mL). 1 H NMR (400MHz, CDCl3)δ8.78(br s, 1H), 7.99(dd, 1H, J=10.4Hz, J=8.8Hz), 7.60(d, 1H, J=7.6Hz), 7.40(d, 1H, J=8.0Hz), 7.18(td, 1H, J=7.6Hz, J=0.8Hz), 7.11(t d, 1H, J=7.6Hz, J=0.8Hz), 6.77(d, 1H, J=1.2Hz), 6.55(dd, 1H, J=8.4Hz, J=2.0Hz), 3.82(t, 4H, J=4.8Hz), 3.57(t, 4H, J=4.8Hz), 13 C NMR (125MHz, DMSO-d6) δ157.86(d, 1 J FC =237.2Hz), 156.31(d, J FC =16.1Hz), 139.26(d, J FC =4.4Hz), 136.47, 131.13(d, J FC=7.2Hz), 128.50, 121.30, 119.71, 119.27, 111.06, 104.14(d, J FC =3.6Hz), 102.11(d, J FC =27.5Hz), 100.23(d, J FC =8.6Hz), 65.71, 44.83, 19 F NMR (376.5MHz, CDCl3)δ-70.37. HRMS(ESI)[M+H] + In C 17 H 17 Calculated value of FN3O: 298.1350, measured value: 298.1396. X-ray quality crystals grew due to the slow evaporation of CDCl3.

[0175] tert-butyl 5-fluoro-2-(2-morpholinopyrimidine-5-yl)-1H-indole-1-carboxylate (compound 70) Compound 42 (0.21g, 7.52×10 -4 mol), compound 24 (0.10g, 4.10×10 -4 mol), and Pd(dppf)Cl2(20 mg, 2.73 × 10⁻¹⁰) -5 The compound (mol) was dissolved in 10 mL of 1,4-dioxane, and then K2CO3 (aq) (2 M, 0.5 mL) was added. The reaction mixture was flushed with N2 (g) for 10 minutes, and then heated overnight at 100°C under N2 (g). The reaction mixture was cooled to ambient temperature, evaporated to dryness, the residue was dissolved in  (30 mL), washed with H2O (10 mL), dried over MgSO4, and filtered. The filtrate was passed through a silica gel pad, the silica was washed with Â, and the filtrate was concentrated to dryness. The residue was purified by radial chromatography (hexane /  v / v 90:10 to 80:20) to obtain compound 70 (0.15 g, 92%) as a solid. 1H NMR (300MHz, CDCl3) δ8.38(s, 2H), 8.13(dd, 1H, J=9.0Hz, J=4.5Hz), 7.20(dd, 1H, J=8.7Hz, J=2.7H z), 7.05 (td, 1H, J=9.0Hz, J=2.7Hz), 6.50 (d, 1H, J=0.3Hz), 3.88 (m, 4H), 3.79 (m, 4H), 1.49 (s, 9H). HRMS(ESI)[M+H] + In C 21 H 24 Calculated value for O3N4F: 399.1827, measured value: 399.1837.

[0176] tert-butyl 5-fluoro-2-(6-morpholinopyridine-3-yl)-1H-indole-1-carboxylate (compound 71) Compound 42 (0.21g, 7.52×10 -4 mol), compound 29 (0.10g, 4.11×10 -4 mol), Pd(dppf)Cl2(20mg, 2.73×10 -5 The mixture was flushed with 1,4-dioxane (10 mL) and K2CO3 (aq) (2 M, 0.5 mL) under N2 (g) for 10 minutes, then stirred overnight at 100°C under N2 (g). The reaction mixture was cooled to ambient temperature, evaporated to dryness, the residue was dissolved in siRNA (30 mL), washed with H2O (10 mL), dried over MgSO4, and filtered. The filtrate was passed through a silica gel pad, the silica was washed with siRNA, and the filtrate was concentrated to dryness. The residue was purified by radial chromatography (hexane / siRNA v / v 90:10 to 80:20) to obtain compound 71 (0.12 g, 73%) as a solid. 1H NMR (300MHz, CDCl3) δ8.26(d, 1H, J=2.1Hz), 8.12(dd, 1H, J=9.0Hz, J=4.5Hz), 7.55(dd, 1H, J=8.7Hz, J=1.8Hz), 7.19(dd, 1H, J=8.7Hz, J=2.4H z), 7.03(td, 1H, J=9.0Hz, J=2.4Hz), 6.68(d, 1H, J=8.7Hz), 6.48(s, 1H), 3.85(t, 4H, J=4.8Hz), 3.57(partially isolated t, 4H, J=4.8Hz), 1.43(s, 9H). HRMS(ESI)[M+H] + In C 22 H 25 Calculated value for O3N3F: 398.1875, measured value: 398.1887.

[0177] 4-(5-(5-fluoro-1H-indole-2-yl)pyrimidine-2-yl)morpholine (compound 72) Compound 70 (0.14g, 3.51×10 -4 The compound (mol) was dissolved in TFA (2 mL) and stirred at ambient temperature for 2 hours, after which the TFA was removed by a stream of N2 (g). The residue was neutralized with aqueous NaHCO3 to obtain a solid, which was washed with cold CH2Cl2 to obtain compound 72 (58 mg, 55%) as an off-white solid. 1 H NMR (300MHz, acetone-d6) δ10.80(br s, 1H), 8.83(s, 2H), 7.37(dd, 1H, J=8.7Hz, J=4.5Hz), 7.23(dd, 1H, J=9.6Hz, J=2.4Hz ), 6.88(dd, 1H, J=9.6Hz, J=8.7Hz, J=2.4Hz), 6.79(m, 1H), 3.82(m, 4H), 3.71(m, 4H), 13 C NMR (125MHz, DMSO-d6), 160.33, 157.23(d, 1 J FC =231.4Hz), 154.69, 135.15, 133.59, 128.93(d, J FC =10.3Hz), 115.41, 111.92(d, J FC =9.9Hz), 109.27(d, J FC =25.9Hz), 104.31, 104.12, 97.50(d, JFC =4.7Hz), 65.95, 44.01. HRMS(ESI)[M+H] + In C 16 H 16 ON4F calculated value: 299.1303, measured value: 299.1301.

[0178] 4-(5-(5-fluoro-1H-indole-2-yl)pyridine-2-yl)morpholine (compound 73) Compound 71 (0.12g, 3.02×10 -4 The compound (mol) was dissolved in TFA (2 mL), stirred at ambient temperature for 2 hours, and then the TFA was removed by a stream of N2 (g). The residue was neutralized with aqueous NaHCO3 to obtain a solid, which was placed in a filter and washed with a small amount of acetone to obtain compound 73 (66 mg, 74%) as an off-white solid. 1 H NMR (acetone-d6, 300MHz) δ10.62(s, 1H), 8.65(dd, 1H, J=2.5Hz, J=0.7Hz), 7.98(dd, 1H, J=8.9Hz, J=2.6Hz), 7.34(d d, 1H, J=8.8Hz, J=4.5Hz), 7.21(dd, 1H, J=9.9Hz, J=2.5Hz), 6.85(m, 2H), 6.74(s, 1H), 3.74(m, 4H), 3.56(m, 4H), 13 C NMR (125MHz, DMSO-d6), 158.30, 157.17(d, 1 J FC =231.5Hz), 144.57, 137.81, 134.38, 133.54, 129.07(d, J FC =10.6Hz), 117.78, 111.75(d, J FC =9.9Hz), 108.88(d, J FC =26.0Hz), 106.86, 104.05(d, J FC =23.1Hz), 97.18(d, J FC =4.5Hz), 65.90, 45.01. HRMS(ESI)[M+H] + In C 17 H 17 ON3F calculated value: 298.1350, measured value: 298.1348.

[0179] tert-butyl 6-fluoro-2-(2-fluoro-6-(piperidine-1-yl)pyridine-3-yl)-1H-indole-1-carboxylate (compound 74) 1,4-Dioxane (25 mL) was purged with N2 (g) for 1 hour, and compound 52 (0.25 g, 9.65 × 10⁻⁶) was dissolved. -4 mol), compound 4 (0.41 g, 1.03 mmol, 1.1 equiv.), Na2CO3 (0.51 g, 4.81 mmol, 5 equiv.), and Pd(dppf)Cl2 (89 mg, 1.22 × 10⁻¹⁰) -4 After flushing (mol, 0.1 equiv.) with N2(g) for 10 minutes, 1,4-dioxane was added. The reaction mixture was stirred under reflux under N2(g) for 3 hours, then cooled to ambient temperature, filtered through Celite, and washed with ethyl acetate. The filtrate was concentrated to obtain a brown oil, then removed with hexane to obtain a brown residue, which was dissolved in CH2Cl2 and poured onto dry silica (55 mm h × 45 mm id) and eluted under vacuum. An orange syrup (0.43 g) was obtained from hexane (50 mL), hexane / ethyl acetate / Net3 v / v / v 90:8:2 (100 mL), 75:20:5 (100 mL), and 50:45:5 (50 mL). Purification by radial chromatography (2mm silica) with hexane / siRNA / NEt3v / siRNA 95:4:1 (100mL) and 90:8:2 (50mL) yielded an off-white foamy substance (0.16g). Further purification by radial chromatography (2mm silica) with %CH2Cl2 / hexane-25% (50mL), 50% (75mL), 75% (50mL), and CH2Cl2 (25mL) yielded an off-white solid (0.14g). Final purification by radial chromatography (2mm silica) with hexane / siRNA / siRNA / NEt3v / siRNA 90:8:2 (125mL) and 75:20:5 (25mL) yielded compound 74 (0.13g, 33%) as a white solid. 1H NMR (300MHz, CDCl3) δ7.95(dd,1H,J=10.8Hz, J=2.1Hz), 7.54(dd, 1H, J=9.6Hz, J=8.4Hz), 7.44(dd, 1H, J=8.4Hz, J=5 .4Hz), 6.99(td, 1H, J=8.7Hz, J=2.4Hz), 6.54(d, 1H, J=7.5Hz), 6.49(s, 1H), 3.57(m, 4H), 1.67(s, 6H), 1.44(s, 9H), 13 C NMR (100MHz, CDCl3)δ161.06(d, J FC =238.0Hz), 159.46(d, J FC =234.5Hz), 158.04(d, J FC =16.2Hz), 150.10, 141.80(d, J FC =4.8Hz), 137.51(d, J FC =12.8Hz), 134.52(t, J FC =4.3Hz), 125.55(d, J FC =1.1Hz), 120.95(d, J FC =9.8Hz), 111.16(d, J FC =24.1Hz), 109.84, 103.46(d, J FC =31.3Hz), 103.11(d, J FC =28.6Hz), 102.25(d, J FC =3.8Hz), 84.08, 46.45, 27.93, 25.55, 24.84. HRMS(ESI)[M+H] + In C 23 H 26 Calculated value of F2N3O2: 414.1988, measured value: 414.1989.

[0180] 6-Fluoro-2-(2-Fluoro-6-(piperidine-1-yl)pyridine-3-yl)-1H-indole (Compound 75) Compound 74 (92mg, 2.23×10 -4The solution was dissolved in TFA (2.2 mL, 28.6 mmol, 128 equiv.) and stirred at ambient temperature for 20 minutes. This mixture was then poured into a mixture of NaHCO3 (2.68 g, 31.90 mmol, 1.1 equiv. TFA), H2O (55 mL), and CH2Cl2 (55 mL). After stirring the mixture for 10 minutes, the layers were separated, and the H2O layer was extracted with CH2Cl2 (25 mL). The combined CH2Cl2 layers were washed with H2O (50 mL) and brine (50 mL) and dried on MgSO4. The solution was concentrated and poured onto dry silica (55 mm h × 45 mm id) and eluted under vacuum. An off-white / yellowish-brown solid (68 mg) was obtained from hexane (50 mL), hexane / siRNA / NET3 v / v / v 75:20:5 (200 mL), and 50:45:5 (150 mL). This solid was suspended in CHCl3 (1 mL), filtered, washed with CHCl3 (1 mL x 3), then with hexane (5 mL), and dried under vacuum to obtain compound 75 (49 mg, 70%) as a white solid. 1 H NMR (300MHz, DMSO-d6) δ11.36(s, 1H), 8.08(dd, 1H, J=10.8Hz, J=8.7Hz), 7.49(dd, 1H, J=8.7Hz, J=5.7Hz), 7.13(dd, 1H, J=9.9Hz, J=1.8Hz), 6.84(m, 2H), 6.69(s, 1H), 3.56(m, 4H), 1.59(m, 6H), 13 C NMR (125MHz, DMSO-d6) δ158.71(d, 1 J FC =232.9Hz), 157.84(d, 1 J FC =234.8Hz), 157.05(d, J FC =16.3Hz), 139.06(d, J FC =4.4Hz), 136.33(d, J FC =12.9Hz), 132.23(dd, J FC =7.3Hz, J FC =3.6Hz), 125.34, 120.53(d, J FC =10.0Hz), 107.61(d, J FC =24.3Hz), 103.91(d, J FC =3.5Hz), 100.40(d, JFC =27.9Hz), 99.70(d, J FC =8.0Hz), 97.03(d, J FC =25.8Hz), 45.41, 24.92, 24.06. HRMS(ESI)[M+H] + In C 18 H 18 Calculated value for F2N3: 314.1463, measured value: 314.1454.

[0181] (1-(tert-butoxycarbonyl)-6-fluoro-1H-indole-2-yl)boronic acid (compound 76) Compound 104 (2.35 g, 9.99 mmol) was dissolved in THF (100 mL) under N2 (g), cooled to 0°C, and then triisopropyl borate (2.82 g, 14.99 mmol) was added. LDA (2 M THF / heptane / ethylbenzene, 6.5 mL, 13 mmol) was added dropwise over 10 minutes, and the reaction mixture was stirred at 0°C for 2 hours, after which the reaction was stopped with 1 M HCl (aq). The reaction mixture was extracted with RINKAN (100 mL × 3), the combined extract was washed with H2O (2 × 50 mL), dried over MgSO4, filtered, and evaporated to dryness. The residue was recrystallized from 1:2 v / v hexane / RINKAN to obtain compound 76 (1.78 g, 64%) as a white solid. 1 H NMR (300MHz, CDCl3) δ7.72(dd, 1H, J=11.1Hz, J=2.4Hz), 7.52(dd, 1H, J=8.4Hz, J=5.7Hz), 7.45(d, 1H, J=0.6Hz), 7.02(td, 1H, J=8.7Hz, J=2.4Hz) 6.88(br s, 2H), 1.75(s, 9H).

[0182] tert-butyl 6-fluoro-2-(5-morpholinopyrazine-2-yl)-1H-indole-1-carboxylate (compound 77) Compound 76 (0.21g, 7.52×10 -4 mol), compound 30 (0.10g, 4.10×10 -4 mol), Pd(dppf)Cl2(20mg, 2.73×10 -5The mixture was flushed with N2(g) for 10 minutes with 1,4-dioxane (10 mL) and K2CO3(aq)(2M, 0.5 mL), then stirred overnight at 100°C under N2(g). The reaction mixture was cooled to ambient temperature, evaporated to dryness, the residue was dissolved in  (30 mL), washed with H2O (10 mL), dried over MgSO4, and filtered. The filtrate was passed through a silica gel pad, the silica was washed with Â, and the filtrate was concentrated to dryness. The residue was purified by radial chromatography (hexane /  90:10 to 80:20) to obtain compound 77 (78 mg, 48%) as a solid. 1 H NMR (300MHz, CDCl3) δ8.27(d, 1H, J=1.2Hz), 8.17(d, 1H, J=1.5Hz), 7.88(dd, 1H, J=10.8Hz, J=2.4Hz), 7.48(dd, 1H, J=8.7Hz, J=5.7Hz), 7.01(td, 1H, J=9.0Hz, J=2.4Hz), 6.69(s, 1H), 3.86(apparent t, 4H, J=4.8Hz), 3.62(apparent t, 4H, J=4.8Hz), 1.44(s, 9H). HRMS(ESI)[M+H] + In C 21 H 24 Calculated value for O3N4F: 399.1827, measured value: 399.1829.

[0183] tert-butyl 6-fluoro-2-(4-fluoro-6-morpholinopyridine-3-yl)-1H-indole-1-carboxylate (compound 78) Compound 76 (0.34g, 1.22mmol), Compound 57 (0.15g, 5.75×10 -4 mol), Pd(dppf)Cl2(28mg, 3.83×10 -5 Flushing was performed with 1,4-dioxane (30 mL) and K2CO3 (aq) (2 M, 0.7 mL) under N2 (g) for 5 minutes, then stirred overnight at 100°C under N2 (g) (TLC showed that unreacted compound 57 remained). Additional compound 76 (0.15 g, 5.37 × 10⁶) was added. -4Add (mol) and stir the reaction mixture at 100°C overnight under N2(g), then cool to ambient temperature and evaporate to dryness. Dissolve the residue in H2O and extract with RINKAN (10 ml × 3). Dry the combined extract over MgSO4 and filter. Pass the filtrate through a silica gel pad, wash the silica with RINKAN, and remove the solvent to obtain the residue. This residue was purified by radial chromatography (hexane / RINKAN v / v 99:1 to 80:20) to obtain compound 78 (0.15 g, 63%) as a solid. 1 ¹H NMR (500 MHz, CDCl3) δ 8.20 (d, 1H, J=10.5 Hz), 7.92 (dd, 1H, J=10.5 Hz, J=1.5 Hz), 7.47 (dd, 1H, J=9.0 Hz, J=5.5 Hz), 7.01 (td, 1H, J=9.0 Hz, J=2.5 Hz), 6.53 (s, 1H), 6.36 (d, 1H, J=12.5 Hz), 3.83 (t, 4H, J=5.0 Hz), 3.56 (partially separated m, 4H), 1.47 (s, 9H). HRMS (ESI) [M+H] + In C 22 H 24 Calculated value for O3N3F2: 416.1780, measured value: 416.1793.

[0184] tert-butyl 6-fluoro-2-(5-fluoro-6-(piperidine-1-yl)pyridine-3-yl)-1H-indole-1-carboxylate (compound 79) Compound 76 (60mg, 2.15×10 -4 mol), compound 58 (53mg, 2.05×10 -4 mol), Pd(dppf)Cl2(10mg, 1.37×10 -5 The mixture was flushed with N2(g) for 5 minutes, then stirred overnight at 100°C under N2(g) (TLC showed that unreacted compound 58 remained). Additional compound 76 (64 mg, 2.29 × 10⁻⁶) was added. -4The mixture was mixed with (mol) and stirred overnight under N2(g) at 100°C, then cooled to ambient temperature and evaporated to dryness. The residue was dissolved in H2O and extracted with RINKAN (10 ml x 3). The combined extracts were dried over MgSO4 and filtered. The filtrate was passed through a silica gel pad, the silica was washed with RINKAN, and the solvent was removed to obtain the residue. This residue was purified by radial chromatography (hexane / RINKAN v / v 99:1 to 80:20) to obtain compound 79 (59 mg, 70%) as a solid. 1 H NMR(300MHz,CDCl3)δ8.00(m, 1H), 7.89(dd, 1H, J=10.8Hz, J=1.8Hz), 7.41(m, 1H), 7.21(dd, 1H, J=13. 8Hz, J=1.8Hz), 6.96(tt, 1H, J=9.0Hz, J=2.1Hz), 6.47(s, 1H), 3.45(m, 4H), 1.64(m, 6H), 1.39(s, 9H). HRMS(ESI)[M+H] + In C 23 H 26 Calculated value of O2N3F2: 414.1988, measured value: 414.1988.

[0185] tert-butyl 6-fluoro-2-(5-fluoro-6-morpholinopyridine-3-yl)-1H-indole-1-carboxylate (compound 80) Compound 76 (0.19g, 6.81×10 -4 mol), compound 59 (77mg, 2.95×10 -4 mol), Pd(dppf)Cl2(12mg, 1.64×10 -5The mixture was flushed with N2(g) for 10 minutes with 1,4-dioxane (10 mL) and K2CO3(aq)(2M, 0.5 mL), then stirred overnight at 100°C under N2(g). The reaction mixture was cooled to ambient temperature, evaporated to dryness, and the residue was dissolved in H2O and extracted with RINKAN (10 ml × 3). The combined extracts were dried over MgSO4, filtered, and the filtrate was passed through a silica gel pad, washing the silica with RINKAN. The solvent was removed to obtain the residue, which was purified by radial chromatography (hexane / RINKAN v / v 95:5 to 80:20) to obtain compound 80 (86 mg, 70%) as a solid. 1 H NMR (400MHz, CDCl3) δ8.07(s, 1H), 7.92(d, 1H, J=10.8Hz), 7.46(dd, 1H, J=8.8Hz, J=5.6Hz), 7.30(d, 1H, J=13. 6Hz), 7.02 (apparent unseparated td, 1H, J=8.8Hz), 6.53(s, 1H), 3.86(t, 4H, J=4.4Hz), 3.54(t, 4H, J=4.4Hz), 1.45(s, 9H). HRMS(ESI)[M+H] + In C 22 H 24 Calculated value for O3N3F2: 416.1780, measured value: 416.1781.

[0186] 4-(5-(6-fluoro-1H-indole-2-yl)pyrazine-2-yl)morpholine (compound 81) Compound 77 (78mg, 1.96×10 -4 The compound (mol) was dissolved in TFA (2 mL), stirred at ambient temperature for 2 hours, and then the TFA was removed with a stream of N2 (g). The residue was neutralized with aqueous NaHCO3 to obtain a solid, which was washed with cold CH2Cl2 to obtain compound 81 (58 mg, 99%) as an off-white solid. 1H NMR (500MHz, acetone-d6) δ10.75(br s, 1H), 8.72(d, 1H, J=1.0Hz), 8.26(d, 1H, J=1.5Hz), 7.55(dd, 1H, J=9.0Hz, J=5.5Hz), 7.23(dd, 1H, J=10. 0Hz, J=2.0Hz), 6.84(ddd, 1H, J=10.0Hz, J=8.5Hz, J=2.0Hz), 3.79(t, 4H, J=5.0Hz), 3.62(t, 4H, J=5.0Hz), 13 C NMR (125MHz, DMSO-d6) 159.06(d, 1 J FC =234.3Hz), 153.41, 138.40, 136.93(d, J FC =13.2Hz), 136.29, 135.17, 129.82, 125.49, 121.08(d, J FC =9.6Hz), 107.89(d, J FC =24.9Hz), 98.09, 97.51(d, J FC =26.0Hz), 65.81, 44.54. HRMS(ESI)[M+H] + In C 16 H 16 ON4F calculated value: 299.1303, measured value: 299.1302.

[0187] 4-(4-fluoro-5-(6-fluoro-1H-indole-2-yl)pyridine-2-yl)morpholine (compound 82) Compound 78 (0.15g, 3.61×10 -4 The compound (mol) was dissolved in TFA (2 mL), stirred at ambient temperature for 1.5 hours, and then the TFA was removed with a stream of N2 (g). The residue was neutralized with aqueous NaHCO3 to obtain a solid, which was washed with cold CH2Cl2 to obtain compound 82 (0.11 g, 97%) as an off-white solid. 1¹H NMR (300MHz, CDCl3) δ 8.71 (br s, 1H), 8.61 (d, 1H, J=11.1Hz), 7.52 (dd, 1H, J=8.7Hz, J=5.4Hz), 7.08 (dd, 1H, J=9.6Hz, J=2.1Hz), 6.89 (ddd, 1H, J=9.9Hz, J=8.7Hz, J=2.1Hz), 6.80 (dd, 1H, J=1.2Hz), 6.40 (d, 1H, J=14.7Hz), 3.85 (apparent t, 4H, J=4.8Hz), 3.57 (apparent t, 4H, J=4.8Hz), 13 C NMR (125MHz, DMSO-d6) 166.26(d, 1 J FC =257.9Hz), 159.88(d, J FC =4.5Hz), 158.99(d, J FC =241.6Hz), 147.42(d, J FC =4.9Hz), 136.47(d, J FC =12.7Hz), 130.55, 125.23, 120.82(d, J FC =10.1Hz), 107.83(d, J FC =24.4Hz), 107.36(d, J FC =10.9Hz), 100.68(d, J FC =7.0Hz), 97.18(d, J FC =25.7Hz), 93.74(d, J FC =22.6Hz), 65.83, 45.03. HRMS(ESI)[M+H] + In C 17 H 16 ON3F2 calculated value: 316.1256, measured value: 316.1246.

[0188] 6-Fluoro-2-(5-Fluoro-6-(piperidine-1-yl)pyridine-3-yl)-1H-indole (Compound 83) Compound 79 (59mg, 1.43×10 -4The compound (mol) was dissolved in TFA (1 mL) and stirred at ambient temperature for 2.5 hours, after which the TFA was removed by a stream of N2 (g). The residue was neutralized with aqueous NaHCO3 to obtain a solid, which was purified by radial chromatography (hexane / Âi v / v 95:5 to 80:20) to obtain compound 83 (30 mg, 67%) as an off-white solid. 1 H NMR (300MHz, acetone-d6) δ10.74(br s,1H), 8.47(t, 1H, J=1.8Hz), 7.78(dd, 1H, J=14.7Hz, J=2.1Hz), 7.53(dd, 1H, J=8.7Hz, J=5.4Hz), 7.10(dd, 1H, J=9.9Hz, J=2.4Hz), 6.86(m, 1H), 6.84(ddd, 1H, J=9.9Hz, J=8.4Hz, J=2.4Hz), 3.51(m, 4H), 1.67(m, 6H), 13 C NMR (125MHz, DMSO-d6) 159.12(d, J FC =235.1Hz), 149.17(d, J FC =255.3Hz), 148.54(d, J FC =6.4Hz), 139.22(d, J FC =4.5Hz), 137.06(d, J FC =12.7Hz), 135.17, 125.53, 120.91(d, J FC =10.1Hz), 120.73, 119.83(d, J FC =20.8Hz), 107.98(d, J FC =24.4Hz), 98.56, 97.22(d, J FC =25.5Hz), 48.37(d, J FC =5.7Hz), 25.49, 24.31. HRMS(ESI)[M+H] + In C 18 H 18 Calculated value for N3F2: 314.1463, measured value: 314.1455.

[0189] 4-(3-fluoro-5-(6-fluoro-1H-indole-2-yl)pyridine-2-yl)morpholine (compound 84) Compound 80 (80mg, 1.93×10 -4The compound (mol) was dissolved in TFA (2 mL) and stirred at ambient temperature for 1.5 hours, after which the TFA was removed by a stream of N2 (g). The residue was neutralized with aqueous NaHCO3 to obtain a solid, which was washed with cold CH2Cl2 to obtain compound 84 (60 mg, 98%) as an off-white solid. 1 ¹H NMR (300MHz, acetone-d6) δ 10.79 (br s, 1H), 8.51 (apparent t, 1H, J=1.8Hz), 7.84 (dd, 1H, J=14.4Hz, J=1.8Hz), 7.54 (dd, 1H, J=8.7Hz, J=5.4Hz), 7.11 (apparent dd, 1H, J=9.9Hz, J=2.4Hz), 6.89 (m, 1H), 6.85 (ddd, 1H, J=9.9Hz, J=8.7Hz, J=2.4Hz), 3.78 (apparent t, 4H, J=4.8Hz), 3.50 (apparent t, 4H, J=4.8Hz), 13 C NMR (125MHz, DMSO-d6) 159.01(d, J FC =235.2Hz), 149.21(d, J FC =255.4Hz), 147.86(d, J FC =6.8Hz), 139.13(d, J FC =4.7Hz), 136.94(d, J FC =12.7Hz), 134.79(d, J FC =3.2Hz), 125.33, 121.48(d, J FC =3.0Hz), 120.90(d, J FC =10.1Hz), 119.98(d, J FC =20.6Hz), 107.98(d, J FC =24.4Hz), 98.82, 97.17(d, J FC =25.5Hz), 65.97, 47.67(d, J FC (=5.4Hz). HRMS(ESI)[M+H] + In C 17 H 16 ON3F2 calculated value: 316.1256, measured value: 316.1253.

[0190] tert-butyl 2-(4-fluoro-6-morpholinopyridine-3-yl)-1H-indole-1-carboxylate (compound 85) Compound 6 (0.30g, 1.15mmol), Compound 57 (0.15g, 5.75×10 -4 mol), Pd(dppf)Cl2(28mg, 3.83×10 -5 30 mL of 1,4-dioxane (mol) and 0.7 mL of K2CO3 (aq) (2 M) were flushed with N2 (g) for 10 minutes, stirred overnight under N2 (g) at 100°C, then cooled to ambient temperature and evaporated to dryness. The residue was dissolved in H2O and extracted with RINKAN (10 ml x 3). The extracts were combined, dried over MgSO4, and filtered. The filtrate was passed through a silica gel pad, and the silica was washed with RINKAN. The solvent was removed to obtain the residue, which was purified by radial chromatography (hexane / RINKAN v / v 95:5 to 80:20) to obtain compound 85 (0.16 g, 70%) as a solid. 1 ¹H NMR (300MHz, CDCl3) δ 8.22 (d, 1H, J=10.5Hz), 8.19 (d, 1H, J=7.5Hz), 7.56 (dm, 1H, J=7.5Hz), 7.33 (ddd, 1H, J=8.4Hz, J=7.2Hz, J=1.5Hz), 7.24 (td, 1H, J=7.5Hz, J=1.2Hz), 6.57 (s, 1H), 6.36 (d, 1H, J=12.9Hz), 3.83 (apparent t, 4H, J=4.8Hz), 3.56 (apparent t, 4H, J=4.8Hz), 1.48 (s, 9H). HRMS (ESI) [M+H] + In C 22 H 25 Calculated value for O3N3F: 398.1875, measured value: 398.1887.

[0191] tert-butyl 2-(5-fluoro-6-(piperidine-1-yl)pyridine-3-yl)-1H-indole-1-carboxylate (compound 86) Compound 6 (90mg, 3.45×10 -4 mol), compound 58 (90mg, 3.47×10 -4 mol), Pd(dppf)Cl2(19mg, 2.60×10 -5After flushing with N2(g) for 5 minutes with 1,4-dioxane (10 mL) and K2CO3(aq) (2 M, 0.5 mL), the mixture was stirred overnight at 100°C under N2(g) (TLC showed that unreacted compound 58 remained). Additional compound 6 (85 mg, 3.26 × 10⁶) was added. -4 Add (mol) and stir the reaction mixture under N2(g) at 100°C for 5 hours, then cool to ambient temperature and evaporate to dryness. Dissolve the residue in H2O and extract with RINKAN (10 ml x 3). Combine the extracts, dry on MgSO4, and filter. Pass the filtrate through a silica gel pad and wash the silica with RINKAN. Remove the solvent to obtain the residue, which was purified by radial chromatography (hexane / RINKAN v / v 95:5 to 80:20) to obtain compound 86 (0.12 g, 87%) as a solid. 1 ¹H NMR (300 MHz, CDCl3) δ 8.19 (d, 1H, J=8.4 Hz), 8.07 (t, 1H, J=1.5 Hz), 7.54 (apparently unseparated dd, 1H, J=7.8 Hz), 7.33 (partially separated ddd, 1H, J=8.1 Hz, J=7.2 Hz, J=1.2 Hz), 7.26 (indistinct at m-CHCl3 resonance, 3H), 6.55 (s, 1H), 3.50 (m, 4H), 1.69 (m, 6H), 1.44 (s, 9H). HRMS (ESI) [M+H] + In C 23 H 27 Calculated value of O2N3F: 396.2082, measured value: 396.2082.

[0192] tert-butyl 2-(5-fluoro-6-morpholinopyridine-3-yl)-1H-indole-1-carboxylate (compound 87) Compound 6 (0.17g, 6.51×10 -4 mol), compound 59 (75mg, 2.87×10 -4 mol), Pd(dppf)Cl2 (11mg, 1.50×10 -5The mixture was flushed with N2(g) for 5 minutes with 1,4-dioxane (10 mL) and K2CO3(aq)(2M, 0.5 mL), then stirred overnight at 100°C under N2(g). The reaction mixture was cooled to ambient temperature and evaporated to dryness. The residue was dissolved in H2O and extracted with ethyl acetate (10 ml × 3). The extract was dried over MgSO4 and filtered. The filtrate was passed through a silica gel pad, and the silica was washed with ethyl acetate. The solvent was removed to obtain the residue, which was purified by radial chromatography (hexane / ethyl acetate v / v 95:5 to 80:20) to obtain compound 87 (92 mg, 80%) as an off-white solid. 1 H NMR (400MHz, CDCl3) δ8.18(d, 1H, J=8.4Hz), 8.09(s, 1H), 7.55(d, 1H, J=7.6Hz), 7.33(m, 2H), 7.2 6 (obscured by m-CHCl3 resonance, 1H), 6.57 (s, 1H), 3.86 (t, 4H, J=4.4Hz), 3.54 (t, 4H, J=4.4Hz), 1.45 (s, 9H). HRMS(ESI)[M+H] + In C 22 H 25 Calculated value for O3N3F: 398.1875, measured value: 398.1863.

[0193] 4-(4-fluoro-5-(1H-indole-2-yl)pyridine-2-yl)morpholine (compound 88) Compound 85 (54mg, 1.36×10 -4 The compound (mol) was dissolved in TFA (2 mL), stirred at ambient temperature for 2 hours, and then the TFA was removed with a stream of N2 (g). The residue was neutralized with aqueous NaHCO3 to obtain a solid, which was washed with cold CH2Cl2 to obtain compound 88 (30 mg, 74%) as an off-white solid. 1¹H NMR (300MHz, acetone-d6) δ 10.52 (br s, 1H), 8.66 (d, 1H, J=11.4Hz), 7.56 (d, 1H, J=8.1Hz), 7.42 (d, 1H, J=8.1Hz), 7.10 (partially separated ddd, 1H, J=8.1Hz, J=6.9Hz, J=1.2Hz), 7.02 (partially separated ddd, 1H, J=7.8Hz, J=7.2Hz, J=1.2Hz), 6.81 (m, 1H), 6.71 (d, 1H, J=14.7Hz), 3.76 (apparent t, 4H, J=4.8Hz), 3.59 (apparent t, 4H, J=4.8Hz), 13 C NMR (125MHz, DMSO-d6) δ166.33(d, J FC =257.8Hz), 159.93(d, J FC =11.4Hz), 147.61(d, J FC =3.8Hz), 136.49, 129.80, 128.42, 121.48, 119.78, 119.32, 111.12, 107.52(d, J FC =10.1Hz), 100.69(d, J FC =6.3Hz), 93.66(d, J FC =22.5Hz), 65.81, 45.00. HRMS(ESI)[M+H] + In C 17 H 17 ON3F calculated value: 298.1350, measured value: 298.1343.

[0194] 2-(5-fluoro-6-(piperidine-1-yl)pyridine-3-yl)-1H-indole (compound 89) Compound 86 (0.11g, 2.78×10 -4 The compound (mol) was dissolved in TFA (2 mL) and stirred at ambient temperature for 2 hours, after which the TFA was removed by a stream of N2 (g). The residue was neutralized with aqueous NaHCO3 to obtain a solid, which was washed with cold CH2Cl2 to obtain compound 89 (78 mg, 95%) as an off-white solid. 1¹H NMR (300MHz, acetone-d6) δ 10.61 (br s, 1H), 8.50 (t, 1H, J=1.8Hz), 7.80 (dd, 1H, J=14.7Hz, J=1.8Hz), 7.54 (apparent d, 1H, J=8.1Hz), 7.38 (dd, 1H, J=8.7Hz, J=0.6Hz), 7.10 (ddd, 1H, J=8.1Hz, J=6.9Hz, J=1.2Hz), 7.02 (ddd, 1H, J=8.1Hz, J=6.9Hz, J=1.2Hz), 6.85 (m, 1H), 3.50 (m, 4H), 1.67 (m, 6H), 13 C NMR (125MHz, DMSO-d6) 149.03(d, J FC =255.2Hz), 148.34(d, J FC =6.4Hz), 139.27(d, J FC =4.6Hz), 137.00, 134.25, 128.59, 121.49, 120.81(d, J FC =2.8Hz), 119.82(d, J FC =20.8Hz), 119.80, 119.43, 111.08, 98.44, 48.25(d, J FC =5.8Hz), 25.37, 24.19. HRMS(ESI)[M+H] + In C 18 H 19 N3F calculated value: 296.1558, measured value: 296.1550.

[0195] 4-(3-fluoro-5-(1H-indole-2-yl)pyridine-2-yl)morpholine (compound 90) Compound 87 (90mg, 2.26×10 -4 The compound (mol) was dissolved in TFA (2 mL), stirred at ambient temperature for 1 hour, and then the TFA was removed by a stream of N2 (g). The residue was neutralized with aqueous NaHCO3 to obtain a solid, which was purified by radial chromatography to obtain compound 90 (60 mg, 89%) as a solid. 1¹H NMR (300MHz, acetone-d6) δ 10.65 (br s, 1H), 8.54 (apparent t, 1H, J=1.8Hz), 7.86 (dd, 1H, J=14.4Hz, J=1.8Hz), 7.55 (d, 1H, J=7.8Hz), 7.39 (partially separated dd, 1H, J=8.1Hz, J=0.9Hz), 7.11 (ddd, 1H, J=8.7Hz, J=6.9Hz, J=1.2Hz), 7.02 (partially separated ddd, 1H, J=8.1Hz, J=7.8Hz, J=1.2Hz), 6.88 (m, 1H), 3.78 (apparent t, 4H, J=4.8Hz), 3.50 (apparent t, 4H, J=4.8Hz), 13 C NMR (125MHz, DMSO-d6) 149.29(d, J FC =255.4Hz), 147.87(d, J FC =6.9Hz), 139.24(d, J FC =4.8Hz), 137.09, 136.93, 133.83(d, J FC =10.2Hz), 128.49(d, J FC =10.4Hz), 121.65, 120.08(d, J FC =20.5Hz), 119.88, 119.49, 111.10, 98.78, 66.01, 47.73(d, J FC (=5.4Hz). HRMS(ESI)[M+H] + In C 17 H 17 ON3F calculated value: 298.1350, measured value: 298.1348.

[0196] tert-butyl 2-(3-fluoro-5-(piperidine-1-yl)pyridine-2-yl)-1H-indole-1-carboxylate (compound 91) Compound 6 (0.22g, 8.43×10 -4 mol), compound 60 (90mg, 4.19×10 -4 mol), Pd(dppf)Cl2(19mg, 2.60×10 -5Flushing was performed with 1,4-dioxane (10 mL) and K2CO3 (aq) (2 M, 0.5 mL) under N2 (g) for 5 minutes, followed by stirring overnight at 100°C under N2 (g) (TLC showed that unreacted compound 60 remained). Additional compound 6 (90 mg, 3.45 × 10⁻⁶) was added. -4 Add (mol) and stir overnight at 100°C under N2(g). The reaction mixture was cooled to ambient temperature and evaporated to dryness. The residue was dissolved in H2O and extracted with Â(10 ml × 3). The extract was dried over MgSO4 and filtered. The filtrate was passed through a silica gel pad, and the silica was washed with Â(.). The solvent was removed to obtain the residue, which was purified by radial chromatography (hexane / Â(v / v) 95:5 to 80:20) to obtain compound 91 (75 mg, 45%) as a solid. 1 H NMR (300MHz, CDCl3) δ8.19 (dd, 1H, J=8.4Hz, J=0.6Hz), 8.15 (dd, 1H, J=2.7Hz, J=1.5Hz), 7.57 (d, 1H, J=7.5Hz), 7.33 (ddd, 1H, J=8.4Hz, J=7.2Hz, J=1. 2Hz), 7.23 (apparent td - partially obscured by CHCl3 resonance, 1H, J=7.5Hz, J=0.9Hz), 6.91(dd, 1H, J=12.6Hz, J=2.4Hz), 6.78(s, 1H), 3.28(m, 4H), 1.70(m, 6H), 1.42(s, 9H). HRMS(ESI)[M+H] + In C 23 H 27 Calculated value of O2N3F: 396.2082, measured value: 396.2082.

[0197] 2-(3-fluoro-5-(piperidine-1-yl)pyridine-2-yl)-1H-indole (compound 92) Compound 91 (75mg, 1.90×10 -4 The compound (mol) was dissolved in TFA (1 mL) and stirred at ambient temperature for 1 hour, after which the TFA was removed by a stream of N2 (g). The residue was neutralized with aqueous NaHCO3 to obtain a solid, which was washed with cold CH2Cl2 to obtain compound 92 (44 mg, 78%) as an off-white solid. 1¹H NMR (300MHz, acetone-d6) δ 10.60 (br s, 1H), 8.20 (t, 1H, J=2.4Hz), 7.58 (partially separated dd, 1H, J=8.1Hz), 7.55 (partially separated dd, 1H, J=8.1Hz, J=0.9Hz), 7.19 (dd, 1H, J=14.7Hz, J=2.4Hz), 7.12 (ddd, 1H, J=8.1Hz, J=7.2Hz, J=1.2Hz), 7.01 (ddd, 1H, J=7.8Hz, J=7.2Hz, J=1.2Hz), 6.94 (m, 1H), 3.37 (m, 4H), 1.70 (m, 6H), 13 C NMR (125MHz, DMSO-d6) 157.07(d, J FC =258.7Hz), 147.40(d, J FC =5.4Hz), 136.20, 132.75(d, J FC =7.2Hz), 132.42, 128.70, 127.30(d, J FC =12.2Hz), 121.70, 120.22, 119.15, 111.69, 108.77(d, J FC =22.6Hz), 100.67(d, J FC =11.9Hz), 48.18, 24.72, 23.64. HRMS(ESI)[M+H] + In C 18 H 19 N3F calculated value: 296.1558, measured value: 296.1562.

[0198] tert-butyl 2-(3-fluoro-5-morpholinopyridine-2-yl)-1H-indole-1-carboxylate (compound 93). Compound 6 (0.17g, 6.51×10 -4 mol), compound 61 (78mg, 3.60×10 -4 mol), Pd(dppf)Cl2 (18mg, 2.46×10 -5The mixture was flushed with N2(g) for 5 minutes with 1,4-dioxane (10 mL) and K2CO3(aq)(2M, 0.5 mL), then stirred overnight at 100°C under N2(g). The reaction mixture was cooled to ambient temperature and evaporated to dryness. The residue was dissolved in H2O and extracted with ethyl acetate (10 ml × 3). The extract was dried over MgSO4 and filtered. The filtrate was passed through a silica gel pad, and the silica was washed with ethyl acetate. The solvent was removed to obtain the residue, which was purified by radial chromatography (hexane / ethyl acetate v / v 95:5 to 80:2) to obtain the recovered compound 61 (58 mg, 74%) and compound 93 (29 mg, 20%) as solids. 1 ¹H NMR (400MHz, CDCl3) δ 8.18 (d, 1H, J=8.4Hz), 8.15 (s, 1H), 7.58 (d, 1H, J=7.6Hz), 7.34 (apparent t, 1H, J=7.6Hz), 7.24 (apparent t, 1H, J=7.2Hz), 6.92 (dd, 1H, J=12.0Hz, J=2.0Hz), 6.81 (s, 1H), 3.90 (t, 4H, J=4.8Hz), 3.26 (t, 4H, J=4.8Hz), 1.43 (s, 9H). HRMS (ESI) [M+H] + In C 22 H 25 Calculated value for O3N3F: 398.1875, measured value: 398.1865.

[0199] tert-butyl 6-fluoro-2-(3-fluoro-5-morpholinopyridine-2-yl)-1H-indole-1-carboxylate (compound 94) Compound 76 (0.19g, 6.81×10 -4 mol), compound 61 (87mg, 4.02×10 -4 mol), Pd(dppf)Cl2(19mg, 2.60×10 -5 Flushing was performed with N2(g) for 5 minutes with 1,4-dioxane (10 mL) and K2CO3(aq) (2 M, 0.5 mL), followed by stirring overnight at 100°C under N2(g) (TLC showed that unreacted compound 61 remained). Additional compound 76 (93 mg, 3.33 × 10⁻⁶) was added. -4Add (mol) and stir the reaction mixture under N2(g) at 100°C for 5 hours, then cool to ambient temperature and evaporate to dryness. Dissolve the residue in H2O and extract with Â(10 ml × 3). Combine the extracts, dry over MgSO4, and filter. Pass the filtrate through a silica gel pad and wash the silica with Â. Remove the solvent to obtain the residue, which was purified by radial chromatography (hexane / Â(v / v) 95:5 to 80:20) to obtain the recovered compound 61 (23 mg, 26%) and compound 94 (96 mg, 57%) as solids. 1 ¹H NMR (400 MHz, CDCl3) δ 8.15 (s, 1H), 7.91 (partially separated dd, 1H, J=10.8 Hz, J=1.2 Hz), 7.49 (dd, 1H, J=8.4 Hz, J=5.6 Hz), 7.00 (apparent td, 1H, J=9.6 Hz, J=1.6 Hz), 6.92 (apparent dd, 1H, J=12.4 Hz, J=1.6 Hz), 6.76 (s, 1H), 3.90 (t, 4H, J=4.8 Hz), 3.26 (t, 4H, J=4.8 Hz), 1.43 (s, 9H). HRMS (ESI) [M+H] + In C 22 H 24 Calculated value for O3N3F2: 416.1780, measured value: 416.1767.

[0200] tert-butyl 2-(4-fluoro-5-morpholinopyridine-2-yl)-1H-indole-1-carboxylate (compound 95) Compound 6 (85mg, 3.26×10 -4 mol), compound 62 (78mg, 3.60×10 -4 mol), Pd(dppf)Cl2 (18mg, 2.46×10 -51,4-dioxane (5 mL), and K2CO3 (aq) (2 M, 0.5 mL) were stirred overnight at 100°C under N2 (g). The reaction mixture was cooled to ambient temperature, evaporated to dryness, and the residue was dissolved in H2O and extracted with  (10 ml × 3). The combined extracts were dried over MgSO4, filtered, and the filtrate was passed through a silica gel pad and washed with Â. The solution was concentrated to dry, and the residue was purified by radial chromatography (hexane /  v / v 95:5 to 70:30) to obtain recovered compound 62 (32 mg, 41%) and compound 95 (45 mg, 35%). 1 ¹H NMR (300MHz, CDCl3) δ 8.28 (d, 1H, J=11.1Hz), 8.15 (d, 1H, J=8.1Hz), 7.57 (d, 1H, J=7.8Hz), 7.35 (partially separated ddd, 1H, J=8.4Hz, J=7.2Hz, J=1.2Hz), 7.25 (m-partially obscured by CHCl3 resonance, 1H), 7.19 (d, 1H, J=12.9Hz), 6.75 (s, 1H), 3.90 (apparent t, 4H, J=4.5Hz), 3.21 (apparent t, 4H, J=4.5Hz), 1.42 (s, 9H). HRMS (ESI) [M+H] + In C 22 H 25 Calculated value for O3N3F: 398.1875, measured value: 398.1865.

[0201] tert-butyl 6-fluoro-2-(4-fluoro-5-morpholinopyridine-2-yl)-1H-indole-1-carboxylate (compound 96) Compound 76 (0.23g, 8.24×10 -4 mol), compound 62 (97mg, 4.48×10 -4 mol), Pd(dppf)Cl2 (18mg, 2.46×10 -510 mL of 1,4-dioxane (mol) and 0.5 mL of K₂CO₃ (aq) (2 M) were flushed with N₂ (g) for 5 minutes, then stirred under N₂ (g) and heated overnight at 100°C. The reaction mixture was cooled to ambient temperature, evaporated to dryness, and the residue was dissolved in H₂O and extracted with ÃO (10 ml × 3). The combined extracts were dried over MgSO₄, filtered, and the filtrate was passed through a silica gel pad and washed with ÃO. The solution was concentrated until dry, and the residue was purified by radial chromatography (hexane / ÃO v / v 95:5 to 80:20) to obtain recovered compound 62 (68 mg, 70%) and compound 96 (29 mg, 16%) as solids. 1 H NMR (300MHz, CDCl3) δ8.27(d, 1H, J=10.8Hz), 7.88(dd, 1H, J=10.5Hz, J=2.4Hz), 7.48(dd, 1H, J=8.7Hz, J=5.4Hz), 7.18(d, 1H, J=12.9Hz), 7 .01(td, 1H, J=9.0Hz, J=2.4Hz), 6.70(s, 1H), 3.90(m-AA'XX', 4H, J=4.8Hz, J=1.8Hz), 3.21(m-AA'XX', 4H, J=4.8Hz, J=1.8Hz), 1.41(s, 9H). HRMS(ESI)[M+H] + In C 22 H 24 Calculated value for O3N3F2: 416.1780, measured value: 416.1767.

[0202] 4-(5-fluoro-6-(1H-indole-2-yl)pyridine-3-yl)morpholine (compound 97) Compound 93 (29mg, 7.30×10 -5 The compound (mol) was dissolved in TFA (1 mL), stirred at ambient temperature for 2 hours, and then the TFA was removed with a stream of N2 (g). The residue was neutralized with aqueous NaHCO3 to obtain a solid, which was washed with cold CH2Cl2 to obtain compound 97 (17 mg, 78%) as an off-white solid. 11H NMR (400 MHz, acetone-d6) δ 10.65 (br s, 1H), 8.22(t, 1H, J=2.0Hz), 7.60(unseparated dq, 1H, J=8.0Hz), 7.56(partially separated dq, 1H, J=8.0Hz, J=0.8Hz), 7.24(dd, 1H, J=14.4Hz, J=2.4Hz), 7.14(ddd, 1H, J=8.8Hz, J=7.2Hz, J=1.2Hz), 7.02(ddd, 1H, J=7.8Hz, J=7.2Hz, J=1.2Hz), 6.98(dm, 1H, J=4.4Hz), 3.81(apparent t, 4H, J=4.8Hz), 3.32(partially separated m-AA'XX', 4H, J=4.8Hz), 13 C NMR (125MHz, DMSO-d6) 156.94(d, J FC =258.6Hz), 147.20(d, J FC =4.3Hz), 136.24, 132.53(d, J FC =6.4Hz), 132.14, 128.65, 128.46(d, J FC =10.9Hz), 121.88, 120.33, 119.22, 111.74, 108.95(d, J FC =22.1Hz), 101.06(d, J FC =11.9Hz), 65.69, 47.23. HRMS(ESI)[M+H] + In C 17 H 17 ON3F calculated value: 298.1350, measured value: 298.1353.

[0203] 4-(5-fluoro-6-(6-fluoro-1H-indole-2-yl)pyridine-3-yl)morpholine (compound 98) Compound 94 (31mg, 7.46×10 -5 The compound (mol) was dissolved in TFA (1 mL), stirred at ambient temperature for 1 hour, and then the TFA was removed with a stream of N2 (g). The residue was neutralized with aqueous NaHCO3 to obtain a solid, which was washed with cold CH2Cl2 to obtain compound 98 (23 mg, 98%) as an off-white solid. 1¹H NMR (300MHz, acetone-d6) δ 10.77 (br s, 1H), 8.21 (t, 1H, J=1.8Hz), 7.58 (dd, 1H, J=8.7Hz, J=5.4Hz), 7.30 (dd, 1H, J=10.2Hz, J=2.4Hz), 7.23 (dd, 1H, J=14.4Hz, J=2.4Hz), 6.96 (dm, 1H, J=4.2Hz), 6.85 (ddd, 1H, J=9.9Hz, J=8.7Hz, J=2.4Hz), 3.81 (apparent t, 4H, J=4.8Hz), 3.32 (apparent t, 4H, J=4.8Hz), 13 C NMR (125MHz, DMSO-d6) 159.18(d, J FC =235.3Hz), 156.82(d, J FC =258.8Hz), 147.26(d, J FC =5.5Hz), 136.24(d, J FC =13.2Hz), 133.33 (undisclosed m), 132.13 (d, J FC =2.8Hz), 128.15(d, J FC =11.9Hz), 125.49, 121.46(d, J FC =10.2Hz), 108.93(d, J FC =22.8Hz), 107.83(d, J FC =24.5Hz), 101.06(d, J FC =12.1Hz), 97.51(d, J FC =25.7Hz), 65.69, 47.21. HRMS(ESI)[M+H] + In C 17 H 16 ON3F2 calculated value: 316.1256, measured value: 316.1258.

[0204] 4-(4-fluoro-6-(1H-indole-2-yl)pyridine-3-yl)morpholine (compound 99) Compound 95 (40mg, 1.01×10 -4The compound (mol) was dissolved in TFA (1.5 mL), stirred at ambient temperature for 1.5 hours, and then the TFA was removed with a stream of N2 (g). The residue was neutralized with aqueous NaHCO3 to obtain a solid, which was washed with hexane and cold CH2Cl2 to obtain compound 99 (29 mg, 97%) as an off-white solid. 1 ¹H NMR (300 MHz, acetone-d6) δ 10.73 (br s, 1H, 1.2Hz), 7.07(m, 1H), 7.02(ddd, 1H, J=7.8Hz, J=7.2Hz, J=0.9Hz), 3.82(m-AA'XX', 4H, J=4.8Hz, J=1.8Hz), 3.19(m-AA'XX', 4H, J=4.8Hz, J=1.8Hz), 13 C NMR (125MHz, DMSO-d6) 160.55(d, J FC =258.7Hz), 146.18(d, J FC (=8.0Hz), 140.75 (unseparated d), 137.10, 136.37 (unseparated d), 134.62 (d, J FC =5.2Hz), 128.33, 122.15, 120.43, 119.43, 111.87, 107.55(d, J FC =18.4Hz), 100.24, 65.99, 50.08. HRMS(ESI)[M+H] + In C 17 H 17 ON3F calculated value: 298.1350, measured value: 298.1347.

[0205] 4-(4-fluoro-6-(6-fluoro-1H-indole-2-yl)pyridine-3-yl)morpholine (compound 100) Compound 96 (29mg, 6.98×10 -5(0 mol) was dissolved in TFA (1 mL) and stirred at ambient temperature for 2 h, then the TFA was removed under a stream of N2(g). The residue was neutralized with aqueous NaHCO3 to give a solid, which was washed with cold CH2Cl2 to afford compound 100 (21 mg, 95%) as an off-white solid. 1 1H NMR (300 MHz, acetone-d6) δ 10.86 (br s, 1H), 8.27 (d, 1H, J = 11.1 Hz), 7.71 (d, 1H, J = 13.8 Hz), 7.57 (dd, 1H, J = 8.7 Hz, J = 5.4 Hz), 7.26 (dd, 1H, J = 9.9 Hz, J = 2.4 Hz), 7.08 (m, 1H), 6.85 (ddd, 1H, J = 9.9 Hz, J = 8.7 Hz, J = 2.4 Hz), 3.82 (m-AA’XX’, 4H, J = 4.8 Hz, J = 1.8 Hz), 3.19 (m-AA’XX’, 4H, J = 4.8 Hz, J = 1.8 Hz), 13 13C NMR (125 MHz, DMSO-d6) 160.56 (d, J FC = 258.8 Hz), 159.30 (d, J FC = 235.7 Hz), 145.85 (d, J FC = 8.3 Hz), 140.71 (d, J FC = 3.0 Hz), 137.16 (dd, J FC = 3.6 Hz), 137.03 (d, J FC = 13.2 Hz), 134.69 (d, J FC = 5.8 Hz), 125.17, 121.59 (d, J FC = 10.2 Hz), 108.11 (d, J FC = 24.5 Hz), 107.46 (d, J FC = 18.5 Hz), 100.26, 97.63 (d, J FC = 25.8 Hz), 65.99, 50.07 (d, J FC = 2.6 Hz). HRMS (ESI) [M+H] + In 17 C 16 15H16ON3F2, calcd: 316.1256, found: 316.1245.

[0206] tert-butyl 2-(2-(4-(2-(2-fluoroethoxy)ethoxy)piperidine-1-yl)pyrimidine-5-yl)-1H-indole-1-carboxylate (compound 101) Compound 44 (79 mg, 2.00 × 10⁻⁶) -4 Flushing (mol) with N2 (g), dissolved in DMA (1 mL), and cooled to 0°C. NaH (60%, 26 mg, 6.50 × 10⁻⁶) -4 Add 3.2 mol(equiv.) and stir the mixture under N2(g) at 0°C for 5 minutes. Then add 110 mg (4.19 × 10⁻¹) of 2-(2-fluoroethoxy)ethyl 4-methylbenzenesulfonate in 0.5 mL of DMA. -4 A solution of 2.1 mol (equiv.) was added. The reaction mixture was stirred under N2 (g) at ambient temperature for 2 hours, then H2O (5 drops) was added, the mixture was stirred for 10 minutes, and then poured into a mixture of H2O (20 mL), siRNA (20 mL), and hexane (10 mL). The layers were mixed and separated, the organic layer was washed with H2O (10 mL × 2) and brine (10 mL), and dried on MgSO4. The solvent was removed to obtain a yellow residue (138 mg), which was dissolved in CH2Cl2 and poured onto dry silica (55 cm h × 45 cm id) and eluted under vacuum. From CH2Cl2 (100 mL), %MeOH / CH2Cl2-1% (100 mL), 2% (100 mL), 3% (100 mL), and 5% (100 mL), a faint brown syrup (126 mg) was obtained. By purification using radial chromatography (1 mm silica), CH2Cl2 (50 mL), %MeOH / CH2Cl2-1% (50 mL), and 2% (25 mL), compound 101 (32 mg, 33%) was obtained as a colorless residue. 11H NMR (300 MHz, CDCl3) δ 8.48 (s, 2H), 7.62 (d, 1H, J = 7.8 Hz), 7.41 (d, 1H, J = 8.1 Hz), 7.24 (td, 1H, J = 8.1 Hz, J = 0.9 Hz), 7.15 (partially resolved td, 1H, J = 7.8 Hz, J = 0.9 Hz), 6.50 (s, 1H), 4.86 (apparent septet, 1H, J = 4.2 Hz), 4.52 (apparent t, 1H, J = 4.2 Hz), 4.39 (m, 3H), 4.30 (t, 2H, J = 6.0 Hz), 3.80 (t, 2H, J = 6.0 Hz), 3.55 (m, 4H), 2.06 (m, 2H), 1.77 (m, 2H), 1.52 (s, 9H).

[0207] 2-(2-(4-(2-(2-Fluoroethoxy)ethoxy)piperidin-1-yl)pyrimidin-5-yl)-1H-indole (Compound 102) Compound 101 (31 mg, 6.40×10 <所提供的文本中“ -5 ”后面似乎缺少内容,请确认是否完整。这里暂时保留原样。mol) and TFA (0.65 mL, 8.4 mmol, 132 equiv.) were stirred for 20 minutes and then poured into a mixture of NaHCO3 (0.79 g, 9.40 mmol, 1.1 equiv. TFA), H2O (25 mL) and CH2Cl2 (25 mL). The mixture was stirred for 5 minutes, the layers were separated, and the aqueous layer was extracted with CH2Cl2 (5 mL × 2). The combined CH2Cl2 layers were washed with brine (20 mL), dried over MgSO4, concentrated, poured onto dry silica (33 mm h × 33 mm i.d.), and eluted under vacuum. %MeOH / CH2Cl2 - 1% (50 mL), 2% (75 mL), 3% (100 mL) gave Compound 102 (16 mg, 65%) as an off-white solid. 1 1H NMR (MHz, CDCl3) δ 8.48 (s, 2H), 7.62 (d, 1H, J = 7.6 Hz), 7.41 (d, 1H, J = 8.0 Hz), 7.24 (td, 1H, J = 8.0 Hz, J = 0.8 Hz), 7.15 (partially resolved td, 1H, J = 7.6 Hz, J = 0.8 Hz), 6.50 (s, 1H), 4.48 (dt, 2H, J = 13.4 Hz, J = 4.4 Hz), 4.44 (dt, 2H, 2 J HF=48.0Hz, J=4.4Hz), 4.30(t, 2H, J=6.0Hz), 4.01(apparent septet, 1H, J=4.4Hz), 3.80(t, 2H, J=6.0Hz), 3.55(dt, 2H, 3 J HF =29.2Hz, J=4.4Hz), 3.42(ddd, 2H, J=13.4Hz, J=10.0Hz, J=3.6Hz), 2.01(m, 2H), 1.58(m, 2H). HRMS(ESI)[M+H] + In C 21 H 26 Calculated value of FN4O2: 385.2034, measured value: 385.2041.

[0208] 4-(5-(6-fluoro-1-methyl-1H-indole-2-yl)pyrimidine-2-yl)morpholine (compound 103) Compound 2 (21mg, 7.04×10 -5 After flushing with N2(g) (mol), the solution was dissolved in DMA(1mL) and cooled to 0°C. NaH(60%, 9.6mg, 2.40×10) -4 Add mol (3.4 equiv.), stir the reaction mixture under N2(g) at 0°C for 18 minutes, then add MeOTs (26.5 mg, 1.42 × 10⁻⁶). -4 (mol, 2 equiv.) was added. The reaction mixture was heated to ambient temperature and stirred for 2 hours, then H2O (1 drop) was added. The reaction mixture was combined with alkyl (15 mL), hexane (5 mL), and H2O (10 mL), the layers were mixed and separated, the organic layer was washed with H2O (10 mL × 2) and brine (10 mL), and dried on MgSO4. The solvent was removed to obtain an off-white solid, which was dissolved in CH2Cl2, poured onto dry silica (33 mm h × 33 mm id), and eluted under vacuum. Compound 103 (19 mg, 86%) was obtained as an off-white solid from %MeOH / CH2Cl2-1% (100 mL), 2% (50 mL), and 3% (50 mL). 1H NMR (400MHz, CDCl3) δ8.46(s, 2H), 7.52(dd, 1H, J=8.4Hz, J=5.4Hz), 7.02(dd, 1H, J=10.0Hz, J= 2.0Hz), 6.91(td, 1H, J=9.0Hz, J=2.0Hz), 6.49(s, 1H), 3.88(m, 4H), 3.81(m, 4H), 3.67(s, 3H). HRMS(ESI)[M+H] + In C 17 H 18 FN4O calculated value: 313.1459, measured value: 313.1455.

[0209] tert-butyl 6-fluoro-1H-indole-1-carboxylate (compound 104) 6-Fluorindole (3.28 g, 24.27 mmol), Boc2O (6.59 g, 30.19 mmol, 1.2 equiv.), i-Pr2NEt (7 mL, 40.2 mmol, 1.7 equiv.), DMAP (0.31 g, 2.54 mmol, 0.1 equiv.), and CH2Cl2 (210 mL) were stirred under N2 (g) at ambient temperature for 16 hours. The reaction mixture was concentrated to obtain a yellow / orange oil, hexane was added, and the mixture was purified by vacuum flash chromatography on silica (16 cm h × 4 cm id) with hexane (100 mL), %CH2Cl2 / hexane-10% (200 mL), and 25% (400 mL) to obtain compound 104 (5.67 g, 99%) as a colorless oil. 1 H NMR(300MHz, CDCl3)δ7.86(br d, 1H, J=10.2Hz), 7.56(d, 1H, J=3.6Hz), 7.46(dd, 1H, J=8.4Hz, J=5.4Hz), 6.98(td, 1H, J=9.0Hz, J=2.4Hz), 6.53(d, 1H, J=3.6Hz), 1.67(s, 9H).

[0210] Example 3. In vitro binding assay

[0211] All screened compounds were measured by analytical HPLC and showed a purity of over 95%. Binding assays were performed according to the method described in a previous publication (Graham, TJA, et al, Journal of Medicinal Chemistry 2023, 66, 10628-10638), and the results are shown in Table 2. [Table 2]

[0212] Example 4. In vitro competition (K i ) Assay.

[0213] Equilibrium inhibition constant (K) of the compound iThe values ​​were measured against tritium-labeled radioactive ligands using a publicly available method (Klunk et al., “The binding of 2-(4'-methylaminophenyl)benzothiazole to postmortem brain homogenates is dominant by the amyloid component.”. J Neurosci 2003, 23, 2086-2092). This assay used postmortem human brain tissue from AD, PSP, and CBD patients obtained from the UCSF Neurodegenerative Disease Brain Bank, and included only 3R / 4R-tau and amyloid-beta aggregates (AD tissue) or 4R-tau aggregates (PSP and CBD tissue), which were frequently identified at autopsy, and did not include other detectable aggregated amyloid species. Briefly, the tissue was homogenized on ice using a glass homogenizer in 300 mg / mL ice-cold pH 7.0 phosphate-buffered saline (PBS), then diluted 30-fold with PBS to 10 mg / mL, and a second homogenization was performed using a Brinkmann Polytron homogenizer before storage at -80°C. The frozen brain tissue was thawed and diluted 10-fold with PBS to 1 mg / mL. The concentration of the unlabeled competing compound (~400 μM in stock solution) was measured by quantitative NMR in DMSO (0.25% DMSO in the final assay vial). An appropriate concentration of the unlabeled competing compound (range 0.1–1000 nM) in 400 μL of PBS buffer was mixed with tritium-labeled radioligand (final concentration of radioligand ~1 nM) in 500 μL of PBS. The assay was initiated by adding 100 μL of 1 mg / mL brain tissue homogenate to achieve a final concentration of 100 μg tissue / mL. After incubation at room temperature for 60 minutes, the conjugation mixture was filtered through a Brandel M-24R cell harvester (Gaithersburg, MD, USA) using a Whatman GF / B glass filter and rapidly washed four times with 3 mL of PBS buffer. After thoroughly vortexing the filter, the counts were measured using Cytoscint-ES with a liquid scintillation counter.Complete (100%) inhibition of specific binding was defined as the number of counts substituted by 1 μM of unlabeled competing substance. All assays were performed in triplicate at each concentration. K. i The value represents 50% inhibition of radioactive ligand binding (IC). 50 This was measured by the concentration of the inhibitor that produced the effect. i =IC 50 / 2.

[0214] Example 5. Binding affinity (K d ) Assay.

[0215] The tritium-labeled radioactive ligand binding assay uses an equilibrium dissociation constant (K). dTo determine the values, AD, PSP, or CBD brain homogenates were used, following a slightly modified version of a previously described procedure (Klunk et al., “Uncharged thioflavin-T derivatives bind to amyloid-beta protein with high affinity and readily enter the brain.” Life Sciences 2001, 69(13), 1471-1484). Briefly, homogenized cortical cells derived from AD, PSP, or CBD brain (10 mg / mL in PBS (pH=7.0)) were thawed from frozen dispensing samples (-80°C) and diluted 10-fold with PBS to 1 mg / mL. The unlabeled test substance was dissolved in DMSO at 400 μM and diluted with PBS at 20 μM to obtain 5% DMSO / PBS. The remaining serial dilutions (typically 6 μM to 4 nM) were performed using 5% DMSO / PBS to maintain a constant DMSO concentration in the final assay. 50 μL of these solutions were combined with 50 μL of tritium-labeled test compound and 800 μL of PBS to obtain a final assay concentration of 0.25% DMSO, ~1 nM of tritium-labeled compound, and 0.2~1000 nM of unlabeled compound. The assay was initiated by adding 100 μL of 1 mg / mL of brain homogenate to achieve a final concentration of 100 μg tissue / mL. After incubation at room temperature for 60 minutes, the conjugation mixture was filtered through a Brandel M-24R cell harvester (Gaithersburg, MD) using a Whatman GF / B glass filter and rapidly washed three times with 3 mL of PBS. The filters were thoroughly vortexed and allowed to stand overnight before being counted with Cytoscint-ES. All assays were performed in at least three consecutive sets. The concentration of the bound compound was determined from the radioactivity retained on the filter, after correcting for the unsubstituted radioactivity (defined as the radioactivity remaining in the ~1 μM unlabeled compound) and the specific activity of the tritium-labeled compound after dilution with various concentrations of unlabeled compounds. d The value represents the slope of the Scatchard plot of the bound radioactive ligand value relative to binding / release at different ligand concentrations (slope = -1 / K). d It was decided by [the organization / organization].

[0216] Example 6. X-ray crystal structure data [Table 3] [Table 4] [Table 5] [Table 6]

[0217] Example 7. In a tauopathy tissue sample [ 3 H]1([ 3 H]Z-3540) and [ 3 H]2([ 3 Radiosynthesis and in vitro evaluation of H]Z-2340.

[0218] Z-3540 (Enamine Z3777013540, Compound 1) and Z-2340 (Enamine Z4169252340, Compound 2) were identified by molecular similarity fingerprint search of an enamine compound library based on CBD-2115. 3 H]Z-3540 and [ 3 [H]Z-2340 was synthesized and evaluated by binding assays in human CBD, PSP, and AD tissue.

[0219] In human tissue, [ 3 H]Z-3540's K d teeth,[ 3 Compared to H]CBD-2115, AD was 3 times lower and CBD was 7 times lower. 3 [H]Z-3540 was found to selectively bind to aggregated tau more than β-amyloid and α-synuclein.

[0220] Example 8. Radiochemistry

[0221] The X-ray crystal structures of radiolabeling precursors 65 and 66 are shown in Figures 2 and 3, respectively. 18 F]68 and [ 18 The radioactive labeling of F]69 was performed manually in a lead-shielded hot cell using a manipulator arm. 18 [F]Fluoride is trapped in Waters Sep-Pak Plus Light QMA, and then eluted with Et4NHCO3(aq) to obtain Et4N 18 F was obtained and azeotropically dried with CH3CN. Nitro precursor 65 or 66 was dissolved in DMSO and dried in a sealed 5 mL V vial. 18 It was added to F and heated at ~140°C for 30 minutes, and then the crude reaction mixture was purified by semi-preparative HPLC. Radioactive tracer [ 18 F]68 was obtained in 10% EtOH / 0.9% sterile saline at pH 5 with an uncorrected radiochemical yield of 4.4% (1.18 GBq (31.9 mCi)), radiochemical purity of 99.4%, molar activity Am = 0.13 GBq / nmol (3.5 mCi / nmol). Radioactive tracer [ 18 F]69 was obtained in a 10% EtOH / 0.9% sterile physiological saline solution at pH 5 with an uncorrected radiochemical yield of 11.5% (3.49 GBq (94.3 mCi)), radiochemical purity of 98.9%, and molar activity Am = 0.05 GBq / nmol (1.4 mCi / nmol), with logD = 1.96 ± 0.03 (n=4). The radiochemical identity of the radioactive tracer was confirmed by co-injection with an unlabeled standard substance by analytical HPLC.

[0222] [ka]

[0223] 2-(2-[ 18 F]Fluoro-6-(piperidine-1-yl)pyridine-3-yl)-1H-indole([ 18 F]68) Et4NHCO3 (70 mg) was dissolved in Milli-Q H2O (10 mL) to obtain a 7 mg / mL solution. Waters Sep-Pak Plus Light QMA (WAT023525) was washed with Et4NHCO3 (aq) solution (8 mL, 7 mg / mL), then with Milli-Q H2O (10 mL), and then with air (25 mL). Waters C 18 Sep-Pak (WAT020515) was washed with EtOH (10 mL) and then with Milli-Q H2O (10 mL). Carrier-free [ 18 F]F - teeth, 18 O(p,n) 18 It is produced by the F reaction (Siemens Eclipse HP cyclotron), [ 18 It was transferred to a lead-shielded hot cell in [O]H2O. 18 F]F - (26.6 GBq (719 mCi)) was trapped in the QMA, eluted with Et4NHCO3 (aq) solution (1 mL, 7 mg / mL), then eluted with CH3CN (1 mL) (0.45 GBq (12.2 mCi) remained in the QMA), and collected in a 5 mL V-vial. The solvent was evaporated under argon flow at 110°C, and then Et4N 18 F was paroxysmal added to CH3CN (1 mL x 3) and azeotropically dried under argon at 110°C. Compound 65 (3.8 mg) was dissolved in DMSO (0.6 mL) and dried Et4N. 18 Add to F, cap the V-vial, mix gently for about 10 seconds, and after the solution turned pale pink, heat at ~140°C for 30 minutes. Cool the V-vial for ~1 minute, dilute the solution with HPLC solvent (0.5 mL, 55:45 v / v CH3CN / 0.1M NH4HCO2 pH 4.2), load into the HPLC loop, wash the V-vial with HPLC solvent (0.5 mL), load the solvent into the HPLC loop, and purify the sample by prep-HPLC (Phenomenex Gemini, 250 × 10 mm + guard, 5 mL / min for 6 minutes, then 10 mL / min). Collect the HPLC radioactive peak at ~24-25 minutes, dilute with Milli-Q H2O (50 mL), and C 18The Sep-Pak was passed through and washed with Milli-Q H2O (10 mL). The radioactive tracer was eluted from the Sep-Pak with EtOH (1 mL) and then 0.9% sterile saline (9 mL), passed through a 0.2 micrometer filter, and collected in a sterile vial, 18 F]68 (1.18 GBq (31.9 mCi), 4.4% radiochemical yield (uncorrected for decay), 99.4% radiochemical purity (Phenomenex Gemini, 5 μm, NX-C18 110 Å, 100×4.6 mm, 60:40 v / v CH3CN / 0.1 M NH4HCO2 pH 4.2, 1 mL / min, 254 nm), Am = 0.13 GBq / nmol (3.5 mCi / nmol), pH 5) was obtained. Radiochemical identity was confirmed by co-injection with 68. The endotoxin level of the final formulation was <2.00 EU / mL (Charles Rivers Endosafe®).

[0224] 4-(6- 18 F]fluoro-5-(1H-indol-))pyridin-2-yl)morpholine( 18 F]69)( 18 F]JSS20-183A) Et4NHCO(registered trademark)(70 mg) was dissolved in Milli-Q H2O (10 mL) to obtain a 7 mg / mL solution. Waters Sep-Pak Plus Light QMA (WAT023525) was washed with Et4NHCO3(aq) solution (8 mL, 7 mg / mL), Milli-Q H2O (10 mL), and then air (25 mL). Waters C 18 Sep-Pak (WAT020515) was washed with EtOH (10 mL) and then with Milli-Q H2O (10 mL). Carrier-free 18 F]F - was 18 O(p,n) 18 F reaction (Siemens Eclipse HP cyclotron) was generated, 18 O]H2O was transferred to a lead-shielded hot cell. 18 F]F -(30.3 GBq (818 mCi)) was trapped on the QMA and eluted with Et4NHCO3 (aq) solution (1 mL, 7 mg / mL), then CH3CN (1 mL), then air (1 mL) (0.3 GBq (8.0 mCi) remained on the QMA), and collected in a 5 mL V-vial. The solvent was evaporated and removed under an argon stream at 110°C, then Et4N 18 F was paroxysmal added to CH3CN (1 mL x 3) and azeotropically dried at 110°C under an argon stream. Compound 66 (2.5 mg) was dissolved in DMSO (0.5 mL) and dried Et4N. 18 Adding F (the solution turned pale purple), capped the V-vial and heated at ~140°C for 30 minutes. Cool the V-vial for ~1 minute, dilute the solution with HPLC solvent (0.5 mL, 45:55 v / v CH3CN / 0.1M NH4HCO2 pH 4.2), load into the HPLC loop, wash the V-vial with HPLC solvent (0.5 mL), load the solvent into the HPLC loop, and purify the sample by prep-HPLC (Phenomenex Gemini, 250 × 10 mm + guard, 5 mL / min for 6 minutes, then 8 mL / min). Collect the HPLC radioactive peak at ~20.4-22 minutes, dilute with Milli-Q H2O (50 mL), and C 18 The radioactive tracer was passed through Sep-Pak and washed with Milli-Q H2O (10 mL). The radioactive tracer was eluted from Sep-Pak with EtOH (1 mL), then with 0.9% sterile physiological saline (9 mL), passed through a 0.2 micrometer filter, and collected in a sterile vial. [ 18 F]69 was obtained with a radiochemical yield of 3.49 GBq (94.3 mCi), a radiochemical yield of 11.5% (uncorrected for attenuation), and a radiochemical purity of 98.9% (Phenomenex Gemini, 5 μm, NX-C18 110 Å, 100 × 4.6 mm, 45:55 v / v CH3CN / 0.1M NH4HCO2 pH 4.2, 1 mL / min, 254 nm), Am = 0.05 GBq / nmol (1.4 mCi / nmol), pH 5, logD = 1.96 ± 0.03 (n=4). Radiochemical identity was confirmed by co-injection with 69. The endotoxin level of the final formulation was <2.00 EU / mL (Charles Rivers Endosafe®).

[0225] Example 9. [ 3 H]69([ 3 Binding affinity of [H]JSS20-183A)

[0226] [ 3 H]69([ 3 The binding affinity of [H]JSS20-183A) was measured using postmortem brain tissue (Table 7). 3 H]69([ 3 [H]JSS20-183A) shows high affinity in postmortem human AD, PSP, CBD, and Pick's disease brain tissue, as well as in P301L mouse brain tissue. 3 H]69([ 3 [H]JSS20-183A) showed low affinity in postmortem brain tissue from Parkinson's disease (PD), indicating that 69 does not strongly bind to α-synuclein. In human AD postmortem brain tissue, compound 69 is [ 3 Since it does not compete with [H]PiB, it was shown that 69 binds to the PiB binding site on amyloid-beta. [Table 7]

[0227] Example 10. Brain PET imaging test

[0228] [ 18 F]68 (Figure 5) and [ 18 The brain uptake and washout of F]69 (Figure 6) were evaluated using PET imaging studies of the brain in rhesus monkeys. 18 The ratios of F68 at 2 minutes and 60 minutes were 1.2 (whole brain), 1.3 (cerebral cortex), and 2.0 (cerebellum). 18 The ratios of F69 at 2 minutes and 60 minutes were 3.0 (whole brain), 3.7 (cerebral cortex), and 3.8 (cerebellum).

[0229] Example 11. Autoradiography

[0230] In postmortem human PSP, CBD, and AD brain tissue [3 Specific binding of [H]69 was demonstrated by autoradiography (Figure 7). Specific binding was barely observed in postmortem human brain control tissue.

[0231] While specific embodiments have been illustrated and described, it should be understood that modifications and alterations are possible in accordance with the ordinary art of those skilled in the art without departing from the broader embodiments defined in the following claims.

[0232] Embodiments described herein as exemplary may also be suitably implemented without any elements, limitations, or restrictions not specifically disclosed herein. Therefore, terms such as “comprising,” “including,” and “containing” should be read broadly and without limitation. Furthermore, the terms and expressions used herein are for illustrative purposes only and not intended to be limiting. The use of such terms and expressions is not intended to exclude equivalents of the exhibited and described features or any part thereof, but it should be recognized that various modifications are possible within the scope of the claimed technology. Additionally, the expression “consisting essentially of” should be understood to include the specifically described elements and any additional elements that do not substantially affect the fundamental and novel characteristics of the claimed technology. The expression “consisting of” excludes elements that are not specified.

[0233] This disclosure is not limited in terms of the specific embodiments described in this application. It will be apparent to those skilled in the art that many modifications and variations can be made without departing from the spirit and scope of this disclosure. In addition to those enumerated herein, functionally equivalent methods and compositions within the scope of this disclosure will be apparent to those skilled in the art from the foregoing description. Such changes and variations are intended to be included in the appended claims. This disclosure is limited only by the wording of the appended claims and the entire scope of the equivalents to which the claims are entitled. It should be understood that this disclosure is not limited to specific methods, reagents, compounds, or compositions, and that these may naturally change. It should also be understood that the terms used herein are for the purpose of describing specific embodiments only and are not intended to limit them.

[0234] In addition, if any feature or aspect of this disclosure is described from the perspective of the Markush group, a person skilled in the art will recognize that this disclosure is also described from the perspective of any individual member or subgroup of a member of the Markush group.

[0235] As will be understood by those skilled in the art, for all purposes, and particularly in terms of clarity of description, all scopes disclosed herein also encompass any possible sub-scopes and combinations thereof. Any enumerated scope can be readily recognized as fully explaining and enabling that the same scope can be decomposed into at least equal halves, thirds, quarters, fifths, tenths, etc. As a non-limiting example, each scope discussed herein can readily be decomposed into a lower third, a middle third, an upper third, etc. Also, as will be understood by those skilled in the art, all expressions such as “up to,” “at least,” “greater than,” and “less than” refer to a scope that includes the number mentioned and can then be decomposed into sub-scopes as described above. Finally, as will be understood by those skilled in the art, a scope includes its individual members.

[0236] All publications, patent applications, issued patents, and other documents referenced herein are incorporated by reference in the same manner as if each individual publication, patent application, issued patent, or other document were specifically and individually indicated to be incorporated by reference in whole. Definitions contained in the text incorporated by reference are excluded to the extent that they conflict with the definitions in this disclosure.

[0237] Other embodiments are described in the following claims.

Claims

1. Compounds having the structure of formula I or formula II, or pharmaceutically acceptable salts, solvates, hydrates, tautomers, or stereoisomers thereof. 【Chemistry 1】 【Chemistry 2】 Here, X １ is N, O, or S, X ２ , X ３ , X ４ , X ５ Each of these is independently either N or CH, X ６ is CH or O, R １ is H, halogen, CN, Me ３ N ＋ X ７ － , Et ３ N ＋ X ７ － , Pr ３ N ＋ X ７ － , Bu ３ s N ＋ X ７ － , or other suitable ammonium salts, OH, OMs, OTs, OTf, O - nosylate, O - brosylate, or other suitable sulfonate species, SnMe ３ , SnEt ３ , SnPr ３ , SnBu ３ , or other suitable trialkyltin species, I(O - acyl) ２ , ArI ＋ X ７ － , IR ７ , S(O)Ar, SO ２ Ar , BF ３ K , B(OH) ２ , B(OMe) ２ , B(OEt) ２ , B(Opr) ２ , B(O iPr) ２ [ ２ , B(O iPr) ３ Li, B - pinacol, B - neopentyl glycol, or other suitable boron species, provided that when R １ is OH, R １ is not substituted at the 5 - position of the indole ring, Here, X ７ － is the appropriate counterion, and here, ArI ＋ X ７ － is a suitable iodonium species known in the field, where IR ７ This is a suitable iodonium ylide known in the field, R ２ is H, alkyl, tert-butoxycarbonyl, tosyl, benzyl, p-methoxybenzyl, CBz, Fmoc, or other suitable nitrogen protecting group. R ３ H, halogen, NO ２ Me ３ N ＋ X ７ － , Et ３ N ＋ X ７ － , Pr ３ N ＋ X ７ － , Bu ３ N ＋ X ７ － or other suitable ammonium salts, OH, OMs, OTs, OTf, O-nosilates, O-brosilates, or other suitable sulfonate species, SnMe ３ SnEt ３ SnPr ３ SnBu ３ or other suitable trialkyltin species, I(O-acyl) ２ , ArI ＋ X ７ － IR ７ ,S(O)Ar,SO ２ Ar, BF ３ K, B (OH) ２ , B(OMe) ２ , B(OEt) ２ B (Opr) ２ , B (OiPr) ２ , B (OiPr) ３ Li, β-pinacol, β-neopentyl glycol, or other suitable boron species, Here, X ７ － is a suitable counterion, where ArI ＋ X ７ － is a suitable iodonium species known in the art, where IR ７ is a suitable iodonium iridium known in the art, R ４ is H, halogen, OH, alkoxy, carbonyl, or OEtOEtF, R ５ is H, halogen, OH, OMs, OTs, OTf, O-nosilate, O-brosilate, or other suitable sulfonate species. R ６ These are H, halogen, OH, O-acyl, O-alkyl, OBoc, or other suitable oxygen-protecting species. or R ５ and R ６ is epoxide: 【Transformation 3】 Forming, m and n are independently either 0 or 1, and o is 0, 1, or 2.

2. The compound according to claim 1, having the following structure. 【Chemistry 4-1】 【Chemistry 4-2】

3. The compound according to claim 1, having the following structure. 【Transformation 5】

4. The compound according to any one of the preceding claims, wherein the compound selectively binds to 4R tau.

5. The compound according to any one of the preceding claims, wherein the compound comprises one or more detection labels.

6. The compound according to claim 5, wherein the detection label is selected from the group consisting of radionuclides, positron-emitting nuclides, gamma-ray-emitting nuclides, and fluorescent labels.

7. where the detection label is ２ H, ３ H, １８ F, １２３ I, １２４ I, １２５ I, １３１ I, １１ C, １３ C, １４ C, １３ N, １５ N, １５ O, ７５ Br, ７６ Br and ７７ the compound according to claim 5 or claim 6, selected from the group consisting of Br.

8. The aforementioned detection indicator １８ The compound according to any one of claims 5 to 7, wherein F.

9. A compound according to any one of the claims, for use in the detection of disorders or diseases associated with tau aggregates.

10. The compound for use according to claim 9, wherein the disease or disorder is a disease or disorder associated with 4R tauopathy or aggregates of 4R tau.

11. The compound for use according to claim 9 or 10, wherein the disease or disorder is a neurodegenerative disease or disorder.

12. The compound for use according to claim 8 or 10, wherein the disorder or disease is selected from the group consisting of Alzheimer's disease, progressive supranuclear palsy, Pick's disease, frontotemporal dementia (FTLDs), frontotemporal dementia with parkinsonism-17 (FTDP-17), primary age-related tauopathy (PART) dementia, argyurophilic grain disease, glial globular tauopathy, chronic traumatic encephalopathy (CTE), corticobasal syndrome (CBS), and corticobasal degeneration.

13. The compound for use according to any one of claims 9 to 12, wherein the detection is performed by one or more of positron emission tomography (PET), single-photon emission computed tomography (SPECT), magnetic resonance imaging, or autoradiography.

14. The compound for use according to claim 13, wherein the detection is performed by positron emission tomography (PET).

15. A method for diagnosing a disease or disorder in a subject, or a method for monitoring the progression of a disease or disorder in a subject, the method includes the following: (a) A step of administering to a subject a detectable amount of the isotope-labeled compound of formula I or formula II, or a pharmaceutically acceptable salt, solvate, hydrate, tautomer or stereoisomer thereof: 【Transformation 6】 【Transformation 7】 Here, X １ is N, O, or S, X ２ , X ３ , X ４ , X ５ Each of these is independently either N or CH, X ６ is CH or O, R １ H, halogen, CN, Me ３ N ＋ X ７ － , Et ３ N ＋ X ７ － , Pr ３ N ＋ X ７ － , Bu ３ N ＋ X ７ － or other suitable ammonium salts, OH, OMs, OTs, OTf, O-nosilates, O-brosilates, or other suitable sulfonate species, SnMe ３ SnEt ３ SnPr ３ SnBu ３ or other suitable trialkyltin species, I(O-acyl) ２ , ArI ＋ X ７ － IR ７ ,S(O)Ar,SO ２ Ar, BF ３ K, B (OH) ２ , B(OMe) ２ , B(OEt) ２ B (Opr) ２ , B (OiPr) ２ , B (OiPr) ３ Li, β-pinacol, β-neopentyl glycol, or other suitable boron species, however R １ If is OH, then R １ It is a substance that is not substituted at the 5th position of the indole ring. Here, X ７ － is the appropriate counterion, and here, ArI ＋ X ７ － is a suitable iodonium species known in the field, where IR ７ This is a suitable iodonium ylide known in the field, R ２ is H, alkyl, tert-butoxycarbonyl, tosyl, or other suitable nitrogen protecting group. R ３ H, halogen, NO ２ Me ３ N ＋ X ７ － , Et ３ N ＋ X ７ － , Pr ３ N ＋ X ７ － , Bu ３ N ＋ X ７ － or other suitable ammonium salts, OH, OMs, OTs, OTf, O-nosilates, O-brosilates, or other suitable sulfonate species, SnMe ３ SnEt ３ SnPr ３ SnBu ３ or other suitable trialkyltin species, I(O-acyl) ２ , ArI ＋ X ７ － IR ７ ,S(O)Ar,SO ２ Ar, BF ３ K, B (OH) ２ , B(OMe) ２ , B(OEt) ２ B (Opr) ２ , B (OiPr) ２ , B (OiPr) ３ Li, β-pinacol, β-neopentyl glycol, or other suitable boron species, Here, X ７ － is the appropriate counterion, and here, ArI ＋ X ７ － is a suitable iodonium species known in the field, where IR ７ This is a suitable iodonium ylide known in the field, R ４ is H, halogen, OH, alkoxy, carbonyl, or OEtOEtF, R ５ is H, halogen, OH, OMs, OTs, OTf, O-nosilate, O-brosilate, or other suitable sulfonate species. R ６ These are H, halogen, OH, O-acyl, O-alkyl, OBoc, or other suitable oxygen-protecting species. or R ５ and R ６ is epoxide: 【Transformation 8】 Forming, m and n are independently either 0 or 1, and o is 0, 1, or 2. And, (b) A step to detect the binding of a compound to tau aggregates in a patient.

16. The method according to claim 15, wherein the disease or disorder is related to tau aggregates.

17. The method according to claim 16, wherein the disease or disorder is a disease or disorder associated with 4R tauopathy or 4R tau aggregates.

18. The method according to claim 16 or 17, wherein the disease or disorder is a neurodegenerative disease or disorder.

19. The method according to any one of claims 15 to 18, wherein the disease or disorder is selected from the group consisting of Alzheimer's disease, progressive supranuclear palsy, Pick's disease, frontotemporal dementia (FTLDs), frontotemporal dementia with parkinsonism-17 (FTDP-17), primary age-related tauopathy (PART) dementia, argyurophilic grain disease, glial globular tauopathy, chronic traumatic encephalopathy (CTE), corticobasal syndrome (CBS), and corticobasal degeneration.

20. The aforementioned compound, ２ H, ３ H, １８ F, １２３ I, １２４ I, １２５ I, １３１ I, １１ C, １３ C, １４ C, １３ N, １５ N, １５ O, ７５ Br, ７６ Br and ７７ The method according to any one of claims 15 to 19, wherein the material is isotope-labeled with Br.

21. The method according to any one of claims 15 to 20, wherein the detection is performed by one or more of positron emission tomography (PET), single-photon emission computed tomography (SPECT), magnetic resonance imaging, or autoradiography.

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