Nucleoside LINE-1 inhibitors

Abstract

The present disclosure provides a LINE-1 inhibitor and a method of treating or preventing a disease, disorder, or condition in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of a LINE-1 inhibitor.

Description

Technical Field

[0001] The present disclosure provides LINE-1 inhibitors and methods of treating or preventing a disease, disorder or condition caused by a LINE-1 retrotransposon in a subject by administering a LINE-1 inhibitor or a pharmaceutical composition thereof or a tautomer thereof to the subject. Background Art

[0002] Long interspersed element-1 (LINE-1 or L1) retrotransposons form the only autonomously active transposon family in humans. They are expressed and moved in the germline, in embryonic stem cells, and in early embryos, but are silent in most somatic tissues. LINE-1 plays an important role in individual genome variation through insertion mutations and sequence transduction, which occasionally causes genetic diseases and disorders. For example, in many neural disorders, retrotransposon activity increases. For example, see Terry and Devine, Front. Genet., January 08, 2020, https: / / doi.org / 10.3389 / fgene.2019.01244 and WO 2020 / 154656.

[0003] LINE-1 elements encode two proteins, ORF1p and ORF2p, which are essential for their mobility. ORF1p is an RNA-binding protein with nucleic acid chaperone activity. ORF2p has endonuclease and reverse transcriptase activities. These proteins and LINE-1 RNA assemble into ribonucleoprotein particles (LINE-1 RNPs) - the core of the retrotransposition mechanism. LINE-1 RNPs mediate the synthesis of new LINE-1 copies after target DNA cleavage and the reverse transcription of LINE-1 RNA at the target site. LINE-1 elements utilize cellular host factors to complete their life cycle, however, several cellular pathways also limit the cellular accumulation of LINE-1 RNPs and their harmful activities. For example, see Pizarro and Cristofari (2016) Front. Cell Dev. Biol. 4: 14. doi: 10.3389 / fcell.2016.00014. There is a need in the art for LINE-1 inhibitors to treat or prevent diseases, disorders or conditions caused by LINE-1 retrotransposition in subjects. Summary of the invention

[0004] In one aspect, the present disclosure provides compounds of Table 1 (see below), and pharmaceutically acceptable salts or solvates thereof, and / or tautomers thereof; compounds of Table 2 (see below), and pharmaceutically acceptable salts or solvates thereof, and / or tautomers thereof; and compounds of Table 3 (see below), and pharmaceutically acceptable salts or solvates thereof, and / or tautomers thereof. The compounds in Tables 1-3, and pharmaceutically acceptable salts or solvates thereof, and / or tautomers thereof, are collectively referred to as "compounds of the present disclosure" or individually as "compounds of the present disclosure".

[0005] In another aspect, the disclosure provides a method for treating or preventing a disease, disorder or condition in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of a compound of the disclosure. Exemplary diseases, disorders or conditions include, but are not limited to, neurodegenerative diseases, autoimmune diseases, and age-related diseases.

[0006] In another aspect, the disclosure provides a method of treating or preventing a disease, disorder or condition caused by a pathophysiological retrotransposon-associated process in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of a compound of the disclosure.

[0007] In another aspect, the present disclosure provides a method of treating or preventing symptoms of a disease, disorder or condition caused by a pathophysiological retrotransposon-associated process in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of a compound of the present disclosure or a pharmaceutical composition thereof.

[0008] In another aspect, the present disclosure provides a method of inhibiting a LINE-1 retrotransposition event, such as a somatic LINE-1 insertion, that causes a disease, disorder or condition in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of a compound of the present disclosure, or a pharmaceutical composition thereof.

[0009] In another aspect, the present disclosure provides a method for treating a disease, disorder or condition in a subject in need thereof, the method comprising (a) determining the presence or absence of a biomarker, such as retrotransposon RNA, retrotransposon reverse transcriptase or retrotransposon DNA, such as LINE-1 RNA, overexpression of ORF1p or ORF2p, in a biological sample taken from the subject; and (b) if overexpression of the biomarker is present in the biological sample, administering to the subject a therapeutically effective amount of a compound of the present disclosure or a pharmaceutical composition thereof.

[0010] In another aspect, the disclosure provides a method for identifying whether a subject having a disease, condition or disorder is a candidate for treatment with a compound of the disclosure or a pharmaceutical composition thereof, the method comprising (a) determining the presence or absence of a biomarker, such as overexpression of a retrotransposon RNA, a retrotransposon reverse transcriptase, or a retrotransposon DNA, such as LINE-1 RNA, ORF1p, or ORF2p, in a biological sample taken from the subject; (b) if overexpression of the biomarker is present, identifying the subject as a candidate for treatment; or (c) if overexpression of the biomarker is not present, identifying the subject as not a candidate for treatment.

[0011] In another aspect, the disclosure provides a method for predicting a treatment outcome in a subject suffering from a disease, condition or disorder, the method comprising determining the presence or absence of a biomarker, such as retrotransposon RNA, retrotransposon reverse transcriptase or retrotransposon DNA, such as overexpression of LINE-1 RNA ORF1p or ORF2p, in a biological sample taken from the subject, wherein (a) the presence of overexpression of the biomarker in the biological sample indicates that administration of a compound of the disclosure or a pharmaceutical composition thereof to the subject will likely result in a favorable treatment response; and (b) the absence of overexpression of the biomarker in the biological sample indicates that administration of a compound of the disclosure or a pharmaceutical composition thereof to the subject will likely result in an unfavorable treatment response.

[0012] In another aspect, the present disclosure provides methods comprising administering a therapeutically effective amount of a compound of the present disclosure or a pharmaceutical composition thereof to a subject in need thereof, wherein: (a) the subject suffers from a disease, disorder or condition; and (b) the disease, disorder or condition is characterized by overexpression of a biomarker, such as retrotransposon RNA, retrotransposon reverse transcriptase, or retrotransposon DNA, such as LINE-1 RNA, ORF1p, or ORF2p).

[0013] In another aspect, the disclosure provides a kit comprising a compound of the disclosure or a pharmaceutical composition thereof and instructions for administering the compound or composition to a subject suffering from a disease, condition, or disorder caused by a pathophysiological retrotransposon-associated process.

[0014] In another aspect, the present disclosure provides a compound of the present disclosure or a pharmaceutical composition thereof for use in treating or preventing a disease, disorder or condition.

[0015] In another aspect, the present disclosure provides a compound of the present disclosure or a pharmaceutical composition thereof for use in treating or preventing a disease, disorder or condition caused by a pathophysiological retrotransposon-related process.

[0016] In another aspect, the disclosure provides a compound of the disclosure or a pharmaceutical composition thereof for use in treating or preventing a symptom of a disease, disorder or condition caused by a pathophysiological retrotransposon-associated process.

[0017] In another aspect, the present disclosure provides the use of a compound of the present disclosure or a pharmaceutical composition thereof for the preparation of a medicament for treating or preventing a disease, disorder or condition.

[0018] In another aspect, the present disclosure provides the use of a compound of the present disclosure or a pharmaceutical composition thereof for the preparation of a medicament for treating or preventing a disease, disorder or condition caused by a pathophysiological retrotransposon-related process.

[0019] In another aspect, the present disclosure provides the use of a compound of the present disclosure or a pharmaceutical composition thereof for the preparation of a medicament for treating or preventing a symptom of a disease, disorder or condition.

[0020] In another aspect, the disclosure provides the use of a compound of the disclosure or a pharmaceutical composition thereof for the preparation of a medicament for treating or preventing a symptom of a disease, disorder or condition caused by a pathophysiological retrotransposon-related process. DETAILED DESCRIPTION

[0021] I. Compounds of the Disclosure

[0022] In one embodiment, the compound of the present disclosure is a compound of Table 1, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0023] Table 1

[0024]

[0025]

[0026]

[0027]

[0028]

[0029]

[0030] In another embodiment, the compound of the present disclosure is a compound of Table 2, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0031] Table 2

[0032]

[0033]

[0034]

[0035]

[0036]

[0037]

[0038]

[0039]

[0040]

[0041]

[0042]

[0043]

[0044]

[0045]

[0046]

[0047]

[0048]

[0049]

[0050]

[0051]

[0052]

[0053]

[0054]

[0055]

[0056]

[0057] In one embodiment, the compound of the present disclosure is a compound of Table 3, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0058] Table 3

[0059]

[0060] Table 1-3 provides the chemical structures and Relevant chemical names generated by Professional Edition 20.1.1.125. If there is any ambiguity between their chemical structures and chemical names, the compounds of the present disclosure are defined by their structures.

[0061] In another embodiment, the present disclosure provides a pharmaceutical composition comprising a compound of the present disclosure, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof, and one or more pharmaceutically acceptable excipients.

[0062] II. Treatment Methods and Uses

[0063] The compounds of the present disclosure inhibit LINE-1 retrotransposition. Thus, these compounds can be used to treat diseases, disorders or conditions in which, for example, LINE-1 retrotransposition plays a pathogenic role.

[0064] The compounds of the present disclosure or pharmaceutical compositions thereof can be administered to a subject in need thereof, e.g., a subject already suffering from a disease, condition, or disorder; a subject suspected of suffering from a disease, condition, or disorder; or a subject at risk of suffering from a disease, condition, or disorder. When a compound of the present disclosure is administered to a subject at risk of suffering from a disease, condition, or disorder, the purpose is to attempt to avoid the disease, condition, or disorder in the subject, e.g., by preventing or reducing the LINE-1 retrotransposition activity that causes the disease, condition, or disorder.

[0065] In one embodiment, the present disclosure provides a method of treating or preventing a disease, disorder or condition in a subject in need thereof, and / or a method of treating or preventing a symptom of a disease, disorder or condition, comprising administering to the subject a therapeutically effective amount of a Compound of the Disclosure or a pharmaceutical composition thereof.

[0066] In another embodiment, the present disclosure provides a method of treating a disease, disorder or condition in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of a Compound of the Disclosure or a pharmaceutical composition thereof.

[0067] In another embodiment, the present disclosure provides a method of preventing a disease, disorder or condition in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a compound of the present disclosure or a pharmaceutical composition thereof.

[0068] In another embodiment, the present disclosure provides a method of treating a symptom of a disease, disorder or condition in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a Compound of the Disclosure or a pharmaceutical composition thereof.

[0069] In another embodiment, the present disclosure provides a method of preventing symptoms of a disease, disorder or condition in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a Compound of the Disclosure or a pharmaceutical composition thereof.

[0070] In another embodiment, the present disclosure provides a Compound of the Disclosure, or a pharmaceutical composition thereof, for use in treating or preventing a disease, disorder or condition and / or treating or preventing a symptom of a disease, disorder or condition in a subject.

[0071] In another embodiment, the present disclosure provides a Compound of the Disclosure, or a pharmaceutical composition thereof, for use in treating a disease, disorder, or condition in a subject.

[0072] In another embodiment, the present disclosure provides a Compound of the Disclosure, or a pharmaceutical composition thereof, for use in preventing a disease, disorder, or condition in a subject.

[0073] In another embodiment, the present disclosure provides a Compound of the Disclosure, or a pharmaceutical composition thereof, for use in treating a symptom of a disease, disorder, or condition in a subject.

[0074] In another embodiment, the present disclosure provides a Compound of the Disclosure, or a pharmaceutical composition thereof, for use in preventing a symptom of a disease, disorder, or condition in a subject.

[0075] In another embodiment, the present disclosure provides the use of a Compound of the Disclosure or a pharmaceutical composition thereof in the preparation of a medicament for treating or preventing a disease, disorder or condition and / or treating or preventing a symptom of a disease, disorder or condition in a subject.

[0076] In another embodiment, the present disclosure provides the use of a Compound of the Disclosure, or a pharmaceutical composition thereof, in the preparation of a medicament for treating a disease, disorder or condition in a subject.

[0077] In another embodiment, the present disclosure provides use of a Compound of the Disclosure or a pharmaceutical composition thereof in the preparation of a medicament for preventing a disease, disorder or condition in a subject.

[0078] In another embodiment, the present disclosure provides the use of a Compound of the Disclosure, or a pharmaceutical composition thereof, in the preparation of a medicament for treating a symptom of a disease, disorder, or condition in a subject.

[0079] In another embodiment, the present disclosure provides the use of a Compound of the Disclosure, or a pharmaceutical composition thereof, in the preparation of a medicament for preventing a symptom of a disease, disorder, or condition in a subject.

[0080] In another embodiment, the subject is (a) not infected with HIV, (b) not suspected of being infected with HIV, (c) not receiving HIV treatment, and / or (d) not receiving HIV prophylactic treatment.

[0081] In another embodiment, the disclosure provides a method of inhibiting a disease, disorder or condition-causing LINE-1 retrotransposition event in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of a compound of the disclosure.

[0082] III. Diseases, Disorders and Conditions

[0083] Without wishing to be bound by any particular theory, the compounds of the present disclosure are useful for treating or preventing a disease, disorder, or condition in a subject because they inhibit LINE-1 retrotransposition.

[0084] In one embodiment, as described in Example 35, the compounds of the present disclosure inhibited the expression of luciferase in an in vitro HeLa cell-based dual-luciferase assay at a half-maximal inhibitory concentration (IC 50 ) inhibits human LINE-1 retrotransposition activity, see below. See also Jones et al., (2008) PLoS ONE 3(2):e1547.doi:10.1371 / journal.pone.0001547; Xie et al., (2011) Nucleic Acids Res.39(3):e16.doi:10.1093 / nar / gkq1076; Kopera et al., Methods Mol Biol 1400:139-156 (2016). In another embodiment, IC 50 In another embodiment, IC 50 In another embodiment, IC 50In another embodiment, IC 50 In another embodiment, IC 50 In another embodiment, IC 50 In another embodiment, IC 50 It is less than 0.005μM.

[0085] In one embodiment, the disease, disorder or condition and / or symptom(s) thereof is caused by a pathophysiological retrotransposon-associated process, wherein the disease, disorder or condition is not cancer and / or is not an infectious disease.

[0086] In another embodiment, the disease, disorder or condition is a neurodegenerative disease. For example, see Dugger and Dickson, Cold Spring Harb Perspect Biol 2016; 9: a028035. Exemplary neurodegenerative diseases include, but are not limited to, Alzheimer's disease, amyotrophic lateral sclerosis (ALS), Parkinson's disease, dementia with Lewy bodies (DLB), multiple system atrophy (MSA), Huntington's disease, frontotemporal lobar degeneration (FTLD), mild cognitive impairment (MCI), corticobasal degeneration (CDB), progressive supranuclear palsy (PSP), Rett syndrome, peripheral degenerative disease or Aicardi Goutières syndrome (AGS). In another embodiment, frontotemporal lobar degeneration is frontotemporal lobar dementia. For example, see Mohandas and Rajmohan, Indian J Psychiatry 51 (Suppl 1): S65-S69 (2009).

[0087] In another embodiment, symptoms of a neurodegenerative disease include, but are not limited to, memory loss, forgetfulness, apathy, anxiety, agitation, loss of inhibition, or mood changes.

[0088] In another embodiment, the disease, disorder or condition is an autoimmune disease. For example, see Wang et al., J Intern Med 278:369-395 (2016). Exemplary autoimmune diseases include, but are not limited to, lupus, rheumatoid arthritis (RA), Sjögren's syndrome or multiple sclerosis (MS).

[0089] In another embodiment, symptoms of an autoimmune disease include, but are not limited to, fatigue, muscle aches, redness, swelling, low-grade fever, difficulty concentrating, numbness and tingling in the hands and feet, hair loss, or rash.

[0090] In another embodiment, the disease, disorder or condition is an age-related disease. For example, Franceschi et al., Front.Med.5:61.doi:10.3389 / fmed.2018.00061; De Cecco et al., Nature 5666:73-78 (2019); US2021 / 0106586. Exemplary age-related diseases include, but are not limited to, Alzheimer's disease, Parkinson's disease, atherosclerosis, osteoarthritis, osteoporosis, rheumatoid arthritis (RA), macular degeneration, peripheral degenerative diseases, or skin aging. In one embodiment, the subject with an age-related disease is at least 40 years old. In one embodiment, the subject with an age-related disease is at least 45 years old. In one embodiment, the subject with an age-related disease is at least 50 years old. In one embodiment, the subject with an age-related disease is at least 55 years old. In one embodiment, the subject with an age-related disease is at least 60 years old. In one embodiment, the subject with an age-related disease is at least 65 years old. In one embodiment, the subject with an age-related disease is at least 70 years old. In one embodiment, the subject suffering from an age-related disease is at least 75 years old.

[0091] In another embodiment, the disease, disorder or condition is autism spectrum disorder (ASD), cardiovascular dysfunction, hearing loss, hematopoietic stem cell function, pulmonary fibrosis, schizophrenia, or vision loss.

[0092] In another embodiment, the disease, disorder or condition is wound healing in a subject in need thereof.

[0093] In another embodiment, the disease, disorder or condition is tissue regeneration in a subject in need thereof.

[0094] In another embodiment, the disease, disorder or condition is Alzheimer's disease.

[0095] In another embodiment, the symptoms of Alzheimer's disease are any one or more of memory loss, misplacing items, forgetting the names of places or objects, repetition problems, poor dexterity, confusion, disorientation, obsessions, compulsions, delusions, aphasia, sleep disorders, mood swings, depression, anxiety, frustration, agitation, difficulty performing spatial tasks, agnosia, difficulty getting out of bed, weight loss, speech loss, short-term memory loss, or long-term memory loss, and combinations thereof.

[0096] In another embodiment, the symptom(s) of Alzheimer's disease are determined using the Alzheimer's Disease Assessment Scale (ADAS-cog), the Clinician Impression of Change Scale (CIBIC-plus), or the Activities of Daily Living Scale (ADL).

[0097] In another embodiment, disease, disorder or illness is Alzheimer's disease, and one or more optional therapeutic agents are applied to the subject. In another embodiment, optional therapeutic agent is donepezil, galantamine, rivastigmine, memantine, bapizumab, ABBV-8E12, CTS-21166, verubecestat (MK-8931), lanabecestat (lanabecestat, AZD3293), LY2886721, nicotinamide, MPTOG211 or adacanumab-avwa.

[0098] In another embodiment, the disease, disorder or condition is amyotrophic lateral sclerosis.

[0099] In another embodiment, the disease, disorder or condition is amyotrophic lateral sclerosis and one or more optional therapeutic agents are administered to the subject. In another embodiment, the optional therapeutic agent is edaravone, riluzole, raltegravir, curcumin, curcumin derivatives, chicoric acid, chicoric acid derivatives, 3,5-dicaffeoylquinic acid, 3,5-dicaffeoylquinic acid derivatives, aurintricarboxylic acid, aurintricarboxylic acid derivatives, caffeic acid phenethyl ester, caffeic acid phenethyl ester derivatives, tyrphostin, tyrphostin derivatives, quercetin, quercetin derivatives, S-1360, zintevir (AR-177), L-870812 and L-25 870810, MK-0518, BMS-538158 or GSK364735C.

[0100] In another embodiment, the disease, disorder or condition is ataxia-telangiectasia.

[0101] In another embodiment, the disease, disorder or condition is age-related macular degeneration, systemic lupus erythematosus, IFN-related autoimmune diseases, such as rheumatoid arthritis, psoriasis, vitiligo, hypothyroidism, hyperthyroidism, idiopathic thrombocytopenic purpura, autoimmune hemolytic anemia, myasthenia gravis, Addison's disease, celiac disease, polymyositis or superimposed autoimmune hepatitis, Fanconi anemia, idiopathic pulmonary fibrosis or cardiovascular disease. In another embodiment, systemic disease is age-related macular degeneration. In another embodiment, systemic disease is systemic lupus erythematosus. In another embodiment, systemic disease is IFN-related autoimmune diseases, such as psoriasis. In another embodiment, systemic disease is Fanconi anemia. In another embodiment, systemic disease is idiopathic pulmonary fibrosis. In another embodiment, systemic disease is cardiovascular disease.

[0102] In another embodiment, the disease, disorder or condition is Rett syndrome, AGS, ataxia-telangiectasia, ASD, schizophrenia, cocaine or methamphetamine abuse, FTLD or ALS. See, e.g., Terry and Devine, Front. Genet., Jan. 08, 2020, https: / / doi.org / 10.3389 / fgene.2019.01244.

[0103] In one embodiment, a Compound of the Disclosure is administered as a single agent to a subject suffering from a disease, disorder, or condition.

[0104] In another embodiment, the compounds of the present disclosure are administered to a subject suffering from a disease, disorder or condition in combination with one or more optional therapeutic agents. et al., Pharmaceuticals 2018, 11, 44; doi:10.3390 / ph11020044, Optional therapeutic agents for treating neurodegenerative diseases.

[0105] In another embodiment, the Compounds of the Disclosure are co-administered with one optional therapeutic agent to a subject suffering from a disease, disorder, or condition. In another embodiment, the Compounds of the Disclosure are co-administered with two optional therapeutic agents to a subject suffering from a disease, disorder, or condition. In another embodiment, the Compounds of the Disclosure are co-administered with three optional therapeutic agents to a subject suffering from a disease, disorder, or condition.

[0106] The Compounds of the Disclosure and one or more optional therapeutic agents may be co-administered under one or more of the following conditions: in different cycles, different durations, different concentrations, different routes of administration, and the like.

[0107] In one embodiment, a Compound of the Disclosure and one or more optional therapeutic agents are co-administered to a subject as part of a single pharmaceutical composition.

[0108] In another embodiment, the compounds of the present disclosure and one or more optional therapeutic agents are co-administered to a subject individually, e.g., as two or more separate pharmaceutical compositions. In this case, two separate pharmaceutical compositions, one comprising the compounds of the present disclosure and one comprising the optional therapeutic agent, are administered to the subject. The separate pharmaceutical compositions can be administered to the subject, e.g., in different cycles, for different durations, or by the same or different routes of use, e.g., the compounds of the present disclosure can be administered orally, and optionally the therapeutic agent can be administered intravenously.

[0109] In another embodiment, a Compound of the Disclosure is administered to a subject prior to one or more optional therapeutic agents, e.g., 0.5, 1, 2, 3, 4, 5, 10, 12, or 18 hours, 1, 2, 3, 4, 5, or 6 days, or 1, 2, 3, or 4 weeks prior to administration of one or more optional therapeutic agents.

[0110] In another embodiment, a Compound of the Disclosure is administered to a subject after one or more optional therapeutic agents, for example, 0.5, 1, 2, 3, 4, 5, 10, 12, or 18 hours, 1, 2, 3, 4, 5, or 6 days, or 1, 2, 3, or 4 weeks after administration of one or more optional therapeutic agents.

[0111] In another embodiment, a Compound of the Disclosure and one or more optional therapeutic agents are administered simultaneously.

[0112] In one embodiment, a Compound of the Disclosure is administered to a subject according to a continuous dosing schedule.

[0113] In one embodiment, a Compound of the Disclosure is administered to a subject according to an intermittent dosing regimen.

[0114] In one embodiment, the Compounds of the Disclosure are administered orally to a subject.

[0115] The therapeutic methods provided herein include administering a compound of the present disclosure to a subject suffering from a disease, disorder or condition in an amount effective to achieve its intended purpose. Although individual needs vary, determining the optimal range of the effective amount of each component belongs to the technical scope of the art. Typically, the administration amount of the compound of the present disclosure is from about 0.01 mg / kg to about 500 mg / kg, from about 0.05 mg / kg to about 100 mg / kg, from about 0.05 mg / kg to about 50 mg / kg, or from about 0.05 mg / kg to about 10 mg / kg. In one embodiment, the compound of the present disclosure is administered once a day. In another embodiment, the compound of the present disclosure is administered twice a day. In one embodiment, the compound of the present disclosure is administered three times a day. In one embodiment, the compound of the present disclosure is administered four times a day. These dosages are exemplary, but there are individual situations where higher or lower dosages are warranted, which are within the scope of the present disclosure. In practice, the physician determines the actual dosing regimen that best suits a single subject, which may vary with the age, weight and response of a particular subject.

[0116] A unit dose may include from about 0.01 mg to about 1000 mg, such as about 1 mg to about 500 mg, such as about 1 mg to about 250 mg, such as about 1 mg to about 100 mg of a Compound of the Disclosure. For example, a unit oral dose of a compound of the present disclosure may include, e.g., 1 mg, 2 mg, 3 mg, 4 mg, 5 mg, 6 mg, 7 mg, 8 mg, 9 mg, 10 mg, 11 mg, 12 mg, 13 mg, 14 mg, 15 mg, 16 mg, 17 mg, 18 mg, 19 mg, 20 mg, 21 mg, 22 mg, 23 mg, 24 mg, 25 mg, 26 mg, 27 mg, 28 mg, 29 mg, 30 mg, 31 mg, 32 mg, 33 mg, 34 mg, 35 mg, 36 mg, 37 mg, 38 mg, 39 mg, 40 mg, 41 mg, 42 mg, 43 mg, 44 mg, 45 mg, 46 mg, 47 mg, 48 mg, 49 mg. The unit dose can be once or multiple times a day, for example, as one or more tablets or capsules. The unit dose may also be administered to the subject by any suitable route, for example, orally, by IV, inhalation or subcutaneously. In practice, the physician determines the actual dosing regimen most suitable for an individual subject, which may vary with the age, weight and response of the particular subject.

[0117] In one embodiment, the compound of the present disclosure is administered to a subject in an amount of about 0.1 mg to about 100 mg, once a day, twice a day, three times a day, or four times a day. In another embodiment, the compound of the present disclosure is administered to a subject in an amount of from about 1 mg to about 50 mg per day.

[0118] In one embodiment, the Compound of the Disclosure is administered to a subject in a single dose. In another embodiment, the Compound of the Disclosure is administered to a subject in two separate doses. In another embodiment, the Compound of the Disclosure is administered to a subject in three separate doses. In another embodiment, the Compound of the Disclosure is administered to a subject in four separate doses.

[0119] The compounds of the present disclosure can be administered to a subject in the form of a raw chemical or as part of a pharmaceutical composition containing a compound of the present disclosure in combination with a suitable pharmaceutically acceptable carrier. Such a carrier can be selected from pharmaceutically acceptable excipients, vehicles and adjuvants. The terms "pharmaceutically acceptable carrier", "pharmaceutically acceptable vehicle" or "pharmaceutically acceptable vehicle" include any standard pharmaceutical carrier, solvent, surfactant or vehicle. Suitable pharmaceutically acceptable vehicles include aqueous vehicles and non-aqueous vehicles. Standard pharmaceutical carriers and their formulations are described in Remington's Pharmaceutical Sciences, Mack Publishing Co., Easton, PA, 19th edition, 1995.

[0120] A pharmaceutical composition comprising a Compound of the Disclosure may contain from about 0.01 to 99% by weight, e.g., from about 0.25 to 75% by weight, of a Compound of the Disclosure, e.g., about 1%, about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, or about 75% by weight of a Compound of the Disclosure.

[0121] The compounds of the present disclosure or pharmaceutical compositions comprising the compounds of the present disclosure can be administered to a subject by any suitable route, for example, by oral, buccal, inhalation, sublingual, rectal, vaginal, intracisternal or intrathecal administration by lumbar puncture, transurethral, ​​nasal, transdermal, i.e., transdermal, or parenteral (including intravenous, intramuscular, subcutaneous, intracoronary, intradermal, intramammary, intraperitoneal, intraarticular, intrathecal, retrobulbar, intrapulmonary injection and / or surgical implantation at a specific site). The dosage form depends on the route of administration. The dosage form includes, but is not limited to, tablets, lozenges, slow-release lozenges, capsules, liquid solutions, liquid suspensions, oral / nasal sprays, transdermal patches, dissolvable films, ointments, sustained-release or controlled-release implants, mouthwashes and gargles, gels, shampoos, hair gels and shampoos, and suppositories, as well as solutions suitable for administration by intravenous infusion, and suitable suspensions for administration by subcutaneous injection, and suitable powders for reconstitution. Parenteral administration can be accomplished using a needle and syringe or using other techniques known in the art. In one embodiment, the compounds of the present disclosure are administered orally to the subject. In one embodiment, the compounds of the present disclosure are administered subcutaneously to the subject. In one embodiment, the compounds of the present disclosure are administered intravenously to the subject.

[0122] The compounds of the present disclosure and pharmaceutical compositions comprising the compounds of the present disclosure can be administered to any subject that can experience the beneficial effects of inhibiting LINE-1 retrotransposition activity. The term "subject" as used herein refers to any person or animal that is in need of or can benefit from the administration of a compound of the present disclosure. The most important of such subjects are mammals, such as humans, although the methods and compositions provided herein are not intended to be so limited. Other subjects include veterinary animals, such as cattle, sheep, pigs, horses, dogs, cats, etc. In one embodiment, the subject is a human. In one embodiment, the subject is an animal. In another embodiment, the subject is a human suffering from a disease, condition, or disorder that is responsive to LINE-1 inhibition.

[0123] The pharmaceutical preparations provided herein are manufactured by means of conventional mixing, granulation, dragee preparation, dissolution or lyophilization processes. Thus, the pharmaceutical preparations for oral use can be obtained by mixing the active compound with a solid excipient, optionally after adding suitable auxiliaries if desired or necessary, grinding the resulting mixture and processing the E mixture of the granules to obtain tablets or dragee cores.

[0124] Suitable excipients are especially fillers, such as sugars, for example lactose or sucrose, mannitol or sorbitol, cellulose preparations and / or calcium phosphates, for example tricalcium phosphate or calcium hydrogen phosphate, and adhesives such as starch paste, using for example corn starch, wheat starch, rice starch, potato starch, gelatin, tragacanth, methylcellulose, hydroxypropyl methylcellulose, sodium carboxymethylcellulose and / or polyvinyl pyrrolidone. If necessary, disintegrants can be added, such as the above-mentioned starch and carboxymethyl starch, cross-linked polyvinyl pyrrolidone, agar or alginic acid or its salt, such as sodium alginate. Auxiliary agents can be suitable flow regulators and lubricants. Suitable auxiliary agents include, for example silicon dioxide, talcum powder, stearic acid or its salt, such as magnesium stearate or calcium stearate, and / or polyethylene glycol. The lozenge core is equipped with a suitable coating, and if necessary, the coating resists gastric juice. For this reason, concentrated sugar solutions can be used, which can optionally contain gum arabic, talcum powder, polyvinyl pyrrolidone, polyethylene glycol and / or titanium dioxide, lacquer solutions and suitable organic solvents or solvent mixtures. In order to produce a coating resistant to gastric juice, a solution of a suitable cellulose formulation is used, such as acetylcellulose phthalate or hydroxypropylmethylcellulose phthalate. Dyes or pigments can be added to tablets or dragee coatings, for example, for identification or in order to characterize the combination of active compound dosages.

[0125] Other pharmaceutical preparations that can be used orally include push-fit capsules made of gelatin, and soft, sealed capsules made of gelatin and a plasticizer such as glycerol or sorbitol. The push-fit capsules may contain the active compound in the form of granules, which may be mixed with fillers such as lactose, binders such as starches and / or lubricants such as talc or magnesium stearate and optionally stabilizers. In soft capsules, the active compound is in one embodiment dissolved or suspended in a suitable liquid, such as a fatty oil or liquid paraffin. Stabilizers may also be added.

[0126] Possible pharmaceutical preparations that can be used rectally include, for example, suppositories, which consist of a combination of one or more active compounds with a suppository base. Suitable suppository bases are, for example, natural or synthetic triglycerides or paraffins. In addition, gelatin rectal capsules can also be used, which consist of a combination of an active compound with a base. Possible base materials include, for example, liquid triglycerides, polyethylene glycols or paraffins.

[0127] Suitable formulations for parenteral administration include water-soluble forms of the active compound, such as aqueous solutions and alkaline solutions of water-soluble salts. In addition, suspensions of the compounds of the present disclosure may be administered to subjects. Suitable lipophilic solvents or vehicles include fatty oils, such as sesame oil, or synthetic fatty acid esters, such as ethyl oleate or triglycerides or polyethylene glycol-400. Aqueous injection suspensions may contain substances that increase the viscosity of the suspension, such as sodium carboxymethylcellulose, sorbitol and / or dextran. Optionally, the suspension may also contain stabilizers and other additives.

[0128] A therapeutically effective amount of a compound of the present disclosure, formulated in accordance with standard pharmaceutical practice, is administered to a subject in need thereof. Whether such treatment is indicated depends on the individual case, and upon medical evaluation (diagnosis), which takes into account the presence of signs, symptoms, and / or disorders, the risk of developing specific signs, symptoms, and / or disorders, and other factors.

[0129] Pharmaceutical compositions include those in which the compounds of the present disclosure are administered in effective amounts to achieve their intended purpose. The exact formulation, route of administration, and dosage are determined by a single physician based on the diagnosed condition or disease. Dosage amounts and intervals can be adjusted individually to provide levels of the compounds of the present disclosure sufficient to maintain a therapeutic effect.

[0130] The toxicity and therapeutic efficacy of the compounds of the present disclosure can be determined by standardized drug procedures in cell cultures or experimental animals, for example, for determining the maximum tolerated dose (MTD) of the compound, which is defined as the highest dose that does not cause toxicity in a subject. The dose ratio between the maximum tolerated dose and the therapeutic effect is the therapeutic index. Depending on the dosage form employed and the route of administration used, the dose varies within this range. The determination of a therapeutically effective amount is well within the capabilities of those skilled in the art, particularly in light of the detailed disclosure provided herein.

[0131] The therapeutically effective amount of the compound of the present disclosure required for therapy varies with the characteristics of the disease being treated, the length of time of activity required, and the age and condition of the subject, and is ultimately determined by the attending physician. For example, the dosage amount and interval can be adjusted individually to provide a plasma level of the compound of the present disclosure sufficient to maintain the desired therapeutic effect. The desired dose can be conveniently administered as a single dose, or as multiple doses at appropriate intervals, such as one, two, three, four or more sub-doses per day.

[0132] IV. Kit

[0133] In another embodiment, the disclosure provides a kit comprising a compound of the disclosure, or a pharmaceutical composition thereof, and instructions for administering the compound or composition to a subject suffering from a disease, disorder, or condition.

[0134] In another embodiment, the present disclosure provides kits comprising a compound of the present disclosure or a pharmaceutical composition thereof, packaged in a manner to facilitate their use in practicing the methods of the present disclosure.

[0135] In one embodiment, the kit includes a compound of the present disclosure or a pharmaceutical composition thereof, packaged in a container such as a sealed bottle or tube, affixed to the container or a label contained in the kit, the label describing the use of the compound or composition to implement the method of the present disclosure. In one embodiment, the compound or composition is packaged in unit dosage form. The kit may include a single dose or multiple doses of a compound of the present disclosure, or a pharmaceutical composition thereof.

[0136] In another embodiment, a kit includes a Compound of the Disclosure, or a composition thereof, and one or more optional therapeutic agents.

[0137] V. Biomarkers

[0138] In another embodiment, the present disclosure provides a method of treating a subject having a disease, condition or disorder, the method comprising (a) determining the presence or absence of a biomarker in a biological sample taken from the subject; and (b) if the biomarker is present in the biological sample, administering to the subject a therapeutically effective amount of a compound of the present disclosure.

[0139] The term "biomarker" as used herein refers to any biological compound that can be detected and / or quantified in a subject or in a biological sample obtained from a subject, such as a gene, protein, protein fragment, peptide, polypeptide, nucleic acid, etc., or a chromosomal abnormality, such as a chromosomal translocation. A biomarker can be a whole intact molecule, or it can be a part or fragment thereof. In one embodiment, the expression level of a biomarker is measured. The expression level of a biomarker can be measured, for example, by detecting the protein or RNA (e.g., mRNA) level of a biomarker. In some embodiments, a part or fragment of a biomarker can be detected or measured, for example, by an antibody or other specific binding agent. In some embodiments, the measurable aspects of a biomarker are related to a given state of a subject, such as the age of the subject. For biomarkers detected at the protein or RNA level, such measurable aspects can include, for example, the presence, absence or concentration of a biomarker in a subject, i.e., the expression level, or a biological sample obtained from a subject. For biomarkers detected at the nucleic acid level, such measurable aspects can include, for example, the allele version of a biomarker or the mutation type, mutation rate and / or mutation degree of a biomarker, also referred to herein as mutation state.

[0140] For biomarkers detected based on protein or RNA expression levels, the expression levels measured between different phenotypic states can be considered to be different, for example, if the average or median expression levels of the biomarkers in different groups are calculated to be statistically significant. Common statistical significance tests include t-tests, ANOVA, Kruskal-Wallis, Wilcoxon, Mann-Whitney, microarray significance analysis, odds ratios, etc., in particular. Biomarkers alone or in combination provide a measure of the relative likelihood that a subject belongs to one or another phenotypic state. Therefore, they are particularly useful as markers of disease and indicators that a specific therapeutic treatment regimen may bring beneficial patient outcomes. The term "overexpression" indicates that the expression level of a biomarker in a subject with a disease, disorder, or disorder is higher than the average or median expression level of a biomarker in, for example, a normal, non-diseased subject.

[0141] Biomarkers include, but are not limited to, retrotransposon RNA, retrotransposon reverse transcriptases such as ORF1p, ORF2p, and / or retrotransposon DNA. In one embodiment, the measurable aspect of a biomarker is its expression state. In another embodiment, the measurable aspect of a biomarker is an elevated biomarker level. In one embodiment, the measurable aspect of a biomarker is its mutation state.

[0142] In one embodiment, the biomarker is the expression level of LINE-1. Methods for determining the expression level of LINE-1 are described in US2020 / 0253888 and can include, for example, determining the level of ORF1p, determining the level of LINE-1 mRNA, determining the amount of LINE-1 in a cell sample of a subject, or determining the level of ORF2p, or a combination thereof.

[0143] In one embodiment, the biomarker is expression of a retrotransposon RNA that is differentially present in a subject in one phenotypic state, such as a subject with an age-related disease or a neurodegenerative disease, compared to another phenotypic state, such as a normal non-diseased subject or a subject with a disease, disorder or condition without overexpression of the retrotransposon RNA. In one embodiment, the biomarker is overexpression of a retrotransposon RNA.

[0144] In one embodiment, the biomarker is LINE-1 RNA expression, which is differentially present in a subject in one phenotypic state, such as a subject with an age-related disease or a neurodegenerative disease, compared to another phenotypic state, such as a normal non-diseased subject or a subject with a disease, disorder or condition that does not overexpress LINE-1 RNA. In one embodiment, the biomarker is overexpression of LINE-1 RNA.

[0145] In another embodiment, the biomarker is a retrotransposon reverse transcriptase that is differentially present in a subject in one phenotypic state, such as a subject with an age-related disease or a neurodegenerative disease, compared to another phenotypic state, such as a normal non-diseased subject or a subject with a disease, disorder or condition that does not overexpress the retrotransposon reverse transcriptase. In one embodiment, the biomarker is overexpression of a retrotransposon reverse transcriptase.

[0146] In another embodiment, the biomarker is ORF1p expression, which is differentially present in a subject in one phenotypic state, such as a subject with an age-related disease or a neurodegenerative disease, compared to another phenotypic state, such as a normal non-diseased subject or a subject with a disease, disorder or condition that does not overexpress ORF1p. In one embodiment, the biomarker is overexpression of ORF1p.

[0147] In another embodiment, the biomarker is ORF2p expression that is differentially present in a subject of one phenotypic state, such as a subject with an age-related disease or a neurodegenerative disease, compared to another phenotypic state, such as a normal non-diseased subject or a subject with a disease, disorder or condition that does not overexpress ORF2p. In one embodiment, the biomarker is overexpression of ORF2p.

[0148] Biomarker standards (products) can be determined in advance, simultaneously or after obtaining a biological sample from a subject. For example, the biomarker standards used with the methods described herein can include data from samples of subjects without neurodegenerative diseases; data from samples of subjects with neurodegenerative diseases. Comparisons can be made to establish predetermined threshold biomarker standards for different categories of subjects, such as sick subjects and non-sick subjects. Standards (products) can be run in the same test, or can be known standards from previous tests.

[0149] In one embodiment, the biomarker is the expression of retrotransposon DNA, which is differentially present in a subject of one phenotypic state, such as a subject with an age-related disease or a neurodegenerative disease, compared to another phenotypic state, such as a normal non-diseased subject or a subject with a disease, disorder or condition that does not overexpress retrotransposon DNA. In one embodiment, the biomarker is overexpression of retrotransposon DNA. In another embodiment, the biomarker is overexpression of retrotransposon nuclear DNA. In another embodiment, the biomarker is overexpression of retrotransposon cytoplasmic DNA.

[0150] In one embodiment, the biomarker is LINE-1 DNA expression, which is differentially present in a subject of one phenotypic state, such as a subject with an age-related disease or a neurodegenerative disease, compared to another phenotypic state, such as a normal non-diseased subject or a subject with a disease, disorder or condition that does not overexpress LINE-1 DNA. In one embodiment, the biomarker is overexpression of LINE-1 DNA. In another embodiment, the biomarker is overexpression of LINE-1 nuclear DNA. In another embodiment, the biomarker is overexpression of LINE-1 cytoplasmic DNA.

[0151] If the mean or median expression level or mutation level of the biomarker is calculated to be different between the groups, i.e., higher or lower, then the biomarker is differentially present between the different phenotypic state groups. Thus, the biomarker provides an indication that a subject, e.g., a subject with ALS, belongs to one phenotypic state or another.

[0152] In addition to a single biological compound, such as LINE-1 RNA, the term "biomarker" as used herein means a group, set or array that includes multiple biological compounds. For example, a combination of LINE-1 RNA overexpression and ORF1p overexpression can comprise a biomarker, or overexpression of LINE-1 RNA and LINE-1 DNA can comprise a biomarker. The term "biomarker" can include one, two, three, four, five, six, seven, eight, nine, ten, fifteen, twenty, twenty-five, thirty or more biological compounds. In an embodiment, the biomarker includes one, two or three biological compounds.

[0153] The expression level or mutation status of a biomarker in a subject can be determined using any of the many methods known in the art. Any method known in the art for quantifying specific proteins and / or detecting biomarker expression in a patient or biological sample, such as LINE-1 RNA expression, ORF1p expression and / or ORF2p expression, or any other (one or more) biomarker expression or mutation level can be used in the methods of the present disclosure. Examples include, but are not limited to, PCR (polymerase chain reaction) or RT-PCR, flow cytometry, Northern blot, Western blot, ELISA (enzyme-linked immunosorbent assay), RIA (radioimmunoassay), gene chip analysis of RNA expression, immunohistochemistry, or immunofluorescence. For example, see Slagle, Cancer 83: 1401 (1998). Certain embodiments of the present disclosure include methods in which biomarker RNA expression (transcription) is determined. Other embodiments of the present disclosure include methods in which protein expression in a biological sample is determined. For example, see Harlow et al., Antibodies: A Laboratory Manual, Cold Spring Harbor Laboratory, Cold Spring Harbor, NY, (1988); Ausubel et al., Current Protocols in Molecular Biology, John Wiley & Sons, New York, 3rd edition (1995); Kamel and Al-Amodi, Genomics Proteomics Bioinformatics 15: 220-235 (2017). For Northern blot or RT-PCR analysis, RNA is isolated from tissue samples using RNase-free techniques. Such techniques are well known in the art.

[0154] In one embodiment of the present disclosure, a biological sample is obtained from a subject and the biological sample is assayed to determine the expression or mutation status of a biomarker.

[0155] In another embodiment of the present disclosure, Northern blot analysis of biomarker transcription in tumor cell samples is performed. Northern analysis is a standard method for detecting and / or quantifying mRNA levels in samples. Initially, RNA is isolated from the sample to be determined using Northern blot analysis. In the analysis, RNA samples are first separated by size by electrophoresis in an agarose gel under denaturing conditions. The RNA is then transferred to a membrane, cross-linked and hybridized with a labeled probe. Typically, Northern hybridization involves in vitro polymerization of radiolabeled or non-isotope labeled DNA, or the generation of oligonucleotides as hybridization probes. Typically, before probe hybridization, the membrane to which the RNA sample is fixed is pre-hybridized or blocked to prevent the probe from coating the membrane, thereby reducing non-specific background signals. After hybridization, unhybridized probes are typically removed by washing with multiple replacement buffers. The stringency of washing and hybridization conditions can be designed, selected and implemented by any ordinary practitioner in the art. Detection is accomplished using a detectably labeled probe and a suitable detection method. Radiolabeled and non-radiolabeled probes and their use are well known in the art. The presence and / or relative expression level of the biomarker to be determined can be quantified using, for example, densitometry.

[0156] In another embodiment, RT-PCR is used to determine biomarker expression and / or mutation status. RT-PCR allows real-time detection of the progress of PCR amplification of the target gene. The design of primers and probes required for detecting the expression and / or mutation status of the biomarkers of the present disclosure is within the skill range of ordinary practitioners in the art. For example, RT-PCR can be used to determine the level of RNA encoding the biomarkers of the present disclosure in a tissue sample. In an embodiment of the present disclosure, RNA is isolated from a biological sample under the condition of no RNase and then converted to DNA by treatment with reverse transcriptase. The method of converting RNA to DNA by reverse transcriptase is well known in the art. Descriptions of PCR are provided in the following references: Mullis et al., Cold Spring Harbor Symp. Quant. Biol. 51:263 (1986); EP 50,424; EP 84,796; EP 258,017; EP 237,362; EP 201,184; U.S. Pat. Nos. 4,683,202; 4,582,788; 4,683,194.

[0157] RT-PCR probes rely on the 5'-3' nuclease activity of the DNA polymerase used for PCR to hydrolyze oligonucleotides hybridized with the target amplicon (biomarker gene). RT-PCR probes are oligonucleotides with a fluorescent reporter dye connected to the 5' end and a quenching structure part coupled to the 3' end (or vice versa). These probes are intended to hybridize with the internal region of the PCR product. In the unhybridized state, the proximity of the fluorescent and quenching molecules prevents the detection of the fluorescent signal from the probe. During PCR amplification, when the polymerase copies the template bound to the RT-PCR probe, the 5'-3' nuclease activity of the polymerase cleaves the probe. This decouples the fluorescent dye and the quenching dye, and FRET no longer occurs. Therefore, fluorescence increases in each cycle in a manner proportional to the amount of probe cleavage. Commercially available equipment using conventional and traditional techniques can be used to measure or track the fluorescent signal emitted by the reaction over time.

[0158] In another embodiment of the present disclosure, the expression of proteins encoded by biomarkers is detected by western blot analysis. Western blot (also known as immunoblotting) is a method for protein detection in a given sample of a tissue homogenate or extract. It uses gel electrophoresis to separate denatured proteins by mass. The proteins are then transferred from the gel to a membrane (e.g., nitrocellulose or polyvinylidene fluoride (PVDF)), where they are detected using a primary antibody that specifically binds to the protein. The bound antibody can then be detected by a secondary antibody coupled to a detectable marker (e.g., biotin, horseradish peroxidase, or alkaline phosphatase). Detection of a secondary marker signal indicates the presence of the protein.

[0159] In another embodiment of the present disclosure, the expression of proteins encoded by biomarkers is detected by enzyme-linked immunosorbent assay (ELISA). In one embodiment of the present disclosure, a "sandwich ELISA" includes coating a plate with a capture antibody; adding a sample, wherein any antigen present is bound to the capture antibody; adding a detection antibody that also binds to the antigen; adding an enzyme-linked secondary antibody that binds to the detection antibody; and adding a substrate that is converted to a detectable form by an enzyme on the secondary antibody. Detection of a signal from the secondary antibody indicates the presence of a biomarker antigen protein.

[0160] In another embodiment of the present disclosure, by s single molecule array assay (Simoa TM ) detects the expression of proteins encoded by biomarkers, such as ORF1p and ORF2p.

[0161] In another embodiment of the present disclosure, the expression of proteins encoded by biomarkers, such as ORF1p, ORF2p, is detected by droplet digital ELISA (ddELISA). LINE-1 / ORF1p protein in serum can be measured using ddELISA. See Cohen et al., ACS Nano 14:9491-9501 (2020).

[0162] VI. Definitions

[0163] As used herein, the term "pathophysiological retrotransposon-related process" refers to a disordered physiological process associated with the abnormal retrotransposition activity of at least one retrotransposon. Exemplary retrotransposons include, but are not limited to, LINE-1 and human endogenous retroviruses (HERVs), such as HERV-K and HERV-E. For example, see Saleh et al., (2019) Front. Neurol. 10: 894. doi: 10.3389 / fneur.2019.00894. Diseases, disorders or conditions caused by pathophysiological retrotransposon-related processes include, but are not limited to, neurodegenerative diseases, autoimmune diseases, age-related diseases, autism spectrum disorders (ASD), cardiovascular dysfunction, hearing loss, hematopoietic stem cell function, pulmonary fibrosis, schizophrenia or vision loss.

[0164] As used herein, the term "pathophysiological LINE-1-related process" refers to a disordered physiological process associated with abnormal LINE-1 (L1) retrotransposition activity. For example, see Saleh et al., (2019) Front. Neurol. 10:894. doi: 10.3389 / fneur.2019.00894; Zhao et al., PLoS Genet 15(4):e1008043. https: / / doi.org / 10.1371 / journal.pg en.1008043; Bundo et al., Neuron 81:306-313 (2014).

[0165] As used herein, the term "LINE-1 retrotransposition event causing a disease, disorder or condition" refers to any pathogenic factor associated with LINE-1 retrotransposition that causes or promotes a pathological condition, such as a disease or disorder, in a subject, such as aberrant transcription, alternative splicing, insertion mutations, DNA damage, chromosomal translocations, increased LINE-1 RNA expression, ORF1p (40 kDa RNA binding protein), ORF2p (~150 kDa protein with endonuclease (EN) and reverse transcriptase (RT) activities). See, e.g., Beck et al., Annu Rev Genomics Hum Genet 12: 187-215 (2011); Pizarro and Cristofari (2016) Front. Cell Dev. Biol. 4: 14, https: / / doi.org / 10.3389 / fcell.2016.00014. In one embodiment, the LINE-1 retrotransposition event is a somatic LINE-1 insertion. In another embodiment, the LINE-1 retrotransposition event is increased expression of LINE-1 RNA in the subject.

[0166] As used herein, the term "tautomer" refers to each of two or more isomers of a compound, which coexist in an equilibrium state and interconvert by intramolecular atoms, such as the migration of hydrogen or groups. Certain compounds of the present disclosure may exist in tautomeric forms. Where tautomers may exist, the present disclosure includes all tautomeric forms. For example, as shown in Figure 1, lactim and lactam tautomers are included, and both amino and imino tautomers are included.

[0167] Figure 1

[0168]

[0169] Likewise, as shown in Chart 2, both lactim and lactam tautomers are included, and both amino and imino tautomers are included.

[0170] Figure 2

[0171]

[0172] The equilibrium arrows in Diagrams 1 and 2 are not intended to show the position of the equilibrium, but are only intended to show that an equilibrium exists between the two tautomeric forms.

[0173] The term "biological sample" as used herein refers to any tissue or liquid from a subject suitable for detecting a biomarker. Examples of useful biological samples include, but are not limited to, biopsy tissue and / or cells, such as lymph glands, inflammatory tissues, tissues and / or cells involved in illnesses or diseases, blood, plasma, serous fluid, cerebrospinal fluid, saliva, urine, lymph, cerebrospinal fluid, etc. Other suitable biological samples will be familiar to those of ordinary skill in the relevant field. Any technique known in the art can be used to analyze the biological compounds of biological samples, such as the expression level of LINE-1 RNA, ORF1p protein, ORF2p protein). Such techniques include, but are not limited to, polymerase chain reaction (PCR) methodology, reverse transcription polymerase chain reaction (RT-PCR) methodology, or cytoplasmic light chain immunofluorescence combined with fluorescence in situ hybridization (cIg-FISH). Biological samples can be obtained using techniques that are completely within the common sense of clinical practitioners. In one embodiment of the present disclosure, the biological sample includes a tissue or blood sample.

[0174] Unless otherwise stated, the terms "a / an", "the" and similar references in the context of describing the present disclosure (particularly in the context of the claims) should be interpreted as covering both the singular and the plural. Unless otherwise stated herein, the recitation of ranges of values ​​herein is intended only to serve as a shorthand method of individually referring to each individual value falling within the range, and each individual value is incorporated into the specification as if it were individually recorded herein. Unless otherwise stated, the use of any and all examples or exemplary language provided herein, such as "such as", is intended to better illustrate the present disclosure rather than to limit the scope of the present disclosure. No language in the specification should be interpreted as indicating that any unclaimed element is essential to the practice of the present disclosure.

[0175] As used herein, the term "about" includes ±10% of the cited number. Thus, "about 10" means 9 to 11.

[0176] As used herein, the terms "treat", "treating", "treatment", and the like refer to the elimination, reduction, or amelioration of a disease, disorder, or condition, and / or symptoms associated therewith. Although not excluded, treatment of a disease, disorder, or condition does not require the complete elimination of the disease, disorder, or condition and / or the symptom(s) associated therewith. However, in one embodiment, administration of a compound of the present disclosure results in the complete elimination of the disease and associated symptoms.

[0177] As used herein, the terms "prevent" (preventing / prevention) and the like refer to methods of preventing the onset of a disease, disorder or condition and / or symptoms associated therewith, or preventing a subject from developing a disease, disorder or condition. The term "prevention" also includes delaying the onset of a disease, disorder or condition and / or its associated symptoms (one or more), and reducing the risk of a subject developing a disease, disorder or condition. The term "prevention" also includes "prophylactic treatment," which refers to reducing the likelihood of a subject who does not have but is at risk or susceptible to developing a disease, disorder or condition again or a recurrence of a disease, disorder or condition, or the likelihood of a previously controlled disease, disorder or condition recurring. The term "prevention" also includes delaying or reversing the potential pathological progression of a disease, disorder or condition, for example, mutations caused by somatic LINE-1 insertions.

[0178] As used herein, the term "therapeutically effective amount" refers to an amount of a compound of the present disclosure and optionally one or more optional therapeutic agents, sufficient to cause an improvement in one or more symptoms of a disease, disorder, or condition, or to prevent the progression of a disease, disorder, or condition, or to cause regression of a disease, disorder, or condition. For example, a therapeutically effective amount would refer to an amount of a compound of the present disclosure that causes a therapeutic response, such as delaying the progression of a disease, disorder, or condition in a subject by at least about 2%, at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or at least about 100%, or more.

[0179] The term "container" refers to any receptacle and closure suitable for storage, transport, distribution, and / or handling of the compounds of the present disclosure. Non-limiting exemplary containers include vials, ampoules, bottles, and syringes.

[0180] The term "insert" refers to the information that accompanies a drug product that provides a description of how to administer the product, along with the safety and effectiveness data needed to allow physicians, pharmacists, and subjects to make an informed decision about the product's use. The package insert is often considered the "label" for the drug.

[0181] In some embodiments, two or more therapeutic agents can have a synergistic effect when administered in combination. As used herein, the terms "synergistic", "synergistic", "synergistically" and derivatives thereof, such as in "synergistic effect" or "synergistic combination" or "synergistic composition", refer to situations where the biological activity of the combination of a medicament and at least one additional therapeutic agent is greater than the sum of the biological activities of the corresponding medicaments when administered alone. For example, as used herein, the term "synergistically effective" refers to an interaction between a compound of the present disclosure and another therapeutic agent that results in a total effect of the drug that is greater than the sum of the individual effects of each drug. Berenbaum, Pharmacological Reviews 41:93-141 (1989).

[0182] Synergy can be expressed as a "synergy index (SI)", which can generally be determined by the method described in FC Kull et al., Applied Microbiology 9, 538 (1961) by determining the following ratio:

[0183] Q a Q A +Q b Q B =Synergy Index (SI)

[0184] in:

[0185] Q A is the concentration of component A acting alone that produces the endpoint with respect to component A;

[0186] Q a is the concentration of component A in the mixture that produces the endpoint;

[0187] Q B is the concentration of the B component acting alone that produces the endpoint with respect to the B component; and

[0188] Q b is the concentration of component B in the mixture that produces the endpoint.

[0189] Usually, when Q a / Q A and Q b / Q B When the sum is greater than 1, antagonism is indicated. When the sum is equal to 1, an additive effect is indicated. When the sum is less than 1, synergism is demonstrated. The lower the SI, the greater the synergistic effect exhibited by the particular mixture. Thus, the activity of a "synergistic combination" is higher than the expected activity based on the individual components observed when used alone. In addition, a "synergistically effective amount" of a component refers to the amount of the component required to cause a synergistic effect, for example, in another therapeutic agent present in the composition.

[0190] As used herein, the terms "intermittent dosage administration," "intermittent dosing regimen," and similar terms refer to the administration of a Compound of the Disclosure to a subject, ie, not continuously.

[0191] The intermittent dosing of the compounds of the present disclosure can maintain or achieve the efficacy of continuous administration, but with less side effects, for example, less weight loss. Useful intermittent dosing regimens in the present disclosure include any discontinuous dosing regimens that provide a therapeutically effective amount of the compounds of the present disclosure to a subject in need thereof. The intermittent dosing regimen can use the compounds of the present disclosure at equal, lower or higher doses than those used in the continuous dosing regimen. The advantages of intermittent dosing of the compounds of the present disclosure include, but are not limited to, improving safety, reducing toxicity, for example, reducing weight loss, increasing exposure, improving efficacy and / or increasing subject compliance. These advantages can be achieved when the compounds of the present disclosure are administered as a single agent or in combination with one or more optional therapeutic agents. On the day when the compounds of the present disclosure are administered to the subject as planned, they can be administered in a single dose or in separate doses, for example, once a day, twice a day, three times a day, four times a day or more. It can also be administered by any suitable route, for example, orally, intravenously or subcutaneously. In one embodiment, the compounds of the present disclosure are administered once (QD) or twice (BID) to the subject on the day when the compounds are administered as planned.

[0192] In one embodiment, the Compounds of the Disclosure are administered according to a continuous dosing schedule. In another embodiment, the Compounds of the Disclosure are administered according to an intermittent dosing schedule.

[0193] The phrase "in combination with..." used in conjunction with administering a compound of the present disclosure and one or more optional therapeutic agents to a subject means that the compound of the present disclosure and one or more optional therapeutic agents can be administered to the subject together, such as as part of a single pharmaceutical composition or formulation, or separately, such as as part of two or more separate pharmaceutical compositions or formulations. Thus, the phrase "in combination with..." used in conjunction with administering a compound of the present disclosure and one or more optional therapeutic agents to a subject is intended to include the use of the compound of the present disclosure and one or more optional therapeutic agents in a sequential manner, wherein the compound of the present disclosure and one or more optional therapeutic agents are administered to the subject at different times, as well as administration in a simultaneous or substantially simultaneous manner, such as less than 30 minutes apart. Simultaneous administration can be accomplished, for example, by administering to the subject a single capsule having a fixed ratio of each compound of the present disclosure and one or more optional therapeutic agents or a plurality of individual capsules of each compound of the present disclosure and one or more optional therapeutic agents. The sequential or substantially simultaneous administration of the compound of the present disclosure and one or more optional therapeutic agents can be accomplished by any appropriate route, including but not limited to oral routes, intravenous routes, subcutaneous routes, intramuscular routes, and the like. The compounds of the present disclosure and one or more optional therapeutic agents can be administered by the same route or by different routes. For example, one or more optional therapeutic agents and the compounds of the present disclosure can be administered orally. Alternatively, for example, the compounds of the present disclosure can be administered orally, and one or more optional therapeutic agents can be administered by intravenous injection. The compounds of the present disclosure and one or more optional therapeutic agents can also be administered alternately. In one embodiment, the compounds of the present disclosure and one or more optional therapeutic agents are administered separately to the subject, for example as part of two or more separate pharmaceutical compositions or formulations.

[0194] VII. Specific Implementation Methods

[0195] The present disclosure provides the following specific embodiments.

[0196] Embodiment 1. A method of treating or preventing a disease, disorder or condition caused by a pathophysiological retrotransposon-related process, and / or treating or preventing a symptom of a disease, disorder or condition in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of a compound of:

[0197] (i) Table 1, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof;

[0198] (ii) Table 2, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof; or

[0199] (iii) Table 3, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0200] Embodiment 2. A method of inhibiting a LINE-1 retrotransposition event that causes a disease, disorder or condition in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of a compound of:

[0201] (i) Table 1, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof;

[0202] (ii) Table 2, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof; or

[0203] (iii) Table 3, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0204] Embodiment 3. A method of treating a disease, condition or disorder in a subject, the method comprising:

[0205] (a) determining the presence or absence of overexpression of retrotransposon RNA, retrotransposon reverse transcriptase or retrotransposon DNA in a biological sample taken from a subject; and

[0206] (b) if there is overexpression of retrotransposon RNA, retrotransposon reverse transcriptase, or retrotransposon DNA in the biological sample, administering to the subject a therapeutically effective amount of a compound:

[0207] (i) Table 1, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof;

[0208] (ii) Table 2, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof; or

[0209] (iii) Table 3, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0210] Embodiment 4. A method of identifying whether a subject having a disease, condition or disorder is a candidate for treatment with a compound of:

[0211] (i) Table 1, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof,

[0212] (ii) Table 2, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof; or

[0213] (iii) Table 3, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0214] The method includes:

[0215] (a) determining the presence or absence of overexpression of retrotransposon RNA, retrotransposon reverse transcriptase or retrotransposon DNA in a biological sample taken from a subject; and

[0216] (b) identifying the subject as a candidate for treatment if there is overexpression of retrotransposon RNA, retrotransposon reverse transcriptase, or retrotransposon DNA; or

[0217] (c) if there is no overexpression of retrotransposon RNA, retrotransposon reverse transcriptase, or retrotransposon DNA, identifying the subject as not a candidate for treatment.

[0218] Embodiment 5. A method of predicting a treatment outcome in a subject having a disease, condition or disorder, the method comprising determining the presence or absence of overexpression of a retrotransposon RNA, a retrotransposon reverse transcriptase or a retrotransposon DNA in a biological sample taken from the subject, wherein:

[0219] (a) the presence of overexpression of retrotransposon RNA, retrotransposon reverse transcriptase, or retrotransposon DNA in a biological sample indicates that administration of a compound to the subject may result in a beneficial therapeutic response:

[0220] (i) Table 1, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof;

[0221] (ii) Table 2, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof; or

[0222] (iii) Table 3, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof; and

[0223] (b) the absence of overexpression of retrotransposon RNA, retrotransposon reverse transcriptase, or retrotransposon DNA in the biological sample indicates that administration of the following compound to the subject may cause an adverse therapeutic response:

[0224] (i) Table 1, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof;

[0225] (ii) Table 2, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof; or

[0226] (iii) Table 3, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0227] Embodiment 6. A method comprising administering to a subject a therapeutically effective amount of a compound of:

[0228] (i) Table 1, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof:

[0229] (ii) Table 2, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof; or

[0230] (iii) Table 3, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof,

[0231] in:

[0232] (a) the subject suffers from a disease, condition or disorder; and

[0233] (b) The disease, condition or disorder is characterized by overexpression of retrotransposon RNA, retrotransposon reverse transcriptase or retrotransposon DNA.

[0234] Embodiment 7. The method of any one of Embodiments 1-6, comprising a compound of Table 1, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0235] Embodiment 8. The method of any one of Embodiments 1-6, comprising a compound of Table 2, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0236] Embodiment 9. The method of embodiment 7, wherein the compound of Table 1 is compound No. 1, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0237] Embodiment 10. The method of Embodiment 7, wherein the compound of Table 1 is Compound No. 3, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0238] Embodiment 11. The method of Embodiment 7, wherein the compound of Table 1 is Compound 8, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0239] Embodiment 12. The method of Embodiment 7, wherein the compound of Table 1 is Compound No. 9, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0240] Embodiment 13. The method of embodiment 12, wherein the compound of Table 1 is compound No. 4, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0241] Embodiment 14. The method of any one of Embodiments 1-6, comprising a compound of Table 3, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0242] Embodiment 15. The method of any one of Embodiments 1-14, for treating a disease, disorder or condition in a subject.

[0243] Embodiment 16. The method of any one of Embodiments 1-14, for preventing a disease, disorder or condition in a subject.

[0244] Embodiment 17. The method of any one of Embodiments 1-14, for treating a symptom of a disease, disorder or condition in a subject.

[0245] Embodiment 18. The method of any one of Embodiments 1-14, for preventing symptoms of a disease, disorder, or condition in a subject.

[0246] Embodiment 19. The method of any one of embodiments 1-18, wherein the disease, disorder or condition is a neurodegenerative disease.

[0247] Embodiment 20. The method of embodiment 19, wherein the neurodegenerative disease is Alzheimer's disease, amyotrophic lateral sclerosis, Parkinson's disease, dementia with Lewy bodies, multiple system atrophy, Huntington's disease, frontotemporal lobar degeneration, mild cognitive impairment, corticobasal degeneration, progressive supranuclear palsy, Rett syndrome, peripheral degenerative disease or Aicardi Goutières syndrome.

[0248] Embodiment 21. The method of any one of embodiments 1-18, wherein the disease, disorder or condition is an autoimmune disease.

[0249] Embodiment 22. The method of embodiment 11, wherein the autoimmune disease is lupus, rheumatoid arthritis, Sjögren's syndrome, or multiple sclerosis.

[0250] Embodiment 23. The method of any one of embodiments 1-18, wherein the disease, disorder or condition is an age-related disease.

[0251] Embodiment 24. The method of embodiment 23, wherein the age-related disease is Alzheimer's disease, Parkinson's disease, atherosclerosis, osteoarthritis, osteoporosis, rheumatoid arthritis, macular degeneration, peripheral degenerative disease or skin aging.

[0252] Embodiment 25. The method of any one of embodiments 1-18, wherein the disease, disorder or condition is autism spectrum disorder (ASD), cardiovascular dysfunction, hearing loss, hematopoietic stem cell function, pulmonary fibrosis, schizophrenia, or vision loss.

[0253] Embodiment 26. The method of any one of embodiments 1-18, wherein the disease, disorder or condition is progressive supranuclear palsy.

[0254] Embodiment 27. The method of any one of Embodiments 1-18, wherein the disease, disorder or condition is amyotrophic lateral sclerosis.

[0255] Embodiment 28. The method of any one of embodiments 1-18, wherein the disease, disorder or condition is Aicardi-Goutières syndrome.

[0256] Embodiment 29. The method of any one of embodiments 1-3 or 5-28, further comprising administering to the subject one or more optional therapeutic agents.

[0257] Embodiment 30. The method of any one of embodiments 1-29, wherein the subject (a) is not infected with HIV; (b) is not suspected of being infected with HIV; (c) is not receiving HIV treatment; and / or (d) is not receiving HIV prophylactic treatment.

[0258] Embodiment 31. The method of any one of embodiments 1-30, wherein the compound inhibits human LINE-1 retrotransposition activity in an in vitro HeLa cell-based dual-luciferase assay at a half-maximal inhibitory concentration of less than 1 μM.

[0259] Embodiment 32. A kit for implementing the method of any one of claims 1 to 31, comprising the following compounds:

[0260] (i) Table 1, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof:

[0261] (ii) Table 2, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof; or

[0262] (iii) Table 3, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof, and instructions for administering the compound to a subject suffering from a disease, condition, or disorder.

[0263] Embodiment 33. The kit of Embodiment 32 comprises a compound of Table 1, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0264] Embodiment 34. The kit of Embodiment 32 comprises a compound of Table 2, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0265] Embodiment 35. The kit of Embodiment 33, wherein the compound of Table 1 is Compound No. 1, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0266] Embodiment 36. The kit of embodiment 33, wherein the compound of Table 1 is compound No. 3, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof. .

[0267] Embodiment 37. The kit of Embodiment 33, wherein the compound of Table 1 is Compound No. 8, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0268] Embodiment 38. The kit of Embodiment 33, wherein the compound of Table 1 is Compound No. 9, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0269] Embodiment 39. The kit of Embodiment 33, wherein the compound of Table 1 is Compound No. 4, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0270] Embodiment 40. The kit of Embodiment 32 comprises a compound of Table 3, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0271] Embodiment 41. The following compound:

[0272] (i) Table 1, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof, or a pharmaceutical composition thereof;

[0273] (ii) Table 2, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof, or a pharmaceutical composition thereof; or

[0274] (iii) Table 3, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof, or a pharmaceutical composition thereof,

[0275] They are useful for treating or preventing a disease, disorder or condition caused by a pathophysiological retrotransposon-associated process and / or treating or preventing the symptoms of said disease, disorder or condition in a subject in need thereof.

[0276] Embodiment 42. The following compound:

[0277] (i) Table 1, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof, or a pharmaceutical composition thereof;

[0278] (ii) Table 2, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof, or a pharmaceutical composition thereof; or

[0279] (iii) Table 3, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof, or a pharmaceutical composition thereof,

[0280] They are useful for inhibiting a disease, disorder or condition-causing LINE-1 retrotransposition event in a subject in need thereof.

[0281] Embodiment 43. The following compound:

[0282] (i) Table 1, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof, or a pharmaceutical composition thereof;

[0283] (ii) Table 2, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof, or a pharmaceutical composition thereof; or

[0284] (iii) Table 3, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof, or a pharmaceutical composition thereof,

[0285] They are useful for treating a disease, condition or disorder characterized by overexpression of retrotransposon RNA, retrotransposon reverse transcriptase or retrotransposon DNA.

[0286] Embodiment 44. The compound of any one of Embodiments 41-43, comprising a compound of Table 1, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0287] Embodiment 45. The compound of any one of Embodiments 41-43, comprising a compound of Table 2, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0288] Embodiment 46. The compound of Embodiment 44, wherein the compound of Table 1 is Compound No. 1, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0289] Embodiment 47. The compound of Embodiment 44, wherein the compound of Table 1 is Compound No. 3, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0290] Embodiment 48. The compound of Embodiment 44, wherein the compound of Table 1 is Compound No. 8, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0291] Embodiment 49. The compound of Embodiment 44, wherein the compound of Table 1 is Compound No. 9, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0292] Embodiment 50. The compound of Embodiment 44, wherein the compound of Table 1 is Compound No. 4, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0293] Embodiment 51. The compound of any one of Embodiments 41-43, comprising a compound of Table 3, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0294] Embodiment 52. The compound or pharmaceutical composition for use as described in any one of Embodiments 41 or 44-51 for treating a disease, disorder or condition in a subject.

[0295] Embodiment 53. The compound or pharmaceutical composition for use as described in any one of Embodiments 41 or 44-51 for preventing a disease, disorder or condition in a subject.

[0296] Embodiment 54. The compound or pharmaceutical composition for use as described in any one of Embodiments 41 or 44-51 for treating a symptom of a disease, disorder or condition in a subject.

[0297] Embodiment 55. The compound or pharmaceutical composition for use as described in any one of Embodiments 41 or 44-51 for preventing symptoms of a disease, disorder or condition in a subject.

[0298] Embodiment 56. The compound or pharmaceutical composition for use as described in any one of Embodiments 41-55, wherein the disease, disorder or condition is a neurodegenerative disease.

[0299] Embodiment 57. The compound or pharmaceutical composition for the use described in Embodiment 56, wherein the neurodegenerative disease is Alzheimer's disease, amyotrophic lateral sclerosis, Parkinson's disease, dementia with Lewy bodies, multiple system atrophy, Huntington's disease, frontotemporal lobar degeneration, mild cognitive impairment, corticobasal degeneration, progressive supranuclear palsy, Rett syndrome, peripheral degenerative disease or Aicardi-Goutières syndrome.

[0300] Embodiment 58. The compound or pharmaceutical composition for use as described in any one of Embodiments 41-55, wherein the disease, disorder or condition is an autoimmune disease.

[0301] Embodiment 59. The compound or pharmaceutical composition for use according to Embodiment 58, wherein the autoimmune disease is lupus, rheumatoid arthritis, Sjögren's syndrome, or multiple sclerosis.

[0302] Embodiment 60. The compound or pharmaceutical composition for use according to any one of Embodiments 41-55, wherein the disease, disorder or condition is an age-related disease.

[0303] Embodiment 61. The compound or pharmaceutical composition for use according to Embodiment 60, wherein the age-related disease is Alzheimer's disease, Parkinson's disease, atherosclerosis, osteoarthritis, osteoporosis, rheumatoid arthritis, macular degeneration, peripheral degenerative disease or skin aging.

[0304] Embodiment 62. The compound or pharmaceutical composition for use as described in any one of Embodiments 41-55, wherein the disease, disorder or condition is autism spectrum disorder (ASD), cardiovascular dysfunction, hearing loss, hematopoietic stem cell function, pulmonary fibrosis, schizophrenia or vision loss.

[0305] Embodiment 63. The compound or pharmaceutical composition for use as described in any one of Embodiments 41-55, wherein the disease, disorder or condition is progressive supranuclear palsy.

[0306] Embodiment 64. The compound or pharmaceutical composition for use as described in any one of Embodiments 41-55, wherein the disease, disorder or condition is amyotrophic lateral sclerosis.

[0307] Embodiment 65. The compound or pharmaceutical composition for use according to any one of Embodiments 41-55, wherein the disease, disorder or condition is Aicardi-Goutières syndrome.

[0308] Embodiment 66. The compound or pharmaceutical composition for use according to any one of Embodiments 41-55, wherein one or more optional therapeutic agents are to be administered to the subject.

[0309] Embodiment 67. The compound or pharmaceutical composition for use according to any one of Embodiments 41-66, wherein the subject (a) is not infected with HIV virus; (b) is not suspected of being infected with HIV virus; (c) is not receiving HIV virus treatment; and / or (d) is not receiving HIV virus prevention treatment.

[0310] Embodiment 68. The compound or pharmaceutical composition for use according to any one of Embodiments 41-67, wherein the compound inhibits human LINE-1 retrotransposition activity at a half-maximal inhibitory concentration of less than 1 μM in an in vitro HeLa cell-based dual luciferase assay.

[0311] Embodiment 69. The compound or pharmaceutical composition for use according to Embodiment 68, wherein the compound inhibits human LINE-1 retrotransposition activity at a half-maximal inhibitory concentration of less than 0.25 μM in an in vitro HeLa cell-based dual-luciferase assay.

[0312] Embodiment 70. The compound for use according to any one of Embodiments 41-69.

[0313] Embodiment 71. The pharmaceutical composition for use according to any one of Embodiments 41-69.

[0314] Embodiment 82. Use of the following compounds:

[0315] (i) Table 1, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof;

[0316] (ii) Table 2, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof, or a pharmaceutical composition thereof; or

[0317] (iii) Table 3, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof, or a pharmaceutical composition thereof,

[0318] For the preparation of a medicament for treating or preventing a disease, disorder or condition caused by a pathophysiological retrotransposon-associated process and / or treating or preventing a symptom of a disease, disorder or condition in a subject in need thereof.

[0319] Embodiment 83. Use of the following compounds:

[0320] (i) Table 1, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof;

[0321] (ii) Table 2, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof, or a pharmaceutical composition thereof; or

[0322] (iii) Table 3, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof, or a pharmaceutical composition thereof,

[0323] For the preparation of a medicament for inhibiting a LINE-1 retrotransposition event causing a disease, disorder or condition in a subject in need thereof.

[0324] Embodiment 84. Use of the following compounds:

[0325] (i) Table 1, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof;

[0326] (ii) Table 2, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof, or a pharmaceutical composition thereof; or

[0327] (iii) Table 3, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof, or a pharmaceutical composition thereof,

[0328] For use in the preparation of a medicament for treating a disease, condition or disorder characterized by overexpression of retrotransposon RNA, retrotransposon reverse transcriptase or retrotransposon DNA.

[0329] Embodiment 85. The use of any one of Embodiments 82-84, comprising a compound of Table 1, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0330] Embodiment 86. The use of Embodiment 85, comprising a compound of Table 2, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0331] Embodiment 87. The use described in Embodiment 86, wherein the compound of Table 1 is Compound No. 1, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0332] Embodiment 88. The use described in Embodiment 87, wherein the compound of Table 1 is Compound No. 3, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0333] Embodiment 89. The use described in Embodiment 88, wherein the compound of Table 1 is Compound No. 8, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0334] Embodiment 90. The use described in Embodiment 89, wherein the compound of Table 1 is Compound No. 9, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0335] Embodiment 91. The use described in Embodiment 90, wherein the compound of Table 1 is Compound No. 4, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0336] Embodiment 92. The use of any one of Embodiments 82-84, comprising a compound of Table 3, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

[0337] Embodiment 93. The use of any one of Embodiments 82 or 85-92, for treating a disease, disorder or condition in a subject.

[0338] Embodiment 94. The use of any one of Embodiments 82 or 85-92, for preventing a disease, disorder or condition in a subject.

[0339] Embodiment 95. The use of any one of Embodiments 82 or 85-92, for treating a symptom of a disease, disorder or condition in a subject.

[0340] Embodiment 96. The use of any one of Embodiments 82 or 85-92, for preventing symptoms of a disease, disorder or condition in a subject.

[0341] Embodiment 97. The use of any one of Embodiments 82-96, wherein the disease, disorder or condition is a neurodegenerative disease.

[0342] Embodiment 98. The use of embodiment 97, wherein the neurodegenerative disease is Alzheimer's disease, amyotrophic lateral sclerosis, Parkinson's disease, dementia with Lewy bodies, multiple system atrophy, Huntington's disease, frontotemporal lobar degeneration, mild cognitive impairment, corticobasal degeneration, progressive supranuclear palsy, Rett syndrome, peripheral degenerative disease or Aicardi-Goutières syndrome.

[0343] Embodiment 99. The use of any one of Embodiments 82-96, wherein the disease, disorder or condition is an autoimmune disease.

[0344] Embodiment 100. The use of embodiment 99, wherein the autoimmune disease is lupus, rheumatoid arthritis, Sjögren's syndrome, or multiple sclerosis.

[0345] Embodiment 101. The use of any one of Embodiments 82-96, wherein the disease, disorder or condition is an age-related disease.

[0346] Embodiment 102. The use of embodiment 101, wherein the age-related disease is Alzheimer's disease, Parkinson's disease, atherosclerosis, osteoarthritis, osteoporosis, rheumatoid arthritis, macular degeneration, peripheral degenerative disease or skin aging.

[0347] Embodiment 103. The use of any one of embodiments 82-96, wherein the disease, disorder or condition is autism spectrum disorder (ASD), cardiovascular dysfunction, hearing loss, hematopoietic stem cell function, pulmonary fibrosis, schizophrenia, or vision loss.

[0348] Embodiment 104. The use of any one of Embodiments 82-96, wherein the disease, disorder or condition is progressive supranuclear palsy.

[0349] Embodiment 105. The use of any one of Embodiments 82-96, wherein the disease, disorder or condition is amyotrophic lateral sclerosis.

[0350] Embodiment 106. The use of any one of Embodiments 82-96, wherein the disease, disorder or condition is Aicardi-Goutières syndrome.

[0351] Embodiment 107. The use of any one of Embodiments 82-96, wherein one or more optional therapeutic agents are administered to the subject.

[0352] Embodiment 108. The use of any one of Embodiments 82-107, wherein the subject (a) is not infected with HIV virus; (b) is not suspected of being infected with HIV virus; (c) is not receiving HIV virus treatment; and / or (d) is not receiving HIV virus prevention treatment.

[0353] Embodiment 109. The use of any one of Embodiments 82-108, wherein the compound inhibits human LINE-1 retrotransposition activity at a half-maximal inhibitory concentration of less than 1 μM in an in vitro HeLa cell-based dual-luciferase assay.

[0354] Embodiment 110. The method of any one of embodiments 3-6, wherein the retrotransposon RNA is LINE-1 RNA.

[0355] Embodiment 111. The method of any one of embodiments 3-6, wherein the retrotransposon reverse transcriptase is ORF2p.

[0356] Embodiment 112. The method of any one of embodiments 3-6, wherein the retrotransposon DNA is LINE-1 DNA.

[0357] Embodiment 113. The compound for use according to embodiment 43, wherein the retrotransposon RNA is LINE-1 RNA.

[0358] Embodiment 114. The compound for use according to embodiment 43, wherein the retrotransposon reverse transcriptase is ORF2p.

[0359] Embodiment 115. The compound for use according to embodiment 43, wherein the retrotransposon DNA is LINE-1 DNA.

[0360] Embodiment 116. The use of embodiment 84, wherein the retrotransposon RNA is LINE-1 RNA.

[0361] Embodiment 117. The use of embodiment 84, wherein the retrotransposon reverse transcriptase is ORF2p.

[0362] Embodiment 118. The use of embodiment 84, wherein the retrotransposon DNA is LINE-1 DNA.

[0363] Embodiment 119. The method of any one of embodiments 1-18, wherein the disease, disorder or condition is ataxia-telangiectasia.

[0364] Embodiment 120. The method of any one of embodiments 1-18, wherein the disease, disorder or condition is age-related macular degeneration.

[0365] Embodiment 121. The method of any one of embodiments 1-18, wherein the disease, disorder or condition is systemic lupus erythematosus.

[0366] Embodiment 122. The method of any one of embodiments 1-18, wherein the disease, disorder or condition is an IFN-related autoimmune disease.

[0367] Embodiment 123. The method of embodiment 122, wherein the IFN-related autoimmune disease is psoriasis.

[0368] Embodiment 124. The method of any one of embodiments 1-18, wherein the disease, disorder or condition is Fanconi anemia.

[0369] Embodiment 125. The method of any one of Embodiments 1-18, wherein the disease, disorder or condition is idiopathic pulmonary fibrosis.

[0370] Embodiment 126. The method of any one of embodiments 1-18, wherein the disease, disorder or condition is cardiovascular disease.

[0371] Embodiment 127. The compound for use according to any one of Embodiments 41-55, wherein the disease, disorder or condition is ataxia-telangiectasia.

[0372] Embodiment 128. The compound for use as described in any one of Embodiments 41-55, wherein the disease, disorder or condition is age-related macular degeneration.

[0373] Embodiment 129. The compound for use as described in any one of Embodiments 41-55, wherein the disease, disorder or condition is systemic lupus erythematosus.

[0374] Embodiment 130. The compound for use as described in any one of Embodiments 41-55, wherein the disease, disorder or condition is an IFN-related autoimmune disease.

[0375] Embodiment 131. The compound for use according to embodiment 130, wherein the IFN-related autoimmune disease is psoriasis.

[0376] Embodiment 132. The compound for use according to any one of Embodiments 41-55, wherein the disease, disorder or condition is Fanconi anemia.

[0377] Embodiment 133. The compound for use as described in any one of Embodiments 41-55, wherein the disease, disorder or condition is idiopathic pulmonary fibrosis.

[0378] Embodiment 134. The compound for use as described in any one of Embodiments 41-55, wherein the disease, disorder or condition is cardiovascular disease.

[0379] Embodiment 135. The use of any one of Embodiments 82-96, wherein the disease, disorder or condition is ataxia-telangiectasia.

[0380] Embodiment 136. The use of any one of Embodiments 82-96, wherein the disease, disorder or condition is age-related macular degeneration.

[0381] Embodiment 137. The compound for use as described in any one of Embodiments 82-96, wherein the disease, disorder or condition is systemic lupus erythematosus.

[0382] Embodiment 138. The compound for use as described in any one of Embodiments 82-96, wherein the disease, disorder or condition is an IFN-related autoimmune disease.

[0383] Embodiment 139. The compound for use according to embodiment 138, wherein the IFN-related autoimmune disease is psoriasis.

[0384] Embodiment 140. The compound for use according to any one of Embodiments 82-96, wherein the disease, disorder or condition is Fanconi anemia.

[0385] Embodiment 141. The compound for use as described in any one of Embodiments 82-96, wherein the disease, disorder or condition is idiopathic pulmonary fibrosis.

[0386] Embodiment 142. The compound for use as described in any one of Embodiments 82-96, wherein the disease, disorder or condition is cardiovascular disease.

[0387] Embodiment 143. The method, kit, compound for use, or use of any one of embodiments 1, 32, 41, or 82, wherein the disease, disorder, or condition is not (i) cancer; or (ii) an infectious disease.

[0388] Example

[0389] The compounds of Tables 1 and 2 can be prepared as described in the following examples and, for example, Nomura et al., J. Med. Chem. 42:2901-2908 (1999); Ohrui et al., J. Med. Chem. 43:4516-4525 (2000), Japanese Patent No. 6767011 and / or U.S. Patent No. 10,933,067.

[0390] The abbreviations in Table 4 may be used in the examples.

[0391] Table 4

[0392]

[0393]

[0394]

[0395] The following LC-MS method can be used in the examples.

[0396] Method A: UPLC-MS method: Waters Acquity UPLC CSH C18, 1.8 μm, 2.1 x 30 mm, at 40 °C; 5% to 100% B in 5.2 minutes; hold 100% B for 1.8 minutes, run time = 7.0 min, flow rate 0.9 mL / min; eluent: A = Milli-Q H 2 O + 10 mM ammonium formate pH = 3.8; B = MeCN. Waters Acquity UPLC system. UV detector = Waters Acquity PDA, 198-360 nm. MS detector = Waters SQD ESI.

[0397] Method B: Waters Acquity CSH C18, 3.5 μm, 4.6 x 30 mm, at 40 °C; Iso 5% B for 0.5 min, 5% to 100% B in 5 min; hold 100% B for 1.5 min, run time = 7.0 min, flow rate 0.9 mL / min; eluent: A = Milli-Q H 2 O + 10 mM ammonium formate pH = 3.8; B = MeCN. Waters Alliance 2695 system. UV detection: Waters 2996 PDA, 198-360 nm. MS detector: Waters ZQ 2000, ESI

[0398] Method C: SHIMADZU LC20-MS2010: MERCK, RP-18e 25-2 mm, at 50°C; 1.5 ML / 4 L TFA in water (solvent A), and 0.75 ML / 4 L TFA in acetonitrile (solvent B), using an elution gradient of 5%-95% (solvent B) in 0.7 minutes, and maintaining at 95% for 0.4 minutes, flow rate 1.5 ml / min, run time = 1.5 min; UV detector = 220 nm, 254 nm; MS detector = ESI.

[0399] Method D: LCMS-BT: SHIMADZU LC20-MS2020: X bridge Shield RP-18, 5 μm, 2.1*50 mm, at 50°C; 0.8 mL / 4 L NH 3 ·H 2 O in water (solvent A) and acetonitrile (solvent B), using an elution gradient of 0%-30% (solvent B) in 2 minutes and holding at 30% for 0.48 minutes, flow rate 1 ml / min, run time = 3 min; UV detector = 220 nm, 254 nm; MS detector = ESI

[0400] Method E: LCMS-AN: Agilent LC1200-MS6110: Xtimate C18 2.1*30mm, 3μm, 50°C; 1.5ML / 4L TFA in water (solvent A) and 0.75ML / 4L TFA in acetonitrile (solvent B), using an elution gradient of 0%-60% (solvent B) in 0.9min and holding at 60% for 0.6min, flow rate 1.2ml / min, run time = 2min; UV detector = 220nm; MS detector = ESI.

[0401] Method F: LCMS-CI: Agilent LC1200-MS6110: Xbridge Shield RP-18, 5um, 2.1*50mm; at 30°C; water (4L) + NH 3 ·H 2 O (0.8 mL) (solvent A) and acetonitrile (solvent B), using an elution gradient of 0%-60% (solvent B) in 2 min and holding at 60% for 0.48 min, flow rate 1 mL / min, run time = 3.0 min; UV detector = 220 nm, 254 nm; MS detector = ESI.

[0402] Method G: UPLC-MS method: Waters Acquity UPLC CSH C18, 1.8 μm, 2.1 x 30 mm, at 40 °C; 5%-100% B in 2.0 min; 100% B hold 0.7 min, run time = 2.7 min, flow rate 0.9 mL / min; eluent: A = Milli-Q H 2 O + 10 mM ammonium formate pH = 3.8; B = MeCN. Waters Acquity UPLC system. UV detector = Waters Acquity PDA, 198-360 nm. MS detector = Waters SQD ESI.

[0403] Method H: UPLC-MS method: Waters Acquity UPLC Agilent Poroshell 120EC C18, 1.8 μm, 2.1 x 250 mm, at 40 °C; 5% to 100% B in 5.0 min; hold 100% B for 2.0 min, run time = 7.0 min, flow rate 0.5 mL / min; eluent: A = Milli-Q H 2 O + 10 mM ammonium formate pH = 3.8; B = MeCN. Waters Acquity UPLC system. UV detector = Waters Acquity PDA, 198-360 nm. MS detector = Waters SQDESI.

[0404] Example 1

[0405] Synthesis of 1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-vinyltetrahydrofuran-2-yl)-5-methylpyrimidine-2,4(1H,3H)-dione (Compound No. 62)

[0406]

[0407] 1-((2R,4S,5R)-5-ethynyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-5-methylpyrimidine-2,4(1H,3H)-dione was prepared according to WO 2007 / 038507 A2.

[0408] 1-((2R,4S,5R)-5-ethynyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-5-methylpyrimidine-2,4(1H,3H)-dione (18.0 mg, 67.6 μmol) was dissolved in 9:1 EtOAc / pyridine (653 μl / 22.5 μl) in a 5 mL vial with a rubber septum. Solid Lindlar catalyst (18.0 mg, 67.6 μmol) was then added and the reaction mixture was purged with bubbling H using a balloon. 2 The reaction mixture was stirred for 30 min until complete conversion to the title compound was observed by LC-MS. Filter and rinse with EtOAc. The desired product was purified by flash chromatography on silica gel (EtOAc) to yield 1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-vinyltetrahydrofuran-2-yl)-5-methylpyrimidine-2,4(1H,3H)-dione (11 mg, 58%). 12 H 16 N 2 O 5 LC-MS (ESI) m / z calculated value: 268.11, found value: 267.3 [MH] - . 1 H NMR: (400MHz, CD 3 OD)9.01(s,1H),7.23(t,J=5.7Hz,1H),6.99(dd,J=17.3,10.9Hz,1H).6.51(dd,J=17.3,2.0Hz,1H),6.30(dd,J= 10.9, 1.2Hz, 1H), 5.63 (t, J = 7.4Hz, 1H), 4.65 (qAB, J = 11.9, 5.0Hz, 2H), 3.30 (dd, J = 7.4, 5.8Hz, 2H), 2.93 (s, 3H).

[0409] Example 2

[0410] Synthesis of 5-bromo-1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-vinyltetrahydrofuran-2-yl)pyrimidine-2,4(1H,3H)-dione (Compound No. 64)

[0411]

[0412] 5-Bromo-1-((2R,4S,5R)-5-ethynyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)pyrimidine-2,4(1H,3H)-dione was prepared according to WO 2007 / 038507 A2.

[0413] 5-Bromo-1-((2R,4S,5R)-5-ethynyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)pyrimidine-2,4(1H,3H)-dione (3.3 mg, 9.97 μmol) was dissolved in 9:1 EtOAc / pyridine (96.3 μl / 3.32 μl) in a 5 mL vial with a rubber septum. Solid Lindela catalyst (3.30 mg, 9.97 μmol) was then added and the reaction mixture was purged with bubbling H using a balloon. 2 The reaction mixture was stirred for 30 min until complete conversion to the title compound was observed by LC-MS. Filter and rinse with EtOAc. The desired product was purified by flash chromatography on silica gel (EtOAc) to yield 5-bromo-1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-vinyltetrahydrofuran-2-yl)pyrimidine-2,4(1H,3H)-dione (2.70 mg, 81%). 11 H 13 Bn 2 O 5 LC-MS (ESI) m / z calculated value: 332.00, found value: 331.2 [MH] - . 1 H NMR (400MHz, CD 3 OD)δ8.67(s,1H),6.12(t,J=5.3Hz,1H),5.92(dd,J=17.3,10.9Hz,1H),5.46(dd,J=17.3,1.9Hz,1H),5.28(dd,J=1 0.9, 1.9Hz, 1H), 4.55 (t, J = 7.6Hz, 1H), 3.69 (d, J = 12.0Hz, 1H), 3.53 (d, J = 12.0Hz, 1H), 2.27 (dd, J = 7.7, 5.5Hz, 2H).

[0414] Example 3

[0415] Synthesis of 1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-vinyltetrahydrofuran-2-yl)-5-methoxypyrimidine-2,4(1H,3H)-dione (Compound No. 6)

[0416]

[0417] Step 1. Preparation of 1-((2R,4S,5R)-5-ethynyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-5-methoxypyrimidine-2,4(1H,3H)-dione according to WO 2007 / 038507 A2.

[0418] At 25 °C in H 2 To a mixture of 4-methylbenzoic acid [(2R, 3S, 5R) -2-ethynyl-5- (5-methoxy-2,4-dioxo-pyrimidin-1-yl) -3- (4-methylbenzoyl) oxy-tetrahydrofuran-2-yl] methyl ester (15 mg, 28.93 μmol) in MeOH (2 mL) was added Lindela catalyst (7 mg) in one portion under atmosphere (15 psi). The resulting mixture was stirred at 25 ° C for 1 hour. The mixture was then filtered and the filtrate was concentrated to provide a white solid compound (15 mg, 99%).

[0419] Step 2. To a mixture of 4-methylbenzoic acid [(2R,3S,5R)-5-(5-methoxy-2,4-dioxo-pyrimidin-1-yl)-3-(4-methylbenzoyl)oxy-2-vinyl-tetrahydrofuran-2-yl]methyl ester (15 mg, 28.82 μmol) in MeOH (5 mL) was added NaOMe (156 μg, 2.88 μmol) in one portion at 25°C. After stirring at 25°C for 1 hour, the mixture was concentrated under reduced pressure. The residue was purified by preparative HPLC (basic) to provide the title compound (5 mg, 62%) as a white solid. LCMS (ESI): m / z 307.0 (M+Na) + . 1 H NMR (400 MHz, DMSO-d 6 )δ11.28(s,1H),7.77(s,1H),6.17-6.07(m,1H),5.97-5.85(m,1H),5.44(t,J=4.8Hz,1H),5.39-5.31(m,1H),5.28(d,J=5.2Hz,1H ),5.23-5.15(m,1H),4.54-4.46(m,1H),3.65-3.55(m,1H),3.59(s,3H),3.43-3.36(m,1H),2.24-2.15(m,1H),2.13-2.04(m,1H).

[0420] Example 4

[0421] Synthesis of 1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-vinyltetrahydrofuran-2-yl)-2,4-dioxo-1,2,3,4-tetrahydropyrimidine-5-carbonitrile (Compound No. 69)

[0422]

[0423] 1-((2R,4S,5R)-5-ethynyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-2,4-dioxo-1,2,3,4-tetrahydropyrimidine-5-carbonitrile was prepared according to WO 2007 / 038507 A2 using 5-pyrimidinecarbonitrile. 1-((2R,4S,5R)-5-ethynyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-2,4-dioxo-1,2,3,4-tetrahydropyrimidine-5-carbonitrile (8.9 mg, 32.1 μmol) was dissolved in MeOH (321 μl) in a 5 mL vial with a rubber septum. Solid Lindela catalyst (8.9 mg, 32.1 μmol) was then added and the reaction mixture was purged with a balloon through bubbling H 2 The reaction mixture was stirred for 30 min until complete conversion to the title compound was observed by LC-MS. Filter and rinse with EtOAc. The desired product was purified by flash chromatography on silica gel (EtOAc) to yield 1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-vinyltetrahydrofuran-2-yl)-2,4-dioxo-1,2,3,4-tetrahydropyrimidine-5-carbonitrile (1.36 mg, 13%). 12 H 13 N 3 O 5 LC-MS (ESI) m / z calculated value: 279.1, found value 278.1 [MH] - . 1 H NMR (400MHz, CD 3 OD)δ9.13(d,J=4.5Hz,1H),6.21-5.98(m,1H),5.92(dd,J=17.3,11.0Hz,1H),5.46(dd,J=17.3,1.9Hz,1H),5.29 (dd,J=11.0,1.9Hz,1H),4.60-4.45(m,1H),3.72(dd,J=11.9,6.9Hz,1H),3.65-3.54(m,1H),2.48-2.22(m,2H).

[0424] Example 5

[0425] Synthesis of 4-amino-5-bromo-1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-vinyltetrahydrofuran-2-yl)pyrimidin-2(1H)-one (Compound No. 90)

[0426]

[0427] 4-Amino-5-bromo-1-((2R,4S,5R)-5-ethynyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)pyrimidin-2(1H)-one was prepared according to WO 2007 / 038507 A2.

[0428] 4-Amino-5-bromo-1-((2R,4S,5R)-5-ethynyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)pyrimidin-2(1H)-one (64.0 mg, 194 μmol) was dissolved in MeOH (1.94 mL) in a 5 mL vial with a rubber septum. Solid Lindela catalyst (64 mg, 194 μmol) was then added and the reaction mixture was purged with a balloon via bubbling H 2 The reaction mixture was stirred for 30 min until complete conversion to the title compound was observed by LC-MS. Filter and wash with MeOH. The desired product was purified by flash chromatography on silica gel using 0-100% 2-propanol in EtOAc to yield 4-amino-5-bromo-1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-vinyltetrahydrofuran-2-yl)pyrimidin-2(1H)-one (31.6 mg, 45%). 11 H 14 Bn 3 O 4 LC-MS (ESI) m / z calculated value: 331.02, found value 334.0 [M+3H] + . 1 H NMR (400MHz, CD 3 OD)δ8.70(s,1H),6.06(dd,J=6.9,3.0Hz,1H),6.02-5.82(m,1H),5.46(dd,J=17.3,1.8Hz,1H),5.36-5.19(m,1H),4.59-4.43(m,1 H),4.44-4.36(m,1H),3.70(dd,J=12.0,5.7Hz,1H),3.55(d,J=12.0Hz,1H),2.38-2.24(m,1H),2.18(ddd,J=13.4,7.1,3.0Hz,1H).

[0429] Example 6

[0430] Synthesis of 2-amino-9-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-vinyltetrahydrofuran-2-yl)-1H-purin-6(9H)-one (Compound No. 9)

[0431]

[0432] 2-Amino-9-((2R,4S,5R)-5-ethynyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-1H-purin-6(9H)-one was prepared according to WO 2007 / 038507 A2.

[0433] 2-Amino-9-((2R,4S,5R)-5-ethynyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-1H-purin-6(9H)-one (125 mg, 429 μmol) was dissolved in EtOAc / pyridine (4.15 mL / 143 μl) in a 5 mL vial with a rubber septum. Solid Lindela catalyst (125 mg, 429 μmol) was then added and the reaction mixture was purified by bubbling H with a balloon. 2 The reaction mixture was stirred for 30 min until complete conversion to the title compound was observed by LC-MS. Filter and rinse with MeOH. SFC was performed using IC, 10x250mm5um column and 25% MeOH, 0.1% NH 4 OH, 75% CO 2 The desired product was purified by an isocratic gradient of 10 mL / min at a flow rate of 10 mL / min and a run time of 20 min to yield 2-amino-9-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-vinyltetrahydrofuran-2-yl)-1H-purin-6(9H)-one (2.62 mg, 2%). 12 H 15 N 5 O 4 LC-MS (ESI) m / z calculated value: 293.1, found value 292.2 [MH] - . 1 HNMR (400MHz, CD 3OD)δ8.17(d,J=99.1Hz,1H),6.28(dd,J=7.9,3.6Hz,1H),6.04(ddd,J=17.3,13.6,11.1Hz,1H),5.37 -5.08(m,2H),4.47(dd,J=6.5,2.9Hz,1H),3.58-3.37(m,2H),3.04-2.84(m,1H),2.37-2.23(m,1H).

[0434] Example 7

[0435] Synthesis of 4-amino-5-bromo-1-((4S,5R)-5-ethyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)pyrimidin-2(1H)-one (Compound No. 70)

[0436]

[0437] 4-Amino-5-bromo-1-((4S,5R)-5-ethynyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)pyrimidin-2(1H)-one was synthesized according to the procedure described in WO 2007 / 038507 A2.

[0438] 4-Amino-5-bromo-1-((4S,5R)-5-ethynyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)pyrimidin-2(1H)-one (5.45 mg, 16.4 μmol) was dissolved in MeOH in a 5 mL vial with a rubber septum. Solid PtO was then added. 2 (0.500 mg, 2.20 μmol), the reaction mixture was treated with H 2 The reaction mixture was stirred for 30 min with a balloon purge. The reaction mixture was stirred until complete conversion to the title compound was observed by LC-MS. Filter and wash with MeOH, remove the solvent under reduced pressure. Purify the crude mixture by column chromatography on silica gel (EtOAc / hep 1:1 to 100% EtOAc) to give 4-amino-5-bromo-1-((4S,5R)-5-ethyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)pyrimidin-2(1H)-one (0.750 mg, 13%) as an off-white powder mixture of anomers. 11 H 15 Bn 3 O 4 LC-MS (ESI) m / z calculated value: 332.03, found value 332.2 [MH] - . 1 H NMR (400MHz, CD 3OD)δ8.55(s,1H),6.11-6.03(m,1H),4.47-4.34(m,1H),4.12-4.00(m,1H),3.70(d,J=11.7Hz,1H),3.61-3.44(m,1H),2.43(ddd,J=26.1, 18.8, 13.1Hz, 1H), 2.23 (ddd, J=13.8, 6.9, 5.5Hz, 1H), 1.86 (dd, J=15.3, 7.5Hz, 1H), 1.68 (t, J=7.6Hz, 1H), 0.96 (tt, J=22.6, 11.3Hz, 3H).

[0439] Example 8

[0440] Synthesis of 1-((4S,5R)-5-ethyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-5-methoxypyrimidine-2,4(1H,3H)-dione (Compound No. 68)

[0441]

[0442] 1-((4S,5R)-5-ethynyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-5-methoxypyrimidine-2,4(1H,3H)-dione was prepared according to the procedure described in WO 2007 / 038507 A2.

[0443] 1-((4S,5R)-5-ethynyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-5-methoxypyrimidine-2,4(1H,3H)-dione (7.20 mg, 28.7 μmol) was dissolved in MeOH in a 5 mL vial with a rubber septum. Solid Lindela catalyst (7.20 mg, 28.7 μmol) was then added and the reaction mixture was heated with H 2 The reaction mixture was stirred for 30 min until complete conversion to the title compound was observed by LC-MS. Filtration and washing with MeOH and removal of the solvent under reduced pressure provided 4-amino-1-((4S,5R)-5-ethyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-5-methoxypyrimidin-2(1H)-one (5.75 mg, 66%) as a mixture of white powder anomers. 12 H1 8 N 3 O 5 LC-MS (ESI) m / z calculated value: 284.13, found value 284.4 [MH] - . 1 H NMR (400MHz, CD3 OD)δ7.82(d,J=22.7Hz,1H),6.38-6.07(m,1H),4.55-4.10(m,1H),3.64-3.37(m,2H),3.33(s,3H),2.33(td,J =7.0,6.0Hz,1H),1.95-1.75(m,1H),1.62(dd,J=41.8,7.6Hz,1H),1.54(m,1H),0.96(dt,J=11.9,7.6Hz,3H).

[0444] Example 9

[0445] Synthesis of (2R,3S,5R)-5-(6-(cyclopropylamino)-9H-purin-9-yl)-2-(hydroxymethyl)-2-vinyltetrahydrofuran-3-ol (Compound No. 94)

[0446]

[0447] ((2R,3S,5R)-5-(6-chloro-9H-purin-9-yl)-2-ethynyl-2-(((4-methylbenzoyl)oxy)methyl)tetrahydrofuran-3-yl 4-methylbenzoate was synthesized using N-cyclopropyl-9H-purin-6-amine according to the procedure described in WO 2007 / 038507 A2.

[0448] Step 1. At 25°C in N 2 To a mixture of 4-methylbenzoic acid [(2R, 3S, 5R) -5- (6-chloropurin-9-yl) -2-ethynyl -3- (4-methylbenzoyl) oxy-tetrahydrofuran-2-yl] methyl ester (0.05 g, 94.17 μmol) and cyclopropylamine (8 mg, 141.25 μmol) in t-BuOH (2 mL) was added DIEA (24 mg, 188.34 μmol) in one portion. The resulting mixture was stirred at 90 ° C for 16 hours. After cooling to room temperature, the mixture was concentrated under reduced pressure. The residue was purified by preparative TLC (50% EtOAc in petroleum ether) to provide a colorless oily compound (45 mg, 87%). 1 H NMR (400 MHz, CDCl 3)δ8.66(s,1H),8.24(s,1H),8.06-8.00(m,2H),7.87-7.80(m,2H),7.30(d,J=8.0Hz,2H),7.19(d,J=8.0Hz,2H),6.65-6.55(m,1H),6.21 -6.13(m,1H),4.91(d,J=12.0Hz,1H),4.63(d,J=12.0Hz,1H),3.47-3.30(m,1H),3.00-2.90(m,1H),2.74(s,1H),2.44(d,J=16.0Hz,6H).

[0449] Step 2. Heat the mixture at 25°C in H 2 To a solution of [(2R,3S,5R)-5-[6-(cyclopropylamino)purin-9-yl]-2-ethynyl-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl 4-methylbenzoate (45 mg, 81.58 μmol) in MeOH (2 mL) was added Lindela catalyst (17 mg) in one portion under (15 psi). The resulting mixture was stirred at 25 °C for 1 hour. The mixture was directly filtered and concentrated to provide the compound as a white solid (45 mg, 99%).

[0450] Step 3. To a mixture of [(2R,3S,5R)-5-[6-(cyclopropylamino)purin-9-yl]-3-(4-methylbenzoyl)oxy-2-vinyl-tetrahydrofuran-2-yl]methyl 4-methylbenzoate (45 mg, 81.29 μmol) in MeOH (5 mL) was added NaOMe (439 μg, 8.13 μmol) in one portion at 25°C. After stirring for 1 hour at 25°C, the mixture was concentrated under reduced pressure. The residue was purified by preparative HPLC (acetonitrile 0-30% / 0.225% formic acid in water) to provide the title compound (13.6 mg, 53%) as a white solid. 1 H NMR (400MHz, DMSO-d6) δ8.37(s,1H),8.24(s,1H),7.96(s,1H),6.40-6.30(m,1H),6.03-5.91(m,1H),5.44-5.32(m,2H),5.28(d,J=5.2Hz, 1H),5.23-5.16(m,1H),4.68-4.61(m,1H),3.56-3.41(m,2H),2.66-2 .61(m,1H),2.31-2.21(m,1H),0.76-0.68(m,2H),0.65-0.55(m,2H).

[0451] Example 10

[0452] Synthesis of (2R,3S,5R)-5-(6-(cyclopropylamino)-2-fluoro-9H-purin-9-yl)-2-(hydroxymethyl)-2-vinyltetrahydrofuran-3-ol (Compound No. 5)

[0453]

[0454] 4-Methylbenzoic acid [(2R,3S,5R)-5-[6-(cyclopropylamino)-2-fluoro-purin-9-yl]-2-ethynyl-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester was synthesized using N-cyclopropyl-2-fluoro-9H-purin-6-amine according to the procedure described in WO 2007 / 038507 A2.

[0455] Step 1. To a solution of 4-methylbenzoic acid [(2R,3S,5R)-5-[6-(cyclopropylamino)-2-fluoro-purin-9-yl]-2-ethynyl-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester (50 mg, 87.78 μmol) in EtOAc (3 mL) was added Lindela catalyst (18 mg, 8.78 μmol, 10% purity) in one portion at 25°C. The mixture was treated with H 2 Degas 3 times and incubate at 25 °C in 5% HO 2 (15 psi) for 1 h. After this time, the reaction mixture was filtered and the filtrate was concentrated to provide the compound as a white solid (46 mg, 92%). LCMS (ESI): m / z 594.1 (M+Na)+.

[0456] Step 2. To a solution of 4-methylbenzoic acid [(2R,3S,5R)-5-[6-(cyclopropylamino)-2-fluoro-purin-9-yl]-3-(4-methylbenzoyl)oxy-2-vinyl-tetrahydrofuran-2-yl]methyl ester (45 mg, 78.73 μmol) in MeOH (3 mL) was added NH 3 ·H 2 O (1 mL, 6.49 mmol, 25% purity). The mixture was stirred at 40 °C for 16 h. After this time, the reaction mixture was concentrated. The residue was purified by reverse phase HPLC (acetonitrile 1-27% / 0.225% formic acid in water) to give the title compound (14.1 mg, 53%) as a white solid. 1H NMR (400MHz, DMSO-d6) δ8.54(br s,1H),8.37(s,1H),6.28-6.20(m,1H),6.00-5.90(m,1H),5.42-5.36(m,1H),5.30(br d,J=4.8Hz,1H),5.26-5.18(m,1H),5.10(br t,J=5.6Hz,1H),4.71-4.59(m,1H),3.55-3.45(m,2H),2.93(br s,1H),2.61-2.70(m,1H),2.31-2.22(m,1H),0.79-0.60(m,4H).

[0457] Embodiment 11

[0458] Synthesis of (2R,3S,5R)-5-(2-amino-6-(cyclopropylamino)-9H-purin-9-yl)-2-(hydroxymethyl)-2-vinyltetrahydrofuran-3-ol (Compound No. 4)

[0459]

[0460] According to the procedure described in WO 2007 / 038507 A2, N 6 -Cyclopropyl-9H-purine-2,6-diamine synthesis of (2R,3S,5R)-5-(2-amino-6-(cyclopropylamino)-9H-purin-9-yl)-2-ethynyl-2-(hydroxymethyl)tetrahydrofuran-3-ol.

[0461] Step 1. At 25°C in N 2 To a mixture of 4-methylbenzoic acid [(2R, 3S) -5- (2-amino-6-chloro-purine-9-yl) -2-ethynyl -3- (4-methylbenzoyl) oxy-tetrahydrofuran-2-yl] methyl ester (0.20 g, 366.32 μmol) and cyclopropylamine (31 mg, 549.48 μmol) in t-BuOH (15 mL) was added DIEA (95 mg, 732.64 μmol) in one portion. The resulting mixture was stirred at 90 ° C for 16 hours. After cooling to room temperature, the reaction mixture was concentrated under reduced pressure. The residue was purified by preparative TLC (50% EtOAc in petroleum ether) to provide a colorless oily compound (0.04 g, 19%). 1 HNMR (400MHz, CDCl 3): δ8.05-7.97(d,J=8.4Hz,2H),7.90(d,J=8.0Hz,2H),7.59(s,1H),7.30(s,2H) ,7.21(d,J=7.6Hz,2H),6.43-6.37(m,1H),6.20-6.13(m,1H),5.04(d,J=11.6Hz, 1H),4.59(d,J=11.6Hz,1H),3.42-3.33(m,1H),3.13-2.97(m,1H),2.81-2.72(m ,1H),2.68(s,1H),2.43(d,J=16.4Hz,6H),0.96-0.85(m,2H),0.67-0.60(m,2H).

[0462] Step 2. Heat the mixture at 25°C in H 2 To a solution of [(2R,3S,5R)-5-[2-amino-6-(cyclopropylamino)purin-9-yl]-2-ethynyl-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl 4-methylbenzoate (0.02 g, 35.30 μmol) in MeOH (2 mL) was added Lindela catalyst (7 mg) in one portion under (15 psi). The resulting mixture was stirred at 25 °C for 1 hour. The mixture was then filtered and concentrated to provide the compound as a white solid (0.02 g, 99%).

[0463] Step 3. To a solution of 4-methylbenzoic acid [(2R,3S,5R)-5-[2-amino-6-(cyclopropylamino)purin-9-yl]-3-(4-methylbenzoyl)oxy-2-vinyl-tetrahydrofuran-2-yl]methyl ester (0.02 g, 35.17 μmol) in MeOH (5 mL) was added NaOMe (190 μg, 3.52 μmol) in one portion at 25°C and stirred for 18 hours. The mixture was then concentrated. The residue was purified by preparative HPLC (4-34% acetonitrile / 0.05% NH 3 ·H 2 The residue was purified by evaporation (4% by weight, 0.04% by weight, in water) to provide the title compound (14 mg, 53%) as a white solid. 1H NMR (400MHz, DMSO-d6) δ7.95 (s, 1H), 7.37 (s, 1H), 6.17 (t, J = 6.4Hz, 1H), 6.0 3-5.90(m,1H),5.81(s,2H),5.43(s,1H),5.37-5.31(m,1H),5.22(s,1H),5. 20-5.15(m,1H),4.58(t,J=6.4Hz,1H),3.55-3.47(m,1H),3.45-3.38(m,1H) ,2.57-2.52(m,1H),2.22-2.14(m,1H),0.69-0.62(m,2H),0.62-0.55(m,2H).

[0464] Example 12

[0465] Synthesis of 4-amino-5-fluoro-1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-vinyltetrahydrofuran-2-yl)pyrimidin-2(1H)-one (Compound No. 8)

[0466]

[0467] (4S,5R)-5-ethynyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-one was synthesized according to the procedure described in WO 2007 / 038507 A2.

[0468] Step 1. TBSCl (1.06 g, 7.1 mmol) was added to a solution of (4S, 5R)-5-ethynyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-one (1 g, 6.4 mmol) and imidazole (1.31 g, 19.2 mmol) in DMF (30 mL) at 20 ° C. The resulting mixture was stirred for 16 hours at 20 ° C. After this, the reaction was diluted with EtOAc (80 mL) and washed with brine (50 mLx2). Layering was performed and the organic layer was concentrated. The residue was purified by column chromatography on silica gel (20% to 30% EtOAc in petroleum ether) to provide a white solid compound (0.7 g, 40%). 1 H NMR (400 MHz, CDCl 3 )δ4.53-4.50(m,1H),3.95-3.87(m,2H),2.99-2.93(m,1H),2.83(s,1H),2.61-2.55(m,1H),2.39-2.37(m,1H),0.89(s,9H),0.10(s,6H).

[0469] Step 2. NaH (665mg, 16mmol, 60% dispersion in mineral oil) was added to a solution of (4S, 5R)-5-((tert-butyldimethylsilyl)oxy)methyl)-5-ethynyl-4 hydroxydihydrofuran-2(3H)-one (3g, 11mmol) in THF (50mL) at 0°C. After stirring at 0°C for 10min, MOMBr (2.77g, 22mmol) was added. The resulting mixture was stirred for another 3 hours at 25°C. After this, the reaction mixture was poured into water (50mL) and extracted with DCM (50mLx2). The organic layer was washed with brine (50mLx2) and concentrated. The residue was purified by column chromatography on silica gel (10% to 20% EtOAc in petroleum ether) to provide a colorless oily compound (2.1g, 60%). 1 H NMR (400 MHz, CDCl 3 )δ4.81-4.69(m,2H),4.56-4.53(m,1H),3.95-3.85(m,2H),3.42(s,3H),2. 97-2.91(m,1H),2.72(s,1H),2.68-2.62(m,1H),0.89(s,9H),0.10(s,6H).

[0470] Step 3. To a solution of (4S,5R)-5-[[tert-butyl(dimethyl)silyl]oxymethyl]-5-ethynyl-4-(methoxymethoxy)tetrahydrofuran-2-one (1.1 g, 3.5 mmol) in DCM (30 mL) was added DIBAL-H (4.2 mL, 4.2 mmol) dropwise at -70°C. The mixture was stirred for 30 min. After this, the reaction was quenched with methanol (5 mL), washed with aqueous citric acid (10 wt%, 30 mL) and brine (30 mLx2), and concentrated to yield 1.1 g of crude (4S,5R)-5-[[tert-butyl(dimethyl)silyl]oxymethyl]-5-ethynyl-4-(methoxymethoxy)tetrahydrofuran-2-ol as a yellow oil. This was dissolved in DCM (20 mL). Et was added sequentially at 0°C. 3 N (457 mg, 4.52 mmol), DMAP (42 mg) and Ac 2 O (426mg, 4.17mmol). After stirring at 25 ° C for 1 hour, the reaction mixture was diluted with MTBE (100mL), washed with citric acid aqueous solution (10% by weight, 50mL) and brine (50x2 mL), and concentrated under reduced pressure. The residue was purified by column chromatography on silica gel (10% to 30% EtOAc in petroleum ether) to provide a white solid compound (0.9g, 72%).

[0471] Step 4. A mixture of N-(5-fluoro-2-hydroxy-pyrimidin-4-yl)benzamide (100 mg, 0.43 mmol) and BTMSA (219 mg, 1.29 mmol) in MeCN (10 mL) was stirred at 70° C. for 1 hour, then cooled to 20° C., and TMSOTf (148 mg, 0.66 mmol) and a solution of [(4S,5R)-5-[[tert-butyl(dimethyl)silyl]oxymethyl]-5-ethynyl-4-(methoxymethoxy)tetrahydrofuran-2-yl]acetate (0.12 g, 0.33 mmol) in MeCN (5 mL) were added sequentially. After stirring at 20° C. for 3 hours, the reaction mixture was poured into water (50 mL), extracted with EtOAc (50 mL×2), and concentrated. The residue was purified by preparative TLC (30% EtOAc in petroleum ether) to provide the compound as a white solid (40 mg, 22%). 1 HNMR (400MHz, CDCl 3 )δ13.10(s,1H),8.30-8.19(m,3H),7.58-7.54(m,1H),7.48-7.44(m,2H),6.28-6.26(m,1H),4.75-4.71(m,2H),4.43(t,J=7.6Hz,1H),4.10 -4.07(m,1H),3.94-3.91(m,1H),3.42(s,3H),2.76-2.71(m,1H),2.67(s,1H),2.40-2.35(m,1H),0.97(s,9H),0.18(s,6H).LCMS(ESI):m / z 532.5(M+H)+.

[0472] Step 5. A mixture of N-[1-[(2R,4S,5R)-5-[[tert-butyl(dimethyl)silyl]oxymethyl]-5-ethynyl-4-(methoxymethoxy)tetrahydrofuran-2-yl]-5-fluoro-2-oxo-pyrimidin-4-yl]benzamide (10 mg, 18 μmol) and Lindela's catalyst (10 mg) in MeOH (5 mL) was heated at 20 °C under H 2 (15 psi) for 2 hours. The reaction mixture was filtered and concentrated to provide the compound as a white solid (10 mg, 99%). LCMS (ESI): m / z 534.1 (M+H) +.

[0473] Step 6. To a solution of N-[1-[(2R,4S,5R)-5-[[tert-butyl(dimethyl)silyl]oxymethyl]-4-(methoxymethoxy)-5-vinyl-tetrahydrofuran-2-yl]-5-fluoro-2-oxo-pyrimidin-4-yl]benzamide (80 mg, 0.15 mmol) in MeOH (5 mL) was added acetyl chloride (118 mg, 1.5 mmol) at 20°C. The resulting mixture was stirred at 20°C for 16 hours. After this time, the reaction mixture was filtered and concentrated. The residue was purified by preparative HPLC (FA) to provide the title compound (6.7 mg, 16%) as a white solid. 1 H NMR (400 MHz, DMSO-d 6 )δ8.32(d,J=7.6Hz,1H),7.70(s,1H),7.46(s,1H),6.00-5.97(m,1H),5.93-5.85(m,1H),5.38-5.31(m,2H),5.23-5.1 8(m,2H),4.40(t,J=7.6Hz,1H),3.60-3.56(m,1H),3.40-3.37(m,1H),2.13-2.04(m,1H).LCMS(ESI):m / z294.0(M+Na) + .

[0474] Embodiment 13

[0475] Synthesis of (2R,3S,5R)-5-(6-(cyclopropylamino)-9H-purin-9-yl)-2-ethyl-2-(hydroxymethyl)tetrahydrofuran-3-ol (Compound No. 96)

[0476]

[0477] ((2R,3S,5R)-5-(6-chloro-9H-purin-9-yl)-2-ethynyl-2-(((4-methylbenzoyl)oxy)methyl)tetrahydrofuran-3-yl 4-methylbenzoate was synthesized using N-cyclopropyl-9H-purin-6-amine according to the procedure described in WO 2007 / 038507 A2.

[0478] Step 1. At 25°C in N 2To a mixture of 4-methylbenzoic acid [(2R, 3S, 5R) -5- (6-chloropurine-9-yl) -2-ethynyl -3- (4-methylbenzoyl) oxy-tetrahydrofuran-2-yl] methyl ester (0.05 g, 94.17 μmol) and cyclopropylamine (8 mg, 141.25 μmol) in t-BuOH (2 mL) was added DIEA (24 mg, 188.34 μmol) in one portion. The resulting mixture was stirred at 90 ° C for 16 hours. After cooling to room temperature, the mixture was concentrated under reduced pressure. The residue was purified by preparative TLC (50% EtOAc in petroleum ether) to provide a colorless oily compound (45 mg, 87%). 1 H NMR (400 MHz, CDCl 3 )δ8.66(s,1H),8.24(s,1H),8.06-8.00(m,2H),7.87-7.80(m,2H),7.30(d,J=8.0Hz,2H),7.19(d,J=8.0Hz,2H),6.65-6.55(m,1H),6.21 -6.13(m,1H),4.91(d,J=12.0Hz,1H),4.63(d,J=12.0Hz,1H),3.47-3.30(m,1H),3.00-2.90(m,1H),2.74(s,1H),2.44(d,J=16.0Hz,6H).

[0479] Step 2. Heat the mixture at 25°C in H 2 To a mixture of (2R,3S,5R)-5-(6-(cyclopropylamino)-9H-purin-9-yl)-2-ethynyl-2-(((4-methylbenzoyl)oxy)methyl)tetrahydrofuran-3-yl 4-methylbenzoate (0.07 g, 126.91 μmol) in MeOH (5 mL) was added 10% palladium on carbon (2 mg) in one portion at 15 psi. The resulting mixture was stirred at 25 °C for 1 hour. The mixture was then directly filtered and concentrated to provide the compound as a colorless oil (0.07 g, 99%).

[0480] Step 3. To a mixture of 4-methylbenzoic acid [(2R,3S,5R)-5-[6-(cyclopropylamino)purin-9-yl]-2-ethyl-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester (0.07 g, 125.98 μmol) in MeOH (5 mL) was added NaOMe (681 μg, 12.60 μmol) in one portion at 25°C. After stirring at 25°C for 18 hours, the mixture was concentrated under reduced pressure. The residue was purified by preparative HPLC (acetonitrile 5-35% / 0.05% NH 3 ·H 2 O + 10 mM NH4 HCO 3 The residue was purified by evaporation (4% by weight) in water to provide the title compound (10 mg, 49%) as a white solid. 1 H NMR(400MHz,DMSO-d6)δ8.34(s,1H),8.23(s,1H),7.94(s,1H),6.35-6.27(m,1H),5.25-5.12(m,2H),4.46-4.36(m,1H),3.60-3.49(m,1H) ,3.46-3.39(m,1H),2.94-2.84(m,1H),2.31-2.22(m,1H),1.72-1.51(m,2H),0.88(t,J=7.5Hz,3H),0.75-0.68(m,2H),0.64-0.56(m,2H).

[0481] Embodiment 14

[0482] Synthesis of (2R,3S,5R)-5-(2-amino-6-(cyclopropylamino)-9H-purin-9-yl)-2-ethyl-2-(hydroxymethyl)tetrahydrofuran-3-ol (Compound No. 97)

[0483]

[0484] (2R,3S,5R)-5-(2-amino-6-(cyclopropylamino)-9H-purin-9-yl)-2-ethynyl-2-(hydroxymethyl)tetrahydrofuran-3-ol was synthesized according to the procedure described in WO 2007 / 038507 A2 using N6-cyclopropyl-9H-purine-2,6-diamine.

[0485] Step 1. At 25°C in N 2 To a mixture of 4-methylbenzoic acid [(2R, 3S) -5- (2-amino-6-chloro-purine-9-yl) -2-ethynyl -3- (4-methylbenzoyl) oxy-tetrahydrofuran-2-yl] methyl ester (0.20 g, 366.32 μmol) and cyclopropylamine (31 mg, 549.48 μmol) in t-BuOH (15 mL) was added DIEA (95 mg, 732.64 μmol) in one portion. The resulting mixture was stirred at 90 ° C for 16 hours. After cooling to room temperature, the reaction mixture was concentrated under reduced pressure. The residue was purified by preparative TLC (50% EtOAc in petroleum ether) to provide a colorless oily compound (0.04 g, 19%). 1 HNMR (400MHz, CDCl 3): δ8.05-7.97(d,J=8.4Hz,2H),7.90(d,J=8.0Hz,2H),7.59(s,1H),7.30(s,2H) ,7.21(d,J=7.6Hz,2H),6.43-6.37(m,1H),6.20-6.13(m,1H),5.04(d,J=11.6Hz, 1H),4.59(d,J=11.6Hz,1H),3.42-3.33(m,1H),3.13-2.97(m,1H),2.81-2.72(m ,1H),2.68(s,1H),2.43(d,J=16.4Hz,6H),0.96-0.85(m,2H),0.67-0.60(m,2H).

[0486] Step 2. Heat the mixture at 25°C in H 2 To a solution of [(2R, 3S, 5R)-5-[2-amino-6-(cyclopropylamino)purin-9-yl]-2-ethynyl-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl 4-methylbenzoate (0.02 g, 35.30 μmol) in MeOH (5 mL) was added 10% palladium on carbon (24 mg) in one portion under (15 psi). The resulting mixture was stirred at 25 ° C for 1 hour. The mixture was then filtered and concentrated to provide the compound as a white solid (0.02 g, 99%).

[0487] Step 3. To a solution of [(2R,3S,5R)-5-[2-amino-6-(cyclopropylamino)purin-9-yl]-2-ethyl-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl 4-methylbenzoate (0.02 g, 35.05 μmol) in MeOH (5 mL) was added NaOMe (189 μg, 3.50 μmol) in one portion at 25° C. The resulting mixture was stirred at 25° C. for 18 hours to provide the title compound. 1 H NMR(400MHz,DMSO-d6)δ7.91(s,1H),7.34(s,1H),6.21-6.10(m,1H),5.80 (s,2H),5.17-5.26(m,1H),5.10(d,J=4.4Hz,1H),4.43-4.30(m,1H),3.52- 3.45(m,1H),3.44-3.39(m,1H),2.81-2.69(m,1H),2.23-2.13(m,1H),1.69 -1.49(m,2H),0.86(t,J=7.6Hz,3H),0.70-0.63(m,2H),0.61-0.54(m,2H).

[0488] Embodiment 15

[0489] Synthesis of 1-((2R,4S,5R)-5-ethyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-4-hydroxy-2-oxo-1,2-dihydropyrimidine-5-carbonitrile (Compound No. 98)

[0490]

[0491] 4-Methylbenzoic acid [(2R,3S,5R)-5-(5-cyano-2,4-dioxo-pyrimidin-1-yl)-2-ethynyl-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester was synthesized according to the procedure described in WO 2007 / 038507 A2.

[0492] Step 1. Heat the mixture at 25°C in H 2 To a solution of 4-methylbenzoic acid [(2R,3S,5R)-5-(5-cyano-2,4-dioxo-pyrimidin-1-yl)-2-ethynyl-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester (5 mg, 97.37 μmol) in MeOH (3 mL) was added 10% palladium on carbon (10 mg, 83.33 μmol) in one portion under (15 psi). After stirring at 25 °C for 1 hour, the mixture was filtered and concentrated to provide a white oily compound (25 mg, 50%).

[0493] Step 2. To a solution of 4-methylbenzoic acid [(2R,3S,5R)-5-(5-cyano-2,4-dioxo-pyrimidin-1-yl)-2-ethyl-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester (50 mg, 96.61 μmol) in MeOH (5 mL) was added NaOMe (521.94 μg, 9.66 μmol) in one portion at 25°C. After stirring at 25°C for 1 hour, the mixture was concentrated. The residue was purified by preparative HPLC (acetonitrile 3-33% / 0.225% formic acid in water) to provide the title compound (19 mg, 71%) as a white solid. 1 H NMR (400 MHz, DMSO-d 6 )δ8.91(s,1H),5.94(t,J=6.0Hz,1H),5.32-5.25(m,1H),5.17-5.12(m,1H),4.31-4.23(m,1H),3.60-3.52 (m,1H),3.45-3.39(m,1H),2.37-2.22(m,2H),1.65-1.55(m,1H),1.54-1.39(m,1H),0.86(t,J=7.6Hz,3H).

[0494] Example 16

[0495] Synthesis of (9-((2R,4S,5R)-5-ethyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-9H-purin-6-ol (Compound No. 1)

[0496]

[0497] 4-Methylbenzoic acid [(2R,3S,5R)-2-ethynyl-5-(6-hydroxypurin-9-yl)-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester was synthesized using 9H-purin-6-ol according to the procedure described in WO 2007 / 038507 A2.

[0498] Step 1. A mixture of 4-methylbenzoic acid [(2R,3S,5R)-2-ethynyl-5-(6-hydroxypurin-9-yl)-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester (50 mg, 97 μmol) and Lindela's catalyst (20 mg) in MeOH (5 mL) was heated at 20 °C in H 2 (15 psi) for 30 min. After this time, the reaction mixture was filtered and concentrated to provide the compound as a white solid (40 mg, 79%).

[0499] Step 2. To a solution of [(2R, 3S, 5R)-2-ethyl-5-(6-hydroxypurin-9-yl)-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl 4-methylbenzoate (40 mg, 77 μmol) in THF (3 mL) was added NaOMe (2 mg, 38 μmol) in MeOH (2 mL) at 0°C. The resulting mixture was stirred at 25°C for 16 hours. The reaction mixture was then concentrated. The residue was purified by preparative HPLC (basic) to provide the title compound (4.3 mg, 20%) as a white solid. 1 H NMR(400MHz,DMSO-d6)δ8.22(s,1H),7.99(s,1H),6.26-6.23(m,1H),4.41-4.38(m,1H),3.52-3.33(m ,2H),2.83-2.76(m,1H),2.30-2.24(m,1H),1.68-1.53(m,2H),0.87(t,J=7.2Hz,3H).LCMS(ESI):m / z 302.9(M+Na)+

[0500] Embodiment 17

[0501] Synthesis of (2R,3S,5R)-5-(6-(cyclopropylamino)-2-fluoro-9H-purin-9-yl)-2-ethyl-2-(hydroxymethyl)tetrahydrofuran-3-ol (Compound No. 99)

[0502]

[0503] 4-Methylbenzoic acid [(2R,3S,5R)-5-(6-chloro-2-fluoro-purin-9-yl)-2-ethynyl-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester was synthesized according to the procedure described in WO 2007 / 038507 A2 using N-cyclopropyl-2-fluoro-9H-purin-6-amine.

[0504] Step 1. To a solution of 4-methylbenzoic acid [(2R, 3S, 5R)-5-(6-chloro-2-fluoropurin-9-yl)-2-ethynyl-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester (200 mg, 0.364 mmol) in t-BuOH (2 mL) was added DIPEA (63 μl, 0.364 mmol) and cyclopropylamine (22 mg, 0.383 mmol). The reaction mixture was stirred at 90 ° C for 16 h. After cooling to room temperature, the mixture was diluted with water (10 mL), extracted with EtOAc (20 mL), washed with brine (20 mL×2), and purified by Na 2 SO 4 Dried and concentrated.The residue was purified by column chromatography on silica gel (0-33% EtOAc in petroleum ether) to provide the compound as a white solid (140 mg, 67%). 1 H NMR (400 MHz, CDCl 3 )δ8.02(d,J=8.0Hz,2H),7.93(d,J=8.0Hz,2H),7.85(s,1H),7.29(d,J=8.0Hz,2H),7.23(d,J=8.0Hz,2H),6.51 (t,J=6.4Hz,1H),6.08-6.02(m,1H),4.83(d,J=12.0Hz,1H),4.66(d,J=12.0Hz,1H),3.23-3.16(m,1H),3.04(br s,1H),2.96-2.78(m,1H),2.69(s,1H),2.43(d,J=15.2Hz,6H),0.98-0.92(m,2H),0.70-0.64(m,2H).

[0505] Step 2. Heat the mixture at 25°C in H 2To a mixture of 4-methylbenzoic acid [(2R,3S,5R)-5-[6-(cyclopropylamino)-2-fluoro-purin-9-yl]-2-ethynyl-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester (90 mg, 158.01 μmol) in EtOAc (5 mL) was added palladium on carbon (25 mg, 47.40 μmol, 20% purity) in one portion. The mixture was heated at 25° C. under H 2 (15 psi) for 16 h. After this time, the reaction mixture was filtered and the filtrate was concentrated to provide the compound as a white solid (75 mg, 83%).

[0506] Step 3. To a solution of 4-methylbenzoic acid [(2R,3S,5R)-5-[6-(cyclopropylamino)-2-fluoro-purin-9-yl]-2-ethyl-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester (70 mg, 122.03 μmol) in MeOH (3 mL) was added NH 3 ·H 2 O (1 mL, 6.49 mmol, 25% purity). The resulting mixture was stirred at 40 ° C for 16 h. After this time, the reaction mixture was concentrated under reduced pressure. The residue was purified by reverse phase HPLC (acetonitrile 1-27% / 0.225% formic acid in water) to give the title compound (6.5 mg, 16%) as a white solid. 1 H NMR (400MHz, DMSO-d6) δ8.53(brs,1H),8.33(s,1H),6.20(t,J=6.8Hz,1H),5.17(d, J=4.8Hz,1H),4.92(t,J=5.2Hz,1H),4.46-4.36(m,1H),3.53-3.40(m,2H),2.93(br s,1H),2.84-2.76(m,1H),2.36-2.23(m,1H),1.70-1.49(m,2H),0.92-0.83(m,3H),0.75-0.61(m,4H).

[0507] Embodiment 18

[0508] Synthesis of 9-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-vinyltetrahydrofuran-2-yl)-9H-purin-6-ol (Compound No. 100)

[0509]

[0510] 4-Methylbenzoic acid [(2R,3S,5R)-2-ethynyl-5-(6-hydroxypurin-9-yl)-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester was synthesized using 9H-purin-6-ol according to the procedure described in WO 2007 / 038507 A2.

[0511] Step 1. A mixture of 4-methylbenzoic acid [(2R,3S,5R)-2-ethynyl-5-(6-hydroxypurin-9-yl)-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester (50 mg, 98 μmol) and Lindela's catalyst (20 mg) in MeOH (20 mL) was heated at 20 °C in H 2 (15 psi) for 30 min. The reaction mixture was then filtered. The filtrate was concentrated to provide the compound as a white solid (35 mg, 69%).

[0512] Step 2. To a solution of [(2R, 3S, 5R)-5-(6-hydroxypurin-9-yl)-3-(4-methylbenzoyl)oxy-2-vinyl-tetrahydrofuran-2-yl]methyl 4-methylbenzoate (20 mg, 39 μmol) in THF (3 mL) was added NaOMe (1.1 mg, 19 μmol) in MeOH (2 mL) at 0°C. The resulting mixture was stirred at 25°C for 16 hours. After this, the reaction mixture was concentrated. The residue was purified by preparative HPLC (basic) to provide the title compound (4 mg, 36%) as a white solid. 1 HNMR(400MHz,DMSO-d6)δ8.26(s,1H),8.00(s,1H),6.29-6.26(m,1H),5.98-5.92(m,1H),5.40-5.35(m,1H),5. 24-5.18(m,1H),4.62(t,J=6.4Hz,1H)3.52-3.33(m,2H),2.62-2.57(m,1H),2.26-2.21(m,1H).LCMS(ESI):m / z 279.1(M+H)+.

[0513] Embodiment 19

[0514] Synthesis of 5-bromo-1-((2R,4S,5R)-5-ethyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-4-hydroxypyrimidin-2(1H)-one (Compound No. 2)

[0515]

[0516] 4-Methylbenzoic acid [(2R,3S)-5-acetoxy-2-ethynyl-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester was synthesized according to the procedure described in WO 2007 / 038507 A2.

[0517] Step 1. Heat the mixture at 25°C in H 2 To a solution of 4-methylbenzoic acid [(2R, 3S)-5-acetoxy-2-ethynyl-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester (200 mg, 458.24 μmol) in MeOH (10 mL) was added 10% palladium on carbon (9.00 mg) in one portion at (15 psi). The resulting mixture was stirred at 25 °C for 3 hours. After this, the reaction mixture was filtered and concentrated to provide the compound as a white solid (0.2 g, 99%).

[0518] Step 2. Incubate at 25°C in N 2 To a solution of 5-bromo-1H-pyrimidine-2,4-dione (173.4 mg, 908.09 μmol) in MeCN (10 mL) was added BTMSA (309.5 mg, 1.82 mmol) at once. The mixture was then stirred at 70 ° C for 1 hour. After cooling to 25 ° C, TMSOTf (131.2 mg, 590.26 μmol) and 4-methylbenzoic acid [(2R, 3S) -5-acetoxy-2-ethyl-3- (4-methylbenzoyl) oxy-tetrahydrofuran-2-yl] methyl ester (0.2 g, 454.05 μmol) in MeCN (5 mL) were added dropwise at 25 ° C. The resulting mixture was stirred at 25 ° C for 1 hour. After this, the reaction was quenched by adding 10 mL of citric acid aqueous solution (10 wt %) and then extracted with EtOAc (20 mL). The layers were separated and the organic layer was washed with brine (10 mL) and then purified by Na 2 SO 4 Dried, filtered and concentrated.The residue was purified by preparative TLC (50% EtOAc in petroleum ether) to provide the compound as a colorless oil (52 mg, 20%). 1 H NMR (400 MHz, CDCl 3)δ8.25(s,1H),7.99-7.86(m,5H),7.34-7.27(m,4H),6.35-6.26(m,1H),5.85-5.75(m,1H),4.65-4.58(m,1H),4.56-4.48(m, 1H), 2.76-2.66 (m, 1H), 2.55-2.48 (m, 1H), 2.44 (d, J = 8.8Hz, 6H), 2.05-1.91 (m, 1H), 1.90-1.76 (m, 1H), 1.08 (t, J = 7.6Hz, 3H).

[0519] Step 3. To a solution of 4-methylbenzoic acid [(2R, 3S, 5R) -5- (5-bromo-2,4-dioxo-pyrimidin-1-yl) -2-ethyl-3- (4-methylbenzoyl) oxy-tetrahydrofuran-2-yl] methyl ester (0.02 g, 35.00 μmol) in MeOH (5 mL) was added NaOMe (189 μg, 3.50 μmol) in one portion at 25 °C. The resulting mixture was stirred at 25 °C for 1 hour. After this, the mixture was concentrated under reduced pressure. The residue was purified by preparative HPLC (acetonitrile 0-30% / 0.225% formic acid in water) to provide the title compound (5.4 mg, 46%) as a white solid. 1 H NMR (400MHz, DMSO-d6) δ8.47(s,1H),6.03(t,J=6.4Hz,1H),5.27-5.18(m,1H),5.14(d,J=4.8Hz,1H),4.33-4.21(m,1H),3.56-3. 49(m,1H),3.45-3.40(m,1H),2.31-2.24(m,1H),2.22-2.13(m,1H),1.64-1.53(m,1H),1.52-1.41(m,1H),0.85(t,J=7.6Hz,3H).

[0520] Embodiment 20

[0521] Synthesis of (2R,3S,5R)-5-(2-amino-6-methoxy-9H-purin-9-yl)-2-(hydroxymethyl)-2-vinyltetrahydrofuran-3-ol (Compound No. 102)

[0522]

[0523] 4-Methylbenzoic acid [(2R,3S,5R)-5-(2-amino-6-methoxy-purin-9-yl)-2-ethynyl-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester was prepared according to WO 2007 / 038507 A2 using 6-methoxy-9H-purin-2-amine.

[0524] Step 1. Heat the mixture at 25°C in H 2 To a solution of 4-methylbenzoic acid [(2R, 3S, 5R)-5-(2-amino-6-methoxy-purin-9-yl)-2-ethynyl-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester (20 mg, 36.93 μmol) in MeOH (2 mL) was added Lindela catalyst (8 mg) in one portion under (15 psi). The resulting mixture was stirred at 25 °C for 1 hour. The mixture was then filtered and the filtrate was concentrated to provide the compound as a white solid (20 mg, 99%).

[0525] Step 2. To a mixture of 4-methylbenzoic acid [(2R,3S,5R)-5-(2-amino-6-methoxy-purin-9-yl)-3-(4-methylbenzoyl)oxy-2-vinyl-tetrahydrofuran-2-yl]methyl ester (17 mg, 31.27 μmol) in MeOH (5 mL) was added NaOMe (169 μg, 3.13 μmol) in one portion at 25°C. After stirring at 25°C for 1 hour, the mixture was concentrated under reduced pressure. The residue was purified by preparative HPLC (basic) to provide the title compound (2.5 mg, 26%) as a white solid. LCMS (ESI): m / z 308.2 (M+H)+. 1 H NMR(400MHz,CD3OD)δ8.61(s,1H),6.46-6.39(m,1H),6.06-6.95(m,1H),5.62-5.51(m,1H),5.38-5.3 1(m,1H),4.65(t,J=7.6Hz,1H),4.08(s,3H),3.76-3.67(m,1H),3.61-3.56(m,1H),2.53-2.39(m,2H).

[0526] Embodiment 21

[0527] Synthesis of (2R,3S,5R)-5-(2-amino-6-methoxy-9H-purin-9-yl)-2-ethyl-2-(hydroxymethyl)tetrahydrofuran-3-ol (Compound No. 125)

[0528]

[0529] 4-Methylbenzoic acid [(2R,3S,5R)-5-(2-amino-6-methoxy-purin-9-yl)-2-ethynyl-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester was prepared according to WO 2007 / 038507 A2 using 6-methoxy-9H-purin-2-amine.

[0530] Step 1. In H 2 To a mixture of 4-methylbenzoic acid [(2R,3S,5R)-5-(2-amino-6-methoxy-purin-9-yl)-2-ethynyl-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester (0.1 g, 184.65 μmol) in EtOAc (5 mL) was added Pd / C (60 mg, 10% purity) under (15 psi). The mixture was stirred at 25 °C for 3 hours. After this time, the mixture was filtered and concentrated to provide the title compound (80 mg, 79% yield) as a white solid.

[0531] Step 2. To a mixture of 4-methylbenzoic acid [(2R, 3S, 5R) -5- (2-amino-6-methoxy-purine-9-yl) -2-ethyl-3- (4-methylbenzoyl) oxy-tetrahydrofuran-2-yl] methyl ester (80 mg, 146.63 μmol) in MeOH (5 mL) was added NaOMe (1 mg, 14.66 μmol) at 0°C. The resulting mixture was stirred at 25°C for 16 hours. The reaction mixture was then concentrated. The residue was purified by preparative HPLC (basic) to provide the title compound (7 mg, 15%) as a white solid.

[0532] 1 H NMR (400MHz, CD3OD): δ8.50(s,1H),6.41(t,J=6.4Hz,1H),4.53(t,J=5.6Hz,1H),4.08(s,3H),3.70-3.64( m,1H),3.61-3.56(m,1H),2.63-2.50(m,2H),1.85-1.73(m,1H),1.71-1.60(m,1H),1.01(t,J=7.6Hz,3H).

[0533] Embodiment 22

[0534] Synthesis of 1-((2R,4S,5R)-5-ethyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-5-methoxypyrimidine-2,4(1H,3H)-dione (Compound No. 103)

[0535]

[0536] 4-Methylbenzoic acid [(2R,3S,5R)-2-ethynyl-5-(5-methoxy-2,4-dioxo-pyrimidin-1-yl)-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester was prepared according to WO 2007 / 038507 A2.

[0537] Step 1. Heat the mixture at 25°C in H 2 To a mixture of 4-methylbenzoic acid [(2R, 3S, 5R) -2-ethynyl -5- (5-methoxy-2,4-dioxo-pyrimidin-1-yl) -3- (4-methylbenzoyl) oxy-tetrahydrofuran-2-yl] methyl ester (15 mg, 28.93 μmol) in MeOH (5 mL) was added 10% palladium on carbon (9 mg) in one portion at (15 psi). The resulting mixture was stirred at 25 ° C for 1 hour. The mixture was then filtered and the filtrate was concentrated to provide the compound as a white solid (15 mg, 99%).

[0538] Step 2. To a mixture of 4-methylbenzoic acid [(2R, 3S, 5R) -2-ethyl-5- (5-methoxy-2,4-dioxo-pyrimidin-1-yl) -3- (4-methylbenzoyl) oxy-tetrahydrofuran-2-yl] methyl ester (15 mg, 28.71 μmol) in MeOH (5 mL) was added NaOMe (155 μg, 2.87 μmol) in one portion at 25°C. After stirring at 25°C for 18 hours, the mixture was concentrated under reduced pressure. The residue was purified by preparative HPLC (basic) to provide the title compound (2.3 mg, 28%) as a white solid. 1 H NMR (400MHz, CD3OD) δ7.87 (s, 1H), 6.25 (t, J = 6.4Hz, 1H), 4.55-4.47 (m, 1H), 3.76-3.72 (m, 1H), 3.71 (s, 3H), 3.63-3.58 (m, 1H), 2.45-2.27 (m, 2H), 1.79-1.66 (m, 1H), 1.64-1.52 (m, 1H), 0.97 (t, J = 7.6Hz, 3H).

[0539] Embodiment 23

[0540] Synthesis of 5-fluoro-1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-vinyltetrahydrofuran-2-yl)pyrimidine-2,4(1H,3H)-dione (Compound No. 17)

[0541]

[0542] [(2R,3S,5R)-2-ethynyl-5-(5-fluoro-4-hydroxy-2-oxo-pyrimidin-1-yl)-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester was prepared according to WO 2007 / 038507 A2 using 4-methylbenzoic acid 5-fluoropyrimidine-2,4(1H,3H)-dione.

[0543] Step 1. A mixture of 4-methylbenzoic acid [(2R,3S,5R)-2-ethynyl-5-(5-fluoro-4-hydroxy-2-oxo-pyrimidin-1-yl)-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester (30 mg, 59.23 μmol) and Lindela's catalyst (12 mg) in EtOAc (2 mL) was heated at 20 °C under H 2 (15 psi) for 10 min. After this time, the reaction mixture was filtered and the filtrate was concentrated to provide the compound as a white solid (30 mg, 99%). LCMS (ESI): m / z 531.1 (M+Na)+.

[0544] Step 2. A mixture of [(2R, 3S, 5R)-5-(5-fluoro-4-hydroxy-2-oxo-pyrimidin-1-yl)-3-(4-methylbenzoyl)oxy-2-vinyl-tetrahydrofuran-2-yl]methyl 4-methylbenzoate (30 mg, 59 μmol) and NaOMe (3.19 mg, 59 μmol) in MeOH (2 mL) and THF (3 mL) was stirred at 20 ° C for 3 hours. After this, the reaction mixture was concentrated to give the crude product. Purification by preparative HPLC (basic) provided the title compound (4.3 mg, 27%) as a white solid. 1 H NMR(400MHz,DMSO-d6)δ8.26(d,J=7.2Hz,1H),6.06-6.04(m,1H),5.90-5.80(m,1H),5.48(s,1H),5.35- 5.30(m,2H),5.20-5.17(m,1H),4.42-4.31(m,1H),3.68-3.33(m,2H),2.21-2.04(m,2H).LCMS(ESI):m / z 295.0(M+Na)+.

[0545] Embodiment 24

[0546] Synthesis of 1-((2R,4S,5R)-5-ethyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-5-fluoropyrimidine-2,4(1H,3H)-dione (Compound No. 18)

[0547]

[0548] 4-Methylbenzoic acid [(2R,3S,5R)-2-ethynyl-5-(5-fluoro-4-hydroxy-2-oxo-pyrimidin-1-yl)-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester was prepared according to WO 2007 / 038507 A2 using 5-fluoropyrimidine-2,4(1H,3H)-dione.

[0549] Step 1. A mixture of 4-methylbenzoic acid [(2R,3S,5R)-2-ethynyl-5-(5-fluoro-4-hydroxy-2-oxo-pyrimidin-1-yl)-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester (50 mg, 98.72 μmol) and Lindela's catalyst (5 mg) in MeOH (5 mL) and EtOAc (5 mL) was stirred at 20 °C under H 2 (15 psi) for 5 hours. After this time, the reaction mixture was filtered and the filtrate was concentrated to provide the compound as a white solid (40 mg, 79%). LCMS (ESI): m / z 533.1 (M+Na)+.

[0550] Step 2. A mixture of [(2R, 3S, 5R) -2-ethyl-5- (5-fluoro-4-hydroxy-2-oxo-pyrimidin-1-yl) -3- (4-methylbenzoyl) oxy-tetrahydrofuran-2-yl] methyl 4-methylbenzoate (40 mg, 78.35 μmol) and NaOMe (4.23 mg, 78.35 μmol) in MeOH (2 mL) and THF (3 mL) was stirred at 20 ° C for 3 hours. The reaction mixture was then concentrated to generate the crude product. Purification by preparative HPLC (basic) provided the title compound (5.8 mg, 27%) as a white solid. 1 HNMR(400MHz,DMSO-d6)δ8.00(d,J=7.2Hz,1H),6.07(t,J=6.8Hz,1H),5.10-5.08(m,2H),4.26(t,J=6.0Hz,1H) ,3.50-3.30(m,2H),2.18-2.08(m,2H),1.59-1.45(m,2H),0.85(t,J=7.6Hz,3H).LCMS(ESI):m / z297.0(M+Na)+.

[0551] Embodiment 25

[0552] Synthesis of 4-amino-1-((2R,4R,5S)-5-ethynyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)pyrimidin-2(1H)-one (Compound No. 77)

[0553]

[0554] Step 1. Add BTMSA (234 mg, 1.37 mmol) to a mixture of N-(2-oxo-1H-pyrimidin-4-yl)benzamide (148 mg, 687.36 μmol) in MeCN (15 mL). The resulting mixture was heated at 70 ° C for 1 hour. After cooling to ambient temperature, TMSOTf (99 mg, 446.78 μmol) and 4-methylbenzoic acid [(2S, 3R) -5-acetoxy-2-ethynyl-3- (4-methylbenzoyl) oxy-tetrahydrofuran-2-yl] methyl ester (150 mg, 343.68 μmol) were added successively. The resulting mixture was heated at 70 ° C for another 1 hour. After cooling to ambient temperature, the reaction mixture was poured into water (10 mL), extracted with EtOAc (20 mLx2), and concentrated. Through SFC (DAICEL CHIRALPAK AD, (250mm*30mm, 10um); supercritical CO 2 / EtOH+NH 4 The residue was purified by 4% paraformaldehyde (5% CO 2 ) to afford the compound as a white solid (50 mg, 22%). 1 HNMR (400MHz, CDCl 3 ): δ8.71(s,1H),8.49(d,J=7.2Hz,1H),7.97(d,J=8.4Hz,2H),7.92(d,J=7.6Hz,2 H),7.79(d,J=8.4Hz,2H),7.67-7.62(m,1H),7.58-7.52(m,2H),7.30(d,J=8.0Hz ,2H),7.20(d,J=8.0Hz,2H),6.39-6.30(m,1H),5.92-5.85(m,1H),4.70-4.61(m, 2H),3.28-3.14(m,1H),2.79(s,1H),2.73-2.64(m,1H),2.45(s,3H),2.37(s,3H).

[0555] Step 2. To a mixture of 4-methylbenzoic acid [(2S,3R,5R)-5-(4-benzamido-2-oxo-pyrimidin-1-yl)-2-ethynyl-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester (50 mg, 84.52 μmol) in MeOH (2 mL) was added NaOMe (9 mg, 169.03 μmol) at 0°C. The resulting mixture was stirred at 25°C for 16 hours. The reaction mixture was then concentrated. The residue was purified by preparative HPLC (acetonitrile 1-15% / 0.225% formic acid in water) to provide the title compound (3.5 mg, 16%) as a white solid. 1 H NMR (400MHz, DMSO-d6): δ7.95(s,1H),7.37(s,1H),6.17(t,J=6.4Hz,1H),6.03- 5.90(m,1H),5.81(s,2H),5.43(s,1H),5.37-5.31(m,1H),5.22(s,1H),5.20-5. 15(m,1H),4.58(t,J=6.4Hz,1H),3.55-3.47(m,1H),3.45-3.38(m,1H),2.57-2. 52(m,1H),2.22-2.14(m,1H),0.69-0.62(m,2H),0.62-0.55(m,2H)[M+H]+252.1.

[0556] Embodiment 26

[0557] Synthesis of 2-amino-9-[(2R,4R,5S)-5-ethynyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl]-1H-purin-6-one (Compound No. 79)

[0558]

[0559] Step 1. A solution of N-(6-hydroxy-9H-purin-2-yl)-2-methylpropanamide (84 mg, 378.05 μmol) in MeCN (10 mL) was stirred for 2 h. 2 Degas 3 times. Then add (1E)-N-trimethylsilylacetamidine acid trimethylsilyl ester (349 mg, 1.72 mmol). The resulting mixture is stirred at 70 ° C for 1 h and then cooled to room temperature. Add trimethylsilyl trifluoromethanesulfonate (88 mg, 395.23 μmol) and then at 25 ° C under N 2Next, 4-methylbenzoic acid [(2S,3R)-5-acetoxy-2-ethynyl-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester (150 mg, 343.68 μmol) in acetonitrile (2 mL) was added in one portion. The mixture was stirred at 70 ° C for 16 h. After cooling to room temperature, the mixture was diluted with water (10 mL), extracted with EtOAc (20 mL), washed with brine (20 mL), and purified by Na 2 SO 4 Dried, filtered and concentrated.The residue was purified by preparative HPLC (acetonitrile 54-85% / 0.225% formic acid in water) to afford the compound as a white solid (25 mg, 15%). 1 H NMR (400 MHz, CDCl 3 ): δ11.97(s,1H),8.31(s,1H),8.27(br s,1H),7.97(d,J=8.0Hz,2H),7.84-7.73(m,2H),7.30-7.21(m,4H),6.34-6.27(m,1H),5.89-5.81(m,1H),4.73-4.66(m,1H),4.63-4.57 (m,1H),3.15-3.06(m,1H),2.96-2.86(m,1H),2.75-2.71(m,1H),2.67-2.56(m,1H),2.42(d,J=11.6Hz,6H),1.28(dd,J=2.0,6.8Hz,6H).

[0560] Step 2. To a solution of [(2S,3R,5R)-2-ethynyl-5-[6-hydroxy-2-(2-methylpropionylamino)purin-9-yl]-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl 4-methylbenzoate (21 mg, 35.14 μmol) in MeOH (2 mL) was added ammonium hydroxide (30 mg, 105.42 μmol, 28 wt %). The solution was stirred at 60 °C for 16 h. After cooling to room temperature, the reaction mixture was concentrated. The residue was purified by preparative HPLC (acetonitrile 1-20% / 0.225% formic acid in water) to provide the title compound (3.0 mg, 29%) as a white solid. 1H NMR (400MHz, DMSO-d6): δ10.84(br s,1H),8.24(s,1H),6.50-6.41(m,1H),6.15(s,2H),5.52(d,J=5.2Hz,1H),5.36(t,J=5.6Hz ,1H),4.53-4.41(m,1H),3.72-3.53(m,2H),3.51(s,1H),2.46-2.34(m,2H).[M+Na]+313.9.

[0561] Embodiment 27

[0562] Synthesis of (2R,3S,5R)-5-(4-amino-6-fluoro-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-ethynyl-2-(hydroxymethyl)tetrahydrofuran-3-ol (Compound No. 75)

[0563]

[0564] 4-Methylbenzoic acid [(2R,3S,5R)-5-(4,6-diamino-3-methyl-pyrazolo[3,4-d]pyrimidin-1-yl)-2-ethynyl-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester was prepared according to WO 2007 / 038507 A2 using 3-methyl-1H-pyrazolo[3,4-d]pyrimidine-4,6-diamine.

[0565] Step 1. At 0°C under N 2 To a mixture of 4-methylbenzoic acid [(2R,3S,5R)-5-(4,6-diamino-3-methyl-pyrazolo[3,4-d]pyrimidin-1-yl)-2-ethynyl-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester (120 mg, 221.99 μmol) in MeOH (15 mL) was added NaOMe (24 mg, 443.98 μmol) in one portion. The resulting mixture was stirred at 25 °C for 16 hours. The reaction mixture was then concentrated. The results were analyzed by preparative TLC (9% MeOH in dichloromethane (1% NH 4 The residue was purified by 4% paraformaldehyde (5% HCl) to afford the compound as a white solid (60 mg, 89%). LCMS (ESI): m / z 327.1 (M+Na)+.

[0566] Step 2. At 0°C under N 2To a mixture of (2R,3S,5R)-5-(4,6-diamino-3-methyl-pyrazolo[3,4-d]pyrimidin-1-yl)-2-ethynyl-2-(hydroxymethyl)tetrahydrofuran-3-ol (60 mg, 197.17 μmol) in pyridine (2 mL) was added pyridine-hydrofluoride (237 mg, 1.68 mmol, 70% purity) and tert-butyl nitrite (61 mg, 591.51 μmol) dropwise. The resulting mixture was stirred at 0°C for 1 hour. After this, DCM (15 mL) and H 2 The mixture was diluted with 1% O (5 mL). The layers were separated and the aqueous layer was extracted with DCM (10 mL x 3). The combined organic extracts were purified by Na 2 SO 4 Dry, filter, and concentrate under vacuum.The residue was further purified by preparative HPLC (acetonitrile 10-40% / 0.225% formic acid in water) to provide the title compound (1.58 mg, 3%). 1 H NMR (400MHz, DMSO-d6): δ8.24(s,1H),7.51(s,1H),6.37-6.23(m,1H),5.46(d,J=6.0Hz,1H),4.98(t,J=6.0Hz,1 H),4.60-4.47(m,1H),3.61-3.53(m,1H),3.48-3.39(m,2H),2.82-2.69(m,1H),2.53(s,3H),2.40-2.34(m,1H).

[0567] Embodiment 28

[0568] Synthesis of (2R,3S,5S)-5-(4-amino-6-fluoro-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-ethynyl-2-(hydroxymethyl)tetrahydrofuran-3-ol (Compound No. 3)

[0569]

[0570] 4-Methylbenzoic acid [(2R,3S,5S)-5-(4,6-diamino-3-methyl-pyrazolo[3,4-d]pyrimidin-1-yl)-2-ethynyl-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester was prepared according to WO 2007 / 038507 A2 using 3-methyl-1H-pyrazolo[3,4-d]pyrimidine-4,6-diamine.

[0571] Step 1. At 0°C under N 2To a mixture of 4-methylbenzoic acid [(2R,3S,5S)-5-(4,6-diamino-3-methyl-pyrazolo[3,4-d]pyrimidin-1-yl)-2-ethynyl-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester (0.1 g, 184.99 μmol) in MeOH (15 mL) was added NaOMe (20 mg, 369.98 μmol) in one portion. The mixture was stirred at 25 °C for 18 hours. The reaction mixture was then concentrated and analyzed by preparative TLC with 10% MeOH (1% NH 4 The residue was purified by eluting with 4-(4-(4-hydroxy-1-hydroxy- ...

[0572] Step 2. At 0°C under N 2 To a mixture of (2R,3S,5S)-5-(4,6-diamino-3-methyl-pyrazolo[3,4-d]pyrimidin-1-yl)-2-ethynyl-2-(hydroxymethyl)tetrahydrofuran-3-ol (60 mg, 197.17 μmol) in pyridine (2 mL) was added HF-pyridine (237 mg, 1.68 mmol, 70% purity) and tert-butyl nitrite (61 mg, 591.51 μmol) dropwise. The resulting mixture was stirred at 0°C for 1 hour. After this, the mixture was washed with DCM (15 mL) and H 2 The layers were separated and the aqueous layer was extracted with DCM (10 mL). The combined organic extracts were washed with anhydrous Na 2 SO 4 Dry, filter, and concentrate in vacuo.The residue was purified by preparative HPLC (acetonitrile 10-40% / 0.225% formic acid in water) to provide the title compound (5 mg, 8%) as a white solid. 1 H NMR (400MHz, DMSO-d6) δ8.27(s,1H),7.53(s,1H),6.27(t,J=6.4Hz,1H),5.73(d,J=8.8Hz,1H),5.21(t,J=6.4H z,1H),4.38-4.30(m,1H),3.48-3.40(m,3H),2.79-2.71(m,1H),2.68-2.62(m,1H),2.53(s,3H).[M+H]+308.2.

[0573] Embodiment 29

[0574] Synthesis of 4-amino-1-[(2S,5S)-5-ethynyl-5-(hydroxymethyl)-2H-furan-2-yl]-5-fluoro-pyrimidin-2-one (Compound No. 74)

[0575]

[0576] Step 1. A solution of N-(5-fluoro-2-hydroxy-pyrimidin-4-yl)benzamide (59 mg, 253.94 μmol) in acetonitrile (3 mL) was treated with N 2 Degas 3 times. Then add (1E)-N-trimethylsilylacetamidine acid trimethylsilyl ester (129 mg, 634.85 μmol). The reaction mixture is stirred at 70 ° C for 1 h and then cooled to room temperature. Add trimethylsilyl trifluoromethanesulfonate (56 mg, 253.94 μmol) and then at 25 ° C under N 2 Benzoic acid [(2S,4S,5R)-5-benzoyloxy-2-ethynyl-4-(p-toluenesulfonyl)tetrahydrofuran-2-yl]methyl ester (100 mg, 211.62 μmol) in acetonitrile (1 mL) was added. After stirring at 70 ° C for 16 h, the reaction mixture was cooled to room temperature and diluted with water (10 mL), extracted with EtOAc (20 mL), washed with brine (20 mL), and filtered through Na 2 SO 4 Dried, filtered and concentrated.The residue was purified by preparative TLC (25% EtOAc in petroleum ether) to provide the compound as a white solid (21 mg, 17%). 1 H NMR (400 MHz, CDCl 3 )δ8.33-8.25(m,2H),8.09(d,J=7.2Hz,2H),7.73-7.65(m,1H),7.60-7.4 2(m,6H),7.27(s,2H),7.22(d,J=5.6Hz,1H),7.04(d,J=7.6Hz,2H),6.32 -6.13(m,1H),4.69(d,J=11.6Hz,1H),4.55(d,J=12.0Hz,1H),3.63-3.46 (m,1H),2.95-2.82(m,1H),2.79(s,1H),2.53-2.39(m,1H),2.16(s,3H).

[0577] Step 2. To a solution of [(2S,4S,5S)-5-(4-benzamido-5-fluoro-2-oxo-pyrimidin-1-yl)-2-ethynyl-4-(p-tolylsulfanyl)tetrahydrofuran-2-yl]methyl benzoate (40 mg, 68.54 μmol) in MeCN (2 mL) was added sodium [chloro(p-toluenesulfonyl)amino] (22 mg, 95.95 μmol). The reaction mixture was stirred at 50 °C for 2.5 h. Water (10 mL) was added to the reaction mixture, extracted with EtOAc (20 mL), washed with brine (20 mL), and purified by Na 2SO 4 The mixture was dried, filtered and concentrated. The residue was dissolved in n-BuOH (2 mL). The solution was then stirred at 90 ° C for another 3 h. After cooling to room temperature, the reaction mixture was concentrated. The residue was purified by preparative TLC (25% EtOAc in petroleum ether) to provide a white solid compound (39 mg, 51%). LCMS (ESI): m / z 460.0 (M+H) +.

[0578] Step 3. To a solution of [(2S,5S)-2-(4-benzamido-5-fluoro-2-oxo-pyrimidin-1-yl)-5-ethynyl-2H-furan-5-yl]methyl benzoate (20 mg, 43.53 μmol) in MeOH (1 mL) was added MeONa (3 mg, 53.53 μmol) at 0°C. The resulting mixture was stirred at 25°C for 3 h. The reaction was then concentrated. The residue was purified by preparative HPLC (acetonitrile 1-27% / 0.225% formic acid in water) to provide the title compound (5.93 mg, 19%) as a white solid. 1 H NMR (400MHz, DMSO-d6) δ8.38(br s,1H),7.92(d,J=7.2Hz,1H),7.85(br s,1H),7.61(br s,1H),6.92(s,1H),6.33-6.26(m,1H),6.03(d,J=5.6Hz,1H),3.76-3.56(m,3H).[M+Na]+273.8.

[0579] Embodiment 30

[0580] Synthesis of 4-amino-1-((2S,4R,5S)-5-ethynyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)pyrimidin-2(1H)-one (Compound No. 76)

[0581]

[0582] Step 1. To a mixture of N-(2-oxo-1H-pyrimidin-4-yl)benzamide (148 mg, 687.36 μmol) in MeCN (15 mL) was added BTMSA (234 mg, 1.37 mmol). The resulting mixture was heated at 70 ° C for 1 hour. After cooling to ambient temperature, TMSOTf (99 mg, 446.78 μmol) and 4-methylbenzoic acid [(2S, 3R)-5-acetoxy-2-ethynyl-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl] methyl ester (150 mg, 343.68 μmol) were added in sequence. The resulting mixture was heated at 70 ° C for another 1 hour. After cooling to ambient temperature, the reaction mixture was poured into water (10 mL), extracted with EtOAc (20 mL×2), and concentrated. Through SFC (DAICEL CHIRALPAK AD, (250mm*30mm, 10um); supercritical CO 2 / EtOH+NH4OH=45 / 55; 80 mL / min) to afford the compound as a white solid (20 mg, 22%).

[0583] Step 2. To a mixture of 4-methylbenzoic acid [(2S,3R,5S)-5-(4-phenylamino-2-oxo-pyrimidin-1-yl)-2-ethynyl-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester (0.02 g, 33.81 μmol) in MeOH (2 mL) was added NaOMe (4 mg, 67.61 μmol) at 0°C. The resulting mixture was stirred at 25°C for 16 hours. The reaction mixture was then concentrated. The residue was purified by preparative HPLC (acetonitrile 1-17% / 0.225% formic acid in water) to provide the title compound (3.8 mg, 16%) as a white solid. 1 H NMR (400MHz, DMSO-d6) δ8.37(s,2H),7.80(d,J=7.2Hz,1H),7.23-7.07(m,2H),6.20-6.10(m,1H),5.73(d,J=7.6Hz,1H),4.24(d, J=4.8Hz,1H),3.74(d,J=11.2Hz,1H),3.63(d,J=11.2Hz,1H),3.51(s,1H),2.78-2.69(m,1H),1.85-1.77(m,1H).[2M+H]+503.1.

[0584] Embodiment 31

[0585] Synthesis of 2-amino-9-((2S,4R,5S)-5-ethynyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-1H-purin-6(9H)-one (Compound No. 78)

[0586]

[0587] Step 1. A solution of N-(6-hydroxy-9H-purin-2-yl)-2-methylpropanamide (84 mg, 378.05 μmol) in acetonitrile (10 mL) was stirred for 2 h. 2 Degas 3 times. Then add (1E)-N-trimethylsilylacetamidine acid trimethylsilyl ester (349 mg, 1.72 mmol). After stirring at 70 ° C for 1 h, the reaction mixture was cooled to room temperature. Trimethylsilyl trifluoromethanesulfonate (88 mg, 395.23 μmol) was added, and then stirred at 25 ° C under N 2 Next, 4-methylbenzoic acid [(2S,3R)-5-acetoxy-2-ethynyl-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester (150 mg, 343.68 μmol) in acetonitrile (2 mL) was added in one portion. After stirring at 70 °C for 16 h, the reaction mixture was cooled to room temperature and diluted with water (20 mL), extracted with EtOAc (30 mL), washed with brine (20 mL), and purified by Na 2 SO 4 Dried, filtered and concentrated.The residue was purified by preparative HPLC (acetonitrile 54-85% / 0.225% formic acid in water) to afford the compound as a white solid (21 mg, 10%). 1 H NMR (400MHz, CD3OD) δ8.49-8.36(m,2H),8.00(d,J=8.0Hz,2H),7.87(d,J=8.0Hz,2H),7 .32(d,J=8.0Hz,2H),7.34-7.30(m,1H),7.22(d,J=8.0Hz,2H),6.79-6.69(m,1H),6.12- 6.01(m,1H),4.81(d,J=11.6Hz,1H),4.66(d,J=11.6Hz,1H),3.27(s,1H),3.20-3.09(m, 1H),2.98-2.88(m,1H),2.76-2.69(m,1H),2.43(s,3H),2.38(s,3H),1.24-1.21(m,6H).

[0588] Step 2. To a solution of 4-methylbenzoic acid [(2S, 3R, 5S)-2-ethynyl-5-[6-hydroxy-2-(2-methylpropionylamino)purin-9-yl]-3-(4-methylbenzoyl)oxy-tetrahydrofuran-2-yl]methyl ester (21 mg, 35.14 μmol) in methanol (2 mL) was added ammonium hydroxide (300 mg, 1054.2 μmol, 28% purity). The solution was stirred at 60 ° C for 16 h. The reaction solution was then cooled to room temperature and concentrated. The residue was purified by preparative HPLC (acetonitrile 1-20% / 0.225% formic acid in water) to provide the title compound (3.0 mg, 29%) as a white solid. 1 H NMR(400MHz,DMSO-d6)δ10.84(br s,1H),8.24(s,1H),6.50-6.41(m,1H),6.15(s,2H),5.52(d,J=5.2Hz,1H),5.36(t,J=5.6Hz ,1H),4.53-4.41(m,1H),3.72-3.53(m,2H),3.51(s,1H),2.46-2.34(m,2H).[M+Na]+313.9.

[0589] Embodiment 32

[0590] Synthesis of 1-((4S,5R)-5-ethynyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-1H-1,2,4-triazole-3-carboxamide (Compound No. 11)

[0591]

[0592] Step 1: (2R,3S)-5-(3-carbamoyl-1H-1,2,4-triazol-1-yl)-2-ethynyl-2-(((4-methylbenzoyl)oxy)methyl)tetrahydrofuran-3-yl 4-methylbenzoate. To a suspension of 1H-1,2,4-triazole-3-carboxamide (57.7 mg, 504 μmol) in MeCN (3.88 mL) was added N,O-bis(trimethylsilyl)acetamide (590 μL, 2.29 mmol), and the resulting mixture was heated to 70° C. and stirred for about 1 h. The mixture was then cooled to 0°C and a solution of (2R,3S)-5-acetoxy-2-ethynyl-2-(((4-methylbenzoyl)oxy)methyl)tetrahydrofuran-3-yl 4-methylbenzoate (200 mg, 458 μmol) in MeCN (3.88 mL) was added dropwise with stirring, followed by trimethylsilyl trifluoromethanesulfonate (127 μL, 687 μmol). The reaction mixture was then warmed to 70°C, stirred for about 18 h, and then cooled to ambient temperature. The mixture was stirred with saturated NaHCO 3The reaction was quenched with aqueous solution (10 mL), diluted with saturated aqueous NaCl (12 mL), and then the organics were extracted with EtOAc (3 x 30 mL). The combined organics were dried (anhydrous Na 2 SO 4 ), filtered and concentrated under vacuum. 2 The residue was purified on 4% paraffin (eluent: 50-100% EtOAc in heptane) to provide the title compound (132 mg, 270 μmol, 59%) as a yellow oil, a mixture of anomeric forms. For the more polar anomeric form, LC-MS (ESI) m / z 489.2 [M+H]+. LC-MS RT = 1.52 min; Method G. For the less polar anomeric form, LC-MS (ESI) m / z 489.2 [M+H]+. LC-MS RT = 1.54 min; Method G.

[0593] Step 2: 1-((4S,5R)-5-ethynyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-1H-1,2,4-triazole-3-carboxamide. To a solution of (2R,3S)-5-(3-carbamoyl-1H-1,2,4-triazol-1-yl)-2-ethynyl-2-(((4-methylbenzoyl)oxy)methyl)tetrahydrofuran-3-yl 4-methylbenzoate (132 mg, 270 μmol) in MeOH (901 μmol) was added dropwise 28% NH 4 OH aqueous solution (60 μL), and the resulting mixture was stirred at ambient temperature for about 18 h. After this time, another portion of 28% NH 4 OH aqueous solution (60 μL), the mixture was heated to 45 ° C and stirred for about 4 hours. The mixture was then cooled to ambient temperature and directly purified by reverse phase chromatography on C18 (eluent: 0-100% MeCN in 10 mM ammonium formate solution) to provide compound No. 11 (61.3 mg, 243 μmol, 90%) as a yellow oil in the form of a mixture of about 2:1 anomeric compounds. For the major anomeric compound, 1 H NMR (400MHz, CD 3 OD) δ8.72 (s, 1H), 6.30 (dd, J = 7.2, 3.0 Hz, 1H), 4.72 (dd, J = 8.6, 6.8 Hz, 1H), 3.80 (d, J = 12.2 Hz, 1H), 3.72 (d, J = 12.2 Hz, 1H), 3.09 (s, 1H), 2.75 (ddd, J = 13.3, 6.8, 3.0 Hz, 1H), 2.70-2.64 (m, 1H). For the minor anomer, 1 H NMR (400MHz, CD 3OD)δ8.79(s,1H),6.24(dd,J=7.1,3.6Hz,1H),4.47(dd,J=6.7,4.6Hz,1H),3.73(d,J=12.0Hz,1H),3.67(d,J =12.1Hz,1H),3.15(s,1H),2.93(dt,J=13.9,6.9Hz,1H),2.61(ddd,J=14.0,4.6,3.7Hz,1H).LC-MS(ESI)m / z 253.1[M+H]+.LC-MS RT=0.13min; Method A.

[0594] Embodiment 33

[0595] Synthesis of 4-amino-5-bromo-1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-methyltetrahydrofuran-2-yl)pyrimidin-2(1H)-one (Compound No. 15)

[0596]

[0597] Step 1: (2R,3R,4S,5R)-2-(4-amino-5-bromo-2-oxopyrimidin-1(2H)-yl)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-methyltetrahydrofuran-3-yl acetate. 2 Under 4-amino-5-bromo-1H-pyrimidin-2-one (0.38g, 2.00mmol) in MeCN (15mL), N, O-bis (trimethylsilyl) acetamide (1.22g, 6.00mmol) was added at one time. The mixture was then stirred at 70 ° C for 1 hour. After cooling to 25 ° C, TMSOTf (390mg, 1.76mmol) and (3R, 4S, 5R) -4- (benzyloxy) -5- ((benzyloxy) methyl) -5-methyltetrahydrofuran-2,3-diyl diacetate (0.5g, 1.17mmol) were added dropwise at 25 ° C. The resulting mixture was stirred at 25 ° C for 16 hours. The reaction was then quenched by adding 10 wt% aqueous citric acid solution (10mL) and extracted with EtOAc (20mL). The organic layer was washed with brine (15mL) and dried over anhydrous Na 2 SO 4 Dry, filter and concentrate.The residue was purified by column chromatography on silica gel (column height: 250 mm, diameter: 100 mm, 100-200 mesh silica gel, DCM / MeOH=50 / 1) to give the title compound as a yellow oil (0.64 g, 98% yield). 1 H NMR (400 MHz, CDCl 3)δ8.48(s,1H),7.40-7.20(m,10H),6.08(d,J=2.4Hz,1H),5.49-5.45(m,1H),4.63(d,J=12Hz,1H),4.52-4.47(m,1H),4.45-4.39 (m,1H),4.36(d,J=11.6Hz,1H),4.27(d,J=6.0Hz,1H),3.60(d,J=10.4Hz,1H),3.31(d,J=10.4Hz,1H),2.14(s,3H),1.28(s,3H).

[0598] Step 2: 4-amino-1-((2R,3R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-3-hydroxy-5-methyltetrahydrofuran-2-yl)-5-bromopyrimidin-2(1H)-one. To a mixture of (2R,3R,4S,5R)-2-(4-amino-5-bromo-2-oxopyrimidin-1(2H)-yl)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-methyltetrahydrofuran-3-yl acetate (0.638 g, 1.14 mmol) in MeOH (15 mL) was added aqueous NaOH (2M, 2 mL) in one portion at 25°C. The mixture was stirred at 25°C for 30 min. After this time, the reaction mixture was concentrated. The residue was purified by column chromatography on silica gel (column height: 250 mm, diameter: 100 mm, 100-200 mesh silica gel, dichloromethane / MeOH = 100 / 0 to 50 / 1) to afford the title compound (0.5 g, 85% yield) as a white solid. 1 H NMR (400 MHz, CDCl 3 )δ8.25(s,1H),7.40-7.23(m,10H),6.58(s,1H),5.93(d,J=3.6Hz,1H),5.66(s,1H),4.77(d,J=12.0Hz,1H),4.6 0-4.46(m,3H),4.40-4.30(m,1H),4.11(d,J=5.6Hz,1H),3.61-3.51(m,2H),3.37(d,J=10.4Hz,1H),1.35(s,3H).

[0599] Step 3: O-((2R,3R,4S,5R)-2-(4-amino-5-bromo-2-oxopyrimidin-1(2H)-yl)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-methyltetrahydrofuran-3-yl)O-phenylcarbonothioate. To a mixture of 4-amino-1-((2R,3R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-3-hydroxy-5-methyltetrahydrofuran-2-yl)-5-bromopyrimidin-2(1H)-one (0.5 g, 968.27 μmol) in MeCN (15 mL) was added DMAP (355 mg, 2.90 mmol) and then heated at 25 °C under N 2 To the 4-thiazolyl-1-yl thiocyanate (0.54g, 1.2mmol) was added dropwise O-phenyl chlorothiocarbonate (251mg, 1.45mmol) at 25°C. After stirring for 1 hour at 25°C, the mixture was concentrated under reduced pressure. The residue was purified by column chromatography on silica gel (column height: 250mm, diameter: 100mm, 100-200 mesh silica gel, DCM / MeOH=100 / 0 to 99 / 1) to produce the title compound (0.55g, 87% yield) as a light yellow solid. 1 H NMR (400 MHz, CDCl 3 )δ8.32(s,1H),7.49-7.28(m,14H),7.06-7.01(m,2H),6.32(d,J=3.2Hz,1H),6.05-5.95(m,1H),5.68(s,1H),4.75( d,J=11.6Hz,1H),4.63-4.53(m,1H),4.50-4.42(m,3H),3.63(d,J=10.8Hz,1H),3.38(d,J=10.4Hz,1H),1.30(s,3H).

[0600] Step 4: 4-amino-1-((2R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-methyltetrahydrofuran-2-yl)-5-bromopyrimidin-2(1H)-one. 2 To a mixture of O-((2R,3R,4S,5R)-2-(4-amino-5-bromo-2-oxopyrimidin-1(2H)-yl)-4-(benzyloxy)-5-((benzyloxy)-5-methyltetrahydrofuran-3-yl)O-phenylcarbonothioate in toluene (12 mL) was added AIBN (69 mg, 421.42 μmol) and TTMSS (1.05 g, 4.21 mmol) in one portion. The mixture was heated to 110° C. and stirred for 2 hours. The reaction mixture was then cooled to 25° C. and concentrated. The residue was purified by preparative TLC (DCM / MeOH=15 / 1) to give the title compound (170 mg, 40% yield) as a colorless oil.

[0601] Step 5: 4-amino-5-bromo-1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-methyltetrahydrofuran-2-yl)pyrimidin-2(1H)-one (Compound No. 15). 2 To a mixture of 4-amino-1-((2R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-methyltetrahydrofuran-2-yl)-5-bromopyrimidin-2(1H)-one (0.15 g, 299.77 μmol) in DCM (9 mL) was added dropwise BCl 3 (1 M, 2.1 mL). The reaction mixture was stirred at -78 °C for 15 min, then warmed to -40 °C and stirred for another 0.5 h. The mixture was then stirred by adding MeOH (1 mL) and NH 4 The reaction was quenched with 1H-[4-(4-[4-(4-oxo-1-yl)-2-nitropropene]propene (1 mL). After stirring at -40 °C for another 10 min, the mixture was allowed to warm to room temperature and stirred for 10 min. After this, the reaction mixture was directly concentrated. The residue was purified by preparative HPLC (basic) to yield compound No. 15 (2.7 mg, 3% yield) as a white solid. 1 HNMR (400MHz, CD 3 OD)δ8.59(s,1H),6.12-6.03(m,1H),4.36(t,J=6.8Hz,1H),3.67-3.54(m,2H),2.53-2.42(m,1H),2.31-2.21(m,1H),1.17(s,3H).LC-MS(ESI)m / z 661.0[2M+Na]+.LC-MS RT=1.420min; Method D.

[0602] Embodiment 34

[0603] Synthesis of (2R,3S,5R)-5-(4-amino-5-bromo-2-oxopyrimidin-1(2H)-yl)-3-hydroxy-2-(hydroxymethyl)tetrahydrofuran-2-carbonitrile (Compound No. 16)

[0604]

[0605] Step 1: (2R,3R,4S,5R)-2-(4-amino-5-bromo-2-oxopyrimidin-1(2H)-yl)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-cyanotetrahydrofuran-3-yl acetate. To a solution of 4-amino-5-bromo-1H-pyrimidin-2-one (400 mg, 2.1 mmol) in acetonitrile (15 mL) was added N,O-bis(trimethylsilyl)acetamide (1.6 mL, 6.3 mmol), the reaction mixture was stirred at 70° C. for 1 h and cooled to room temperature. Trimethylsilyl trifluoromethanesulfonate (490 μL, 2.7 mmol) was added, followed by (3R, 4S, 5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-cyanotetrahydrofuran-2,3-diyl diacetate (600 mg, 1.4 mmol) in acetonitrile (8 mL). After stirring at 105 °C for 16 h, the reaction was quenched with water (20 mL), extracted with EtOAc (30 mL), washed with brine (20 mL), and purified by Na 2 SO 4 Dried, filtered and concentrated.The residue was purified by silica gel chromatography eluting with 0-3% MeOH in DCM to afford the title product as a brown oil (302 mg, 0.53 mmol, 39% yield). 1 H NMR (400 MHz, CDCl 3 ): δ7.99(s,1H),7.38-7.30(m,8H),7.25-7.18(m,2H),5.97(s,1H),5.62(d,J=5.2Hz,1H),4.69( d,J=11.6Hz,1H),4.51-4.42(m,4H),3.90(d,J=10.8Hz,1H),3.61(d,J=10.8Hz,1H),2.18(s,3H).

[0606] Step 2: (2R,3S,4R,5R)-5-(4-amino-5-bromo-2-oxopyrimidin-1(2H)-yl)-3-(benzyloxy)-2-((benzyloxy)methyl)-4-hydroxytetrahydrofuran-2-carbonitrile. To a solution of (2R,3R,4S,5R)-2-(4-amino-5-bromo-2-oxopyrimidin-1(2H)-yl)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-cyanotetrahydrofuran-3-yl acetate (300 mg, 526.9 μmol) in dioxane (8 mL) was added aqueous NaOH (1 M, 1.0 mL) at 25°C. The resulting reaction mixture was stirred at 25°C for 1 h. The reaction was then quenched with water (5 mL), extracted with EtOAc (10 mL), washed with brine (10 mL), and purified by Na 2 SO 4Dried, filtered and concentrated.The residue was purified by silica gel chromatography eluting with 0-3% MeOH in DCM to afford the title product as a white solid (260 mg, 94% yield).

[0607] Step 3: A solution of (2R,3S,4R,5R)-5-(4-amino-5-bromo-2-oxo-pyrimidin-1-yl)-3-benzyloxy-2-(benzyloxymethyl)-4-hydroxy-tetrahydrofuran-2-carbonitrile (260.0 mg, 493.0 μmol) in acetonitrile (5 mL) was stirred for 1 h. 2 Degas 3 times. Then add DMAP (78.3 mg, 640.9 μmol) and O-phenyl chlorothioate (170.2 mg, 986.0 μmol). The resulting reaction mixture was stirred at 25 ° C for 1 h. After this, the reaction mixture was diluted with water (5 mL), extracted with EtOAc (10 mL), washed with brine (10 mL), and purified by Na 2 SO 4 Dry, filter and concentrate. The residue is purified by silica gel chromatography eluting with 0-3% MeOH in DCM to afford the title product as a brown oil (251.0 mg, 378 μmol, 76% yield). LC-MS (ESI) m / z 663.0 [M+H]+. LC-MS RT=1.025 min; Method C.

[0608] Step 4: (2R,3S,5R)-5-(4-amino-5-bromo-2-oxopyrimidin-1(2H)-yl)-3-(benzyloxy)-2-((benzyloxy)methyl)tetrahydrofuran-2-carbonitrile. A solution of (2R,3S,4R,5R)-5-(4-amino-5-bromo-2-oxopyrimidin-1-yl)-3-benzyloxy-2-(benzyloxymethyl)-4-hydroxy-tetrahydrofuran-2-carbonitrile (250 mg, 376.8 μmol) in toluene (5 mL) was heated to 40 ℃ and 10 ℃ of 30 ℃. 2 The mixture was stirred for 5 min. Bis(trimethylsilyl)silyl-trimethylsilane (281.1 mg, 1.13 mmol) and AIBN (29.0 mg, 176.8 μmol) were added. The reaction mixture was stirred at 110 ° C for 2 h. The reaction mixture was concentrated. The residue was purified by silica gel chromatography eluting with 0-3% MeOH in DCM to provide the title product as a white solid (95.0 mg, 186 μmol, 49% yield).

[0609] Step 5: (2R,3S,5R)-5-(4-amino-5-bromo-2-oxopyrimidin-1(2H)-yl)-3-hydroxy-2-(hydroxymethyl)tetrahydrofuran-2-carbonitrile (Compound No. 16). A solution of (2R,3S,5R)-5-(4-amino-5-bromo-2-oxopyrimidin-1(2H)-yl)-3-(benzyloxy)-2-((benzyloxy)methyl)tetrahydrofuran-2-carbonitrile (95.0 mg, 186 μmol) in DCM (3 mL) was treated with N 2 Degas 3 times and cool to -78 °C, then add BCl 3 (1 M in DCM, 0.5 mL). The resulting reaction mixture was stirred at -45 °C for 0.5 h. The reaction was then quenched with MeOH (1 mL) at -45 °C and warmed to 0 °C. The reaction was washed with 1 mL of NH 3 ·H 2 O (purity 28%) adjusted pH>7 and concentrated to a residue. 3 ·H 2 O+NH 4 HCO 3 The residue was purified by evaporation (4% by weight) in water to afford the title compound (3.5 mg, 10.6 μmol, 6% yield) as a white solid. 1 HNMR (400 MHz, MeOD-d 4 ):δ8.23(s,1H),6.35-6.26(m,1H),4.59(t,J=6.8Hz,1H),4.01-3.86(m,2H),2.55-2.37(m,2H).LC-MS(ESI)m / z 331.0[M+H]+.LC-MS RT=1.312min; Method F.

[0610] Embodiment 35

[0611] Synthesis of 1-((2R,5R)-5-ethynyl-5-(hydroxymethyl)-2,5-dihydrofuran-2-yl)-5-fluoropyrimidine-2,4(1H,3H)-dione (Compound No. 19)

[0612]

[0613] Step 1: Benzoic acid ((2R,4R,5R)-5-(benzoyloxy)-2-ethynyl-4-(p-tolylthio)tetrahydrofuran-2-yl)methyl ester. A solution of TBAF (1M in THF, 15.4 mL, 15.4 mmol) was added to a stirred solution of benzoic acid ((2R,4R,5R)-5-(benzoyloxy)-4-(p-tolylthio)-2-((trimethylsilyl)ethynyl)tetrahydrofuran-2-yl)methyl ester (7.01 g, 12.9 mmol) in THF (50.0 mL) at ambient temperature and the resulting mixture was stirred at this temperature for about 1 h. The reaction was then heated to 40° C. with 20% NH 4 The mixture was quenched with aqueous OAc and concentrated in vacuo. 2 The residue was purified on celite (eluent: 0-30% EtOAc / hexanes) to provide the title compound as a white solid (2.5 g, 5.30 mmol, 41%). LC-MS (ESI) m / z 490.2 [M+H 2 O]+. LC-MS RT=2.02 min; Method A.

[0614] Step 2: ((2R,4R,5R)-2-ethynyl-5-(5-fluoro-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)-4-(p-tolylthio)tetrahydrofuran-2-yl)methyl benzoate. To a flame-dried round-bottom flask equipped with a magnetic stir bar under nitrogen was added 5-fluorouracil (231 mg, 1.78 mmol) and MeCN (3.00 mL). Both HMDS (201 μL, 1.78 mmol) and trimethylsilyl trifluoromethanesulfonate (511 μL, 2.79 mmol) were added and the resulting mixture was stirred at ambient temperature for about 1 h. Solid ((2R,4R,5R)-5-(benzoyloxy)-2-ethynyl-4-(p-tolylthio)tetrahydrofuran-2-yl)methyl benzoate (600 mg, 1.27 mmol) was added in one portion and the resulting mixture was stirred at ambient temperature overnight. The reaction was stirred with K 3 PO 4 (138 mg, 635 μmol) in water (1.00 mL) was quenched. The organics were then extracted with EtOAc and dried (anhydrous Na 2 SO 4 ), filtered and concentrated under vacuum. 2 The residue was purified above (eluent: 0-90% EtOAc in hexanes) to provide the title compound (510 mg, 1.06 mmol, 84%) as a white solid. LC-MS (ESI) m / z 479.3 [MH]-. LC-MS RT = 1.65 min; Method A.

[0615] Step 3: ((2R,5R)-2-ethynyl-5-(5-fluoro-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)-2,5-dihydrofuran-2-yl)methyl benzoate. ((2R,4R,5R)-2-ethynyl-5-(5-fluoro-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)-4-(p-tolylthio)tetrahydrofuran-2-yl)methyl benzoate (510 mg, 1.06 mmol) was stirred and dissolved in MeCN (8.00 mL), and chloramine-T trihydrate (366 mg, 1.27 mmol) and AcOH (6.14 μL, 106 μmol) were added. The reaction mixture was stirred at ambient temperature for about 2.5 h and then quenched by adding 10 wt% NH 4 The reaction was quenched with aqueous OAc (5 mL). The phases were separated and the organic matter was dried (anhydrous Na 2 SO 4 ), filtered and concentrated under vacuum. The residue was dissolved in n-BuOH (8.00 mL) and the resulting solution was heated to 95 °C for about 3 h. The reaction mixture was then cooled to ambient temperature, the volatiles were removed under vacuum and the residue was purified by flash chromatography on SiO 2 The residue was purified on HPLC (eluent: 0-95% EtOAc in hexanes) to provide the title compound (200 mg, 0.56 mmol, 53%) as a clear oil. LC-MS (ESI) m / z 374.4 [M+H 2 O]+. LC-MSRT=0.98 min; Method A.

[0616] Step 4: 1-((2R,5R)-5-ethynyl-5-(hydroxymethyl)-2,5-dihydrofuran-2-yl)-5-fluoropyrimidine-2,4(1H,3H)-dione (Compound No. 19). Benzoic acid ((2R,5R)-2-ethynyl-5-(5-fluoro-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)-2,5-dihydrofuran-2-yl)methyl ester (200 mg, 561 μmol) was dissolved in MeOH (25 mL), stirred at ambient temperature, and DBU (4.3 μL, 28.1 μmol) was added. The resulting mixture was heated to 60 °C until the reaction was determined to be complete by LCMS. The mixture was cooled to ambient temperature, adsorbed onto silica gel, and purified by flash chromatography on SiO 2 Purification on 4% 4% 4% (eluent: 0-10% MeOH in DCM) afforded Compound No. 19 (76 mg, 300 μmol, 54%) as a white solid. 1HNMR (400MHz, DMSO-d6) δ11.92(s,1H),8.01(d,J=7.2Hz,1H),6.87(dt,J=2.0,1.5Hz,1H),6.37(dd,J=5.8,2.0Hz,1H),6.05(dd,J=5.8,1. 3Hz,1H),5.60(t,J=5.8Hz,1H),3.71(dd,J=12.1,5.5Hz,1H),3.70(s,1H),3.61(dd,J=12.1,6.1Hz,1H).LC-MS(ESI)m / z251.3[MH]-.LC-MS RT=0.96min; Method B.

[0617] Embodiment 36

[0618] Synthesis of 1-((2S,5R)-5-ethyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)pyrimidine-2,4(1H,3H)-dione (Compound No. 20)

[0619]

[0620] Step 1: (2R,3S)-5-(5-bromo-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)-2-ethynyl-2-(((4-methylbenzoyl)oxy)methyl)tetrahydrofuran-3-yl 4-methylbenzoate. To a solution of N,O-bis(trimethylsilyl)acetamide (104 μL, 687 μmol) in MeCN (2.00 mL) was added 5-bromouracil (44.7 mg, 229 μmol), and the resulting mixture was heated to 70°C and stirred for about 1 h. Then trimethylsilyl trifluoromethanesulfonate (50.3 μL, 275 μmol) and (2R,3S)-5-acetoxy-2-ethynyl-2-(((4-methylbenzoyl)oxy)methyl)tetrahydrofuran-3-yl 4-methylbenzoate (100 mg, 229 μmol) were added and the reaction mixture was stirred at 70 °C overnight and then cooled to ambient temperature. The reaction was partitioned between water and EtOAc. The combined organics were dried (anhydrous Na 2 SO 4 ), filtered and concentrated under vacuum. 2The residue was purified on 4% paraformaldehyde (eluent: 0-90% EtOAc in heptane) to provide the title compound (121 mg, 213 μmol, 93%) as a clear oil as a mixture of anomeric forms. For the more polar anomeric form, LC-MS (ESI) m / z 568.3 [M+H]+. LC-MS RT = 1.70 min; Method A. For the less polar anomeric form, LC-MS (ESI) m / z 567.3 [MH]-. LC-MS RT = 1.73 min; Method A.

[0621] Step 2: 5-bromo-1-((2S,4S,5R)-5-ethynyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)pyrimidine-2,4(1H,3H)-dione. (2R,3S)-5-(5-bromo-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)-2-ethynyl-2-(((4-methylbenzoyl)oxy)methyl)tetrahydrofuran-3-yl 4-methylbenzoate (454 mg, 213 μmol) was dissolved in MeOH (2.00 mL) and a solution of NaOMe (25 wt% in MeOH, 2.00 mL, 427 μmol) was added with stirring. The reaction mixture was stirred at ambient temperature until the reaction was determined to be complete by LCMS. The reaction mixture was adsorbed onto silica gel and purified by flash chromatography on SiO 2 Purification was carried out as above (eluent: 0-20% MeOH in DCM) to afford the title compound as a mixture of anomers as a white solid (44.9 mg, 13.6 μmol, 64%). Further purification by preparative HPLC afforded the title compound as a single stereoisomer (6.00 mg). 1 H NMR (400MHz, DMSO-d6) δ11.81(br.s,1H),8.32(s,1H),6.09(dd,J=7.2,3.4Hz,1H),5.73(d,J=4.4Hz,1H),5.32(t,J=6.3Hz,1H),4.27(dt ,J=6.1,3.9Hz,1H),3.74(s),3.50(dd,J=11.8,5.9Hz,1H),3.46(dd,J=11.8,6.8Hz,1H),2.74-2.66(m,1H),2.01(dt,J=14.1,3.5Hz,1H).

[0622] Step 3: 5-Bromo-1-((2S,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-vinyltetrahydrofuran-2-yl)pyrimidine-2,4(1H,3H)-dione. In a glass bottle equipped with a magnetic stir bar and covered with a rubber septum, 5-bromo-1-((2S,4S,5R)-5-ethynyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)pyrimidine-2,4(1H,3H)-dione (6.00 mg, 18.1 μmol) was dissolved in MeOH (181 μL) with stirring. Lindela's catalyst (6.00 mg) was then added, and the container was purged with hydrogen from a balloon. The reaction mixture was then stirred under hydrogen (1 atm) at ambient temperature for about 3 h. The reaction mixture was separated by a short stopcock. The mixture was filtered, the filter cake was washed with MeOH, and the filtrate was concentrated under vacuum to provide the crude title compound (4.85 mg, 14.5 μmol, 80%). 1 H NMR (400MHz, CD 3 OD)δ8.45(s,1H),6.34(dd,J=7.8,2.0Hz,1H),6.12(dd,J=17.4,11.2Hz,1H),5.44(dd,J=17.4,1.7Hz,1H),5.35(dd,J=11.2, 1.7Hz, 1H), 4.40 (dd, J = 5.9, 0.9Hz, 1H), 3.51 (d, J = 11.8Hz, 1H), 3.43 (d, J = 11.8Hz, 1H), 2.97-2.89 (m, 1H), 2.06-2.00 (m, 1H).

[0623] Step 4: 1-((2S,4S,5R)-5-ethyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)pyrimidine-2,4(1H,3H)-dione (Compound No. 20). In a glass vial equipped with a magnetic stir bar and covered with a rubber septum, 5-bromo-1-((2S,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-vinyltetrahydrofuran-2-yl)pyrimidine-2,4(1H,3H)-dione (6.04 mg, 18.1 μmol) was dissolved in MeOH (181 μL) under stirring. PtO was then added. 2 (2.06 mg, 9.06 μmol), and the vessel was purged with hydrogen from a balloon. The reaction mixture was then stirred under hydrogen (1 atm) at ambient temperature for about 4 h. The mixture was filtered through a short plug of Celite®, the filter cake was washed with MeOH, and the filtrate was concentrated under vacuum. 2The residue was purified above (eluent: 50-100% EtOAc in heptane, then 100% IPA) to provide Compound No. 20 (2.45 mg, 9.6 μmol, 53%) as an amorphous white solid. 1 H NMR (400MHz, CD 3 OD)δ8.00(d,J=8.1Hz,1H),6.15(dd,J=7.9,2.6Hz,1H),5.67(d,J=8.1Hz,1H),4.22(dd,J=6.0,1.4Hz,1H),3.46(q,J=1 1.8Hz,2H),2.87(ddd,J=14.7,7.9,6.0Hz,1H),2.01-1.92(m,1H),1.59-1.46(m,1H).LC-MS(ESI)m / z255.2[MH]-.LC-MS RT=1.05min; Method H.

[0624] Embodiment 37

[0625] Synthesis of 4-amino-5-bromo-1-((2R,4S,5R)-5-(fluoromethyl)-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)pyrimidin-2(1H)-one (Compound No. 117)

[0626]

[0627] Step 1: (2R,3R,4S,5R)-2-(4-amino-5-bromo-2-oxopyrimidin-1(2H)-yl)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-(fluoromethyl)tetrahydrofuran-3-yl acetate. 2 Under 4-amino-5-bromo-1H-pyrimidin-2-one (0.26g, 1.37mmol) in MeCN (15mL), (1E)-N-trimethylsilylacetamidine acid trimethylsilyl ester (835.14mg, 4.11mmol) was added at one time. The mixture was then stirred at 70°C for 1 hour. After cooling to 25°C, TMSOTf (373.37mg, 1.68mmol) and (2S, 3R, 4S, 5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-(fluoromethyl)tetrahydrofuran-2,3-diyl diacetate (0.5g, 1.12mmol) were added dropwise at 25°C. The resulting mixture was stirred at 25°C for 16 hours. The reaction was then quenched by adding 10 wt% aqueous citric acid solution (10mL) and then extracted with EtOAc (20mL). The organic layer was washed with brine (15 mL) and 2 SO 4Dry, filter and concentrate.The residue was purified by column chromatography on silica gel (column height: 250 mm, diameter: 100 mm, 100-200 mesh silica gel) eluting with 2% MeOH in dichloromethane to give the title compound as a yellow oil (0.326 g, 51% yield). 1 H NMR (400 MHz, CDCl 3 )δ8.26(s,1H),7.40-7.26(m,12H),6.21(d,J=4.0Hz,1H),5.63(s,1H),5.53-5. 45(m,1H),4.72-4.35(m,7H),3.90-3.82(m,1H),3.60-3.53(m,1H),2.11(s,3H).

[0628] Step 2: 4-amino-1-((2R,3R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-(fluoromethyl)-3-hydroxytetrahydrofuran-2-yl)-5-bromopyrimidin-2(1H)-one. To a mixture of (2R,3R,4S,5R)-2-(4-amino-5-bromo-2-oxopyrimidin-1(2H)-yl)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-(fluoromethyl)tetrahydrofuran-3-yl acetate (0.325 g, 563.83 μmol) in MeOH (15 mL) was added aqueous NaOH (2M, 2 mL) in one portion at 25°C. The mixture was stirred at 25°C for 30 min. The reaction mixture was directly concentrated. The residue was purified by column chromatography on silica gel (column height: 250 mm, diameter: 100 mm, 100-200 mesh silica gel) eluting with 2% MeOH in DCM to afford the title compound as a white solid (0.29 g, 96% yield). 1 H NMR (400 MHz, CDCl 3 )δ8.09(s,1H),7.40-7.27(m,10H),6.73(s,1H),5.93(d,J=4.8Hz,1H),5.59(s,1H),4.81-4 .52(m,6H),4.46-4.40(m,1H),4.28(d,J=5.6Hz,1H),3.89-3.76(m,2H),3.68-3.61(m,1H).

[0629] Step 3: O-((2R,3R,4S,5R)-2-(4-amino-5-bromo-2-oxopyrimidin-1(2H)-yl)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-(fluoromethyl)tetrahydrofuran-3-yl)O-phenylcarbonothioate. To a solution of 4-amino-1-((2R,3R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-(fluoromethyl)-3-hydroxytetrahydrofuran-2-yl)-5-bromopyrimidin-2(1H)-one (0.29 g, 542.69 μmol) in MeCN (15 mL) was added DMAP (198.90 mg, 1.63 mmol) and then heated at 25 °C under N 2 The product was stirred for 1 hour at 25 ° C. The mixture was concentrated directly. The residue was purified by column chromatography on silica gel (column height: 250 mm, diameter: 100 mm, 100-200 mesh silica gel) with 1% MeOH in DCM to produce the title compound (0.266 g, 73% yield) as a light yellow solid. 1 H NMR (400 MHz, CDCl 3 )δ8.17(s,1H),7.42-7.28(m,13H),7.05-6.99(m,2H),6.41(d,J=5.2Hz,1H),6.03(t,J=5. 2Hz, 1H), 4.73 (d, J = 11.2Hz, 1H), 4.70-4.50 (m, 6H), 3.91-3.86 (m, 1H), 3.71-3.65 (m, 1H).

[0630] Step 4: 4-amino-1-((2R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-(fluoromethyl)tetrahydrofuran-2-yl)-5-bromopyrimidin-2(1H)-one. 2 To a mixture of O-((2R,3R,4S,5R)-2-(4-amino-5-bromo-2-oxopyrimidin-1(2H)-yl)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-(fluoromethyl)tetrahydrofuran-3-yl)O-phenylcarbonothioate (0.266 g, 396.69 μmol) in toluene (12 mL) was added AIBN (33 mg, 198.35 μmol) and TTMSS (493.21 mg, 1.98 mmol) in one portion. The mixture was heated to 110 °C and stirred for 2 hours. The reaction mixture was then cooled to room temperature and directly concentrated. The residue was purified by preparative TLC (DCM / MeOH=15 / 1) to give the title compound (40 mg, 19% yield) as a colorless oil.

[0631] Step 5: 4-amino-5-bromo-1-((2R,4S,5R)-5-(fluoromethyl)-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)pyrimidin-2(1H)-one (Compound No. 117). Heat at -78 °C under N 2 To a mixture of 4-amino-1-((2R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-(fluoromethyl)tetrahydrofuran-2-yl)-5-bromopyrimidin-2(1H)-one (0.04 g, 77.16 μmol) in DCM (9 mL) was added dropwise BCl 3 (1 M in DCM, 0.5 mL). The reaction mixture was stirred at -78 °C for 15 min, then allowed to warm to -40 °C and stirred for another 0.5 h. The mixture was then heated at -40 °C by adding MeOH (1 mL) and NH 4 The reaction was quenched with 4-(4-(4-(4-oxo-1-yl)-2-nitropropene)-2-nitropropene (10-nitropropene)-4-nitropropene (20-nitropropene)-5-nitropropene (10-nitropropene)-2-nitropropene (20-nitropropene)-5 ... 1 H NMR (400 MHz, CDCl 3 )δ8.42(s,1H),6.24(t,J=6.4Hz,1H),4.68-4.62(m,1H),4.55-4.46(m,2H),3.75-3.71(m,2H),2.55-2.42(m,1H),2.35-2.24(m,1H).

[0632] Embodiment 38

[0633] Synthesis of 5-chloro-1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-methyltetrahydrofuran-2-yl)pyrimidine-2,4(1H,3H)-dione (Compound No. 120)

[0634]

[0635] Step 1: (2R,3R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-2-(5-chloro-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)-5-methyltetrahydrofuran-3-yl acetate. A mixture of 5-chloro-1H-pyrimidine-2,4-dione (0.22 g, 1.50 mmol) and BTMSA (767.51 mg, 4.50 mmol) in MeCN (10 mL) was stirred at 70 °C for 1 hour, and then the reaction mixture was cooled to room temperature. A solution of (0.5 g, 1.17 mmol) in MeCN (10 mL) and TMSOTf (520 mg, 2.34 mmol) were added, and then the mixture was stirred at 30 °C for 16 hours. After this, the reaction was diluted with EtOAc (50 mL), washed with brine (30 mL x 2), and concentrated to give the crude title compound as a colorless oil (0.5 g, yield 83%). LC-MS (ESI) m / z 537.0 [M+Na]+. LC-MS RT = 0.901 min; Method C.

[0636] Step 2: 1-((2R,3R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-3-hydroxy-5-methyltetrahydrofuran-2-yl)-5-chloropyrimidine-2,4(1H,3H)-dione. A mixture of (2R,3R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-2-(5-chloro-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)-5-methyltetrahydrofuran-3-yl acetate (0.5 g, 970.96 μmol) and NaOH (2M, 1 mL) in MeOH (10 mL) was stirred at 20°C for 0.2 h. The reaction was concentrated and diluted with EtOAc (50 mL), washed with brine (30 mL x 2), and then concentrated to provide the crude title compound (0.4 g, yield 87%) as a white solid, which was used directly in the next step.

[0637] Step 3: O-((2R,3R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-2-(5-chloro-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)-5-methyltetrahydrofuran-3-yl)O-phenylcarbonothioate. To a solution of 1-((2R,3R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-3-hydroxy-5-methyltetrahydrofuran-2-yl)-5-chloropyrimidine-2,4(1H,3H)-dione (0.2 g, 422.91 μmol) and DMAP (155 mg, 1.27 mmol) in MeCN (20 mL) was added O-phenyl chlorocarbonothioate (109 mg, 634.36 μmol), followed by stirring at 20°C for 1 hour. The reaction mixture was then diluted with EtOAc (50 mL), washed with brine (30 mL x 2), concentrated to give a crude product. The residue was purified by column chromatography (DCM / MeOH = 100 / 1 to 100 / 5) to afford the title compound (0.23 g, 89% yield) as a colorless solid.

[0638] Step 4: 1-((2R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-methyltetrahydrofuran-2-yl)-5-chloropyrimidine-2,4(1H,3H)-dione. A mixture of O-((2R,3R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-2-(5-chloro-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)-5-methyltetrahydrofuran-3-yl)-O-phenylcarbonothioate (0.33 g, 541 μmol), TTMSS (673.6 mg, 2.71 mmol) and AIBN (44 mg, 270 μmol) in toluene (15 mL) was stirred at 110° C. for 2 h. After cooling to 20° C., the reaction mixture was concentrated to give a crude product. The residue was purified by column chromatography (DCM:MeOH=100 / 1 to 20 / 1) to afford the title compound (0.11 g, 44% yield) as a colorless oil. LC-MS (ESI) m / z 479.0 [M+Na]+. LC-MS RT=0.892 min; Method C.

[0639] Step 5: 5-chloro-1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-methyltetrahydrofuran-2-yl)pyrimidine-2,4(1H,3H)-dione (Compound No. 120). 1-((2R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-methyltetrahydrofuran-2-yl)-5-chloropyrimidine-2,4(1H,3H)-dione (50 mg, 109.43 μmol) and PdCl 2A mixture of (9.7 mg, 54.71 μmol) in MeOH (10 mL) was heated at 26 °C under H 2 (15 psi) for 20 min. Afterwards, the reaction solution was filtered and the filtrate was concentrated to give a crude product. The residue was purified by preparative HPLC (acetonitrile 0-27% / FA-MeCN in water) to provide Compound No. 120 (13.9 mg, 46% yield) as a white solid. 1 H NMR (400 MHz, MeOD-d 4 ): δ8.45(s,1H),6.17-6.14(m,1H),4.41-4.39(m,1H),3.64-3.57(m,2H),2.42-2.34(m,2H),1.16(s,3H).

[0640] Embodiment 39

[0641] Synthesis of 1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-methyltetrahydrofuran-2-yl)-5-methylpyrimidine-2,4(1H,3H)-dione (Compound No. 122)

[0642]

[0643] Step 1: (2R,3R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-methyl-2-(5-methyl-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)tetrahydrofuran-3-yl acetate. 2 Under 40 ℃, to a mixture of 5-methyl-1H-pyrimidine-2,4-dione (0.2 g, 1.59 mmol) in MeCN (15 mL) was added N, O-bis (trimethylsilyl) acetamide (811 mg, 4.76 mmol) at one time. The mixture was then stirred at 70 ° C for 1 hour. After cooling to 25 ° C, TMSOTf (519 mg, 2.33 mmol) and (2S, 3R, 4S, 5R) -4- (benzyloxy) -5- ((benzyloxy) methyl) -5-methyltetrahydrofuran -2,3- diyl diacetate (0.5 g, 1.17 mmol) were added at 25 ° C. The resulting mixture was stirred at 25 ° C for 16 hours. The reaction was then quenched by adding 10 wt % aqueous citric acid solution (10 mL) and then extracted by EtOAc (20 mL). The organic layer was washed with brine (15 mL) and purified by Na 2 SO 4 Drying, filtration and concentration gave the title compound as a yellow oil (0.5 g, 87% yield). LC-MS (ESI) m / z 517.1 [M+Na]+. LC-MS RT=0.876 min; Method C.

[0644] Step 2: 1-((2R,3R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-3-hydroxy-5-methyltetrahydrofuran-2-yl)-5-methylpyrimidine-2,4(1H,3H)-dione. To a mixture of (2R,3R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-methyl-2-(5-methyl-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)tetrahydrofuran-3-yl acetate (0.5 g, 1.01 mmol) in MeOH (15 mL) was added aqueous NaOH (2M, 2 mL) in one portion at 25 °C. The mixture was stirred at 25 °C for 30 min. The reaction mixture was concentrated directly to afford the title compound as a white solid (0.45 g, 98% yield).

[0645] Step 3: O-((2R,3R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-methyl-2-(5-methyl-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)tetrahydrofuran-3-yl)O-phenylcarbonothioate. To a mixture of 1-((2R,3R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-3-hydroxy-5-methyltetrahydrofuran-2-yl)-5-methylpyrimidine-2,4(1H,3H)-dione (0.35 g, 773.48 μmol) in MeCN (15 mL) was added DMAP (283.5 mg, 2.32 mmol) and then heated at 25 °C under N 2 O-phenylthiochloroformate (200 mg, 1.16 mmol) was added dropwise at 25 °C. After stirring for 1 hour at 25 °C, the mixture was directly concentrated. The residue was purified by silica gel chromatography (column height: 250 mm, diameter: 100 mm, 100-200 mesh silica gel, DCM / MeOH=99 / 1) to produce the title compound as a light yellow solid (0.4 g, yield 88%). LC-MS (ESI) m / z 611.1 [M+Na]+. LC-MS RT=0.981 min; Method C.

[0646] Step 4: 1-((2R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-methyltetrahydrofuran-2-yl)-5-methylpyrimidine-2,4(1H,3H)-dione. Heat at 25°C under N 2To a mixture of O-((2R, 3R, 4S, 5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-methyl-2-(5-methyl-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)tetrahydrofuran-3-yl)O-phenylcarbonothioate (0.2 g, 339.75 μmol) in toluene (12 mL) was added AIBN (28 mg, 169.87 μmol) and TTMSS (422 mg, 1.70 mmol) in one portion. The mixture was heated to 110 ° C and stirred for 2 hours. The reaction mixture was then concentrated directly. The residue was purified by preparative TLC (DCM / MeOH=99 / 1) to give the title compound (100 mg, 67% yield) as a colorless oil. LC-MS (ESI) m / z 437.0 [M+Na]+. LC-MS RT=0.886 min; Method C.

[0647] Step 5: 1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-methyltetrahydrofuran-2-yl)-5-methylpyrimidine-2,4(1H,3H)-dione (Compound No. 122). 2 To a mixture of 1-((2R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-methyltetrahydrofuran-2-yl)-5-methylpyrimidine-2,4(1H,3H)-dione (0.08 g, 183.28 μmol) in MeOH (2 mL) was added PdCl at 15 psi. 2 (3 mg). The mixture was stirred at 25°C for 20 min. After that, the mixture was directly filtered and concentrated. The residue was purified by preparative HPLC (FA) to provide compound No. 122 as a white solid (10.2 mg, 22% yield). 1 H NMR (400MHz, CD 3 OD)δ7.90(s,1H),6.21(t,J=6.4Hz,1H),4.40(t,J=6.0Hz,1H),3.66-3.51(m,2H),2.34(t,J=6.4Hz,2H),1.88(s,3H),1.17(s,3H).

[0648] Embodiment 40

[0649] Synthesis of 4-amino-5-chloro-1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-methyltetrahydrofuran-2-yl)pyrimidin-2(1H)-one (Compound No. 131)

[0650]

[0651] Step 1: (2R,3R,4S,5R)-2-(4-amino-5-chloro-2-oxopyrimidin-1(2H)-yl)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-methyltetrahydrofuran-3-yl acetate. 2 Under 4-amino-5-chloro-1H-pyrimidin-2-one (0.21g, 1.44mmol) in MeCN (15mL), N, O-bis (trimethylsilyl) acetamide (881mg, 4.33mmol) was added at one time. The mixture was then stirred at 70°C for 1 hour. After cooling to 25°C, TMSOTf (389mg, 1.75mmol) and (2S, 3R, 4S, 5R) -4- (benzyloxy) -5- ((benzyloxy) methyl) -5-methyltetrahydrofuran-2,3-diyl diacetate (0.5g, 1.17mmol) were added dropwise at 25°C. The resulting mixture was stirred at 25°C for 16 hours. The reaction was then quenched by adding 10 wt% aqueous citric acid solution (10mL), followed by extraction with EtOAc (20mL). The organic layer was washed with brine (15mL) and purified by Na 2 SO 4 Dried, filtered and concentrated. The residue was purified by column chromatography on silica gel (column height: 250 mm, diameter: 100 mm, 100-200 mesh silica gel) eluted with 2% MeOH in DCM to give the title compound as a yellow oil (0.59 g, 98% yield). LC-MS (ESI) m / z 236.1 [M+Na]+. LC-MS RT = 0.846 min; Method C.

[0652] Step 2: 4-amino-1-((2R,3R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-3-hydroxy-5-methyltetrahydrofuran-2-yl)-5-chloropyrimidin-2(1H)-one. To a mixture of (2R,3R,4S,5R)-2-(4-amino-5-chloro-2-oxopyrimidin-1(2H)-yl)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-methyltetrahydrofuran-3-yl acetate (0.55 g, 1.07 mmol) in MeOH (15 mL) was added aqueous NaOH (2M, 2 mL) in one portion at 25°C. The mixture was stirred at 25°C for 30 min. The reaction mixture was then concentrated. The residue was purified by column chromatography on silica gel (column height: 250 mm, diameter: 100 mm, 100-200 mesh silica gel) eluting with 2% MeOH in DCM to give the title compound as a white solid (0.5 g, 99% yield).

[0653] Step 3: O-((2R,3R,4S,5R)-2-(4-amino-5-chloro-2-oxopyrimidin-1(2H)-yl)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-methyltetrahydrofuran-3-yl)O-phenylcarbonothioate. To a mixture of 4-amino-1-((2R,3R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-3-hydroxy-5-methyltetrahydrofuran-2-yl)-5-chloropyrimidin-2(1H)-one (0.5 g, 1.06 mmol) in MeCN (15 mL) was added DMAP (388 mg, 3.18 mmol) and then heated at 25 °C under N 2 The product was stirred for 1 hour at 25 ° C. The mixture was concentrated directly. The residue was purified by column chromatography on silica gel (column height: 250 mm, diameter: 100 mm, 100-200 mesh silica gel) with 1% MeOH in DCM to provide the title compound (0.5 g, 78% yield) in the form of a light yellow solid. 1 H NMR (400 MHz, DMSO-d 6 )δ8.22(s,1H),7.42-7.28(m,13H),7.07-7.01(m,2H),6.33(d,J=3.2Hz,1H),6.02-5.96(m,1H),5.58(s,1H),4.75( d,J=11.6Hz,1H),4.58-4.52(m,1H),4.50-4.40(m,3H),3.64(d,J=10.4Hz,1H),3.38(d,J=10.4Hz,1H),1.31(s,3H).

[0654] Step 4: 4-amino-1-((2R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-methyltetrahydrofuran-2-yl)-5-chloropyrimidin-2(1H)-one. 2To a mixture of O-((2R, 3R, 4S, 5R)-2-(4-amino-5-chloro-2-oxopyrimidin-1(2H)-yl)-4-(benzyloxy)-5-((benzyloxy)-5-methyltetrahydrofuran-3-yl)-5-methyltetrahydrofuran-3-yl)O-phenylcarbonothioate (0.5 g, 822.23 μmol) in toluene (12 mL) was added AIBN (68 mg, 411.11 μmol) and TTMSS (1.02 g, 4.11 mmol) in one portion. The mixture was heated to 110 ° C and stirred for 2 hours. The reaction mixture was then concentrated directly. The residue was purified by preparative TLC (DCM / MeOH=20 / 1) to give the title compound (80 mg, 21% yield) as a colorless oil.

[0655] Step 5: 4-amino-5-chloro-1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-methyltetrahydrofuran-2-yl)pyrimidin-2(1H)-one (Compound No. 131). 2 To a mixture of 4-amino-1-((2R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-methyltetrahydrofuran-2-yl)-5-chloropyrimidin-2(1H)-one (0.02 g, 43.87 μmol) in MeOH (2 mL) was added PdCl at 15 psi. 2 (778 ug). The mixture was stirred at 25° C. for 0.3 h. After that, the mixture was directly filtered and concentrated. The residue was purified by preparative HPLC (acetonitrile 0-27 / 0.225% FA in water) to provide compound No. 131 as a white solid (2.6 mg, 7% yield). 1 H NMR (400MHz, CD 3 OD)δ8.50(s,1H),6.15-6.05(m,1H),4.36(t,J=6.4Hz,1H),3.68-3.51(m,2H),2.55-2.41(m,1H),2.35-2.20(m,1H),1.17(s,3H).

[0656] Embodiment 41

[0657] Synthesis of ((((3R,5R)-5-(6-amino-9H-purin-9-yl)tetrahydrofuran-3-yl)oxy)methyl)phosphonic acid (Compound No. 113)

[0658]

[0659] Step 1: (2R,3R,4S)-2-(6-benzamido-9H-purin-9-yl)-4-(tert-butyldimethylsilyl)oxy)tetrahydrofuran-3-yl benzoate. A mixture of N-(9H-purin-6-yl)benzamide (1.0 g, 4.2 mmol) in acetonitrile (20 mL) was heated to 40 ℃ with N 2 Degas 3 times. Then add N, O-bis (trimethylsilyl) acetamide (2.2g, 10.5mmol). The resulting mixture was stirred at 70 ° C for 1 hour and then cooled to room temperature. After this, trimethylsilyl trifluoromethanesulfonate (873.0mg, 3.9mmol) was added, followed by benzoic acid (3R, 4S) -2-acetoxy-4- (tert-butyldimethylsilyl) oxy) tetrahydrofuran-3-yl ester (1.3g, 3.4mmol) in acetonitrile (10mL). The reaction mixture was stirred at 75 ° C for 16h. After cooling to room temperature, the reaction mixture was diluted with water (30mL), extracted with EtOAc (40mL), washed with brine (30mL), and filtered through Na 2 SO 4 Dry, filter and concentrate.The residue was purified by silica gel chromatography eluting with 0-30% ethyl acetate in petroleum ether to afford the title product as a white solid (0.8 g, 42% yield). 1 H NMR (400 MHz, CDCl 3 )δ8.83(s,1H),8.49(s,1H),8.08-7.93(m,5H),7.56-7.50(m,5H),6.53(s,1H),5.64 (s,1H),4.59-4.52(m,1H),4.50-4.43(m,2H),0.88(s,9H),0.17(s,3H),0.08(s,3H).

[0660] Step 2: (2R,3R,4S)-2-(6-benzoylamino-9H-purin-9-yl)-4-hydroxytetrahydrofuran-3-yl benzoate. To a solution of (2R,3R,4S)-2-(6-benzoylamino-9H-purin-9-yl)-4-(tert-butyldimethylsilyl)oxy)tetrahydrofuran-3-yl benzoate (0.8 g, 1.4 mmol) in THF (10 mL) was added TBAF (1 M, 1.4 mL) in THF (10 mL) at 0°C. The reaction mixture was stirred at 25°C for 1 h. The reaction mixture was then diluted with water (10 mL), extracted with EtOAc (20 mL), washed with brine (20 mL x 2), and filtered through Na 2 SO 4Dried and concentrated.The residue was purified by silica gel chromatography eluting with 0-3% MeOH in DCM to afford the title product as a white solid (0.35 g, 55% yield). 1 H NMR (400 MHz, CDCl 3 )δ9.06(s,1H),8.85(s,1H),8.18(s,1H),8.06(t,J=8.4Hz,4H),7.71-7.63(m,2H),7.58-7.46(m,4H),6. 90(d,J=10.8Hz,1H),6.02(s,1H),5.62(s,1H),4.73-4.66(m,1H),4.43-4.35(m,1H),4.32-4.25(m,1H).

[0661] Step 3: Diisopropyl ((((3S,4R,5R)-5-(6-Benzoylamino-9H-purin-9-yl)-4-hydroxytetrahydrofuran-3-yl)oxy)methyl)phosphonate. A solution of (2R,3R,4S)-2-(6-Benzoylamino-9H-purin-9-yl)-4-hydroxytetrahydrofuran-3-yl benzoate (700.0 mg, 1.6 mmol) and diisopropoxyphosphorylmethyl trifluoromethanesulfonate (516.0 mg, 1.6 mmol) in THF (15 mL) was heated to 40 ℃ with N 20 ℃. 2 Degas 3 times and cool to 0° C. NaH (189.0 mg, 4.7 mmol, 60% purity) was added in several batches and the reaction mixture was stirred at 0° C. for 1 h. The reaction mixture was then washed with saturated NH 4 The mixture was quenched with aqueous Cl solution (10 mL) and extracted with EtOAc (20 mL). The organic layer was washed with brine (20 mL) and purified by Na 2 SO 4 Dried, filtered and concentrated.The residue was purified by silica gel chromatography eluting with 0-3% MeOH in DCM to afford the title product as a white solid (350 mg, 43% yield). 1 H NMR (400 MHz, CDCl 3 )δ9.00(s,1H),8.80(s,1H),8.23(s,1H),8.06-8.00(m,2H),7.66-7.60(m,1H),7.57-7.50(m,2H),6.03(d,J=4.4Hz,1H), 4.91(t,J=4.0Hz,1H),4.81-4.71(m,2H),4.40-4.27(m,3H),4.03-3.96(m,1H),3.85-3.76(m,1H),1.33(d,J=6.0Hz,12H).

[0662] Step 4: O-((2R,3R,4S)-2-(6-Benzoylamino-9H-purin-9-yl)-4-((diisopropoxyphosphoryl)methoxy)tetrahydrofuran-3-yl)O-phenylcarbonothioate. To a solution of diisopropyl (((3S,4R,5R)-5-(6-Benzoylamino-9H-purin-9-yl)-4-hydroxytetrahydrofuran-3-yl)oxy)methyl)phosphonate (350.0 mg, 673 μmol) in MeCN (10 mL) was added DMAP (82.0 mg, 674 μmol) and O-phenylcarbonothioate (151.0 mg, 876 μmol). The reaction mixture was stirred at 25 °C for 16 h. The reaction mixture was then diluted with water (10 mL), extracted with EtOAc (20 mL), washed with brine (20 mL), and purified by Na 2 SO 4 Drying, filtration and concentration provided the crude title compound as a yellow solid (180.0 mg, 41% yield).

[0663] Step 5: Diisopropyl ((((3R,5R)-5-(6-Benzoylamino-9H-purin-9-yl)tetrahydrofuran-3-yl)oxy)methyl)phosphonate. A solution of O-((2R,3R,4S)-2-(6-Benzoylamino-9H-purin-9-yl)-4-(diisopropoxyphosphoryl)methoxy)tetrahydrofuran-3-yl)O-phenylcarbonothioate (180.0 mg, 274 μmol) in toluene (6 mL) was heated with N 2 Purge for 5 min. AIBN (36.0 mg, 219.0 μmol) and TTMSS (341.0 mg, 1.37 mmol) were added. The reaction mixture was stirred at 110 °C for 2 h. After cooling to room temperature, the reaction mixture was concentrated. The residue was purified by silica gel chromatography eluting with 0-3% MeOH in DCM to provide the title compound (90 mg, 65% yield) as a white solid. LC-MS (ESI) m / z 504.0 [M+H] +. LC-MS RT = 0.754 min; Method C.

[0664] Step 6: Diisopropyl ((((3R,5R)-5-(6-amino-9H-purin-9-yl)tetrahydrofuran-3-yl)oxy)methyl)phosphonate. To a solution of diisopropyl ((((3R,5R)-5-(6-benzamido-9H-purin-9-yl)tetrahydrofuran-3-yl)oxy)methyl)phosphonate (90.0 mg, 179 μmol) in MeOH (2 mL) was added NH 4OH (1 mL, 28% purity). The resulting reaction mixture was stirred at 25 ° C for 16 h. The reaction mixture was then concentrated. The residue was purified by silica gel chromatography eluting with 0-3% MeOH in DCM to provide the title compound as a white solid (50 mg, 70% yield). LC-MS (ESI) m / z 400.2 [M+H] +. LC-MS RT = 1.615 min; Method F.

[0665] Step 7: ((((3R,5R)-5-(6-amino-9H-purin-9-yl)tetrahydrofuran-3-yl)oxy)methyl)phosphonic acid (Compound No. 113). To a solution of diisopropyl ((((3R,5R)-5-(6-amino-9H-purin-9-yl)tetrahydrofuran-3-yl)oxy)methyl)phosphonate (50.0 mg, 125 μmol) in MeCN (3 mL) at 25°C was added 2,6-lutidine (107.0 mg, 1.00 mmol). The reaction mixture was cooled to 0°C and TMSBr (153.0 mg, 1.00 mmol) was added. The reaction mixture was stirred at 0°C for 2 h. The reaction was then quenched with 1 M TEAB solution (1 mL) and concentrated. The residue was purified by preparative HPLC (acetonitrile 0-15% / FA in water) to provide Compound No. 113 (2.5 mg, 7% yield) as a white solid. 1 HNMR (400MHz, D 2 O)δ8.49(s,1H),8.22(s,1H),6.40(d,J=6.4Hz,1H),4.47-4.40(m,1H),4.33(d,J=1 0.0Hz,1H),4.11-4.02(m,1H),3.62-3.56(m,2H),2.80-2.56(m,2H).LC-MS(ESI)m / z 315.9[M+H]+.LC-MS RT=0.193min; Method C.

[0666] Embodiment 42

[0667] Synthesis of 5-(aminomethyl)-1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-methyltetrahydrofuran-2-yl)pyrimidine-2,4(1H,3H)-dione (Compound No. 140)

[0668]

[0669] Step 1: (2R,3R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-2-(5-cyano-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)-5-methyltetrahydrofuran-3-acetate. A mixture of 2,4-dioxo-1H-pyrimidine-5-carbonitrile (0.25 g, 1.82 mmol) and BTMSA (932 mg, 5.47 mmol) in MeCN (10 mL) was stirred at 70 °C for 1 hour, and then the reaction was cooled to room temperature. A solution of (2S,3R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-methyltetrahydrofuran-2,3-diyl diacetate (0.5 g, 1.17 mmol) in MeCN (10 mL) and TMSOTf (520 mg, 2.34 mmol) were added, and the mixture was stirred at 30 °C for 16 hours. After this, the reaction was diluted with EtOAc (50 mL), washed with brine (30 mL x 2), and the organic layer was concentrated to give the crude title compound (0.55 g, 93% yield) as a colorless oil. LC-MS (ESI) m / z 528.0 [M+Na]+. LC-MS RT = 0.888 min; Method C.

[0670] Step 2: 1-((2R,3R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-3-hydroxy-5-methyltetrahydrofuran-2-yl)-2,4-dioxo-1,2,3,4-tetrahydropyrimidine-5-carbonitrile. A mixture of (2R,3R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-2-(5-cyano-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)-5-methyltetrahydrofuran-3-yl acetate (0.59 g, 1.17 mmol) and NaOH (2M, 1.20 mL) in MeOH (10 mL) was stirred at 20°C for 0.2 h. The reaction was concentrated and diluted with EtOAc (50 mL), washed with brine (30 mL x 2), and then concentrated to provide the crude title compound (0.5 g, 92% yield) as a white solid, which was used directly in the next step.

[0671] Step 3: O-((2R,3R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-2-(5-cyano-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)-5-methyltetrahydrofuran-3-yl)O-phenylcarbonothioate. To a solution of 1-((2R,3R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-3-hydroxy-5-methyltetrahydrofuran-2-yl)-2,4-dioxo-1,2,3,4-tetrahydropyrimidine-5-carbonitrile (0.5 g, 1.08 mmol) and DMAP (395.39 mg, 3.24 mmol) in MeCN (20 mL) was added O-phenyl chlorothioate (279 mg, 1.62 mmol) and then stirred at 20°C for 1 hour. The reaction solution was diluted with EtOAc (50 mL) and washed with brine (30 mL x 2), then the organic layer was concentrated to give a crude product. The residue was purified by column chromatography (DCM / MeOH=100 / 1 to 100 / 5) to afford the title compound (0.47 g, 73% yield) as a colorless solid.

[0672] Step 4: 1-((2R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-methyltetrahydrofuran-2-yl)-2,4-dioxo-1,2,3,4-tetrahydropyrimidine-5-carbonitrile. A mixture of 1-[(2R,3R,4S,5R)-4-benzyloxy-5-(benzyloxymethyl)-5-methyl-3-phenoxythiocarbonyloxy-tetrahydrofuran-2-yl]-2,4-dioxo-pyrimidine-5-carbonitrile (0.47 g, 783.79 umol), TTMSS (974.48 mg, 3.92 mmol) and AIBN (64.35 mg, 391.89 umol) in toluene (15 mL) was stirred at 110° C. for 2 hours. The reaction was concentrated to give a crude product. The residue was purified by column chromatography (DCM:MeOH=100 / 1 to 20 / 1) to afford the title compound as a colorless oil (0.09 g, 26% yield). LC-MS (ESI) m / z 470.1 [M+Na]+. LC-MS RT=0.882 min; Method C.

[0673] Step 5: 5-(Aminomethyl)-1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-methyltetrahydrofuran-2-yl)pyrimidine-2,4(1H,3H)-dione (Compound No. 140). 1-((2R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-methyltetrahydrofuran-2-yl)-2,4-dioxo-1,2,3,4-tetrahydropyrimidine-5-carbonitrile (0.05 g, 111.74 umol) and PdCl were reacted at 26°C under hydrogen (15 psi).2 A mixture of (9.91 mg, 55.87 umol) in MeOH (10 mL) was stirred for 20 min. The reaction solution was filtered and concentrated to produce a crude product. The residue was purified by preparative HPLC (acetonitrile 0-28% / FA-MeCN in water) to provide compound No. 140 as a white solid (13.08 mg, 43% yield). 1 H NMR (400 MHz, MeOD-d 4 ):δ8.29(s,1H),6.19-6.16(m,1H),4.39-4.36(m,1H),3.81-3.80(m,2H),3.62-3.60(m,2H),2.42-2.32(m,2H),1.19(s,3H).LC-MS(ESI)m / z 293.8[M+Na]+.LC-MS RT=0.168min; Method C.

[0674] Embodiment 43

[0675] Synthesis of 4-amino-5-fluoro-1-((2R,5S)-5-(hydroxymethyl)-5-methyltetrahydrofuran-2-yl)pyrimidin-2(1H)-one (Compound No. 219)

[0676]

[0677] Step 1: (2R,3R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-2-(5-fluoro-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)-5-methyltetrahydrofuran-3-yl acetate. To a solution of 5-fluoro-1H-pyrimidine-2,4-dione (0.9 g, 7.3 mmol) in MeCN (8 mL) was added N,O-bis(trimethylsilyl)acetamide (5.4 mL, 21.9 mmol). The mixture was stirred at 70 °C under nitrogen atmosphere for 1 h. After this time, the reaction mixture was cooled to room temperature. Trimethylsilyl trifluoromethanesulfonate (1.4 mL, 7.9 mmol) and (2S, 3R, 4S, 5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-5-methyltetrahydrofuran-2,3-diyl diacetate (2 g, 4.7 mmol) in MeCN (20 mL) were added. The resulting reaction mixture was stirred at 25 °C under nitrogen atmosphere for 16 h. The reaction was quenched with water (20 mL), extracted with EtOAc (50 x 2 mL), washed with brine (20 mL), and purified by Na 2 SO 4Dry, filter and concentrate to give the crude product. The residue was purified by silica gel chromatography (solvent gradient: 0-3% MeOH in dichloromethane) to provide the title product as a white solid (2.3 g, 99% yield). LC-MS (ESI) m / z 499.3 [M+H]+. LC-MS RT = 0.955 min; Method C.

[0678] Step 2: 1-((2R,3R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-3-hydroxy-5-methyltetrahydrofuran-2-yl)-5-fluoropyrimidine-2,4(1H,3H)-dione. To a solution of (2R,3R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-2-(5-fluoro-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)-5-methyltetrahydrofuran-3-yl acetate (2.3 g, 4.6 mmol) in THF (25 mL) was added aqueous NaOH (2M, 6.9 mL). The mixture was stirred at 25 °C for 1 h. The reaction was then quenched with water (15 mL), extracted with EtOAc (50 mL), washed with brine (10 mL), and purified by Na 2 SO 4 Dry, filter and concentrate to give the crude product. It was purified by silica gel chromatography (100-200 mesh silica gel, 0-50% ethyl acetate in petroleum ether) to provide the title product (2.0 g, 98.8% yield). LC-MS (ESI) m / z 479.1 [M+Na]+. LC-MS RT = 0.898 min; Method C.

[0679] Step 3: 1-((2R,3R,4S,5R)-3,4-dihydroxy-5-(hydroxymethyl)-5-methyltetrahydrofuran-2-yl)-5-fluoropyrimidine-2,4(1H,3H)-dione. 1-((2R,3R,4S,5R)-4-(benzyloxy)-5-((benzyloxy)methyl)-3-hydroxy-5-methyltetrahydrofuran-2-yl)-5-fluoropyrimidine-2,4(1H,3H)-dione (1 g, 2.2 mmol) and PdCl 2 A mixture of (388.5 mg, 2.2 mmol) in MeOH (10 mL) was degassed and heated to 4% CO. 2 Purge 3 times, then incubate at 25°C in H 2 The mixture was stirred for 1 h under atmosphere (15 psi). After this, the reaction mixture was filtered and concentrated to produce a crude product. It was purified by silica gel chromatography (100-200 mesh silica gel, 0-50% ethyl acetate in petroleum ether) to provide the title compound (561.7 mg, 93% yield) as a colorless solid.

[0680] Step 4: ((2R,3R,4S,5R)-4-bromo-5-(5-fluoro-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)-3-hydroxy-2-methyltetrahydrofuran-2-yl)methyl 2-acetoxy-2-methylpropanoate. To a solution of 1-[(2R,3R,4S,5R)-3,4-dihydroxy-5-(hydroxymethyl)-5-methyl-tetrahydrofuran-2-yl]-5-fluoro-pyrimidine-2,4-dione (200 mg, 724.1 umol) in MeCN (4 mL) was added (2-bromo-1,1-dimethyl-2-oxo-ethyl)acetate (1.6 g, 7.2 mmol) at room temperature. The reaction mixture was stirred at 50° C. for 16 h. The reaction was then quenched with water (10 mL), extracted with EtOAc (20 mL), washed with brine (20 mL), and purified by Na 2 SO 4 Dry, filter and concentrate to give the crude product. The residue was purified by silica gel chromatography (100-200 mesh silica gel, 0-25% ethyl acetate in petroleum ether) to provide the title compound (262.5 mg, 95% yield) as a colorless oil. LC-MS (ESI) m / z 510 [M+H]+. LC-MS RT = 0.753 min; Method C.

[0681] Step 5: 2-acetoxy-2-methylpropanoic acid ((2S,5R)-5-(5-fluoro-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)-2-methyl-2,5-dihydrofuran-2-yl)methyl ester. A mixture of 2-acetoxy-2-methylpropanoic acid ((2R,3R,4S,5R)-4-bromo-5-(5-fluoro-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)-3-hydroxy-2-methyltetrahydrofuran-2-yl)methyl ester (40 mg, 78.5 umol), AcOH (44.9 uL, 785.4 umol) and Zn-Cu (30 mg, 232.7 umol) in MeOH (4 mL) was degassed and heated with N 2 Purge 3 times, then incubate at 25 °C under N 2 The reaction was then quenched with water (5 mL), extracted with EtOAc (10 mL), washed with brine (10 mL), and purified by Na 2 SO 4 Drying, filtration and concentration gave the crude product which was purified by silica gel chromatography (solvent gradient: 0-3% MeOH in dichloromethane) to afford the title product (25 mg, 85% yield). LC-MS (ESI) m / z 393.1 [M+Na]+. LC-MS RT=0.773 min; Method C.

[0682] Step 6: 2-acetoxy-2-methylpropanoic acid ((2S,5R)-5-(5-fluoro-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)-2-methyltetrahydrofuran-2-yl)methyl ester. To a solution of 2-acetoxy-2-methylpropanoic acid ((2S,5R)-5-(5-fluoro-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)-2-methyl-2,5-dihydrofuran-2-yl)methyl ester (25 mg, 67.5 umol) in MeOH (4 mL) was added PdCl 2 (12 mg, 67.5 umol). The mixture was heated at 25 °C under H 2 (15 psi) for 1 h. The reaction mixture was filtered and concentrated to give the crude product. It was purified by silica gel chromatography (100-200 mesh silica gel, 0-25% ethyl acetate in petroleum ether) to provide the title compound as a colorless oil (24 mg, 95% yield). LC-MS (ESI) m / z 395.1 [M+Na]+. LC-MS RT = 0.768 min; Method C.

[0683] Step 7: 2-acetoxy-2-methylpropanoic acid ((2S,5R)-5-(4-amino-5-fluoro-2-oxopyrimidin-1(2H)-yl)-2-methyltetrahydrofuran-2-yl)methyl ester. To 2-acetoxy-2-methylpropanoic acid ((2S,5R)-5-(5-fluoro-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)-2-methyltetrahydrofuran-2-yl)methyl ester (50.0 mg, 134.0 μmol), Et 3 To a solution of N (60.0 μL, 402.8 μmol) and DMAP (16.0 mg, 134.0 μmol) in MeCN (3 mL) was added 2,4,6-triisopropylbenzenesulfonyl chloride (82.0 mg, 268.6 μmol). The reaction mixture was stirred at 25 °C for 16 h. NH 3 A solution in MeOH (7M, 1 mL). The reaction mixture was stirred at 25 °C for 16 h. The reaction mixture was quenched with water (10 mL), extracted with EtOAc (20 mL), washed with brine (20 mL), and purified by Na 2 SO 4 Dry, filter and concentrate to give the crude product which was purified by silica gel chromatography (solvent gradient: 0-3% MeOH in dichloromethane) to afford the title product as a yellow solid (30.0 mg, 80% yield). LC-MS (ESI) m / z 394.1 [M+Na]+. LC-MS RT=0.654 min; Method C.

[0684] Step 8: 4-amino-5-fluoro-1-((2R,5S)-5-(hydroxymethyl)-5-methyltetrahydrofuran-2-yl)pyrimidin-2(1H)-one (Compound No. 219). To a solution of 2-acetoxy-2-methylpropanoic acid ((2S,5R)-5-(4-amino-5-fluoro-2-oxopyrimidin-1(2H)-yl)-2-methyltetrahydrofuran-2-yl)methyl ester (30.0 mg, 80.8 μmol) in MeOH (2 mL) was added NaOMe (5.0 mg, 80.8 μmol). The reaction mixture was stirred at 25 °C for 16 h. The mixture was concentrated to give a residue which was purified by preparative HPLC (acetonitrile 0-30% / NH 3 ·H 2 O+NH 4 HCO 3 The product was purified by HPLC (pH 7.0) and purified by HPLC (pH 7.0) to provide Compound No. 219 (9.4 mg, 48% yield) as a white solid. 1 H NMR (400 MHz, MeOD-d 4 )δ8.48(d,J=7.2Hz,1H),6.07-6.01(m,1H),3.73-3.65(m,1H),3.60-3. 53(m,1H),2.60-2.52(m,1H),2.20-2.11(m,1H),2.08-2.00(m,1H),1.75 -1.66(m,1H),1.20(s,3H).LC-MS(ESI)m / z 487.2[2M+H]+.LC-MS RT=1.040min; Method D.

[0685] Embodiment 44

[0686] Analytical characterization of representative compounds of the present disclosure is provided in Table 5.

[0687] Table 5

[0688]

[0689]

[0690]

[0691]

[0692]

[0693]

[0694]

[0695]

[0696]

[0697] Embodiment 33

[0698] Human LINE-1 retrotransposition assay

[0699] Representative compounds of the present disclosure were tested for inhibition of human LINE-1 retrotransposition activity in HeLa cells according to the following procedure.

[0700] HeLa cervical cancer cells were cultured at 37 °C in a humidified 5% CO 2 The cells were cultured in Dulbecco's Modified Eagle's Medium (DMEM) - High Glucose, containing 4500 mg / L glucose, L-glutamine, sodium pyruvate and sodium bicarbonate (Sigma), supplemented with 10% heat-inactivated fetal bovine serum (Thermo Fisher) in an incubator.

[0701] The assay was performed as described (Xie et al., 2011) using the reporter plasmid pYX017 with several modifications. The reporter assay was performed in 96-well white optical bottom plates. HeLa cells were seeded into the wells 24 h prior to transfection and compound treatment so that the cells were approximately 30% confluent on the day of transfection. Different cell plating densities were tested and 2×10 3 The density of cells is optimal.

[0702] Resuspend the compound in DMSO. Prepare a serial dilution (1:3) in DMSO. Prepare medium containing different concentrations of compound by adding 2 μl of compound dilution to 1 mL of medium. The final concentration of DMSO in the medium is 0.2%.

[0703] use Plasmids were transfected into cells using HD transfection reagent (Promega, E2311, Lot 382574 and Lot 397842). Transfection reagent: DNA mixture was prepared in OpiMEM (Thermo Fisher) according to the manufacturer's instructions. Different ratios of transfection reagent to DNA were tested and a 3:1 ratio was determined to be optimal. The medium was removed from the cells and discarded. Transfection reagent: DNA mixture (5 μl) was mixed with medium containing compound (100 μl / well) and added to the cells in each well. Cells were incubated at 37°C / 5% CO 2 Incubate for different incubation times. It was determined that 72h incubation time was optimal.

[0704] Dual-well plates were used according to the manufacturer's instructions for multiwell plates, except that cells were lysed directly on the multiwell plates with 30 μl of passive lysis buffer (PLB) for 20 min at room temperature with gentle shaking to ensure complete cell lysis. Luciferase reporter activity was quantified using a reporter gene assay system (Promega).

[0705] Firefly and Renilla luciferase signals were measured using a SpectraMax i3x multi-mode microplate reader. Firefly and Renilla signals were measured using integration times of 100 ms and 10 ms, respectively. Relative L1 activity was calculated as firefly / Renilla*1000 or firefly / Renilla*10,000. Dose-response inhibition data were fitted to a four-parameter logistic equation using nonlinear regression (using Graphpad Prism 8) to determine the IC of each inhibitor. 50 The results are provided in Table 6.

[0706] Table 6: Inhibition of human LINE-1 activity

[0707]

[0708]

[0709]

[0710]

[0711]

[0712] Given that the compounds, methods, kits and compositions herein are now fully described, those skilled in the art will appreciate that they can be performed within a wide and equivalent range of conditions, formulations and other parameters without affecting the scope of the methods, compounds and compositions provided herein or any embodiment thereof. All patents, patent applications and publications cited herein are fully incorporated herein by reference in their entirety.

Claims

1. A method for treating or preventing a disease, disorder or condition, and / or treating or preventing a symptom of a disease, disorder or condition in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of: (9-((2R,4S,5R)-5-ethyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-9H-purin-6-ol; 5-Bromo-1-((2R,4S,5R)-5-ethyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-4-hydroxypyrimidin-2(1H)-one; (2R,3S,5S)-5-(4-amino-6-fluoro-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-ethynyl-2-(hydroxymethyl)tetrahydrofuran-3-ol; (2R,3S,5R)-5-(2-amino-6-(cyclopropylamino)-9H-purin-9-yl)-2-(hydroxymethyl)-2-vinyltetrahydrofuran-3-ol; (2R,3S,5R)-5-(6-(cyclopropylamino)-2-fluoro-9H-purin-9-yl)-2-(hydroxymethyl)-2-vinyltetrahydrofuran-3-ol; 1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-vinyltetrahydrofuran-2-yl)-5-methoxypyrimidine-2,4(1H,3H)-dione; 1-((2S,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-vinyltetrahydrofuran-2-yl)-2,4-dioxo-1,2,3,4-tetrahydropyrimidine-5-carbonitrile; 4-amino-5-fluoro-1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-vinyltetrahydrofuran-2-yl)pyrimidin-2(1H)-one; 2-amino-9-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-vinyltetrahydrofuran-2-yl)-1H-purin-6(9H)-one; (2R,3S,5R)-5-(2-amino-6-(methylamino)-9H-purin-9-yl)-2-(hydroxymethyl)-2-vinyltetrahydrofuran-3-ol; 1-((4S,5R)-5-ethynyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-1H-1,2,4-triazole-3-carboxamide; 4-amino-1-((2R,3R,4R,5R)-5-ethynyl-3-fluoro-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)pyrimidin-2(1H)-one; (2R,3R,4R,5R)-5-(6-amino-2-fluoro-9H-purin-9-yl)-2-ethynyl-4-fluoro-2-(hydroxymethyl)tetrahydrofuran-3-ol; ((2R,5R)-5-(6-amino-9H-purin-9-yl)-2-ethynyl-2,5-dihydrofuran-2-yl)methanol; 4-amino-5-bromo-1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-methyltetrahydrofuran-2-yl)pyrimidin-2(1H)-one; (2R,3S,5R)-5-(4-amino-5-bromo-2-oxopyrimidin-1(2H)-yl)-3-hydroxy-2-(hydroxymethyl)tetrahydrofuran-2-carbonitrile; 5-Fluoro-1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-vinyltetrahydrofuran-2-yl)pyrimidine-2,4(1H,3H)-dione; 1-((2R,4S,5R)-5-ethyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-5-fluoropyrimidine-2,4(1H,3H)-dione; 1-((2R,5R)-5-ethynyl-5-(hydroxymethyl)-2,5-dihydrofuran-2-yl)-5-fluoropyrimidine-2,4(1H,3H)-dione; 1-((2S,5R)-5-ethyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)pyrimidine-2,4(1H,3H)-dione; 9-((4S,5R)-5-ethynyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-9H-purin-6-ol; 9-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-vinyltetrahydrofuran-2-yl)-9H-purin-6-ol; (2R,3S,5R)-5-(2-amino-6-methoxy-9H-purin-9-yl)-2-ethynyl-2-(hydroxymethyl)tetrahydrofuran-3-ol; ((((3R,5R)-5-(5-methyl-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)tetrahydrofuran-3-yl)oxy)methyl)phosphonic acid; ((((3R,5R)-5-(6-amino-9H-purin-9-yl)tetrahydrofuran-3-yl)oxy)methyl)phosphonic acid; 4-amino-5-bromo-1-((2R,4S,5R)-5-(fluoromethyl)-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)pyrimidin-2(1H)-one; (2R,3S,5R)-5-(6-amino-2-methoxy-9H-purin-9-yl)-2-(hydroxymethyl)-2-vinyltetrahydrofuran-3-ol; 5-Chloro-1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-methyltetrahydrofuran-2-yl)pyrimidine-2,4(1H,3H)-dione; 1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-methyltetrahydrofuran-2-yl)-5-methylpyrimidine-2,4(1H,3H)-dione; (2R,3S,5R)-5-(6-amino-2-methoxy-9H-purin-9-yl)-2-ethyl-2-(hydroxymethyl)tetrahydrofuran-3-ol; 4-amino-5-chloro-1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-methyltetrahydrofuran-2-yl)pyrimidin-2(1H)-one; 4-amino-5-fluoro-1-((2R,3aS,6aR)-6a-(hydroxymethyl)-2,3,3a,6a-tetrahydrofuro[3,2-b]furan-2-yl)pyrimidin-2(1H)-one; 5-(Aminomethyl)-1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-methyltetrahydrofuran-2-yl)pyrimidine-2,4(1H,3H)-dione; (2R,3S,4S,5R)-5-(6-amino-2-fluoro-9H-purin-9-yl)-2-ethyl-2-(hydroxymethyl)tetrahydrofuran-3,4-diol; 4-amino-5-fluoro-1-((2R,5S)-5-(hydroxymethyl)-5-methyltetrahydrofuran-2-yl)pyrimidin-2(1H)-one; (2R,3S,5R)-5-(2-amino-6-(methylamino)-9H-purin-9-yl)-2-(hydroxymethyl)-2-methyltetrahydrofuran-3-ol; (2R,3S,4S,5R)-5-(6-amino-2-fluoro-9H-purin-9-yl)-2-(hydroxymethyl)-2-methyltetrahydrofuran-3,4-diol; or 1-((2R,3R,4S,5R)-3,4-dihydroxy-5-(hydroxymethyl)-5-methyltetrahydrofuran-2-yl)-5-fluoropyrimidine-2,4(1H,3H)-dione, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof, wherein the disease, disorder or condition is a neurodegenerative disease, an autoimmune disease, an age-related disease, an autism spectrum disorder, a cardiovascular dysfunction, hearing loss, hematopoietic stem cell function, pulmonary fibrosis, schizophrenia, visual loss, progressive supranuclear palsy, amyotrophic lateral sclerosis, Aicardi-Goutières syndrome, ataxia-telangiectasia, age-related macular degeneration, systemic lupus erythematosus, IFN-related autoimmune disease, Fanconi anemia, idiopathic pulmonary fibrosis or cardiovascular disease.

2. The method of claim 1, wherein the disease, disorder or condition is a neurodegenerative disease.

3. The method of claim 2, wherein the neurodegenerative disease is Alzheimer's disease, amyotrophic lateral sclerosis, Parkinson's disease, dementia with Lewy bodies, multiple system atrophy, Huntington's disease, frontotemporal dementia (FTD), frontotemporal lobar degeneration, mild cognitive impairment, corticobasal degeneration, progressive supranuclear palsy, Rett syndrome, peripheral degenerative disease, or Aicardi-Goutières syndrome.

4. The method of claim 1, wherein the disease, disorder or condition is an autoimmune disease.

5. The method of claim 4, wherein the autoimmune disease is lupus, rheumatoid arthritis, Sjögren's syndrome, or multiple sclerosis.

6. The method of claim 1, wherein the disease, disorder or condition is an age-related disease.

7. The method of claim 6, wherein the age-related disease is Alzheimer's disease, Parkinson's disease, atherosclerosis, osteoarthritis, osteoporosis, rheumatoid arthritis, macular degeneration, peripheral degenerative disease or skin aging.

8. The method of claim 1, wherein the disease, disorder or condition is autism spectrum disorder, cardiovascular dysfunction, hearing loss, hematopoietic stem cell function, pulmonary fibrosis, schizophrenia, or vision loss.

9. The method of claim 1, wherein the disease, disorder or condition is progressive supranuclear palsy.

10. The method of claim 1, wherein the disease, disorder or condition is amyotrophic lateral sclerosis.

11. The method of claim 1, wherein the disease, disorder or condition is Aicardi-Goutières syndrome.

12. The method of claim 1, wherein the disease, disorder or condition is ataxia-telangiectasia, age-related macular degeneration, systemic lupus erythematosus, IFN-related autoimmune disease, Fanconi anemia, idiopathic pulmonary fibrosis, or cardiovascular disease.

13. The method of any one of claims 1-12, for treating a disease, disorder or condition in a subject.

14. The method of any one of claims 1-12, for preventing a disease, disorder or condition in a subject.

15. The method of any one of claims 1-12, for treating a symptom of a disease, disorder or condition in a subject.

16. The method of any one of claims 1-12, for preventing symptoms of a disease, disorder or condition in a subject.

17. The method according to any one of claims 1 to 16, wherein the compound inhibits human LINE-1 retrotransposition activity in an in vitro HeLa cell-based dual-luciferase assay at a half-maximal inhibitory concentration of less than 1 μM.

18. The method of claim 17, wherein the compound inhibits human LINE-1 retrotransposition activity at a half-maximal inhibitory concentration of less than 0.1 μM in an in vitro HeLa cell-based dual-luciferase assay.

19. The method of any one of claims 1-18, further comprising administering to the subject one or more optional therapeutic agents.

20. A kit for performing the method of any one of claims 1 to 19, comprising the compound, or a pharmaceutically acceptable salt or solvate thereof, or a stereoisomer thereof, or a tautomer thereof, and instructions for administering the compound to a subject.

21. A compound selected from the group consisting of: (9-((2R,4S,5R)-5-ethyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-9H-purin-6-ol; 5-Bromo-1-((2R,4S,5R)-5-ethyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-4-hydroxypyrimidin-2(1H)-one; (2R,3S,5S)-5-(4-amino-6-fluoro-3-methyl-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-ethynyl-2-(hydroxymethyl)tetrahydrofuran-3-ol; (2R,3S,5R)-5-(2-amino-6-(cyclopropylamino)-9H-purin-9-yl)-2-(hydroxymethyl)-2-vinyltetrahydrofuran-3-ol; (2R,3S,5R)-5-(6-(cyclopropylamino)-2-fluoro-9H-purin-9-yl)-2-(hydroxymethyl)-2-vinyltetrahydrofuran-3-ol; 1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-vinyltetrahydrofuran-2-yl)-5-methoxypyrimidine-2,4(1H,3H)-dione; 1-((2S,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-vinyltetrahydrofuran-2-yl)-2,4-dioxo-1,2,3,4-tetrahydropyrimidine-5-carbonitrile; 4-amino-5-fluoro-1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-vinyltetrahydrofuran-2-yl)pyrimidin-2(1H)-one; 2-amino-9-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-vinyltetrahydrofuran-2-yl)-1H-purin-6(9H)-one; (2R,3S,5R)-5-(2-amino-6-(methylamino)-9H-purin-9-yl)-2-(hydroxymethyl)-2-vinyltetrahydrofuran-3-ol; 1-((4S,5R)-5-ethynyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-1H-1,2,4-triazole-3-carboxamide; 4-amino-1-((2R,3R,4R,5R)-5-ethynyl-3-fluoro-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)pyrimidin-2(1H)-one; (2R,3R,4R,5R)-5-(6-amino-2-fluoro-9H-purin-9-yl)-2-ethynyl-4-fluoro-2-(hydroxymethyl)tetrahydrofuran-3-ol; ((2R,5R)-5-(6-amino-9H-purin-9-yl)-2-ethynyl-2,5-dihydrofuran-2-yl)methanol; 4-amino-5-bromo-1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-methyltetrahydrofuran-2-yl)pyrimidin-2(1H)-one; (2R,3S,5R)-5-(4-amino-5-bromo-2-oxopyrimidin-1(2H)-yl)-3-hydroxy-2-(hydroxymethyl)tetrahydrofuran-2-carbonitrile; 5-Fluoro-1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-vinyltetrahydrofuran-2-yl)pyrimidine-2,4(1H,3H)-dione; 1-((2R,4S,5R)-5-ethyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-5-fluoropyrimidine-2,4(1H,3H)-dione; 1-((2R,5R)-5-ethynyl-5-(hydroxymethyl)-2,5-dihydrofuran-2-yl)-5-fluoropyrimidine-2,4(1H,3H)-dione; 1-((2S,5R)-5-ethyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)pyrimidine-2,4(1H,3H)-dione; 9-((4S,5R)-5-ethynyl-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-9H-purin-6-ol; 9-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-vinyltetrahydrofuran-2-yl)-9H-purin-6-ol; (2R,3S,5R)-5-(2-amino-6-methoxy-9H-purin-9-yl)-2-ethynyl-2-(hydroxymethyl)tetrahydrofuran-3-ol; ((((3R,5R)-5-(5-methyl-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)tetrahydrofuran-3-yl)oxy)methyl)phosphonic acid; ((((3R,5R)-5-(6-amino-9H-purin-9-yl)tetrahydrofuran-3-yl)oxy)methyl)phosphonic acid; 4-amino-5-bromo-1-((2R,4S,5R)-5-(fluoromethyl)-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)pyrimidin-2(1H)-one; (2R,3S,5R)-5-(6-amino-2-methoxy-9H-purin-9-yl)-2-(hydroxymethyl)-2-vinyltetrahydrofuran-3-ol; 5-Chloro-1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-methyltetrahydrofuran-2-yl)pyrimidine-2,4(1H,3H)-dione; 1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-methyltetrahydrofuran-2-yl)-5-methylpyrimidine-2,4(1H,3H)-dione; (2R,3S,5R)-5-(6-amino-2-methoxy-9H-purin-9-yl)-2-ethyl-2-(hydroxymethyl)tetrahydrofuran-3-ol; 4-amino-5-chloro-1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-methyltetrahydrofuran-2-yl)pyrimidin-2(1H)-one; 4-amino-5-fluoro-1-((2R,3aS,6aR)-6a-(hydroxymethyl)-2,3,3a,6a-tetrahydrofuro[3,2-b]furan-2-yl)pyrimidin-2(1H)-one; 5-(Aminomethyl)-1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)-5-methyltetrahydrofuran-2-yl)pyrimidine-2,4(1H,3H)-dione; (2R,3S,4S,5R)-5-(6-amino-2-fluoro-9H-purin-9-yl)-2-ethyl-2-(hydroxymethyl)tetrahydrofuran-3,4-diol; 4-amino-5-fluoro-1-((2R,5S)-5-(hydroxymethyl)-5-methyltetrahydrofuran-2-yl)pyrimidin-2(1H)-one; (2R,3S,5R)-5-(2-amino-6-(methylamino)-9H-purin-9-yl)-2-(hydroxymethyl)-2-methyltetrahydrofuran-3-ol; (2R,3S,4S,5R)-5-(6-amino-2-fluoro-9H-purin-9-yl)-2-(hydroxymethyl)-2-methyltetrahydrofuran-3,4-diol; and 1-((2R,3R,4S,5R)-3,4-dihydroxy-5-(hydroxymethyl)-5-methyltetrahydrofuran-2-yl)-5-fluoropyrimidine-2,4(1H,3H)-dione, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof.

22. A pharmaceutical composition comprising the compound of claim 21, or a pharmaceutically acceptable salt or solvate thereof, or a tautomer thereof, and one or more pharmaceutically acceptable excipients.

23. A kit comprising a compound according to claim 21, or a pharmaceutically acceptable salt or solvate thereof, or a stereoisomer thereof, or a tautomer thereof, and instructions for administering the compound to a subject, and optionally comprising one or more optional therapeutic agents.

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