WO2014143799A2 - Substituted xanthines and methods of use thereof - Google Patents

Substituted xanthines and methods of use thereof Download PDF

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Publication number
WO2014143799A2
WO2014143799A2 PCT/US2014/027920 US2014027920W WO2014143799A2 WO 2014143799 A2 WO2014143799 A2 WO 2014143799A2 US 2014027920 W US2014027920 W US 2014027920W WO 2014143799 A2 WO2014143799 A2 WO 2014143799A2
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Prior art keywords
alkyl
mmol
methyl
dione
purine
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PCT/US2014/027920
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French (fr)
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WO2014143799A4 (en
WO2014143799A9 (en
WO2014143799A3 (en
Inventor
Bertrand Chenard
Randall Gallaschun
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Hydra Biosciences, Inc.
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Priority to UAA201507774A priority Critical patent/UA117470C2/en
Priority to CN201480023267.0A priority patent/CN105143229B/en
Priority to JP2016502662A priority patent/JP6360149B2/en
Priority to MYPI2015702631A priority patent/MY189912A/en
Priority to ES14722452.1T priority patent/ES2636835T7/en
Priority to KR1020157029667A priority patent/KR102227629B1/en
Priority to MX2015011617A priority patent/MX370372B/en
Priority to NZ711718A priority patent/NZ711718A/en
Priority to DK14722452.1T priority patent/DK2970303T6/en
Priority to CA2899646A priority patent/CA2899646C/en
Priority to EA201591615A priority patent/EA028815B1/en
Priority to EP14722452.1A priority patent/EP2970303B3/en
Priority to PL14722452T priority patent/PL2970303T6/en
Application filed by Hydra Biosciences, Inc. filed Critical Hydra Biosciences, Inc.
Priority to LTEP14722452.1T priority patent/LT2970303T/en
Priority to RS20170566A priority patent/RS56066B2/en
Priority to SG11201506479TA priority patent/SG11201506479TA/en
Priority to AU2014228206A priority patent/AU2014228206C1/en
Priority to SI201430241T priority patent/SI2970303T1/en
Priority to BR112015021164-0A priority patent/BR112015021164B1/en
Publication of WO2014143799A2 publication Critical patent/WO2014143799A2/en
Publication of WO2014143799A3 publication Critical patent/WO2014143799A3/en
Publication of WO2014143799A9 publication Critical patent/WO2014143799A9/en
Publication of WO2014143799A4 publication Critical patent/WO2014143799A4/en
Priority to ZA2015/05414A priority patent/ZA201505414B/en
Priority to IL240216A priority patent/IL240216B/en
Priority to PH12015502141A priority patent/PH12015502141B1/en
Priority to HK16105298.0A priority patent/HK1217329A1/en
Priority to HRP20170840TT priority patent/HRP20170840T4/en
Priority to CY20171100647T priority patent/CY1119042T1/en

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D473/00—Heterocyclic compounds containing purine ring systems
    • C07D473/02—Heterocyclic compounds containing purine ring systems with oxygen, sulphur, or nitrogen atoms directly attached in positions 2 and 6
    • C07D473/04—Heterocyclic compounds containing purine ring systems with oxygen, sulphur, or nitrogen atoms directly attached in positions 2 and 6 two oxygen atoms
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P13/00—Drugs for disorders of the urinary system
    • A61P13/12—Drugs for disorders of the urinary system of the kidneys
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A61P25/02—Drugs for disorders of the nervous system for peripheral neuropathies
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A61P25/08—Antiepileptics; Anticonvulsants
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A61P25/18—Antipsychotics, i.e. neuroleptics; Drugs for mania or schizophrenia
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A61P25/22—Anxiolytics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A61P25/24—Antidepressants
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A61P25/28—Drugs for disorders of the nervous system for treating neurodegenerative disorders of the central nervous system, e.g. nootropic agents, cognition enhancers, drugs for treating Alzheimer's disease or other forms of dementia
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D473/00—Heterocyclic compounds containing purine ring systems
    • C07D473/02—Heterocyclic compounds containing purine ring systems with oxygen, sulphur, or nitrogen atoms directly attached in positions 2 and 6
    • C07D473/04—Heterocyclic compounds containing purine ring systems with oxygen, sulphur, or nitrogen atoms directly attached in positions 2 and 6 two oxygen atoms
    • C07D473/06—Heterocyclic compounds containing purine ring systems with oxygen, sulphur, or nitrogen atoms directly attached in positions 2 and 6 two oxygen atoms with radicals containing only hydrogen and carbon atoms, attached in position 1 or 3
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D487/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00
    • C07D487/02—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00 in which the condensed system contains two hetero rings
    • C07D487/04—Ortho-condensed systems
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D487/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00
    • C07D487/12—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00 in which the condensed system contains three hetero rings

Definitions

  • ion channel proteins exist to mediate ion flux across cellular membranes.
  • the proper expression and function of ion channel proteins is . essential for the maintenance of cell function, intracellular communication, and the like. Numerous diseases are the result of misregulation of membrane potential or aberrant calcium handling. Given the central importance of ion channels in modulating membrane potential and ion flux in cells, identification of agents that can promote or inhibit particular ion channels are of great interest as research tools and as possible therapeutic agents.
  • the present invention provides methods of treating a TRPC5 mediated disorder in a subject, comprising administering to the sub ect a compound of Formula I:
  • the present invention further rovides compounds of Formula 1(a):
  • the present invention further provides compounds of Formula II:
  • the present invention further provides compounds of Formula III:
  • compositions comprising a compound of Formula 1(a), II or III, and a pharmaceutically acceptable carrier.
  • the present invention further provides methods of treating a TRPC5 mediated disorder in a subject, e.g. a human, comprising administering to the subject a compound or composition of a compound of Formula 1(a), II or Formula III, or a pharmaceutically acceptable salt thereof.
  • the present invention provides methods, compounds and compositions for treating conditions such as a neuropsychiatric disorder, a neurodegenerative disorder, nephropathy, and seizure disorder by modulating the activity of the transient receptor potential cation channel subfamily C, member 5 (TRPC5), which can exist in homomultimeric form as well as heteromultimeric form with other ion channels such as TRPC1 or TRPC3 (i.e., TRPC5-TRPC1 and TRPC 1 -TRPC3-TRPC5).
  • TRPC5-TRPC1 and TRPC 1 -TRPC3-TRPC5 The compounds described herein modulate the function of TRPC5 by inhibiting a TRPC5-mediated ion flux or by inhibiting the inward current, the outward current, or both currents mediated by TRPC5.
  • the inhibition of a particular current is the ability to inhibit or reduce such current (e.g., inward and/or outward) in an in vitro or an in vivo assay.
  • the activation of a particular current is the ability to activate or increase such current (e.g., inward and/or outward) in an in vitro or an in vivo assay.
  • the invention relates to a method for treating a condition for which reduced
  • TRPC5 activity can reduce the severity of the condition, by administering a TRPC5 antagonist, such as a compound as described herein that inhibits TRPC5-mediated current and/or TRPC5- mediated ion flux.
  • a TRPC5 antagonist such as a compound as described herein that inhibits TRPC5-mediated current and/or TRPC5- mediated ion flux.
  • Described herein are compounds, which are TRPC5 antagonists that have a measured IC50 for inhibition of TRPC5 of 10 nanomolar or less.
  • the compounds described herein, which are TRPC5 antagonists inhibit one or both of inward and outward TRPC5-mediated currents with an IC 50 10 nanomolar or less.
  • the compounds described herein inhibit at least 95% of TRPC5-mediated current or TRPC5- mediated ion flux when administered at 1 micromolar or less.
  • the compounds described herein, which are TRPC5 antagonists can be used to inhibit a function of TRPC5, for example a TRPC5-mediated current and/or a TRPC5- mediated ion flux.
  • the compounds described herein can be used to inhibit a TRPC5 mediated current in vitro, for example in cells in culture.
  • the compounds described herein can be used to inhibit a TRPC5 mediated current in vivo.
  • the compounds described herein inhibit both an inward and an outward TRPC5- mediated current.
  • Another aspect of the invention features a pharmaceutical preparation suitable for use in a human patient, or for veterinary use, comprising an effective amount of a compound described herein (or a salt thereof, or a solvate, hydrate, oxidative metabolite or prodrug of the compound or its salt), and one or more pharmaceutically acceptable excipients.
  • the invention further contemplates the use of the compounds described herein in the manufacture of a medicament or pharmaceutical preparation to treat or reduce the symptoms of any of the diseases or conditions provided in the specification.
  • the compounds described herein can be used for treating a particular disease or condition can be formulated for administration via a route appropriate for the particular disease or condition.
  • the compounds described herein can be administered alone or in combination with another therapeutic agent.
  • the compounds described herein can be administered conjointly with one or more of an anti-inflammatory agent, anti-acne agent, anti-wrinkle agent, anti-scarring agent, anti-psoriatic agent, anti-proliferative agent, anti-fungal agent, anti-viral agent, anti-septic agent, anti-migraine agent, keratolytic agent, or a hair growth inhibitor.
  • the compounds described herein can be administered topically, orally, transdermally, rectally, vaginally, parentally, intranasally, intrapulmonary, intraocularly, intravenously, intramuscularly, intraarterially, intrathecally, intracapsularly, intraorbitally, intracardiacly, intradermal ly, intraperitoneally, transtracheally, subcutaneously, subcuticularly, intraarticularly, subcapsularly, subarachnoidly, intraspinally, intrasternally, sublingually, or by inhalation.
  • the compounds described herein can be administered topically.
  • the compounds described herein can be administered orally.
  • the compounds described herein can be administered administered parentally.
  • FIG. 1 is a plot depicting NMRI mouse immobility after administration of 0.5%
  • Methylcellulose (MC) (4000 CPS) orally, or Compound 260 at oral doses of 0.3, ) , 3 and 10 mg/kg one hour prior to testing, or desipramine by intraperitoneal injection of 30 mg/kg, 30 minutes prior to testing.
  • FIGs. 2A and 2B are plots depicting activity in the Vogel Conflict Test Model for Wistar- Kyoto rats after administration of 0.5% Methylcellulose (4000 CPS), or Compound 260 at doses of 0.3, 1 , 3 and 10 mg/kg orally 1 hour prior to testing, or Midazolam 1 mg/kg by intraperitoneal injection, 30 minutes prior to testing.
  • FIGs. 3A-3D are plots depicting anxiolytic effects of Wistar-Kyoto rats employed in an elevated-plus maze assay using Midazolam lmg/kg dose and 0.5% methylcellulose as positive and negative controls, respectively, as well as Compound 260 at doses of 0.3, 1 , 3 and 10 mg/kg.
  • FIGs. 3A and 3B are plots depicting % entries of the rats in the open arm portions of the maze.
  • FIG. 3C is a plot depicting entries of the rats in the closed arm portion of the maze.
  • FIG. 3D is a plot depicting the distance travelled (in centimeters) by the rats.
  • FIGs. 4A and 4B are line graphs depicting stimulant effects on NMRI mice after administration of phencyclidine (PCP) at 5mg/kg delivered subcutaneously or with 0.5% methylcellulose (MC) (vehicle) delivered subcutaneously (FIG. 4A), or with Compound 260 at doses of 0.3, 1 , 3 and 10 mg/kg in MRI mice (FIG. 4B).
  • PCP phencyclidine
  • MC 0.5% methylcellulose
  • Compound 260 at doses of 0.3, 1 , 3 and 10 mg/kg in MRI mice
  • Cation channels such as TRPC5 modulate the flux of calcium and sodium ions across cellular membranes.
  • Sodium and calcium influx leads to a depolarization of the cell. This increases the probability that voltage-gated ion channels will reach the threshold required for activation.
  • activation of non-selective cation channels can increase electrical excitability and increase the frequency of voltage-dependent events.
  • Voltage-dependent events include, but are not limited to, neuronal action potentials, cardiac action potentials, smooth muscle contraction, cardiac muscle contraction, and skeletal muscle contraction.
  • Non-selective cation channels such as TRPC5
  • Calcium influx caused by the activation of non-selective cation channels such as TRPC5 also alters the intracellular free calcium concentration.
  • Calcium is a ubiquitous second messenger molecule within the cell and the alterations in intracellular calcium levels have profound effects on signal transduction and gene expression.
  • activation of non-selective cation channels such as TRPC5 can lead to changes in gene expression and cellular phenotype.
  • Gene expression events include, but are not limited to, production of mRNAs encoding cell surface receptors, ion channels, and kinases. These changes in gene expression can lead to hyperexcitability in that cell.
  • TRP Transient receptor potential
  • TRPC5 forms homomultimeric structures such as tetramers (i.e., TRPC5 homomultimers) and
  • TRPC5-TRPC1 heteromultimers such as tetramers
  • TRPC5-TRPC1 heteromultimers i.e., TRPC5-TRPC1 heteromultimers
  • TRPC5 when identifying a modulator of TRPC5 such as a TRPC5 antagonist, the term TRPC5 is used generically so as to include either or both of a TRPC5 homomultimer or a heteromultimer (e.g., TRPC5-TPRC1 or TRPC5-TRPC4 heteromultimer).
  • TRPC5 in the literature include the following: Nature. 2008 Jan. 3; 451 (7174):69-72; Mol Pharmacol. 2008 January; 73 (l):42-9; J Biol Chem. 2007 Nov.
  • Modulating the function of TRPC5 proteins provides a means of modulating calcium homeostasis, sodium homeostasis, membrane polarization, and/or intracellular calcium levels
  • compounds that can modulate TRPC5 function are useful in many aspects, including, but not limited to, maintaining calcium homeostasis, modulating intracellular calcium levels, modulating membrane polarization, and treating or preventing diseases, disorders, or conditions associated with calcium and/or sodium homeostasis or dyshomeostasis.
  • the invention provides methods of treating a TRPC5 mediated disorder in a subject comprising administering to the sub ect a compound of Formula I:
  • R 1 is C]-C 6 alkyl, C 2 -C 6 alkenyl or C 2 -C 6 alkynyl, each of which is optionally substituted with 1-4 R 5 ;
  • R 2 is Cf-C 6 alkyl, C C 6 heteroalkyl, C 2 -C 6 alkenyl, C 2 -C 6 alkynyl, Ci-C 6 haloalkyl, halo, Ci-C 6 haloalkoxy, hydroxyl, Ci-C 6 alkoxy, C 3 -C 7 cycloalkyloxy, C 6 -Cio aryl, C 6 -Ci 0 aryloxy, C 7 - Ci6 arylalkoxy, amino, Ci-C 6 akylamino, C 2 -C] 2 dialkylamino, -S-, -S-Ci-C 6 alkyl, -S(O)-, - S(0) 2 -, heterocycloalkyl, heteroaryl, heteroaryloxy, sulfonamidyl, amido, urea, sulfonylurea, acyl, nitro, cyano, wherein each of Q-C 6
  • R 4 is Cj-C 6 alky], C
  • R 6 , R 7 , and R 8 are each independently C
  • each R 9 is independently Ci-C 6 alkyl, Ci-C 6 heteroalkyl, Ci-C 6 haloalkyl, Cj-C 6 haloalkoxy, heterocycloalkyl, C6-Ci 0 aryl, heteroaryl, C -Cio cycloalkylalkyl, heterocycloalkyl- Ci-C 6 alkyl, C 7 -Ci 6 arylalkyl, heteroaryl-Ci-C 6 alkyl, halo, hydroxyl, Cj-C 6 alkoxy, C 6
  • the TRPC5 mediated disorder is selected from the group consisting of: a neuropsychiatric disorder, a neurodegenerative disorder, nephropathy, and seizure disorder.
  • R 1 is Ci-C 6 alkyl.
  • R 1 is C]-C 6 alkyl and R 5 is independently C 6 -Cio aryl or heteroaryl.
  • R 1 is C]-C 6 alkyl and R 5 is independently phenyl optionally substituted with 1 -3 R 9 .
  • R 1 is C
  • R 2 is C 6 -Cio aryl, C 6 -Ci 0 aryloxy or heteroaryl oxy. In some embodiments, R is C 6 -Cio aryloxy.
  • R is Cj-C alkoxy.
  • R is Ci-C 6 alkyl.
  • R is Ci-C 6 alkyl and R is independently C 6 -Cio aryl or heteroaryl, optionally substituted with 1-3 R 9 .
  • R 2 is Ci-C 6 alkyl and R 6 is independently phenyl, optionally substituted with 1 -3 R 9 .
  • R is or Ci-C 6 akylamino.
  • R is -S(O)- or -S(0) 2 -.
  • R 3 is Ci-C 6 alkyl, C 2 -C 6 hydroxyalkyl, or Ci-C alkoxy.
  • R 3 is C 2 -C 6 hydroxyalkyl, e.g., hydroxypropyl.
  • R is hydroxypropyl
  • R 4 is Ci-C 6 alkyl.
  • R 5 is independently C 6 -C 10 aryl, heteroaryl, C 3 -C 7 cycloalkyl, or hetero cycloalkyl.
  • R 5 is phenyl, pyridyl, thiazolyl, pyrimidinyl, or oxazolyl.
  • R 5 is phenyl
  • R 5 is pyridyl
  • R 5 is thiazolyl
  • R 9 is independently Ci-C 6 alkyl, Ci-C 6 alkoxy, halo, Ci-C 6 haloalkyl, C[-C 6 haloalkoxy, or heterocycloalkyl.
  • R 2 is C 6 -Ci 0 aryl or C6-Ci 0 aryloxy
  • R 6 is independently Ci-C 6 alkyl, Ci-C 6 haloalkyl, Cj-C 6 or haloalkoxy.
  • R 2 is C 6 -Cjo aryloxy and R 6 is independently Ci-C 6 haloalkyl or C i -C 6 or haloalkoxy.
  • R 2 is C 6 -Cio aryloxy and R 6 is -CF 3 or -OCF 3 .
  • the invention rovides compounds of Formula T(a):
  • Ci-C 6 alkyl C2-C 6 alkenyl or C 2 -C6 alkynyl, each of which is optionally substituted with 1 -4 R 5 ;
  • R 2 is Ci-C 6 alkyl, Ci-C 6 heteroalkyl, C 2 -C 6 alkenyl, C 2 -C 6 alkynyl, C r C 6 haloalkyi, halo, Ci-C 6 haloalkoxy, hydroxyl, C t -C 6 alkoxy, C 3 -C 7 cycloalkyloxy, C 6 -Ci 0 aryl, C 6 -Cio aryloxy, C 7 - C i6 arylalkoxy, amino, Cj-C 6 akylamino, C2-C12 dialkylamino, -S-, -S-C]-C 6 alkyl, -S(O)-, - S(0) 2 -, heterocycloalkyl, heteroaryl, heteroaryloxy, sulfonamidyl, amido, urea, sulfonylurea, acyl, nitro, cyano, wherein each of
  • R is C 2 -C 6 hydroxyalkyl or Ci-C 6 heteroalkyl
  • R 4 is Ci-C 6 alkyl, Cj-C heteroalkyl, C 2 -C 6 alkenyl or C 2 -C 6 alkynyl, each of which is optionally substituted with 1-4 R 8 ;
  • R 5 , R 6 , and R 8 are each independently Ci-C 6 alkyl, Ci-C 6 heteroalkyl, halo, Q-Q haloalkyi, Ci-C 6 haloalkoxy, hydroxyl, Ci-C 6 alkoxy, amino, Ci-C 6 alkylamino, C 2 -C] 2 dialkylamino, cyano, nitro, amido, Ci-C alkylamido, C 2 -Ci 2 dialkylamido, -S-, -S(0) 2 -,
  • Ci-C 6 haloalkyi Ci-C 6 haloalkoxy, hydroxyl, Cj-C 6 alkoxy, amino, Ci-C 6 alkylamino, C 2 -C) 2 dialkylamino, amido, C]-C 6 alkylamido, C 2 -C 12 dialkylamido, -S-, -S(0) 2 -, -C(0)0-, -C(O)-, -C(0)0-Ci-C 6 alkyl, C 3 -C 7 cycloalkyl, C 6 -C ]0 aryl, heterocycloalkyl, or heteroaryl is optionally substituted with 1-3 R 9 ; and
  • each R 9 is independently Ci-C 6 alkyl, Ci-C 6 heteroalkyl, Ci-C 6 haloalkyi, Ci-C 6 haloalkoxy, heterocycloalkyl, C6-C )0 aryl, heteroaryl, C 4 -Cio cycloalkyl alkyl, heterocycloalkyl- Ci-C 6 alkyl, C 7 -C 16 arylalkyl, heteroaryl-Ci-C 6 alkyl, halo, hydroxyl, Ci-C 6 alkoxy, C 6 -C) 0 aryloxy, C 7 -C( 6 arylalkoxy, C 2 -C 8 alkoxyalkoxyl, amino, Ci-C akylamino, C 2 -Cj 2 dialkylamino, C]-C 6 akyl-amino-C]-C 6 akyl, C C 6 akyl-amino-C]-C 6 akyl, C C
  • R 1 is Ci-C 6 alkyl and R 5 is phenyl, pyridyl, thiazolyl, pyrimidinyl, or oxazolyl, e.g., phenyl, pyridiyl, or thiazolyl.
  • R 1 is C)-C 6 alkyl and R 5 is independently phenyl optionally substituted with 1 -3 R 9 .
  • R 1 is Cj-C 6 alkyl and R 5 is independently pyridyl, e.g., 2-pyridyl, 3-pyridyl, or 4 pyridyl, optionally substituted with 1-3 R 9 .
  • R 2 is C 6 -Cio aryl or C 6 -Cio aryloxy
  • R 6 is independently Ci-C 6 alkyl, Ci-C 6 haloalkyl, C]-C 6 or haloalkoxy.
  • R 2 is Ci-C 6 alkyl and R 6 is independently C 6 -C
  • R 2 is Cj-C 6 alkyl and R 6 is independently phenyl, optionally substituted with 1-3 R 9 .
  • R 3 is C 2 -C 6 hydroxyalkyl, e.g., hydroxypropyl.
  • inve ormula II in another aspect, the inve ormula II:
  • Ring A is phenyl, pyridyl, thiazolyl, pyrimidinyl, or oxazolyl;
  • R 2 is Ci-C 6 alkyl, Ci-C 6 heteroalkyl, C 2 -C 6 alkenyl, C 2 -C 6 alkynyl, C r C 6 haloalkyl, halo, Ci-C 6 haloalkoxy, hydroxyl, Ci-C 6 alkoxy, C 3 -C 7 cycloalkyloxy, C 6 -Cio aryl,.
  • R 3 is C r C 6 alkyl, Ci-C 6 heteroalkyl, C 2 -C 6 alkenyl, C 2 -C 6 alkynyl, C 2 -C 6 hydroxyalkyl, or Ci-C 6 alkoxy, each of which is optionally substituted with 1 -4 R ;
  • R 4 is C]-C 6 alkyl, Ci-C 6 heteroalkyl, C 2 -C alkenyl or C 2 -C 6 alkynyl, each of which is optionally substituted with 1-4 R 8 ;
  • R , R , and R are each independently Ci-C alkyl, Ci-C 6 heteroalkyl, halo, Ci-C 6 haloalkyl, Ci-C 6 haloalkoxy, hydroxyl, C[-C 6 alkoxy, amino, Ci-C 6 alkylamino, C 2 -C
  • each R 9 is independently Ci-C 6 alkyl, Ci-C 6 heteroalkyl, Ci-C 6 haloalkyl, Ci-C 6 haloalkoxy, heterocycloalkyl, C 6 -Ci 0 aryl, heteroaryl, C4-C10 cycloalkylalkyl, heterocycloalkyl- Ci-C 6 alkyl, C 7 -Ci 6 arylalky], heteroaryl-Ci-C alkyl, halo, hydroxyl, Ci-C 6 alkoxy, C 6 -Cio aryloxy
  • n 1 or 2;
  • n 1 , 2, or 3.
  • Ring A is phenyl or thiazolyl.
  • Ring A is pyridyl, e.g. 2-pyridyl, 3-pyridyl, or 4-pyridyl.
  • R 2 is CpC 6 alkyl and R 6 is independently C 6 -C] 0 aryl or heteroaryl, optionally substituted with 1 -3 R 9 .
  • R 2 is Ci-C 6 alkyl and R 6 is independently phenyl, optionally substituted with 1 -3 R 9 . In some embodiments, R 3 is hydroxypropyl.
  • the invention rovides compound of Formula 111:
  • R is C
  • R 3 is Ci-C 6 heteroalkyl or C 2 -C 6 hydroxyalkyl
  • R 4 is Ci-C 6 alkyl
  • R 6 is independently C]-C 6 alkyl, halo, Ci-C 6 haloalkyl, C)-C 6 haloalkoxy, or Ci-C 6 alkoxy;
  • each R a is Ci-C 6 alkyl, d-C 6 haloalkyl, halo;
  • n 1 or 2;
  • n 1 , 2, or 3.
  • R 3 is hydroxypropyl
  • R a is independently chloro, fluoro, or methyl.
  • the invention provides methods of treating a TRPC5 mediated disorder in a subject, the method comprising administering to the subject a compound or composition of any one of claims 23 to 33, to thereby treat the subject.
  • the TRPC5 mediated disorder is selected from the group consisting of: a neuropsychiatric disorder, a neurodegenerative disorder, nephropathy, and seizure disorder.
  • exemplary compounds of the invention include the compounds described in Table A and in the Examples. Table A
  • substituents of compounds of the invention are disclosed in groups or in ranges. It is specifically intended that the invention include each and every individual subcombination of the members of such groups and ranges.
  • the term "Ci -6 alkyl” is specifically intended to individually disclose methyl, ethyl, C 3 alkyl, C 4 alkyl, C 5 alkyl, and C 6 alkyl.
  • each variable can be a different moiety selected from the Markush group defining, the variable.
  • the two R groups can represent different moieties selected from the Markush group defined for R.
  • acyl refers to the group (C[-C 6 alkyl)-C(O)-.
  • alkyl by itself or as part of another substituent, means, unless otherwise stated, a straight or branched chain, and can have a number of carbon atoms optionally designated (i.e. , Ci-C 6 means one to six carbons).
  • saturated hydrocarbon groups include, but are not limited to, groups such as methyl, ethyl, n-propyl, isopropyl, n-butyl, t-butyl, isobutyl, sec-butyl, n-pentyl, isopentyl, homologs and isomers of, for example, n-pentyl, n-hexyl, and the like.
  • alkenyl can be a straight or branched hydrocarbon chain, containing at least one double bond, and having from two to six carbon atoms (i.e. C 2 -C alkenyl).
  • alkenyl groups include, but are not limited to, groups such as ethenyl (i.e., vinyl), prop-l-enyl (i.e., allyl), but-l -enyl, pent-l-enyl, penta-l ,4-dienyl, and the like.
  • alkoxy can be a straight chain or branched alkoxy group having from one to six carbon atoms (i.e., Ci-C 6 alkoxy).
  • alkoxy groups include, but are not limited to, groups such as methoxy, ethoxy, propyloxy, isopropyloxy, butyloxy, isobutyloxy, tert-butyloxy, pentyloxy, or hexyloxy, and the like.
  • alkynyl can be a straight or branched hydrocarbon chain, containing at least one triple bond, having from two to six carbon atoms (i.e. C 2 -C 6 alkynyl).
  • alkynyl groups include, but are not limited to, groups such as ethynyl, propynyl, butynyl, pentynyl, hexynyl, and the like.
  • amide or “amido” refers to a chemical moiety with the formula - C(0)NR a - or -NR a C(0)- wherein R a is H or Ci-C 6 alkyl.
  • amino or “amine” refers to a -NH 2 radical group.
  • alkylamino refers to a group of formula -NH(alkyl), wherein the alkyl group each has 1 to 6 carbons.
  • dialkylamino refers to a group of formula -N(alkyl) 2 , wherein the two alkyl groups each independently has, 1 to 6 carbons.
  • aryl refers to a polyunsaturated, aromatic, hydrocarbon moiety which can be a single ring or multiple rings (e.g., 1 to 2 rings) which are fused together or linked covalently, having from six to twelve carbon atoms (i.e. C 6 -C] 2 aryl).
  • Non-limiting examples of aryl groups include phenyl, 1 -naphthyl, 2-naphthyl, and 4-biphenyl.
  • arylalkyl refers to an (aryl)alkyl— radical wherein aryl and alkyl moieties are as disclosed herein.
  • aryloxy refers to -O-(aryl), wherein the heteroaryl moiety is as defined herein.
  • arylalkoxy refers to -O-(arylalkyl), wherein the heteroaryl moiety is as defined herein.
  • cyano refers to a -CN radical
  • cycloalkyl refers to a monocyclic or polycyclic radical that contains only carbon and hydrogen, and may be saturated, or partially unsaturated.
  • Cycloalkyl groups include groups having from 3 to 10 ring atoms (i.e. C 3 -C] 0 cycloalkyl). Examples of cycloalkyl groups include, but are not limited to, groups such as cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cycloseptyl, cyclooctyl, cyclononyl, cyclodecyl, norbornyl, and the like.
  • C3-C7 cycloalkyloxy refers to -0-(C 3 -C 7 cycloalkyl), wherein the
  • C3-C7cycloalkyl moiety is as defined herein.
  • halo or halogen
  • halo independently or as part of another substituent, mean, unless otherwise stated, a fluorine, chlorine, bromine, or iodine atom.
  • halide by itself or as part of another substituent, refers to a fluoride, chloride, bromide, or iodide atom.
  • haloalkyl and haloalkoxy can include alkyl and alkoxy structures that are substituted with one or more halo groups or with combinations thereof.
  • fluoroalkyl and fluoroalkoxy include haloalkyl and haloalkoxy groups, respectively, in which the halo is fluorine.
  • heteroalkyl can include an optionally substituted alkyl, which has one or more skeletal chain atoms selected from an atom other than carbon, e.g., oxygen, nitrogen, sulfur, phosphorus or combinations thereof.
  • a numerical range may be given, e.g. C]-C 6 heteroalkyl which refers to the number of carbons in the chain, which in this example includes 1 to 6 carbon atoms.
  • a -CH2OCH2CH3 radical is referred to as a "C 3 " heteroalkyl. Connection to the rest of the molecule may be through either a heteroatom or a carbon in the heteroalkyl chain.
  • heteroaryl refers to a 5- to 14-membered aromatic radical (e.g., C2-C 13 heteroaryl) that includes one or more ring heteroatoms selected from nitrogen, oxygen and sulfur, and which may be a monocyclic or bicyclic ring system.
  • Bivalent radicals derived from univalent heteroaryl radicals whose names end in "-yl” by removal of one hydrogen atom from the atom with the free valence are named by adding "-idene" to the name of the corresponding univalent radical, e.g., a pyridyl group with two points of attachment is a pyridylidene.
  • heteroaryl refers to an aromatic group in which at least one of the skeletal atoms of the ring is a nitrogen atom.
  • the polycyclic heteroaryl group may be fused or non-fused.
  • the heteroatom(s) in the heteroaryl radical is optionally oxidized.
  • One or more nitrogen atoms, if present, are optionally quatemized.
  • the heteroaryl is attached to the rest of the molecule through any atom of the ring(s).
  • heteroaryl groups include without limitation, pyridyl, pyrimidinyl, pyrazinyl, pyridazinyl, triazinyl, furyl (furanyl), quinolyl, isoquinolyl, thienyl, imidazolyl, thiazolyl, indolyl, pyrryl, oxazolyl, benzofuryl, benzothienyl, benzthiazolyl, isoxazolyl, pyrazolyl, triazolyl, tetrazolyl, indazolyl, 1 ,2,4-thiadiazolyl, isothiazolyl, benzothienyl, purinyl, carbazolyl, benzimidazolyl, indolinyl, and the like.
  • heteroaryloxy refers to -O-(heteroaryl), wherein the heteroaryl moiety is as defined herein.
  • heterocycloalkyl can be a stable 3- to 18-membered non-aromatic ring radical that comprises two to twelve carbon atoms and from one to six heteroatoms selected from nitrogen, oxygen and sulfur.
  • heterocycloalkyl groups include, but are not limited to, groups such as dioxolanyl, thienyl [l ,3]dithianyl, decahydroisoquinolyl, imidazolinyl, imidazolidinyl, isothiazolidinyl, isoxazolidinyl, morpholinyl, octahydroindolyl,
  • octahydroisoindolyl 2-oxopiperazinyl, 2-oxopiperidinyl, 2-oxopyrrolidinyl, oxazolidinyl, piperidinyl, piperazinyl, 4-piperidonyl, pyrrolidinyl, pyrazolidinyl, quinuclidinyl, thiazolidinyl, tetrahydrofuryl, trithianyl, tetrahydropyranyl, thiomorpholinyl, thiamorpholinyl,
  • hydroxy or “hydroxyl” refers to -OH.
  • hydroxyalkyl refers to an alkyl group having 1 to 6 carbon atoms, which is substituted with a hydroxyl group, e.g., hydroxypropyl.
  • cyano refers to -CN.
  • nitro refers to -N0 2 .
  • urea refers to -NR a -C(0)-NR a 2 or -NR a -C(0)NR a -, wherein R a is H or
  • sulfonylurea refers to -S(0) 2 -NR a -C(0)-NR a - or
  • sulfonamidyl refers to -S(0) 2 -NR a - or -NR a -S(0) 2 -, wherein R a is H or C,-C 6 alkyl.
  • TRPC5 ion channels as described herein include homomultimeric and heteromultimeric structures (e.g., homomultimeric TRPC5 and heteromeric TRPC5-TRPC1 or TRPC5-TRPC4).
  • TRPC5 antagonists include inhibitors having any combination of the structural and/or functional properties disclosed herein.
  • an "effective amount" of, e.g., a TRPC5 antagonist, with respect to the subject methods of inhibition or treatment refers to an amount of the antagonist in a preparation which, when applied as part of a desired dosage regimen brings about a desired clinical or functional result.
  • an effective amount of a TRPC5 antagonist for use in the methods of the present invention includes an amount of a TRPC5 antagonist effective to decrease one or more in vitro or in vivo function of a TRPC5 channel.
  • Exemplary functions include, but are not limited to, membrane polarization (e.g., an antagonist may promote hyperpolarization of a cell), ion flux, ion concentration in a cell, outward current, and inward current.
  • an effective amount is an amount sufficient to inhibit a TRPC5-mediated current and/or the amount sufficient to inhibit TRPC5 mediated ion flux.
  • the TRPC5 anagonists for use in the methods of the present invention may be characterized according to their activity, or lack of activity, against one or more other ion channels.
  • inhibition of a function of such other ion channels is defined similarly.
  • inhibition of an ion channel or an activity of an ion channel means the antagonist inhibits one or more functional activities of the other ion channel.
  • Such functions include the current mediated by the particular ion channel, ion flux, or membrane polarization.
  • preventing is art-recognized, and when used in relation to a condition, such as a local recurrence, a disease such as cancer, a syndrome complex such as heart failure or any other medical condition, is well understood in the art, and includes administration of a composition which reduces the frequency of, or delays the onset of, symptoms of a medical condition in a subject relative to a subject which does not receive the composition.
  • prevention of cancer includes, for example, reducing the number of detectable cancerous growths in a population of patients receiving a prophylactic treatment relative to an untreated control population, and/or delaying the appearance of detectable cancerous growths in a treated population versus an untreated control population, e.g., by a statistically and/or clinically significant amount.
  • Prevention of an infection includes, for example, reducing the number of diagnoses of the infection in a treated population versus an untreated control population, and/or delaying the onset of symptoms of the infection in a treated population versus an untreated control population.
  • Prevention of pain includes, for example, reducing the magnitude of, or alternatively delaying, pain sensations experienced by subjects in a treated population versus an untreated control population.
  • prodrug is intended to encompass compounds that, under physiological conditions, are converted into the therapeutically active agents of the present invention.
  • a common method for making a prodrug is to include selected moieties that are hydrolyzed under physiological conditions to reveal the desired molecule.
  • the prodrug is converted by an enzymatic activity of the host animal.
  • TRPC5 refers to a compound having a molecular weight less than about 2500 amu, preferably less than about 2000 amu, even more preferably less than about 1500 amu, still more preferably less than about 1000 amu, or most preferably less than about 750 amu.
  • TRPC5 includes homomultimeric structures (e.g., homomultimeric TRPC5) and heteromultimeric structures (e.g.,
  • oxidative metabolite is intended to encompass compounds that are produced by metabolism of the parent compound under normal physiological conditions. Specifically, an oxidative metabolite is formed by oxidation of the parent compound during metabolism. For example, a thioether group may be oxidized to the corresponding sulfoxide or sulfone.
  • solvate refers to a compound formed by solvation (e.g., a compound formed by the combination of solvent molecules with molecules or ions of the solute).
  • hydrate refers to a compound formed by the union of water with the parent compound.
  • treating includes prophylactic and/or therapeutic treatments.
  • prophylactic or therapeutic treatment is art-recognized and includes administration to the host of one or more of the subject compositions. If it is administered prior to clinical manifestation of the unwanted condition (e.g., disease or other unwanted state of the host animal) then the treatment is prophylactic, (i.e., it protects the host against developing the unwanted condition), whereas if it is administered after manifestation of the unwanted condition, the treatment is therapeutic, (i.e., it is intended to diminish, ameliorate, or stabilize the existing unwanted condition or side effects thereof).
  • the unwanted condition e.g., disease or other unwanted state of the host animal
  • the compounds are small organic or inorganic molecules, e.g., with molecular weights less than 7500 amu, preferably less than 5000 amu, and even more preferably less than 2000, 1500, 1000, or 600 amu. Such compounds can bind to and inhibit a function of TRPC5.
  • the compounds are nucleic acids, for example, TRPC5 antisense oligonucleotides or TRPC5 RNAi constructs. Such compounds can inhibit the expression of TRPC5, thereby inhibiting the activity of TRPC5.
  • Other exemplary compounds that may act as inhibitors include ribozymes and peptide fragments.
  • Contemplated equivalents of the compounds described above include compounds which otherwise correspond .thereto, and which have the same general properties thereof (e.g., the ability to antagonize TRPC5 activity), wherein one or more simple variations of substituents are made which do not adversely affect the efficacy of the compound.
  • the compound of the present invention may be prepared by the methods illustrated in the general reaction schemes as, for example, described below, or by modifications thereof, using readily available starting materials, reagents and conventional synthesis procedures. In these reactions, it is also possible to make use of variants which are in themselves known, but are not mentioned here.
  • permissible hydrocarbons include acyclic and cyclic, branched and unbranched, carbocyclic and heterocyclic, aromatic and nonaromatic organic compounds which can be substituted or unsubstituted.
  • the compounds described herein can be asymmetric (e.g., having one or more stereocenters). All stereoisomers, such as enantiomers and diastereomers, are intended unless otherwise indicated.
  • An example method includes fractional recrystallizaion using a "chiral resolving acid" which is an optically active, salt-forming organic acid.
  • Suitable resolving agents for fractional recrystallization methods are, for example, optically active acids, such as the D and L forms of tartaric acid, diacetyltartaric acid, dibenzoyltartaric acid, mandelic acid, malic acid, lactic acid or the various optically active camphorsulfonic acids such as ⁇ -camphorsulfonic acid.
  • Other resolving agents suitable for fractional crystallization methods include
  • stereoisomerically pure forms of ct-methylbenzylamine e.g., 5 * and R forms, or diastereomerically pure forms
  • 2-phenylglycinol norephedrine, ephedrine, N-methylephedrine, cyclohexylethylamine, 1 ,2-diaminocyclohexane, and the like.
  • Resolution of racemic mixtures can also be carried out by elution on a column packed with an optically active resolving agent (e.g., dinitrobenzoylphenyl glycine).
  • an optically active resolving agent e.g., dinitrobenzoylphenyl glycine
  • Suitable elution solvent composition can be determined by one skilled in the art.
  • Compounds of the invention also include tautomeric forms, such as keto-enol tautomers.
  • Compounds of the invention can also include all isotopes of atoms occurring in the intermediates or final compounds.
  • the compound of the invention may be radiolabeled with radioactive isotopes, such as for example tritium ( 3 H) or carbon- 4 ( 14 C). All isotopic variations, whether radioactive or not, are intended to be encompassed within the scope of the present invention.
  • Compounds of the invention can exist in unsolvated forms as well as solvated forms, including hydrated forms. In general, the solvated forms are equivalent to unsolvated forms and are encompassed within the scope of the present invention.
  • a compound of formula (I) may exist in multiple crystalline or amorphous forms. In general, all physical forms are equivalent for the uses contemplated by the present invention and are intended to be within the scope of the present invention.
  • salts includes salts of a compound of the invention which are prepared with relatively nontoxic acids or bases.
  • Base addition salts can be obtained by contacting the neutral form of a compound of the invention with a sufficient amount of the desired base, either neat or in a suitable inert solvent.
  • Examples of pharmaceutically acceptable base addition salts include sodium, potassium, calcium, ammonium, organic amino, or magnesium salt, or a similar salt.
  • Acid addition salts can be obtained by contacting the neutral form of compound of the invention with a sufficient amount of the desired acid, either neat or in a suitable inert solvent.
  • pharmaceutically acceptable acid addition salts include those derived from inorganic acids like hydrochloric, hydrobromic, nitric, carbonic,
  • salts of amino acids such as arginate and the like
  • salts of organic acids like glucuronic or galactunoric acids and the like
  • the neutral forms of compound of the invention is preferably regenerated by contacting the salt with a base or acid and isolating the parent compound in the conventional manner.
  • the parent form of the compound differs from the various salt forms in certain physical properties, such as solubility in polar solvents, but otherwise the salts are equivalent to the parent form of the compound for the purposes of the present invention.
  • low enough pyrogen activity refers to a preparation that does not contain a pyrogen in an amount that would lead to an adverse effect (e.g., irritation, fever, inflammation, diarrhea, respiratory distress, endotoxic shock, etc.) in a subject to which the preparation has been administered.
  • an adverse effect e.g., irritation, fever, inflammation, diarrhea, respiratory distress, endotoxic shock, etc.
  • the term is meant to encompass preparations that are free of, or substantially free of, an endotoxin such as, for example, a lipopolysaccharide (LPS).
  • LPS lipopolysaccharide
  • the invention provides methods and compositions for antagonizing a function of a TRPC5 channel in vitro or in vivo. Exemplary functions include, but are not limited to, TRPC5-mediated current.
  • the invention provides methods for treating a disease or disorder or condition by administering compound of the invention.
  • the compound of formula (1) selectively inhibits the expression level and/or activity of a T PC5 protein. In other words, in certain embodiment, the compound of the invention inhibits the activity of a TRPC5 protein preferentially in comparison to the activity of one or more other ion channels.
  • the compound of the invention can be used for preventing or treating anxiety and fear-related disorders (see, e.g., Riccio et al. (2009) Cell 137:761-72).
  • disorders include post-traumatic stress disorder, panic disorder, agoraphobia, social phobias, generalized anxiety disorder, panic disorder, social anxiety disorder, obsessive- compulsive disorder, and separation anxiety.
  • post-traumatic stress disorder panic disorder, agoraphobia, social phobias
  • generalized anxiety disorder panic disorder
  • social anxiety disorder obsessive- compulsive disorder
  • separation anxiety separation anxiety.
  • a compound of the invention is also useful for the treatment of Parkinson's disease, epilepsy, memory disorders, stroke, seizure, and mood disorders.
  • Mood disorders include depression (e.g., major depression, psychiatric depression, dysthymia, and postpartum depression) and bipolar disorder (e.g., bipolar I, bipolar II, and cyclothymia).
  • Memory disorders are conditions associated with any memory loss and may result from Alzheimer's disease, amnesia, aphasia, atherosclerosis, brain injury or disorder, brain tumor, chronic fatigue syndrome, Creutzfedt- Jacob disease, dissociative amnesia, depression, fuge amnesia,
  • Huntington's disease learning disorders, sleeping disorders, multiple personality disorder, pain, post-traumatic stress disorder, schizophrenia, sports injuries, stroke, and Wernicke-Korsakoff syndrome.
  • Pain Treatment of Pain, Sensitivity to Pain and Touch, or Pain-Related Diseases or Disorders
  • a compound of the invention is used to treat or ameliorate pain.
  • exemplary classes of pain that can be treated using a compound of formula (I)in include, but are not limited to nociceptive pain, inflammatory pain, and neuropathic pain.
  • the pain can be chronic or acute.
  • a compound of the invention may be particularly useful in the treatment of pain associated with cancer, osteoarthritis, rheumatoid arthritis, post-herpetic neuralgia, burns, and other indications detailed above.
  • additional exemplary indications for which a compound of the invention can be used include oral pain, pelvic pain, Fabry's disease, complex regional pain syndrome, pancreatitis, and fibromyalgia syndrome.
  • a compound of the invention may also be used in connection with prevention or treatment of sensitivity to pain and touch. Pain or sensitivity to pain and touch may be indicated in a variety of diseases, disorders or conditions, including, but not limited to, diabetic neuropathy, breast pain, psoriasis, eczema, dermatitis, burn, post-herpetic neuralgia (shingles), nociceptive pain, peripheral neuropathic and central neuropathic pain, chronic pain, cancer and tumor pain, spinal cord injury, crush injury and trauma induced pain, migraine, cerebrovascular and vascular pain, sickle cell disease pain, rheumatoid arthritis pain, musculoskeletal pain including treating signs and symptoms of osteoarthritis and rheumatoid arthritis, orofacial and facial pain, including dental, temperomandibular disorder, and cancer related, lower back or pelvic pain, surgical incision related pain, inflammatory and non-inflammatory pain, visceral pain, psychogenic pain and soft tissue inflammatory pain, fibromyalgia
  • the oral pain is due to an injury to the mouth, jaw, lips, gums, or teeth.
  • the oral pain is due to oral surgery, for example, surgery for cancer, tooth extraction, or jaw remodeling.
  • Other conditions that may lead to oral ulcers, and thus oral pain include, but are not limited to chickpox, herpes zoster, infectious mononucleosis, syphilis, tuberculosis, acute necrotizing gingivitis, and burning mouth syndrome.
  • Fibromyalgia is a widespread musculoskeletal pain and fatigue disorder. Fibromyalgia is characterized by pain in the muscles, ligaments, and tendons. The condition affects more women than men, and occurs in people of all ages. Overall, FMS is estimated to afflict 3-6% of the population. Patients have described the pain associated with fibromylagia as deep muscular aching, throbbing, shooting, and stabbing. The pain sometimes includes an intense burning sensation. The pain and stiffness are often worse in the morning or after repetitive use of a particular muscle group.
  • fibromylagia a chronic myelogenous fibromylagia .
  • Other symptoms of fibromylagia include gastrointestinal symptoms. Irritable bowel syndrome and IBS-like symptoms such as constipation, diarrhea, frequent abdominal pain, abdominal gas, and nausea occur in roughly 40 to 70% of FMS patients. Acid reflux or gastroesophogeal reflux disease (GERD) occurs at a similar frequency.
  • GFD gastroesophogeal reflux disease
  • CRPS Complex Regional Pain Syndrome
  • RSD reflex sympathetic dystrophy
  • CRPS is a chronic, painful, and progressive neurological condition that affects skin, muscles, joints, and bones. The syndrome usually develops in an injured limb, such as a broken leg or following surgery. However, many cases involve only a minor injury, such as a sprain, and sometimes no precipitating injurious event can be identified. CRPS involves continuous, intense pain that is disproportionate to the severity of the injury. The pain worsens, rather than improves, over time.
  • CRPS can affect a variety of regions of the body, it most often affects the arms, legs, hands, or feet. Often the pain begins in one portion of a limb, but spreads over time to include the entire limb or even to include a different limb. Typical features include dramatic changes in the color and temperature of the skin over the affected limb or body part,
  • the compounds disclosed herein can also be used to treat endometriosis and the pain associated therewith.
  • Neurodegenerative diseases and disorders include but are not limited to Alzheimer's disease (AD), Parkinson's disease, Huntington's disease, amyotrophic lateral sclerosis (ALS), and other brain disorders caused by trauma or other insults including aging.
  • AD Alzheimer's disease
  • Parkinson's disease Huntington's disease
  • ALS amyotrophic lateral sclerosis
  • other brain disorders caused by trauma or other insults including aging.
  • Mechanisms associated with calcium signaling may be altered in many
  • fibroblasts or T-lymphocytes from patients with AD have consistently displayed an increase in Ca release from intracellular stores compared to controls (Ito et al. (1994) Proc. Natl. Acad. Sci. U.S.A. 91 :534-538; Gibson et al. (1996) Biochem. Biophys. ACTA 1316:71-77; Etchenberrigaray et al. (1998) Neurobiology of Disease, 5:37-45). Consistent with these observations, mutations in presenilin genes (PSl or PS2) associated with familial AD (FAD) have been shown to increase InsP3 -mediated Ca 2+ release from internal stores (Guo et al.
  • TRPC5 is also expressed in the podocyte of the kidney. It has been proposed that there is an antagonistic regulation of actin dynamics and cell in the podocytes by TRPC5 and TRPC6 (Tian et al., (2010) Science Signaling). Thus, inhibiting TRPC5 may impact the reaction of the podocyte to injury.
  • the present invention provides compounds of the invention for use in vitro and in vivo.
  • the present invention also provides compositions and pharmaceutical compositions comprising a compound of formula (I) that inhibits TRPC5 activity.
  • the compound of the invention is selective.
  • the compound of the invention inhibits TRPC5 activity preferentially over the activity of other ion channels.
  • the compound of formula (I) inhibits TRPC5 activity preferentially over TRPVl, TRPV2, TRPV3, TRPV4, TRPC3, TRPC6, TRPC7, TRPA1 , and/or TRPM8 activity.
  • the compound of formula (I) inhibits the activity of TRPC5 and also inhibits the activity of one or more of TRPC4, TRPVl , TRPV2, TRPV3, TRPV4, TRPC3, TRPC6, TRPC7, TRPA1 , and TRPM8.
  • a compound of the invention can be used alone or in combination with other compounds
  • pharmaceutically active agents examples include, but are not limited to, anti-depressants, anti-anxiety agents, anti-epileptic agents, antiinflammatory agents (e.g., NSAIDS, bradykinin receptor antagonists, hormones and autacoids such as corticosteroids), or anti-migraine agents. Certain active agents belong to more than one category.
  • a compound of the invention is conjointly administered with an analgesic.
  • suitable analgesics include, but are not limited to, opioids, glucocorticosteroids, nonsteroidal anti-inflammatories, naphthylalkanones, oxicams, para-aminophenol derivatives, propionic acids, propionic acid derivatives, salicylates, fenamates, fenamate derivatives, pyrozoles, and pyrozole derivatives.
  • analgesic compounds include, but are not limited to, codeine, hydrocodone, hydromorphone, levorpharnol, morphine, oxycodone, oxymorphone, butorphanol, dezocine, nalbuphine, pentazocine, etodolac, indomethacin, sulindac, tolmetin, nabumetone, piroxicam, acetaminophen, fenoprofen, flurbiprofen, ibuprofen, ketoprofen, naproxen, diclofenac, oxaprozin, aspirin, diflunisal, meclofenamic acid, mefanamic acid, prednisolone, and dexamethasone.
  • Preferred analgesics are non-steroidal anti- inflammatories and opioids (preferably moiphine).
  • a compound of the invention can be administered in conjunction with a therapeutic whose administration causes pain.
  • a compound of the invention can be administered in conjunction with an anesthetic, to reduce the pain caused by the administration of the anaesthetic.
  • a compound of the invention can also be administered in conjunction with a chemotherapeutic agent, to reduce the pain caused by administration of the chemotherapeutic agent.
  • a compound of the invention is conjointly administered with a non-steroidal anti-inflammatory.
  • Suitable non-steroidal anti-inflammatory compounds include, but are not limited to, piroxicam, diclofenac, etodolac, indomethacin, ketoralac, oxaprozin, tolmetin, naproxen, flubiprofen, fenoprofen, ketoprofen, ibuprofen, mefenamic acid, sulindac, apazone, phenylbutazone, aspirin, celecoxib and rofecoxib.
  • a compound of the invention While it is possible for a compound of the invention to be administered alone, it is preferable to administer the compound as a pharmaceutical formulation, where the compound is combined with one or more pharmaceutically acceptable excipients or carriers.
  • the compound of the invenyion may be formulated for administration in any convenient way for use in human or veterinary medicine.
  • the compound of the invention may be a prodrug, e.g., capable of being converted to an active compound in a physiological setting.
  • phrases "pharmaceutically acceptable” is employed herein to refer to those compounds, materials, compositions, and/or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit/risk ratio.
  • Examples of pharmaceutically acceptable carriers include: (1 ) sugars, such as lactose, glucose and sucrose; (2) starches, such as corn starch and potato starch; (3) cellulose, and its derivatives, such as sodium carboxymethyl cellulose, ethyl cellulose and cellulose acetate; (4) powdered tragacanth; (5) malt; (6) gelatin; (7) talc; (8) excipients, such as cocoa butter and suppository waxes; (9) oils, such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil and soybean oil; (10) glycols, such as propylene glycol; (1 1 ) polyols, such as glycerin, sorbitol, marmitol and polyethylene glycol; (12) esters, such as ethyl oleate and ethyl laurate; (13) agar; (14) buffering agents, such as magnesium hydroxide and aluminum hydroxide; (15)
  • antioxidants examples include: (1) water soluble antioxidants, such as ascorbic acid, cysteine hydrochloride, sodium bisulfate, sodium
  • antioxidants such as ascorbyl palmitate, butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), lecithin, propyl gallate, alpha-tocopherol, and the like
  • metal chelating agents such as citric acid
  • EDTA ethylenediamine tetraacetic acid
  • sorbitol sorbitol
  • tartaric acid tartaric acid
  • phosphoric acid and the like.
  • Solid dosage forms can include one or more pharmaceutically acceptable carriers, such as sodium citrate or dicalcium phosphate, and/or any of the following: (1) fillers or extenders, such as starches, lactose, sucrose, glucose, mannitol, and/or silicic acid; (2) binders, such as, for example, carboxymethylcellulose, alginates, gelatin, polyvinyl pyrrolidone, sucrose and/or acacia; (3) humectants, such as glycerol; (4) disintegrating agents, such as agar-agar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates, and sodium carbonate; (5) solution retarding agents, such as paraffin; (6) absorption accelerators, such as quaternary ammonium compounds; (7) wetting agents, such as, for example, cetyl alcohol and glycerol
  • Liquid dosage forms can include pharmaceutically acceptable emulsions,
  • the liquid dosage forms may contain inert diluents commonly used in the art, such as, for example, water or other solvents, solubilizing agents and emulsifiers, such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1 ,3-butylene glycol, oils (in particular, cottonseed, groundnut, corn, germ, olive, castor and sesame oils), glycerol, tetrahydrofuryl alcohol, polyethylene glycols and fatty acid esters of sorbitan, and mixtures thereof.
  • inert diluents commonly used in the art, such as, for example, water or other solvents, solubilizing agents and emulsifiers, such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol
  • Suspensions in addition to the compound of the invention, may contain suspending agents as, for example, ethoxylated isostearyl alcohols, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum metahydroxide, bentonite, agar-agar and tragacanth, and mixtures thereof.
  • suspending agents as, for example, ethoxylated isostearyl alcohols, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum metahydroxide, bentonite, agar-agar and tragacanth, and mixtures thereof.
  • Ointments, pastes, creams and gels may contain, in addition to the compound of the invention, excipients, such as animal and vegetable fats, oils, waxes, paraffins, starch, tragacanth, cellulose derivatives, polyethylene glycols, silicones, bentonites, silicic acid, talc and zinc oxide, or mixtures thereof.
  • excipients such as animal and vegetable fats, oils, waxes, paraffins, starch, tragacanth, cellulose derivatives, polyethylene glycols, silicones, bentonites, silicic acid, talc and zinc oxide, or mixtures thereof.
  • Powders and sprays can contain, in addition to the compound of the invention, excipients such as lactose, talc, silicic acid, aluminum hydroxide, calcium silicates and polyamide powder, or mixtures of these substances.
  • Sprays can additionally contain customary propellants, such as chlorofluorohydrocarbons and volatile unsubstituted hydrocarbons, such as butane and propane.
  • the formulations may conveniently be presented in unit dosage form and may be prepared by any methods well known in the art of pharmacy.
  • the amount of a compound of the invention which can be combined with a carrier material to produce a single dosage form will vary depending upon the host being treated, the particular mode of administration.
  • the amount of a compound of the invention that can be combined with a carrier material to produce a single dosage form will generally be that amount of the compound which produces a therapeutic effect. Generally, out of one hundred percent, this amount will range from about 1 percent to about ninety-nine percent of a compound of the invention, preferably from about 5 percent to about 70 percent, most preferably from about 10 percent to about 30 percent.
  • the tablets, and other solid dosage forms of the pharmaceutical compositions disclosed herein, such as dragees, capsules, pills and granules, may optionally be scored or prepared with coatings and shells, such as enteric coatings and other coatings well known in the
  • compositions may also be formulated so as to provide slow or controlled release of the active ingredient therein using, for example, hydroxypropylmethyl cellulose in varying proportions to provide the desired release profile, other polymer matrices, liposomes and/or microspheres. They may be sterilized by, for example, filtration through a bacteria- retaining filter, or by incorporating sterilizing agents in the form of sterile solid compositions that can be dissolved in sterile water, or some other sterile injectable medium immediately before use. These compositions may also optionally contain pacifying agents and may be of a composition that they release the active ingredient(s) only, or preferentially, in a certain portion of the gastrointestinal tract, optionally, in a delayed manner. Examples of embedding
  • compositions that can be used include polymeric substances and waxes.
  • the active ingredient can also be in micro-encapsulated form, if appropriate, with one or more of the above-described excipients.
  • the active compound may be mixed under sterile conditions with a
  • pharmaceutically acceptable carrier and with any preservatives, buffers, or propellants that may be required.
  • the formulations disclosed herein can be delivered via a device.
  • Exemplary devices include, but are not limited to, a catheter, wire, stent, or other intraluminal device. Further exemplary delivery devices also include a patch, bandage, mouthguard, or dental apparatus.
  • Transdermal patches have the added advantage of providing controlled delivery of a compound of the invention to the body. Such dosage forms can be made by dissolving or dispersing the compound of the invention in the proper medium.
  • Absorption enhancers can also be used to increase the flux of the compound across the skin. The rate of such flux can be controlled by either providing a rate controlling membrane or dispersing the compound in a polymer matrix or gel.
  • Ophthalmic formulations are also contemplated as being within the scope of this invention.
  • the absorption of the drug in order to prolong the effect of a drug, it is desirable to slow the absorption of the drug from subcutaneous or intramuscular injection. This may be accomplished by the use of a liquid suspension of crystalline or amorphous material having poor water solubility. The rate of absorption of the drug then depends upon its rate of dissolution, which, in turn, may depend upon crystal size and crystalline form. Alternatively, delayed absorption of a parenterally administered drug form is accomplished by dissolving or suspending the drug in an oil vehicle.
  • Injectable depot forms are made by forming microencapsule matrices of the subject compounds in biodegradable polymers such as polylactide-polyglycolide. Depending on the ratio of drug to polymer, and the nature of the particular polymer employed, the rate of drug release can be controlled. Examples of other biodegradable polymers include poly(orthoesters) and poly(anhydrides). Depot injectable formulations are also prepared by entrapping the drug in liposomes or microemulsions that are compatible with body tissue.
  • a compound of the invention When a compound of the invention is administered as a pharmaceutical, to humans and animals, it can be given per se or as a pharmaceutical composition containing, for example, 0.1 to 99.5% (more preferably, 0.5 to 90%) of the compound of the invention in combination with a pharmaceutically acceptable carrier.
  • the formulations can be administered topically, orally, transdermally, rectally, vaginally, parentally, intranasally, intrapulmonary, intraocularly, intravenously, intramuscularly, intraarterially, intrathecally, intracapsularly, intraorbitally, intracardiacly, intradermally, intraperitoneal ly, transtracheal ly, subcutaneously, subcuticularly, intraarticularly, subcapsularly, subarachnoid ⁇ , intraspinally, intrasternally, sublingually, or by inhalation.
  • Actual dosage levels of the the compound of the invention in the pharmaceutical compositions of this invention may be varied so as to obtain an amount of the active ingredient that is effective to achieve the desired therapeutic response for a particular patient, composition, and mode of administration, without being toxic to the patient.
  • the selected dosage level will depend upon a variety of factors including the activity of the compound of the invention disclosed herein employed, or the ester, salt or amide thereof, the route of administration, the time of administration, the rate of excretion of the particular compound being employed, the duration of the treatment, other drugs, compounds and/or materials used in combination with the particular compound employed, the age, sex, weight, condition, general health and prior medical history of the patient being treated, and like factors well known in the medical arts.
  • a physician or veterinarian having ordinary skill in the art can readily determine and prescribe the effective amount of the pharmaceutical composition required.
  • the physician or veterinarian could start doses of the compound of the invention in the
  • composition at levels lower than that required in order to achieve the desired therapeutic effect and gradually increase the dosage until the desired effect is achieved.
  • a suitable daily dose of a compound of the invention will be that amount of the compound that is the lowest dose effective to produce a therapeutic effect. Such an effective dose will generally depend upon the factors described above.
  • intravenous, intracerebro ventricular and subcutaneous doses of the compounds of the invention for a patient will range from about 0.0001 to about 100 mg per kilogram of body weight per day.
  • the dose can be 0.1 -50, 0.1 -25, 0.5-10, 1 -10, or 5-10 mg/kg.
  • the effective daily dose of the compound of the invention may be administered as two, three, four, five, six or more sub-doses administered separately at appropriate intervals throughout the day, optionally, in unit dosage forms.
  • a compound of the invention which antagonizes TRPC5 function may be useful in the prophylaxis and treatment of any of the foregoing injuries, diseases, disorders, or conditions.
  • its efficacy can be readily tested in one or more animal models.
  • numerous well known animal models exist.
  • One or more suitable animal models e.g., suitable in light of the particular indication) can be selected.
  • Fear-related behaviors can be measured as described, e.g., in Riccio et al. Pain behaviors can be studied using various agents or procedures to simulate pain resulting from injuries, diseases, or other conditions. Blackburn-Munro (2004) Trends in Pharmacological Sciences 25: 299-305 (see, for example, Table 1). Behavioral characteristics of challenged animals can then be observed. Compounds or procedures that may reduce pain in the animals can be readily tested by observing behavioral characteristics of challenged animals in the presence versus the absence of the test compound(s) or procedure.
  • Exemplary behavioral tests used to study chronic pain include tests of spontaneous pain, allodynia, and hyperalgesia. Id. To assess spontaneous pain, posture, gait, nocifensive signs (e.g., paw licking, excessive grooming, excessive exploratory behavior, guarding of the injured body part, and self-mutilation) can be observed. To measure evoked pain, behavioral responses can be examined following exposure to heat (e.g., thermal injury model).
  • heat e.g., thermal injury model
  • Exemplary animal models of pain include, but are not limited to, the Chung model, the carageenan induced hyperalgesia model, the Freund's complete adjuvant induced hyperalgesia model, the thermal injury model, the formalin model and the Bennett Model.
  • the Chung model of neuropathic pain involves ligating one or more spinal nerves. Chung et al. (2004) Methods Mol Med 99: 35-45; Kim and Chung (1992) Pain 50: 355-363. Ligation of the spinal nerves results in a variety of behavioral changes in the animals including heat hyperalgesia, cold allodynia, and ongoing pain. Compounds that antagonize TRPC5 can be administered to ligated animals to assess whether they diminish these ligation-induced behavioral changes in comparison to that observed in the absence of compound.
  • Useful anxiety and depression models include the maternal separation model, the elevated plus-maze model, the forced swim test, the tail suspension test, the light/dark preference model, the light-enhanced startle model, and the ultrasonic vocalization model.
  • Useful seizure models include but are not limited to maximal electric shock (MES), acoustic startle in susceptible animals (eg DBA mice), and chemical induced seizure (with proconvulsant compounds such as pilocarpine, pentalene tetrazole, kainic acid, N-methyl-D- aspartic acid).
  • MES maximal electric shock
  • acoustic startle in susceptible animals eg DBA mice
  • chemical induced seizure with proconvulsant compounds such as pilocarpine, pentalene tetrazole, kainic acid, N-methyl-D- aspartic acid.
  • Useful models of kidney function include the LPS-induced proteinuria (waiting for a reference for others).
  • Patch clamp experiments permit the detection of currents through the TRPC5 channel in the cell line described above.
  • a glass electrode is brought into contact with a single cell and a high-resistance (gigaohm) seal is established with the cell membrane.
  • the membrane is then ruptured to achieve the whole-cell configuration, permitting control of the voltage of the cell membrane and measurement of currents flowing across the membrane using the amplifier attached to the electrode and resulting in the replacement of cytoplasm with the pipette solution.
  • a perfusion system permits control of the extracellular solution, including the addition of blockers and activators of the current.
  • the current can be activated by including 1 .4 ⁇ free Ca 2+ in the pipette (intracellular) solution, and 80 ⁇ LaCl 3 in the extracellular solution.
  • TRPC5 cells were induced 20-48 hours, removed from growth plates, and replated at low density (to attain good single-cell physical separation) on glass coverslips for measurement. In some cases, cells were grown in low density overnight on glass coverslips. Patch clamp recordings were made in the whole-cell mode with a holding potential of -40 mV. Every 5 seconds, a voltage ramp was applied from -120 to +100 mV, 400 ms in duration. Currents elicited were quantified at -80 mV and +80 mV.
  • the internal solution consisted of 140 mM cesium aspartate, 10 mM HEDTA, 2 mM CaCl 2 , 2.27 mM MgCl 2 and 10 mM HEPES, pH 7.2, with 1 ,400 nM calculated free Ca 2+ .
  • the external solution consisted of 150 mM NaCl, 4.5 mM KCl, 1 mM MgCl 2 , 2 mM CaCl 2, 10 mM HEPES, 10 mM glucose, 1 mM EGTA, pH 7.4.
  • TRPC5 current was induced only in TRPC5-expressing cells and not in parental HEK293 TREx cells. Removal of the LaCl 3 stimulus causes most of the current to go away. Potential blockers were tested for ability to block both inward and outward currents in the continued presence of LaCl 3 .
  • IC 50 of a compound of the invention was estimated by testing the compound at 5 ⁇ and 500 nM. When 5 ⁇ of a compound showed no block, IC50 was estimated as >10 ⁇ . When 5 ⁇ of a compound showed 50% or less block, a rough estimate of IC 50 in the range of 5-10 ⁇ could be made. IC50 for a compound of Formula I or Formula II between 500 nM and 5 ⁇ was similarly estimated.
  • a compound described herein may be tested for its ability to block both inward and outward currents through the TRPC5 channel, e.g., by an assay as described in Example 1.
  • IC 50 of a compound of the invention was estimated by testing the compound at 5 ⁇ and 500 nM. When 5 ⁇ of a compound showed no block, IC 50 was estimated as >10 ⁇ . When 5 ⁇ of a compound showed 50% or less block, a rough estimate of IC 5 o in the range of 5-10 ⁇ could be made. IC 50 for a compound of the invention between 500 nM and 5 ⁇ was similarly estimated. Exemplary compounds are shown in Table B below. As shown in Table B, "A" refers to an 1C 50 ⁇ 100 nM.
  • B refers to an IC 50 between 100 nM and 500 nM.
  • C refers to an IC 5 o between 500 nM and 1000 nM.
  • D refers to an IC 50 between 1 ⁇ and 2 ⁇ .
  • E refers to an 1C 50 between 2 ⁇ and 10 ⁇ .
  • F refers to agonist compounds.
  • ND refers to compounds wherein the IC50 was not determined.
  • the high throughput assay depended on detection of the rise in intracellular Ca concentration ([Ca ],) following channel activation in cells inducibly expressing the TRPC5 channel.
  • Ca 2+ rise was quantified with the use of fluorescent Ca 2+ indicators that were loaded into cells and thereafter indicated the [Ca 2+ ]j Ca 2+ influx following activation of the TRPC5 channel. Compounds inhibiting the [Ca ]j rise were considered hits for further investigation.
  • the commercially available HEK293/TREx line Invitrogen was stably transfected with a TRPC5 construct and screened by conventional calcium imaging to find clones with TRPC5 expression following stimulation with 1 g/ml tetracycline.
  • HBSS Hank's Balanced Salt Solution
  • 0.185 g/1 D-glucose 0.9767 g/1 MgS0 4 (anhydrous)
  • 0.4 g/1 KCl 0.06 g/1 KH 2 P0 4 (anhydrous)
  • 0.35 g/1 NaHC0 3 8.0 g/1 NaCl, and 0.04788 g/1 Na 2 HP0 4 (anhydrous); pH 7.4 was then added to each well.
  • cells were assayed using the Hamamatsu FDSS 6000 system, which permitted illumination alternately at 340 nm and 380 nm for Fura-2 experiments, or at 485 nm for Fluo4 experiments. Frames were acquired at a rate of 0.2 Hz. During the assay, the plates were continuously vortexed, with pipette mixing of wells following addition of each reagent.
  • Negative controls consisted of HEK293/TREx TRPC5 cells exposed to high Ca 2+ solution, but no compound.
  • Positive control conditions consisted of addition of 2-APB, a promiscuous blocker of TRPC5 and other channels, to columns 23 and 24 of the plates, to a final
  • efficacy can be represented as % inhibition in the presence (of a given concentration of compound) versus the absence of compound or in comparison to a control compound.
  • efficacy can be represented as % inhibition of ion flux in the presence versus the absence of compound.
  • Exemplary compounds are shown in Table X below. As shown in Table X, "A” refers to an IC 50 ⁇ 100 nM. "B” refers to an IC 50 between 100 nM and 500 nM. “C” refers to an IC 50 between 500 nM and 1000 nM. “D” refers to an IC 50 between 1 ⁇ and 2 ⁇ . “E” refers to an IC 50 between 2 ⁇ and 10 ⁇ . “F” refers to agonist compounds. “ND” refers to compounds wherein the 1C 50 was not determined.
  • Reactions were monitored via TLC on silica gel plates (either 60 HSGF254 percolated plates (0.15-0.2 mm Si0 2 ) or Baker-flex IB2-F TLC plates), and visualized using UV light (254 nm or 365 nm) and/or staining with a solution of DNP (12 g, 2,4-dinitrofenylhydrazin, 60 mL concentrated H 2 S0 , 80 ml H 2 0, 200 mL ethanol) and subsequent heating or monitored via LCMS.
  • DNP 2,4-dinitrofenylhydrazin
  • Microwave reactions were carried out with a Biotage Smith Synthesizer.
  • LCMS were performed on a SHIMADZU LCMS-2010EV instrument using one of two sets of conditions.
  • Dess Martin [or Dess Martin periodinane] l,l,l-triacetoxy-l ,l -dihydro-l ,2-benziodoxol- 3(lH)-one
  • Step 1 8-bromo-7-ethyl-l -(3-hydroxypropyl)-3-methyl-l H-purine-2,6(3H,7H)-dione
  • Step 2 8-bromo-7-ethyl-3-methyl-l -(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-l H-purine- 2,6(3H,7H)-dion
  • Step 1 8-bromo-3-meth -lH-purine-2,6(3H,7H)-dio
  • Step 1 8-bromo-3-methyl-7-((2-(trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H,7H)-dione
  • Step 2 3 -methyl -8-(3 -(tri fluoromethoxy)phenoxy)-7-((2-(trimethy 1 si ly l)ethoxy)methyl)- 1 H- purine-2,6(3H,7H)-dione
  • Step 3 3-methyl-l-(2-(2-(tetrahydro-2H-pyran-2-yloxy)ethoxy)ethyl)-8-(3- (trifluoromethoxy)phenoxy)-7-((2-(trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H,7H)-dione
  • Step 4 1 -(2-(2-hydroxyethoxy)ethyl)-3-methyl-8-(3-(trifluoromethoxy)phenox
  • Step 1 3-methyl-l-(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-8-(3-(trifluoromethoxy)phenoxy)- 7-((2-(trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H,7H)-dione
  • Step 2 1 -(3-hydroxypropyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)- dione
  • Step 1 8-bromo-3.-methyl- 1 -(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-7-((2
  • Step 2 8-bromo-l -(3-hydroxypropyl)-3-methyl-lH-purine-2,6(3H,7H)-dione
  • Step 2 3-methyl-l -(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-8-(2-(3- (trifluoromethoxy)phenoxy)ethoxy)- 1 H-purine-2,6(3 H,7H)-dione
  • Step 3 7-benzyl- -((2-(trimethylsilyl)ethoxy)methyl)-l H-purine-2,6(3H,7H)-dione
  • Step 4 7-benzyl-8-chloro-3-((2-(trimethylsilyl)ethoxy)methyl)-l H-purine-2,6(3H,7H)-dione
  • Step 5 7-benzyl-8-chloro-l-(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-3-((2- (trimethyls)
  • Step 1 7-benzyl-8-chloro-l -(3-hydroxypropyl)-lH-purine-2,6(3H,7H)-dione
  • Step 2 7-benzyl-8-chloro-3-ethyl-l -(3-hydroxypropyl)-lH-purine-2,6(3H,7H)-dione
  • Step 3 7-benzyl- -ethoxy-3 -ethyl- 1 -(3 -hydroxypropyl)- 1 H-purine-2,6(3H,7H)-dione
  • Step 4 8-ethox -3-ethy]-l -(3-hydroxypropyl)-lH-purine-2,6(3H,7H)-dione
  • Step 1 l ,4-dioxaspiro[4.5]decane (JF-000357-022) (CH 2 OH) 2 ⁇ _ o ⁇
  • Step 1 8-(2-fluoro-5-(trifluoromethoxy)benzyl)-l-(4-methoxybenzyl)-3-methyl-lH- purine-2,6(3H,7H)-dione
  • Step 2 7-(4-chlorobenzyl)-8-(2-fluoro-5-(trifluoromethoxy)benzyl)- 1 -(4- methoxybenzyl)-3-
  • Step 3 7-(4-chlorobenzyl)-8-(2-fluoro-5-(trifluoromethoxy)benzyl)-3-methyl-l H-purine-
  • Step 2 7-benzy -8-(3-(trifluoromethoxy)phenoxy)-lH-purine-2 ) 6(3H,7H)-dione
  • Step 3 7-benzyl-3-ethyl-8-(3-(trifluoromethoxy)phenoxy)-l H-purine-2,6(3H,7H)-dione
  • Step 1 N-(6-amino-3-(4-methoxybenzyl)-l -methyl-2,4-dioxo-l ,2,3,4-tetrahydropyrimidin-5- yl)-2-(3-(trifluoromethoxy)phenyl)acetamide
  • Step 2 1 -(4-methoxybenzyl)-3-methyl-8-(3-(trifluoromethoxy)benzyl)-lH-purine-2,6(3H,7H)- dione
  • Step 2 ethyl 3-(5-methylthiazol-2-yl)propanoate -Sy Pd/c, H, s v
  • Step 1 1 -(4-methoxybenzyl)-3-methyl-7-((5-methylpyridin-2-yl)methy])-8-(3- (trifluoromethox
  • Step 1 8-azido-7-(4-chlorobenzyl)-3-methyl-l-(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-lH- purine-2,6(3H,7 -dione
  • Step 2 8-amino-7-(4-chlorobenzyl)-3-methy]-l -(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-lH- purine-2,6(3H,7H)-dione
  • Step 1 8-bromo-l-(3-hydroxybutyl)-3-methyl-7-((2-(trimethylsilyl)ethoxy)methyl) purine-2,6(3H,7H)-dione
  • Step 3 l-(3-hydroxybutyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-7-((2- (trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H,7H)-dione
  • Step 1 (E)-N'-(l -(4-methoxybenzyl)-3-methyl-2,6-dioxo- 1,2,3, 6-tetrahydropyrimidin-4- yl)-N,N-dimethylformimidami
  • Step 4 1 -(4-methoxybenzyl)-3-methyl-8-(3-(3-(trifluoromethoxy)phenoxy)propyl)-l H- purine-2,6(3H,7H)-dione
  • Step 2 4-chlorobenzyl)-lH-purine-2,6( -dione
  • Step 4 8-chloro-7-(4-chlorobenzyl)-3-'((2-(trimethylsilyl)ethoxy)methyl)-lH-purine- 2,6(3H,7H)-dione
  • Step 5 7-benzyl-l -(3-(tert-butyldimethylsilyloxy)propyl)-8-chloro-3-((2- (trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H,7H)-dione
  • Step 6 chloro-7-(4-chlorobenzyl)-l -(3-hydroxypropyl)- l H-purine-2,6(3H,7H)-dione
  • Steps 1 and 2 8-bromo-7-(4-fluorobenzyl)-3 -methyl- 1 -(3 -(tetrahydro-2H-pyran-2- yloxy)propyl)-lH-purine-2,6(3H,7H)-dione
  • Step 3 8-bromo-7-(4-fluorobenzyl)-l -(3-hydroxypropyl)-3-methyl-l H-purine- 2,6(3H,7H)-dione
  • Step 2 7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-2,3,6,7-tetrahydro-lH-purine-8-sulfonyl chloride

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Abstract

Compounds, compositions and methods are described for inhibiting the TRPC5 ion channel and disorders related to TRPC5.

Description

SUBSTITUTED XANTHINES AND METHODS OF USE THEREOF
CLAIM OF PRIORITY
This application claims priority to U.S. S.N. 61/789,724, filed March 15, 2013, the contents of which are incorporated herein by reference.
BACKGROUND
A variety of ion channel proteins exist to mediate ion flux across cellular membranes. The proper expression and function of ion channel proteins is . essential for the maintenance of cell function, intracellular communication, and the like. Numerous diseases are the result of misregulation of membrane potential or aberrant calcium handling. Given the central importance of ion channels in modulating membrane potential and ion flux in cells, identification of agents that can promote or inhibit particular ion channels are of great interest as research tools and as possible therapeutic agents.
SUMMARY OF THE INVENTION
The present invention provides methods of treating a TRPC5 mediated disorder in a subject, comprising administering to the sub ect a compound of Formula I:
Figure imgf000003_0001
Formula I or a pharmaceutically acceptable salt thereof, wherein constituent members are provided herein.
The present invention further rovides compounds of Formula 1(a):
Figure imgf000003_0002
Formula 1(a)
or a pharmaceutically acceptable salt thereof, wherein constituent members are provided herein.
The present invention further provides compounds of Formula II:
Figure imgf000004_0001
Formula II
or a pharmaceutically acceptable salt thereof, wherein constituent members are provided herein.
The present invention further provides compounds of Formula III:
Figure imgf000004_0002
Formula III
or a pharmaceutically acceptable salt thereof, wherein constituent members are provided herein.
The present invention further provides compositions comprising a compound of Formula 1(a), II or III, and a pharmaceutically acceptable carrier.
The present invention further provides methods of treating a TRPC5 mediated disorder in a subject, e.g. a human, comprising administering to the subject a compound or composition of a compound of Formula 1(a), II or Formula III, or a pharmaceutically acceptable salt thereof.
The present invention provides methods, compounds and compositions for treating conditions such as a neuropsychiatric disorder, a neurodegenerative disorder, nephropathy, and seizure disorder by modulating the activity of the transient receptor potential cation channel subfamily C, member 5 (TRPC5), which can exist in homomultimeric form as well as heteromultimeric form with other ion channels such as TRPC1 or TRPC3 (i.e., TRPC5-TRPC1 and TRPC 1 -TRPC3-TRPC5). The compounds described herein modulate the function of TRPC5 by inhibiting a TRPC5-mediated ion flux or by inhibiting the inward current, the outward current, or both currents mediated by TRPC5. The inhibition of a particular current is the ability to inhibit or reduce such current (e.g., inward and/or outward) in an in vitro or an in vivo assay. The activation of a particular current is the ability to activate or increase such current (e.g., inward and/or outward) in an in vitro or an in vivo assay.
In one aspect, the invention relates to a method for treating a condition for which reduced
TRPC5 activity can reduce the severity of the condition, by administering a TRPC5 antagonist, such as a compound as described herein that inhibits TRPC5-mediated current and/or TRPC5- mediated ion flux. Described herein are compounds, which are TRPC5 antagonists that have a measured IC50 for inhibition of TRPC5 of 10 nanomolar or less. In certain embodiments, the compounds described herein, which are TRPC5 antagonists inhibit one or both of inward and outward TRPC5-mediated currents with an IC50 10 nanomolar or less. In certain embodiments, the compounds described herein inhibit at least 95% of TRPC5-mediated current or TRPC5- mediated ion flux when administered at 1 micromolar or less.
In another aspect, the compounds described herein, which are TRPC5 antagonists can be used to inhibit a function of TRPC5, for example a TRPC5-mediated current and/or a TRPC5- mediated ion flux. In some embodiments, the compounds described herein can be used to inhibit a TRPC5 mediated current in vitro, for example in cells in culture. In other embodiments, the compounds described herein can be used to inhibit a TRPC5 mediated current in vivo. In certain embodiments, the compounds described herein inhibit both an inward and an outward TRPC5- mediated current.
Another aspect of the invention features a pharmaceutical preparation suitable for use in a human patient, or for veterinary use, comprising an effective amount of a compound described herein (or a salt thereof, or a solvate, hydrate, oxidative metabolite or prodrug of the compound or its salt), and one or more pharmaceutically acceptable excipients. The invention further contemplates the use of the compounds described herein in the manufacture of a medicament or pharmaceutical preparation to treat or reduce the symptoms of any of the diseases or conditions provided in the specification. The compounds described herein can be used for treating a particular disease or condition can be formulated for administration via a route appropriate for the particular disease or condition.
The compounds described herein can be administered alone or in combination with another therapeutic agent. For instance, the compounds described herein can be administered conjointly with one or more of an anti-inflammatory agent, anti-acne agent, anti-wrinkle agent, anti-scarring agent, anti-psoriatic agent, anti-proliferative agent, anti-fungal agent, anti-viral agent, anti-septic agent, anti-migraine agent, keratolytic agent, or a hair growth inhibitor.
The compounds described herein can be administered topically, orally, transdermally, rectally, vaginally, parentally, intranasally, intrapulmonary, intraocularly, intravenously, intramuscularly, intraarterially, intrathecally, intracapsularly, intraorbitally, intracardiacly, intradermal ly, intraperitoneally, transtracheally, subcutaneously, subcuticularly, intraarticularly, subcapsularly, subarachnoidly, intraspinally, intrasternally, sublingually, or by inhalation.
In some embodiments, the compounds described herein can be administered topically.
In some embodiments, the compounds described herein can be administered orally.
In some embodiments, the compounds described herein can be administered administered parentally.
BRIEF DESCRIPTION OF THE DRAWINGS
FIG. 1 is a plot depicting NMRI mouse immobility after administration of 0.5%
Methylcellulose (MC) (4000 CPS) orally, or Compound 260 at oral doses of 0.3, ) , 3 and 10 mg/kg one hour prior to testing, or desipramine by intraperitoneal injection of 30 mg/kg, 30 minutes prior to testing.
FIGs. 2A and 2B are plots depicting activity in the Vogel Conflict Test Model for Wistar- Kyoto rats after administration of 0.5% Methylcellulose (4000 CPS), or Compound 260 at doses of 0.3, 1 , 3 and 10 mg/kg orally 1 hour prior to testing, or Midazolam 1 mg/kg by intraperitoneal injection, 30 minutes prior to testing.
FIGs. 3A-3D are plots depicting anxiolytic effects of Wistar-Kyoto rats employed in an elevated-plus maze assay using Midazolam lmg/kg dose and 0.5% methylcellulose as positive and negative controls, respectively, as well as Compound 260 at doses of 0.3, 1 , 3 and 10 mg/kg. FIGs. 3A and 3B are plots depicting % entries of the rats in the open arm portions of the maze. FIG. 3C is a plot depicting entries of the rats in the closed arm portion of the maze. FIG. 3D is a plot depicting the distance travelled (in centimeters) by the rats.
FIGs. 4A and 4B are line graphs depicting stimulant effects on NMRI mice after administration of phencyclidine (PCP) at 5mg/kg delivered subcutaneously or with 0.5% methylcellulose (MC) (vehicle) delivered subcutaneously (FIG. 4A), or with Compound 260 at doses of 0.3, 1 , 3 and 10 mg/kg in MRI mice (FIG. 4B).
DETAILED DESCRIPTION OF THE INVENTION
Cation channels such as TRPC5 modulate the flux of calcium and sodium ions across cellular membranes. Sodium and calcium influx leads to a depolarization of the cell. This increases the probability that voltage-gated ion channels will reach the threshold required for activation. As a result, activation of non-selective cation channels can increase electrical excitability and increase the frequency of voltage-dependent events. Voltage-dependent events include, but are not limited to, neuronal action potentials, cardiac action potentials, smooth muscle contraction, cardiac muscle contraction, and skeletal muscle contraction.
Calcium influx caused by the activation of non-selective cation channels such as TRPC5 also alters the intracellular free calcium concentration. Calcium is a ubiquitous second messenger molecule within the cell and the alterations in intracellular calcium levels have profound effects on signal transduction and gene expression. Thus, activation of non-selective cation channels such as TRPC5 can lead to changes in gene expression and cellular phenotype. Gene expression events include, but are not limited to, production of mRNAs encoding cell surface receptors, ion channels, and kinases. These changes in gene expression can lead to hyperexcitability in that cell.
Transient receptor potential (TRP) horhomeric TRPC5 ion channels are signal
04- transduction gated, Ca -permeable channels predominantly expressed in the neurons. TRPC5 forms homomultimeric structures such as tetramers (i.e., TRPC5 homomultimers) and
heteromultimeric structures such as tetramers (i.e., TRPC5-TRPC1 heteromultimers). Unless expressly stated otherwise, when the term TRPC5 is used herein, for example, when identifying a modulator of TRPC5 such as a TRPC5 antagonist, the term TRPC5 is used generically so as to include either or both of a TRPC5 homomultimer or a heteromultimer (e.g., TRPC5-TPRC1 or TRPC5-TRPC4 heteromultimer). Examples of TRPC5 in the literature include the following: Nature. 2008 Jan. 3; 451 (7174):69-72; Mol Pharmacol. 2008 January; 73 (l):42-9; J Biol Chem. 2007 Nov. 16; 282 (46):33868-78; Biochem Biophys Res Commun. 2008 Jan. 1 1 ; 365 (2):239- 45; J Biol Chem. 2006 Nov. 3; 281 (44):33487-96; Eur J Pharmacol. 2005 Mar. 14; 510 (3):217- 22; J Biol Chem. 2006 Feb. 24; 281 (8):4977-82; Biochem Soc Trans. 2007 February; 35 (Pt l): 101-4; Handb Exp Pharmacol. 2007; (179): 109-23; J Biol Chem. 2005 Mar. 25; 280
(12):10997-1006; J Physiol. 2006 Jan. 15; 570 (Pt 2):219-35; and Nat Neurosci. (2003) 6: 837-
45.
Modulating the function of TRPC5 proteins provides a means of modulating calcium homeostasis, sodium homeostasis, membrane polarization, and/or intracellular calcium levels, and compounds that can modulate TRPC5 function are useful in many aspects, including, but not limited to, maintaining calcium homeostasis, modulating intracellular calcium levels, modulating membrane polarization, and treating or preventing diseases, disorders, or conditions associated with calcium and/or sodium homeostasis or dyshomeostasis.
In one aspect, the invention provides methods of treating a TRPC5 mediated disorder in a subject comprising administering to the sub ect a compound of Formula I:
Figure imgf000008_0001
Formula I or a pharmaceutically acceptable salt thereof, wherein:
R1 is C]-C6 alkyl, C2-C6 alkenyl or C2-C6 alkynyl, each of which is optionally substituted with 1-4 R5;
R2 is Cf-C6 alkyl, C C6 heteroalkyl, C2-C6 alkenyl, C2-C6 alkynyl, Ci-C6 haloalkyl, halo, Ci-C6 haloalkoxy, hydroxyl, Ci-C6 alkoxy, C3-C7 cycloalkyloxy, C6-Cio aryl, C6-Ci0 aryloxy, C7- Ci6 arylalkoxy, amino, Ci-C6 akylamino, C2-C]2 dialkylamino, -S-, -S-Ci-C6 alkyl, -S(O)-, - S(0)2-, heterocycloalkyl, heteroaryl, heteroaryloxy, sulfonamidyl, amido, urea, sulfonylurea, acyl, nitro, cyano, wherein each of Q-C6 alkyl, Ci-C6 heteroalkyl, C2-C6 alkenyl, C2-C6 alkynyl, C|-C6 haloalkyl, Ci-C6 haloalkoxy, hydroxyl, Ci-C6 alkoxy, C3-C7 cycloalkyloxy, C6-Cio aryl, C6-Cio aryloxy, C7-Ci6 arylalkoxy, amino, Ci-C6 akylamino, C2-Ci2 dialkylamino, -S-, -S-Ci-C alkyl, -S(O)-, -S(0)2-, heterocycloalkyl, heteroaryl, heteroaryloxy, sulfonamidyl, amido, urea, sulfonylurea, acyl, is optionally substituted with 1 -3 R6; R3 is Ci-C6 alkyl, Ci-C6 heteroalkyl, C3-C7 cycloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, C2- C6 hydroxyalkyl, or Ci-C6 alkoxy, each of which is optionally substituted with 1 -4 R7;
R4 is Cj-C6 alky], C|-C6 heteroalkyl, C2-C6 alkenyl or C2-C6 alkynyl, each of which is optionally substituted with 1 -4 R8;
R6, R7, and R8 are each independently C|-C6 alkyl, Ci-C6 heteroalkyl, halo, Ci-C6 haloalkyl, C|-C6 haloalkoxy, hydroxyl, Ci-C6 alkoxy, amino, C)-C6 alkylamino, C2-C)2 dialkylamino, cyano, nitro, amido, Ci-C alkylamido, C2-C12 dialkylamido, -S-, -S(0)2-,
-C(0)0-, -C(O)-, -C(0)0-C,-C6 alkyl, C3-C7 cycloalkyl, C6-C10 aryl, heterocycloalkyl, or heteroaryl, wherein each of Ci-C6 alkyl, Ci-C6 heteroalkyl, Ci-C6 haloalkyl, Ci-C6 haloalkoxy, hydroxyl, Ci-C6 alkoxy, amino, Ci-C6 alkylamino, C2-C)2 dialkylamino, amido, Ci-C6
alkylamido, C2-C]2 dialkylamido, -S-, -S(0)2-, -C(0)0-, -C(O)-, -C(0)0-Ci-C6 alkyl, C3-C7 cycloalkyl, C6-Cio aryl, heterocycloalkyl, or heteroaryl is optionally substituted with 1 -3 R9; and each R9 is independently Ci-C6 alkyl, Ci-C6 heteroalkyl, Ci-C6 haloalkyl, Cj-C6 haloalkoxy, heterocycloalkyl, C6-Ci0 aryl, heteroaryl, C -Cio cycloalkylalkyl, heterocycloalkyl- Ci-C6 alkyl, C7-Ci6 arylalkyl, heteroaryl-Ci-C6 alkyl, halo, hydroxyl, Cj-C6 alkoxy, C6-C]0 aryloxy, C7-C16 arylalkoxy, C2-C8 alkoxyalkoxyl, amino, Ci-C6 akylamino, C2-C]2 dialkylamino, Ci-C6 akyl-amino-Ci-C6 akyl, Ci-C6 akyl-amino-C2-Ci2 dialkyl, -S-, -S-Ci-Q alkyl, -S(0)2-Ct- C6 alkyl, sulfonamidyl, amido, urea, sulfonylurea, acyl, -C(O)-C6-C]0 aryl, -NHC(O)-C6-Ci0 aryl, -C(0)NH-C6-Cio aryl, -C(0)OH, -C(0)0-Ci-C6 alkyl, -C(0)-C!-C6 alkyl acyl, nitro, or cyano; to thereby treat the subject.
In some embodiments, the TRPC5 mediated disorder is selected from the group consisting of: a neuropsychiatric disorder, a neurodegenerative disorder, nephropathy, and seizure disorder.
In some embodiments, R1 is Ci-C6 alkyl.
In some embodiments, R1 is C]-C6 alkyl and R5 is independently C6-Cio aryl or heteroaryl.
In some embodiments, R1 is C]-C6 alkyl and R5 is independently phenyl optionally substituted with 1 -3 R9.
In some embodiments, R1 is C|-C6 alkyl and R5 is independently pyridyl, e.g., 2-pyridyl, 3-pyridyl, or 4 pyridyl, optionally substituted with 1 -3 R9.
In some embodiments, R2 is C6-Cio aryl, C6-Ci0 aryloxy or heteroaryl oxy. In some embodiments, R is C6-Cio aryloxy.
In some embodiments, R is Cj-C alkoxy.
In some embodiments, R is Ci-C6 alkyl.
In some embodiments, R is Ci-C6 alkyl and R is independently C6-Cio aryl or heteroaryl, optionally substituted with 1-3 R9.
In some embodiments, R2 is Ci-C6 alkyl and R6 is independently phenyl, optionally substituted with 1 -3 R9.
In some embodiments, R is or Ci-C6 akylamino.
In some embodiments, R is -S(O)- or -S(0)2-.
In some embodiments, R3 is Ci-C6 alkyl, C2-C6 hydroxyalkyl, or Ci-C alkoxy.
In some embodiments, R3 is C2-C6 hydroxyalkyl, e.g., hydroxypropyl.
In some embodiments, R is hydroxypropyl.
In some embodiments, R4 is Ci-C6 alkyl.
In some embodiments, R5 is independently C6-C10 aryl, heteroaryl, C3-C7 cycloalkyl, or hetero cycloalkyl.
In some embodiments, R5 is phenyl, pyridyl, thiazolyl, pyrimidinyl, or oxazolyl.
In some embodiments, R5 is phenyl.
In some embodiments, R5 is pyridyl.
In some embodiments, R5 is thiazolyl.
In some embodiments, R9 is independently Ci-C6 alkyl, Ci-C6 alkoxy, halo, Ci-C6 haloalkyl, C[-C6 haloalkoxy, or heterocycloalkyl.
In some embodiments, R2 is C6-Ci0 aryl or C6-Ci0 aryloxy, and R6 is independently Ci-C6 alkyl, Ci-C6 haloalkyl, Cj-C6 or haloalkoxy.
In some embodiments, R2 is C6-Cjo aryloxy and R6 is independently Ci-C6 haloalkyl or C i -C6 or haloalkoxy.
In some embodiments, R2 is C6-Cio aryloxy and R6 is -CF3 or -OCF3.
In another aspect, the invention rovides compounds of Formula T(a):
Figure imgf000010_0001
Formula 1(a) or a pharmaceutically acceptable salt thereof, wherein:
1 is Ci-C6 alkyl, C2-C6 alkenyl or C2-C6 alkynyl, each of which is optionally substituted with 1 -4 R5;
R2 is Ci-C6 alkyl, Ci-C6 heteroalkyl, C2-C6 alkenyl, C2-C6 alkynyl, CrC6 haloalkyi, halo, Ci-C6 haloalkoxy, hydroxyl, Ct-C6 alkoxy, C3-C7 cycloalkyloxy, C6-Ci0 aryl, C6-Cio aryloxy, C7- C i6 arylalkoxy, amino, Cj-C6 akylamino, C2-C12 dialkylamino, -S-, -S-C]-C6 alkyl, -S(O)-, - S(0)2-, heterocycloalkyl, heteroaryl, heteroaryloxy, sulfonamidyl, amido, urea, sulfonylurea, acyl, nitro, cyano, wherein each of Ci-C6 alkyl, Ci-C6 heteroalkyl, C2-C6 alkenyl, C2-C6 alkynyl, Ci-C6 haloalkyi, Ci-C6 haloalkoxy, hydroxyl, Ci-C6 alkoxy, C3-C7 cycloalkyloxy, C6-Ci0 aryl, C -Ci0 aryloxy, C7-Ci6 arylalkoxy, amino, Ci-C6 akylamino, C2-Cj2 dialkylamino, -S-, -S-Ci-C6 alkyl, -S(O)-, -S(0)2-, heterocycloalkyl, heteroaryl, heteroaryloxy, sulfonamidyl, amido, urea, sulfonylurea, acyl, is optionally substituted with 1 -3 R6;
R is C2-C6 hydroxyalkyl or Ci-C6 heteroalkyl;
R4 is Ci-C6 alkyl, Cj-C heteroalkyl, C2-C6 alkenyl or C2-C6 alkynyl, each of which is optionally substituted with 1-4 R8;
R5, R6, and R8 are each independently Ci-C6 alkyl, Ci-C6 heteroalkyl, halo, Q-Q haloalkyi, Ci-C6 haloalkoxy, hydroxyl, Ci-C6 alkoxy, amino, Ci-C6 alkylamino, C2-C]2 dialkylamino, cyano, nitro, amido, Ci-C alkylamido, C2-Ci2 dialkylamido, -S-, -S(0)2-,
-C(0)0-, -C(O)-, -C(0)0-Ci-C6 alkyl, C3-C7 cycloalkyl, C6-C10 aryl, heterocycloalkyl, or heteroaryl, each of Ci-C6 alkyl, C!-C6 heteroalkyl, Ci-C6 haloalkyi, Ci-C6 haloalkoxy, hydroxyl, Cj-C6 alkoxy, amino, Ci-C6 alkylamino, C2-C)2 dialkylamino, amido, C]-C6 alkylamido, C2-C12 dialkylamido, -S-, -S(0)2-, -C(0)0-, -C(O)-, -C(0)0-Ci-C6 alkyl, C3-C7 cycloalkyl, C6-C]0 aryl, heterocycloalkyl, or heteroaryl is optionally substituted with 1-3 R9; and
each R9 is independently Ci-C6 alkyl, Ci-C6 heteroalkyl, Ci-C6 haloalkyi, Ci-C6 haloalkoxy, heterocycloalkyl, C6-C)0 aryl, heteroaryl, C4-Cio cycloalkyl alkyl, heterocycloalkyl- Ci-C6 alkyl, C7-C16 arylalkyl, heteroaryl-Ci-C6 alkyl, halo, hydroxyl, Ci-C6 alkoxy, C6-C)0 aryloxy, C7-C(6 arylalkoxy, C2-C8 alkoxyalkoxyl, amino, Ci-C akylamino, C2-Cj2 dialkylamino, C]-C6 akyl-amino-C]-C6 akyl, C C6 akyl-amino-C2-Ci2 dialkyl, -S-, -S-C,-C6 alkyl, -S(0)2-C C6 alkyl, sulfonamidyl, amido, urea, sulfonylurea, acyl, -C(O)-C -Ci0 aryl, -NHC(O)-C6-C)0 aryl, -C(0)NH-C6-C,o aryl, -C(0)OH, -C(0)0-Ci -C6 alkyl, -C(0)-Ci-C6 alkyl acyl, nitro, or cyano.
In some embodiments, R1 is Ci-C6 alkyl and R5 is phenyl, pyridyl, thiazolyl, pyrimidinyl, or oxazolyl, e.g., phenyl, pyridiyl, or thiazolyl.
In some embodiments, R1 is C)-C6 alkyl and R5 is independently phenyl optionally substituted with 1 -3 R9.
In some embodiments, R1 is Cj-C6 alkyl and R5 is independently pyridyl, e.g., 2-pyridyl, 3-pyridyl, or 4 pyridyl, optionally substituted with 1-3 R9.
In some embodiments, R2 is C6-Cio aryl or C6-Cio aryloxy, and R6 is independently Ci-C6 alkyl, Ci-C6 haloalkyl, C]-C6 or haloalkoxy.
In some embodiments, R2 is Ci-C6 alkyl and R6 is independently C6-C|0 aryl or heteroaryl, optionally substituted with 1-3 R9.
In some embodiments, R2 is Cj-C6 alkyl and R6 is independently phenyl, optionally substituted with 1-3 R9.
In some embodiments, R3 is C2-C6 hydroxyalkyl, e.g., hydroxypropyl.
In another aspect, the inve ormula II:
Figure imgf000012_0001
Formula II
or a pharmaceutically acceptable salt thereof, wherein:
Ring A is phenyl, pyridyl, thiazolyl, pyrimidinyl, or oxazolyl;
R2 is Ci-C6 alkyl, Ci-C6 heteroalkyl, C2-C6 alkenyl, C2-C6 alkynyl, CrC6 haloalkyl, halo, Ci-C6 haloalkoxy, hydroxyl, Ci-C6 alkoxy, C3-C7 cycloalkyloxy, C6-Cio aryl,. C6-Cio aryloxy, C7- Ci6 arylalkoxy, amino, Ci-C6 akylamino, C2-Ci2 dialkylamino, -S-, -S-C|-C6 alkyl, -S(O)-, - S(0)2-, heterocycloalkyl, heteroaryl, heteroaryloxy, sulfonamidyl, amido, urea, sulfonylurea, acyl, nitro, cyano, wherein each of Ci-C6 alkyl, Ci-C6 heteroalkyl, C2-C6 alkenyl, C2-C6 alkynyl, Ci-C6 haloalkyl, Ci-C6 haloalkoxy, hydroxyl, Ci-C6 alkoxy, C3-C7 cycloalkyloxy, C6-Cjo aryl, C6-C|0 aryloxy, C7-Ci6 arylalkoxy, amino, Ci-C6 akylamino, C2-Cj2 dialkylamino, -S-, -S-Ci-C6 alkyl, -S(O)-, -S(0)2-, heterocycloalkyl, heteroaryl, heteroaryloxy, sulfonamidyl, amido, urea, sulfonylurea, acyl, is optionally substituted with 1 -3 R6;
R3 is CrC6 alkyl, Ci-C6 heteroalkyl, C2-C6 alkenyl, C2-C6 alkynyl, C2-C6 hydroxyalkyl, or Ci-C6 alkoxy, each of which is optionally substituted with 1 -4 R ;
R4 is C]-C6 alkyl, Ci-C6 heteroalkyl, C2-C alkenyl or C2-C6 alkynyl, each of which is optionally substituted with 1-4 R8;
7 ft
R , R , and R are each independently Ci-C alkyl, Ci-C6 heteroalkyl, halo, Ci-C6 haloalkyl, Ci-C6 haloalkoxy, hydroxyl, C[-C6 alkoxy, amino, Ci-C6 alkylamino, C2-C|2 dialkylamino, cyano, nitro, amido, Ci-C6 alkylamido, C2-Ci2 dialkylamido, -S-, -S(0)2-,
-C(0)0-, -C(O)-, -C(0)0-C C6 alkyl, C3-C7 cycloalkyl, C6-Ci0 aryl, heterocycloalkyl, or heteroaryl, wherein each of Ci-C6 alkyl, C|-C6 heteroalkyl, Ci-C6 haloalkyl, Ci-C6 haloalkoxy, hydroxyl, Ci-C6 alkoxy, amino, C]-C6 alkylamino, C2-C)2 dialkylamino, amido, C]-C
alkylamido, C2-Ci2 dialkylamido, -S-, -S(0)2-, -C(0)0-, -C(O)-, -C(0)0-Ci-C6 alkyl, C3-C7 cycloalkyl, C6-C10 aryl, heterocycloalkyl, or heteroaryl is optionally substituted with 1-3 R9; each R9 is independently Ci-C6 alkyl, Ci-C6 heteroalkyl, Ci-C6 haloalkyl, Ci-C6 haloalkoxy, heterocycloalkyl, C6-Ci0 aryl, heteroaryl, C4-C10 cycloalkylalkyl, heterocycloalkyl- Ci-C6 alkyl, C7-Ci6 arylalky], heteroaryl-Ci-C alkyl, halo, hydroxyl, Ci-C6 alkoxy, C6-Cio aryloxy, C7-Ci6 arylalkoxy, C2-C8 alkoxyalkoxyl, amino, Ci-C6 akylamino, C2-Ci2 dialkylamino, Ci-C6 akyl-amino-Ci-C6 akyl, C|-C6 akyl-amino-C2-C12 dialkyl, -S-, -S-Ci-C6 alkyl, -S(0)2-C Ce alkyl, sulfonamidyl, amido, urea, sulfonylurea, acyl, -C(O)-C6-Ci0 aryl, -NHC(0)-C6-Cio aryl, -C(0)NH-C6-Cio aryl, -C(0)OH, -C(0)0-Ci-C6 alkyl, -C(0)-CrC6 alkyl acyl, nitro, or cyano; each Ra is Ci-C6 alkyl, Ci-C6 haloalkyl, halo;
n is 1 or 2; and
m is 1 , 2, or 3.
In some embodiments, Ring A is phenyl or thiazolyl.
In some embodiments, Ring A is pyridyl, e.g. 2-pyridyl, 3-pyridyl, or 4-pyridyl.
In some embodiments, R2 is CpC6 alkyl and R6 is independently C6-C]0 aryl or heteroaryl, optionally substituted with 1 -3 R9.
In some embodiments, R2 is Ci-C6 alkyl and R6 is independently phenyl, optionally substituted with 1 -3 R9. In some embodiments, R3 is hydroxypropyl.
In another aspect, the invention rovides compound of Formula 111:
Figure imgf000014_0001
Formula III
or a pharmaceutically acceptable salt thereof, wherein:
R is C|-C6 alkoxy or C6-Ci0 aryloxy substituted with 1-3 R ;
R3 is Ci-C6 heteroalkyl or C2-C6 hydroxyalkyl;
R4 is Ci-C6 alkyl;
R6 is independently C]-C6 alkyl, halo, Ci-C6 haloalkyl, C)-C6 haloalkoxy, or Ci-C6 alkoxy;
each Ra is Ci-C6 alkyl, d-C6 haloalkyl, halo;
n is 1 or 2; and
m is 1 , 2, or 3.
In some embodiments, R3 is hydroxypropyl.
In some embodiments, Ra is independently chloro, fluoro, or methyl.
In another aspect, the invention provides methods of treating a TRPC5 mediated disorder in a subject, the method comprising administering to the subject a compound or composition of any one of claims 23 to 33, to thereby treat the subject.
In some embodiments, the TRPC5 mediated disorder is selected from the group consisting of: a neuropsychiatric disorder, a neurodegenerative disorder, nephropathy, and seizure disorder.
In certain embodiments, exemplary compounds of the invention include the compounds described in Table A and in the Examples. Table A
Figure imgf000015_0001
Figure imgf000016_0001
Figure imgf000017_0001
Figure imgf000018_0001
16
Figure imgf000019_0001
Figure imgf000020_0001
Figure imgf000021_0001
Figure imgf000022_0001
Figure imgf000023_0001
Figure imgf000024_0001
Figure imgf000025_0001
Figure imgf000026_0001
Figure imgf000027_0001
Figure imgf000028_0001
Figure imgf000029_0001
Figure imgf000030_0001
Figure imgf000031_0001
Figure imgf000032_0001
Figure imgf000033_0001
Figure imgf000034_0001
Figure imgf000035_0001
Figure imgf000036_0001
Figure imgf000037_0001
Figure imgf000038_0001
Figure imgf000039_0001
Figure imgf000040_0001
Figure imgf000041_0001
Figure imgf000042_0001
Figure imgf000043_0001
Figure imgf000044_0001
Figure imgf000045_0001
Figure imgf000046_0001
Figure imgf000047_0001
Figure imgf000048_0001
Figure imgf000049_0001
Figure imgf000050_0001
Figure imgf000051_0001
Figure imgf000052_0001
Figure imgf000053_0001
Figure imgf000054_0001
Figure imgf000055_0001
Figure imgf000056_0001
Figure imgf000057_0001
Figure imgf000058_0001
Figure imgf000059_0001
Figure imgf000060_0001
Figure imgf000061_0001
Figure imgf000062_0001
60
Figure imgf000063_0001
Figure imgf000064_0001
Figure imgf000065_0001
Figure imgf000066_0001
Figure imgf000067_0001
Figure imgf000068_0001
66
Figure imgf000069_0001
Figure imgf000070_0001
Figure imgf000071_0001
Figure imgf000072_0001
Figure imgf000073_0001
Figure imgf000074_0001
Figure imgf000075_0001
Figure imgf000076_0001
Figure imgf000077_0001
Figure imgf000078_0001
Figure imgf000079_0001
326
327
1
328
r
329
1
330
1
Figure imgf000081_0001
Figure imgf000082_0001
Figure imgf000083_0001
Figure imgf000084_0001
Figure imgf000085_0001
Figure imgf000086_0001
Figure imgf000087_0001
Figure imgf000088_0001
Figure imgf000089_0001
Figure imgf000090_0001
Figure imgf000091_0001
Figure imgf000092_0001
Figure imgf000093_0001
Figure imgf000094_0001
Figure imgf000095_0001
Figure imgf000096_0001
Figure imgf000097_0001
416
417
418
419
420
Figure imgf000099_0001
Figure imgf000100_0001
Figure imgf000101_0001
Figure imgf000102_0001
Figure imgf000103_0001
Figure imgf000104_0001
Figure imgf000105_0001
Figure imgf000106_0001
Figure imgf000107_0001
Figure imgf000108_0001
Figure imgf000109_0001
Figure imgf000110_0001
Figure imgf000111_0001
Figure imgf000112_0001
Figure imgf000113_0001
Figure imgf000114_0001
Figure imgf000115_0001
Figure imgf000116_0001
Figure imgf000117_0001
Figure imgf000118_0001
Figure imgf000119_0001
Figure imgf000120_0001
Figure imgf000121_0001
Figure imgf000122_0001
Figure imgf000123_0001
Figure imgf000124_0001
Figure imgf000125_0001
Figure imgf000126_0001
Figure imgf000127_0001
Figure imgf000128_0001
Figure imgf000129_0001
Figure imgf000130_0001
Figure imgf000131_0001
Figure imgf000132_0001
Figure imgf000133_0001
Figure imgf000134_0001
Figure imgf000135_0001
Figure imgf000136_0001
Figure imgf000137_0001
Figure imgf000138_0001
Figure imgf000139_0001
Figure imgf000140_0001
Figure imgf000141_0001
Figure imgf000142_0001
Figure imgf000143_0001
Figure imgf000144_0001
Definitions
At various places in the present specification, substituents of compounds of the invention are disclosed in groups or in ranges. It is specifically intended that the invention include each and every individual subcombination of the members of such groups and ranges. For example, the term "Ci-6 alkyl" is specifically intended to individually disclose methyl, ethyl, C3 alkyl, C4 alkyl, C5 alkyl, and C6 alkyl.
For compounds of the invention in which a variable appears more than once, each variable can be a different moiety selected from the Markush group defining, the variable. For example, where a structure is described having two R groups that are simultaneously present on the same compound; the two R groups can represent different moieties selected from the Markush group defined for R. It is further appreciated that certain features of the invention, which are, for clarity, described in the context of separate embodiments, can also be provided in combination in a single embodiment. Conversely, various features of the invention which are, for brevity, described in the context of a single embodiment, can also be provided separately or in any suitable subcombination.
As used herein, "acyl" refers to the group (C[-C6 alkyl)-C(O)-.
As used herein, "alkyl," by itself or as part of another substituent, means, unless otherwise stated, a straight or branched chain, and can have a number of carbon atoms optionally designated (i.e. , Ci-C6 means one to six carbons). Examples of saturated hydrocarbon groups include, but are not limited to, groups such as methyl, ethyl, n-propyl, isopropyl, n-butyl, t-butyl, isobutyl, sec-butyl, n-pentyl, isopentyl, homologs and isomers of, for example, n-pentyl, n-hexyl, and the like.
As used herein, "alkenyl" can be a straight or branched hydrocarbon chain, containing at least one double bond, and having from two to six carbon atoms (i.e. C2-C alkenyl). Examples of alkenyl groups, include, but are not limited to, groups such as ethenyl (i.e., vinyl), prop-l-enyl (i.e., allyl), but-l -enyl, pent-l-enyl, penta-l ,4-dienyl, and the like.
As used herein, "alkoxy" can be a straight chain or branched alkoxy group having from one to six carbon atoms (i.e., Ci-C6 alkoxy). Examples of alkoxy groups, include, but are not limited to, groups such as methoxy, ethoxy, propyloxy, isopropyloxy, butyloxy, isobutyloxy, tert-butyloxy, pentyloxy, or hexyloxy, and the like.
As used herein, "alkynyl" can be a straight or branched hydrocarbon chain, containing at least one triple bond, having from two to six carbon atoms (i.e. C2-C6 alkynyl). Examples of alkynyl groups, include, but are not limited to, groups such as ethynyl, propynyl, butynyl, pentynyl, hexynyl, and the like.
As used herein, "amide" or "amido" refers to a chemical moiety with the formula - C(0)NRa- or -NRaC(0)- wherein Ra is H or Ci-C6 alkyl.
As used herein, "amino" or "amine" refers to a -NH2 radical group.
As used herein, "alkylamino" refers to a group of formula -NH(alkyl), wherein the alkyl group each has 1 to 6 carbons.
As used herein, the term "dialkylamino" refers to a group of formula -N(alkyl)2, wherein the two alkyl groups each independently has, 1 to 6 carbons. As used herein, "aryl" refers to a polyunsaturated, aromatic, hydrocarbon moiety which can be a single ring or multiple rings (e.g., 1 to 2 rings) which are fused together or linked covalently, having from six to twelve carbon atoms (i.e. C6-C]2 aryl). Non-limiting examples of aryl groups include phenyl, 1 -naphthyl, 2-naphthyl, and 4-biphenyl.
As used herein, "arylalkyl" refers to an (aryl)alkyl— radical wherein aryl and alkyl moieties are as disclosed herein.
As used herein, "aryloxy" refers to -O-(aryl), wherein the heteroaryl moiety is as defined herein.
As used herein, "arylalkoxy" refers to -O-(arylalkyl), wherein the heteroaryl moiety is as defined herein.
As used herein, "carboxyl" refers to a -(C=0)OH radical.
As used herein, "cyano" refers to a -CN radical.
As used herein, "cycloalkyl" refers to a monocyclic or polycyclic radical that contains only carbon and hydrogen, and may be saturated, or partially unsaturated. Cycloalkyl groups include groups having from 3 to 10 ring atoms (i.e. C3-C]0 cycloalkyl). Examples of cycloalkyl groups include, but are not limited to, groups such as cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cycloseptyl, cyclooctyl, cyclononyl, cyclodecyl, norbornyl, and the like.
As used herein, "C3-C7 cycloalkyloxy" refers to -0-(C3-C7 cycloalkyl), wherein the
C3-C7cycloalkyl moiety is as defined herein.
As used herein, "halo" or "halogen," independently or as part of another substituent, mean, unless otherwise stated, a fluorine, chlorine, bromine, or iodine atom. The term "halide" by itself or as part of another substituent, refers to a fluoride, chloride, bromide, or iodide atom.
As used herein, "haloalkyl" and "haloalkoxy" can include alkyl and alkoxy structures that are substituted with one or more halo groups or with combinations thereof. For example, the terms "fluoroalkyl" and "fluoroalkoxy" include haloalkyl and haloalkoxy groups, respectively, in which the halo is fluorine.
As used herein, "heteroalkyl" can include an optionally substituted alkyl, which has one or more skeletal chain atoms selected from an atom other than carbon, e.g., oxygen, nitrogen, sulfur, phosphorus or combinations thereof. A numerical range may be given, e.g. C]-C6 heteroalkyl which refers to the number of carbons in the chain, which in this example includes 1 to 6 carbon atoms. For example, a -CH2OCH2CH3 radical is referred to as a "C3" heteroalkyl. Connection to the rest of the molecule may be through either a heteroatom or a carbon in the heteroalkyl chain.
As used herein, "heteroaryl" refers to a 5- to 14-membered aromatic radical (e.g., C2-C 13 heteroaryl) that includes one or more ring heteroatoms selected from nitrogen, oxygen and sulfur, and which may be a monocyclic or bicyclic ring system. Bivalent radicals derived from univalent heteroaryl radicals whose names end in "-yl" by removal of one hydrogen atom from the atom with the free valence are named by adding "-idene" to the name of the corresponding univalent radical, e.g., a pyridyl group with two points of attachment is a pyridylidene. An N- containing "heteroaromatic" or "heteroaryl" moiety refers to an aromatic group in which at least one of the skeletal atoms of the ring is a nitrogen atom. The polycyclic heteroaryl group may be fused or non-fused. The heteroatom(s) in the heteroaryl radical is optionally oxidized. One or more nitrogen atoms, if present, are optionally quatemized. The heteroaryl is attached to the rest of the molecule through any atom of the ring(s). Examples of heteroaryl groups include without limitation, pyridyl, pyrimidinyl, pyrazinyl, pyridazinyl, triazinyl, furyl (furanyl), quinolyl, isoquinolyl, thienyl, imidazolyl, thiazolyl, indolyl, pyrryl, oxazolyl, benzofuryl, benzothienyl, benzthiazolyl, isoxazolyl, pyrazolyl, triazolyl, tetrazolyl, indazolyl, 1 ,2,4-thiadiazolyl, isothiazolyl, benzothienyl, purinyl, carbazolyl, benzimidazolyl, indolinyl, and the like.
As used herein, "heteraryloxy" refers to -O-(heteroaryl), wherein the heteroaryl moiety is as defined herein.
As used herein, "heterocycloalkyl" can be a stable 3- to 18-membered non-aromatic ring radical that comprises two to twelve carbon atoms and from one to six heteroatoms selected from nitrogen, oxygen and sulfur. Examples of heterocycloalkyl groups include, but are not limited to, groups such as dioxolanyl, thienyl [l ,3]dithianyl, decahydroisoquinolyl, imidazolinyl, imidazolidinyl, isothiazolidinyl, isoxazolidinyl, morpholinyl, octahydroindolyl,
octahydroisoindolyl, 2-oxopiperazinyl, 2-oxopiperidinyl, 2-oxopyrrolidinyl, oxazolidinyl, piperidinyl, piperazinyl, 4-piperidonyl, pyrrolidinyl, pyrazolidinyl, quinuclidinyl, thiazolidinyl, tetrahydrofuryl, trithianyl, tetrahydropyranyl, thiomorpholinyl, thiamorpholinyl,
1 -oxo-thiomorpholinyl, 1,1 -dioxo-thiomorpholinyl, and the like.
As used herein, "hydroxy" or "hydroxyl" refers to -OH. As used herein, "hydroxyalkyl" refers to an alkyl group having 1 to 6 carbon atoms, which is substituted with a hydroxyl group, e.g., hydroxypropyl.
As used herein, "cyano" refers to -CN.
■ As used herein, "nitro" refers to -N02.
As used herein, "urea" refers to -NRa-C(0)-NRa 2 or -NRa-C(0)NRa-, wherein Ra is H or
CrC6 alkyl.
As used herein, "sulfonylurea" refers to -S(0)2-NRa-C(0)-NRa- or
-NRa-C(0)-NRa-S02-, wherein Ra is H or d-C6 alkyl.
As used herein, "sulfonamidyl" refers to -S(0)2-NRa- or -NRa-S(0)2-, wherein Ra is H or C,-C6 alkyl.
The terms "antagonist" and "inhibitor" are used interchangeably to refer to an agent that decreases or suppresses a biological activity, such as to repress an activity of an ion channel, such as TRPC5. TRPC5 ion channels as described herein include homomultimeric and heteromultimeric structures (e.g., homomultimeric TRPC5 and heteromeric TRPC5-TRPC1 or TRPC5-TRPC4). TRPC5 antagonists include inhibitors having any combination of the structural and/or functional properties disclosed herein.
An "effective amount" of, e.g., a TRPC5 antagonist, with respect to the subject methods of inhibition or treatment, refers to an amount of the antagonist in a preparation which, when applied as part of a desired dosage regimen brings about a desired clinical or functional result. Without being bound by theory, an effective amount of a TRPC5 antagonist for use in the methods of the present invention includes an amount of a TRPC5 antagonist effective to decrease one or more in vitro or in vivo function of a TRPC5 channel. Exemplary functions include, but are not limited to, membrane polarization (e.g., an antagonist may promote hyperpolarization of a cell), ion flux, ion concentration in a cell, outward current, and inward current. Compounds that antagonize TRPC5 function include compounds that antagonize an in vitro or in vivo functional activity of TRPC5. When a particular functional activity is only readily observable in an in vitro assay, the ability of a compound to inhibit TRPC5 function in that in vitro assay serves as a reasonable proxy for the activity of that compound. In certain embodiments, an effective amount is an amount sufficient to inhibit a TRPC5-mediated current and/or the amount sufficient to inhibit TRPC5 mediated ion flux. The TRPC5 anagonists for use in the methods of the present invention may be characterized according to their activity, or lack of activity, against one or more other ion channels. When other ion channels are referred to, inhibition of a function of such other ion channels is defined similarly. For example, inhibition of an ion channel or an activity of an ion channel means the antagonist inhibits one or more functional activities of the other ion channel. Such functions include the current mediated by the particular ion channel, ion flux, or membrane polarization.
The term "preventing" is art-recognized, and when used in relation to a condition, such as a local recurrence, a disease such as cancer, a syndrome complex such as heart failure or any other medical condition, is well understood in the art, and includes administration of a composition which reduces the frequency of, or delays the onset of, symptoms of a medical condition in a subject relative to a subject which does not receive the composition. Thus, prevention of cancer includes, for example, reducing the number of detectable cancerous growths in a population of patients receiving a prophylactic treatment relative to an untreated control population, and/or delaying the appearance of detectable cancerous growths in a treated population versus an untreated control population, e.g., by a statistically and/or clinically significant amount. Prevention of an infection includes, for example, reducing the number of diagnoses of the infection in a treated population versus an untreated control population, and/or delaying the onset of symptoms of the infection in a treated population versus an untreated control population. Prevention of pain includes, for example, reducing the magnitude of, or alternatively delaying, pain sensations experienced by subjects in a treated population versus an untreated control population.
The term "prodrug" is intended to encompass compounds that, under physiological conditions, are converted into the therapeutically active agents of the present invention. A common method for making a prodrug is to include selected moieties that are hydrolyzed under physiological conditions to reveal the desired molecule. In other embodiments, the prodrug is converted by an enzymatic activity of the host animal.
The term "small molecule" refers to a compound having a molecular weight less than about 2500 amu, preferably less than about 2000 amu, even more preferably less than about 1500 amu, still more preferably less than about 1000 amu, or most preferably less than about 750 amu. The terms "TRPC5", "TRPC5 protein", and "TRPC5 channel" are used interchangeably throughout the application. Unless expressly stated, the term TRPC5 includes homomultimeric structures (e.g., homomultimeric TRPC5) and heteromultimeric structures (e.g.,
heteromultimeric TRPC5-TRPC1 ).
The term "oxidative metabolite" is intended to encompass compounds that are produced by metabolism of the parent compound under normal physiological conditions. Specifically, an oxidative metabolite is formed by oxidation of the parent compound during metabolism. For example, a thioether group may be oxidized to the corresponding sulfoxide or sulfone.
The term "solvate" as used herein, refers to a compound formed by solvation (e.g., a compound formed by the combination of solvent molecules with molecules or ions of the solute).
The term "hydrate" as used herein, refers to a compound formed by the union of water with the parent compound.
The term "treating" includes prophylactic and/or therapeutic treatments. The term
"prophylactic or therapeutic" treatment is art-recognized and includes administration to the host of one or more of the subject compositions. If it is administered prior to clinical manifestation of the unwanted condition (e.g., disease or other unwanted state of the host animal) then the treatment is prophylactic, (i.e., it protects the host against developing the unwanted condition), whereas if it is administered after manifestation of the unwanted condition, the treatment is therapeutic, (i.e., it is intended to diminish, ameliorate, or stabilize the existing unwanted condition or side effects thereof).
The terms "compound" and "agent" are used interchangeably to refer to the
inhibitors/antagonists of the invention. In certain embodiments, the compounds are small organic or inorganic molecules, e.g., with molecular weights less than 7500 amu, preferably less than 5000 amu, and even more preferably less than 2000, 1500, 1000, or 600 amu. Such compounds can bind to and inhibit a function of TRPC5. In certain other embodiments, the compounds are nucleic acids, for example, TRPC5 antisense oligonucleotides or TRPC5 RNAi constructs. Such compounds can inhibit the expression of TRPC5, thereby inhibiting the activity of TRPC5. Other exemplary compounds that may act as inhibitors include ribozymes and peptide fragments.
Contemplated equivalents of the compounds described above include compounds which otherwise correspond .thereto, and which have the same general properties thereof (e.g., the ability to antagonize TRPC5 activity), wherein one or more simple variations of substituents are made which do not adversely affect the efficacy of the compound. In general, the compound of the present invention may be prepared by the methods illustrated in the general reaction schemes as, for example, described below, or by modifications thereof, using readily available starting materials, reagents and conventional synthesis procedures. In these reactions, it is also possible to make use of variants which are in themselves known, but are not mentioned here.
For purposes of this invention, the chemical elements are identified in accordance with the Periodic Table of the Elements, CAS version, Handbook of Chemistry and Physics, 67th Ed., 1986-87, inside cover. Also for purposes of this invention, the term "hydrocarbon" is
contemplated to include all permissible compounds having at least one hydrogen and one carbon atom. In a broad aspect, the permissible hydrocarbons include acyclic and cyclic, branched and unbranched, carbocyclic and heterocyclic, aromatic and nonaromatic organic compounds which can be substituted or unsubstituted.
The compounds described herein can be asymmetric (e.g., having one or more stereocenters). All stereoisomers, such as enantiomers and diastereomers, are intended unless otherwise indicated. Compounds of the present invention that contain asymmetrically substituted carbon atoms can be isolated in optically active or racemic forms. Methods on how to prepare optically active forms from optically active starting materials are known in the art, such as by resolution of racemic mixtures or by stereoselective synthesis. Many geometric isomers of olefins, C=N double bonds, and the like can also be present in the compounds described herein, and all such stable isomers are contemplated in the present invention. Cis and trans geometric isomers of the compounds of the present invention are described and may be isolated as a mixture of isomers or as separated isomeric forms.
Resolution of racemic mixtures of compounds can be carried out by any of numerous methods known in the art. An example method includes fractional recrystallizaion using a "chiral resolving acid" which is an optically active, salt-forming organic acid. Suitable resolving agents for fractional recrystallization methods are, for example, optically active acids, such as the D and L forms of tartaric acid, diacetyltartaric acid, dibenzoyltartaric acid, mandelic acid, malic acid, lactic acid or the various optically active camphorsulfonic acids such as β-camphorsulfonic acid. Other resolving agents suitable for fractional crystallization methods include
stereoisomerically pure forms of ct-methylbenzylamine (e.g., 5* and R forms, or diastereomerically pure forms), 2-phenylglycinol, norephedrine, ephedrine, N-methylephedrine, cyclohexylethylamine, 1 ,2-diaminocyclohexane, and the like.
Resolution of racemic mixtures can also be carried out by elution on a column packed with an optically active resolving agent (e.g., dinitrobenzoylphenyl glycine). Suitable elution solvent composition can be determined by one skilled in the art. Compounds of the invention also include tautomeric forms, such as keto-enol tautomers.
Compounds of the invention can also include all isotopes of atoms occurring in the intermediates or final compounds. For example, the compound of the invention may be radiolabeled with radioactive isotopes, such as for example tritium (3H) or carbon- 4 (14C). All isotopic variations, whether radioactive or not, are intended to be encompassed within the scope of the present invention.
Compounds of the invention can exist in unsolvated forms as well as solvated forms, including hydrated forms. In general, the solvated forms are equivalent to unsolvated forms and are encompassed within the scope of the present invention. A compound of formula (I) may exist in multiple crystalline or amorphous forms. In general, all physical forms are equivalent for the uses contemplated by the present invention and are intended to be within the scope of the present invention.
The term "pharmaceutically acceptable salts" includes salts of a compound of the invention which are prepared with relatively nontoxic acids or bases. Base addition salts can be obtained by contacting the neutral form of a compound of the invention with a sufficient amount of the desired base, either neat or in a suitable inert solvent. Examples of pharmaceutically acceptable base addition salts include sodium, potassium, calcium, ammonium, organic amino, or magnesium salt, or a similar salt. Acid addition salts can be obtained by contacting the neutral form of compound of the invention with a sufficient amount of the desired acid, either neat or in a suitable inert solvent. Examples of pharmaceutically acceptable acid addition salts include those derived from inorganic acids like hydrochloric, hydrobromic, nitric, carbonic,
monohydrogencarbonic, phosphoric, monohydrogenphosphoric, dihydrogenphosphoric, sulfuric, monohydrogensulfuric, hydriodic, or phosphorous acids and the like, as well as the salts derived from relatively nontoxic organic acids like acetic, trifluoroacetic, propionic, isobutyric, maleic, malonic, benzoic, succinic, suberic, fumaric, lactic, mandelic, phthalic, benzensulfonic, p- tolylsulfonic, citric, tartaric, methanesulfonic, and the like. Also included are the salts of amino acids such as arginate and the like, and salts of organic acids like glucuronic or galactunoric acids and the like (see, for example, Berge et al., "Pharmaceutical Salts", Journal of
Pharmaceutical Science, 1977, 66, 1-19).
The neutral forms of compound of the invention is preferably regenerated by contacting the salt with a base or acid and isolating the parent compound in the conventional manner. The parent form of the compound differs from the various salt forms in certain physical properties, such as solubility in polar solvents, but otherwise the salts are equivalent to the parent form of the compound for the purposes of the present invention.
The term "low enough pyrogen activity", with reference to a pharmaceutical preparation, refers to a preparation that does not contain a pyrogen in an amount that would lead to an adverse effect (e.g., irritation, fever, inflammation, diarrhea, respiratory distress, endotoxic shock, etc.) in a subject to which the preparation has been administered. For example, the term is meant to encompass preparations that are free of, or substantially free of, an endotoxin such as, for example, a lipopolysaccharide (LPS).
Diseases, Disorders, or Conditions Related to TRPC5 Function
In certain embodiments, the invention provides methods and compositions for antagonizing a function of a TRPC5 channel in vitro or in vivo. Exemplary functions include, but are not limited to, TRPC5-mediated current. In certain embodiments, the invention provides methods for treating a disease or disorder or condition by administering compound of the invention. In other embodiments, the compound of formula (1) selectively inhibits the expression level and/or activity of a T PC5 protein. In other words, in certain embodiment, the compound of the invention inhibits the activity of a TRPC5 protein preferentially in comparison to the activity of one or more other ion channels.
Treatment of Anxiety and Fear-Related Disorders
In certain embodiments, the compound of the invention can be used for preventing or treating anxiety and fear-related disorders (see, e.g., Riccio et al. (2009) Cell 137:761-72).
Examples of such disorders include post-traumatic stress disorder, panic disorder, agoraphobia, social phobias, generalized anxiety disorder, panic disorder, social anxiety disorder, obsessive- compulsive disorder, and separation anxiety. Memory, Motion and Mood Disorders
A compound of the invention is also useful for the treatment of Parkinson's disease, epilepsy, memory disorders, stroke, seizure, and mood disorders. Mood disorders include depression (e.g., major depression, psychiatric depression, dysthymia, and postpartum depression) and bipolar disorder (e.g., bipolar I, bipolar II, and cyclothymia). Memory disorders are conditions associated with any memory loss and may result from Alzheimer's disease, amnesia, aphasia, atherosclerosis, brain injury or disorder, brain tumor, chronic fatigue syndrome, Creutzfedt- Jacob disease, dissociative amnesia, depression, fuge amnesia,
Huntington's disease, learning disorders, sleeping disorders, multiple personality disorder, pain, post-traumatic stress disorder, schizophrenia, sports injuries, stroke, and Wernicke-Korsakoff syndrome.
Treatment of Pain, Sensitivity to Pain and Touch, or Pain-Related Diseases or Disorders
In certain embodiments, a compound of the invention is used to treat or ameliorate pain. Exemplary classes of pain that can be treated using a compound of formula (I)include, but are not limited to nociceptive pain, inflammatory pain, and neuropathic pain. The pain can be chronic or acute.
A compound of the invention may be particularly useful in the treatment of pain associated with cancer, osteoarthritis, rheumatoid arthritis, post-herpetic neuralgia, burns, and other indications detailed above. To further illustrate, additional exemplary indications for which a compound of the invention can be used include oral pain, pelvic pain, Fabry's disease, complex regional pain syndrome, pancreatitis, and fibromyalgia syndrome.
A compound of the invention may also be used in connection with prevention or treatment of sensitivity to pain and touch. Pain or sensitivity to pain and touch may be indicated in a variety of diseases, disorders or conditions, including, but not limited to, diabetic neuropathy, breast pain, psoriasis, eczema, dermatitis, burn, post-herpetic neuralgia (shingles), nociceptive pain, peripheral neuropathic and central neuropathic pain, chronic pain, cancer and tumor pain, spinal cord injury, crush injury and trauma induced pain, migraine, cerebrovascular and vascular pain, sickle cell disease pain, rheumatoid arthritis pain, musculoskeletal pain including treating signs and symptoms of osteoarthritis and rheumatoid arthritis, orofacial and facial pain, including dental, temperomandibular disorder, and cancer related, lower back or pelvic pain, surgical incision related pain, inflammatory and non-inflammatory pain, visceral pain, psychogenic pain and soft tissue inflammatory pain, fibromyalgia-related pain, and reflex sympathetic dystrophy, and pain resulting from kidney stones or urinary tract infection.
The foregoing are merely exemplary of diseases and conditions that cause or lead to inflammation, lesions, ulcers, or other sources of oral pain. In other embodiments, the oral pain is due to an injury to the mouth, jaw, lips, gums, or teeth. In other embodiments, the oral pain is due to oral surgery, for example, surgery for cancer, tooth extraction, or jaw remodeling. Other conditions that may lead to oral ulcers, and thus oral pain, include, but are not limited to chickpox, herpes zoster, infectious mononucleosis, syphilis, tuberculosis, acute necrotizing gingivitis, and burning mouth syndrome.
Fibromyalgia (FMS; fibromyalgia syndrome) is a widespread musculoskeletal pain and fatigue disorder. Fibromyalgia is characterized by pain in the muscles, ligaments, and tendons. The condition affects more women than men, and occurs in people of all ages. Overall, FMS is estimated to afflict 3-6% of the population. Patients have described the pain associated with fibromylagia as deep muscular aching, throbbing, shooting, and stabbing. The pain sometimes includes an intense burning sensation. The pain and stiffness are often worse in the morning or after repetitive use of a particular muscle group.
Additionally, varying levels of fatigue ranging from mild to incapacitating are often associated with fibromylagia. Other symptoms of fibromylagia include gastrointestinal symptoms. Irritable bowel syndrome and IBS-like symptoms such as constipation, diarrhea, frequent abdominal pain, abdominal gas, and nausea occur in roughly 40 to 70% of FMS patients. Acid reflux or gastroesophogeal reflux disease (GERD) occurs at a similar frequency.
Complex Regional Pain Syndrome (CRPS; also known as chronic regional pain syndrome) is a chronic pain condition. CRPS was formerly known as reflex sympathetic dystrophy (RSD). CRPS is a chronic, painful, and progressive neurological condition that affects skin, muscles, joints, and bones. The syndrome usually develops in an injured limb, such as a broken leg or following surgery. However, many cases involve only a minor injury, such as a sprain, and sometimes no precipitating injurious event can be identified. CRPS involves continuous, intense pain that is disproportionate to the severity of the injury. The pain worsens, rather than improves, over time.
Although CRPS can affect a variety of regions of the body, it most often affects the arms, legs, hands, or feet. Often the pain begins in one portion of a limb, but spreads over time to include the entire limb or even to include a different limb. Typical features include dramatic changes in the color and temperature of the skin over the affected limb or body part,
accompanied by intense burning pain, skin sensitivity, sweating, and swelling.
The compounds disclosed herein can also be used to treat endometriosis and the pain associated therewith.
Neurological or Neurodegenerative Diseases and Disorders
Neurodegenerative diseases and disorders include but are not limited to Alzheimer's disease (AD), Parkinson's disease, Huntington's disease, amyotrophic lateral sclerosis (ALS), and other brain disorders caused by trauma or other insults including aging.
Mechanisms associated with calcium signaling may be altered in many
neurodegenerative diseases and in disorders resulting from brain injury. For example, fibroblasts or T-lymphocytes from patients with AD have consistently displayed an increase in Ca release from intracellular stores compared to controls (Ito et al. (1994) Proc. Natl. Acad. Sci. U.S.A. 91 :534-538; Gibson et al. (1996) Biochem. Biophys. ACTA 1316:71-77; Etchenberrigaray et al. (1998) Neurobiology of Disease, 5:37-45). Consistent with these observations, mutations in presenilin genes (PSl or PS2) associated with familial AD (FAD) have been shown to increase InsP3 -mediated Ca2+ release from internal stores (Guo et al. (1996) Neuro Report, 8:379-383; Leissring et al. (1999) J. Neurochemistry, 72: 1061-1068; Leissring et al. (1999) J. Biol. Chem. 274 (46):32535-32538; Leissring et al. (2000) J. Cell Biol. 149 (4):793-797; Leissring et al. (2000) Proc. Natl. Acad. Sci. U.S.A. 97 (15):8590-8593). Furthermore, mutations in PSl or PS2 associated with an increase in amyloidogenic amyloid β peptide generation in AD are reported to be associated with a decrease in intracellular calcium level (Yoo et al. (2000) Neuron, 27 (3):561 -572).
Experimental traumatic brain injury has been shown to initiate massive disturbances in Ca2+ concentrations in the brain that may contribute to further neuronal damage. Intracellular Ca2+ may be elevated by many different ion channels. It has been further shown that channel blockers may be beneficial in the treatment of neurological motor dysfunction when
administered in the acute posttraumatic period (Cheney et al. (2000) J. Neurotrauma, 17 (1 ):83- 91). Seizure
Excitotoxicity of a variety of origins leads to seizures. Commonly excess neuronal firing can drive seizure activity. Compounds that reduce the hyperexcitability of relevant neuronal populations have significant potential in reducing seizure activity. Compounds of the invention that inhibit TRPC5 may reduce hyperexcitability and thus reduce seizure activity.
Proteinuric Kidney Disease
TRPC5 is also expressed in the podocyte of the kidney. It has been proposed that there is an antagonistic regulation of actin dynamics and cell in the podocytes by TRPC5 and TRPC6 (Tian et al., (2010) Science Signaling). Thus, inhibiting TRPC5 may impact the reaction of the podocyte to injury.
Combination Therapy
The present invention provides compounds of the invention for use in vitro and in vivo.
The present invention also provides compositions and pharmaceutical compositions comprising a compound of formula (I) that inhibits TRPC5 activity. In certain embodiments, the compound of the invention is selective. In other words, in certain embodiments, the compound of the invention inhibits TRPC5 activity preferentially over the activity of other ion channels. In certain embodiments, the compound of formula (I) inhibits TRPC5 activity preferentially over TRPVl, TRPV2, TRPV3, TRPV4, TRPC3, TRPC6, TRPC7, TRPA1 , and/or TRPM8 activity. For example, in certain embodiments, the compound of formula (I) inhibits the activity of TRPC5 and also inhibits the activity of one or more of TRPC4, TRPVl , TRPV2, TRPV3, TRPV4, TRPC3, TRPC6, TRPC7, TRPA1 , and TRPM8.
A compound of the invention can be used alone or in combination with other
pharmaceutically active agents. Examples of such other pharmaceutically active agents include, but are not limited to, anti-depressants, anti-anxiety agents, anti-epileptic agents, antiinflammatory agents (e.g., NSAIDS, bradykinin receptor antagonists, hormones and autacoids such as corticosteroids), or anti-migraine agents. Certain active agents belong to more than one category.
In certain embodiments, a compound of the invention is conjointly administered with an analgesic. Suitable analgesics include, but are not limited to, opioids, glucocorticosteroids, nonsteroidal anti-inflammatories, naphthylalkanones, oxicams, para-aminophenol derivatives, propionic acids, propionic acid derivatives, salicylates, fenamates, fenamate derivatives, pyrozoles, and pyrozole derivatives. Examples of such analgesic compounds include, but are not limited to, codeine, hydrocodone, hydromorphone, levorpharnol, morphine, oxycodone, oxymorphone, butorphanol, dezocine, nalbuphine, pentazocine, etodolac, indomethacin, sulindac, tolmetin, nabumetone, piroxicam, acetaminophen, fenoprofen, flurbiprofen, ibuprofen, ketoprofen, naproxen, diclofenac, oxaprozin, aspirin, diflunisal, meclofenamic acid, mefanamic acid, prednisolone, and dexamethasone. Preferred analgesics are non-steroidal anti- inflammatories and opioids (preferably moiphine).
In some embodiments, a compound of the invention can be administered in conjunction with a therapeutic whose administration causes pain. For example, a compound of the invention can be administered in conjunction with an anesthetic, to reduce the pain caused by the administration of the anaesthetic. A compound of the invention can also be administered in conjunction with a chemotherapeutic agent, to reduce the pain caused by administration of the chemotherapeutic agent.
In certain embodiments, a compound of the invention is conjointly administered with a non-steroidal anti-inflammatory. Suitable non-steroidal anti-inflammatory compounds include, but are not limited to, piroxicam, diclofenac, etodolac, indomethacin, ketoralac, oxaprozin, tolmetin, naproxen, flubiprofen, fenoprofen, ketoprofen, ibuprofen, mefenamic acid, sulindac, apazone, phenylbutazone, aspirin, celecoxib and rofecoxib.
Pharmaceutical Compositions
While it is possible for a compound of the invention to be administered alone, it is preferable to administer the compound as a pharmaceutical formulation, where the compound is combined with one or more pharmaceutically acceptable excipients or carriers. The compound of the invenyion may be formulated for administration in any convenient way for use in human or veterinary medicine. In certain embodiments, the compound of the invention may be a prodrug, e.g., capable of being converted to an active compound in a physiological setting.
The phrase "pharmaceutically acceptable" is employed herein to refer to those compounds, materials, compositions, and/or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit/risk ratio.
Examples of pharmaceutically acceptable carriers include: (1 ) sugars, such as lactose, glucose and sucrose; (2) starches, such as corn starch and potato starch; (3) cellulose, and its derivatives, such as sodium carboxymethyl cellulose, ethyl cellulose and cellulose acetate; (4) powdered tragacanth; (5) malt; (6) gelatin; (7) talc; (8) excipients, such as cocoa butter and suppository waxes; (9) oils, such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil and soybean oil; (10) glycols, such as propylene glycol; (1 1 ) polyols, such as glycerin, sorbitol, marmitol and polyethylene glycol; (12) esters, such as ethyl oleate and ethyl laurate; (13) agar; (14) buffering agents, such as magnesium hydroxide and aluminum hydroxide; (15) alginic acid; (16) pyrogen-free water; (17) isotonic saline; (18) Ringer's solution; (19) ethyl alcohol; (20) phosphate buffer solutions; (21) cyclodextrins such as Captisol®; and (22) other non-toxic compatible substances employed in pharmaceutical formulations.
Examples of pharmaceutically acceptable antioxidants include: (1) water soluble antioxidants, such as ascorbic acid, cysteine hydrochloride, sodium bisulfate, sodium
metabisulfite, sodium sulfite and the like; (2) oil-soluble antioxidants, such as ascorbyl palmitate, butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), lecithin, propyl gallate, alpha-tocopherol, and the like; and (3) metal chelating agents, such as citric acid,
ethylenediamine tetraacetic acid (EDTA), sorbitol, tartaric acid, phosphoric acid, and the like.
Solid dosage forms (e.g., capsules, tablets, pills, dragees, powders, granules and the like) can include one or more pharmaceutically acceptable carriers, such as sodium citrate or dicalcium phosphate, and/or any of the following: (1) fillers or extenders, such as starches, lactose, sucrose, glucose, mannitol, and/or silicic acid; (2) binders, such as, for example, carboxymethylcellulose, alginates, gelatin, polyvinyl pyrrolidone, sucrose and/or acacia; (3) humectants, such as glycerol; (4) disintegrating agents, such as agar-agar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates, and sodium carbonate; (5) solution retarding agents, such as paraffin; (6) absorption accelerators, such as quaternary ammonium compounds; (7) wetting agents, such as, for example, cetyl alcohol and glycerol monostearate; (8) absorbents, such as kaolin and bentonite clay; (9) lubricants, such a talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate, and mixtures thereof; and (10) coloring agents.
Liquid dosage forms can include pharmaceutically acceptable emulsions,
microemulsions, solutions, suspensions, syrups and elixirs, in addition to the compound of the invention, the liquid dosage forms may contain inert diluents commonly used in the art, such as, for example, water or other solvents, solubilizing agents and emulsifiers, such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1 ,3-butylene glycol, oils (in particular, cottonseed, groundnut, corn, germ, olive, castor and sesame oils), glycerol, tetrahydrofuryl alcohol, polyethylene glycols and fatty acid esters of sorbitan, and mixtures thereof.
Suspensions, in addition to the compound of the invention, may contain suspending agents as, for example, ethoxylated isostearyl alcohols, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum metahydroxide, bentonite, agar-agar and tragacanth, and mixtures thereof.
Ointments, pastes, creams and gels may contain, in addition to the compound of the invention, excipients, such as animal and vegetable fats, oils, waxes, paraffins, starch, tragacanth, cellulose derivatives, polyethylene glycols, silicones, bentonites, silicic acid, talc and zinc oxide, or mixtures thereof.
Powders and sprays can contain, in addition to the compound of the invention, excipients such as lactose, talc, silicic acid, aluminum hydroxide, calcium silicates and polyamide powder, or mixtures of these substances. Sprays can additionally contain customary propellants, such as chlorofluorohydrocarbons and volatile unsubstituted hydrocarbons, such as butane and propane.
The formulations may conveniently be presented in unit dosage form and may be prepared by any methods well known in the art of pharmacy. The amount of a compound of the invention which can be combined with a carrier material to produce a single dosage form will vary depending upon the host being treated, the particular mode of administration. The amount of a compound of the invention that can be combined with a carrier material to produce a single dosage form will generally be that amount of the compound which produces a therapeutic effect. Generally, out of one hundred percent, this amount will range from about 1 percent to about ninety-nine percent of a compound of the invention, preferably from about 5 percent to about 70 percent, most preferably from about 10 percent to about 30 percent.
The tablets, and other solid dosage forms of the pharmaceutical compositions disclosed herein, such as dragees, capsules, pills and granules, may optionally be scored or prepared with coatings and shells, such as enteric coatings and other coatings well known in the
pharmaceutical-formulating art. They may also be formulated so as to provide slow or controlled release of the active ingredient therein using, for example, hydroxypropylmethyl cellulose in varying proportions to provide the desired release profile, other polymer matrices, liposomes and/or microspheres. They may be sterilized by, for example, filtration through a bacteria- retaining filter, or by incorporating sterilizing agents in the form of sterile solid compositions that can be dissolved in sterile water, or some other sterile injectable medium immediately before use. These compositions may also optionally contain pacifying agents and may be of a composition that they release the active ingredient(s) only, or preferentially, in a certain portion of the gastrointestinal tract, optionally, in a delayed manner. Examples of embedding
compositions that can be used include polymeric substances and waxes. The active ingredient can also be in micro-encapsulated form, if appropriate, with one or more of the above-described excipients.
Dosage forms for the topical or transdermal administration of a compound of the .
invention include powders, sprays, ointments, pastes, creams, lotions, gels, solutions, patches and inhalants. The active compound may be mixed under sterile conditions with a
pharmaceutically acceptable carrier, and with any preservatives, buffers, or propellants that may be required.
The formulations disclosed herein can be delivered via a device. Exemplary devices include, but are not limited to, a catheter, wire, stent, or other intraluminal device. Further exemplary delivery devices also include a patch, bandage, mouthguard, or dental apparatus. Transdermal patches have the added advantage of providing controlled delivery of a compound of the invention to the body. Such dosage forms can be made by dissolving or dispersing the compound of the invention in the proper medium. Absorption enhancers can also be used to increase the flux of the compound across the skin. The rate of such flux can be controlled by either providing a rate controlling membrane or dispersing the compound in a polymer matrix or gel.
Ophthalmic formulations, eye ointments, drops, solutions and the like, are also contemplated as being within the scope of this invention.
In some cases, in order to prolong the effect of a drug, it is desirable to slow the absorption of the drug from subcutaneous or intramuscular injection. This may be accomplished by the use of a liquid suspension of crystalline or amorphous material having poor water solubility. The rate of absorption of the drug then depends upon its rate of dissolution, which, in turn, may depend upon crystal size and crystalline form. Alternatively, delayed absorption of a parenterally administered drug form is accomplished by dissolving or suspending the drug in an oil vehicle.
Injectable depot forms are made by forming microencapsule matrices of the subject compounds in biodegradable polymers such as polylactide-polyglycolide. Depending on the ratio of drug to polymer, and the nature of the particular polymer employed, the rate of drug release can be controlled. Examples of other biodegradable polymers include poly(orthoesters) and poly(anhydrides). Depot injectable formulations are also prepared by entrapping the drug in liposomes or microemulsions that are compatible with body tissue.
When a compound of the invention is administered as a pharmaceutical, to humans and animals, it can be given per se or as a pharmaceutical composition containing, for example, 0.1 to 99.5% (more preferably, 0.5 to 90%) of the compound of the invention in combination with a pharmaceutically acceptable carrier.
The formulations can be administered topically, orally, transdermally, rectally, vaginally, parentally, intranasally, intrapulmonary, intraocularly, intravenously, intramuscularly, intraarterially, intrathecally, intracapsularly, intraorbitally, intracardiacly, intradermally, intraperitoneal ly, transtracheal ly, subcutaneously, subcuticularly, intraarticularly, subcapsularly, subarachnoid^, intraspinally, intrasternally, sublingually, or by inhalation.
Dosages
Actual dosage levels of the the compound of the invention in the pharmaceutical compositions of this invention may be varied so as to obtain an amount of the active ingredient that is effective to achieve the desired therapeutic response for a particular patient, composition, and mode of administration, without being toxic to the patient.
The selected dosage level will depend upon a variety of factors including the activity of the compound of the invention disclosed herein employed, or the ester, salt or amide thereof, the route of administration, the time of administration, the rate of excretion of the particular compound being employed, the duration of the treatment, other drugs, compounds and/or materials used in combination with the particular compound employed, the age, sex, weight, condition, general health and prior medical history of the patient being treated, and like factors well known in the medical arts.
A physician or veterinarian having ordinary skill in the art can readily determine and prescribe the effective amount of the pharmaceutical composition required. For example, the physician or veterinarian could start doses of the compound of the invention in the
pharmaceutical composition at levels lower than that required in order to achieve the desired therapeutic effect and gradually increase the dosage until the desired effect is achieved.
In general, a suitable daily dose of a compound of the invention will be that amount of the compound that is the lowest dose effective to produce a therapeutic effect. Such an effective dose will generally depend upon the factors described above. Generally, intravenous, intracerebro ventricular and subcutaneous doses of the compounds of the invention for a patient will range from about 0.0001 to about 100 mg per kilogram of body weight per day. For example, the dose can be 0.1 -50, 0.1 -25, 0.5-10, 1 -10, or 5-10 mg/kg.
If desired, the effective daily dose of the compound of the invention may be administered as two, three, four, five, six or more sub-doses administered separately at appropriate intervals throughout the day, optionally, in unit dosage forms.
Disease and Injury Models
A compound of the invention which antagonizes TRPC5 function may be useful in the prophylaxis and treatment of any of the foregoing injuries, diseases, disorders, or conditions. In addition to in vitro assays of the activity of the compound of the invention, its efficacy can be readily tested in one or more animal models. By way of example, numerous well known animal models exist. One or more suitable animal models (e.g., suitable in light of the particular indication) can be selected.
Fear-related behaviors can be measured as described, e.g., in Riccio et al. Pain behaviors can be studied using various agents or procedures to simulate pain resulting from injuries, diseases, or other conditions. Blackburn-Munro (2004) Trends in Pharmacological Sciences 25: 299-305 (see, for example, Table 1). Behavioral characteristics of challenged animals can then be observed. Compounds or procedures that may reduce pain in the animals can be readily tested by observing behavioral characteristics of challenged animals in the presence versus the absence of the test compound(s) or procedure.
Exemplary behavioral tests used to study chronic pain include tests of spontaneous pain, allodynia, and hyperalgesia. Id. To assess spontaneous pain, posture, gait, nocifensive signs (e.g., paw licking, excessive grooming, excessive exploratory behavior, guarding of the injured body part, and self-mutilation) can be observed. To measure evoked pain, behavioral responses can be examined following exposure to heat (e.g., thermal injury model).
Exemplary animal models of pain include, but are not limited to, the Chung model, the carageenan induced hyperalgesia model, the Freund's complete adjuvant induced hyperalgesia model, the thermal injury model, the formalin model and the Bennett Model. The Chung model of neuropathic pain (without inflammation) involves ligating one or more spinal nerves. Chung et al. (2004) Methods Mol Med 99: 35-45; Kim and Chung (1992) Pain 50: 355-363. Ligation of the spinal nerves results in a variety of behavioral changes in the animals including heat hyperalgesia, cold allodynia, and ongoing pain. Compounds that antagonize TRPC5 can be administered to ligated animals to assess whether they diminish these ligation-induced behavioral changes in comparison to that observed in the absence of compound.
Useful anxiety and depression models include the maternal separation model, the elevated plus-maze model, the forced swim test, the tail suspension test, the light/dark preference model, the light-enhanced startle model, and the ultrasonic vocalization model.
Useful seizure models include but are not limited to maximal electric shock (MES), acoustic startle in susceptible animals (eg DBA mice), and chemical induced seizure (with proconvulsant compounds such as pilocarpine, pentalene tetrazole, kainic acid, N-methyl-D- aspartic acid).
Useful models of kidney function include the LPS-induced proteinuria (waiting for a reference for others).
Examples
Method A: Patch Clamp Experiments
Patch clamp experiments permit the detection of currents through the TRPC5 channel in the cell line described above. In normal whole-cell patch clamp recordings, a glass electrode is brought into contact with a single cell and a high-resistance (gigaohm) seal is established with the cell membrane. The membrane is then ruptured to achieve the whole-cell configuration, permitting control of the voltage of the cell membrane and measurement of currents flowing across the membrane using the amplifier attached to the electrode and resulting in the replacement of cytoplasm with the pipette solution. A perfusion system permits control of the extracellular solution, including the addition of blockers and activators of the current. The current can be activated by including 1 .4 μΜ free Ca2+ in the pipette (intracellular) solution, and 80 μΜ LaCl3 in the extracellular solution.
TRPC5 cells were induced 20-48 hours, removed from growth plates, and replated at low density (to attain good single-cell physical separation) on glass coverslips for measurement. In some cases, cells were grown in low density overnight on glass coverslips. Patch clamp recordings were made in the whole-cell mode with a holding potential of -40 mV. Every 5 seconds, a voltage ramp was applied from -120 to +100 mV, 400 ms in duration. Currents elicited were quantified at -80 mV and +80 mV. The internal solution consisted of 140 mM cesium aspartate, 10 mM HEDTA, 2 mM CaCl2, 2.27 mM MgCl2 and 10 mM HEPES, pH 7.2, with 1 ,400 nM calculated free Ca2+. The external solution consisted of 150 mM NaCl, 4.5 mM KCl, 1 mM MgCl2, 2 mM CaCl2, 10 mM HEPES, 10 mM glucose, 1 mM EGTA, pH 7.4. Upon addition of LaCl3, TRPC5 current was induced only in TRPC5-expressing cells and not in parental HEK293 TREx cells. Removal of the LaCl3 stimulus causes most of the current to go away. Potential blockers were tested for ability to block both inward and outward currents in the continued presence of LaCl3.
IC50 of a compound of the invention was estimated by testing the compound at 5 μΜ and 500 nM. When 5 μΜ of a compound showed no block, IC50 was estimated as >10 μΜ. When 5 μΜ of a compound showed 50% or less block, a rough estimate of IC50 in the range of 5-10 μΜ could be made. IC50 for a compound of Formula I or Formula II between 500 nM and 5 μΜ was similarly estimated.
A compound described herein may be tested for its ability to block both inward and outward currents through the TRPC5 channel, e.g., by an assay as described in Example 1. For example, IC50 of a compound of the invention was estimated by testing the compound at 5 μΜ and 500 nM. When 5 μΜ of a compound showed no block, IC50 was estimated as >10 μΜ. When 5 μΜ of a compound showed 50% or less block, a rough estimate of IC5o in the range of 5-10 μΜ could be made. IC50 for a compound of the invention between 500 nM and 5 μΜ was similarly estimated. Exemplary compounds are shown in Table B below. As shown in Table B, "A" refers to an 1C50 < 100 nM. "B" refers to an IC50 between 100 nM and 500 nM. "C" refers to an IC5o between 500 nM and 1000 nM. "D" refers to an IC50 between 1 μΜ and 2 μΜ. "E" refers to an 1C50 between 2 μΜ and 10 μΜ. "F" refers to agonist compounds. "ND" refers to compounds wherein the IC50 was not determined.
Table B
Figure imgf000166_0001
Figure imgf000167_0001
Figure imgf000168_0001
Figure imgf000169_0001
Figure imgf000170_0001
Figure imgf000171_0001
Figure imgf000172_0001
Figure imgf000173_0001
Figure imgf000174_0001
Figure imgf000175_0001
Figure imgf000176_0001
Figure imgf000177_0001
Figure imgf000178_0001
Figure imgf000179_0001
Figure imgf000180_0001
Figure imgf000181_0001
Figure imgf000182_0001
Figure imgf000183_0001
Figure imgf000184_0001
Figure imgf000185_0001
Figure imgf000186_0001
Figure imgf000187_0001
Figure imgf000188_0001
Figure imgf000189_0001
Figure imgf000190_0001
Figure imgf000191_0001
Figure imgf000192_0001
Figure imgf000193_0001
Figure imgf000194_0001
Figure imgf000195_0001
Figure imgf000196_0001
Figure imgf000197_0001
Figure imgf000198_0001
Figure imgf000199_0001
224 A
1
226 Β
227 Β
228 D
229 Ε
1
Figure imgf000201_0001
Figure imgf000202_0001
Figure imgf000203_0001
Figure imgf000204_0001
Figure imgf000205_0001
260 A
262 A
1
263 A 65 B 66 A
1,
Figure imgf000207_0001
Figure imgf000208_0001
Figure imgf000209_0001
Figure imgf000210_0001
Figure imgf000211_0001
Figure imgf000212_0001
Figure imgf000213_0001
Figure imgf000214_0001
Figure imgf000215_0001
Figure imgf000216_0001
Figure imgf000217_0001
326 A
327 A
1
329 A 31 A
1
33 A
1
Figure imgf000219_0001
Figure imgf000220_0001
Figure imgf000221_0001
Figure imgf000222_0001
Figure imgf000223_0001
Figure imgf000224_0001
Figure imgf000225_0001
Figure imgf000226_0001
Figure imgf000227_0001
Figure imgf000228_0001
Figure imgf000229_0001
227
Figure imgf000230_0001
Figure imgf000231_0001
Figure imgf000232_0001
Figure imgf000233_0001
522 A
1
523 A
1.
524 A
1
525 A
527 A
Figure imgf000235_0001
Figure imgf000236_0001
Figure imgf000237_0001
Figure imgf000238_0001
Figure imgf000239_0001
Figure imgf000240_0001
Figure imgf000241_0001
23.9
Figure imgf000242_0001
595 A
596 A
V-,
597 A
598 A
599 A
Figure imgf000244_0001
Figure imgf000245_0001
Figure imgf000246_0001
Figure imgf000247_0001
Figure imgf000248_0001
Figure imgf000249_0001
Figure imgf000250_0001
Figure imgf000251_0001
Method B: High Thoughput Screening Assay
The high throughput assay depended on detection of the rise in intracellular Ca concentration ([Ca ],) following channel activation in cells inducibly expressing the TRPC5 channel. Ca2+ rise was quantified with the use of fluorescent Ca2+ indicators that were loaded into cells and thereafter indicated the [Ca2+]j Ca2+ influx following activation of the TRPC5 channel. Compounds inhibiting the [Ca ]j rise were considered hits for further investigation. The commercially available HEK293/TREx line (Invitrogen) was stably transfected with a TRPC5 construct and screened by conventional calcium imaging to find clones with TRPC5 expression following stimulation with 1 g/ml tetracycline. These cells were maintained in the growth medium recommended by the manufacturer supplemented with 100 μg /ml hygromycin to promote retention of the TRPC5 construct. After growing to near confl uency, cells were plated at a density of -35,000 cells/well in 384 well CellBind plates (Corning) in the presence of 1 μg/ml tetracycline, and allowed to grow for 20-30 hrs. A nearly confluent monolayer resulted. Cells were then loaded with Ca2+ dye: Fura-2/AM or Fluo4/AM was added to the wells to a final concentration of 4 μΜ or 0.5 μΜ, respectively, and incubated for 80 min or 60 min, respectively, at room temperature. Supernatant was then removed from the cells by inverting plates with a sharp flick, and 25 μΐ Hank's Balanced Salt Solution (HBSS; 0.185 g/1 D-glucose, 0.9767 g/1 MgS04 (anhydrous), 0.4 g/1 KCl, 0.06 g/1 KH2P04 (anhydrous), 0.35 g/1 NaHC03, 8.0 g/1 NaCl, and 0.04788 g/1 Na2HP04 (anhydrous); pH 7.4) was then added to each well. Following -0.5 hour for recovery from loading, cells were assayed using the Hamamatsu FDSS 6000 system, which permitted illumination alternately at 340 nm and 380 nm for Fura-2 experiments, or at 485 nm for Fluo4 experiments. Frames were acquired at a rate of 0.2 Hz. During the assay, the plates were continuously vortexed, with pipette mixing of wells following addition of each reagent.
For the screening assay, 26μ1 of a diluted compound stock (at 50 μΜ) was added to each well for 2 minutes following the collection of a short (4 frame) baseline. 13 μΐ 62 mM high- Ca2+ Ringer solution (4.17 ml of normal ringer (with 2mM Ca2+) plus 5.83 ml of isotonic calcium ringer (105mM Ca2+; in this ringer all sodium has been replaced with calcium)) was then added to each well, achieving a final concentration of 14 mM Ca2+ and 10 μΜ test compound. Data was collected for ~3 minutes following addition of high Ca2+ Ringer, where the fluorescent intensity (for Fluo4) and the F340/F380 ratio (for Fura-2) were proportional to the [Ca ]i
Negative controls consisted of HEK293/TREx TRPC5 cells exposed to high Ca2+ solution, but no compound. Positive control conditions consisted of addition of 2-APB, a promiscuous blocker of TRPC5 and other channels, to columns 23 and 24 of the plates, to a final
concentration of 200 μΜ. These controls defined a screening window, and "hits" were defined as those compounds inhibiting the fluorescence response by at least 40%. 1C50 values were determined for compounds defined as "hits." The Fluo4 cell-based fluorescence assay was used to determine the intracellular Ca2+ concentration in the presence of varying drug concentration. Final concentrations of compounds tested were 20 μΜ, 6.667 μΜ, 2.222 μΜ, 0.741 μΜ, 0.247 μΜ, 0.082 μΜ, and 0.027 μΜ. Compounds were tested in triplicate at all concentrations.
Standard software was used to fit IC50 curves.
Additionally or alternatively, efficacy can be represented as % inhibition in the presence (of a given concentration of compound) versus the absence of compound or in comparison to a control compound. For example, efficacy can be represented as % inhibition of ion flux in the presence versus the absence of compound. Exemplary compounds are shown in Table X below. As shown in Table X, "A" refers to an IC50 < 100 nM. "B" refers to an IC50 between 100 nM and 500 nM. "C" refers to an IC50 between 500 nM and 1000 nM. "D" refers to an IC50 between 1 μΜ and 2 μΜ. "E" refers to an IC50 between 2 μΜ and 10 μΜ. "F" refers to agonist compounds. "ND" refers to compounds wherein the 1C50 was not determined.
Figure imgf000253_0001
Figure imgf000254_0001
Figure imgf000255_0001
Figure imgf000256_0001
Figure imgf000257_0001
Figure imgf000258_0001
Figure imgf000259_0001
Figure imgf000260_0001
Figure imgf000261_0001
Figure imgf000262_0001
Figure imgf000263_0001
Figure imgf000264_0001
Figure imgf000265_0001
Figure imgf000266_0001
Figure imgf000267_0001
Figure imgf000268_0001
Figure imgf000270_0001
Figure imgf000271_0001
Figure imgf000272_0001
Figure imgf000273_0001
121 ' E
122 F
123 D
1
124 ND
126 B
Figure imgf000275_0001
Figure imgf000276_0001
Figure imgf000277_0001
Figure imgf000278_0001
Figure imgf000279_0001
Figure imgf000280_0001
165 E
166
167 ND
169 A
1.
170 A
Figure imgf000282_0001
177 ND
1
178 E
179 D
0
180 C
181 C
Figure imgf000284_0001
Figure imgf000285_0001
Figure imgf000286_0001
Figure imgf000287_0001
Figure imgf000288_0001
Figure imgf000289_0001
Figure imgf000290_0001
Figure imgf000291_0001
Figure imgf000292_0001
Figure imgf000293_0001
Figure imgf000294_0001
Figure imgf000295_0001
Figure imgf000296_0001
262 B
!
263 A
1;
265 B
1
266 A
1
267 B
Figure imgf000298_0001
Figure imgf000299_0001
Figure imgf000300_0001
Figure imgf000301_0001
Figure imgf000302_0001
Figure imgf000303_0001
301 A
302 A
i
303 A
1
304 A
1
305 A
1
Figure imgf000305_0001
Figure imgf000306_0001
Figure imgf000307_0001
Figure imgf000308_0001
Figure imgf000309_0001
Figure imgf000310_0001
Figure imgf000311_0001
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Figure imgf000315_0001
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Figure imgf000318_0001
Figure imgf000319_0001
Figure imgf000320_0001
Figure imgf000321_0001
Figure imgf000322_0001
Figure imgf000323_0001
Figure imgf000324_0001
Figure imgf000325_0001
Figure imgf000326_0001
Figure imgf000327_0001
Figure imgf000328_0001
Figure imgf000329_0001
Figure imgf000330_0001
Figure imgf000331_0001
Figure imgf000332_0001
Figure imgf000333_0001
Figure imgf000334_0001
Figure imgf000335_0001
Figure imgf000336_0001
Figure imgf000337_0001
Figure imgf000338_0001
Figure imgf000339_0001
Figure imgf000340_0001
Figure imgf000341_0001
Figure imgf000342_0001
Figure imgf000343_0001
Figure imgf000344_0001
Figure imgf000345_0001
Figure imgf000346_0001
Figure imgf000347_0001
Figure imgf000348_0001
Figure imgf000349_0001
Figure imgf000350_0001
Figure imgf000351_0001
Figure imgf000352_0001
Figure imgf000353_0001
Figure imgf000354_0001
Figure imgf000355_0001
Figure imgf000356_0001
Figure imgf000357_0001
Figure imgf000358_0001
Figure imgf000359_0001
Figure imgf000360_0001
Figure imgf000361_0001
Figure imgf000362_0001
Figure imgf000363_0001
Figure imgf000364_0001
Figure imgf000365_0001
Example 2:
General Procedures
All reagents were purchased from commercial suppliers and used without further purification unless otherwise stated. THF was continuously refluxed and freshly distilled from sodium and benzophenone under nitrogen, DCM was continuously refluxed and freshly distilled from Ca¾ under nitrogen.
Reactions were monitored via TLC on silica gel plates (either 60 HSGF254 percolated plates (0.15-0.2 mm Si02) or Baker-flex IB2-F TLC plates), and visualized using UV light (254 nm or 365 nm) and/or staining with a solution of DNP (12 g, 2,4-dinitrofenylhydrazin, 60 mL concentrated H2S0 , 80 ml H20, 200 mL ethanol) and subsequent heating or monitored via LCMS.
Microwave reactions were carried out with a Biotage Smith Synthesizer.
LCMS were performed on a SHIMADZU LCMS-2010EV instrument using one of two sets of conditions. LCMS conditions one: (Chromolith SpeedROD, RP-18e column, 50x4.6 mm, mobile phase: Solvent A: CH3CN/H20 /HCOOH=l 0/90/0:05, Solvent B: CH3CN/H20
/HCOOH=90/10/0.05, 0.8min@ 10% B, 2.7min gradient (10-95% B), then 0.8min@95%B, Flow rate: 3mL/min, temperature: 40°C). LCMS conditions two: (Zorbax , 3.5 micron, 2.1 x 50 mm CI 8 column. Mobile phase: Solvent A: 0.1% formic acid /acetonitrile Solvent B: 0.1% formic acid / water. Gradient: 5% to 95%» B using a 5 min or 8 min runtime).
Preparative HPLC were performed either on a SHIMADZU LC-8A instrument. (Column: YMC Pack ODS-A (150*30mm, ΙΟμηι)) or LC-6AD (Column: Shim=Pack PREP-ODS-H
(250*20mm, Ι Ομηι)) with UV detection which were controlled by LC solution Chemstation software. H20 (0.1 % HCOOH) and methanol (MeCN) as mobile phase at the indicated flow rate.
Analytical HPLC were performed on a SHIMADZU LC-2010A instrument. (Chromolith SpeedROD, RP-18e, 50x4.6 mm, mobile phase: Solvent A: CH3CN/H20 /HCOOH=l 0/90/0.05, Solvent B: CH3CN/H20 /HCOOH=90/10/0.05, 0.8min@ 10% B, 2.7min gradient (10-95% B), then 0.8min@95%B, Flow rate: 3mL/min, temperature: 40°C).
'H-NMR spectra were recorded on either a Bruker Avance II 400MHz or a Varian Unity Inova 400 MHz instrument. Chemical shifts (δ) are reported in ppm relative to tetramethylsilane (δ = 0.000 ppm) and the spectra were calibrated to the residual solvent signal of chloroform (δ = 7.26), Dimethyl sulfoxide (δ = 2.50), methanol (δ = 3.30). Data for lH-NMR spectra are reported as follows: chemical shift (multiplicity, number of hydrogens). Abbreviations are as follows: s (singlet), d (doublet), t (triplet), q (quartet), quint (quintet), m (multiplet), br (broad). List of abbreviations and terms
BPO benzoyl peroxide
CDI carbonyldiimidazole
Chromatography compound purification using silica gel
Concentrated [or concentrated at reduced pressure] solvent removal with the aid of a rotary evaporation device
DCM dichloromethane
DBU l,8-diazabicyclo[5.4.0]undec-7-ene
Dess Martin [or Dess Martin periodinane] l,l,l-triacetoxy-l ,l -dihydro-l ,2-benziodoxol- 3(lH)-one
Dilute HC1 IN hydrochloric acid
DMF N,N-dimethylformamide
DMAP 4-dimethylaminopyridine
Dried, referring to removal of residual water from organic solutions implies the use of an inorganic drying agent such as sodium sulfate
Dried in vacuo [or dried under vacuum] residual solvent removal with the aid of a vacuum pump
DMSO dimethyl sulfoxide
Eaton's reagent 7.7 wt% phosphorus pentoxide in methanesulfonic acid
EDCI l-ethyl-3-(3-dimethylaminopropyl)carbodiimide)
Evaporated solvent removal with the aid of a rotary evaporation device
h hour
HMDS hexamethyldisilazane
LAH lithium aluminum hydride
MCPBA 3-chloroperoxybenzoic acid
min minutes
n-BuLi n-Butyllithium
NBS N-bromosuccinimide
NCS N-chlorosuccinimide
NMP N-methylpyrolidinone Oxone potassium peroxomonosulfate
Pd/C palladium on activated carbon
Pd-dppf l, -Bis(diphenylphosphino)ferrocene-palladium(II)dichloride dichloromethane complex
PMB 4-methoxybenzyl
PPTS pyridinium p-toluenesulfonate
Preparative TLC preparative thin layer chromatography
SEM (trimethylsilyl)ethoxy)methyl
TBAI tetrabutyl ammonium iodide
TBAF tetrabutylammonium fluoride
TBAH tetrabutylammonium hydroxide
TEA triethylamine
TFA trifluoroacetic acid
THF tetrahydrafuran
tic thin layer chromatography on silica gel
X-phos 2-(dicyclohexylphosphino)-2',4',6'-triisopropylbiphenyl
Purine numbering
Figure imgf000368_0001
Preparation of intermediates
Intermediate 1 2-( -(2-chloroethoxy)ethoxy)tetrahydro-2H-pyran
Figure imgf000369_0001
To a solution of 2-(2-chloroethoxy)ethanol (10 g, 0.071 mol) in DCM (150 mL) was added PPTS (891 mg, 3.6 mmol), then dihydropyran (10.2 g, 0.107 mol) was added dropwise at 0 °C. The mixture was stirred at room temperature for 16 h. It was partitioned between DCM and water. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give a crude product which was purified by silica gel chromatography eluted with petroleum ether to give 2-(2-bromoethoxy)tetrahydro-2H-pyran(l 1.9 g, 79.8% yield) as light yellow oil. lH- NMR (CDC13) δ 4.65-4.67 (t, 1H), 3.86-3.92 (m, 2H), 3.78-.381 (m, 2H), 3.71-3.73 (t, 2H), 3.64-3.67(t, 2H), 1.77-1.87 (m, 2H), 1.51-1.64(m, 6H).
Intermediate 2 2-(2-bromoethoxy)tetrahydro-2H-pyran
HO ^o'
Br THPO^-
Br
PPTS, DCM
To a solution of 2-bromoethanol (15 g, 0.12 mol) in DCM (150 mL) was added PPTS (891 mg, 3.6 mmol), then dihydropyran (10.6 g, 0.126 mol) was added dropwise at 0 °C. The mixture was stirred at room temperature for 4 h. It was partitioned between DCM and water. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give a crude product which was purified by column chromatography eluting with petroleum ether/ethyl acetate (50: 1) to give 2-(2-bromoethoxy)tetrahydro-2H-pyran(18.0 g, 72.0% yield) as a light yellow oil.
Intermediate 3 2-(3-bromopropoxy)tetrahydro-2H-pyran
PPTS.DCM To a solution of 3-bromopropan-l -ol (8 g, 57.9 mmol) in DCM(100 mL) was added PPTS (891 mg, 3.6 mmol), then dihydropyran (7.3 g, 86.9 mmol) was added dropwise at 0 °C. The mixture was stirred at room temperature overnight. It was partitioned between DCM and water. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give a crude product which was purified by column chromatography eluting with petroleum ether/ethyl acetate (50: 1) to give 2-(3-bromopropoxy)tetrahydro-2H-pyran (8.9 g, 69.0% yield) as a light yellow oil. -(chloromethyl)-5-meth lthiazole
Figure imgf000370_0001
(5-methylthiazol-2-yl)methanol
VSV_ Rr
Figure imgf000370_0002
To a solution of n-BuLi (8.4 ml, 13.48 mmol) in THF (30 mL) was added 2-bromo-5- methylthiazole(2.0 g, 1 1.23 mmol) dropwise under a nitrogen atmosphere at -70 °C ; then it was stirred at this temperature for 1.5 h. DMF (1.3 ml, 16.85 mmol) was added dropwise under nitrogen atmosphere at -70 °C. The resulting mixture was stirred at this temperature for 1 h. Then the mixture was quenched with aqueous saturated ammonium chloride (5 mL), the mixture was partitioned between ethyl acetate and water. The organic layers were combined, dried over sodium sulfate, filtered and concentrated to give a yellow oil. The yellow oil was dissolved in methanol (15 ml), sodium borohydride (512 mg, 13.48 mmol) was added portionwise under a nitrogen atmosphere at -60 °C. The mixture was stirred at this temperature for 1 h. The reaction mixture was quenched with acetone and concentrated. The residue was partitioned between ethyl acetate and water. The organic layers were dried over sodium sulfate, filtered and concentrated, then purified by silica gel chromatography eluting with petroleum/ ethyl acetate= 3: 1 to give thiazol-2-ylmethanol (1.3 g, 90.3%) as brown oil . LCMS retention time 0.366 min; LCMS MH+ 130.
Step 2 2-(chloromethyl)-5-methylthiazole
Figure imgf000371_0001
To a solution of (5-methylthiazol-2-yl)methanol (0.5 g, 3.87 mmol) in DCM (5 mL) was added thionyl chloride (0.19 ml, 2.6 mmol) at 0 °C, then the mixture was stirred at room temperature for 2 h. The solvent was concentrated to give 2-(chloromethyl)-5-methylthiazole (570 mg) as a yellow oil which was used without purification. LCMS retention time 0.912 min; LCMS MH+ 148.
Intermediate 5 -(3-(trifluoromethoxy)phenoxy)ethanol
Figure imgf000371_0002
To a solution of 3-(trifluoromethoxy)phenol(3 g, 16.8 mmol) in DMF (30 mL) was added 2- bromoethanol (3.16 g, 25.3 mmol), potassium carbonate(4.65 g, 33.7 mmol). The mixture was heated at 80 °C overnight. The mixture was cooled, partitioned between ethyl acetate and water. The organic layers were combined, dried over sodium sulfate, filtered and concentrated to give a crude product which was purified by a column chromatography eluting with petroleum ether and ethyl acetate(10: l) to give 2-(3-(trifluoromethoxy)phenoxy)ethanol (3.5 g, 94.5 % yield) as a yellow oil. 'H-NMR (DMSO-i/6) δ 7.28-7.30 (t, 1H), 6.81 -6.86 (m, 2H), 6.78 (s, 1 H), 4.07-4.09 (t, 2H), 3.95-3.99 (m, 2H), 2.58 (t, 1H).
Intermediate 6 8-bromo-7-ethyl-3-methyl-l -(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-lH- purine-2,6(3H,7H)-dione
Figure imgf000371_0003
Step 1 8-bromo-7-ethyl-l -(3-hydroxypropyl)-3-methyl-l H-purine-2,6(3H,7H)-dione
Figure imgf000371_0004
To a solution of 8-bromo- l-(3-hydroxypropyl)-3-methyl-lH-purine-2,6(3H,7H)-dione(0.4 g, 1.32 mmol, product of intermediate 16 step 2) in DMF (10 mL) was added iodoethane (0.25 g, 1.58 mmol) and potassium carbonate (0.27 g, 1.98 mmol). The reaction was heated at 50 °C for 2 h. The mixture was cooled and partitioned between ethyl acetate and water. The combined organic layer was dried over sodium sulfate, filtered and concentrated to give 8-bromo-7-ethyl-l - (3-hydroxypropyl)-3 -methyl- l H-purine-2,6(3H,7H)-dione (0.4 g, 91 .7% yield) as white solid. LCMS retention time 1.674 min; LCMS MH+ 331.
Step 2 8-bromo-7-ethyl-3-methyl-l -(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-l H-purine- 2,6(3H,7H)-dion
Figure imgf000372_0001
To a solution of 8-bromo-7-ethyl-l -(3-hydroxypropyl)-3-methyl-lH-purine-2,6(3H,7H)- dione(0.4 g, 1.2 mmol) in DCM (10 mL) was added PPTS (27 mg, 0.1 mmol); then dihydropyran (0.15 g, 1.8 mmol) was added dropwise at 0 °C. The mixture was stirred at room temperature overnight. The mixture was partitioned between DCM and water. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give a a crude product which was purified by a column chromatography eluting with petroleum ether/ethyl acetate (3 : 1 ) to give 8-bromo-7-ethyl-3 -methyl- 1 -(3-(tetrahydro-2H-pyran-2-yloxy)propyl)- 1 H-purine- 2,6(3H,7H)-dione (0.38 g,76.9% yield) as white solid. LCMS retention time 1.871 min; LCMS MH+-THP 331. Intermediate 7 7-(4-chlorobenzyl)-8-mercapto-3-methyI-l-(3-(tetrahydro-2H-pyran-2- yloxy)propy -lH-purine-2,6(3H,7H)-dione
Figure imgf000372_0002
To a solution of 8-bromo-7-(4-chlorobenzyl)-3-methyl-l -(3-(tetrahydro-2H-pyran-2- yloxy)propyl)-l H-purine-2,6(3H,7H)-dione (400 mg,0.781 mmol, intermediate 14) in DMF (10 mL) was added sodium sulfide nonahydrate (375 mg, 1.563 mmol). The reaction was stirred at 85 °C for 16 h. The mixture was cooled, diluted with ethyl acetate and water, and the phases were separated. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give 7-(4-chlorobenzyl)-8-mercapto-3-methyl-l -(3-(tetrahydro-2H-pyran-2- yloxy)propyl)-lH-purine-2,6(3H,7H)-dione (230 mg, 63.4%) as orange oil. LCMS retention time 1.517 min; LCMS MH+-THP 381.
Intermediate 8 8-bromo-7-(4-chlorobenzyl)-3 -methyl- lH-purine-2,6(3H,7H)-dione
Figure imgf000373_0001
Step 1 8-bromo-3-meth -lH-purine-2,6(3H,7H)-dio
Figure imgf000373_0002
To a solution of 3 -methyl- lH-purine-2,6(3H,7H)-dione (11.3 g, 6.8 mmol) in acetic acid (300 mL) was added sodium acetate (8.37 g, 13.6 mmol) followed by drop- wise addition of bromine (13.04 g, 8.2 mmol) at 50 °C. After addition, the mixture was stirred at 65 °C for 3 h. The mixture was cooled and poured into ice-water (500 g), and the product precipitated. The slurry was filtered and the filter cake was washed with water twice and dried under vacuum to give 8- bromo-3 -methyl- l H-purine-2,6(3H,7H)- dione (16.1 g, 96.6% yield) as light yellow solid.
LCMS retention time 0.541 min; LCMS MH+ 245.
Step 2 8-bromo-7-(4- ne
Figure imgf000373_0003
To a solution of 8-bromo-3-methyl-lH-purine-2,6(3H,7H)-dione (10. 8 g, 4.42 mmol) in DMF (200 mL) was added l-(bromomethyl)-4-chlorobenzene (10 g, 4.86 mmol) followed by potassium carbonate (9.16 g, 6.63 mmol). The resulting mixture was stirred at 45 °C for 2 h . The mixture was diluted with ethyl acetate (300 mL) and washed with brine (200 mL). The layers were separated and the organic slurry was filtered and the filter cake was washed with ice cold ethanol, dried under vacuum to give 8-bromo-7-(4-chlorobenzyl)-3-methyl- lH-purine- 2,6(3H,7H)-dione (10.6 g, 64.9% yield) as white solid. LCMS retention time 1 .637 min; LCMS MH+ 369.
Intermediate 9 7-(4-chlorobenzyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine- 2,6(3I-l,7H)-dione
Figure imgf000374_0001
•The title compound was prepared using the method of intermediate 12, step 2 to give 7-(4- chlorobenzyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-l H-purine-2,6(3H,7H)-dione (1.97 g, 80.1% yield) as white solid. LCMS retention time 1.724 min, LCMS MH+ 467.
Intermediate 10 8-chloro-7-(4-chlorobenzyl)-l-methyl-lH-purine-2,6(3H,7 -dione
Figure imgf000374_0002
Step 1 6-amino-3-methylpyrimidine-2,4(lH,3H)-dione
Figure imgf000374_0003
To a solution of the 6-aminopyrimidine-2,4(lH,3H)-dione (15 g, 1 18 mmol) in 1 ,1 ,1 ,3,3,3- hexamethyldisilazane (50 mL) was added ammonium sulfate(0.671g, 5 mmol), then the resulting mixture was heated to reflux with stirring for 6 h. The mixture was concentrated to give a light-yellow solid. This solid was combined with acetonitrile (50 mL) and iodomethane (15 mL, 250 mmol) was added. The resulting mixture was stirred at 40 °C for 16 h. Then the mixture was concentrated, neutralized with saturated sodium bicarbonate to pH =7, filtered, and the filter cake was washed with brine and ethanol, dried under vacuum to give 6-amino-3- methylpyrimidine-2,4 (lH,3H)-dione (7.1 g, 42.6 % yield) as yellow solid. LCMS MH+ 142. Step 2 6-amino-3-methyl-5-nitrosopyrimidine-254(lH,3H)-dione
Figure imgf000375_0001
To a solution of 6-amino-3-methylpyrimidine-2,4(lH,3H)-dione (6 g, 42.6 mmol) in acetic acid (50 mL) was added a solution of sodium nitrite (6.8 g, 98.6 mmol) in water (20 mL) dropwise, then the mixture was stirred at room temperature for 1 h. The mixture was filtered, the filter cake was washed with water and ethanol and dried under vacuum to give 6-amino-3-methyl-5- nitrosopyrimidine-2,4(l H,3H)-dione as violet solid, which was used directly in the next step . Step 3 5,6-diamin -3-methylpyrimidine-2,4(lH,3H)-dione
Figure imgf000375_0002
To a solution of 6-amino-3-methyl-5-nitrosopyrimidine-2,4(lH>3H)-dione in ammonium hydroxide
(100 mL) was added sodium hydrosulfite (6.8 g, 39.1mmol) in small portions at 70 °C, the mixture was stirred at 70 °C for 1 h. The mixture was concentrated and ice-water was added. The slurry was filtered and the filter cake was washed with water and ethanol and dried under vacuum to give 5,6-diamino-3-methylpyrimidine-2,4(lH,3H)-dione (2.0 g, 30.1 % yield over two steps) as yellow solid, LCMS MH+ 157. Step 4 6-amino-5-(4-chlorobenzylamino)-3-methylpyrimidine-2,4(lH,3H)-dione
Figure imgf000375_0003
To a solution of 5,6-diamino-3-methylpyrimidine-2,4(lH,3H)-dione (0.8 g, 5.13 mmol) in water (10 mL) and acetic acid (0.5 mL) was added 4-chlorobenzaldehyde(0.72 g, 5.14 mmol), the mixture was stirred at room temperature for 5 h. The mixture was cooled to 0 °C, sodium cyanoborohydride (0.3 g, 4.76 mmol) was added and the resulting mixture was stirred at room temperature overnight. The mixture was concentrated and filtered, and the filter cake was washed with methanol, dried in vacuo to give 6-amino-5-(4-chlorobenzylamino)-3- methylpyrimidine-2,4(l H53H)-dione (0.6 g, 42.9% yield) as yellow solid. LCMS MH+
Step 5 6-amino-5-(4-chlorobenzylamino)-3-methylpyrimidine-2,4(l H,3H)-dione
Figure imgf000376_0001
To a solution of 6-amino-5-(4-chlorobenzylamino)-3-methylpyrimidine-2,4(lH,3H)-dione (0.56 g, 2.0 mmol) in toluene (10 mL) was added trimethyl orthoformate (20 n L, 183 mmol), the mixture was stirred at 1 10 °C for 5 h. Then the mixture was concentrated and filtered, the filter cake was washed with diethyl ether and dried under vacuum to give 6-amino-5-(4- chlorobenzylamino)-3-methylpyrimidine-2,4(lH,3H)-dione (0.5 g, 86.2% yield) as light-yellow solid. Mass spec: 291 (M+H).
Step 6 8-chloro-7-(4-chl dione
Figure imgf000376_0002
To a solution of 6-amino-5-(4-chlorobenzylamino)-3-methylpyrimidine-2,4(lH,3H)-dione (0.5 g, 1.72mmol) in THF (20 ml) was added NCS (0.3 g, 2.26 mmol), the mixture was stirred at room temperature for 16 h. Then the mixture was concentrated, diluted with ethyl acetate and water, and the phases were separated. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give product 8-chloro-7-(4-chlorobenzyl)-l-methyl- lH-purine-2,6(3H,7H)-dione (500 mg, 89.3% yield) as light yellow solid. LCMS MH+ 325.
Intermediate 11 8-bromo-7-(4-chlorobenzyl)-3-methyl-l -(2-(2-(tetrahydro-2H-pyran-2- yloxy)ethoxy)ethyl)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000376_0003
To a solution of 8-bromo-7-(4-chlorobenzyl)-3-methyl-lH-purine-2,6(3H,7H)-dione (0.2 g, 0.54 mmol, intermediate 8) in DMF (3 mL) were added 2-(2-bromoethoxy)tetrahydro-2H-pyran (169 mg, 0.81 mmol, intermediate 1), potassium carbonate (150 mg, 1 .08 mmol) and a catalytic amount of TBAI .The mixture was stirred at 50°C overnight. It was partitioned between ethyl acetate and water. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give crude product, which was washed with ethanol and dried under vacuum to give 8-bromo-7-(4-chlorobenzyl)-3-methyl- 1 -(2-(tetrahydro-2H-pyran-2-yloxy)ethyl)- 1 H- purine-2,6(3H,7H)-dione(260 mg, 88.8%) as white solid. LCMS retention time 1.712 min, LCMS MH+-THP 459.
Intermediate 12 1 -(2-(2-hydiOxyethoxy)ethyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)- urine- -
Figure imgf000377_0001
Step 1 8-bromo-3-methyl-7-((2-(trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H,7H)-dione
Figure imgf000377_0002
To a solution of 8-bromo-3-methyl-lH-purine-2,6(3H,7H)-dione (3.5 g, 14.3 mmol, intermediate 8 step 1) in DMF (30 mL) was added potassium carbonate (3.9 g, 28.6 mmol). Then 2- (trimethylsilyl)ethoxymethyl chloride (2.37 g, 14.3 mmol) was added dropwise 0 °C , it was stirred room temperature for 3 h. The mixture was partitioned between ethyl acetate and water. The organic layers were combined, dried over sodium sulfate, filtered and concentrated to give a crude product which was washed with ethanol and dried in vacuo to give 8-bromo-3-methyl-7- ((2-(trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H,7H)-dione (4.0 g, 74.9%) as a white solid. LCMS retention time 1.458 min, LCMS MH+ 377.
Step 2 3 -methyl -8-(3 -(tri fluoromethoxy)phenoxy)-7-((2-(trimethy 1 si ly l)ethoxy)methyl)- 1 H- purine-2,6(3H,7H)-dione
Figure imgf000378_0001
To a solution of 8-bromo-3-methyl-7-((2-(trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H,7H)- dione (2.5 g, 6.65 mmol) in DMF (30 mL) were added 3-(trifluoromethoxy)phenol (1.78 g, 9.97 mmol) and potassium carbonate (3.9 g, 28.6 mmol). It was heated at 80 °C overnight. The mixture was cooled, partitioned between ethyl acetate and water. The organic layers were combined, dried over sodium sulfate, filtered and concentrated to give a crude product which was washed with ethanol and dried in vacuo to give 3-methyl-8-(3-(trifluoromethoxy)phenoxy)- 7-((2-(trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H,7H)-dione (1.7 g, 54.3%) as a white solid. LCMS retention time 1.850 min, LCMS MH+ 473.
Step 3 3-methyl-l-(2-(2-(tetrahydro-2H-pyran-2-yloxy)ethoxy)ethyl)-8-(3- (trifluoromethoxy)phenoxy)-7-((2-(trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H,7H)-dione
Figure imgf000378_0002
To a solution of 3-methyl-8-(3-(trifluoromethoxy)phenoxy)-7-((2-
(trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H,7H)-dione (1.2 g, 2.54 mmol) in DMF (15 mL) were added 2-(2-(2-chloroethoxy)ethoxy)tetrahydro-2H-pyran (0.79 g, 3.82 mmol, intermediate 1), potassium carbonate (0.7 g, 5.08mmol), TBAI (2 mg, 0.02 mmol).It was heated at 80 °C overnight. The mixture was cooled, partitioned between ethyl acetate and water. The organic layers were combined, dried over sodium sulfate, filtered and concentrated to give a crude product (1.6 g, 98.1 %) as a yellow oil which was used without purification. LCMS retention time 2.179min, LCMS MH+-THP 561 .
Step 4 1 -(2-(2-hydroxyethoxy)ethyl)-3-methyl-8-(3-(trifluoromethoxy)phenox
purine-2,6(3H,7H)-dione
Figure imgf000379_0001
To a solution of 3-methyl-l -(2-(2-(tetrahydro-2H-pyran-2-yloxy)ethoxy)ethyl)-8-(3- (trifluoromethoxy)phenoxy)-7-((2-(trimethylsilyl)ethoxy)methyl)-l l-l-purine-2,6(3H,7H)- dione(0.64 g, 1.16 mmol,) in ethanol (1 mL) was added HC1 (3 mL); then it was refluxed overinight. The solvent was concentrated to give a crude product which was purified by a column chromatography to give l -(2-(2-hydroxyethoxy)ethyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H37H)-dione (420 mg, 84% yield) as a white solid. LCMS retention time 1.209 min, LCMS MH+ 431. Intermediate 13 8-bromo-7-(4-chlorobenzyl)-3-methyl-l -(2-(tetrahydro-2H-pyran-2- yloxy)ethyl)-lH-purine-2,6(3H,7H)-dione
Figure imgf000379_0002
To a solution of 8-bromo-7-(4-chlorobenzyl)-3-methyl-lH-purine-2,6(3H,7H)-dione (2.0 g, 5.4 mmol, intermediate 8) in DMF(10 mL) was added 2-(2-bromoethoxy)tetrahydro-2H-pyran (1.35 g, 6.5 mmol, intermediate 2), potassium carbonate (1.49 g, 10.8 mmol).The mixture was stirred at 50°C overnight. It was partitioned between ethyl acetate and water. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give a crude product which was washed with ethanol and dried under vacuum to give 8-bromo-7-(4-chlorobenzyl)-3- methyl-l-(2-(tetrahydro-2H-pyran-2-yloxy)ethyl)-lH-purine-2,6(3H,7H)-dione (2.10 g, 74.4%) as a white solid. LCMS retention time 1.669 min LCMS MH+-THP 415. Intermediate 14 8-bromo-7-(4-chlorobenzyl)-3-methyl-l -(3-(tetrahydro-2H-pyran-2- yloxy)propyl)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000380_0001
To a solution of 8-bromo-7-(4-chlorobenzyl)-3-methyl-l H-purine-2,6(3H,7H)-dione(0.5 g, 1.35 mmol, intermediate 8) in DMF (5 mL) was added 2-(3-bromopropoxy)tetrahydro-2H-pyran(0.45 g, 2.03 mmol, intermediate 3), potassium carbonate (0.37 g, 2.7 mmol).The mixture was stirred at 50°C for 3 h; then it was partitioned between ethyl acetate and water. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give a crude product which was washed with ethanol and dried under vacuum to give 8-bromo-7-(4-chlorobenzyl)-3- methyl-l-(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-lH-purine-2,6(3H,7H)-dione(0.5 g, 72.2%) as a white solid. LCMS retention time 1.732 min, LCMS MBT^-THP 429. Intermediate 15 l-(3-hydroxypropyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine- 2,6(3H,7H)-dione
Figure imgf000380_0002
Step 1 3-methyl-l-(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-8-(3-(trifluoromethoxy)phenoxy)- 7-((2-(trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H,7H)-dione
Figure imgf000380_0003
To a solution of 3-methyl-8-(3-(trifluoromethoxy)phenoxy)-7-((2-
(tnmethylsilyl)ethoxy)rnethyl)-l H-purine-2,6(3H,7H)-dione (1.0 g, 2.12 mmol, intermediate 12, step 2) in DMF (10 mL) was added 2-(3-bromopropoxy)tetrahydro-2H-pyran (0.57 g, 2.54 mmol, intermediate 3) and potassium carbonate (0.88 g, 6.36 mmol). The reaction was heated at 50 °C for 3 h. The mixture was cooled, partitioned between ethyl acetate and water. The organic layers were combined, dried over sodium sulfate, filtered and concentrated to give a crude product which was purified by column chromatography eluting with petroleum ether/ethyl acetate (1 : 3) to give 3-methyl-l -(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-8-(3- (trifluoromethoxy)phenoxy)-7-((2-(trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H,7H)-dione (0.9 g, 70.0 % yield) as a white solid. LCMS retention time 2.270 min, LCMS MH+-THP 531.
Step 2 1 -(3-hydroxypropyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)- dione
Figure imgf000381_0001
To a solution of 3-methyl- 1 -(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-8-(3-
(trifluoromethoxy)phenoxy)-7-((2-(trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H,7H)-dione (0.9 g, 1.5 mmol) in ethanol (10 mL) was added HCl (2 mL), then it was refluxed overnight. The solvent was concentrated to give a crude product which was collected, washed with ethanol and dried in vacuo to give l -(3-hydroxypropyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH- purine-2,6(3H,7H)-dione (0.5 g, 80% yield) as a white solid. 'H-NMR (DMSO-dd) δ 13.52 (br, IH), 7.55-7.59(t, IH), 7.47 (s, IH), 7.36-7.38 (d, IH), 7.28-7.30 (d, IH), 4.45 (br, IH), 3.89-3.93 (t, 2H), 3.39-3.43 (t, 2H), 3.42 (s, 3H), 1.66-1.69 (m, 2H). LCMS retention time 1.308 min, LCMS MH+ 401.
Intermediate 16 8-bromo-l-(3-hydroxypropyl)-3-methyl-7-((5-methylthiazol-2-yl)methyl)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000382_0001
Step 1 8-bromo-3.-methyl- 1 -(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-7-((2
(trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H,7H)-dione
Figure imgf000382_0002
To a solution of 8-bromo-3-methyl-7-((2-(trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H,7H)- dione(4.0 g, 10.6 mmol, intermediate 12 step 1) in DMF (50 mL) was added 2-(3- bromopropoxy)tetrahydro-2H-pyran (2.6 g, 1 1.6 mmol, intermediate 3), potassium carbonate (2.4 g, 17.4 mmol). It was heated at 60 °C overnight. The mixture was cooled, partitioned between ethyl acetate and water. The organic layers were combined, dried over sodium sulfate, filtered and concentrated to give a crude product (5.2 g, 94.5 %) as a yellow oil which was used directly for the next step had LCMS MH+ 18.
Step 2 8-bromo-l -(3-hydroxypropyl)-3-methyl-lH-purine-2,6(3H,7H)-dione
Figure imgf000382_0003
To a solution of 8-bromo-3 -methyl- 1 -(3 -(tetrahydro-2H-pyran-2-yloxy)propyl)-7-((2
(trimethylsilyl)ethoxy)methyl)-l H-purine-2,6(3H,7H)-dione (4.2 g, 8.12 mmol) in ethanol (30 mL) was added HCl (6 mL). The mixture was refluxed overnight. The solvent was removed to give a crude product which was washed with ethanol and dried in vacuo to 8-bromo-l-(3- hydroxypropyl)-3-methyl-lH-purine-2,6(3H,7H)-dione(2.0 mg, 81.3 % yield) as a white solid. LCMS MH+ 303. Step 3 8-bromo-l -(3-hydroxypropyl)-3-methyl-7-((5-methylthiazol-2-yl)methyl)-l H-purine- 2,6(3H,7H)-dione
Figure imgf000383_0001
To a solution of 8-bromo-l-(3-hydroxypropyl)-3-methyl-l H-purine-2,6(3H,7H)-dione (0.5 g, 1.65 mmol) in DMF (50 mL) was added 2-(chloromethyl)-5-methylthiazole (0.27 g, 1.81 mmol, intermediate 4), potassium carbonate (0.34 g, 2.48 mmol) and TBAI (2 mg, 0.02 mmol). The mixture was heated at 60 °C for 3 h. The mixture was cooled; then partitioned between ethyl acetate and water. The organic layers were combined, dried over sodium sulfate, filtered and concentrated. The residue was purified by a column chromatography eluting with
DCM/methanol(60: l to 30: 1 ) to give 8-bromo-l-(3-hydroxypropyl)-3-methyl-7-((5- methylthiazol-2-yl)methyl)-lH-purine-2,6(3H,7H)-dione (0.4 g, 58.4 %) as a yellow oil which was used without further purification. LCMS MH+ 416.
Intermediate 17 3-methyl-l -(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-8-(2-(3- (trifluoromethoxy)phenoxy)ethoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000383_0002
Step 1 3-methyl-l-(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-8-(2-(3-
(trifluoromethoxy)phenoxy)ethoxy)-7-((2-(trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H,7H)- dione
Figure imgf000383_0003
To a solution of 2-(3-(trifluoromethoxy)phenoxy)ethanol (0.64 g, 2.9 mmol, intermediate 5) in THF (15 mL) was added sodium hydride (0.23 g, 9.67 mmol) at 0 °C. it was stirred at 0 °C for 30 min, then 8-bromo-3-methyl-l-(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-7-((2- (trimethylsilyl)ethoxy)methy])- lH-purine-2,6(3H,7H)-dione (1 .0 g, 1.93 mmol, intermediate 16 step 1) was added. The mixture was stirred 16 h. The reaction was quenched with aq. ammonium chloride (2 mL) at 0 °C; then it was partitioned between ethyl acetate and water. The organic phases were combined, dried over sodium sulfate, filtered and concentrated to give a crude product which was used without purification. LCMS retention time 2.144; LCMS MH+-THP 575.
Step 2 3-methyl-l -(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-8-(2-(3- (trifluoromethoxy)phenoxy)ethoxy)- 1 H-purine-2,6(3 H,7H)-dione
Figure imgf000384_0001
To a solution of 3-methyl-l-(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-8-(2-(3- (trifluoromethoxy)phenoxy)ethoxy)-7-((2-(trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3HJ7H)- dione (1 .2 g, 1.82 mmol) in THF (15 mL) was added TBAF (3.7 mL, 3.64 mmol). The mixture was stirred at 80°C overnight. Then it was cooled, partitioned between ethyl acetate and water. The organic layers were combined, washed with water, dried over sodium sulfate, filtered and concentrated to give a crude product which was purified by column chromatography eluting with petroleum ether and ethyl acetate(2: l) to give 3-methyl-l-(3-(tetrahydro-2H-pyran-2- yloxy)propyl)-8-(2-(3-(trifluoromethoxy)phenoxy)ethoxy)-lH-purine-2,6(3H,7H)-dione(900 mg, 93.7% yield) as a white solid. LCMS retention time 1.613; LCMS MH+-THP 445.
Intermediate 18 7-benzyl-8-chloro-l-(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-3-((2-
Figure imgf000384_0002
Step 1 2-amino-7-benzyl-lH-purin-6(7H)-one
Figure imgf000385_0001
To a solution of 2-amino-9-((2R,3S,4R,5S)-3,4-dihydroxy-5-(hydroxymethyl)tetrahydrofuran-2- yl)-l H-purin-6(9H)-one ( 14 g,49.47 mmol) in DMSO (50 mL) was added
(bromomethyl)benzene (10 g, 59.36 mmol). The mixture was stirred at 50 °C overnight. Then the mixture was cooled to room temperature and HCI (50 mL, 10% w.w) was added and the mixture was stirred at 70 °C for 2 h. The reaction was cooled to room temperature, filtered and the filter cake was washed with water and ethanol and dried under vacuum to give 2-amino-7-benzyl-l H- purin-6(7H)-one (10 g, 74.2% yield) as grey solid. LCMS retention time 0.529 min; LCMS MH+ 242.
Step 2 2-amino-7-benzyl-lH-purin-6(7H)-one
Figure imgf000385_0002
To a solution of 2-amino-7-benzyl-l H-purin-6(7H)-one (6.8 g, 28.33 mmol) in acetic acid (80 mL) and water (10 mL) was added a solution of sodium nitrite (1 .95 g, 28.26 mmol) in water (10 mL) at 50 °C dropwise, the mixture was stirred at 50 °C for 1 h. Then the mixture was cooled to room temperature and stirred for another 1.5 h. The mixture was filtered; then the filter cake was washed with water and ethanol, dried in vacuo to give 2-amino-7-benzyl-lH-purin- 6(7H)-one (6.5 g, 94.9% yield) as yellow solid. LCMS retention time 0.502 min; LCMS MH+ 243.
Step 3 7-benzyl- -((2-(trimethylsilyl)ethoxy)methyl)-l H-purine-2,6(3H,7H)-dione
Figure imgf000385_0003
To a solution of 2-amino-7-benzyl-l H-purin-6(7H)-one (6.5 g, 26.86mmol) in DMF(50 mL) was added potassium carbonate (5.6 g, 40.58 mmol), followed by 2-(trimethylsilyl)ethoxymethyl chloride (4.45 g, 27.08 mmol) at 0 °C. The resulting mixture was stirred at 60 °C for 16 h. The mixture was diluted with water and filtered. The filter cake was washed with water twice, dried in vacuo to give 7-benzyl-3-((2-(trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H,7H)-dione (10.8 g, 100% yield) as yellow oil. LCMS retention time 1.523 min; LCMS MH+-58 315.
Step 4 7-benzyl-8-chloro-3-((2-(trimethylsilyl)ethoxy)methyl)-l H-purine-2,6(3H,7H)-dione
Figure imgf000386_0001
To a solution of 7-benzyl-3-((2-(trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H,7H)-dione (6.0 g, 16.13 mmol) in DMF (50 mL) was added NCS (3.23 g, 24.19 mmol) and the resulting mixture was stirred at room temperature for 8 h. The mixture was diluted with ethyl acetate and water, and the phases were separated. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give 7-benzyl-8-chloro-3-((2-(trimethylsilyl)ethoxy)methyl)- lH-purine-2,6(3H,7H)-dione (5.1 g, 77.9% yield) as light yellow oil. Product tic [petroleum ether/ethyl acetate (15: 1) iodine detection, Rf 0.6)
Step 5 7-benzyl-8-chloro-l-(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-3-((2- (trimethyls
Figure imgf000386_0002
To a solution of 7-benzyl-8-chloro-3-((2-(trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H,7H)- dione (5 g, 12.32 mmol) in DMF (30 mL) was added 2-(3-bromopropoxy)tetrahydro-2H-pyran (3.60 g, 16.22 mmol, intermediate 14 step 1), followed by potassium carbonate (3.4 g, 24.64 mmol). The mixture was stirred at 65 °C overnight. The mixture was diluted with ethyl acetate and water, and the phases were separated. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give 7-benzyl-8-chloro-l -(3-(tetrahydro-2FI-pyran-2- yloxy)propyl)-3-((2-(trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H,7H)-dione (6.3 g, 93.3% yield) as yellow oil. LCMS retention time 3.574 min; LCMS MNa+ 571 . Intermediate 19 l -(4-(bromomethyl)phenyl)ethanone
Figure imgf000387_0001
To a solution ofl-p-tolylethanone (600 mg, 4.47 mmol) in carbon tetrachloride (15 mL) was added N-bromosuccinimide (955 mg, 5.37 mmol) and BPO (31 mg, 0.13 mmol). The mixture was refluxed for 3 h; then it was cooled , and filtered. The filtrate was concentrated to give l -(4- (bromomethyl)phenyl)ethanone (683 mg, 71%) as a brown oil. LCMS MH+ 213.
Intermediate 20 2-(3-(4-methyloxazol-2-yl) henyl.)acetic acid
Figure imgf000387_0002
Step 1 2-(3-(4-methyloxazol-2-yl)phenyl)acetonitrile
Figure imgf000387_0003
A solution of 3-(cyanomethyl)benzamide (200 mg, 1.25 mmol) in l-chloropropan-2-one (2 mL) was microwave irradiated at 120 °C for 20 min in a sealed tube. The mixture was concentrated to give crude product (1 10 mg), which was directly used to the next reaction without purification. LCMS MH+ 199.
2-(3-(4-meth loxazol-2- l hen l acetic acid
Figure imgf000387_0004
To a solution of 2-(3-(4-methyloxazol-2-yl)phenyl)acetonitrile (110 mg, 0.555 mmol) in 1 ,4- dioxane (3 mL) was added 6N HC1 (1 mL). Then the mixture was stirred at 80 °C for 2 h. The mixture was concentrated and purified via silica gel chromatography eluted with DCM/ methanol (20 :1 ) to give 2-(3-(4-methyloxazol-2-yl)phenyl)acetic acid (85 mg, 70.5%) as yellow syrup. LCMS retention time 0.920min, LCMS MH+ 218.
Intermediate 21 l-(4-chlorophenyl)ethyl methanesulfonate
Figure imgf000388_0001
The title compound was prepared using the method of intermediate 34 to givel -(4- chlorophenyl)ethyl methanesulfonate (258 mg, 23.6% yield) as yellow oil, which was used to the next reaction without purification.
Intermediate 22 2-(bromometh l)-5-methylthiazole
Figure imgf000388_0002
To a solution of 2,5-dimethylthiazole (200 mg, 1.77 mmol) in carbon tetrachloride (5 mL) was added N BS (377 mg, 2.12 mmol) , followed by BPO (20 mg, 0.083 mmol). Then the mixture was heated to reflux for 4 h with stirring. The mixture was cooled to room temperature and filtered, the filtrate was concentrated to give crude product 2-(bromomethyl)-5-methylthiazole (260 mg, 76.9%) as yellow oil, which was used without purification.
Intermediate 23 2-(chloromethyl)thiazole
Step 1 thiazol-2-ylmethanol
Figure imgf000388_0003
To a solution of n-BuLi (8.4 ml , 1.6 mol/1 , 13.4 mmol) in THF (30 mL) was added 2- bromothiazole (377 mg, 2.12 mmol) dropwise under nitrogen atmosphere at -70 °C, and the mixture was stirred at the temperature for 1 h. Then DMF (1.4 ml, 18.3 mmol) was added into the solution dropwise under nitrogen atmosphere at -70 °C. The resulting mixture was stirred at the temperature for 1 h. Then the mixture was quenched with aqueous saturated ammonium chloride, diluted with ethyl acetate and water, and the phases were separated. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give yellow oil. The yellow oil was dissolved in methanol (15 ml) , cooled to -60 °C, ans sodium borohydride (463 mg , 12.2 mmol) was added portionwise under nitrogen atmosphere. The mixture was stirred at the temperature for 1 h. The reaction was quenched with acetone and concentrated. The residue was diluted with ethyl acetate and water, and the phases were separated. The organic layer was dried over sodium sulfate, filtered and concentrated, then purified by silica gel chromatography eluting with petroleum/ ethyl acetate= 3: 1 to give thiazol-2-ylmethanol (230 mg, 16.4 % yield) as brown oil . LCMS MH+ 1 16. Step 2 2-(chloromethyl)thiazole
Figure imgf000389_0001
To a solution of thiazol-2-ylmethanol (230 mg, 2.0 mmol)in DCM (5 mL) was added thionyl chloride (0.19 ml, 2.6 mmol) at 0 °C and the mixture was stirred at room temperature for 1 h. The mixture was diluted with DCM and water, and the phases were separated. The organic layer was dried over sodium sulfate, filtered and concentrated to give 2-(chloromethyl)thiazole (240 mg, crude), which was used without purification.
Intermediate 24 5-(chloromethyl)-2-methylpyrimidine Step 1 (2-methylpyrimidin-5-yl)methanol
Figure imgf000389_0002
To a solution of 2-methylpyrimidine-5-carboxylic acid (150 mg, 1.23 mmOl) in ethanol (5 mL) was added sodium borohydride(93 mg, 2.46 mmol). The mixture was stirred at room temperature for 3 h. It was quenched with aqueous HC1 (2 N, 2 mL), extracted with DCM, dried over sodium sulfate, filtered and concentrated give the yellow oil product (2-methylpyrimidin-5-yl)methanol (95 mg, 62.6%). LCMS MH+ 125.
5-(chloromethyl)-2-methylpyrimidine
Figure imgf000389_0003
A solution of (2-methylpyrimidin-5-yl)methanol (95 mg, 0.77 mmol) in thionyl chloride(l was stirred at room temperature for 1 h. The mixture was concentrated to dryness and used without purification. LCMS MH+ 143. Int rmediate 25 3- -methyloxazol-2-yl)phenol
Figure imgf000390_0001
Step 1 2-(3-methoxyphen l -4-meth loxazole
Figure imgf000390_0002
To a solution of 3-methoxybenzamide (1.5 g, 9.9 mmol) in toluene (15 mL) was added 1- chloropropan-2-one (1.37 g, 14.9 mmol), and the mixture was stirred at reflux for 16 h. The mixture was cooled and concentrated to give crude product, which was purified via silica gel chromatography eluted with ethyl acetate/petroleum ether (1 : 5) to give 2-(3-methoxyphenyl)-4- methyloxazole (1.21 g, 64.6% yield) as yellow syrup. LCMS MH+ 190.
Step 2 3-(4-methyloxazol- -yl)phenol
Figure imgf000390_0003
A solution of 2-(3-methoxyphenyl)-4-methyloxazole (1.1 g, 5.79 mmol) in aqueous hydrogen bromide (10 mL, 48% w/w) was heated to 100 °C for 16 h. The mixture was concentrated to give 3-(4-methyloxazol-2-yl)phenol (0.97 g, crude), which was used without purification. LCMS MH+ 176. ne
Figure imgf000390_0004
Step 1 7-benzyl-8-chloro-l -(3-hydroxypropyl)-lH-purine-2,6(3H,7H)-dione
Figure imgf000391_0001
To a solution of 7-benzyl-8-chloro-l -(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-3-((2- (trimethylsilyl)ethoxy)methyl)-l H-purine-256(3H,7H)-dione (500 mg,0.912 mmol, intermediate 18) in ethyl alcohol (20 mL) was added concentrated HC1 (5 mL). The mixture was stirred at 80 °C overnight. Then the mixture was concentrated, neutralized with saturated sodium bicarbonate and extracted with ethyl acetate. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give 7-benzyl-8-chloro-l-(3-hydroxypropyl)-l H-purine- 2,6(3 H,7H)-dione (280 mg, 91.9% yield) as yellow oil. LCMS retention time 1.710 min; LCMS MH+ 335.
Step 2 7-benzyl-8-chloro-3-ethyl-l -(3-hydroxypropyl)-lH-purine-2,6(3H,7H)-dione
Figure imgf000391_0002
To a solution of 7-benzyl-8-chloro-l-(3-hydroxypropyl)-l H-purine-2,6(3H,7H)-dione (150 mg, 0.449 mmol) in DMF (5 mL) was added iodoethane (0.16 g, 0.13 mmol), followed by potassium carbonate (0.4 g, 2.90 mmol). The mixture was stirred at 40 °C for 4 h. The reaction was cooled and diluted with ethyl acetate. The organic phase was washed with water and brine; then it was dried over sodium sulfate, filtered and concentrated to give 7-benzyl-8-chloro-3-ethyl-l-(3- hydroxypropyl)-lH-purine-2,6(3H,7H)-dione (0.15 g, 92.3% yield) as yellow oil. LCMS retention time 1.296 min; LCMS MH+ 363.
Step 3 7-benzyl- -ethoxy-3 -ethyl- 1 -(3 -hydroxypropyl)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000391_0003
A solution of sodium (50 mg, 2.17 mmol) in ethyl alcohol (10 mL) was stirred at 40 °C under nitrogen until the sodium was consumed. Then 7-benzyl-8-chloro-3-ethyl-l -(3-hydroxypropyl)- l H-purine-2,6(3H,7H)-dione (150 mg, 0.414 mmol) was added at 0 °C and the mixture was stirred at 40 °C for 4 h. The mixture was quenched with ice-water (15 mL) and concentrated. The aqueous residue was extracted with ethyl acetate. The organic phase was dried over sodium sulfate, filtered and concentrated to give 7-benzyl-8-ethoxy-3-ethyl-l -(3-hydroxypropyl)-l H- purine-2,6(3H,7H)-dione (120 mg, 77.9% yield) as yellow solid.LCMS retention time 1.488 min; LCMS MH+ 373.
Step 4 8-ethox -3-ethy]-l -(3-hydroxypropyl)-lH-purine-2,6(3H,7H)-dione
Figure imgf000392_0001
7-benzyl-8-ethoxy-3-ethyl-l-(3-hydroxypropyl)-l H-purine-2,6(3H,7H)-dione (0.12 g, 0.323 mmol) was dissolved in ethanol (20 mL) and the mixture was degassed and refilled with nitrogen three times. Ammonium formate (0.5 g, 7.94 mmol) and 10% Pd/C (30 mg) were added. The mixture was again degassed and refilled with nitrogen three times; then it was warmed to 80 °C and stirred overnight. The mixture was cooled and filtered, and the filter cake was washed with methanol. The filtrate was concentrated to give 8-ethoxy-3-ethyl-l -(3-hydroxypropyl)-lH- purine-2,6(3H,7H)-dione (70 mg, 77.1%> yield) as white solid. LCMS retention time 0.413 min; LCMS MH+ 283.
Intermediate 27 3-(morpholinomethyl)phenol
Figure imgf000392_0002
To a solution of 3-hydroxybenzaldehyde (1 g, 8.19 mmol) in methanol (15 mL) was added morpholine (1.42 g, 16.4 mmol). Then the mixture was stirred at room temperature for 1 h. The mixture was cooled to -5 °C, and sodium borohydride (403 mg, 10.6 mmol) was added in small portions. The resulting mixture was stirred at room temperature for 4 h. The mixture was quenched with diluted hydrochloride acid and concentrated. The aqueous solution was washed with ethyl acetate and then made basic by addition of ammonium hydroxide. The basic aqueous slurry was extracted with ethyl acetate and this organic layer was dried and concentrated to give 3-(morpholinomethyl)phenol (0.76 g, 48.2%) as white solid. LCMS retention time 0.871 min, LCMS MH+ 194.
Intermediate 28a and 28b 2-chloro-5-hydroxybenzaldehyde (28a) and 4-chloro-3- hydroxybenzaldehyde (
Figure imgf000393_0001
To a solution of 3-hydroxybenzaldehyde (1 g, 10 mmol) in acetonitrile (50 mL) was added p- toluenesulfonic acid (3.4 g, 20 mmol) portionwise. The mixture was stirred at room temperature for 5 min, and NCS (1.33 g, 10 mmol) was added and the resulting mixture was stirred at room temperature for 2 h. The mixture was quenched with aqueous sodium thiosulfate, and diluted with ethyl acetate and brine. The organic layer was separated, dried, and concentrated to give the crude products which were purified by silica gel chromatography eluting with petroleum ether /ethyl acetate (10: 1 to 5: 1) to give 2-chloro-5-hydroxybenzaldehyde (340 mg, 21.7% yield intermediate 28a) as yellow solid; 'H-NMR (CDC13) δ 10.43 (s, 1H), 7.54-7.56 (d, 1H), 7.30- 7.37 (m, 2H) and 4-chloro-3-hydroxybenzaldehyde (310 mg, 19.7% yield, intermediate 28b) as yellow solid. ^- MR (CDC13) δ 10.43 (s, 1H), 7.40 (d, 1H), 7.33 (d, 1H), 7.06-7.09 (dd, 1H).
Intermediate 29 4-chloro-3-(morpholinomethyl)phenol
Figure imgf000393_0002
To a mixture of 2-chloro-5-hydroxybenzaldehyde (200 mg, 1.27 mmol, intermediate 28a) and morpholine (280 mg, 3.21 mmol) in methanol (10 mL) was added 2 drops of acetic acid and the mixture was stirred at room temperature for 2 h. To the mixture, sodium borohydride (97 mg, 2.56 mmol ) was added and the resulting mixture was stirred at room temperature for 4 h. The mixture was quenched with dilute hydrochloride acid and concentrated to give crude product. This crude material was purified by silica gel chromatography eluting with petroleum ether/ethyl acetate (5:1 to 1 : 1) to give 4-chloro-3-(morpholinomethyl)phenol (150 mg, 51.8% yield) as white solid. LCMS retention time 0.378; LCMS MH+ 228. Intermediate 30 4-
Figure imgf000394_0001
The title compound was prepared using the method of intermediate 29 starting with intermediate 28b but using sodium triacetoxyborohydride as the reducing agent to give 4-chloro-3- (morpholinomethyl)phenol (190 mg, 68.1% yield) as yellow solid. LCMS retention time 0.347; LCMS MH+ 228.
Intermediate -(chloromethyl)-5-methylisoxazole
Figure imgf000394_0002
(5 -methylisoxazol-3 -yl)methanol
Figure imgf000394_0003
To a solution of ethyl 5-methylisoxazole-3-carboxylate (500 mg, 3.22 mmol) in ethanol (8 mL) was added sodium borohydride (244 mg, 6.44 mmol) in portions at 0 °C. After addition, the mixture was stirred at room temperature for 16 h. The mixture was quenched with diluted hydrochloride acid. The mixture was extracted with ethyl acetate. The organic phase was washed with brine, dried and concentrated to give (5 -methylisoxazol-3 -yl)methanol (370 mg, 100% yield) as yellow oil, which was used directly in the next reaction without purification. LCMS retention time 0.391 min; LCMS MH+ 1 14. Step 2 3-(chIoromethyl)-5-methylisoxazole
P-N SOCI, P-N
DC
To a solution of (5-methylisoxazol-3-yl)methanol (370 mg, 3.27 mmol) in DCM (5 mL) was added thionyl chloride (5 mL) dropwise. The resulting mixture was stirred at room temperature for 16 h. The mixture was concentrated to give 3-(chloromethyl)-5-methylisoxazole (350 mg, crude) as brown oil, which was used without purification. LCMS retention time 0.768 min; LCMS MH+ 132.
Intermediate 32 2-(2-ethyl-l ,3-dioxolan-2-yl)ethyl methanesulfonate
Figure imgf000395_0001
Step 1 methyl 2-(2-ethyl-l,3-dioxolan-2-yl)acetate
Figure imgf000395_0002
To a solution of methyl 3-oxopentanoate (3 g, 23.1 mmol) in toluene (30 mL) was added ethane- 1,2-diol (3 mL, 70.2 mmol) and p-toluenesulfonic acid (500 mg, 2.91 mmol), the resulting mixture was stirred at 120 °C overnight. Then the mixture was concentrated and partitioned between ethyl acetate and aqueous saturated sodium bicarbonate solution. The phases were separated and the aqueous layer was extracted with ethyl acetate. The combined organic phase was washed with brine and dried over sodium sulfate; then it was filtered and concentrated to give methyl 2-(2-ethyl-l,3-dioxolan-2-yl)acetate (1.7 g, 42.5%) as yellow oil which was used without purification. Product tic [petroleum ether/ethyl acetate (15: 1) developed with iodine, Rf 0.6]
Step 2 2-(2-ethyl-l ,3-dioxolan-2-yl)ethanol
Figure imgf000395_0003
To a solution of methyl 2-(2-ethyl-l ,3-dioxolan-2-yl)acetate (1 g, 5.75 mmol) in anhydrous THF (40 mL) was added LAH (900 mg, 23.7 mmol) at 0 °C under nitrogen atmosphere, the resulting mixture was stirred at 90 °C overnight. The mixture was cooled to room temperature, and ethyl acetate was added to the mixture. The resulting slurry was filtered and the filtrate was concentrated to dryness. The crude product was purified by silica gel chromatography eluting with DCM/ methanol (100: 1 to 20: 1) to give 2-(2-ethyl-l ,3-dioxolan-2-yl)ethanol (233 mg, 27.4%) as yellow oil that was used without purification. Step 3 2-(2-ethyl-l ,3-dioxolan-2-yl)ethyl methanesulfonate
Figure imgf000396_0001
To a solution of 2-(2-ethyJ-l ,3-dioxoIan-2-yl)ethanol (233 mg, 1.59 mmol) and TEA (327 mg, 3.2 mmol) in DCM (10 mL) was added methanesulfonyl chloride (0.23 ml, 2.92 mmol) at 0 °C. The resulting mixture was stirred at room temperature for 3 h. The mixture was diluted with DCM and water, and the phases were separated. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give 2-(2-ethyl-l ,3-dioxolan-2-yl)ethyl methanesulfonate (500 mg, 100%) as yellow oil that was used without purification. Product tic [DCM/methanol (100: 1 ) developed with potassium permanganate, Rf 0.6]
Intermediate 33 3-hydroxybutyl 4-methylbenzenesulfonate
Figure imgf000396_0002
To a solution of butane- 1 ,3 -diol (1 g, 1 1.1 mmol), TEA (4.6 mL, 33.2 mmol) and DMAP (30 mg, 0.25 mmol) in DCM (20 mL) was added p-toluenesulfonyl chloride (2.75 g, 14.4 mmol) at 0 °C. The resulting mixture was stirred at room temperature for 16 h. The mixture was diluted with DCM and water, and the phases were separated. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give 3-hydroxybutyl 4-methylbenzenesulfonate (3 g, 1 00% yield) as yellow oil, which was used without purification. . Product tic
[DCM/methanol (100: 1) developed with potassium permanganate, Rf 0.4]
Intermediate 34 2-(2-
Figure imgf000396_0003
To a solution of 2-(2-methoxyethoxy)ethanol (300 mg, 2.5 mmol) in DCM (5 mL) was added methanesulfonyl chloride (370 mg, 2.6 mmol) dropwise at 0°C. After stirred at this temperature for 5 min, TEA (505 mg, 5.0 mmol) was added. It was stirred at room temperature overnight. The mixture was partitioned between DCM and water. The organic layers were combined, dried over sodium sulfate, filtered and concentrated to give a crude oil product, which was used without purification. Produt tic [petroleum ether/ethyl acetate [1 : 1) developed with iodine, Rf
0.5] Intermediate 35 3-(3-(trifluoromethoxy)phenoxy)propan-l -ol
Figure imgf000397_0001
To a solution of 3-(trifluoromethoxy)phenol (0.3 g, 1.68 mmol) in DMF (10 mL) was added 3- bromopropan-l-ol (0.47 g, 3.37 mmol) and potassium carbonate (0.47 g, 3.37 mmol). The reaction was heated at 80 °C overnight. The mixture was cooled and partitioned between ethyl acetate and water. The organic layer was dried over sodium sulfate, filtered and concentrated to give a crude product which was purified by column chromatography eluting with petroleum ether and ethyl acetate(10: l) to give 3-(3-(trifluoromethoxy)phenoxy)propan-l -ol (0.32 g, 80.6 % yield) as yellow oil.
Intermediate 36 3-(2-hydroxyethoxy)benzaldehyde
Figure imgf000397_0002
The title compound was prepared using the method of intermediate 5 except the reaction was carried out at 140 °C to give 3-(2-hydroxyethoxy)benzaldehyde (1 .1 g, 73.4% yield) as yellow oil which was used without purification. 'if-NMR (DMSO-</6) δ 9.98(s, 1H), 7.53-7.51(m, 2H), 7.44-7.43(m, 1H), 7.30-7.29(m, 1H), 4.95-4.93(t, 1H), 4.08-4.06(t, 2H), 3.77-3.73(q, 2H)
Intermediate 37 3 -(3 -(trifluoromethoxy)phenyl)propan- 1 -ol
Figure imgf000397_0003
Step 1 3-(3-(trifluoromethoxy)phenyl)prop-2-yn-l-ol
Figure imgf000397_0004
To a solution of l -iodo-3-(trifluoromethoxy)benzene (0.5 g, 1.73 mmol) in toluene (10 mL) was added prop-2-yn-l-ol (0.1 5 g, 2.62 mmol) and morpholine (0.5 mL). Then cuprous iodide (4 mg, 0.02mmol) and bis(triphenylphosphine)palladium(II) chloride (14 mg, 0.02mmol) were added to the mixture under nitrogen. The reaction was heated at 80 °C overnight. The mixture was cooled and partitioned between ethyl acetate and water. The combined organic layer was dried over sodium sulfate, filtered and concentrated to give a crude product which was purified by column chromatography eluting with ethyl acetate/petroleum ether (1 :6 to 1 :3) to give 3-(3- (trifluoromethoxy)phenyl)prop-2-yn-l-ol (0.3 g, 80.2 %) as a yellow oil. LCMS retention time 1.552 min.
Step 2 3-(3-(trifluoromethoxy)phenyl)propan-l-ol
Figure imgf000398_0001
To a solution of 3-(3-(trifluoromethoxy)phenyl)prop-2-yn-l-ol (0.3 g, 1.39 mmol) in THF (10 mL) was added LAH (63 mg, 1.68 mmol) portionwise at 0 °C. The reaction was stirred at 0 °C for 3 h. The reaction was carefully quenched by dropwise addition of ethyl acetate at 0° C and the mixture was partitioned between ethyl acetate and water. The combined organic layer was dried over sodium sulfate, filtered and concentrated to give a crude product which was purified by column chromatography eluting with ethyl acetate/petroleum ether (1 :4 to 1 : 1) to give 3-(3- (trifluoromethoxy)phenyl)propan-l-ol (0.26 g, 85.2 %) as a yellow oil which was used without purification. 'H-NMR (CD3OD) δ 7.37-7.39(d, 1H), 7.31-7.35(m, 1 H), 7.15-7.17 (d, 1H), 4.07- 4.1 l (t, 2H), 2.72-2.76(t, 2H), 1.83-1.90 (m, 2H). -(cyclohexyloxy)ethanol
Figure imgf000398_0002
Step 1 l ,4-dioxaspiro[4.5]decane (JF-000357-022)
Figure imgf000398_0003
(CH2OH)2 \_ o^
To a solution of cyclohexanone (10 g, 0.102 mol) in ethylene glycol (50 mL) was added iodine (3.88 g, 15.3 mmol) and the resulting mixture was stirred at room temperature overnight. The mixture was diluted with ethyl acetate and extracted with saturated aqueous sodium thiosulfate. The organic phase was dried and concentrated to give a crude product, which was purified via silica gel chromatography eluting with ethyl acetate /petroleum ether (1 : 8) to give 1 ,4- dioxaspiro[4.5]decane (6.8 g, 46.9% yield) as light oil. 'H-NMR (CDC13) δ 3.93(s, 4Η), 1.58- 1.59 (d, 8Η), 1.39-1 .41 (m, 2Η).
Step 2 2-(cyclohexyloxy)ethanol
Figure imgf000399_0001
To the slurry of zirconium tetrachloride (10.65 g, 45.7 mmol) in THF (100 mL) was added sodium borohydride (4.3 g, 1 14.3 mmol) in small portions at room temperature. A solution of l,4-dioxaspiro[4.5]decane (6.5 g, 45.7 mmol) in THF (20 mL) was added dropwise and the resulting mixture was stirred at room temperature overnight. The reaction was carefully quenched with IN HC1 and extracted with ethyl acetate. The organic phase was washed with brine, dried and concentrated to give a crude product, which was purified via silica gel chromatography eluting with petroleum ether/ethyl acetate (3: 2) to give 2-
(cyclohexyloxy)ethanol (3.9 g, 59.2% yield) as light yellow oil. 'H-NMR (CDC13) δ 3.68-3.70 (t, 2Η), 3.53-3.56 (m, 2Η), 3.27-3.29 (m, 1Η), 2.54 (br, 1Η), 1.89-1.93 (m, 2Η), 1.70-1.73 (m, 2Η), 1.51-1.54 (m, 1 Η), 1.17-1.28 (m, 5Η).
Intermediate 39 7-(4-chlorobenzyl)-8-(2-fluoro-5-(trifluoromethoxy)benzyl)-3-methyl-lH- purine-2,6(3H,7H)-dione
Figure imgf000399_0002
Step 1 8-(2-fluoro-5-(trifluoromethoxy)benzyl)-l-(4-methoxybenzyl)-3-methyl-lH- purine-2,6(3H,7H)-dione
Figure imgf000399_0003
To a solution of 5,6-diamino-3-(4-methoxybenzyl)-l-methylpyrimidine-2,4(lH,3H)-dione (80 mg, 0.289 mmol, intermediate 59) in ethanol (2 mL) was added 2-(2-fluoro-5- (trifluoromethoxy)phenyl)acetic acid (68.8 mg, 0.289 mmol) followed by EDCI (66.5 mg, 0.347 mmol) and the mixture was stirred at room temperature for 2 h. The reaction was partitioned between ethyl acetate and brine. The organic phase was dried and concentrated. The residue was dissolved in ethanol (3 mL) and 1 mM sodium hydroxide (1 mL) was added. The resulting mixture was stirred at reflux for 3 h. The reaction was concentrated and residue was partitioned between ethyl acetate and brine. The organic layer was dried and concentrated to give a crude solid product, which was triturated with ethanol, collected and dried to afford 8-(2-fluoro-5- (trifiuoromethoxy)benzyl)- l-(4-methoxybenzyl)-3-methyl-l H-purine-2,6(3H,7H)-dione (67 mg, 48.4% yield) as a light yellow solid. LCMS retention time 1.61 1 min; LCMS MH+ 479.
Step 2 7-(4-chlorobenzyl)-8-(2-fluoro-5-(trifluoromethoxy)benzyl)- 1 -(4- methoxybenzyl)-3-
Figure imgf000400_0001
To a solution of 8-(2-fluoro-5-(trifluoromethoxy)benzyl)-l -(4-methoxybenzyl)-3-methyl-lH- purine-2,6(3H,7H)-dione (67 mg, 0.14 mmol) in DMF (3 mL) was added l-(bromomethyl)-4- chlorobenzene (43.2 mg, 0.21 mmol) followed by potassium carbonate (38.7 mg, 0.28 mmol). The reaction was heated to 60 °C for 3 h. The reaction was partitioned between ethyl acetate and brine. The organic phase was dried and concentrated to give a crude product, which was purified by silica gel chromatography eluting with DCM/methanol (60: 1) to give 7-(4-chlorobenzyl)-8- (2-fluoro-5-(trifluoromethoxy)benzyl)-l-(4-methoxybenzyl)-3-methyl-lH-purine-2,6(3H,7H)- dione (71.2 mg, 84.3% yield) as a white solid. LCMS retention time 2.375 min; LCMS MH+ 603.
Step 3 7-(4-chlorobenzyl)-8-(2-fluoro-5-(trifluoromethoxy)benzyl)-3-methyl-l H-purine-
2,6(3H,7H)-dione
Figure imgf000400_0002
To a solution of 7-(4-chlorobenzyl)-8-(2-fluoro-5-(trifluoromethoxy)benzyl)-l -(4- methoxybenzyl)-3-methyl-lH-purine-2,6(3H,7H)-dione (71 mg, 0.1 18 mmol) in DCM (1 mL) was added TFA (0.3 mL) and trifluoromethanesulfonic acid (0.2 mL). The reaction was stirred at room temperature for 16 h. The mixture was concentrated and the residue was partitioned between DCM and aqueous sodium bicarbonate. The organic layer was dried and concentrated to give 7-(4-ch]orobenzyl)-8-(2-fluoro-5-(trifluoromethoxy)benzyl)-3-methyl-lH-purine- 2,6(3H,7H)-dione (39 mg, 68.4% yield) as a yellow syrup, which was directly without purification. LCMS retention time 1.686 min; LCMS ΜΙ- 483.
Intermediate 40 (5-methyloxazol-2-yl)methanol
Figure imgf000401_0001
Step 1 ethyl 2-(2-hydroxypropylamino)-2-oxoacetate
Figure imgf000401_0002
To a mixture of l-aminopropan-2-ol (2 g, 26.6 mmol) and TEA (4.03 g, 39.9 mmol) in DCM (50 mL) was added ethyl 2-chloro-2-oxoacetate (4.36 g, 31.95 mmol) dropwise at 0 °C. The mixture was stirred at room temperature for 16 h. The mixture was diluted with DCM and extracted with brine. The organic layer was dried and concentrated to give a crude product which was purified by silica gel chromatography eluting with petroleum ether/ethyl acetate (2: 1) to give ethyl 2-(2-hydroxypropylamino)-2-oxoacetate (2.19 g, 51% yield) as yellow syrup.
LCMS retention time 0.348 min, LCMS MH+ 176.
Step 2 ethyl 2-oxo-2-(2-oxopropylamino)acetate
Figure imgf000401_0003
To a solution of ethyl 2-(2-hydroxypropylamino)-2-oxoacetate (2.1 g, 13 mmol) in DCM (30 mL) was added Dess-Martin periodinane (6.2 g, 14.6 mmol) in portions at 0 °C and the resulting mixture was stirred at room temperature for 16 h. The reaction was diluted with DCM and extracted with saturated aqueous sodium bicarbonate solution and brine. The organic layer was dried and concentrated to give crude product, which was purified via silica gel chromatography eluting with DCM/methanol (60: 1) to give ethyl 2-oxo-2-(2-oxopropylamino)acetate (1.7 g, 81.2% yield) as yellow syrup. LCMS retention time 0.392 min, LCMS MH+ 174. Step 3 ethyl 5-methyloxazole-2-carbox late
Figure imgf000402_0001
To a solution of ethyl 2-oxo-2-(2-oxopropylamino)acetate (1.7 g, 9.8 mmol) in toluene (10 mL) was added phosphorus oxychloride (2 mL). The mixture was heated to reflux for 3 h. The reaction was concentrated to dryness and the residue was taken up in ethyl acetate (20 mL). This organic phase was washed with saturated aqueous sodium bicarbonate solution and brine, dried and concentrated to give a crude product which was purified via silica gel chromatography eluting with petroleum ether/ ethyl acetate (4: 1) to give ethyl 5-methyloxazole-2-carboxylate (0.68 g, 44.1 % yield) as yellow syrup. Vf-NMR (CDCI3) δ 6.97 (s, 1H), 4.44-4.49 (q, 2H), 2.43 (s, 3H), 1.42-1.46 (t, 2H). LCMS retention time 0.643 min; LCMS MH+ 156.
Step 4 (5-methyloxazol- -yl)methanol
Figure imgf000402_0002
To a solution of ethyl 5-methyloxazole-2-carboxylate (0.68 g, 4.38 mmol) in anhydrous THF (10 mL) was added LAH (250 mg, 6.59 mmol) in portions at 0 °C. The resulting mixture was stirred at room temperature for 16 h. The mixture was quenched with ethyl acetate and filtered. The filter cake was washed several times with ethyl acetate and the combined filtrate was
concentrated to give crude product which was purified via silica gel chromatography eluting with DCM /methanol (65: 1) to give (5-methyloxazol-2-yl)methanol (0.33 g, 66.6% yield) as yellow syrup. LCMS retention time 0.328 min; LCMS MH+ 114.
Intermediate 41 5-(chloromethyl)isoxazole
Figure imgf000402_0003
To a solution of isoxazole-5-carboxylic acid (1.0 g, 8.8 mmol,) in THF (10 mL) was added borane-THF complex (26.4 mL,26.4 mmol) at 0 °C. The reaction was stirred at room
temperature until the substrate was consumed. The reaction was quenched with ethanol (5 mL) at 0 °C. The reaction mixture was partitioned between ethyl acetate and water. The combined organic phase was dried over sodium sulfate, filtered and concentrated to give a crude product which was purified by column chromatography eluting with petroleum ether/ ethyl acetate (2: 1 to give isoxazol-5-ylmethanol (670 mg, 77.0% yield) as a light yellow oil. LCMS retention time 0.329 min; LCMS MH+ 100.
Isoxazol-5-ylmethanol (50 mg, 0.5 mmol,) was dissolved in thionyl chloride (1 mL) at 0 °C. The reaction was stirred at room temperature until the substrate was consumed. The reaction was concentrated to give 5-(chloromethyl)isoxazole as a brown solid which was used without purification. LCMS retention time 0.349 min; LCMS MH+ 1 18.
Intermediate 42 (1 -( sulfonate
Figure imgf000403_0001
To a solution of cyclopropane- 1 ,1 -diyldimethanol (0.1 g, 0.98 mmol) in DCM (5 mL) was added TEA (0.15 g, 1.44 mmol). Methanesulfonyl chloride (0.1 1 g, 0.98 mmol) was added dropwise at 0 °C. The reaction was stirred at room temperature for 2h. The mixture was partitioned between DCM and water. The combined organic layer was dried over sodium sulfate, filtered and concentrated to give a crude product which was used without purification.
Intermediate 43 2-(l-hydroxycyclopentyl)ethyl methanesulfonate
Figure imgf000403_0002
Step 1 tert-butyl 2-(l -hydro ycyclopentyl)acetate
Figure imgf000403_0003
To a solution of tert-butyl acetate (5.0 g, 42.8 mmol,) in THF (20 mL) was added lithium diisopropylamide (21.4 mL, 42.8 mmol) at -60 °C. After stirred at -60 °C for 30 min,
cyclopentanone(3.0 g, 35.7 mmol) was added at -60 °C. The reaction was stiired 1 h; then it was quenched with aqueous ammonium chloride (5 mL) at -60 °C and the mixture was warmed to room temperature. The reaction was partitioned between ethyl acetate and water. The combined organic layer was dried over sodium sulfate, filtered and concentrated to give a crude product which was purified by column chromatography eluting with petroleum ether/ ethyl acetate (3: 1) to give tert-butyl 2-(l -hydroxycyclopentyl)acetate (5.0 g, 70.1 % yield) as a yellow oil. Step 2 l -(2-hydroxyethyl)cyclopentanol
Figure imgf000404_0001
To a solution of tert-butyl 2-(l-hydroxycyclopentyl)acetate (0.5 g, 2.5 mmol) in THF (10 mL) was added lithium aluminium hydride (0.28 g, 7.5 mmol) portionwise at 0°C. The reaction was stirred at room temperature overnight. The mixture was partitioned between ethyl acetate and water. The combined organic layer was dried over sodium sulfate, filtered and concentrated to give a crude product which was purified by column chromatography eluting with petroleum ether/ ethyl acetate (1 : 1) to give l-(2-hydroxyethyl)cyclopentanol (0.33 g, 100%) as a yellow oil. Step 3 2-(l-hydroxycyclopentyl)ethyl methanesulfonate
Figure imgf000404_0002
To a solution of l-(2-hydroxyethyl)cyclopentanol (19 0 mg,1.46 mmol) in dry dichloromethane (2 mL) was added methanesulfonyl chloride (200.3 mg, 1.75 mmol) dropwise at 0 °C , followed by TEA (295 mg, 2.92 mmol). Then the mixture was stirred at room temperature 1 h. The mixture was partitioned between ethyl acetate and water. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give crude product (50mg, 31.1 % . yield), which was used without purification.
Intermediate 44 2-(l-h droxycyclobutyl)ethyl methanesulfonate
Figure imgf000404_0003
The title product was prepared using the method of intermediate 43. Intermediate 45 4-bromo-2-methylbutan-2-ol
Figure imgf000405_0001
To a solution of methyl 3-bromopropanoate (0.2 g, 1.2 mmol,) in THF (5 mL) was added methyl magnesium bromide (2.4 mL, 2.4 mmol) at 0 °C. The mixture was stirred at this temperature until the substrate was consumed based on tic [petroleum ether/ethyl acetate (3: 1); product Rf 0.3]. The reaction was quenched with ammonium chloride (2 mL) at 0 °C. The mixture was partitioned between ethyl acetate and water. The combined organic layer was dried over sodium sulfate, filtered and concentrated, to give a crude product which was purified by a column chromatography eluting with petroleum ether/ ethyl acetate (3: 1 ) to give 4-bromo-2- methylbutan-2-ol (0.15 g, 75.1 % yield) as a yellow oil.
Intermediate 46 l-(2-bromoethyl)cyclopropanol
o
ElMgBr, Ti(Q'Pr)4 V .
B O ' — Br-^ ^QH
To a solution of ethyl 3-bromopropanoate (0.5 g, 2.76 mmol,) in THF (5 mL) was added titanium tetra-isopropanolate (0.8 mL, 0.27 mmol) at 0 °C. Ethyl magnesium bromide (8.27 mL, 8.27 mmol) was added to the mixture at 0 °C. The reaction was stirred at this temperature 2 h. The reaction was quenched with aqueous ammonium chloride (2 mL) at 0 °C. The mixture was partitioned between ethyl acetate and water. The combined organic layer was dried over sodium sulfate, filtered and concentrated to give a crude product which was purified by column chromatography eluting with petroleum ether/ethyl acetate (5: 1) to give l-(2- bromoethyl)cyclopropanol (0.35 g, 77.7% yield) as a yellow oil.
Intermediate 47 7-benzyl-3-ethyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)- dione
Figure imgf000405_0002
Step 1 7-benzyl-8-(3-(trifluoromethoxy)phenoxy)-3-((2-(trimethylsilyl)ethoxy):
1 H-purine-2,6(3H,7H)-dione
Figure imgf000406_0001
To a solution of 7-benzyl-8-chloro-3-((2-(trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H,7H)- dione (500 mg, 1.232 mmol, product of intermediate 18, step 4) in DMF (3 raL) was added 3- (trifluoromethoxy)phenol (280 mg, 1.55 mmol), followed by potassium carbonate (400 mg, 2.90 mmol). The mixture was stirred at 80 °C overnight. The reaction was cooled and partitioned between ethyl acetate and water. The organic phase was washed with brine, dried over sodium sulfate, and concentrated to give 7-benzyl-8-(3-(trifluoromethoxy)phenoxy)-3-((2- (trimethylsilyl)ethoxy)methyl)-l H-purine-2,6(3H,7H)-dione (0.59 g, 87.3% yield) as brown oil. LCMS retention time 2.157 min; LCMS MH+ 549.
Step 2 7-benzy -8-(3-(trifluoromethoxy)phenoxy)-lH-purine-2)6(3H,7H)-dione
Figure imgf000406_0002
To a solution of 7-benzyl-8-(3-(trifluoromethoxy)phenoxy)-3-((2-(trimethylsilyl)ethoxy)methyl)- lH-purine-2,6(3H,7H)-dione (0.59 g, 1.07 mmol) in ethanol (20 mL) was added concentrated HC1 (2 mL). The reaction was stirred at 80 °C for 16 h. The reaction was concentrated and partitioned between ethyl acetate and saturated sodium bicarbonate. The organic phase was washed with brine, dried over sodium sulfate, and concentrated to give 7-benzyl-8-(3- (trifluoiOmethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (390 mg, 87.1% yield) as yellow oil. LCMS retention time 1.543 min; LCMS MH+ 419.
Step 3 7-benzyl-3-ethyl-8-(3-(trifluoromethoxy)phenoxy)-l H-purine-2,6(3H,7H)-dione
Figure imgf000406_0003
To a solution of 7-benzyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (390 mg, 0.93 mmol) in DMF (3 mL) was added iodoethane (150 mg, 0.962 mmol), followed by potassium carbonate (350 mg, 2.54 mmol). The reaction was stirred at 40 °C for 2 h. The mixture was cooled and partitioned between ethyl acetate and water. The organic phase was washed with brine, dried over sodium sulfate, and concentrated to give 7-benzyl-3-ethyl-8-(3-
(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (271 mg, 65.2% yield) as yellow solid. LC S retention time 1.765 min; LCMS MH+ 447. -ethoxy-3-ethyl-l-(3-hydroxypropyl)-lH-purine-2,6(3H,7H)-dione
Figure imgf000407_0001
7-benzyl-8-ethoxy-3-ethyl-l-(3-hydroxypropyl)-lH-purine-2,6(3H,7H)-dione (0.12 g, 0.323 mmol) was dissolved in ethanol (20 mL) and degassed and refilled with nitrogen three times. Ammonium formate (0.5 g, 7.94 mmol) and 10% Pd/C (30 mg) were added and the mixture was again degassed and refilled with nitrogen three times. The reaction was stirred at 80 °C overnight. Then the mixture was cooled, filtered and the filter cake was washed with methanol. The filtrate was concentrated to give 8-ethoxy-3-ethyl-l -(3-hydroxypropyl)-lH-purine- 2,6(3H,7H)-dione (70 mg, 77.1 % yield) as white solid; LCMS retention time 0.413 min; LCMS MH+ 283.
Intermediate 48 l-(4-methoxybenzyl)-3-methyl-8-(3-(trifluoromethoxy)benzyl)-lH- purine-2,6(3H,7H)-dione
Figure imgf000407_0002
Step 1 N-(6-amino-3-(4-methoxybenzyl)-l -methyl-2,4-dioxo-l ,2,3,4-tetrahydropyrimidin-5- yl)-2-(3-(trifluoromethoxy)phenyl)acetamide
Figure imgf000408_0001
To a solution of 5,6-diamino-3-(4-methoxybenzyl)-l -methylpyrimidine-2,4(lH,3H)-dione (50 mg, 0.1 81 mmol, intermediate 59) in ethanol (3 ml) was added 2-(3-
(trifiuoromethoxy)phenyl)acetic acid (40 mg, 0.181 mmol) and EDO (52 mg, 0.271 mmol) The mixture was stirred at room temperature for 2 h. The reaction was diluted with water, filtered and the solids were rinsed with water. The solid residue was dissolved in ethyl acetate, dried over sodium sulfate, filtered and concentrated to give N-(6-amino-3-(4-methoxybenzyl)- l -methyl-2,4- dioxo-1 ,2,3,4-tetrahydropyrimidin-5-yl)-2-(3-(trifluoromethoxy)phenyl)acetamide (70 mg, 80.1 %) as light gray solid. LCMS retention time 1.239 min; LCMS MH+ 479.
Step 2 1 -(4-methoxybenzyl)-3-methyl-8-(3-(trifluoromethoxy)benzyl)-lH-purine-2,6(3H,7H)- dione
Figure imgf000408_0002
To a solution of N-(6-amino-3-(4-methoxybenzyl)-l -methyl-2,4-dioxo-l,2,3,4- tetrahydropyrimidin-5-yl)-2-(3-(trifluoromethoxy)phenyl)acetamide (70 mg, 0.146 mmol) in ethanol (7 ml) was added 2N sodium hydroxide (1 ml). The reaction was stirred at 80 °C for 3 h. The reaction was neutralized with aqueous ammonium chloride, filtered and the solids were rinsed with water. The solid residue was dissolved in ethyl acetate, dried over sodium sulfate, filtered and concentrated to give l -(4-methoxybenzyl)-3-methyl-8-(3-(trifluoromethoxy)benzyl)- lH-purine-2,6(3H,7H)-dione (60 mg, 89.1%) as light yellow solid. LCMS retention time 1.499 min; LCMS MH+ 461.
Intermediate 49 3-(5-methylthiazol-2-yl)propanoic acid
Figure imgf000409_0001
Step 1 (E)-ethyl 3-(5-methylthiazol-2-yl)acrylate
Figure imgf000409_0002
To a solution of n-BuLi (4.2 ml, 6.74 mmol) in THF (15 mL) was added 2-bromo-5- methylthiazole (1.0 g, 5.65 mmol) dropwise under nitrogen at -70 °C. The reaction was stirred at this temperature for 1.5 h. DMF (0.65 ml, 8.43 mmol) was added dropwise at -70 °C. The reaction was maintained at this temperature 1 h. The reaction was quenched with aqueous saturated ammonium chloride (5 mL) and warmed to room temperature. The mixture was partitioned between ethyl acetate and water. The combined organic layer was dried over sodium sulfate, filtered and concentrated to give the aldehyde intermediate as a yellow oil which was used without purification.
To a suspension of sodium hydride (0.27 g, 6.78 mmol) in THF (10 mL) was added ethyl 2- (diethoxyphosphoryl)acetate (1.52 g, 6.78 mmol) dropwise at 0 °C. After the reaction was stirred 30 min, a solution of the aldehyde in THF (3 mL) was added dropwise at 0 °C. The reaction was stirred at room temperature overnight. The mixture was quenched with aqueous saturated ammonium chloride (5 mL) and partitioned between ethyl acetate and water. The combined organic layer was dried over sodium sulfate, filtered and concentrated to give a yellow oil which was purified by column chromatography eluting with petroleum ether /ethyl acetate(4: l)to give (E)-ethyl 3-(5-methylthiazol-2-yl)acrylate (600 mg, 54.1% yield) as a light yellow oil. LCMS retention time 1.410 min; LCMS MH+ 198.
Step 2 ethyl 3-(5-methylthiazol-2-yl)propanoate -Sy Pd/c, H, sv
Figure imgf000410_0001
To a solution of (E)-ethyl 3-(5-methylthiazol-2-yl)acrylate (200 mg, 1.01 mmol,) in methanol (5 mL) was added 5% Pd/C (20 mg). The reaction was purged with nitrogen and then hydrogen and then was stirred under a hydrogen atmosphere (balloon). The reaction was filtered. The filtrate was concentrated to give ethyl 3-(5-methylthiazol-2-yl)propanoate (200 mg, 99% yield) as a light yellow oil. LCMS retention time 1.33 min; LCMS MH+ 200.
Step 3 3-(5-methylthiazol- -yl)propanoic acid
Figure imgf000410_0002
To a solution of ethyl 3-(5-methylthiazol-2-yl)propanoate (0.2 g, lmmol) in THF/water (3 mL/3 mL) was added lithium hydroxide (61 mg, 1.5 mmol) portionwise at room temperature. The resulting mixture was stirred for 3 h. The solvent was evaporated and the residue was acidified with cone. HC1 to adjust the pH to 1-2. The mixture was partitioned between ethyl acetate and water. The combined organic layer was dried over sodium sulfate and concentrated to afford 3- (5-methylthiazol-2-yl)propanoic acid (0.17 g, 87.7% yield) as a colorless oil. LCMS retention time 0.436 min; LCMS MH+ 172.
Intermediate 50 2-(chloromethyl)-5-methylpyridine hydrochloride
Figure imgf000410_0003
Step 1 2,5-dimethylpyridine 1 -oxide
Figure imgf000410_0004
To a solution of 2,5-dimethylpyridine (2.24 g, 21 mmol) in CHC13 (20 mL) was added MCPBA (4.31 g, 25 mmol) in portions over 30 min. The mixture was stirred at room temperature overnight. Calcium hydroxide (4g, 54 mmol) was added to quench the reaction and the mixture was stirred vigorously for 3 h . The mixture was filtered through a Celite pad. The filtrate was . concentrated and dried under vacuum to give 2, 5-dimethylpyridine 1 -oxide (2.5g, 97.1 % yield) as white solid, which was used without purification. CMS retention time 0.442 min; LCMS MH+ 124.
Step 2 (5-methylpyridin-2-yl)methanol
Figure imgf000411_0001
2, 5-Dimethylpyridine 1 -oxide (2.5g, 20.3 mmol) was added to acetic anhydride (8.2mL) at 100 °C over a period of 30 min. Then the mixture was refluxed 1 h. The mixture was cooled to room temperature and carefully quenched with ethanol (1 lmL). The reaction mixture was
concentrated. The residue was treated with IN HC1 (6 mL) and refluxed 1 h. The reaction was concentrated and the residue was partitioned between ethyl acetate and water. The organic phase was dried over sodium sulfate, filtered and concentrated to give (5-methylpyridin-2-yl) methanol (2g, 80.0% yield) as a light yellow oil. LCMS MH+ 124.
2-(chloromethyl)- -methylpyridine hydrochloride
Figure imgf000411_0002
A solution of (5-methylpyridin-2-yl)methanol in SOCl2 (10 mL) was refluxed 30 min. The mixture was concentrated and dried in vacuo to give 2-(chloromethyl)-5-methylpyridine hydrochloride (2. lg, 91.3% yield) as white solid. LCMS MH+ 142.
Intermediate 51 3-methyl-7-((5-methylpyridin-2-yl)methyl)-8-(3-(trifluoromethoxy)benzyl)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000411_0003
Step 1 1 -(4-methoxybenzyl)-3-methyl-7-((5-methylpyridin-2-yl)methy])-8-(3- (trifluoromethox
Figure imgf000412_0001
To a solution of l-(4-methoxybenzyl)-3-methyl-8-(3-(trifluoromethoxy)benzyl)-lH-purine- 2,6(3H,7H)- dione (1 10 mg, 0.238 mmol, intermediate 48) in DMF (4 mL) was added 2- (chloromethyl)-5-methylpyridine (33.8 mg, 0.238 mmol, intermediate 50), followed by potassium carbonate (49.5 mg, 0.358 mmol). The reaction was stirred at 55 °C for 2 h. The mixture was partitioned between ethyl acetate and water. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give a white solid product (120mg, 88.8% yield) which was used without purification. LCMS MH+ 566.
Step 2 3-methyl-7-((5-methylpyridin-2-yl)methyl)-8-(3-(trifluoromethoxy)benzyl)-lH-purine- 2,6(3H,7H)-dione
Figure imgf000412_0002
To a solution of 1 -(4-methoxybenzyl)-3 -methyl-7-((5 -methylpyridin-2-yl)methyl)-8-(3 -
(trifluoromethoxy) benzyl)-lH-purine-2,6(3H,7H)-dione (120 mg, 0.212 mmol) in DCM (1 .4 mL) was added triflic acid (0.7 mL) and TFA (0.7 mL). The reaction was stirred at room temperature overnight. The mixture was partitioned between ethyl acetate and water. The organic phase was washed with saturated NaHC03 and brine, dried over sodium sulfate, filtered and concentrated to give crude product, which was purified by silica gel chromatography eluting with petroleum/ethyl acetate (10: 1 to 1 :1) to give 3-methyl-7-((5-methylpyridin-2-yl)methyl)-8-(3- (trifluoromethoxy)benzyl)-lH-purine-2,6(3H,7H)-dione (60mg, 63.5% yield) as white solid. LCMS MPT 446. Intermediate 52 4,4,4-trifluorobutyl methanesulfonate MsCI
HO CF3 MsO CF3
TEA/DCM
To a solution of 4,4,4-trifluorobutan-l -ol (100m g, 83 mmol) in dry DCM (2 mL) was added methanesulfonyl chloride (107.3 mg, 72 mmol) and then by TEA (1 58 mg, 216 mmol) at 0 °C. The reaction was stirred at room temperature for 1 h. The reaction was partitioned between ethyl acetate and water. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give a yellow oil product (50mg, 31.1 % yield), which was used without purification. Product tic [petroleum ether/ethyl acetate (3: 1) Rf 0.4]
Intermediate 53 8-amino-7-(4-chlorobenzyl)-3-methyl-l-(3-(tetrahydro-2H-pyran-2- yloxy)propyl)-lH-purine-2,6(3H,7H)-dione
Figure imgf000413_0001
Step 1 8-azido-7-(4-chlorobenzyl)-3-methyl-l-(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-lH- purine-2,6(3H,7 -dione
Figure imgf000413_0002
To a solution of 8-bromo-7-(4-chlorobenzyl)-3-methyl-l-(3-(tetrahydro-2H-pyran-2- yloxy)propyl)-lH-purine-2,6(3H,7H)-dione (0.28 g, 0.4 mmol, intermediate 14) in DMSO (10 mL) was added sodium azide (65 mg, 1 mmol). The reaction was heated at 65 °C for 2.5 h. The mixture was cooled and partitioned between ethyl acetate and water. The combined organic layer was dried over sodium sulfate, filtered and concentrated to give a crude product which was used without purification. LCMS retention time 1.800 min; LCMS MH+-THP 390.
Step 2 8-amino-7-(4-chlorobenzyl)-3-methy]-l -(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-lH- purine-2,6(3H,7H)-dione
Figure imgf000414_0001
8-azido-7-(4-chlorobenzyl)-3-methyl-l -(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-lH-purine- 2,6(3H,7H)-dione (0.19 g, 0.4 mmol) was dissolved in THF (5 mL) and 10% Pd/C (20 mg) was added. The reaction was blanketed in a hydrogen atmosphere (balloon) and stirred overnight at room temperature. The mixture was filtered through celite, the filter cake was washed with methanol. The filtrate was concentrated in vacuo to give a 8-amino-7-(4-chlorobenzyl)-3- methyl-l-(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-lH-purine-2,6(3H,7H)-dione (0.2 g, 100% yield) as a white solid. LCMS retention time 1.324 min; LCMS MH+-THP 364 Intermediate 54 S nthesis of 2-(chloromethyl)-4-methylthiazole
Figure imgf000414_0002
Step 1 Synthesis of (4-methylthiazol-2-yl)methanol
1) "BuLi NaBH4
1/ 2) DMF J ^CH0 X ~\
To a solution of 4-methylthiazole (1.0 g, 10.1 mmol) in THF (30 mL) was added n-BuLi (7.56 mL, 13.48 mmol) dropwise under a nitrogen atmosphere at -60 °C. The reaction was stirred for 1 h; then DMF (1.4 ml, 18.2 mmol) was added dropwise while maintaining -60 °C. The resulting mixture was stirred at this temperature for 30 min. The reaction was quenched with aqueous saturated ammonium chloride (5 mL) and partitioned between ethyl acetate and water. The combined organic layer was dried over sodium sulfate, filtered and concentrated to give a yellow oil. This oil was dissolved in methanol (15 ml) and sodium borohydride (460 mg, 12.1 mmol) was added portionwise under nitrogen atmosphere at -60 °C. The mixture was stirred at this temperature for 1 h. The reaction mixture was quenched with acetone, warmed to room temperature, and concentrated. The residue was partitioned between ethyl acetate and water. The combined organic layer was dried over sodium sulfate, filtered and concentrated. The residue was purified by silica gel chromatography eluting with petroleum ether/ ethyl acetate (3 : 1) to give (4-methylthiazol-2-yl)methanol (1 .3 g, 90.3%) as brown oil . LCMS retention time 0.375 min; LCMS MH+ 130.
Step 2 Synthesis of 2-(chloromethyl)-4-methylthiazole
Figure imgf000415_0001
To a solution of (4-methylthiazol-2-yl)methanol (0.5 g, 3.87 mmol) in DCM (5 mL) was added thionyl chloride (0.19 ml, 2.6 mmol) at 0 °C. The mixture was stirred at room temperature for 2 hour. The reaction was concentrated to give 2-(chloromethyl)-4-methylthiazole (570 mg, crude) as yellow oil, which was used without purification. LCMS retention time 0.895 min; LCMS MH+ 148
Intermediate 55 2-(chloromethyl)-5-methylpyrazine
Figure imgf000415_0002
To a solution of 2,5-dimethylpyrazine (500 mg, 4.62 mmol) in carbon tetrachloride (7 mL) was added NCS (679 mg, 5.09 mmol) followed by BPO (20 mg) and the mixture was heated to 80 °C for 6 hours. The mixture was diluted with DCM and extracted with saturated aqueous sodium sulfite solution and brine. The organic layer was dried and concentrated to give a crude product which was purified via silica gel chromatography eluting with petroleum ether/ ethyl acetate (1 :0 to 15: 1) to give 2-(chloromethyl)-5-methylpyrazine (133 mg, 20.19% yield) as yellow oil. LCMS retention time 0.557 min; LCMS MH+ 143.
Intermediate 56 8-bromo-7-(4-chlorobenzyl)-l-(2-(l-hydroxycyclopropyl)ethyl)-3-methyl- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000415_0003
To a solution of 8-bromo-7-(4-chlorobenzyl)-3-methyl-l H-purine-2,6(3H,7H)-dioiie (150 mg, 0.405 mmol, intermediate 8) in DMF (5 mL) was added l-(2-bromoethyl)cyclopropanol (94 mg, 0.568 mmol) followed by potassium carbonate (84 mg, 0.609 mmol) and TBAI (catalytic amount). The resulting mixture was stirred at 50 °C overnight. The reaction was diluted with ethyl acetated and washed with brine. The organic phase was dried and concentrated to give crude product which was purified via silica gel chromatography eluting with petroleum ether/ ethyl acetate (5: 1 to 2: 1) to give 8-bromo-7-(4-chlorobenzyl)-l-(2-(l -hydroxycyclopropyl)ethyl)- 3-methyl-lH-purine-2,6(3H,7H)-dione (1 50 mg, 81.6% yield) as light yellow solid. LCMS retention time 1.523 min; LCMS MH+ 455.
Intermediate 57 1 -(3-hydroxybutyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH- purine-2 -dione
Figure imgf000416_0001
Step 1 8-bromo-l-(3-hydroxybutyl)-3-methyl-7-((2-(trimethylsilyl)ethoxy)methyl) purine-2,6(3H,7H)-dione
Figure imgf000416_0002
To a solution of 8-bromo-3-methyl-7-((2-(trimethylsilyl)ethoxy)methyl)-l H-purine-2,6(3H,7H)- dione (3 g, 8 mmol, product of intermediate 12, step 1 ) in DMF (15 mL) was added 3- hydroxybutyl 4-methylbenzenesulfonate (2.3 g, 8.8 mmol, intermediate 33) followed by potassium carbonate (2.2 g, 16 mmol) and TBAI (catalytic amount). The resulting mixture was stirred at 80 °C for 4 h. The reaction was diluted with ethyl acetated and washed with brine. The organic phase was dried and concentrated to give 8-bromo-l -(3-hydroxybutyl)-3-methyl-7-((2- (trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H,7H)-dione (3.5 g, 97.9% yield) as yellow solid. LCMS retention time 1.428 min; LCMS ΜΗ+ 448.
Step 3 l-(3-hydroxybutyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-7-((2- (trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H,7H)-dione
Figure imgf000417_0001
To a solution of 8-brorno-l -(3-hydroxybutyl)-3-methyl-7-((2-(trimethylsilyl)ethoxy)methyl)-lH- purine-2,6(3H,7H)-dione (1 g, 2.2 mmol) in DNF (10 mL) was added 3- (trifluoromethoxy)phenol (0.6 g, 3 mmol) followed by potassium carbonate (0.6 g, 4.4 mmol). The reaction was stirred at 80 °C for 4 h. The mixture was diluted wuith ethyl acetate and washed with brine. The organic phase was dried and concentrated to give l -(3-hydroxybutyl)-3- methyl-8-(3-(trifluoromethoxy)phenoxy)-7-((2-(trimethylsilyl)ethoxy)methyl)-l H-purine- 2,6(3H,7H)-dione (1.1 g, 91 .8% yield) as yellow syrup. LCMS retention time 2.036 min; LCMS MH+ 545.
Step 3 Synthesis of l -(3-hydroxybutyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine- 2,6(3H,7H)-dione
Figure imgf000417_0002
To a solution of l -(3-hydroxybutyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-7-((2- (trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H,7H)-dione (1.1 g, 2.02 mmol) in THF (5 mL) was added TBAF (5 mL, lmmol/L in THF) and the mixture was heated at reflux for 16 h. The mixture was cooled and diluted with ethyl acetate. The organic phase was washed with brine and saturated aqueous ammonium chloride; then it was dried and concentrated to give a crude product which was purified by silica gel chromatography eluting with DCM/ methanol (45: 1) to give (701 mg, 83.8% yield) as white solid. LCMS retention time 1.372 min; LCMS MH+ 415. Intermediate 58 l-(2-(l -hydroxycyclopropyl)ethyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-lH- urine-2,6(3H,7H)-dione
Figure imgf000417_0003
The title compound was prepared using the method of intermediate 57 and purified by silica gel chromatography eluting with petroleum ether/ ethyl acetate (1 : 1) to give l -(2-(l- hydroxycyclopropyl)ethyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-l H-purine-2,6(3H,7H)- dione (400 mg, 87.0% yield) as white solid. LCMS retention time 1.457 min; LCMS MH+ 427.
Intermediate 59 5,6-diamino-3-(4-methoxybenzyl)-l -methylpyrimidine-2,4(l H,3H)-dione
Figure imgf000418_0001
Step 1 (E)-N'-(l -(4-methoxybenzyl)-3-methyl-2,6-dioxo- 1,2,3, 6-tetrahydropyrimidin-4- yl)-N,N-dimethylformimidami
Figure imgf000418_0002
To a solution of 6-amino-l-methylpyrimidine-2,4(lH,3H)-dione (13.1 g, 92.9 mmol) in DMF (200 mL) was added 1 , 1 -dimethoxy-N,N-dimethylmethanamine (13 mL, 97.9 mmol) and the mixture was stirred at 40 °C for 3 h. Then l-(chloromethyl)-4-methoxybenzene (15.05 mL, 1 1 1.5 mmol) was added followed by potassium carbonate (25.64 g, 0.186 mmol) and DMF (100 mL), the resulting mixture was stirred at 80 °C for 48 h. The reaction was cooled and diluted with ethyl acetate. The organic phase was washed with brine, dried and concentrated to give a crude product which was purified by silica gel chromatography eluting with methanol/ DCM (1 :80 to 1 :20) to give (E)-N'-(l -(4-methoxybenzyl)-3-methyl-2>6-dioxo-l,2,3,6-tetrahydropyrimidin-4- yl)-N,N-dimethylformimidamide (7.8 g, 26.5% yield) as yellow solid. LCMS retention time 1.023 min; LCMS MH+ 317.
Step 2 6-amino-3-(4-methoxybenzyl)-l-methylpyrimidine-2,4(lH,3H)-dione
Figure imgf000418_0003
To a solution of (E)-N'-(l -(4-methoxybenzyl)-3-methyl-2,6-diOxo-l ,2,3,6-tetrahydropyrimidin- 4-yl)-N,N-dimethylformimidamide (7.8 g, 24.7 mmol) in methanol (90 mL) was added ammonium hydroxide (155 mL, 28%) wt/wt) and the mixture was stirred at room temperature for 48 h. The reaction was concentrated to give crude product which was purified by silica gel chromatography eluting with methanol/ DCM (1 : 15) to give 6-amino-3-(4-methoxybenzyl)-l - methylpyrimidine-2,4(l H,3H)-dione (4.3 g, 66.6%) as yellow solid. LCMS retention time 0.445 min; LCMS MH+ 262.
Step 3 6-amino-3-(4-methoxybenzyl)-l -methyl-5-nitrosopyrimidine-2,4(lH,3H)-dione
Figure imgf000419_0001
To a solution of 6-amino-3-(4-methoxybenzyl)-l -methylpyrimidine-2,4(l H,3H)-dione (4.3 g, 16.45 mmol) in water (20 mL) and acetic acid (20 mL) was added sodium nitrite (3.4 g, 49.4 mmol) in small potions at 70 °C. After addition, the mixture was stirred at 70 °C for 4 h. The reaction was cooled and concentrated. The solids were collected, washed with water and dried under vacuum to give 6-amino-3-(4-methoxybenzyl)-l-methyl-5-nitrosopyrimidine-2,4(lH,3H)- dione (4.1 g, 85.9% yield) as purple solid. LCMS retention time 0.885 min; LCMS MH+ 291. Step 4 5,6-diamino-3-(4-methoxybenzyl)-l-methylpyrimidine-2,4(lH,3H)-dione
Figure imgf000419_0002
To a solution of 6-amino-3-(4-methoxybenzyl)-l-methyl-5-nitrosopyrimidine-2,4(lH,3H)-dione (4.1 g, 14.1 mmol) in ammonium hydroxide (100 mL, 14% wt/wt) was added sodium dithionite (4.9 g, 28.2 mmol) in small portions at 60 °C, and the mixture was stirred at this temperature for 4 h. The reaction was cooled and was diluted with water. The solids were collected, rinsed with water and dried under vacuum to give 5,6-diamino-3-(4-methoxybenzyl)-l -methylpyrimidine- 2,4(1 H,3H)-dione (3.5 g, 89.8% yield) as a light yellow solid. LCMS retention time 0.404 min; LCMS MH+ 277.
Intermediate 60 l-(4-methoxybenzyl)-3-methyl-8-(3-(3-
Figure imgf000419_0003
Step 1 ethyl 3-(3-(trifluoromethoxy)phenoxy)propanoate
Figure imgf000420_0001
To a solution of ethyl 4-bromobutanoate (2 g, 10.26 mmol) in acetone (20 mL) was added 3- (trifluoromethoxy)phenol (1.52 g, 8.54 mmol), followed by potassium carbonate (3.5 g, 25.36 mmol). The reaction was stirred at room temperature overnight. The mixture was partitioned between DCM and water. The organic phase was washed with brine, dried over sodium sulfate, and concentrated to give a crude product which was purified by silica gel chromatography eluting with petroleum/ ethyl acetate (10: 1 to 5: 1) to give ethyl 3-(3- (trifluoromethoxy)phenoxy)propanoate (2.3 g, 92.1 % yield) as a colorless oil.
Step 2 3-(3-(trifl
Figure imgf000420_0002
To a solution of ethyl 3-(3-(trifluoromethoxy)phenoxy)propanoate (1.0 g, 3.42 mmol) in THF (8 mL) was added a solution of lithium hydroxide(164 mg, 6.83 mmol) in water (4 mL). The reaction was stirred at room temperature overnight. The reaction was neutralized with diluted HC1 and extracted with ethyl acetate. The organic phase was washed with brine, dried over sodium sulfate, and concentrated to give 3-(3-(trifluoromethoxy)phenoxy)propanoic acid (0.9 g, 99.1% yield) as colorless oil which was used without purification.
Step 3 N-(6-amino-3-(4-methoxybenzyl)-l-methyl-2,4-dioxo-l ,2,3,4-tetrahydropyrimidin-5-yl)- 4-(3-(trifluoromethoxy)phenoxy)butanamide
Figure imgf000420_0003
To a solution of 5,6-diamino-3-(4-methoxybenzyl)-l -methylpyrimidine-2,4(lH,3H)-dione (100 mg, 0.362 mmol, Intermediate 59) in ethanol (3 mL) was added 3-(3- (trifluoromethoxy)phenoxy)propanoic acid (96 mg, 0.364 mmol) and EDCI (105 mg, 0.546 mmol). The reaction was stirred at room temperature for 3 h. Then the mixture was diluted with water and the precipitate was collected by filtration and rinsed with water. The solids were then dissolved in ethyl acetate. This organic solution was dried over sodium sulfate and concentrated to give N-(6-amino-3-(4-methoxybenzyl)-l -methyl-2,4-dioxo-l ,2,3,4-tetrahydropyrimidin-5-yl)- 4-(3-(trifluoromethoxy)phenoxy)butanamide (150 mg, 79.3%) as a yellow solid. LCMS retention time 1.422 min; LCMS MH+ 523.
Step 4 1 -(4-methoxybenzyl)-3-methyl-8-(3-(3-(trifluoromethoxy)phenoxy)propyl)-l H- purine-2,6(3H,7H)-dione
Figure imgf000421_0001
To a solution of N-(6-amino-3-(4-methoxybenzyl)-l -methyl-2,4-dioxo-l , 2,3,4- tetrahydropyrimidin-5-yl)-4-(3-(trifluoromethoxy)phenoxy)butanamide (150 mg, 0.287 mmol) in ethanol (7 mL) was added 2N NaOH(l mL). The mixture was stirred at 80 °C for 3 h. The reaction was neutralized with saturated ammonium chloride and the product precipitated. The solids were collected and washed with water. The solids were dissolved in ethyl acetate, dried over sodium sulfate and concentrated to give l -(4-methoxybenzyl)-3 -methyl -8-(3 -(3- (trifluoromethoxy)phenoxy)propyl)-lH-purine-2,6(3H,7H)-dione (130 mg, 89.7%) as a light yellow solid. LCMS retention time 1.635 min; LCMS MH+ 505.
Intermediate 61 8-chloro-7-(4-chlorobenzyl)-l -(3-hydroxypropyl)-lH-purine-2,6(3H,7H)- dione
Figure imgf000421_0002
Step 1 2-amino-7-(4-chlorobenzyl)-lH-purin-6(7H)-one
Figure imgf000422_0001
To a solution of 2-amino-9-((2R,3S,4R,5S)-3,4-dihydroxy-5-(hydroxymethyl)tetrahydrofuran-2- yl)-lH-purin-6(9H)-one (14 g,49.47 mmol) in DMSO (50 mL) was added l -chloro-4- (chloromethyl)benzene (10 g, 62.16 mmol). The reaction was stirred at 50 °C overnight. The mixture was cooled to room temperature and HCI (50 mL, 10% w.w) was added. The reaction was stirred at 70 °C for 2 h. The reaction was cooled and the precipitate was collected and washed with water and ethanol. The product was dried under vacuum to give 2-amino-7-(4- chlorobenzyl)-lH-purin-6(7H)-one (10 g, 74.1 % yield) as grey solid. LCMS retention time 0.529 min; LCMS MH+ 276.
Step 2 7- 4-chlorobenzyl)-lH-purine-2,6( -dione
Figure imgf000422_0002
To a solution of 2-amino-7-(4-chlorobenzyl)-lH-purin-6(7H)-one (6.8 g, 24.7 mmol) in acetic acid (80 mL) and water (10 mL) was added a solution of sodium nitrite (3.4g, 24.7 mmol) in water (10 mL) dropwise at 50 °C. The reaction was stirred at 50 °C for 1 h. The mixture was cooled to room temperature and stirred for another 1 .5 h. The precipitate was collected, washed with water and ethanol, and dried in vacuo to give 7-(4-chlorobenzyl)-lH-purine-2,6(3H,7H)- dione (6.5 g, 95.3% yield) as yellow solid. LCMS retention time 0.635min; LCMS MH+ 277. Step 3 8-chloro-7-(4-chlorobenzyl)-lH-purine-2,6(3H,7H)-dione
Figure imgf000422_0003
To a solution of 7-(4-chlorobenzy])-l H-purine-2,6(3H,7H)-dione (2.2 g, 7.97 mmol) in THF (30 mL) was added NCS (1.1 g, 8.27 mmol) at 0 °C in portions. Then the resulting mixture was stirred at room temperature until it became a clear solution. The reaction was immediately quenched with ice-water and concentrated. The residue was extracted with ethyl acetate. The organic phase was washed with saturated sodium bicarbonate and brine, dried over sodium sulfate, and concentrated to give a crude product. This material was purified by silica gel chromatography eluting with DCM/ methanol (50: 1 to 20: 1) to give 8-chloro-7-(4- chlorobenzyl)-l H-purine-2,6(3H,7H)-dione (450 mg, 1 8.2% yield) as white solid. LCMS retention time 1.071 ; LCMS ΜΙ- -58 31 1.
Step 4 8-chloro-7-(4-chlorobenzyl)-3-'((2-(trimethylsilyl)ethoxy)methyl)-lH-purine- 2,6(3H,7H)-dione
Figure imgf000423_0001
To a solution of 8-chloro-7-(4-chlorobenzyl)-lH-purine-2,6(3H,7H)-dione (450 mg, 1.45 mmol) in DMF (20 mL) was added potassium carbonate (0.4 g, 2.90 mmol), followed by 2- (trimethylsilyl)ethoxymethyl chloride (0.7 mL, 3.96 mmol) at 0 °C. The resulting mixture was stirred at 60 °C for 16 h. The mixture was partitioned between ethyl acetate and water. The combined organic phase was washed with brine, dried over sodium sulfate, and concentrated to give crude product. This material was purified by silica gel chromatography eluting with petroleum/ ethyl acetate (3 : 1 to 1 : 1) to give 8-chloro-7-(4-chlorobenzyI)-3-((2- (trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H,7H)-dione (250 mg, 39.2% yield) as yellow oil. LCMS retention time 1 .886; LCMS MH+-28 413.
Step 5 7-benzyl-l -(3-(tert-butyldimethylsilyloxy)propyl)-8-chloro-3-((2- (trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H,7H)-dione
Figure imgf000423_0002
To a solution of 8-chloro-7-(4-chlorobenzyl)-3-((2-(trimethylsilyl)ethoxy)methyl)-lH-purine- 2,6(3H,7H)-dione (250 mg, 0.568 mmol) in DMF (30 mL) was added (3-bromopropoxy)(tert- butyl)dimethylsilane (0.2 g,0.794 mmol), followed by potassium carbonate (160 mg, 1.13 mmol). The mixture was stirred at 70 °C overnight. The reaction was cooled and partitioned between ethyl acetate and water. The combined organic phase was washed with brine, dried over sodium sulfate, and concentrated to give 7-benzyl-l -(3-(tert-butyldimethylsilyloxy)propyl)- 8-chloro-3-((2-(trimethylsilyl)ethoxy)methyl)-l H-purine-2,6(3H,7H)-dione (340 mg, 97.4%. yield) as colorless oil. LCMS retention time 4.675 min; LCMS MH+ 613.
Step 6 -chloro-7-(4-chlorobenzyl)-l -(3-hydroxypropyl)- l H-purine-2,6(3H,7H)-dione
Figure imgf000424_0001
To a solution of 7-benzyl- l-(3-(tert-butyldimethylsiIyloxy)propyI)-8-chloro-3-((2- (trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H,7H)-dione (340 mg, 0.56 mmol) in ethyl alcohol (20 mL) was added concentrated HC1 (2 mL). The mixture was stirred at 80 °C overnight. The reaction was concentrated, neutralized with saturated sodium bicarbonate, extracted with ethyl acetate. The organic phase was washed with brine, dried over sodium sulfate, and concentrated to give 8-chloro-7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-lH-purine- 2,6(3H,7H)-dione (180 mg, 87.5% yield) as colorless oil. LCMS retention time 1.990 min; LCMS MH+ 369.
Intermediate 62 8-bromo-7-(4-fluorobenzyl)-3-methyl-l -(3-hydroxypropyl)-lH-purine- 2,6(3H,7H)-dione
Figure imgf000424_0002
Steps 1 and 2 8-bromo-7-(4-fluorobenzyl)-3 -methyl- 1 -(3 -(tetrahydro-2H-pyran-2- yloxy)propyl)-lH-purine-2,6(3H,7H)-dione
Figure imgf000425_0001
The title compound was prepared using the methods of intermediate 8, step 2 and intermediate 14 to give 8-bromo-7-(4-fluorobenzyl)-3 -methyl- 1 -(3 -(tetrahydro-2H-pyran-2-yloxy)propyl)- 1 H- purine-2,6(3H,7H)-dione (26 g, 64.3% yield) as white solid. LCMS retention time 1.694 min; LCMS MH+-THP 41 1.
Step 3 8-bromo-7-(4-fluorobenzyl)-l -(3-hydroxypropyl)-3-methyl-l H-purine- 2,6(3H,7H)-dione
Figure imgf000425_0002
To a solution of 8-bromo-7-(4-fluorobenzyl)-3-methyl-l -(3-(tetrahydro-2H-pyran-2- yloxy)propyl)-lH-purine-2,6(3H,7H)-dione (26 g, 52.5 mmol) in ethanol (500 mL) was added concentrated HCl (50 mL) and the mixture was stirred at room temperature for 1 h. The mixture was concentrated to half its volume and the product precipitated. The solids were collected and washed with water and ethanol; then dried under vaccum to give 8-bromo-7-(4-fluorobenzyl)-l- (3-hydroxypropyl)-3-methyl-lH-purine-2,6(3H,7H)-dione (18.1 g, 83.9% yield) as white solid. LCMS retention time 1.243 min; LCMS MH+ 41 1.
Intermediate 63 8-bromo-7-(4-chlorobenzyl)-l -(3-hydroxypropyl)-3-methyl-lH-purine- 2,6(3H,7H)-dione
Figure imgf000425_0003
The title compound was prepared using the method of intermediate 62 to give 8-bromo-7-(4- chlorobenzyl)-l -(3 -hydroxypropyl)-3 -methyl- lH-purine-2,6(3H,7H)-dione (21 g, 93.1 % yield) as white solid. LCMS retention time 1.375 min; LCMS MH+ 429. Intermediate 64 7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-2,3,6,7-tetrahydro-l H-purine-{ sulfonyl chloride
Figure imgf000426_0001
Step 1 7-(4-chlorobenzyl)-8-mercapto-3-methyl-lH-purine-2,6(3H,7H)-dione
Figure imgf000426_0002
The title compound was prepared using the method of intermediate 7 to give 7-(4-chlorobenzyl)- 8-mercapto-3 -methyl- lH-purine-2,6(3H,7H)-dione (710 mg, 81.5% yield) as yellow solid.
LCMS retention time 0.973 min; LCMS MH+ 323.
Step 2 7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-2,3,6,7-tetrahydro-lH-purine-8-sulfonyl chloride
Figure imgf000426_0003
To a solution of 7-(4-chlorobenzyl)-8-mercapto-3 -methyl- l H-purine-2,6(3H,7H)-dione (400 mg, 1.24 mmol) in concentrated sulfuric acid (5 mL) was added aqueous sodium hypochlorite solution (5 mL, 5% active chlorine) drop wise at 0 °C and the mixture was stirred at room temperature for 15 min. The mixture was poured into ice-water and extracted with ethyl acetate. The organic layer was washed with brine, dried and concentrated to give 7-(4-chlorobenzyl)-3- methyl-2,6-dioxo-2,3,6,7-tetrahydro-lH-purine-8-sulfonyl chloride (230 mg, 47.6% yield) as brown solid, which was directly used immediately without purification. The following intermediates 65-71 were prepared using the method of intermediate 5, ntermediate 65 2-(3,5-bis(trifluoromethyl)phenoxy)ethanol
Figure imgf000427_0001
Light yellow oil, mg, 87.6% yield. TLC petroleum ether/ethyl acetate (1 :2), UV detection, Rf=0.4
In -(3-chlorophenoxy)ethanol
Figure imgf000427_0002
Light yellow oil, 350 mg, 89.1% yield. TLC petroleum ether/ethyl acetate (1 :2), UV detection, Rf=0.4
Intermediate 67 2-(m-tolyloxy)ethanol
Figure imgf000427_0003
Light yellow oil, 357 mg, 91% yield. TLC petroleum ether/ethyl acetate (1 :2), UV detection, RfM).35
Intermediat 68 2-(3-(trifluoromethyl)phenoxy)ethanol
Figure imgf000427_0004
Light yellow oil, 332 mg, 81.9% yield. TLC petroleum ether/ethyl acetate (1 :2), UV detection, Rf ).4
Intermediate 69 2- 4-(trifluoromethoxy)phenoxy)ethanol
Figure imgf000427_0005
Light yellow oil, 100 mg, 78.9% yield. TLC petroleum ether/ethyl acetate (1 :2), UV detection, Rf=0.4 Intermediate 70 2-(2-(trifluoromethoxy)phenoxy)ethanol
Figure imgf000428_0001
Light yellow oil, 105 mg, 85% yield. TLC petroleum ether/ethyl acetate (1 :2), UV detection, Rf=0.4 - 4-chloro-3-(trifluoromethyl)phenoxy)ethanol
Figure imgf000428_0002
Light yellow oil, 100 mg, 81% yield. TLC petroleum ether/ethyl acetate (1 :2), UV detection, Rf-0.4 -hydroxycyclopentyl)methyl methanesulfonate
Figure imgf000428_0003
2-(hydroxymethyl)cyclopentanol
Figure imgf000428_0004
To a solution of ethyl 2-oxocyclopentanecarboxylate (1 g, 6.41 mmol) in anhydrous THF (40 mL) was added lithium bis(trimethylsilyl)arnide (6.41 mL, 6.41 mmol, lmmol/L in THF) at 0 °C under nitrogen. The reaction was stirred at 0 °C for 40 min. Then LAH (500 mg, 13.2 mmol) was added in portions into the above solution at 0 °C. The reactin was stirred at 0 °C for 1 h. The mixture was quenched with ice-water, filtered and the filter cake washed with ethyl acetate. The filtrate was washed with brine, dried over sodium sulfate, and concentrated to give a crude product, which was purified by silica gel chromatography eluting with DCM/ methanol (30: 1 to 10: 1) to give 2-(hydroxymethyl)cyclopentanol (40 mg, 5.4%) as yellow oil. 'H-NMR (CDCI3) δ 4.03-4.05(m, 1H), 3.76-3.80 (q, 1H), 3.54-3.59 (t, 1H), 1.57-2.01 (m, 7H). Step2 (2-hydroxycyclopentyl)methyl methanesulfonate
Figure imgf000429_0001
The title compound was prepared using the method of intermediate 42 to give (2- hydroxycyclopentyl)methyl methanesulfonate (50 mg, 73.5% yield) as yellow oil.
Intermediate 73 l -(3-(benzyloxy)cyclobutyl)-8-chloro-7-(4-chlorobenzyl)-3-methyl-lH- purine-2,6(3 -dione
Figure imgf000429_0002
Step 1 ethyl 4-(3-(3-(benzyloxy)cyclobutyl)ureido)-l -(4-chlorobenzyl)-lH-imidazole-5- carboxylate
Figure imgf000429_0003
To a solution of 3-(benzyloxy)cyclobutanecarboxylic acid (442 mg, 2.14 mmol) in toluene (10 mL) was added diphenylphosphoryl azide (589 mg, 2.14 mmol) followed by TEA (216 mg, 2.14 mmol). The reaction was stirred at room temperature for 30 min. Ethyl 4-amino-l -(4- chlorobenzyi)-l H-imidazole-5-carboxylate (300 mg, 1 .07 mmol) was added to the mixture and the resulting mixture was stirred at 1 10 °C for 6 h. The reaction was partitioned between ethyl acetate and brine. The organic phase was dried and concentrated to give a crude product, which was purified by silica gel chromatography eluting with DCM/ methanol (50: 1) to give ethyl 4- (3-(3-(benzyloxy)cyclobutyl)ureido)-l -(4-chlorobenzyl)-lH-imidazole-5-carboxylate (310 mg, 59.9 yield) as yellow solid. LCMS retention time 1 .909 min; LCMS ΜΙ- 483.
Step 2 1 -(3-(benzyloxy)cyclobutyl)-7-(4-chlorobenzyl)-l H-purine-2,6(3H,7H)-dione
Figure imgf000430_0001
To a solution of ethyl 4-(3-(3-(benzyloxy)cyclobutyl)ureido)-l -(4-chlorobenzyl)-l H-imidazole- 5-carboxylate (310 mg, 0.64 mmol) in ethanol (50 mL) was added freshly prepared sodium ethoxide (87.4 mg, 1.28 mmol) and the reaction was heated at reflux for 4 h. The reaction was cooled and concentrated. The residue was partitioned between ethyl acetate and brine. The organic layer was dried and concentrated to give l-(3-(benzyloxy)cyclobutyl)-7-(4- chIorobenzyl)-lH-purine-2,6(3H,7H)-dione (240 mg, 85.5% yield) as a yellow solid. LCMS retention time 1.764 min; LCMS MH+ 437.
Step 3 1 -(3-(benzyloxy)cyclobutyl)-7-(4-chlorobenzyl)-lH-purine-2,6(3H,7H)-dione
Figure imgf000430_0002
To a solution of l-(3-(benzyloxy)cyclobutyl)-7-(4-chlorobenzyl)-3-methyl- lH-purine- 2,6(3H,7H)-dione (240 mg, 0.62 mmol) in DMF (5 mL) was added iodomethane (105.3 mg, 0.74 mmol) followed by potassium carbonate (128 mg, 0.93 mmol) and the reaction was stirred at room temperature for 16 h. The reaction was partitioned between ethyl acetate and brine. The organic phase was dried and concentrated to give l-(3-(benzyloxy)cyclobutyl)-7-(4- chlorobenzyl)-3-methyl-lH-purine-2,6(3H,7H)-dione (220 mg, 78.9% yield) as a yellow solid. LCMS retention time 1.914 min; LCMS MH+ 451.
Step 4 l-(3-(benzyloxy)cyclobutyl)-8-chloro-7-(4-chlorobenzyl)-3-methyl-lH-purine- 2,6(3H,7H)-dione
Figure imgf000431_0001
To a solution of l -(3-(benzyloxy)cyclobutyl)-7-(4-chlorobenzyl)-3-methyl- l H-purine- 2,6(3H,7H)-dione (220 mg, 0.49 mmol) in DMF (3 mL) was added NCS (71.7 mg, 0.54 mmol) in portions at 0 °C. The reaction was stirred at room temperature for 4 h. The reaction was partitioned between ethyl acetate and saturated aqueous sodium bicarbonate. The organic phase was washed with brine, dried and concentrated to give a crude product, which was purified by silica gel chromatography eluting with DCM/ methanol (60: 1) to give l -(3- (benzyloxy)cyclobutyl)-8-chloro-7-(4-chlorobenzyl)-3-methyl-lH-purine-2,6(3H,7H)-dione (170 mg, 71.8% yield) as yellow solid. LCMS retention time 2.079 min; LCMS MH+ 485.
Intermediate 74 3 -hydroxycyclopentyl methanesulfonate
Figure imgf000431_0002
The title compound was prepared using the method of intermediate 42 step 1 to give 3- hydroxycyclopentyl methanesulfonate (100 mg, 89.1%> yield) as yellow oil which was used without characterization.
Intermediate 75 benzyl 3-(methylsulfonyloxy)cyclobutanecarboxylate
Figure imgf000431_0003
The title compound was prepared using the method of intermediate 42 to give benzyl 3- (methylsulfonyloxy)cyclobutanecarboxylate (0.71 g, 79.8% yield) as yellow solid which was used without characterization.
Intermediate 7 - chlorometh l -5-fluoro ridine
Figure imgf000431_0004
Step 1 (5-fluoropyridin-3-yl)methanol
Figure imgf000432_0001
To a solution of 5-fIuoronicotinic acid (1.0 g, 7.09 mmol) in THF (10 mL) was added TEA (0.9 mL, 7.73 mmol), followed by ethyl chloroformate (0.6 mL, 7.73 mmol) at 0 °C. The reaction was stirred at room temperature 2 h; then it was filtered. The residue was washed with a small amount of THF. The filtrate was chilled to 0 °C and sodium borohydride (0.67 g, 17.73 mmol) was added, followed by dropwise addition of water (5 mL). The reaction was stirred at room temperature overnight. The reaction was partitioned between ethyl acetate and water. The organic layers were combined, dried over sodium sulfate, and concentrated to give a crude product which was used purified by a silica gel chromatography to give (5-fluoropyridin-3- yl)methanol (197 mg, 21.8% yield) as a colorless oil. LCMS retention time 0.375 min; LCMS MH+ 128.
Step 2 3 -(chloromethyl -5 -fluoropyridine
Figure imgf000432_0002
To a solution of (5-fluoropyridin-3-yl)methanol (0.19 g, 1.50 mmol) in DCM (5 mL) was added thionyl chloride (0.19 ml, 2.6 mmol) at 0 °C. The reaction was stirred at room temperature for 2 h. The reaction was concentrated and the residue was dried in vacuo to yield 3-(chloromethyl)-5- fiuoropyridine (166 mg) which was used without characterization. Intermediate 77 8-Bromo-l -(3-((tert-butyldimethylsilyl)oxy)propyI)-7-(4-chlorobenzyl)-3- meth l-l H-purine-2,6(3H,7H)-dione
Figure imgf000432_0003
8-Bromo-7-(4-chlorobenzyl)-3-methyl-lH-purine-2,6(3H,7H)-dione (33.4 g, 90.4 mmol, intermediate 8), potassium carbonate (25.0 g, 180.8 mmol) and TBAI (0.225 g) were combined in DMF(460 mL). To the mixture was added (3-bromopropoxy)(tert-butyl)dimethylsilane (25.17 g, 99.4 mmol) and the reaction was heated at 100°C for 4 h. The reaction was cooled to room temperature, diluted with water (1.2 L) and extracted with ethyl acetate (3 x 400 mL). The combined organic extracts were washed with IN lithium chloride (2 x 500 mL), dried with magnesium sulfate, filtered and the solvent removed under reduced pressure to give a light golden oil. The oil was purified using 2 x 120 g silica gel columns eluted with 10% ethyl acetate / hexanes to yield 8-bromo-l -(3-((tert-butyldimethylsilyl)oxy)propyl)-7-(4-chlorobenzyl)-3- methyl- l H-purine-2,6(3H,7H)-dione (38.3 g, 78% yield) as an off-white solid. LCMS retention time = 5.083 min and 99% purity, LCMS MH+ 543. 'HNMR (CDC13) δ 7.29-7.34 (m, 4H), 5.50 (s, 2H), 4.08 (t, 2H, J= 8 Hz), 3.70 (t, 2H, J = 8 Hz), 3.53 (s, 3H), 1.83-1.90 (m, 2H), 0.86 (s, 9H), 0.02 (s, 6H).
Intermediate 78 8-chloro-7-(4-chlorobenzyl)-3-ethyl-l -(3-hydroxypropyl)-lH-purine- 2,6(3H,7H)-dione
Figure imgf000433_0001
Step 1 7-allyl-2-amino-lH-purin-6(7H)-one
Figure imgf000433_0002
To a solution of 2-amino-9-((2R,3S,4R,5S)-3,4-dihydroxy-5-(hydroxymethyl)tetrahydrofuran-2- yl)-lH-purin-6(9H)-one (50 g, 176.7 mmol) in DMSO (150 mL) was added allyl bromide (36 mL, 420 mmol) , the mixture was stirred at room temperature overnight under nitrogen atmosphere. The reaction was chilled to 0 °C concentrated HC1 (100 mL) was added and the reaction was stirred at 70 °C for 4 h. The reaction was cooled to room temperature, poured into water (800 mL), and neutralized with 6N sodium hydroxide to pH 7-8. The solids that formed were collected and the filter cake was washed with water and ethanol, then dried under vacuum to give 7-allyl-2-amino-l H-purin-6(7H)-one (20 g, 59.2% yield) as yellow solid. LCMS retention time 0.330 min; LCMS ΜΙ-Γ 192.
Step 2 7-allyl-l -purine-2,6(3H,7H)-dione
Figure imgf000434_0001
To a solution of 7-allyl-2-amino-lH-purin-6(7H)-one (20 g, 104.7 mmol) in acetic acid (500 mL) and water (70 mL) was added a solution of sodium nitrite (30 g, 434.8 mmol) in water (70 mL) dropwise at 50 °C. The reaction was stirred at 50 °C for 1 h. The reaction was concentrated and poured into ice- water. The solids that formed were filtered and the filter cake was washed with water and ethanol, then dried under vaccum to give7-allyl-l H-purine-2,6(3H,7H)-dione (15 g, 74.4% yield) as yellow solid. LCMS retention time 0.555 min; LCMS MH+ 193. Step 3 7-allyl-3-ethyl-lH-purine-2,6(3H,7H)-dione
Figure imgf000434_0002
To a solution of 7-allyl-lH-purine-2,6(3H,7H)-dione (15 g, 78.13 mmol) in DMF (200 mL) was added potassium carbonate (23.7 g, 171.74 mmol), followed by dropwise addition of iodoethane (3.6 mL, 44.54 mmol). The reaction was stirred at room temperature overnight. The reaction was concentrated and partitioned between ethyl acetate and water. The organic phase was washed with brine and dried over sodium sulfate, and concentrated to give a crude product which was purified by silica gel chromatography eluting with petroleum/ ethyl acetate (6: 1 to 3: 1) to give 7- allyl-3-ethyl-lH-purine-2,6(3H,7H)-dione (7 g, 40.7% yield) as yellow solid. LCMS retention time 0.402 min; LCMS MH+ 221.
Step 4 7-allyl-8-chloro-3-ethyl-lH-purine-2,6(3H,7H)-dione
Figure imgf000435_0001
To a solution of 7-allyl-3-ethyl-l H-purine-2,6(3H,7H)-dione (7 g, 31 .82 mmol) in DMF (50 mL) was added NCS (5 g, 37.45 mmol) in portions at 0 °C. The resulting mixture was stirred at room temperature for 5 h under nitrogen. The reaction was partitioned between ethyl acetate and water. The organic phase was washed with brine, dried over sodium sulfate, and concentrated to give 7- allyl-8-chloro-3-ethyl-l H-purine-2,6(3H,7H)-dione (1 1.8 g, -50% pure) as light yellow oil.
LCMS retention time 0.764min; LCMS MFf 255.
Step 5 7-allyl-l-(3-(tert-butyldimethylsilyloxy)propyl)-8-chloro-3-ethyl-lH-purine- 2,6(3H,7H)-dione
Figure imgf000435_0002
To a solution of 7-allyl-8-chloro-3-ethyl-lH-purine-2,6(3H,7H)-dione (6.8 g, 26.77mmol) in DMF (50 mL) was added (3-bromopropoxy)(tert-butyl)dimethylsilane (8 g,31.75 mmol), followed by potassium carbonate (5 g, 36.23 mmol) and TBAI (5 mg, 0.014 mmol). The reaction was stirred at 60 °C overnight. The reaction was cooled to room temperature and partitioned between ethyl acetate and water. The organic phase was washed with brine, dried over sodium sulfate, and concentrated to give a crude product which was purified by silica gel
chromatography eluting with petroleum/ ethyl acetate (15: 1 to 1 : 1) to give 7-allyl-l-(3-(tert- butyldimethylsilyloxy)propyl)-8-chloro-3-ethyl-lH-purine-2,6(3H,7H)-dione (5.88 g, 51.6% yield) as a colorless oil. LCMS retention time 2.224 min; LCMS MH+ 427.
Step 6 -allyl-8-chloro-3-ethyl-l-(3-hydroxypropyl)-lH-purine-2,6(3H,7H)-dione
Figure imgf000435_0003
To a solution of 7-allyl-l -(3-(tert-butyldimethylsilyloxy)propyl)-8-chloro-3-ethyl-lH-purine- 2,6(3H,7H)-dione (3.8 g, 8.92 mmol) in ethanol (20 mL) was added concentrated HC1 (2 mL). The mixture was stirred at room temperature for 0.5 h. The mixture was concentrated and partitioned between ethyl acetate and saturated sodium bicarbonate. The organic phase was washed with brine and dried over sodium sulfate, and concentrated to give 7-allyl-8-chloro-3- ethyl-l -(3-hydroxypropyl)-lH-purine-2,6(3H,7H)-dione (3.8 g) as yellow oil, which was used without purification. LCMS retention time 0.999 min; LCMS MH 313.
Step 7 8-chloro-3-ethyl-l-(3-hydroxypropyl)-l H-purine-2,6(3H,7H)-dione
Figure imgf000436_0001
To a solution of 7-allyl-8-chloro-3-ethyl-l -(3-hydroxypropyl)-lH-purine-2,6(3H,7H)-dione (3.8 g, 12.18 mmol) in THF (40 mL) and DMSO (10 mL) was added
tetrakis(triphenylphosphine)palladium(0) (800 mg, 0.693 mmol) and morpholine (5.4 mL, 62.07 mmol). The reaction was degassed and refilled with nitrogen 3 times. The reaction was stirred at room temperature overnight. The mixture was poured into water (20 mL) and ethyl acetate (100 mL). The solid precipitate was collected, washed with ethyl acetate, and dried under vacuum to give 8-chloro-3-ethyl-l-(3-hydroxypropyl)-lH-purine-2,6(3H,7H)-dione (2.2 g, 66.4% yield)as grey solid. LCMS retention time 0.404 min; LCMS MH+ 273. Step 8 8-chloro-7-(4-chlorobenzyl)-3-ethyl-l-(3-hydroxypropyl)-lH-purine-2,6(3H,7H)- dione
Figure imgf000436_0002
To a solution of 8-chloro-3-ethyl-l-(3-hydroxypropyl)-l H-purine-2,6(3H,7H)-dione (2.2 g, 8.09 mmol) in DMF (10 mL) was added 1 -(bromomethyl)-4-chlorobenzene (2.47 g, 12.1 1 mmol), followed by potassium carbonate (2.23 g, 16.18 mmol) and TBAI (15 mg, 0.042 mmol). The reaction was stirred at 50 °C for 2 h. The reaction was cooled to room temperature and partitioned between ethyl acetate and water. The organic phase. was washed with brine and dried over sodium sulfate, and concentrated to give a crude product which was purified by silica gel chromatography eluting with petroleum/ ethyl acetate (5 : 1 to 1 :2) to give 8-chloro-7-(4- chlorobenzyl)-3-ethyl- l -(3-hydroxypropyl)-ll-l-purine-2,6(3H,7H)-dione (2.3 g, 71 .8% yield) as white solid. LCMS retention time 1 .485 min; LCMS MH+ 397.
Intermediate 79 3,3,3 -trifluoropropyl methanesulfonate
MsCI
ΌΗ OMs
TEA, DCM
To a solution of 3,3,3-trifluoropropan-l-ol (500 mg, 4.38 mmol) in DCM (10 mL) was added TEA (1.22 mL, 8.77 mmol) followed by dropwise addition of methanesulfonyl chloride (0.51 mL, 6.58 mmol) at 0 °C. The resulting mixture was stirred at room temperature for 2 h. The reaction was diluted with DCM and the organic phase washed with brine, dried and concentrated to give crude product which was purified via silica gel chromatography eluting with petroleum ether/ethyl acetate (5: 1) to give 3,3,3-trifluoropropyl methanesulfonate (490 mg, 58.2% yield) as light yellow oil. TLC (petroleum ether/ethyl acetate=3: l), Rf=0.6.
Intermediate 80 3-methyl-8-(3-(trifluoromethoxy)phenoxy)-l -(3,3,3-trifluoropropyl)- 1 H- purine-2,6(3H,7H)-dione
Figure imgf000437_0001
Step 1 3-methyl-8-(3-(trifluoromethoxy)phenoxy)- 1 -(3,3,3-trifluoropropyl)-7-((2-
(trimethylsilyl)ethoxy)methyl)-l H-purine-2,6(3H,7H)-dione
Figure imgf000437_0002
To a solution of 3-methyl-8-(3-(trifluoromethoxy)phenoxy)-7-((2- (trimethylsilyl)ethoxy)methyl)-l H-purine-2,6(3H,7H)-dione (500 mg, 1.06 mmol, intermediate 12, step 2) in DMF (6 mL) was added 3,3,3-trifluoropropyl methanesulfonate (305 mg, 1.59 mmol) followed by potassium carbonate (439 mg, 3.17 mmol). The reaction was stirred at 60 °C overnight. The mixture was diluted with ethyl acetate and extracted with brine. The organic phase was dried and concentrated to give 3-methyl-8-(3-(trifluoromethoxy)phenoxy)-l-(3,3,3- trifluoropropyl)-7-((2-(trimethylsilyl)ethoxy)methyl)-l H-purine-2,6(3H,7H)-dione (510 mg, 84.6% yield) as yellow syrup. LCMS retention time 2.275 min; LCMS MH+ 569.
Step 2 3-methyl-8-(3-(trifluoromethoxy)phenoxy)- l -(3,3,3-trifluoropropyl)-l H-purine-
2,6(3H,7H)-dione
Figure imgf000438_0001
The title compound was prepared as intermediate 17, step 2 to give 3-methyl-8-(3- (trifluoromethoxy)phenoxy)-l-(3,3,3-trifluoropropyl)-lH-purine-2,6(3H,7H)-dione (228 mg, 56.3% yield) as white solid. LCMS retention time 1.626 min; LCMS MH+ 439.
Intermediate 81 8-Bromo-7-isopentyl-3-methyl-lH-purine-2,6(3H,7H)-dione
Figure imgf000438_0002
8-Bromo-3-methyl-lH-purine-2,6(3H,7H)-dione (2.0 g, 8.16 mmol), diisopropylethylamine (2.13 mL, 12.24 mmol) and l -bromo-3-methylbutane (0.98 mL, 8.16 mmol) were combined in DMF (20 mL) and stirred at room temperature for 15 h. The reaction was diluted with water (200 mL) and extracted with ethyl acetate (3 x 100 mL). The combined extracts were washed with 1 N LiCl (2 x 100 mL), dried with magnesium sulfate, filtered and evaporated under reduced pressure to give 8-bromo-7-isopentyl-3-methyl-17 -purine-2,6(3H,7H)-dione (2.2 g, 78% yield) as a white solid. LCMS retention time = 2.654 min and 98% purity, LCMS MH+ 315.
Intermediate 82 8-Bromo-l -(3-((tert-butyldimethylsilyl)oxy)propyl-7-isopentyl-3-methyl- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000439_0001
8-BiOmo-7-isopentyl-3-methyl-l -purine-2,6(3H,7H)-dione (2.0 g, 6.35 mmol), (3- bromoprpoxy)(tert-butyl)dimethylsilane (1.77 g, 6.98 mmol intermediate 81), potassium carbonate (1 .76 g, 12.70 mmoL) and TBAI (0.020 g) were combined in DMF (45 mL) and stirred at room temperature for 6 h. The reaction was diluted with water (200 mL) and extracted with ethyl acetate (3 x 100 mL). The combined extracts were washed with 1 N LiCl (2 x 100 mL), dried with magnesium sulfate, filtered and evaporated under reduced pressure to leave a golden oil. The oil was purified using a 40 g silica gel flash column eluted with 10% ethyl acetate / hexanes to give 8-bromo-l-(3-((tert-butyldimethylsily!)oxy)propyl-7-isopentyl-3- methyl-lH-purine-2,6(3H,7H)-dione (2.73 g, 88% yield) as a clear oil: LCMS retention time = 5.271 minutes and 92% purity, LCMS MH+ = 489..
Intermediate 83 7-Benzyl-8-bromo-l-(3-((tert-butyldimethylsilyl)oxy)propyl)-3-methyl- -purine-2,6(3H,7H)-dione
Figure imgf000439_0002
The title compound was prepared using the methods of intermediates 81 and 82. Light golden oil, 15.5g, 100% yield: LCMS retention time = 4.872 minutes and 96% purity, LCMS MH+ = 509. Examples
Example 1 7-(4-chlorobenzyl)-3-methyl-8-(3-(trifluoromethyl)phenoxy)-lH-purine- 2,6(3 H,7H)-dione
Figure imgf000440_0001
To a solution of 8-bromo-7-(4-chlorobenzyl)-3-methyl-lH-purine-2,6(3H,7H)-dione (240 mg, 0.649 mmol, intermediate 8) in dimethyl formamide (5 mL) was added 3-(trifluoromethyl)phenol (105 mg, 0.649 mmol) followed by potassium carbonate (107 mg, 0.779 mmol). The resulting mixture was stirred at 80 °C for 6 h. The mixture was diluted with ethyl acetate (15 mL) and extracted with brine and saturated aqueous ammonium chloride solution. Then the organic phase was dried and concentrated to give a crude solid product which was collected and washed with ethanol to give 7-(4-chlorobenzyl)-3-methyl-8-(3-(trifluoromethyl)phenoxy)-lH-purine- 2,6(3H,7H)-dione (237 mg, 81.1% yield) as white solid. 'H-NMR (OMSO-d6) δ 1 1. 24 (s, 1H), 7.77 (s, lH), 7.69-7.71 (m, 3H), 7.43 (s, 4H), 5.42 (s, 2H), 3.22 (s, 3H). LCMS retention time 2.306 min; LCMS MH+ 451.
Example 2 7-(4-chlorobenzyl)-l -ethyl-3-methyl-8-(3-(trifluoromethyl)phenoxy)-lH-purine- 2,6(3 H,7H)-dione
Figure imgf000440_0002
To a solution of 7-(4-chlorobenzyl)-3-methyl-8-(3-(trifluoromethyl)phenoxy)-lH-purine- 2,6(3H,7H)-dione (50 mg, 0.1 1 mmol, example 1) in DMF (1 mL) was added iodoethane (18.7 mg, 0.12 mmol) followed by potassium carbonate (23 mg, 0.16 mmol). Then the mixture was stirred at 50 °C for 3 h. The mixture was diluted with ethyl acetate and extracted with brine and saturated aqueous ammonium chloride solution. The organic phase was dried and concentrated to give crude solid product which was collected and washed with ethanol to give 7-(4- chlorobenzyl)-l-ethyl-3-methyl-8-(3-(trifluoromethyl)phenoxy)-lH-purine-2,6(3H,7H)-dione (15 mg, 26.7% yield) as white solid. 'H-NMR (OMSO-d6) δ 7.77 (s, 1 H), 7.69-7.73 (m, 3H), 7.43 (s, 4H), 5.46 (s, 2H), 3.91 -3.93 (q, 2H), 3.29 (s, 3H), 1 .1 1 - 1 . 14 (t, 3H). LCMS retention time 3.109 min; LCMS MH+ 479.
Example 3 2-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-(trifluoromethyl)phenoxy)-2,3,6,7- tetrahydro- 1 H-purin- 1 -yl)-N-methylacetamide
Figure imgf000441_0001
The title compound was prepared using the method of example 2 except a catalytic amount of TBAI was added to give 2-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3- (trifluoromethyl)phenoxy)-2,3,6,7-tetrahydro-lH-purin-l -yl)-N-methylacetamide (36 mg, 62.1 % yield) as white solid. 'H-NMR (DMSO-^) δ 7.97-7.98 (m, 1H), 7.79 (s, 1H), 7.69-7.74 (m, 3H), 7.44 (s, 4H), 5.45 (s, 2H), 4.43 (s, 2H), 3.91 -3.93 (q, 2H), 3.29 (s, 3H), 2.58-2.59 (d, 3H). LCMS retention time 2.547 min; LCMS MH"1' 522. Example 4 7-(4-chlorobenzyl)-3-methyl- l -((5-methylisoxazol-3-yl)methyl)-8-(3- (trifluoromethyl)phen- -oxy)-l H-purine-2,6(3H,7H)-dione
Figure imgf000441_0002
The title compound was prepared using the method of example 2 except a catalytic amount of TBAI was added to give 7-(4-chlorobenzyl)-3-methyl-l -((5-methylisoxazol-3-yl)methyl)-8-(3- (trifluoromethyl)phenoxy)-l H-purine-2,6(3H,7H)-dione (29 mg, 39.9% yield) as white solid. XH- NMR (DMSO-i¾ δ 7.79 (s, 1 H), 7.69-7.73 (m, 3H), 7.44 (s, 4H),6.12 (s, l H), 5.45 (s, 2H), 5.07 (s, 1 H), 3.30 (s, 3H), 2.35 (S, 3H). LCMS retention time 2.920 min; LCMS MH+ 546.
Example 5 7-(4-chlorobenzyl)- l -(2-hydroxyethyl)-3-methyl-8-(3-(trifluoromethyl)phenoxy)- l H-purine-2,6(3H,7H)-dione
Figure imgf000442_0001
To a solution of 7-(4-chlorobenzyl)-3-methyl-8-(3-(trifluoromethyl)phenoxy)-l H-purine- 2,6(3H,7H)-dione (60 mg, 0.133 mmol, example 1 ) in DMF (1 mL) was added 2-bromoethanol (16.5 mg, 0.133 mmol) and cesium carbonate (86.6 mg, 0.266 mmol). The resulting mixture was subjected to microwave irradiation at 120 °C for 20 min in a sealed tube. The mixture was diluted with ethyl acetate (5 mL) and extracted with saturated aqueous ammonium chloride solution. The organic phase was dried and concentrated to give crude product which was purified preparative HPLC to give 7-(4-chlorobenzyl)-l-(2-hydroxyethyl)-3-methyl-8-(3- (trifluoromethyl)phenoxy)-lH-purine-2,6(3H,7H)-dione (19 mg, 28.8% yield) as white solid. f-NMR ' (DMSO-i¼) δ 7.76 (s, IH), 7.69-7.71 (m, 3H), 7.43 (s, 4H), 5.45 (s, 2H), 4.77-4.78 (d, 1 H), 3.95-3.98 (t, 2H), 3.51 -3.54 (t, 2H), 3.28 (s, 3H). LCMS retention time 2.608 min; LCMS MH+ 495.
Example 6 7-(4-chlorobenzyl)-l-(2-(dimethylamino)ethyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)- lH-purine-2,6(3H,7H)-dione hydrochloride
Figure imgf000442_0002
Step 1 l-(2-bromoethyl)-7-(4-chlorobenzyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH- purine-2,6(3 -dione
Figure imgf000442_0003
To a solution of 7-(4-chlorobenzyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine- 2,6(3 H,7H)-dione (100 mg, 0.215 mmol, Intermediate 9) in DMF (3 mL) was added 1 ,2- dibromoethane (100 mg, 0.532 mmol), followed by potassium carbonate (60 mg, 0.429 mmo The mixture was stirred at 80 °C overnight. The mixture was diluted with ethyl acetate and water, and the phases were separated. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give crude product, which was purified by recrystallization from ethanol to give l -(2-bromoethyl)-7-(4-chlorobenzyl)-3-methyl-8-(3- (trifluoiOmethoxy)phenoxy)-lH-purine-2J6(3H,7H)-dione (120 mg, 97.6% yield) as white solid. LCMS retention time 2.103 min; LCMS MH+ 573.
Step 2 7-(4-chlorobenzyl)-l-(2-(dimethylamino)ethyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-l H-purine-2,6(3H,7H)-dione hydrochloride
Figure imgf000443_0001
l-(2-bromoethyl)-7-(4-chlorobenzyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-l H-purine- 2,6(3 H,7H)-dione (120 mg, 0.0.210 mmol) was dissolved in aqueous dimethylamine (3 mL), then the mixture was heated to 100 °C in a sealed tube with stirring overnight. The mixture was diluted with ethyl acetate and water, and the phases were separated. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give crude product, which was purified by preparative HPLC and lyophilized to give 7-(4-chlorobenzyl)-l -(2- (dimethylamino)ethyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione hydrochloride (29 mg, 25.6% yield) as white solid. 'H-NMR (DMSO-t¼) δ 9.67(s, 1H), 7.64- 7.60(t, 1H), 7.47-7.41(m, 6H), 7.36-7.34(m, 1H), 5.46(s, 2H), 4.23-4.21(m, 2H), 3.37-3.34(m, 2H), 3.31(s, 3H), 2.87-2.86(d, 6H). LCMS retention time 2.174 min; LCMS MH+ 538. The following examples 7a through 7k were prepared using the method of example 6, step 1.
Example 7a 7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-l -purine-2,6(3H,7H)-dione
Figure imgf000443_0002
The title compound was prepared using the method of example 6, step 1 and purified via preparative HPLC to give 7-(4-chlorobenzyl)-l -(3-hydroxypropyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-l H-purine-2,6(3HJ7H)-dione (10 mg, 17.3% yield) as white solid. 'H-NMR (CD3OD) δ 7.57-7.53(t, 1H), 7.46-7.44(d, 2H), 7.37-7.33(m, 4H); 7.26-7.24(d, 1 H), 5.49(s, 2H), 4.13-4.09(t, 1 H), 3.64-3.60(t, 2H), 3.42(s, 3H), 1.89-1.86(m, 2H). LCMS retention time 3.059 min; LCMS ΜΗ+ 525.
Example 7b 2-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-(trifluoromethoxy)phenoxy)- 2,3,6,7-tetrahydro-lH-purin-l - -N,N-dimethylacetamide
Figure imgf000444_0001
The title compound was prepared with 2-chloro-N,N-dimethylacetamide and purified via preparative HPLC to give 2-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-
(trifluoromethoxy)phenoxy)-2,3,6,7-tetrahydro-l H-purin-1 -yl)-N,N-dimethylacetamide (20 mg, 17.2% yield) as white solid. *H-NMR (400 Hz, DMSO-ik) δ= 7.64-7.59(t, IH), 7.55(s, 1H), 7.48-7.40(m, 5H), 7.35-7.33(d, IH), 5.42(s, 2H), 4.69(s, 2H), 3.30(s, 3H), 3.07(s, 3H), 2.84(s, 3H). LCMS retention time 3.079 min; LCMS MH+ 552.
Example 7c 7-(4-chlorobenzyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-l-(3,3,3- trifluoropropyl)-lH-purine-2,6(3H,7H)-dione
Figure imgf000444_0002
The title compound was prepared with 1,1 ,1 -trifluoro-3-iodopropane to give 7-(4-chlorobenzyl)- 3-methyl-8-(3-(trifluoromethoxy)phenoxy)-l-(3,3,3-trifluoropropyl)-lH-purine-2,6(3H,7H)- dione (30 mg, 35.6% yield) as white solid. 'H-NMR (DMSO-^) δ 7.64-7.59(t, I H), 7.50(s, IH), 7.45-7.41(m, 5H), 7.35-7.33(d, IH), 5.44(s, 2H), 4.15-4.12(t, 2H), 3.22(s, 3H), 2.63- 2.55(m, 2H). LCMS retention time 3.575 min; LCMS MH+ 563. Example 7d 7-(4-chlorobenzyl)-l -(4-hydroxybutyl)-3 -methyl
(trifluoromethoxy)phenoxy - 1 H-purine-2,6(3H,7H)-dione
Figure imgf000445_0001
The title compound was prepared using the method of example 58, step 1 except the reaction was facilitated by microwave irradiation at 120 °C for 45 min to give 7-(4-chlorobenzyl)-l -(4- hydroxybuty])-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (10 mg, 8.9% yield) as white solid. 'H-NMR (DMSO-<¾) δ 7.63-7.58(t, 1Η), 7.49(s, 1Η), 7.44-7.42(m, 5Η), 7.38-7.32(d, 1 Η), 5.44(s, 2Η), 4.41(s, 1 Η), 3.89-3.86(1, 2Η), 3.39-3.34(m, 2Η), 3.29(s, 3Η), 1.61-1.54(m, 2Η), 1.45-1.38(m, 2Η). LCMS retention time 3.053 min; LCMS MH+ 539.
Example 7e 7-(4-chlorobenzyl)-l-(5-hydroxypentyl)-3-methyl-8-(3- (trifluoromethoxy)phe
Figure imgf000445_0002
The title compound was prepared with 5-bromopentan-l -ol to give 7-(4-chlorobenzyl)-l -(5- hydroxypentyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (30 mg, 20.9% yield) as white solid. ^H-NMR (DMSO-< 6) δ 7.63-7.58(t, IH), 7.49(s, IH), 7.44- 7.42(m, 5H), 7.34-7.32(d, IH), 5.44(s, 2H), 4.40-3.78(t, I H), 3.88-3.84(t, 2H), 3.40-3.36(m, H), 3.29(s, 3H), 1.56-1.52(m, 2H), 1.46-1.42(m, 2H), 1.31-1.27(m, 2H). LCMS retention time 3.057 min; LCMS MH+ 553.
Example 7f 7-(4-chlorobenzyl)-l -(2-hydroxy-3-methoxypropyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000446_0001
The title compound was prepared with l -chloro-3-methoxypropan-2-ol and with heating to 130 °C. White solid, 70 mg, 58.8% yield: 'H-NMR (400 Hz, CD3OD) δ 7.57-7.53(t, I H), 7.47- 7.44(d, 2H), 7.37-7.33(m, 4H), 7.26-7.24(d, IH), 5.49(s, 2H), 4.24-4.19(m, IH), 4.12-4.1 l (m, IH), 4.04-3.99(m, IH), 3.45-3.42(m, 5H), 3.36-3.30(d, 3H). LCMS retention time 3.044 min; LCMS MH+ 555.
Example 7g 7-(4-chlorobenzyl)-l-(2-methoxyethyl)-3-methyl-8-(3- (trifluoromemoxy)phenoxy)- -purine-2, 6(3H,7H)-dione
Figure imgf000446_0002
The title compound was prepared with 1 -bromo-2-methoxyethane and purified via preparative HPLC. White solid, 40 mg, 50.9% yield: 1 H-NMR (DMSO-<¾ δ 7.63-7.59(t, 1Η), 7.50(s, 1Η), 7.45-7.43(m, 5Η), 7.34-7.32(d, 1Η), 5.44(s, 2Η), 4.08-4.05(t, 2Η), 3.88-3.84(t, 2Η), 3.52-3.49(t, 2Η), 3.29(s, 3Η), 3.24(s, 3Η). LCMS retention time 3.322 min; LCMS ΜΗ*~ 525.
Example 7h 7-(4-chlorobenzyl)-l-(isoxazol-5-ylmethyl)-3-methyl-8-(3- (tri fluoromethoxy)phenoxy)- 1 H-
Figure imgf000446_0003
The title compound was prepared with 5-(chloromethyl)isoxazole and the reaction was catalyzed with TBAI. The crude product was purified by preparative HPLC to give7-(4-chlorobenzyl)-l- (isoxazol-5-ylmethyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-l H-purine-2, 6(3H,7H)-dione( 21 mg, 8.9% yield) as white solid. 'H-NMR (DMSO-<¾ δ 8.50-8.49(d, 1 Η), 7.64-7.59(t, 1Η), 7.51 (s, I H), 7.46-7.43(m, 5H), 7.36-7.33(d, IH), 6.39-6.38(d, I H), 5.44(s, 2H), 5.22(s, 2H), 3.32(s, 3H). LCMS retention time 3.269 min; LCMS MH+ 548.
Example 7i N-tert-butyl-l -(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3- (trifluoromethoxy)phenoxy)-2,3 ,6,7-telrahydro- 1 H-purin- 1 -yl)methanesulfonamide
Figure imgf000447_0001
The title compound was prepared with N-tert-butyl-l-chloromethanesulfonamide. White solid, 180 mg, 68.3% yield: 'H-NMR (DMSO-^) δ 7.64-7.59(t, IH), 7.49(s, I H), 7.45-7.39(m, 6H), 7.35-7.33(d, I H), 7.14(s, IH), 5.45(s, 2H), 5.21(s, 2H), 3.31(s, 3H), 1.28(s, 9H).LCMS retention time 3.426 min; LCMS MH+ 616.
Example 7j 7-(4-chlorobenzyl)-l-(2-hydroxyethyl)-3-methyl-8-(3- (trifluoromethoxy)phe -purine-2,6(3H,7H)-dione
Figure imgf000447_0002
The title compounds prepared with 2-bromoethanol. White solid, 11 mg, 16.7% yield: 'H-NMR (DMSO-i¾ δ 7.58-7.62 (m, I H), 7.49 (s, I H), 7.41-7.43 (m, 5H), 7.31-7.32 (d, IH), 5.44 (s, 2H), 4.76-4.79 (t, IH), 3.95-3.98 (t, 2H), 3.50-3.54 (t, 2H), 3.28 (s, 3H). LCMS retention time 2.690 min; LCMS MH+ 51 1. Example 7k 7-(4-chlorobenzyl)-l -(3-hydroxybutyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000447_0003
The title compound was prepared with 3-hydroxybutyl-4-methylbenzenesulfonate (intermediate 33). White solid, 70 mg, 61 .9% yield: '/ -NMR (DMSO-i¾ δ 7.59-7.55(t,lH), 7.46- 7.39(d,6H), 7.30-7.28(d,lH), 5.41(s,2H), 4.49-4.48(d,lH), 4.02-3.97(m, l H), 3.84-3.79(m, l H), 3.64-3.61 (m, lH), 3.26(s,3H), 1.59-1 .52(m,2H), 1.06-1.05(d,3H). LCMS retention time 3.159 min; LCMS MH+ 539.
Example 8 3-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-(trifluoromethoxy)phi
2,3,6,7-tetrahydro- 1 H-purin- 1 -yl)propanenitrile
Figure imgf000448_0001
To a solution of 7-(4-chlorobenzyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine-
2,6(3H,7H)-dione (100 mg, 0.215 mmol, Intermediate 9) in a mixed solution of THF (3mL) and acetonitrile (5mL) was added TBAH (0.05 ml, 0.193 mmol) and acrylonitrile (40 mg, 0.642 mmol). The mixture was stirred at 60 °C overnight. Then the mixture was diluted with DCM and water, and the phases were separated. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give crude product. This crude material was purified by silica gel chromatography eluting with petroleum/ ethyl acetate (3:1 to 1 : 1) to give 3-(7-(4- chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-(trifluoromethoxy)phenoxy)-2,3,6,7-tetrahydro-lH- purin-l -yl)propanenitrile (75 mg, 68.8%) as white solid. 'H-NMR (OMSO-d6) δ 7.63-7.59(t, IH), 7.50(s, IH), 7.45-7.40(m, 5H), 7.35-7.33(d, IH), 5.45(s, 2H), 4.16-4.13(t, 2H), 3.31(s, 3H), 2.89-2.86(t, 2H). LCMS retention time 3.248 min; LCMS MH+ 520.
Example 9 2-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-(trifluoromethoxy)phenoxy)- 2,3,6, -tetrahydro-l H-purin- l-yl)acetic acid
Figure imgf000448_0002
Step 1 tert-butyl 2-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-(trifluoromethoxy)phenoxy)- 2,3 ,6,7-tetrahydro- 1 H-purin- 1 -yl)acetate
The title compound was prepared using the method of example 6, step 1 to give 200 mg (100% yield) as yellow oil which was used without purification. LCMS M-'Bu+H 525. Step 2 2-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-(trifluoromethoxy)phenoxy)-2,3,6,7- tetrahydro- 1 H-purin- 1 -yl)acetic acid
Figure imgf000449_0002
To a solution of tert-butyl 2-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3- (trifluoromethoxy)phenoxy)-2,3,6,7-tetrahydro-lH-purin-l -yl)acetate (200 mg, 0.342 mmol) in DCM (5ml) was added trifluoroacetic acid(2 ml, 26.9 mmol) at 0 °C. The mixture was stirred at 0 °C to room temperature for 3 h. Then the mixture was concentrated, diluted with DCM and water. The phases were separated and the organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give 2-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3- (trifluoromethoxy)phenoxy)-2,3,6,7-tetrahydro-lH-purin-l-yl)acetic acid (180 mg, 99.9% yield) as white solid. 'H-NMR (DMSO-c¾ δ 13.06(s, I H), 7.64-7.59(t, IH), 7.55(s, IH), 7.48-7.43(m, 5H), 7.36-7.34(d, I H), 5.43(s, 2H), 4.53(s, 2H), 3.37(s, 3H). LCMS retention time 2.994 min; LCMS MH+ 525.
Example 10 7-(4-chlorobenzyl)-3-methyl-l -(2-(4-methylpiperazin-l -yl)-2-oxoethyl)-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000449_0003
To a solution of 2-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-(trifluoromethoxy)phenoxy)- 2,3,6,7-tetrahydro-l H-purin- l -yl)acetic acid (80 mg, 0.152 mmol, example 9) in NMP (2mL) was added TEA (0.1 mL, 0.614 mmol), 1 -methylpiperazine (0.07 mL, 0.614 mmol) and 0-(7- aza-l H-benzotriazol-l-y^-NjNjN'jN'-tetramethyluronium hexafluorophosphate (87 mg, 0.231 mmol); then the mixture was stirred at 60 °C for 16 h. Then the mixture was diluted with ethyl acetate and water, the phases were separated. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give crude product. This crude material was purified by recrystallization from methanol to give 7-(4-chlorobenzyl)-3-methyl-l -(2-(4- methylpiperazin-l -yl)-2-oxoethyl)-8-(3-(trifluoromethoxy)phenoxy)-l H-purine-2,6(3H,7H)- dione (25 mg, 27.5% yield) as white solid. 'H-NMR (DMSO-<¾ δ 7.62-7.59(t, 1H), 7.55(s, 1H), 7.49-7.39(m, 5H), 7.36-7.34(d, 1 H), 5.43(s, 2H), 4.71 (s, 2H), 3.55-3.48(m, 4H), 3.30(s, 3H), 2.46-2.36(m, 4H), 2.21(s, 3H). LCMS retention time 2.298 min; LCMS MH+ 607.
Example 11 7-(4-chlorobenzyl)-l-(3-(dimethylamino)-2-oxopropyl)-3-methyl-8-(3-
Figure imgf000450_0001
The title compound was prepared using the 2 step method of example 6 to give 7-(4- chlorobenzyl)-l-(3-(dimethylamino)-2-oxopropyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)- lH-purine-2,6(3H,7H)-dione hydrochloride (20 mg, 25.6% yield) as white solid. JH-NMR (DMSO-4 δ 10.05(s, 1H), 7.65-7.61 (t, 1 H), 7.54(s, 1 H), 7.48-7.40(m, 5H), 7.37-7.35(d, 1 H), 5.43(s, 2H),4.85(s, 2H), 4.52(s, 2H), 3.32(s, 3H), 2.80(s, 6H). LCMS retention time 2.352 min; LCMS MH+ 566. '
Example 12 7-(4-chlorobenzyl)-3-methyl-l -(2-(4-methylpiperazin-l -yl)ethyl)-8-(3- (trifluoromethoxy)phenoxy)-l H-purine-2,6(3H,7H)-dione
Figure imgf000450_0002
The title compound was prepared using the 2 step method of example 6 with 1-methylpiperazine to give 7-(4-chlorobenzyl)-3 -methyl- 1 -(2-(4-methylpiperazin- 1 -yl)ethyl)-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (40 mg, 38.7% yield) as white solid. 'H-NMR (CD3OD) δ 7.59-7.55(t, 1 Η), 7.46-7.34(m, 6Η), 7.27-7.25(d, 2Η), 5.51(s, 2Η), 4.19- 4.16(t, 2Η), 3.43(s, 3Η), 3.33-3.32(m, 4Η), 3.03-3.93(bs, 4Η), 2.82-2.73(t, 2Η), 2.69(s, 3Η). LCMS retention time 2.310 min; LCMS MH+ 593.
Example 13 7-(4-chlorobenzyl)-l -(2,3-dihydroxypropyl)-3-methyl-8-(3- (trifluoromethyl)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000451_0001
Step 1 7-(4-chlorobenzyl)-3-methyl-l-(oxiran-2-ylmethyl)-8-(3-(trifluoiOmethoxy)phenoxy)- lH-purine-2,6(3H,7H)-dione
Figure imgf000451_0002
The title compound was prepared using the method of example 6, step 1 to give 7-(4- chlorobenzyl)-3-methyl-l-(oxiran-2-ylmethyl)-8-(3-(trifluoromethoxy)phenoxy)-lH-purine- 2,6(3H,7H)-dione (220 mg, 100% yield) as yellow solid which was used without purification. LCMS MH+ 523.
Step 2 7-(4-chlorobenzyl)-l -(2,3-dihydroxypropyl)-3-methyl-8-(3-(trifluoromethyl)phenoxy)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000451_0003
To a solution of 7-(4-chlorobenzyl)-3 -methyl- l -(oxiran-2-ylmethyl)-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (140 mg, 0.268 mmol) in 1 ,4-dioxane (10 mL) was added TFA (0.1 1 mL, 1.48 mmol) and water (2 mL); then the mixture was stirred at 80 °C for 2 h. The mixture was concentrated; then purified by silica gel chromatography eluting with DCM/ methanol (50: 1 to 20 : 1) to give 7-(4-chlorobenzyl)-l -(2,3-dihydroxypropyl)-3- methyl-8-(3-(trifluoromethyl)phenoxy)-l H-purine-2,6(3H,7H)-dione (70 mg, 48.4% yield) as white solid. 'H-NMR (DMSO-< ) δ 7.63-7.58(t, 1 H), 7.48(s, 1H), 7.44-7.41(m, 5H), 7.34- 7.3 l(d, 1H), 5.44(s, 2H), 4.68-4.67(d, 1H), 4.55-4.52(m, 1H), 4.01-3.98(m, 1H), 3.84-3.80(m, 2H), 3.33(s, 2H), 3.29(s, 3H). LCMS retention time 2.81 lmin; LCMS MH+ 541.
Example 14 7-(4-chlorobenzyl)-l -(3-hydroxy-2-methoxypropyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000452_0001
SStteepp 11 ll--((33--((tteerrtt--bbuuttyyllddiimmeetthhyyllssiillyyllooxxyy))--22--hhyyddrrooxxyypprrooppyyll))--77--((44--cchhlloorroobbeennzzyyll))--33--mmeetthhyyll--88--((33--
((ttrriiflfluuoorroommeetthhooxxyy))pphheennooxxyy))-- 11 HH--ppuurriinnee--22,,66((33HH,,77HH))--ddiioonnee
Figure imgf000452_0002
To a solution of 7-(4-chlorobenzyl)-l-(2,3-dihydroxypropyl)-3-methyl-8-(3- (trifluoromethyl)phenoxy)-l H-purine-2,6(3H,7H)-dione (100 mg, 0.185 mmol, product of example 13) in DCM (10 mL) was added imidazole (1 1 mg, 0.16 mmol) and DMAP (10 mg, 0.08 mmol) at 0 °C, then tert-butylchlorodimethylsilane ( 40 mg, 0.27mmol) was added in one portion . The mixture was stirred at room temperature overnight. The mixture was diluted with DCM and water, and the phases were separated. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give crude l -(3-(tert-butyldimethylsilyloxy)-2- hydroxypropyl)-7-(4-chlorobenzyJ)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-l.H-purine- 2,6(3H,7H)-dione (180 mg, 100% yield) as white solid. LCMS MH+ 655.
Step 2 l-(3-(tert-butyldimethylsilyloxy)-2-methoxypropyl)-7-(4-chlorobenzyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-l H-purine-2,6(3H,7H)-dione
Figure imgf000453_0001
To a slurry of sodium hydride (240 mg, 9.6 mmol)in anhydrous THF (10 ml) was added dropwise a solution of l-(3-(tert-butyldimethylsilyloxy)-2-hydroxypropyl)-7-(4-chlorobenzyl)-3- methy3-8-(3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (124 mg, 0.19 mmol) in THF (3 mL) at 0 °C under nitrogen . The reaction was stirred at 0 °C for 10 miniutes. Then iodomethane (1.2 ml, 19.4 mmol) was added dropwise and the resulting mixture was allowed to warm from 0 °C to room temperature over 2 h. The mixture was quenched with ice-water (5 mL). The mixture was concentrated, diluted with ethyl acetate and water, and the organic layer was separated. The organic layer was dried over sodium sulfate, filtered and concentrated to give crude product (150 mg, 100% yield) as yellow oil. LCMS MH+ 669.
Step 3 7-(4-chlorobenzyl)-l -(3-hydroxy-2-methoxypropyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000453_0002
To a solution of l-(3-(tert-butyldimethylsilyloxy)-2-methoxypropyl)-7-(4-chlorobenzyl)-3- methyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (150 mg, 0.22 mmol) in ethyl alcohol (5 mL) was added concentrated HC1 (0.1 mL). The reaction was stirred for 10 miniutes . Then it was concentrated and purified by preparative HPLC to give 7-(4- chlorobenzyl)-l -(3-hydroxy-2-methoxypropyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH- purine-2,6(3H,7H)-dione (18 mg, 16.2% yield) as white solid. 'H-NMR (DMSO-i¾ δ 7.63- 7.59(t, I H), 7.50(s, I H), 7.45-7.43(m, 5H), 7.34-7.32(d, I H), 5.44(s, 2H), 4.65-4.62(t, IH), 4.12- 4.08(q, IH), 3.84-3.79(q, IH), 3.53-3.41 (m, 2H), 3.29-3.28(d, 6H). LCMS retention time 3.089 min; LCMS MH+ 555.
Example 15 7-(4-chlorobenzyl)- l-(3-(dimethylamino)-2-hydroxypropyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000454_0001
To a solution of 7-(4-chlorobenzyl)-3-methyl-l-(oxiran-2-ylmethyl)-8-(3-
(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (100 mg, 0.192 mmol, example 13, step 1) in THF (10 mL) was added dimethylamine hydrochloride (160 mg, 1.98 mmol) and lithium perchlorate (20 mg, 0.189 mmol). The mixture was stirred at 25 °C in a sealed tube for 2 h. The mixture was diluted with ethyl acetate and water. The phases were separated and the organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give crude product. This crude material was purified via preparative HPLC to give 7-(4- chlorobenzyl)-l-(3-(dimethylamino)-2-hydroxypropyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (50 mg, 46.2% yield) as white solid. 'H-NMR (CD3OD) δ 8.48(s, IH), 7.59-7.54(t, IH), 7.47-7.45(d, 2H), 7.38-7.34(m, 4H), 7.27- 7.25(d, IH), 5.51(s, 2H), 4.34-4.31 (m, IH), 4.23-4.18(m, IH), 4.07-4.02(m, IH), 3.44(s, IH), 3.22-3.2 l (d, 2H), 2.90(s, 6H). LCMS retention time 2.227 min; LCMS MH+ 568.
Example 16 7-(4-chlorobenzyl)-l -(3-methoxy-2-oxopropyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000454_0002
To a solution of 7-(4-chlorobenzyl)-l -(2-hydroxy-3-methoxypropyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-l H-purine-2,6(3H,7H)-dione (40 mg, 0.072 mmol, example 7f) in methyl sulfoxide (4 mL) was added acetic anhydride(37 mg, 0.363 mmol) dropwise. Then the mixture was stirred at 25°C for 16 h. The mixture was partitioned between ethyl acetate and water and the organic phase was washed with brine, dried over sodium sulfate, filtered and was concentrated to give crude product. This crude material was further purified preparative HPLC to give 7-(4-chlorobenzyl)-l-(3-methoxy-2-oxopropyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)- 1 H- purine-2,6(3H,7H)-dione (12 mg, 30.2%) as white solid. yH- NMR (CD3OD) δ 7.56-7.52(t, 1 H), 7.42-7.39(d, 2H), 7.36-7.3 l (m, 4H), 7.24-7.22(d, 1H), 5.45(s, 2H), 4.92(s, 2H), 4.24(s, 2H), 3.46(s, 3H), 3.39(s, 3H). LCMS retention time 3.210 min; LCMS MH+ 553.
Example 17 N-(2-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-(trifluoromethoxy)phenoxy)- 2,3,6,7-tetrahydro-lH-purin-l-yl)ethyl)propane-2-sulfonamide
Figure imgf000455_0001
Step 1 7-(4-chlorobenzyl)-l-(2-(l,3-dioxoisoindolin-2-yl)ethyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000455_0002
The title compound was prepared using the method of example 6, step 1 with 2-(2- bromoethyl)isoindoline-l ,3-dione to give 7-(4-chlorobenzyl)-l -(2-(l ,3-dioxoisoindolin-2- yl)ethyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-l H-purine-2,6(3H,7H)-dione (150 mg, 72.9% yield) as a white solid. LCMS retention time 2.045 min; LCMS MH+ 640.
Step 2 l -(2-aminoethyl)-7-(4-chlorobenzyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-l H- purine-2,6(3 H,7 -dione
Figure imgf000456_0001
To a solution of 7-(4-chlorobenzyl)-l -(2-(l ,3-dioxoisoindolin-2-yl)ethyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (100 mg, 0.157 mmol) in ethanol (10 mL) was added hydrazine hydrate(2 ml) dropwise, then the mixture was stirred at 80°C for 2 h. The mixture was concentrated to give l -(2-aminoethyl)-7-(4-chlorobenzyl)-3-methyl-8-(3-
(trifluoromethoxy)phenoxy)-lH-purine-256(3H,7H)-dione (100 mg, 100%) as yellow oil. LCMS MH+ 510.
Step 3 N-(2-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-(trifluoromethoxy)phenoxy)- 2,3,6,7-tetrahydro-lH-purin-l -yl)ethyl)propane-2-sulfonamide
Figure imgf000456_0002
To a solution of l -(2-aminoethyl)-7-(4-chlorobenzyl)-3-methyl-8-(3-
(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (100 mg, 0.197 mmol) and TEA (40 mg, 0.393 mmol) in DCM (10 mL) was added propane-2-sulfonyl chloride (42 mg, 0.295 mmol) dropwise at 0 °C. The resulting mixture was stirred at 40 °C for 16 h. The mixture was diluted with DCM and water, and the phases were separated. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give crude product. This crude material was recrystallized from ethanol to give N-(2-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3- (trifluoromethoxy)phenoxy)-2,3,6,7-tetrahydro-l H-purin-l-yl)ethyl)propane-2-sulfonamide (20mg, 16.5% yield) as white solid. !H-NMR (DMSO-c¾ δ 7.63-7.59(t, 1H), 7.48(s, 1 H), 7.45-7.41(m, 5H), 7.35-7.33(d, 1 H), 7.26-7.18(m, 1 H), 5.45(s, 2H), 4.01 -3.97(t, 1H), 3.29(s, 611). LCMS retention time 3.195 min; LCMS MH+ 616.
Example 18 (7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-(trifluoromethoxy)phenoxy)-2,3,6,7- tetrahydro-l H-purin-1 -yl)methanesulfonamide
Figure imgf000457_0001
A solution of N-tert-butyl-l -(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3- (trifluoromethoxy)phenoxy)-2,3 ,6,7-tetrahydro- 1 H-purin- 1 -yl)methanesul fonamide ( 120 mg, 0.195 mmol, example 7i) in TFA (2 ml, 26.9 mmol) was stirred at 50 °C for 16 h. The mixture was concentrated and then purified by silica gel chromatography eluting with DCM/ methanol (40: 1) to give (7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-(trifluoromethoxy)phenoxy)-2,3,6,7- tetrahydro-1 H-purin- l-yl)methanesulfonamide (79 mg, 72.4% yield) as white solid. lH-NMR (DMSO-<fc) δ 7.65-7.61(t, 1H), 7.49(s, 1H), 7.47-7.41(m, 6H), 7.36-7.35(d, lH), 7.05(s, 2H), 5.45(s, 2H), 5.23(s, 2H), 3.3 l(s, 3H). LCMS retention time 3.004 min; LCMS MH+ 560.
Example 19 l-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-(trifluoromethoxy)phenoxy)- 2,3,6,7-tetrah dro- 1 H- urin- 1 - l -N-meth lmethanesulfonamide
Figure imgf000457_0002
To a solution of 7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-(trifluoromethoxy)phenoxy)- 2,3,6,7-tetrahydro-l H-purin- l-yl)methanesulfonamide (0.1 g, 0.18 mmol, example 1 8) in DMF (3 mL) was added iodomethane (16 mg, 0.18 mmol) and potassium carbonate(36 mg, 0.26 mmol). The reaction was stirred at room temperature overnight. The mixture was partitioned between ethyl acetate and water. The organic layers were combined, dried over sodium sulfate, filtered and concentrated. The crude product was purified by preparative-HPLC to give l -(7-(4- chlorobenzyl)-3-methyl-2,6-dioxo-8 (3-(trifluoromethoxy)-phenoxy)-2,3,6,7-tetrahydro-lH- purin-l -yl)-N-methylmethanesulfonamide (23 mg, 22.3% yield) as a white solid. 'H-NMR (DMSO-4 δ 7.63(t, I H), 7.50(d, I H), 7.41 -7.46(m, 4H), 7.34-7.37(m, IH), 7.19-7.23(m, IH), 5.45(s, 2H), 5.20(s, 2H), 3.3 l(s, 3H), 2.60(d, 3H).LCMS retention time 3.097 min; LCMS MH+ 574
Example 20 8-(3,5-bis(trifluoromethyl)phenoxy)-7-(4-chlorobenzyl)-3-ethyl-l -methyl-lH- purine-2,6(3H,7H)-dione
Figure imgf000458_0001
Step 1 7-(4-chlorobenzyl)-l -(2-(2-ethyl-l ,3-dioxolan-2-yl)ethyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-l H-purine-2,6(3H,7H)-dione
Figure imgf000458_0002
The title compound was prepared using the method of example 6, step 1 with intermediate 32 to give 7-(4-chlorobenzyl)-l-(2-(2-ethyl-l ,3-dioxolan-2-yl)ethyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (360 mg, 100% yield) as white solid. LCMS MH+ 595.
Step 2 7-(4-chIorobenzyl)-3 -methyl- 1 -(3-oxopentyl)-8-(3-(trifluoromethoxy)phenoxy)-lH- purine-2,6 -dione
Figure imgf000458_0003
The title compound was prepared using the method of example 14, step 3 to give 7-(4- chlorobenzyl)-3-methyl-l-(3-oxopentyl)-8-(3-(trifluoromethoxy)phenoxy)-lH-purine- 2,6(3H,7H)-dione (60 mg, 64.2% yield) as white solid. 'H-NMR DMSO-d6) δ 7.59-7.55(t, IH), 7.46-7.39(d, 6H), 7.31-7.29(d, IH), 5.39(s, 2H), 4.06-4.03(m, 2H), 3.26(s, 3H), 2.69-2.65(t, 2H), 2.48-2.42(t, 2H), 0.91-0.87(t, 3H). LCMS retention time 3.384 min; LCMS MH+ 551. Example 21 7-(4-chlorobenzyl)-l -(3 -hydroxypentyl)-3 -methyl- 8-(3- (trifluoromethoxy)phenoxy)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000459_0001
To a solution of 7-(4-chlorobenzyl)-3-methyl-l -(3-oxopentyl)-8-(3-(trifluoromethoxy)phenoxy)- lH-purine-2,6(3H,7H)-dione (150 mg, 0.273 mmol, example 20) in methanol (20 mL) was added sodium borohydride (40 mg, 1.05 mmol) at 0 °C under nitrogen. The mixture was stirred at 0°C for 2 h. The mixture was diluted with ethyl acetate and water, and the phases were separated. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated. The crude product was purified by preparative HPLC to give 7-(4-chlorobenzyl)-l -(3- hydroxypentyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (50 mg, 33.5% yield) as white solid. 'H-NMR (DMSO-cfc) δ 7.63-7.58(t, 1H), 7.49(s, 1H), 7.44- 7.42(m, 5H), 7.34-7.3 l(d, 1H), 5.44(s, 2H), 4.48-4.46(d, 1H), 4.08-4.01(m, 1H), 3.89-3.83(m, 1H), 3.42-3.37(m, 1H), 3.29(s, 3H), 1.63-1.55(m, 2H), 1.44-1.31 (m, 2H), 0.87-0.84(t, 3H).
LCMS retention time 3.372 min; LCMS MH+ 553.
Example 22 7-(4-chlorobenzyl)-l -(3-hydroxybutyl)-3-methyl-8-(6-methylpyridin-3-yloxy)-lH- purine-2,6(3H,7H)-dione
Figure imgf000459_0002
Step 1 8-(4-chloro-3-(trifluoromethyl)phenoxy)-7-(4-chlorobenzyl)-3 -methyl- lH-purine-
2,6(3H,7H)-dione
Figure imgf000459_0003
The title compound was prepared from intermediate 8 using the method of intennediate 12, step 2 to give 8-(4-chloro-3-(trifluoromethyl)phenoxy)-7-(4-chlorobenzyl)-3-methyl-lH-purine- 2,6(3H,7H)-dione( 77 mg, 39.5% yield) as white solid. LCMS retention time 1.809 min; LCMS MH+ 485
Step 2 7-(4-chlorobenzyl)-l-(3-hydroxybutyl)-3-methyl-8-(6-methylpyridin-3-yloxy)-l H- purine-2,6(3H,7H)-dio
Figure imgf000460_0001
The title compound was prepared using the method of example 6, step 1 to give 7-(4- chlorobenzyl)-l -(3-hydroxybutyl)-3-methyl-8-(6-methylpyridin-3-yloxy)-lH-purine- 2,6(3H,7H)-dione (20 mg, 24.4% yield) as white solid. 'H-NMR (DMSO-^) δ 7.94-7.93(d, 1 H), 7.86-7.84(d, 1H), 7.79-7.77(m, 1H), 7.46(s, 4H), 5.44(s, 2H), 3.94-3.8'9(m, 2H), 3.29(s, 3H), 1.14-1.10(t, 3H).LCMS retention time 3.400 min; LCMS MH+ 513
Example 23 3-benzyl-7-(4-chlorobenzyl)-l-methyl-8-(3-(trifluoromethyl)phenoxy)-lH-purine- 2,6(3 -dione
Figure imgf000460_0002
Step 1 3 -benzyl-8-chloro-7- 4-chlorobenz l - 1 -meth l- 1 H- urine-2 6(3H,7H)-dione
Figure imgf000460_0003
To a solution of 8-chloro-7-(4-chlorobenzyl)- l -methyl- lH-purine-2,6(3H,7H)-dione (50 mg, 0.154 mmol, intermediate 10) in DMF (3 niL) was added (bromomethyl)benzene (50 mg, 0.29 mmol) followed by potassium carbonate (40 mg, 0.29 mmol). The mixture was stirred at 40 °C overnight. The mixture was diluted with ethyl acetate and water, and the phases were separated. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated. The crude product was purified by silica gel chromatography eluting with petroleum/ ethyl acetate (5: 1 to 3: 1) to give 3-benzyl-8-chloro-7-(4-chlorobenzyI)- l -methyl- l H-purine- 2,6(3H,7H)-dione (60 mg, 93.6% yield) as white solid. LCMS retention time 1.893 min; LCMS MH+ 415.
Step 2 3-benzyl-7-(4-chlorobenzyl)-l-methyl-8-(3-(trifluoromethyl)phenoxy)-lH-purine- 2,6(3H,7H)-dione
Figure imgf000461_0001
To a solution of 3-benzyl-8-chloro-7-(4-chlorobenzyl)-l -methyl- l H-purine-2,6(3H,7H)-dione (60 mg, 0.145 mmol) in DMF (3 mL) was added 3-(trifluoromethyl)phenol (35 mg, 0.215 mmol) followed by potassium carbonate (40 mg, 0.29 mmol). The mixture was stirred at 100 °C for 2 h. The mixture was diluted with ethyl acetate and water, and the phases were separated. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated. The give crude product was purified by preparative HPLC to give 3-benzyl-7-(4-chlorobenzyl)-l - methyl-8-(3-(trifluoromethyl)phenoxy)-lH-purine-2,6(3H,7H)-dione (31 mg, 41.7% yield) as white solid. 'H-NMR (CDC13) δ 7.65(s, 1H), 7.58-7.57(m, 2H),7.45-7.41(m, 5H), 7.33-7.27(m, 5H), 5.42(s, 2H), 5.09(s, 2H), 3.41(s, 3H). LCMS retention time 3.649 min; LCMS (M+H) 541.
The following examples 24a through 24m were prepared using the 2 step method of example 23.
Example 24a 7-(4-chlorobenzyl)-3-(2-hydroxyethyl)-l-methyl-8-(3-(trifluoromethyl)phenoxy)- -purine-2,6(3H,7H)-dione
Figure imgf000461_0002
White solid, 7 mg, 10.1 % yield: 'H-NMR (CDC13) 5 7.57-7.51 (m, 3H), 7.44-7.42(m, 3H), 7.34-7.32(m, 2H), 5.44(s, 2H),4.21-4.18(m, 2H), 3.88-3.84(m, 2H), 3.42(s, 3H). LCMS retention time 3.045 min; LCMS MH+ 495. Example 24b 7-(4-chlorobenzyl)-l -methyl-3-(pyridin-2-ylmethy])-8-(3- (trifluoromethyl)phenoxy)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000462_0001
White solid, 30 mg, 50.4% yield: 1H-NMR (DMSO-< ) δ 8.43-8.41(d, 1H), 7.71 -7.63(m, 5H), 7.49-7.44(m, 4H), 7.28-7.23(m, 2H), 5.46(s, 2H), 5.14(s, 2H), 3.26(s, 3H). LCMS retention time 3.279 min; LCMS MH+ 542.
Example 24c 3-benzyl-7-(4-chlorobenzyl)-l-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH- purine-2,6(3H,7H)-dione
Figure imgf000462_0002
White solid, 15 mg, 10.8% yield: ^-NMR (DMSO-^) δ 7.61 -7.56(m, 1 H), 7.49(bs, 1H),
7.41-7.39(m, 5H), 7.31-7.24(m, 6H), 5.41 (s, 2H), 4.99(s, 2H), 3. 22(s, 3H). LCMS retention time 3.705min; LCMS MH+ 557.
Example 24d 7-(4-chlorobenzyl)-3 -(2-hydroxyethyl)- 1 -methyl-8-(3 - (trifluoromethoxy)phenoxy)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000462_0003
White solid, 58 mg, 18.9% yield: V/-NMR (DMSO-^)5 7.63-7.59(m, I H), 7.49-7.42(m, 6H) , 7.34-7.3 l (m, IH), 5.44(s, 2H), 4.81 -4.78(t, IH), 3.94-3.90(m, 2H), 3.60-3.37(m, 2H), 3.24 (s, 3H). LCMS retention time 3.002min; LCMS MH+ 51 1.
Example 24e 7-(4-chlorobenzyl)-l-methyl-3-(pyridin-2-ylmethyl)-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000463_0001
White solid, 38 mg, 62.0% yield: 'H-NMR (OMSO-d6) δ 8.43-8.42(d, IH), 7.73-7.69(m, IH), 7.56-7.52(t, IH), 7.48-7.43(m, 4H), 7.38-7.34(m, 2H), 7.29-7.23(m, 3H), 5.45(s, 2H), 5.15(s, 2H), 3.26 (s, 3H). LCMS retention time 3.266min; LCMS MH+ 558.
Example 24f 8-(3,5-bis(trifluoromethyl)phenoxy)-7-(4-chlorobenzyl)-3-ethyl-l-methyl-lH- purine-2,6(3H,7H)-dione
Figure imgf000463_0002
White solid, 10 mg, 26.2% yield: 3H-NMR (DMSO-<¾ δ 8.23(s, 2H), 8.05(s, IH), 7.48- 7.39(dd, 4H), 5.42(s, 2H), 3.86-3.84(m, 2H), 3.22(s, 3H), 1.14-1.1 l(t, 3H). LCMS retention time 3.591min; LCMS MH+ 547.
Example 24g 7-(4-chlorobenzyl)-3 -ethyl- l -methyl-8-(3 -(trifluorornethoxy)phenoxy)-lH- purine-2,6(3H,7H)-dione
Figure imgf000463_0003
White solid, 20 mg, 20.4% yield: 'H-NMR (DMSO-t¼) δ 7.61 -7.59(1, I H), 7.52(s, I H), 7.43(s, 5H), 7.33-7.32(d, IH), 5.44(s, 2H), 3.89-3.88(m, 2H), 3.24(s, 3H), 1.17-1.13(t, 3H). LCMS retention time 3.561min; LCMS MH+ 495. Example 24h 7-(4-chlorobenzyl)-3-ethyl-l -methyl-8-(3-(trifluoromethyl)phenoxy)-lH-purine- 2,6(3H,7H)-dione
Figure imgf000464_0001
White solid, 20 mg, 24.5% yield: 'H-NMR (DMSO-i¾ δ 7.79(s, IH), 7.73-7.67(m, 3H), 7.47- 7.42(m, 4H), 5.45(s, 2H), 3.90-3.85(q, 2H), 3.30(s, 3H), 1.24-1.13(t, 3H). LCMS retention time 3.51 lmin; LCMS MH+ 479.
Example 24i 7-(4-chlorobenzyl)-l -methyl-3-(pyridin-4-ylmethyl)-8-(3- (trifluoromethoxy)phenoxy)-lH
Figure imgf000464_0002
White solid, 10 mg, 6.21 % yield: 'H-NMR (DMSO-c¾ δ 8.48-8.46(d, 2Η), 7.59-7.55(1, 1Η), 7.47-7.45(m, 5Η), 7.43-7.39(d, 1 Η), 7.38-7.37(d, 1Η), 7.31 -7.24(m, 2Η), 5.45(s, 2Η), 5.05(s, 2Η), 3.25(s, 3Η). LCMS retention time 2.856min; LCMS MH+ 558.
Example 24j 7-(4-chlorobenzyl)-l -methyl-3-(2-oxopropyl)-8-(3-(trifluoromethoxy)phenoxy)- lH-purine-2,6(3H,7H)-dione
Figure imgf000464_0003
White solid, 1 1 mg, 9.93% yield: 'H-NMR (DMSO-4 δ 7.61-7.59(t, I H), 7.44-7.38(m, 6H), 7.32-7.30(d, I H), 5.44(s, 2H), 4.74(s, 2H), 3.24(s, 3H), 2.16(s, 3H). LCMS retention time 3.2 l Omin; LCMS MH+ 523. Example 24k 7-(4-chlorobenzyl)-3-(2-(2-hydroxyethoxy)ethyl)-l-methyl-8-(3- (trifluoromethoxy)phenoxy)- 1 H
Figure imgf000465_0001
White solid, 31 mg, 45.3% yield: 'H-NMR (DMSO-i¾ δ 7.59-7.56(t, 1Η), 7.44-7.37(m, 6Η), 7.31 -7.29(d, 1Η), 5.42(s, 2Η), 4.01 -3.98(t, 2Η), 3.35(s, 4Η), 3.22(s, 3Η). LCMS retention time 2.969min; LCMS MH+ 555.
Example 241 7-(4-chlorobenzyl)-l-methyl-3-propyl-8-(3-(trifluoromethoxy)phenoxy)-l I- purine-2,6(3H,7H)-dione
Figure imgf000465_0002
White solid, 15 mg, 26.8% yield: 'H-NMR (CDC13) δ 7.49-7.43(m, 3LI), 7.35-7.3 l(m, 3Η),
7.24-7.15(m, 2Η), 5.44(s, 2Η), 3.97-3.93(t, 2Η), 3.43(s, 3Η), 1.76-1.70(m, 2Η), 0.94-0.90(t, 3Η). LCMS retention time 3.526min; LCMS MH+ 509.
Example 24m 7-(4-chlorobenzyl)-3-(isoxazol-5-ylmethyl)-l-methyl-8-(3- (trifluoromethoxy)phenoxy)- lH-purine-2,6(3H,7H)-dione
Figure imgf000466_0001
White solid, 16 mg, 16.3% yield: VJ-NMR (DMSO-i¾) δ 8.33-8.3 l (d, 2H), 7.61 -7.57(t, 1H), 7.45-7.4 l (m, 6H), 7.32-7.29(d, 1H), 5.44(s, 2H), 4.34(bs, 2H), 3.26(s, 3H). LCMS retention time 3.1 14min; LCMS MH+ 548.
Example 25 7-(4-chlorobenzyl)-3-(2-(dimethylamino)ethyl)- 1 -methyl-8-(3- (tr
Figure imgf000466_0002
Step 1
3-(2-bromoethyl)-7-(4-chlorobenzyl)-l-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine- 2,6(3H,7H)-dion
Figure imgf000466_0003
To a solution of 7-(4-chlorobenzyl)-l -methyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine- 2,6(3H,7H)-dione (100 mg, 0.215mmol) in DMF (3 mL) was added 1 ,2-dibromoethane (0.2 mL, 2.32 mmol), followed by potassium carbonate (60 mg, 0.435 mmol). The mixture was stirred at 50 °C for 16 h. The reaction was partitioned between ethyl acetate and water. The organic phase was washed with brine and dried over sodium sulfate, filtered and concentrated to give the crude product (1 10 mg, 91.6% yield) as white solid. LCMS retention time 2.099 min; LCMS MH+ 573. Step 2 7-(4-chlorobenzyl)-3-(2-(dimethylamino)ethyl)-l-methj
(trifluoiOmethox
Figure imgf000467_0001
To a solution of 3-(2-bromoethyl)-7-(4-chlorobenzyl)-l -methyl-8-(3-
(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (1 10 mg, 0.192mmol) in DMF (3 mL) was added dimethylamine hydrochloride (150 mg, 1 .84 mmol), followed by potassium carbonate (100 mg, 0.725 mmol). The mixture was stirred at 40 °C for 16 h; then it was diluted with ethyl acetate and water. The phases were separated and the organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated. The residue was purified by preparative HPLC to give 7-(4-chlorobenzyl)-3-(2-(dimethylamino)ethyl)-l -methyl-8-(3r
(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (15 mg, 13.3% yield) as white solid. 'H-NMR (DMSO-4 δ 9.69(s, 1H), 7.64-7.60(1, 1H), 7.54(s, 1H), 7.49-7.43(m, 5H), 7.35- 7.33(d, 1H), 5.47(s, 2H), 4.21-4.18(t, 2H), 3.39(m, 2H), 3.26(s, 3H), 2.79-2.78(d, 6H). LCMS retention time 3.266min; LCMS MH+ 58.
Example 26 7-(4-chlorobenzyl)-l -methyl-3-(2-oxopropyl)-8-(3-(trifluoromethoxy)phenoxy)- lH-purine- -dione
Figure imgf000467_0002
To a solution of 7-(4-chlorobenzyl)-l -methyl-3-(2-oxopropyl)-8-(3-(trifluoromethoxy)phenoxy)- lH-purine-2,6(3H,7H)-dione (80 mg, 0.153 mmol, example 24j) in ethanol (3 mL) was added sodium borohydride (12 mg, 0.317 mmol) at 0 °C. The resulting mixture was stirred at 0 °C overnight. The mixture was concentrated and the residue was purified by preparative HPLC to give the title product (6 mg, 7.47% yield) as white solid. XH-NMR
Figure imgf000467_0003
δ 7.63-7.58(t, 1H), 7.50(s, 1 H), 7.43-7.41 (m, 5H), 7.33-7.31(d, 1H), 5.44(s, 2H), 4.80-4.79(d, 1H), 4.02- 3.99(bs, 1 H), 3.89-3.84(m, 1 H), 3.69-3.65(dd, 1 H), 3.24(s, 3H), 1.01 -0.99(d, 3H). LCMS retention time 3.095min; LCMS MH+ 525.
Example 27 8-(4-chloro-3-(trifluoromethyl)phenoxy)-7-(4-chlorobenzyl)-l -(2-(2- hydroxyethoxy)ethyl)-3-methyl-lH-purine-2,6(3H,7H)-dione
Figure imgf000468_0001
Step 1 8-(4-chloro-3-(trifluoromethyl)phenoxy)-7-(4-chlorobenzyl)-3-methyl-l -(2-(2- (tetrahydro-2H-pyran-2-yloxy)ethoxy)ethyl)-lH-purine-2,6(3H,7H)-dione
Figure imgf000468_0002
To a solution of 8-bromo-7-(4-chlorobenzyl)-3 -methyl- 1 -(2-(2-(tetrahydro-2H-pyran-2- yloxy)ethoxy)ethyl)-lH-purine-2,6(3H,7H)-dione (150 mg, 0.28mmol, intermediate 1 1) in DMF (5 mL) was added 4-chloro-3-(trifluoromethyl)phenol (81 mg, 0.45 mmol) and potassium carbonate (76 mg, 0.55 mmol). The mixture was stirred at 80°C overnight. The mixture was partitioned between ethyl acetate and water. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give a crude product which was used without purification. LCMS retention time 2.1 12 min; LCMS MH+-THP 573.
Step 2 8-(4-chloro-3-(trifluoromethyl)phenoxy)-7-(4-chlorobenzyl)-l-(2-(2- h droxyethoxy)ethyl)-3-methyl-lH-purine-2,6(3H, -dione
Figure imgf000468_0003
To a solution of 8-(4-chloro-3-(trifluoromethyl)phenoxy)-7-(4-chlorobenzyl)-3-methyl-l-(2-(2- (tetrahydro-2H-pyran-2-yloxy)ethoxy)ethyl)-l H-purine-2,6(3H,7H)-dione (100 mg, 0.15 mmol) in ethanol (5 mL) was added acetyl chloride (0.2 niL) at 0 °C. The mixture was stirred at room temperature for 1 h. The mixture was partitioned between ethyl acetate and water. The organic phase was dried over sodium sulfate, filtered and concentrated to give a crude product, which was purified via preparative HPLC to give 8-(4-chloro-3-(trifluoromethyl)phenoxy)-7-(4- chlorobenzyl)- l -(2-(2-hydroxyethoxy)ethyl)-3-methyl- l H-purine-2,6(3H,7H)-dione (37 mg, 43.1 % yield) as white solid. !H-NMR (DMSO-40 δ 7.95(d, IH), 7.85(d, 1H), 7.77-7.80(m, lH),7.44(s, 4H), 5.44(s, 2H), 4.58-4.61(m, IH), 4.06(t, 2H), 3.59(t, 2H), 3.43-3.46(m, 4H), 3.29(s, 3 ). LCMS retention time 3.070 min; LCMS MH+ 573. The following examples 28a through 28g were prepared using the method of example 27.
Example 28a 8-(3,5-bis(trifluoromethyl)phenoxy)-7-(4-chlorobenzyl)-l -(2-(2- hydroxyethoxy)ethyl)-3-methy -lH-purine-2,6(3H,7H)-dione
Figure imgf000469_0001
White solid, 27 mg, 30.1% yield: JH-NMR (DMSO-^) δ 8.25(s, I H), 8.09(s, IH), 7.48(d, 2H), 7.42(d, 2H) , 5.46(s, 2H), 4.59(s, IH), 4.07(t, 2H), 3.57(t, 2H), 3.40-3.46(m, 4H), 3.29(s, 3H). LCMS retention time 3.150 min; LCMS MH+ 607.
Example 28b 7-(4-chlorobenzyl)-l-(2-(2-hydroxyethoxy)ethyl)-3-methyl-8-(3- (trifluoromethyl)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000469_0002
White solid, 30 mg, 40.5% yield: 'H-NMR (DMSO-c¾ δ 7.78(s, I H), 7.70-7.73(m, 3H), 7.44 (s, 4H), 5.45(s, 2H), 4.59(t, IH), 4.06(t, 2H), 3.60(t, 2H), 3.44(s, 4H), 3.32(s, 3H). LCMS retention time 2.923 min; LCMS MH+ 539. Example 28c 7-(4-chlorobenzyl)-l -(2-(2-hydroxyethoxy)ethyl)-3-methyl-8-(m-tolyloxy)-l H- purine-2,6(3H,7H)-dione
Figure imgf000470_0001
White solid, 21 mg, 29.8% yield: 'H-NMR (DMSO-<¾ δ 7.37-7.46(m, 4H), 7.33(t, IH), 7.09- 7.12(m, 3H), 5.42(s, 2H), 4.58(t, IH), 4.06(t, 2H), 3.56(t, 2H), 3.45(t, 4H), 3.32(s, 3H), 2.32(d, 3H). LCMS retention time 2.801 min; LCMS MH+ 485.
Example 28d 7-(4-chlorobenzyl)-l -(2-(2-hydroxyethoxy)ethyl)-8-(3-methoxyphenoxy)-3- methyl- 1 H-purine-2,6(3 H,7H)-dione
Figure imgf000470_0002
White solid, 26 mg, 37.7% yield: 'H-NMR (DMSO-^) δ 7.38-7.47(m, 4H), 7.36(t, IH), 6.88- 6.90(m, 3H), 5.42(s, 2H), 4.59(t, IH), 4.06(t, 2H), 3.76(s, 3H), 3.56(t, 2H), 3.45(s, 4H), 3.29(s, 3H). LCMS retention time 2.631 min; LCMS MH+ 501.
Example 28e 7-(4-chlorobenzyl)-8-(3-chlorophenoxy)-l-(2-(2-hydroxyethoxy)ethyl)-3-methyl- lH-purine-2,6(3H,7H)-dione
Figure imgf000470_0003
White solid, 33 mg, 36.8% yield: 1 H-NMR (DMSO-<¾ δ 7.48-7.52(m, 2Η), 7.35-7.42(m, 6Η), 5.43(s, 2Η), 4.59(t, 1Η), 4.06(t, 2Η), 3.58(t, 2Η), 3.45(s, 4Η), 3.30(s, 3Η). LCMS retention time 2.856 min; LCMS MH+ 505. Example 28f 7-(4-chlorobenzyl)-8-(3-ethylphenoxy)-l -(2-(2-hydroxyethoxy)ethyl)-3-methyl- lH-purine-2,6(3H,7H)-dione
Figure imgf000471_0001
White solid, 27 mg, 36.8% yield: 'H-NMR (DMSO- d6) δ 7.36-7.46(m, 5H), 7.1 1 -7.15(m, 3H), 5.43(s, 2H), 4.61 (t, 1H), 4.06(t, 2H), 3.56(t, 2H), 3.43-3.47(m, 4H), 3.28(s, 3H), 2.62(q, 2H), 1.17(t, 3H). LCMS retention time 2.909 min; LCMS MH+ 499.
Example 28g 7-(4-chlorobenzyl)-l -(2-(2-hydroxyethoxy)ethyl)-3-methyl-8-(3- (morpholinomethy])phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000471_0002
White solid, 41 mg, 41.1% yield: JH-NMR (DMSO-i¾) δ 7.77 (s, 1H), 7.51 -7.58 (m, 2H), 7.43- 7.46 (m, 5H), 5.43 (s, 2H), 4.32-4.37 (m, 2H), 4.03-4.06(m, 2H), 3.85-3.90 (m, 4H), 3.54-3.57 (t, 2H), 3.44 (m, 4H), 3.27 (s, 3H), 3.00-3.09 (m, 4H). LCMS retention time 1.699 min; LCMS MH+ 570.
Example 29 7-(4-chlorobenzyl)-l-(2-(2-hydroxyethoxy)ethyl)-3-methyl-8-(3- methylbenzyloxy)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000471_0003
Step 1 7-(4-chlorobenzyl)-3-methyl-8-(3-methylbenzyloxy)-l-(2-(2-(tetrahydro-2H-pyran-2- yloxy)ethoxy)ethyl)-lH-purine-2,6(3H,7H)-dione
Figure imgf000471_0004
To a solution of m-tolylmethanol (37 mg, 0.3 mmol) in DMF (2 mL) was added sodium hydride (1 8 mg, 0.45 mmol) at 0 °C. After stirring at 0 °C for 30 min, 8-bromo-7-(4-chlorobenzyl)-3- - methyl- l -(2-(2-(tetrahydro-2H-pyran-2-yloxy)ethoxy)ethyl)-lH-purine-2,6(3H,7H)-dione (80 mg, 0.15 mmol, intermediate 1 1) was added. The mixture was stirred at 0 °C for 1 5 min, then aqueous ammonium chloride solution (2 mL) was added at 0 °C. The mixture was partitioned between ethyl acetate and water. The organic layers were combined, dried over sodium sulfate, filtered and concentrated to give a crude product which was used without purification. LCMS retention time 1.967 min; LCMS MH+-THP 499. Step 2 7-(4-chlorobenzyl)-l -(2-(2-hydroxyethoxy)ethyl)-3-methyl-8-(3-methylbenzyloxy)-lH- puri -2,6(3H,7H)-dione
Figure imgf000472_0001
The title product was prepared using the method of example 27, step 2. White solid, 12 mg, 16.2% yield: H-NMR (DMSC ¾ 5 7.39(d, 2H), 7.25-7.28(m, 3H), 7.15-7.2 l(m, 3H), 5.49(s, 2H) , 5.25(s, 2H), 4.57(d, IH), 4.03(t, 2H), 3.54(t, 2H), 3.44(t, 4H), 3.41(s, 3H). LCMS retention time 2.888 min; LCMS MH+ 499.
The following examples 30a through 30d were prepared using the 2 step method of example 29.
Example 30a 7-(4-chlorobenzyl)-8-(3-chlorobenzyloxy)-l-(2-(2-hydroxyethoxy)ethyl)-3- methyl- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000472_0002
White solid, 15 mg, 19.4% yield: 'H-NMR (DMSO-c¾ δ 7.37-7.44(m, 6H), 7.28(d, 2H), 5.53(s, 2H), 5.28(s, 2H), 4.58(t, IH), 4.03(t, 2H), 3.56(t, 2H), 3.45(t, 4H), 3.39(s, 3H). LCMS retention time 2.894 min; LCMS MH+ 519. Example 30b 7-(4-chlorobenzyl)-l-(2-(2-hydroxyethoxy)ethyl)-3-methyl-8-(3- (trifluoromethyl)benzyloxy - -purine-2,6(3H,7H)-dione
Figure imgf000473_0001
White solid, 18 mg, 21.6% yield: 'H-NMR (DMSO-<¾ δ 7.79(s, 1H), 7.74(d, 2H), 7.63(1, 1H), 7.37 (d, 2H), 7.28(d, 2H), , 5.62(s, 2H), 5.28(s, 2H), 4.58(t, 1H), 4.04(t, 2H), 3.55(t, 2H), 3.44- 3.45(m, 4H), 3.42(s, 3H). LCMS retention time 2.921 min; LCMS MH+ 553.
Example 30c 7-(4-chlorobenzyl)-l -(2-(2-hydroxyethoxy)ethyl)-8-(3-methoxybenzyloxy)-3- methyl-lH-purine-2,6(3H,7H)-dione
Figure imgf000473_0002
White solid, 17 mg, 22.1% yield: 'H-NMR (DMSO-£/0) δ 7.36-7.38(m, 2Η), 7.25-7.32(m, 3Η), 6.91 -6.99(m, 3 Η), 5.50(s, 2Η), 5.25(s, 2Η), 4.60(t, 1Η), 4.03(t, 2Η), 3.73(s, 3Η), 3.54(t, 2Η), 3.42-3.47(m, 7Η). LCMS retention time 2.718 min; LCMS MH+ 515. Example 30d 7-(4-chlorobenzyl)-l -(2-(2-hydroxyethoxy)ethyl)-3-methyl-8-(3-
(trifluoromethoxy) benzyloxy)-l H-purine-2,6(3H,7H)-dione
Figure imgf000473_0003
White solid, 31 mg, 22.1% yield: 'H-NMR (DMSO-^) δ 7.53(t, 1Η), 7.37-7.47(m, 5Η), 7.28 (d, 2Η), 5.58(s, 2Η), 5.28(s, 2Η), 4.58(t, 1Η), 4.04(t, 2Η), 3.54(t, 2Η), 3.44(t, 4Η), 3.42(s, 3Η). LCMS retention time 2.984 min; LCMS ΜΙ- 569.
Example 31 1 -(2-(2-hydroxyethoxy)ethyl)-7-(4-methoxybenzyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000474_0001
To a solution of l -(2-(2-hydroxyethoxy)ethyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH- pu ne-2,6(3H,7H)-dione (50 mg, 0.1 16 mmol, intermediate 12) in DMF (3mL) was added 1 - (bromomethyl)-4-methoxybenzene (35 mg, 0.174 mmol) and potassium carbonate (32 g, 0.232 mmol). The reaction was heated at 50 °C for 3 h. The mixture was cooled and partitioned between ethyl acetate and water. The organic layers were combined, dried over sodium sulfate, filtered and concentrated to give a crude product which was purified by preparative HPLC to give l -(2-(2-hydroxyethoxy)ethyl)-7-(4-methoxybenzyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (14 mg, 22.2 % yield) as white solid. 'iZ-NMR (DMSO-flfc) δ 7.61 (t, IH), 7.48(s, lH),7.32-7.44(m, 4H), 6.91 (d, 2H), 5.37(s, 2H), 4.60(t, IH), 4.08(t, 2H), 3.73(s, 3H), 3.58(t, 2H), 3.46(s, 4H), 3.29(s, 3H). LCMS retention time 2.581 min; LCMS MH+ 551.
The following products 32a through 32h were prepared using the method of example 31.
Example 32a 7-(3-chlorobenzyl)-l -(2-(2-hydroxyethoxy)ethyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)- -purine-2,6(3H,7H)-dione
Figure imgf000474_0002
White solid, 16 mg, 25.0 % yield: 'H-NMR (400 MHz, DMSO-i¾ δ = 7.61 (t, IH), 7.49(d, 2H),7.34-7.44(m, 3H), 7.33 (d, 2H), 5.45(s, 2H), 4.59(t, IH), 4.07(t, 2H), 3.57(t, 2H), 3.45(s, 4H), 3.30(s, 3H). LCMS retention time 2.876 min; LCMS MH+ 555.
Example 32b l -(2-(2-hydroxyethoxy)ethyl)-3-methyl-7-(4-methylbenzyl)-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000475_0001
White solid, 23 mg, 25.0 % yield: 'H-NMR (DMSO-ifc) δ 7.61 (t, I H), 7.47(d, IH), 7.42(dd, lH),7.29-7.34(m, 3H), 7.16(d, 2H), 5.77(s, 2H), 4.60(t, I H), 4.07(t, 2H), 3.57(t, 2H), 3.44- 3.47(m, 4H), 3.30(s, 3H), 2.27(s, 3H). LCMS retention time 2.750 min; LCMS MH+ 535.
Example 32c 1 -(2-(2-hydroxyethoxy)ethyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-7-(4- (trifluoromethyl)benzyl)-l H-purine-2,6(3H,7H)-dione
Figure imgf000475_0002
White solid, 19 mg, 27.9 % yield: 'H-NMR (DMSO-< ) δ 7.74(d, 2H), 7.61(t, 3H), 7.44- 7.48(m, 2H),7.33(d, IH) 5.55(s, 2H), 4.58(q, IH), 4.09(t, 2H), 3.56(t, 2H), 3.45(s, 4H), 3.31(s, 3H). LCMS retention time 2.831 min; LCMS MH+ 589.
Example 32d 7-(4-ethylbenzyl)-l -(2-(2-hydroxyethoxy)ethyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-l -purine-2,6(3H,7H)-dione
Figure imgf000475_0003
White solid, 16 mg, 25.4 % yield: !H-NMR (DMSO-rf6) δ 7.61 (t, I H), 7.46(s, lH),7.42(d, IH), 7.31-7.34(m, 3H), 7.19(d, 2H), 5.40(s, 2H), 4.60(t, IH), 4.07(t, 2H), 3.57(t, 2H), 3.47(s, 4H), 3.29(s, 3H), 2.57(q, 2H), 1.14(t, 3H). LCMS retention time 2.897 min; LCMS MH+ 549. Example 32e l -(2-(2-hydroxyethoxy)ethyl)-7-(3-methoxybenzyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy) -1 H-purine-2,6(3H,7H)-dione
Figure imgf000476_0001
White solid, 10 rag, 25.0 % yield: 'H-NMR (DMSO-c¼) δ 7.61(t, 1H), 7.48(s, 1Η),7.40-7.43 (m, 1H), 7.26-7.34(m, 2H),6.87-6.98 (m, 3H), 5.4 l (s, 2H), 4.60(t, 1 H), 4.07(t, 2H), 3.71 (s, 3H), 3.57(t, 2H), 3.46(s, 4H), 3.30(s, 3H). LCMS retention time 2.644 min; LCMS MH+ 551.
Example 32f l-(2-(2-hydroxyethoxy)ethyl)-3-methyI-7-(3-methylbi
(trifluoromethoxy)phenoxy). -purine-2,6(3H,7H)-dione
Figure imgf000476_0002
White solid, 12 mg, 19.6 % yield: 'H-NMR (DMSO-c/6) S 7.6 l(t, 1Η), 7.46(s, lH),7.40(dd, 1H), 7.33 (d, 1H), 7.24(d, 1H), 7.11-7.19(m, 3H), 5.41 (s, 2H), 4.59(t, 1H), 4.07(t, 2H), 3.57(t, 2H), 3.47(s, 4H), 3.30(s, 3H), 2.27(s, 3H). LCMS retention time 2.726 min; LCMS MH+ 535.
Example 32g l-(2-(2-hydroxyethoxy)ethyl)-3-methyl-7-(3-(trifluoromethoxy)benzyl)-8-(3- (trifluoromethoxy)phenoxy)-l -purine-2,6(3H,7H)-dione
Figure imgf000476_0003
White solid, 22mg, 31.4 % yield: 'H-NMR (DMSO-d6) δ 7.61 (t, 1H), 7.53(t, lH),7.41-7.49(m, 4H), 7.33 (d, 2H), 5.50(s, 2H), 4.59(t, 1H), 4.07(t, 2H), 3.56(t, 2H), 3.46(s, 4H), 3.30(s, 3H). LCMS retention time 2.865 min; LCMS MH+ 605. Example 32h l-(2-(2-hydroxyethoxy)ethyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-7-(3- (trifluoromethyl)benzyl)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000477_0001
White solid, 10 mg, 31.4 % yield: 'H-NMR (DMSO-<¾ δ 7.8 l(s, 1 H), 7.58-7.71 (m, 4H), 7.41 -7.46(m, 2H), 7.33 (d, 2H), 5.55(s, 2H), 4.59(t, 1H), 4.07(t, 2H), 3.57(t, 2H), 3.47(s, 4H), 3.30(s, 3H). LCMS retention time 2.795 min; LCMS MH+ 589.
Example 33 l-(2-(2-aminoethoxy)ethyl)-7-(4-chlorobenzyl)-3-methyl-8-(3- (trifluoro
Figure imgf000477_0002
Step 1 1 -(2-(2-bromoethoxy)ethyl)-7-(4-chlorobenzyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-l H-purine-2,6(3H,7H)-dione
Figure imgf000477_0003
To a solution of 7-(4-chlorobenzyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine- 2,6(3H,7H)-dione(120 mg, 0.26 mmol, intermediate 9) in DMF (3 mL) was added l-bromo-2-(2- bromoethoxy)ethane (77 mg, 0.33 mmol), potassium carbonate (71 mg, 0.51 mmol). The reaction was heated at 60°C overnight. The mixture was cooled and partitioned between ethyl acetate and water. The organic layers were combined, dried over sodium sulfate, filtered and concentrated to give crude product (lOOmg, 70% of purity by HPLC), which was used without purification. LCMS retention time 2.054 min; LCMS MH+ 617 Step 2 l -(2-(2-aminoethoxy)ethyl)-7-(4-chlorobenzyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000478_0001
l -(2-(2-bromoethoxy)ethyl)-7-(4-chlorobenzyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)- l H- purine-2,6(3H,7H)-dione (100 mg, 0.16 mmol) was dissolved in a preformed solution of ammonia in methanol (15 mL). The reaction was heated at 60°C overnight in a sealed tube. The reaction was cooled to room temperature and the solvent was evaporated. The residue was purified by preparative HPLC to give l -(2-(2-aminoethoxy)ethyl)-7-(4-chlorobenzyl)-3-methyl- 8-(3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione(35 mg, 39% yield) as white solid. !H-NMR (CD3OD) δ 7.94(s, 3H), 7.62(t, 1H), 7.41-7.49(m, 6H), 7.34(d, 1H), 5.45 (s, 2H), 4.10(1, 2H), 3.65-3.67(m, 4H), 3.30(s, 3H), 2.94(q, 2H).LCMS retention time 2.323 min; LCMS MH+ 554.
Example 34 7-(4-chlorobenzyl)-l-(2-(2-methoxyethoxy)ethyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2:,6(3H,7H)-dione
Figure imgf000478_0002
The title product was prepared using the method of example 33, step 1 with 2-(2- rnethoxyethoxy)ethyl methanesulfonate (51 mg, 0.26 mmol, intermediate 34) as the alkylating agent. White solid, 44 mg, 45% yield: 'H-NMR (CD3OD) S 7.61(t, 1H), 7.50(s, 1H), 7.40- 7.45(m, 5H), 7.33(d, 1H), 5.44(s, 2H), 4.05(t, 2H), 3.51-3.57(m, 4H), 3.36(s, 2H), 3.29(s, 3H), 3.20(s, 3H).LCMS retention time 3.161 min; LCMS MH+ 569.
Example 35 7-(4-chlorobenzyl)-l-(2-hydroxyethyl)-3-methyl-8-(2-(3- (trifluoromethoxy)phenoxy)ethoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000478_0003
To a solution of 7-(4-chlorobenzyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine- 2,6(3H,7H)-dione (120 mg, 0.257mmol) in DMF (5 mL) was added 2-bromoethanol (128.9 mg, 1.028 mmol) followed by cesium carbonate (167 mg, 0.515 mmol). The resulting mixture was microwave irradiated at 120 °C for 30 min. The reaction was partitioned between ethyl acetate and brine. The combined organic phase was dried and concentrated to give a residue, which was purified via preparative HPLC to give 7-(4-chlorobenzyl)-l -(2-hydroxyethyl)-3-methyl-8-(2-(3- (trifluoromethoxy)phenoxy)ethoxy)-l H-purine-256(3H,7H)-dione (17 mg, 34.7 % yield) as white solid. 'H-NMR (DMSO-<¾ δ 7.43(t, 1 H), 7.27-7.32(m,4 H), 6.95-7.03(m, 3 H), 5.20(s, 2 H) , 4.81 (t, 2 H), 4.75(t, 1 H), 4.41 (t,2 H), 3.92-3.95(m,2 H), 3.47-3.52(m,2 H), 3.31(s,3 H). LCMS retention time 2.731 min; LCMS MH+ 555.
Example 36 7-(4-chlorobenzyl)-l-(2-hydroxyethyl)-3-methyl-8-(2-(3- (trifluoromethoxy)phenoxy)ethoxy)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000479_0001
Step 1 7-(4-chlorobenzyl)-3-methyl-l-(2-(tetrahydro-2H-pyran-2-yloxy)ethyl)-8-(2-(3- (trifluoromethoxy)phenoxy)ethoxy)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000479_0002
To a solution of 2-(3-(trifluoromethoxy)phenoxy)ethanol (61 mg, 0.275 mmol, intermediate 5) in anhydrous THF (10 mL) was added sodium hydride (66 mg, 2.75 mmol)at 0 °C under a nitrogen atmosphere. The mixture was stirred at 0 °C for 20 min; then a solution of 8-bromo-7-(4- chlorobenzyl)-3-methyl- l -(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-lH-purine-2,6(3H,7H)- dione (70 mg, 0.137 mmol, intermediate 14) in THF (3 mL) was added dropwise , and the resulting mixture was stirred at 0 °C to room temperature for 16 h under nitrogen atmosphere. The mixture was quenched with ice-water and partitioned between ethyl acetate and brine. The organic layer was dried over sodium sulfate, filtered and concentrated to give a crude product which was purified via silica gel chromatography eluting with petroleum ether/ ethyl acetate (1 :0 to 2: 1) to give 7-(4-chlorobenzyl)-3 -methyl- l -(2-(tetrahydro-2H-pyran-2-yloxy)ethyl)-8-(2- (3-(trifluoromethoxy)phenoxy)ethoxy)-lH-purine-2,6(3H,7H)-dione (70 mg, 78.2%) as yellow oil. LCMS MH+-THP 569. Step 2 7-(4-chlorobenzyl)-l -(2-hydroxyethyl)-3-methyl-8-(2-(3- (trifluoromethoxy)phenoxy)ethoxy)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000480_0001
To a solution of 7-(4-chlorobenzyl)-3-methyl-l-(2-(tetrahydro-2H-pyran-2-yloxy)ethyl)-8-(2-(3- (trifluoromethoxy)phenoxy)ethoxy)-lH-purine-2,6(3H,7H)-dione (70 mg, 0.107 mmol) in methanol (3 mL) was added concentrated HCl (4 drops). The mixture was stirred for 1 h at. room temperature. The mixture was neutralized with saturated sodium bicarbonate, and extracted with ethyl acetate. The organic phase was dried over sodium sulfate, filtered and concentrated to give a crude product which was recrystallized from ethanol to give 7-(4-chlorobenzyl)-l-(2- hydroxyethyl)-3-methyl-8-(2-(3-(trifluoromethoxy)phenoxy)ethoxy)-lH-purine-2,6(3H,7H)- dione (16 mg, 25.6% yield) as white solid. H-NMR (DMSO-c¾5 7.45-7.41 (t, 1H), 7.32- 7.27(m, 4H), 7.03-6.95(m, 3H), 5.19(s, 2H), 4.81 (s, 2H), 4.49-4.46(t, 1H), 4.41 -4.40(d, 2H), 3.91-3.88(t, 2H), 3.44-3.40(m, 5H), 1.69-1.65(t, 2H). LCMS retention time 3.142 min; LCMS MH+ 569.
Example 37 7-(4-chlorobenzyl)-l -(2-hydroxyethyl)-3-methyl-8-(3-(3- trifluoromethox ) henoxy) ro oxy) - 1 H-purine-2,6(3H,7H)-dione
Figure imgf000480_0002
Step 1 7-(4-chlorobenzyl)-3 -methyl- 1 -(2-(tetrahydro-2H-pyran-2-yloxy)ethyl)
(trifluoromethoxy)phenoxy)propoxy)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000481_0001
To a solution of 3-(3-(trifluoromethoxy)phenoxy)propan-l -ol (95 mg, 0.4 mmol, intermediate 35) in THF (5 mL) was added sodium hydride (19 mg, 0.40 mmol) at 0 °C. After stirred at 0 °C for 30 min, 8-bromo-7-(4-chlorobenzyl)-3-methyl-L-(2-(tetrahydro-2H-pyran-2-yloxy)ethyl)-lH- purine-2,6(3H,7H)-dione(0.1 g, 0.20 mmol, intermediate 13) was added. The mixture was allowed to warm to room temperature and stirred for 16 h. Then the reaction was chilled to 0 °C and aqueous ammonium chloride solution (2 mL) was added. The mixture was partitioned between ethyl acetate and water. The organic layer was dried over sodium sulfate, filtered and concentrated to give crude product which was used without purification. LCMS MH -THP 569.
Step 2 7-(4-chlorobenzyl)-l -(2-hydroxyethyl)-3-methyl-8-(3-(3-(trifluoromethoxy)
phenoxy)prop -purine-2,6(3H,7H)-dione
Figure imgf000481_0002
To a solution of 7-(4-chlorobenzyl)-3 -methyl- l -(2-(tetrahydro-2H-pyran-2-yloxy)ethyl)-8-(3 -(3- (trifluoromethoxy)phenoxy)propoxy)-lH-purine-2,6(3H,7H)-dione (50 mg, 0.09 mmol,) in ethanol (5 mL) was added acetyl chloride (0.2 mL) at 0 °C. The mixture was stirred at room temperature for 1 h; then it was partitioned between ethyl acetate and water. The organic phase was separated and dried over sodium sulfate. This organic layer was then filtered and
concentrated to give a crude product which was purified by preparative HPLC to give 7-(4- chlorobenzyl)-l-(2-hydroxyethyl)-3-methyl-8-(3-(3-(trifluoromethoxy) phenoxy)propoxy)lH- purine-2,6(3H,7H)-dione (20 mg, 39.2% yield) as a white solid. "H-NMR (DMSO-<¾ δ 7.40(t, 1 H), 7.28-7.33(m, 4H), 6.92(dd, 2H), 6.85(s, lH),5.23(s, 2H) , 4.76(t, 1H), 4.62(t, 2H), 4.08(t, 2H), 3.93(t, 2H), 3.49(q, 2H), 3.33(s, 3H), 2.18-2.24(m, 2H). LCMS retention time 3.058 min; LCMS MH+ 569. The following examples 38a tlirough 381 were prepared using the methods of examples 36 and/or 37.
Example 38a 8-(2-(3,5-bis(trifluoromethyl)phenoxy)ethoxy)-7-(4-chlorobenzyl)-l -(2- hydroxyethyl)-3-methyl-lH-purine-2,6(3H,7H)-dione
Figure imgf000482_0001
The title product was prepared using intermediate 65. White solid, 49 mg, 44.6 % yield: lH- NMR (DMSO-<fc) δ 7.66(s, 1H), 7.60(s, 2H),7.23-7.29(m, 4H), 5.20(s, 2H) , 4.85(t, 2H), 4.84(t, 1H), 4.55-4.57(m, 2H), 3.94(t, 2H), 3.49(q, 2H), 3.33(s, 3H). LCMS retention time 3.229 min; LCMS MH* 607.
Example 38b 7-(4-chlorobenzyl)-8-(2-(3-chlorophenoxy)ethoxy)-l-(2-hydroxyethyl)-3- methyl- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000482_0002
The title product was prepared using intermediate 66. White solid, lOmg, 22.2 % yield: lH-
NMR '(DMSO-flfe) £ 7.30-7.34(m, 5H), 7.01-7.04(m, 2H),6.92-6.94(m, 1H), 5.19(s, 2H), 4.74-
4.80(m, 3H), 4.37-4.40 (m, 2H), 3.93(t, 2H), 3.49(q, 2H), 3.34(s, 3H). LCMS retention time
2.837 min; LCMS MH+ 505.
Example 38c 7-(4-chIorobenzyl)-l -(2-hydroxyethyl)-3-methyl-8-(2-(m-tolyloxy)ethoxy)-lH- purine-2,6(3H,7H)-dione
Figure imgf000483_0001
The title product was prepared using intermediate 67. White solid, 12 mg, 27.9 % yield: H- NMR (DMSO-rftf) δ 7.27-7.32(m, 4H), 7.15-7.19(m, lH),6.74-6.79(m, 3H), 5.19(s, 2H) , 4.74- 4.80(m, 3H), 4.32-4.34(m, 2H), 3.94(t, 2H), 3.49(q, 2H), 3.35(s, 3H), 2.27(s, 3H). LCMS retention time 2.676 min; LCMS MH+ 485.
Example 38d 7-(4-chlorobenzyl)-l -(2-hydroxyethyl)-3-methyl-8-(2-(3- (trifluoromethyl)phenoxy)ethoxy)-ll-I-purine-2,6(3H,7H)-dione
Figure imgf000483_0002
The title product was prepared using intermediate 68. White solid, 10 mg, 20.8 % yield: lH- NMR (DMSO-i¾ δ 7.54(t, IH), 7.24-7.33(m, 7H),6.74-6.79(m, 3H), 5.19(s, 2H), 4.81-4.83(m, 2H), 4.75(t, IH), 4.45-4.46(m, 2H), 3.94(t, 2H), 3.49(q, 2H), 3.35(s, 3H). LCMS retention time 2.857 min; LCMS MH+ 539.
Example 38e 8-(2-(4-chloro-3-(trifluoromethyl)phenoxy)ethoxy)-7-(4-chlorobenzyl)-l -(2- hydroxyethyl)-3-methyl-lH-purine-2,6(3H,7H)-dione
Figure imgf000483_0003
The title product was prepared using intermediate 71. White solid, 30 mg, 49.2% yield: λΗ- NMR (DMSO-i¾ 5 7.64-7.62(d, I H), 7.31 -7.26(m, 6H), 5.20(s, 2H), 4.83-4.80(m, 2H), 4.76- 4.73(t, IH), 4.46-4.44(m, 2H), 3.95-3.92(t, 2H), 3.52-3.49(m, 2H), 3.47(s, 3H).LCMS retention time 2.973min; LCMS MH+ 573 Example 38f 7-(4-chlorobenzyl)-l-(2-hydroxyethyl)-3-methyl-8-(2-(2- (trifluoromethoxy)phenoxy)ethoxy)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000484_0001
The title product was prepared using intermediate 70. White solid, 18 mg, 29.7% yield: XH- NMR (DMSO-<¾ 6 7.36-7.34(d, 2H), 7.29-7.24(m, 5H), 7.07-7.03(t, 1H), 5.17(s, 2H), 4.85- 4.83(m, 2H), 4.77-4.74(t, 1 H), 4.46-4.44(m, 2H), 3.95-3.92(t, 2H), 3.51 -3.48(m, 2H), 3.47(s, 3H).LCMS retention time 2.828 min; LCMS MH+ 555.
Example 38g 7-(4-chlorobenzyl)-l-(2-hydroxyethyl)-3-methyl-8-(2-(4- (trifluoromethoxy)phenoxy)e
Figure imgf000484_0002
The title product was prepared using intermediate 69. White solid, 30 mg, 49.4% yield: 1H-
NMR (DMSO-ί ί) δ 7.32-7.26(m, 6H), 7.06-7.03(m, 2H), 5.19(s, 2H) , 4.82-4.80(m, 2H), 4.76- 4.73(t, 1H), 4.38-4.36(m, 2H), 3.95-3.92(t, 2H), 3.52-3.47(m, 2H), 3.39(s, 3H).LCMS retention time 2.908min; LCMS MH+ 555. Example 38h 7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl-8-(3-(3- (trifluoromethoxy)phenyl)propoxy) -lH-purine-2,6(3H,7H)-dione
Figure imgf000484_0003
The title product was prepared using the methods of examples 36 and 37, and using intermediate 37 in the first step. White solid, 12 mg, 21.4% yield: 'H-NMR (CD3OD) δ 7.35-7.49(m, 5 H), 7.07-7.17(m, 3 H), 5.28(s, 2 H), 4.54(t, 2 H), 4.10(t, 2 H), 3.67(t, 2 H), 3.45(s, 2 H), 2.75(t, 2 H), 2.1 1 -2.18(m, 2 H), 1 .83-1.91 (m, 2 H). LC S retention time 3.125 min; LCMS MH+ 567.
Example 38i 7-(4-chlorobenzyl)-l -(3-hydroxypropyl)-3-methyl-8-(3-
Figure imgf000485_0001
The title product was prepared using the methods of examples 36 and 37, and using 3- trifluoromethoxy-benzyl alcohol in the first step. White solid, 12 mg, 16.0 % yield): 7/-NMR (DMSO- ¾) δ 7.27-7.55 (m, 8H), 5.57(s, 2H), 5.28(s, 2H), 4.47(t, 1H), 3.90(t, 2H), 3.56- 3.45(m, 5H), 1.65-1.90(m, 2H). LCMS retention time 2.870 min; LCMS MH+ 539.
Example 38j 7-(4-chJorobenzyl)-l -(3-hydroxypropyl)-3-methyl-8-((5-methylthiazol-2- l methox - 1 H- urine-2 6 3H 7H -dione
Figure imgf000485_0002
The title compound was prepared using the methods of examples 36 and 37, and using (5- methylthiazol-2-yl)methanol (intermediate 4, step 1) in the first step. White solid, 46 mg, 54.1 % yield: 'H-NMR (CDC13) δ 7.50(d,l H), 7.33-7.32(m, 2 H), 7.26-7.28(m, 2 H), 5.74(s, 2 H) , 5.27(s, 2 H), 4.19(t, 2 H), 3.61(t, 1 H), 3.51-3.55(m, 5 H), 2.51(d, 3 H), 1.88- 1.92(m, 2 H). LCMS retention time 2.366 min; LCMS MH+ 476.
Example 38k 7-(4-chlorobenzyl)-l -(3-hydroxypropyl)-3-methyl-8-((5-methyloxazol-2- yl)methoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000485_0003
The title compound was prepared using the methods of examples 36 and 37 and with (5- methyloxazol-2-yl)methanol (intermediate 40). The crude product was purified via preparative HPLC to give 7-(4-chlorobenzyl)- l -(3-hydroxypropyl)-3-methyl-8-((5-methyloxazol-2- yl)methoxy)-l H-purine-2,6(3H,7H)-dione (17 mg, 40.9%) as white solid. 'H-NMR (DMSO- d6) δ 7.38-7.40 (d, 2H), 7.26-7.28 (d, 2H), 6.90 (s, IH), 5.58 (s, 2H), 5.23 (s, 2H), 4.45-4.48 (t, IH), 3.87-3.91 (t, 2H), 3.39-3.44 (t, 2H), 2.28 (s, 3H), 1.65-1.68 (m, 2H). LCMS retention time 2.284 min; LCMS MH+ 460.
Example 381 7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl-8-((5-methylthiazol-2- yl)methoxy)-lH-purine-2,6(3H,7 -dione
Figure imgf000486_0001
The title compound was prepared using the methods of examples 36 and 37 and with (5- methyloxazol-2-yl)methanol (intermediate 40). White solid, 46 mg, 54.1 % yield: 'H-NMR (DMSO-4 δ 7.37-7.39 (d, 2Η), 7.26-7.28 (d, 2Η), 6.90 (s, 1Η), 5.58(s, 2Η), 5.23(s, 2Η), 4.73- 4.76 (t, 1Η), 3.93-3.95 (t, 2Η), 3.47-3.51 (m ,2H), 3.43(s, 3H), 2.28(s, 3H). LCMS retention time 2.194 min; LCMS MH+ 446.
Example 39 7-(4-chlorobenzyl)-8-(2-(3-((dimethylamino)methyl)phenoxy)ethoxy)-l-(2- hydroxyethyl)-3-methyl-lH-purine-2,6(3H,7H)-dione hydrochloride
Figure imgf000486_0002
Step 1 3-(2-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-l -(2-(tetrahydro-2H-pyran-2-yloxy)ethyl)- 2,3,6,7-tetrahydro-lH-purin-8-yloxy)ethoxy)benzaldehyde
Figure imgf000487_0001
The title compound was prepared using the method of example 36, step 1 and using intermediate 36. The product was purified by silica gel chromatography eluting with petroleum/ ethyl acetate (3: 1 to 1 : 1) to give 3-(2-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-l -(2-(tetrahydro-2H-pyran-2- yloxy)ethyl)-2,3,6,7-tetrahydro-lH-purin-8-yloxy)ethoxy)benzaldehyde (64 mg, 36.4% yield) as yellow syrup. LCMS MH+-THP 499.
Step 2 7-(4-chlorobenzyl)-8-(2-(3-((dimethylamino)methyl)phenoxy)ethoxy)-3-methyl-l -(2- (tetrahydro-2H-pyran-2-yloxy)ethyl)-lH-purine-2,6(3H,7H)-dione
Figure imgf000487_0002
To a solution of 3-(2-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo- l -(2-(tetrahydro-2H-pyran-2- yloxy)ethyl)-2,3,6,7-tetrahydro-lH-purin-8-yloxy)ethoxy)benzaldehyde (100 mg, 0.171 mmol) in methanol (3 mL) and THF (3 mL) was added acetic acid (0.2 ml, 3.49 mmol) and
dimethylamine hydrochloride (90 mg, 1.10 mmol) . The mixture was stirred at room temperature for 1 h. Then the mixture was cooled to 0 °C, sodium cyanoborohydride (16.1 mg, 0.256 mmol) was added under a nitrogen atmosphere. The resulting mixture was stirred and allowed to warm to room temperature overnight. The mixture was concentrated and purified via silica gel chromatography eluting with DCM/ methanol (60: 1) to give7-(4-chlorobenzyl)-8-(2-(3- ((dimethylamino)methyl)phenoxy)ethoxy)-3-methyl-l-(2-(tetrahydro-2H-pyran-2-yloxy)ethyl)- lH-purine-2,6(3H,7H)-dione (26.1 mg, 24.9%) as white solid. LCMS MH+ 613.
Step 3 7-(4-chlorobenzyl)-8-(2-(3-((dimethylamino)methyl)phenoxy)ethoxy)-l -(2- hydroxyethyl)-3-methyl- 1 H-purine-2,6(3H,7H)-dione hydrochloride
Figure imgf000488_0001
The title compound was prepared using the method of example 37, step 2 to give 7-(4- chlorobenzyl)-8-(2-(3-((dimethylamino)methyl)phenoxy)ethoxy)-l-(2-hydroxyethyl)-3-methyl- l H-purine-2,6(3H,7H)-dione hydrochloride (13.6 mg, 65.2% yield) as white solid. 'H-NMR (DMSO-<¾ 5 10.1 1(bs, 1H), 7.42-7.38(t, 1H), 7.33-7.28(m, 4H), 7.18(s, 1H), 7.11-7.09(d, 1H), 7.06-7.04(m, 1H), 5.20(s, 2H), 4.85-4.83(m, 2H), 4.76-4.73(t, 1H), 4.39-4.37(m, 2H), 4.22(s, 2H), 3.96-3.92(t, 2H), 3.52-3.46(m, 2H), 3.29(s, 3H), 2.69(s, 6H). LCMS retention time 1.678 min; LCMS MH+ 553. .
Example 40 7-(4-chlorobenzyl)-l -(2-(dimethylamino)ethyl)-3-methyl-8-(2-(3- (trifluoromethoxy)phenoxy)ethoxy)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000488_0002
Step 1 8-bromo-7-(4-chlorobenzyl)-l -(2-(dimethylamino)ethyl)-3 -methyl- lH-purine- 2,6(3H,7H)-dione
Figure imgf000488_0003
To a solution of 8-bromo-7-(4-chlorobenzyl)-3-methyl-lH-purine-2,6(3H,7H)-dione(0.2 g, 0.54 mmol, intermediate 8) in DMF (5 mL) was added 2-chloro-N,N-dimethylethanamine (0.87 mg,
0.83 mmol), potassium carbonate (0.15 g,1.08mmol), and TBAI (2 mg, 0.02 mmol). The reaction was heated at 80 °C overnight. The mixture was cooled and partitioned between ethyl acetate and water. The combined organic layer was dried over sodium sulfate, filtered and concentrated to give a crude product (0.2 g, 84 %) as yellow oil, which was used without purification. LCMS MH+ 442.
Step 2 7-(4-chlorobenzyl)-l-(2-(dimethylamino)ethyl)-3-methyl-8-(2-(3- (trifluoromethoxy)phenoxy)ethoxy)-lH- urine-2,6 3H,7H)-dione
Figure imgf000489_0001
The title compound was prepared using the method of example 36, step 1 and purified by preparative HPLC to give 7-(4-chlorobenzyl)-l-(2-(dimethylamino)ethyl)-3-methyl-8-(2-(3- (triiluoromethoxy)phenoxy)ethoxy)-lH-purine-2,6(3H,7H)-dione (20 mg, 16.4% yield) as white solid. lB-NMR (DMSO-d6) δ 8.18(s. I H), 7.43(t, Hi), 7.26-7.32(m, 4H), 6.93-7.02(m, 3H), 5.20(s, 2H), 4.80-4.82(m, 2H), 4.39-4.41(m, 2H), 3.99(t, 2H), 3.67(s, 3H), 2.58(t, 2H), 2.30(s, 6H). LCMS retention time 2.352 min; LCMS MH+ 582
Example 41 7-(4-chlorobenzyl)-3-methyl-l-propyl-8-(2-(3- (trifluoromethoxy)phenoxy)ethoxy)-l H-purine-2,6(3H,7H)-dione
Figure imgf000489_0002
The title compound was prepared using the 2 step method of example 40 and purified via preparative HPLC to give 7-(4-chlorobenzyl)-3 -methyl- l-propyl-8-(2-(3- (trifluoromethoxy)phenoxy)ethoxy)- 1 H-purine-2,6(3H,7H)-dione (80 mg, 29.7% yield) as white solid. 'H-NMR (DMSO-flfc) δ 7.45-7.41 (t, IH), 7.32-7.26(m, 4H), 7.02-6.94(m, 3H), 5.19(s, 2H), 4.82-4.79(m, 2H), 4.41 -4.39(m, 2H), 3.81-3.78(t, 2H), 1.58-1.49(m, 2H), 0.86-0.83(t, 3H). LCMS retention time 3.571 min; LCMS MH+ 553 Example 42 7-((5-chloropyridin-2-yl)methyl)-l -(3-hydroxypropyl)-3-methyl-8-(2-(3- (trifluoromethoxy)phenoxy)ethoxy)-l H-purine-2,6(3H,7H)-dione
Figure imgf000490_0001
Step 1 7-((5-chloropyridin-2-yl)methyl)-3-methyl- 1 -(3-(tetrahydro-2H-pyran-2-yloxy)propyl)- -(2-(3-(trifluoromethoxy)phenoxy)ethoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000490_0002
To a solution of 3-methyl-l-(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-8-(2-(3- (trifluoromethoxy)phenoxy)ethoxy)-lH-purine-2,6(3H,7H)-dione (60 mg, 0.14 mmol, intermediate 17) in DMF (5 mL) was added 5-chloro-2-(chloromethyl)pyridine (33 mg, 0.20 mmol), potassium carbonate (37 mg, 0.27 mmol) and TBAI (2 mg, 0.02 mmol). The reaction was heated at 50 °C for 8 h. The mixture was cooled and partitioned between ethyl acetate and water. The combined organic layer was dried over sodium sulfate, filtered and concentrated to give crude product, which was used without purification. LCMS MH+-THP 570.
Step 2 7-((5-chloropyridin-2-yl)methyl)-l-(3-hydroxypropyl)-3-methyl-8-(2-(3- (trifluoromethoxy)phenoxy)ethoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000490_0003
To a solution of 7-((5-chloropyridin-2-yl)methyl)-3-methyl-l -(3-(tetrahydro-2H-pyran-2- yloxy)propyl)-8-(2-(3-(trifluoromethoxy)phenoxy)ethoxy)-lH-purine-2,6(3H,7H)-dione (80mg, 0.14 mmol) in ethanol (5 mL) was added acetyl chloride (0.2 mL) at 0 °C The mixture was stirred at room temperature for 1 h. The mixture was partitioned between ethyl acetate and water. The organic phase was dried over sodium sulfate, filtered and concentrated to give crude product which was purified by preparative HPLC to give 7-((5-chloropyridin-2-yl)methyl)-l -(3- hydroxypropyl)-3-methyl-8-(2-(3-(trifluoromethoxy)phenoxy)ethoxy)-lH-purine-2,6(3H,7H)- dione (15 mg, 19.0% yield) as white solid. ' -NMR (CD3OD) δ 8.33(d, 1 H), 7.70(dd, 1H), 7.35(t, 1H), 7.26(d, 1H), 6.87(dd, 1H), 6.76(s, 1H), 5.43(s, 2H), 4.86-4.87(m, 2H), 4.33-4.35(m, 2H), 4.04(t, 2H), 3.56(t, 2H), 3.54(s, 3H), 1.80-1.85(m, 2H). LCMS retention time 2.606 min; LCMS MH+ 570.
The following examples 43a through 43e were prepared using the method of example 42.
Example 43a l-(3-hydroxypropyl)-3-methyl-7-((6-methylpyridin-3-yl)methyl)-8-(2-(3- (trifluoromethoxy)phenoxy)e
Figure imgf000491_0001
White solid, 20 mg, 26.3% yield: '/J-NMR (CD3OD) δ 8.55(s, 1H), 7.95(d, 1H), 7.39(dd, 2H),6.96(dd, 1H), 6.90(d, 1H), 6.81 (s, 1H), 5.37(s, 2H), 4.93-4.95(m, 2H), 4.43-4.45(m, 2H), 4.07(t, 2H), 3.61(t, 2H),3.50(s, 3H), 2.55(s, 3H), 1.84-1.88(m, 2H). LCMS retention time 2.022 min; LCMS MH+ 550.
Example 43b 7-benzyl-l-(3-hydroxypropyl)-3-methyl-8-(2-(3- (trifluoromethoxy)phenoxy)e
Figure imgf000491_0002
White solid, 17 mg, 23.0% yield: JH-NMR (DMSO-<¾ δ 7.43(t, 1H), 7.23-7.32(m, 5H), 6.95- 7.04(m, 3H), 5.20(s, 2H), 4.80-4.82(m, 2H), 4.70(t, 1H), 4.40-4.43(m, 2H), 3.90(t, 2H), 3.33- 3.44(m, 5H), 1.65-1.70(m, 2H). LCMS retention time 2.897 min; LCMS MH+ 535.
Example 43c 1 -(3-hydroxypropyl)-3-methyl-7-((5-methylthiazol-2-yl)melhyl)-8-(2-(3- (trifluoromethoxy)phenoxy)eth
Figure imgf000491_0003
White solid, 20 mg, 43.2% yield: 'H-NMR (CD3OD) δ 7.39-7.35(t, 1H), 7.28(s, l H), 6.93- 6.80(m, 2H), 6.79(s, 1 H), 5.56(s, 2H), 4.89(s, 2H), 4.39-4.36(dd, 2H), 4.10-4.06(t, 2H), 3.61 - 3.58(t, 2H), 3.53(s, 3H), 2.35(s, 3H), 1 .90-1.85(m, 2H). LCMS retention time 2.536 min; LCMS MH+ 556.
Example 43d 7-ethyl-l-(3-hydroxypropyl)-3-methyl-8-(2-(3- (trifluoromethoxy)phenoxy)ethoxy)-lH-purine-2,6(3H,7H)-dione
0 Γ"
White solid, 33 mg, 37.1 % yield: V/-NMR (CD3OD) δ 7.39(t, 1 H), 7.01 (dd, 1H), 6.88- 6.90(m, 2H), 4.87-4.88(m, 2H), 4.44-4.46(m, 2H), 4.08-4.16(m, 4H), 3.61 (t, 2H), 3.51 (s, 3H), 1.80-1.90 (m, 2H), 1.32(t, 3H).LCMS retention time 1.526 min; LCMS MH+ 473.
Example 43e 1 -(2-hydroxyethyl)-3,7-dimethyl-8-(2-(3-(trifluoromethoxy)phenoxy)ethoxy)- lH-purine-2,6(3H,7H)-dione
Figure imgf000492_0001
White solid, 16 mg, 50.1% yield: ^-NMR (DMSO-^) δ 7.41-7.45(t, lH), 7.04-7.06(dd, 1 H), 6.97-7.00(m, 2H), 4.75-4.79(m, 4H), 4.42-4.44(t, 2H), 3.92-3.95 (t, 2H), 3.57 (s, 3H), 3.50-3.52 (t, 2H), 3.47 (s, 3H). LCMS retention time 2.411 min; LCMS MH+ 445.
Example 44 l-(3-hydroxypropyl)-3-methyl-7-((4-methylthiazol-2-yl)methyl)-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000492_0002
To a solution of l-(3-hydroxypropyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-l H-purine- 2,6(3H,7H)-dione (100 mg, 0.25 mmol, intermediate 13) in DMF (5 mL) was added 2- (chloromethyl)-4-methylthiazole (47.9 mg, 0.32 mmol, intermediate 54) followed by potassium carbonate (51.6 mg, 0.37 mmol) and a catalytic amount of TBAI. The mixture was stirred at 60 °C for 4 h. The mixture was diluted with ethyl acetate and extracted with brine and saturated aqueous ammonium chloride solution. Then the organic phase was dried and concentrated to give a crude product which was purified by preparative HPLC to give l -(3-hydroxypropyl)-3- methyl-7-((4-methylthiazol-2-yl)methyl)-8-(3-(trifluoromethoxy)phenoxy)-lH-purine- 2,6(3H,7H)-dione (29 mg, 22.7% yield) as white solid. 'H-NMR DMSO-d6) δ 7.62 (t, IH), 7.41 -7.46 (m, 2H), 7.33 (dd, IH), 7.26 (d, IH), 5.73(s, 2H), 4.49 (t, IH), 3.91(t, 2H), 3.35(t, 2H), 3.31 (s, 3H),2.29(d, 3H), 1.67- 1.71 (m. 2H). LCMS retention time 2.681 min; LCMS MH+ 512.
The following examples 45a through 45m were prepared using the method of example 44.
Example 45a l-(3-hydroxypropyl)-3-methyl-7-(thiazol-2-ylmethyl)-8-(3- (trifiuoromethoxy)phenoxy)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000493_0001
White solid, 42 mg, 38.1% yield: 'H-NMR (DMSO-tfc) δ 7.74-7.77(m, 2H)7.62 (t, IH), 7.41- 7.49(m, 2H), 7.33(d, IH), 5.80(s, 2H) , 4.47(t, IH), 3.91 (t, 2H), 3.43(t, 2H), 3.3 l (s, 3H),2.29(d, 3H), 1. -65- 1.70(111, 2H). LCMS retention time 2.378 min; LCMS MH+ 498. Example 45b l -(3-hydroxypropyl)-3-methyl-7-((5-methylthiazol-2-yl)methyl)-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000493_0002
White solid, 22 mg, 28.9% yield: 'H-NMR (DMSO-4 δ 7.70 (s, IH), 7.63 (t, IH), 7.56 (s, IH), 7.47-7.50(m, IH), 7.34-7.37(m, I H), 5.60(s, 2H), 4.50 (t, IH), 3.95(t, 2H), 3.46(t, 2H), 3.30(s, 3H),2.59(s, 3H), 1.67-1.74(m, 2H). LCMS retention time 2.419 min; LCMS MH+ 512. Example 45c 7-(4-fluorobenzyl)-l-(3-hydroxypropyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)- 1 H
Figure imgf000494_0001
White solid, 23 mg, 30.2% yield: 'H-NMR (CD3OD) δ 7.46-7.55 (m, 3Η), 7.30 (d, 2Η), 7.22 (d, 1Η), 7.07 (t, 2Η), 5.48(s, 2Η) , 4.10 (t, 1Η), 3.60(t, 2Η), 3.40(s, 3Η), 1.84-1.90(m, 2Η). LCMS retention time 2.944 min; LCMS MH+ 509.
Example 45d l-(3-hydroxypropyl)-3-methyl-7-(4-methylbenzyl)-8-(3- (trifluoromethoxy)phenoxy)- 1 H-
Figure imgf000494_0002
White solid, 28 mg, 37.3% yield: 'H-NMR (CD3OD) δ 7.51 (t, 1Η), 7.19-7.31 (m, 4Η), 7. (d, 2Η), 5.44(s, 2Η) , 4.10 (t, 1Η), 3.60(t, 2Η), 3.40(s, 3Η), 2.28(s, 3Η), 1.83-1.90(m, 21 LCMS retention time 3.075 min; LCMS MH+ 505.
Example 45e 4-((l-(3-hydroxypropyl)-3-methyl-2,6-dioxo-8-(3-(trifluoromethoxy)ph^
2,3-dihydro-lH-purin-7(6H)-yl)methyl)benzonitrile
Figure imgf000494_0003
White solid, 16 mg, 20.7 % yield: 'H-NMR (CD3OD) S 7.72 (d, 2Η), 7.51-7.59 (m, 3Η), 7.32 (d, 2Η), 7.22 (d, 1Η), 5.58(s, 2Η) , 4.08(t, 1Η), 3.59(t, 2Η), 3.41(s, 3Η), 1.82-1.87(m, 2Η). LCMS retention time 2.750 min; LCMS MH+ 516. Example 45f 1 -(3-hydroxypropyl)-7-(4-methoxybenzyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000495_0001
White solid, 22 mg, 28.2% yield: V/-NMR (CD3OD) δ 7.52 (t, 2H), 7.38 (d, 2H), 7.20-7.30 (m, 3H), 6.87 (d, 2H), 5.41(s, 2H) , 4.10(t, IH), 3.75(s, 3H), 3.60(t, 2H), 3.40(s, 3H), 1.83- 1.89(m, 2H). LCMS retention time 2.899 min; LCMS MH+ 521.
Example 45g 1 -(3-hydroxypropyl)-3-methyl-7-((6-methylpyridin-3-yl)methyl)-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000495_0002
White solid, 45 mg, 29.7% yield: !H-NMR (DMSO-cfc) δ 8.53-8.52(d, IH), 7.71-7.69(dd, IH), 7.63-7.59(t, I H), 7.52(s, IH), 7.46-7.44(dd, IH), 7.34-7.32(d, IH), 7.25-7.23(d, IH), 5.43(s, 2H), 4.53(bs, IH), 3.95-3.91(t, 2H), 3.46-3.43(t, 2H), 3.28(s, 3H), 2.43(s, 3H), 1.71-1.68(m, 2H). LCMS retention time 1.923 min; LCMS MH+ 506.
Example 45h 7-((5-chloropyridin-2-yl)methyl)-l-(3-hydroxypropyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000495_0003
White solid, 49 mg, 37.3% yield: 'H-NMR DMSO-d6) δ 8.54(s, IH), 7.94-7.97(dd, I H), 7.57- 7.61(t, IH), 7.49-7.51(d, IH), 7.38-7.43(m, 2H), 7.30-7.32(d, IH), 5.58(s, 2H), 4.44-4.47(t, IH), 3.85-3.89(t, 2H), 3.40-3.42(t, 2H), 3.30(s, 3H), 1.63-1 .66(m, 2H). LCMS retention time 2.710 min; LCMS MH+ 526. Example 45i 7-((6-chloropyridin-3-yl)methyl)-l -(3-hydroxypropyl)-3-methyl-8-(3- (trifluoromethoxy)ph dione
Figure imgf000496_0001
White solid, 20 mg, 22.4% yield: 'H-NMR (DMSO-<¾ δ 8.50-8.49(d, IH), 7.90-7.87(dd, IH), 7.63-7.58(t, IH), 7.54-7.45(m, 3H), 7.33-7.3 l(d, IH), 5.48(s, 2H), 3.94-3.90(1, 2H), 3.62-3.59(1, 2H), 3.28(s, 3H), 1.71 -1.67(t, 2H). LCMS retention time 2.625 min; LCMS MH+ 526.
Example 45j 7-ethyl-l -(3-hydroxypropyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH- purine-2,6(3H,7H)-dione
Figure imgf000496_0002
White solid, 40 mg, 54.9% yield: 'H-NMR (DMSO-d6) δ 7.64-7.60(t, IH), 7.55(s, IH), 7.47- 7.45(dd, IH), 7.34-7.32(d, IH), 4.25-4.20(m, 2H), 3.95-3.91(t, 2H), 3.46-3.42(m, 2H), 3.28(s, 3H), 1.72-1.68(m, 2H), 1.39-1.35(t, 3H). LCMS retention time 2.525 min; LCMS MH+ 429. Example 45k l-(3-hydroxypropyl)-3,7-dimethyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine- 2,6(3H,7H)-dione
Figure imgf000496_0003
White solid, 40 mg, 55.1% yield: ^- -NMR (DMSO-i¾ δ 7.64-7.60(t, IH), 7.54(s, I H), 7.47 7.45(dd, IH), 7.32-7.31 (dd, IH), 4.49-4.46(1, IH), 3.94-3.90(t, 2H), 3.78(s, 3H),3.46-3.42(m, 2H), 3.28(s, 3H), 1 1 .73-1.66(m, 2H). LCMS retention time 2.363 min; LCMS MH+ 415.
Example 451 l-(3-hydroxypropyl)-3-methyl-7-((2-methylpyrimidin-5-yl)methyl)-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000497_0001
Light yellow solid, 18 mg, 20.9% yield: 'H-NMR (DMSO-<¾ δ 8.76(s, 2H), 7.64-7.58(m, 2H), 7.50-7.48(m, IH), 7.35-7.32(dd, I H), 5.45(s, 2H), 3.94-3.91(t, 2H), 3.46-3.42(m, 2H), 3.29(s, 3H), 2.96(s, 3H), 1.71 -1.68(m, 2H). LCMS retention time 2.226 min; LCMS MH+ 507.
Example 45m l-(3-hydroxypropyl)-3-methyl-7-((5-methyloxazol-2-yl)methyl)-8-(3- (trifluoromethoxy)phenox - lH-purine-2,6(3H,7H)-dione
Figure imgf000497_0002
Light yellow solid, 15 mg, 20.2% yield: XH-NMR (DMSO-i¾ δ 7.64-7.60(t, IH), 7.45-7;40(m, 2H), 7.34-7.32(d, IH), 6.78(s, IH), 5.59(s, 2H), 4.50(bs, IH), 3.91-3.88(t, 2H), 3.48(s, 2H),
3.3 l(s, 3H), 2.25(s, 3H), 1.71 -1.64(m, 2H). LCMS retention time 2.358 min; LCMS MH+ 496.
Example 46 7-(4-((dimethylamino)methyl)benzyl)-l -(3-hydroxypropyl)-3-methyl-8-(3-
Figure imgf000497_0003
Step 1 7-(4-(bromomethyl)benzyl)-l -(3-hydroxypropyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)- 1 H- urine-2,6(3H,7 -dione
Figure imgf000497_0004
The title compound was prepared using the method of example 44 to give 7-(4- (bromomethyl)benzyl)- l -(3-hydroxypropyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH- purine-2,6(3H,7H)-dione (126 mg, 43.3% yield) as white solid. LCMS retention time 1.758 min; LCMS MH+ 583.
Step 2 7-(4-((dimethylamino)methyl)benzyl)-l -(3-hydroxypropyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-l H-purine-2,6(3H,7H)-dione
Figure imgf000498_0001
The title compound was prepared using the method of example 25 step 2 and purified via preparative HPLC to give 7-(4-((dimethylamino)methyl)benzyl)-l -(3-hydroxypropyl)-3-methyl- 8-(3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H57H)-dione (15 mg, 35.3% yield) as white solid. 1H-NMR (DMSO-c β) δ 10.94(s, IH), 7.62-7.57(m, 3H), 7.50(s, I H), 7.46-7.44(m, 2H), 7.42-7.39(dd, I H), 7.33-7.3 l(d, I H), 5.48(s, 2H), 4.25-4.24(d, 2H), 3.95-3.91(1, 2H), 3.45-3.42(t, 2H), 3.30(s, 3H), 2.65-2.64(d, 6H), 1.71-1.67(m, 2H). LCMS retention time 3.1 83 min; LCMS MH+ 548. Example 47 7-benzyl-3-ethyl-l -(isoxazol-5-ylmethyl)-8-(3-(trifluoromethoxy)phenoxy)-lH- purine-2,6(3H,7I-I)-
Figure imgf000498_0002
To a solution of 7-benzyl-3-ethyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (40 mg, 0.090 mmol, intermediate 47) in DMF (3 mL) was added 5-(chloromethyl)isoxazole (30 mg, 0.256 mmol), followed by potassium carbonate (50 mg, 0.362 mmol) and TBA1 (5 mg, 0.014 mmol). The reaction was stirred at 60 °C overnight. The mixture was cooled and partitioned between ethyl acetate and water. The organic phase was washed with brine, dried over sodium sulfate, and concentrated to give a crude product, which was purified by preparative HPLC to give 7-benzyl-3-ethyl-l -(isoxazol-5-ylmethyl)-8-(3-(trifluoromethoxy)phenoxy)-lH- purine-2,6(3H,7H)-dione (5.9 mg, 12.6% yield) as white solid. 'H-NMR (CD3OD) δ 8.33- 8.32(d, I H), 7.57-7.53(1, 1H), 7.47-7.45(d, 2H), 7.38-7.31 (m, 5H), 7.25-7.23(d, 1 H), 6.34- 6.33(dd, IH), 5.52(s, 2H), 5.36(s, 2H), 4.06-4.01 (q, 2H), 1.27-1.24(t, 3H). LCMS retention time 3.269min; LCMS MH+ 528. Example 48 7-(4-chloro-3-methoxybenzyl)-l -(3-hydroxypropyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-l H-purine-2,6(3H,7H -dione
Figure imgf000499_0001
Figure imgf000499_0002
Step ί 7-(4-chlorobenzyl)-8-(3-hydroxyprop-l -yny])-3-methyl-l -(3-(tetrahydro-2H- pyran-2-yloxy)propyl)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000499_0003
To a solution of 8-bromo-7-(4-chlorobenzyl)-3-methyl-l -(3-(tetrahydro-2H-pyran-2- yloxy)propyl)-l H-purine-2,6(3H,7H)-dione (300 mg, 0.588 mmol, intermediate 14) in anhydrous DMF (12 mL) was added prop-2-yn-l-ol (0.15 ml, 2.545 mmol),
bis(triphenylphosphine)palladium(II) chloride (10 mg, 0.043 mmol), copper iodide (10 mg, 0.053 mmol) (45 mg, 0.402 mmol) and TEA (3 mL, 21.6 mmol). Then the mixture was degassed and refilled with nitrogen 3 times. The reaction was stirred at 85 °C overnight under nitrogen. The mixture was cooled, filtered and the filter cake was washed with ethyl acetate. The filtrate was concentrated to give a crude product, which was purified by silica gel chromatography eluting with DCM / ethyl acetate (3: 1 to 1 : 1) to give 7-(4-chlorobenzyl)-8-(3-hydroxyprop-l - ynyl)-3-methyl-l-(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-lH-purine-2,6(3H,7H)-dione (170 mg, 59.5% yield) as a light yellow solid. LCMS retention time 1.523 min; LCMS MH+ 487.
Step 2 7-(4-chlorobenzyl)-8-(3-hydroxypropyl)-3-methyl-l-(3-(tetrahydro-2H-pyran-2- yloxy)propyl)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000500_0001
A solution of 7-(4-chlorobenzyl)-8-(3-hydroxyprop-l-ynyl)-3-methyl-l -(3-(tetrahydro-2H- pyran-2-yloxy)propyl)- l H-purine-2,6(3H,7H)-dione (120 mg, 0.247 mmol) in methanol (15 mL) was degassed and refilled with nitrogen for 3 times; then 10% Pd/C (30 mg) was added. The mixture was degassed and refilled with hydrogen 3 times and stirred under 45 psi of hydrogen at room temperature for 20 min. The mixture was filtered and the filter cake was washed with methanol. The filtrate was concentrated to give 7-(4-chlorobenzyl)-8-(3-hydroxypropyl)-3- methy[-l-(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-lH-purine-2,6(3H,7H)-dione (100 mg, 82.6% yield) as a grey solid. LCMS retention time 1.447 min; LCMS MH+ 491.
Step 3 7-(4-chlorobenzyl)-8-(3-ethoxypropyl)-3-methyl-l-(3-(tetrahydro-2H-pyran-2- yloxy)propyl)-l -purine-2,6(3H,7H)-dione
Figure imgf000500_0002
To a solution of 7-(4-chlorobenzyl)-8-(3-hydroxypropyl)-3-methyl-l -(3-(tetrahydro-2H-pyran- 2-yloxy)propyl)-lH-purine-2,6(3H,7H)-dione (100 mg, 0.204 mmol) in DMF (3 mL) was added sodium hydride (24.5 mg, 0.612 mmol, 60% dispersion in mineral oil) at 0 °C. The mixture was stirred at room temperature for 30 min. Iodoethane (63.6 mg, 0.408 mmol) was added to the reaction and it was stirred at room temperature for 16 h. The reaction was partitioned between ethyl acetate and brine. The organic phase was dried and concentrated to give a crude product, which was purified by silica gel chromatography eluting with petroleum ether/ ethyl acetate (2: 1 to 1 :1) to give 7-(4-chlorobenzyl)-8-(3-ethoxypropyl)-3-methyl-l -(3-(tetrahydro-2H-pyran-2- yloxy)propyl)-lH-purine-2,6(3H,7H)-dione (96 mg, 90.6% yield) as a yellow solid. LCMS retention time 1.870 min; LCMS MH+ 519.
Step 4 7-(4-chloro-3 -methoxy benzyl)- 1 -(3 -hydroxypropyl)-3 -methyl-8-(3 -
(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000501_0001
The title compound was prepared as example 14, step 3 to give 7-(4-chloro-3-methoxybenzyl)- l - (3-hydroxypropyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (51 mg, 56.5% yield) as a white solid. 'H-NM (CD3OD) δ 7.38-7.35(d, 2H), 7.24-7.22(d, 2H), 5.62(s, 2H), 4.12-4.18(1, 2H), 3.62-3.59(1, 2H), 3.57(s, 3H), 3.46-3.43(m, 4H), 2.86-2.82(t, 2H), 1.98-1.84(m, 4H), 1.18-1.14(t, 3H). LCMS retention time 2.384min; LCMS MH+ 435.
Example 49 7-(4-chlorobenzyl)-l ,8-bis(3-hydroxypropyl)-3-methyl-lH-purine-2,6(3H,7H)- dione
Figure imgf000501_0002
To a solution of 7-(4-chlorobenzyl)-8-(3-hydroxypropyl)-3-methyl-l -(3-(tetrahydro-2H-pyran-2- yloxy)propyl)-lH-purine-2,6(3H,7H)-dione (30 mg, 0.061 mmol, example 48, step 3) in ethyl alcohol (20 mL) was added acetyl chloride (0.2 mL, 2.80 mmol). The reactin was stirred at 0 °C for 10 min. The mixture was concentrated to give a crude product, which was purified by preparative HPLC to give 7-(4-chlorobenzyl)-l ,8-bis(3-hydroxypropyl)-3-methyl-l H-purine- 2,6(3H,7H)-dione (15 mg, 60.6% yield) as a white solid. 'H-NMR (CD3OD) δ 7.37-7.35(d, 2Η), 7.21-7.22(d, 2Η), 5.62(s, 2Η), 4.1 l-4.08(t, 2Η), 3.62-3.59(t, 4Η), 3.56(s, 3Η), 2.87-2.83(1, 2Η), 1.95-1.83(m, 4Η). LCMS retention time 1.801 min; LCMS MlT 407. Example 50 8-(cyclopentyloxy)-l-(3-hydroxypropyl)-3-methyl-7-((4-methylthiazol-2- yl)methyl)-HT-purine-2,6(3H,7H)-dione
Figure imgf000501_0003
Step 1 7-(4-chlorobenzyl)-8-(cyclopentyloxy)-3-methyl-l -(3-(tetrahydro-2H-pyran-2- yloxy)propyl)- 1 -purine-2,6(3H,7H)-dione (GG-0 - 120)
Figure imgf000502_0001
To a solution of cyclopentanol (67 mg, 0.78 mmol) in THF (5 mL) was added sodium hydride (39 mg, 0.98 mmol) at 0 °C. After stirring at 0 °C for 30 min, 8-bromo-7-(4-chlorobenzyl)-3- methyl-l -(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-lH-purine-2,6(3H,7H)-dione (0.2 g, 0.39 mmol, intermediate 14) was added. The mixture was stirred at room temperature for 16 h; then aqueous ammonium chloride solution (2 mL) was added at 0 °C. The reaction mixture was partitioned between ethyl acetate and water. The combined organic layer was dried over sodium sulfate, filtered and concentrated to give a crude solid product. This material was slurried in ice cold ethanol, collected by filtration, and dried under vacuum to give 7-(4-chlorobenzyl)-8- (cyclopentyloxy)-3-methyl-l-(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-l H-purine-2,6(3H,7H)- dione (180 mg, 89.5% yield) as white solid. LCMS retention time 2.085 min; LCMS MH+-THP 433.
Step 2 8-(cyclopentyloxy)-3-methyl-l-(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-lH-purine- 2,6(3H,7H)-dione
Figure imgf000502_0002
To a solution of 7-(4-chlorobenzyl)-8-(cyclopentyloxy)-3-methyl-l -(3-(tetrahydro-2H-pyran-2- yloxy)propyl)-lH-purine-2,6(3H,7H)-dione (180 mg, 0.35 mmol) in methanol (10 mL) was added ammonium formate (220 mg, 3.5 mmol) and 5% Pd/C (20 mg). The reaction was refluxed under nitrogen atmosphere overnight. The mixture was cooled and filtered. The filtrate was concentrated to give 8-(cyclopentyloxy)-3-methyl-l -(3-(tetrahydro-2H-pyran-2-yloxy)propyl)- lH-purine-2,6(3H,7H)-dione(120 mg, 87.6% yield) as white solid. LCMS retention time 1.345 min; LCMS MH+-THP 309. Step 3 8-(cyclopentyloxy)-3-methyl-7-((4-methylthiazol-2-yl)methyl)-l -(3-(tetrahydro-2H- pyran-2-yloxy)propyl)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000503_0001
To a solution of 8-(cyclopentyloxy)-3 -methyl- 1 -(3 -(tetrahydro-2H-pyran-2-yloxy)propyl)-lH- purine-2,6(3H,7H)-dione (0.12 g, 0.31 mmol) in DMF (5 mL) was added 2-(chloromethyl)-4- methylthiazole (91 mg, 0.62 mmol, intermediate 54), potassium carbonate (85 mg, 0.62 mmol), and TBAI (2 mg, 0.02 mmol). The reaction was heated at 50 °C overnight. The mixture was cooled and partitioned between ethyl acetate and water. The combined organic layer was dried over sodium sulfate, filtered and concentrated to give a crude product which was used without purification. LCMS retention time 1.704 min; LCMS MH+-THP 470.
Step 4 8-(cyclopentyloxy)-l -(3-hydroxypropyl)-3-methyl-7-((4-methylthiazol-2-yl)methyl)- lH-purine-2,6(3 -dione
Figure imgf000503_0002
The title compound was prepared using the method of example 48, step 2 to give 8-
(cyclopentyloxy)-l-(3-hydroxypropyl)-3-methyl-7-((4-methylthiazol-2-yl)methyl)-lH-purine- 2,6(3H,7H)-dione (60 mg, 60.4% yield) as a white solid. 'H-NMR (CDC13) δ 6.84(d. IH), 5.55(s, 2H), 5.46-5.50(m, IH), 4.19(t, 2H), 3.69(s, IH), 3.56(s, 3H), 3.52(s, 2H), 2.42(d, 3H), 1.87-1.97(m, 6H), 1.62-1.79(m, 4H). LCMS retention time 2.163 min; LCMS MH+ 420.
The following examples 51a through 51j were prepared following the method of example 50.
Example 51a 7-ethyl-l -(3-hydroxypropyl)-8-isopropoxy-3-methyl-lH-purine-2,6(3H,7H)- dione
Figure imgf000504_0001
White solid, 50 mg,58.8% yield: 'H-NMR (DMSO-<¾ δ 5.15-5.20(m, I H), 4.46(t, 1H), 4.02(q, 2H), 3.90(t, 2H), 3.42(q, 2H), 3.34(s, 3H), 1.66-1.70(m, 2H), 1.38(d, 6H), 1.25(t, 3H). LCMS retention time 1.946 min; LCMS MH+ 31 1.
Example 51b l-(3-hydroxypropyl)-8-isopropoxy-3-methyl-7-propyl- lH-purine-2,6(3H,7H)- dione
Figure imgf000504_0002
White solid, 39 mg,44.3% yield: -NMR (DMSO-c¾ S 5.15-5.21(m, 1H), 4.46(t, 1H), 3.95(t, 2H), 3.90(t, 2H), 3.41 (q, 2H), 3.37(s, 3H), 1.64-1.71(m, 4H), 1.37(d, 6H)5 0.8 l(t, 3H). LCMS retention time 2.149 min; LCMS MH+ 325.
Example 51c 7-benzyl-l -(3-hydroxypropyl)-8-isopropoxy-3-methyl-lH-purine-2,6(3H,7H)- dione
Figure imgf000504_0003
White solid, 26 mg, 31.7% yield: '/7-NMR (CDC13) δ 7.40-7.42(dd, 2H), 7.28-7.36 (m, 3H), 5.27(s, 2H), 4.17-4.20(t, 2H), 3.77(m, 1H), 3.51(m, 5H), 1.88-1.92(m, 2H), 1.43-4.44(d, 6H). LCMS retention time 2.336 min; LCMS MH+ 373.
Example 51d 1 -(3-hydroxypropyl)-8-isopropoxy-3-methyl-7-((6-methylpyridin-3-yl)methyl)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000504_0004
White solid, 50 mg, 30.4% yield: 'H-NMR (DMSO-< 6) δ 8.47(s, 1H), 7.78-7.76(dd, 1 H), 7.31- 7.29(d, 1 H), 5.32(s, 3H), 4.1 l -4.08(t, 2H), 3.62-3.59(1, 2H), 3.50(s, 3H), 2.52(s, 3H), 1.88- 1.85(m, 2H), 1 .45-1.44(d, 6H). LCMS retention time 1.581 min; LCMS MH+ 388. Example 51e 8-ethoxy- l -(3-hydroxypropyl)-3-methyl-7-((6-methylpyridin-3-yl)methyl)-lH- purine-2,6(3H,7H)-dione
Figure imgf000505_0001
White solid, 17 mg, 28.7% yield: !H-NMR (DMSO-<¾ δ 8.47(s, I H), 7.75-7.78(dd, 1H), 7.28- 7.30(d, IH), 5.32 (s, 3H), 4.58-4.63(q, 2H), 4.07-4.10 (t, 2H), 3.58-3.61 (t, 2H), 3.49(s, 3H), 2.52(s, 3H), 1.84-1.88(q, 2H), 1.45-1.48(t, 3H). LCMS retention time 1.371min; LCMS MH+ 374.
Example 51f 8-ethoxy-l-(3-hydroxypropyl)-3-methyl-7-((5-methyloxazol-2-yl)methyl)-lH- purine-2,6(3H,7H)-dione
Figure imgf000505_0002
White solid, 19 mg, 30.6% yield: 'H-NMR (DMSO-^) δ 6.73(s, IH), 5.42 (s, 2H), 4.57- 4.62(q, 2H), 4.04-4.07(t, 2H), 3.56-3.59(t, 2H), 3.53(s, 3H), 2.3 l (s, 3H), 1 .82-1.86(q, 2H), 1.40- 1.44(t, 3H). LCMS retention time 1.516min; LCMS MH+ 364.
Example 51g 7-((5-chloropyridin-2-yl)methyl)-8-ethoxy-l-(3-hydroxypropyI)-3-methyl-lH- purine-2,6(3H,7H)-dione
Figure imgf000505_0003
White solid, 13 mg, 31.2% yield: 'H-NMR (CDC13) δ 8.49(s, 1 H), 7.64-7.66(dd, 1 H), 7.20- 7.22(d, 1H), 5.40(s, 2H), 4.55-4.60(q, 2H), 4.14-4.16 (t, 2H), 3.59-3.61 (t, 1 H), 3.56 (s, 3H), 3.46-3.49 (t, 3H), 1 .86-1.88 (m, 2H), 1 .40-1.42 (t, 2H). LCMS retention time 1.896min; LCMS MH+ 394.
• Example 51h 8-(cyclopentylmethoxy)-7-ethyl-l-(3-hydroxypropyl)-3-methyl-lH-purine- 2,6(3H,7H)-dione
Figure imgf000506_0001
Light yellow solid, 1 1 mg, 20.7% yield: 'H-NMR (DMSO-^) δ 4.45-4.35(t, 1Η), 4.35-4.33(d, 2Η), 4.07-4.02(m, 2Η), 3.92-3.88(t, 2Η), 3.45-3.40(q, 2Η), 3.30(s, 3Η), 2.39-2.32(m, III), 1.80- 1.52(m, 8Η), 1.37-1.24(m, 5Η). LCMS retention time 2.485min; LCMS MH+ 351.
Example 51i 8-(cyclopentylmethoxy)-l -(3-hydroxypropyl)-3-methyl-7-propyl-lH-purine- 2,6(3H,7H)-dione
Figure imgf000506_0002
White solid, 16 mg, 31.5% yield: 'H-NMR (DMSO-i¾ δ 4.45-4.34(t, 1Η), 4.35-4.33(d, 2Η), 3.99-3.96(t, 2Η), 3.91-3.88(t, 2Η), 3.44-3.40(m, 2Η), 3.37(s, 3Η), 2.37-2.34(m, 1Η), 1.78- 1.52(m, 8Η), 1.35-1.31(m, 2Η), 0.84-0.80(t, 3Η). LCMS retention time 2.670min; LCMS MH+ 365.
Example 51j 8-(cyclopentylmethoxy)-l-(3-hydroxypropyl)-3-methyl-7-((6-methylpyridin-3- yl)methyI)-l H-purine-2,6(3H,7H)-dione
Figure imgf000507_0001
White solid, 25 mg, 49.9% yield: 'H-NMR (DMSO-4 δ 8.43(s, I H), 7.61 -7.58(dd, I H), 7.24- 7.22(d, IH), 5.22(s, 2H), 4.47-4.44(t, IH), 4.35-4.34(d, 2H), 3.93-3.89(t, 2H), 3.45-3.38(m, 2H), 3.35(s, 3H), 2.33(s, 3H), 2.13-2.1 l (m, IH), 1 .70-1.51(m, 8H), 1.28-1.25(m, 2H). LCMS retention time 1.761 min; LCMS MH+ 428.
Example 52 7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl-8-(6-methylpyridin-3-yloxy)- lH-purine-2,6(3H,7H)-dione hydrochloride
Figure imgf000507_0002
Step 1 7-(4-chlorobenzyl)-3-methyl-8-(6-methylpyridin-3-yloxy)-l-(3-(tetrahydro-2H-pyran-2- yloxy)propyl -lH-purine-2,6(3H,7H)-dione
Figure imgf000507_0003
To a solution of 8-bromo-7-(4-chlorobenzyl)-3 -methyl- 1 -(3 -(tetrahydro-2H-pyran-2- yloxy)propyl)-l H-purine-2,6(3H,7H)-dione (100 mg, 0.195 mmol, intermediate 14) in in DMF (5 mL) was added 6-methylpyridin-3-ol (24 mg, 0.22 mmol) followed by potassium carbonate (76 mg, 0.55 mmol) and the mixture was stirred at 85 °C for 3 h. The mixture was cooled and partitioned between ethyl acetate and brine. The layers were separated and the organic layer was washed with saturated aqueous ammonium chloride, dried and concentrated to give 7-(4- chlorobenzyl)-3-methyl-8-(6-methylpyridin-3-yloxy)-l-(3-(tetrahydro-2H-pyran-2- yloxy)propyl)-l H-purine-2,6(3H,7H)-dione (79 mg, 75% yield) as yellow solid. LCMS retention time 2.297 min; LCMS MH+ -THP 456.
Step 2 7-(4-chlorobenzyl)- 1 -(3-hydroxypropyl)-3-methyl-8-(6-methylpyridin-3-yloxy)- 1 H- purine-2,6(3H,7H)-dione
Figure imgf000508_0001
A solution of 7-(4-chlorobenzyl)-3-methyl-8-(6-methylpyridin-3-yloxy)-l -(3-(tetrahydro-2H- pyran-2-yloxy)propyl)-l H-purine-2,6(3H,7H)-dione (79 mg, 0.146 mmol) in IN ethanolic HCI (3 mL) was stirred at room temperature for 2 h. The mixture was concentrated to dryness to give a crude product, which was purified via preparative HPLC to give 7-(4-chlorobenzyl)-l -(3- hydroxypropyl)-3-methyl-8-(6-methylpyridin-3-yloxy)-lH-purine-2,6(3H,7H)-dione (35 mg, 52.6% yield) as white solid. 'H-NMR (DMSO-<¼) δ 8.69(s, 1Η), 8.03-8.01 (d, 1 Η), 7.59-7.57(d, 1Η), 7.45(s, 4Η), 5.45(s, 2Η), 4.06-3.98(m, 2Η), 3.45-3.42(t, 2Η), 3.28(s, 3Η), 2.59(s, 3Η), 1 .71 - 1.67(t, 2Η). LCMS retention time 2.207 min; LCMS MH+ 456.
The following examples 53a and 53b were prepared using the method of example 52.
Example 53a 7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl-8-(2-methylpyridin-4-yloxy)- 1 H-purine-2,6(3H,7H)-dione hydrochloride
Figure imgf000508_0002
HCI
White solid, isolated as the HCI salt, 30 mg, 36.6% yield: 'H-NMR (DMSO-<¾) δ 8.48-8.46(d, 1Η), 8.17(s, 1Η), 7.43-7.33(m, 4Η), 7.26-7.22(m, 2Η), 5.42(s, 2Η), 4.52(s, 1Η), 3.95-3.92(t, 2Η), 3.44-3.42(m, 2Η), 3.37(s, 3Η), 2.47(s, 3Η), 1.72-1.69(t, 2Η). LCMS retention time 1.716 min; LCMS MH+ 456.
Example 53b 7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl-8-(5-methylpyridin-3-yloxy)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000509_0001
White solid, 40 mg, 48.8% yield: 'H-NMR (DMSO-efe) δ 8.45-8.44(d, 1H), 8.36(s, 1H), 7.65(s,lH), 7.46-7.4 l(m, 4H), 5.44(s, 2H), 4.50-4.47(t, 1H), 3.94-3.91(1, 2H), 3.46-3.40(m, 2H), 3.28(s, 3H), 2.34(s, 3H), 1.73-1.66(m, 2H). LCMS retention time 2.259 min; LCMS MH+ 456.
Example 54 7-butyl-l -(3-hydroxypropyl)-3-methyl-8-(5-methylpyridin-3-yloxy)-lH-purine- 2,6(3H,7H)-dione
Figure imgf000509_0002
Step 1 3-methyl-8-(5-methylpyridin-3-yloxy)- l-(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-7- ((2-(trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H,7H)-dione
Figure imgf000509_0003
The title compound was prepared using the method of example 52, step 1 from intermediate 16, step 1 to give 3-methyl-8-(5-methylpyridin-3-y[oxy)-l-(3-(tetrahydro-2H-pyran-2-yloxy)propyl)- 7-((2-(trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H,7H)-dione (300 mg, 97.6% yield) as yellow oil. LCMS retention time 1.838 min; LCMS M-THP+H 462.
Step 2 7-(4-chlorobenzyl)- 1 -(3 -hydroxybuty l)-3 -methyl -8 -(6-methylpyridin-3 -yloxy)- 1 H- purine-2,6(3H,7H)-dione
Figure imgf000510_0001
To a solution of 3-methyl-8-(5-methylpyridin-3-yloxy)-l -(3-(tetrahydro-2H-pyran-2- yloxy)propyl)-7-((2-(trimethylsilyl)ethoxy)methyl)-l H-purine-2,6(3H,7H)-dione (300 mg, 0.554 mmol) in ethyl alcohol (10 mL) was added concentrated HC1 (3 mL) and the mixture was stirred for 6 h at reflux. The mixture was cooled, concentrated and filtered to give 7-(4-chlorobenzyl)-l- (3-hydroxybutyl)-3-methyl-8-(6-methylpyridin-3-yloxy)-lH-purine-2,6(3H,7H)-dione (1 59 mg, 86.6% yield) as yellow solid. LCMS retention time 0.356 min; LCMS MH+ 332.
Step 3 7-butyl- 1 -(3-hydroxypropyl)-3-methyl-8-(5-methylpyridin-3 -yloxy)- 1 H-purine- 2,6(3H,7H)-dione
Figure imgf000510_0002
The title compound was prepared using the method of example 44, step 3 to give 7-butyl- 1 -(3- hydroxypropyl)-3-methyl-8-(5-methylpyridin-3-yloxy)-lH-purine-2,6(3H,7H)-dione (30 mg, 42.8% yield) as white solid. JH-NMR (DMSO-40 = 8.48-8.47(d, IH), 8.37-8.36(d, IH), 7.71 (s, I H), 4.51 -4.48(t, IH), 4.22-4.19(t, 2H), 3.94-3.91 (t, 2H), 3.46-3.43(t, 2H), 3.28(s, 3H), 2.36(s, 3H), 1 .81 -1.66(m, 4H), 1.35-1.30(m, 2H), 0.93-0.89(t, 3H). LCMS retention time 1.854 min; LCMS MH+ 388.
Example 55 8-(3-fluorophenoxy)-l-(3-hydroxypropyl)-3-methyl-7-((5-methylthiazol-2- yl)methyl)-lH-purine-2,6(3H,7H)-dione
Figure imgf000510_0003
To a solution of 8-bromo-l -(3-hydroxypropyl)-3-methyl-7-((5-methylthiazol-2-yl)methyl)-lH- purine-2,6(3H,7H)-dione (50 mg,0.12 mmol, intermediate 16) in DMF (3mL) was added 3- fluorophenol (23 mg, 0.17 mmol) and potassium carbonate (75 g, 0.24 mmol). The mixture was stirred at 80°C for 12 h. The mixture was cooled and partitioned between ethyl acetate and water. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give a crude product which was purified by preparative HPLC to give 8-(3-fluorophenoxy)-l -(3- hydroxypropyl)-3-methyl-7-((5-methylthiazol-2-yl)methyl)-l H-purine-2,6(3H,7H)-dione (1 1 mg, 20.75 % yield) as white solid. 'H-NMR (CD3OD) δ 7.42-7.48(m, 2H), 7.18-7.22(m, 2H), 7.06-7.10(m, 1H), 5.76(s, 2H), 4.10 (t, 2H), 3.61(t, 2H), 3.45(s, 3H), 2.47(d, 3H), 1.85-1.90(m, 2H). LCMS retention time 1.273 min; LCMS MH+ 446.
The following examples 56a through 56i were prepared using the method of example 55.
Example 56a 8-(3-chlorophenoxy)-l -(3-hydroxypropyl)-3-methyl-7-((5-methylthiazol-2- yl)methyl)-l H-purine-2,6(3H,7H)-dione
Figure imgf000511_0001
White solid, 17 mg, 30.9 % yield: 'H-NMR (CD3OD) S 7.42-7.47(m, 3 H), 7.30-7.35(m, 2 H), 5.76(s, 2 H) , 4.10 (t, 2 H), 3.61(t, 2 H), 3.44(s,3 H), 2.48(d,3 H),1.85-1.90(m,2 H). LCMS retention time 2.246 min; LCMS MH+ 462.
Example 56b l-(3-hydroxypropyl)-3-methyl-7-((5-methylthiazol-2-yl)methyl)-8-(m-tolyloxy)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000511_0002
White solid, 25 mg, 47.1% yield: 'H-NMR (CD3OD) δ 7.42(d, 1Η), 7.29-7.34(m, 1 Η), 7.09- 7.13(m, 3Η), 5.75(s, 2Η) , 4.10 (t, 2Η), 3.61(t, 2Η), 3.43(s, 3Η), 2.48(d, 3H),2.38(s, '3H),1.84- 1.91(m, 2H). LCMS retention time 2.199 min; LCMS MIT" 442. Example 56c 1 -(3-hydroxypropyl)-8-(3-methoxyphenoxy)-3-methyl-7-((5-methylth yl)methyl)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000512_0001
White solid, 18 mg, 47.1% yield: *H-NMR (CD3OD) δ 7.41 (d, 1H), 7.33(t, 1 H), 6.85-6.91 (m, 3H), 5.74(s, 2H), 4.09 (t, 2H), 3.81(s, 3H), 3.61(t, 2H), 3.43(s, 3H), 2.47(d, 3H), 1.84-1.90(m, 2H). LCMS retention time 2.092 min; LCMS MH+ 458.
Example 56d 1 -(3-hydroxypropyl)-3-methyl-7-((5-methylthiazol-2-yl)methyl)-8-(3- (trifluoromethyl)phenoxy)-lH-purine-2,6 3H,7H)-dione
Figure imgf000512_0002
White solid, 45 mg, 47.1% yield: ^/-NMR (CD3OD) δ 7.64-7.71(m, 4 H), 7.41(d, 1 H), 5.78(s, 2 H) , 4.10 (t, 2 H), 3.61 (t, 2 H), 3.45(s,3 H), 2.47(d,3 H), 1.85-1.91(m,2 H). LCMS retention time 2.349 min; LCMS MH+ 496. Example 56e 8-(4-fluorophenoxy)-l -(3-hydroxypropyl)-3-methyl-7-((5-methylthiazol-2- yl)methyl)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000512_0003
White solid, 23 mg, 43.4% yield: ^-NMR (CD3OD) δ 7.42(d, 1H), 7.35-7.39(m, 2H), 7. 7.22(m, 2H), 5.76(s, 2H) , 4.10 (t, 2H), 3.61 (t, 2H), 3.45(s, 3H), 2.48(d, 3H), 1.85-1.90(m, 2 LCMS retention time 2.108 min; LCMS MH+ 446. Example 56f 1 -(3-hydroxypropyl)-3-methyl-7-((5-methylthiazol-2-yl)methyl)-8-(4- (trifluoromethyl)phenoxy)-lH-purine-2,6 3H,7H)-dione
Figure imgf000513_0001
White solid, 20 mg, 40.4% yield: 'H-NMR (CD3OD) δ 7.78(d, 2H), 7.57(d, 2H), 7.41(d, 1 H), 5.78(s, 2H) , 4.10 (t, 2H), 3.61 (t, 2H), 3.45(s, 3H), 2.48(d, 3H), 1.85-1.90(m, 2H). LCMS retention time 2.386 min; LCMS MH+ 496.
Example 56g l-(3-hydroxypropyl)-3-methyl-7-((5-methylthiazol-2-yl)methyl)-8-(4- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000513_0002
White solid, 19 mg, 31.1% yield: *H-NMR (CD3OD) δ 7.46-7.49(m, 2Η), 7.38-7.42(m, 3Η), 5.77(s, 2Η) , 4.10(t, 2Η), 3.61(t, 2Η), 3.44(s, 3Η), 2.47(s, 3Η), 1.85-1.90(m, 2Η). LCMS retention time 2.432 min; LCMS MH+ 512. Example 56h 8-(4-chlorophenoxy)-l -(3-hydroxypropyl)-3-methyl-7-((5-methylthiazol-2- yl)methyl)-lH-purine-2,6(3H,7H)-dione
Figure imgf000513_0003
White solid, 15 mg, 31.1 % yield: \H-NMR (CD3OD) δ 7.42-7.48(m, 3H), 7.35-7.38(m, 2H), 5.76(s, 2H) , 4.10 (t, 2H), 3.61(t, 2H), 3.43(s, 3H), 2.48(d, 3H), 1.85-1.91(m, 2H). LCMS retention time 2.273 min; LCMS MH+ 462. Example 56i l-(3-hydroxypropyl)-8-(4-methoxyphenoxy)-3-methyl-7-((5-methylthiazol-2- yl)methy])-l H-purine-2,6(3H,7H)-dione
Figure imgf000514_0001
White solid, 13 mg, 31.1% yield: 'H-NMR (CD3OD) δ 7.42 (d, I H), 7.23(dd, 2H), 6.98(dd, 2H), 5.74(s, 2H) , 4.09 (t, 2H), 3.82(s, 3H), 3.60(t, 2H), 3.42(s, 3H), 2.48(d, 3H), 1.85-1.90(m, 2H). LCMS retention time 2.104 min; LCMS MH+ 458.
Example 57 8-ethoxy-l -(3-hydroxypropyl)-3-methyl-7-((5-methylthiazol-2-yl)methyl)-lH- purine-2,6(3H,7H -dione
Figure imgf000514_0002
To a solution of 8-bromo-l-(3-hydroxypropyl)-3-methyl-7-((5-methylthiazol-2-yl)methyl)-lH- purine-2,6(3H,7H)-dione (50 mg,0.12 mmol, intermediate 16) in ethanol (2 mL) was added sodium (10 mg, 0.4 mmol). The mixture was stirred at room temperature for 2 h. The reaction was concentrated and the residue was purified by preparative HPLC to give 8-ethoxy-l -(3- hydroxypropyl)-3-methyl-7-((5-methylthiazol-2-yl)methyl)-lH-purine-2,6(3H,7H)-dione (27 mg,59.3 %) as white solid. l//-NMR (CD3OD) δ 7.38 (d, IH), 5.55(s, 2H) ,4.60(q, 2H), 4.09 (t, 2H), 3.59(t, 2H), 3.52(s, 3H), 2.45(d, 3H), 1.83-1.88(m, 2H), 1.43(d, 3H). LCMS retention time 1.680 min; LCMS ΜΗ' 380.
Example 58 7-ethyl-l -(3-hydroxypropyl)-3-methyl-8-((5-methylthiazol-2-yl)methoxy)-lH- purine-2,6(3H,7H)-dione
Figure imgf000514_0003
The title compound was prepared using the 2 step method of example 36 from intermediate 6. The product was purified via preparative HPLC to give7-ethyl-l -(3-hydroxypropyl)-3-methyl-8- ((5-methylthiazol-2-yl)methoxy)-l H-purine-2,6(3H,7H)-dione (5 mg, 6.9% yield) as white solid. 'H-NMR (DMSO-40 δ 7.53(s, 1 H), 5.76(s, 2H), 4.18(q, 2H), 4.10(t, 2H), 3.6 l (t, 2H), 3.54(s, 3H), 2.52(s, 3H), 1.80-1.90(m, 2H), 1.36(t, 3H). LCMS retention time 1.760 min; LCMS MH+ 380.
Example 59 7-ben2yl-l-(3-hydroxypropyl)-8-(3-(trifluoiOmethoxy)phenoxy)-lH-purine- 2,6( -dione
Figure imgf000515_0001
Step 1 7-benzyl-l -(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-8-(3-(trifluoromethoxy)phenoxy)- 3-((2-(trimeth lsilyl)ethoxy)methyl)-lH-purine-2,6( -dione
Figure imgf000515_0002
To a solution of 7-benzyl-8-chloro-l-(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-3-((2- (trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H,7H)-dione (4.7 g, 8.58 mmol, intermediate 18) in DMF (15 mL) was added 3-(trifluoromethoxy)phenol (1.83 g, 10.28 mmol) and potassium carbonate (2.4 g, 17.39 mmol). The mixture was stirred at 80 °C overnight. The mixture was diluted with ethyl acetate and water, and the phases were separated. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give 7-benzyl-l -(3- (tetrahydro-2H-pyran-2-yloxy)propyl)-8-(3-(trifluoromethoxy)phenoxy)-3-((2-
(trimethylsilyl)ethoxy)methyl)-l H-purine-2,6(3H,7H)-dione (5.3 g, 89.3% yield) as yellow oil. LCMS retention time 3.974 min; LCMS M+Na 713.
Step 2 7-benzyl-l -(3-hydroxypropyl)-8-(3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)- dione
Figure imgf000516_0001
To a solution of 7-benzyl-l -(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-8-(3- (trifluoromethoxy)phenoxy)-3-((2-(trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H
(5.3 g, 7.68 mmol) in ethyl alcohol (20 mL) was added concentrated HC1 (5 mL). The reaction was stirred at 80 °C overnight. The reaction was concentrated and the residue was neutralized with saturated sodium bicarbonate. This aqueous phase was extracted with ethyl acetate and the phases were separated. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give 7-benzyl-l -(3-hydroxypropyl)-8-(3-
(trifluordmethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (4.5 g, 100% yield) as white solid. XH- NMR (DMSO-cfe) δ 1 1.93(s, 1H), 7.62-7.57(t, 1 H), 7.44-7.32(m, 8H), 5.42(s, 2H), 4.49-4.46(t, 1H), 3.89-3.86(t, 2H), 3.46-3.41 (m, 2H), 1.72-1.65(m, 2H). LCMS retention time 2.560 min; LCMS MH+ 477.
Example 60 7-benzyl-3-ethyl-l-(3-hydroxypropyl)-8-(3-(trifluoromethoxy)phenoxy)-l H- purine- -dione
Figure imgf000516_0002
To a solution of 7-benzyl-l -(3-hydroxypropyl)-8-(3-(trifluoromethoxy)phenoxy)-lH-purine- 2,6(3H,7H)-dione (1 .2 g, 2.52 mmol, example 59) in DMF (5 mL) was added iodoethane (0.7 g, 4.49 mmol), followed by potassium carbonate (0.7 g, 5.07 mmol). The reaction was stirred at 80 °C for 2 h; then it was cooled and partitioned between ethyl acetate and water. The phases were separated and the organic phase was washed with brine. The organic layer was dried over sodium sulfate, filtered and concentrated to give 7-benzyl-3-ethyl- l -(3-hydroxypropyl)-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (0.62 g, 49.2% yield) as yellow solid. 'H-NMR (DMSO-t¼) δ 7.62-7.57(t, 1H), 7.50(s, 1H), 7.42-7.3 l(m, 7H), 5.44(s, 2H), 4.51- 4.48(t, IH), 3.95-3.85(m, 4H), 3.47-3.42(q, 2H), 1.73-1 .67(m, 2H), 1 .16-1.13(t, 3H). LCMS retention time 2.974 min; LCMS MH+ 505.
Example 61 3-ethyl-l -(3-hydroxypropyl)-7-((6-methylpyridin-3-yl)methyl)-8-(3- (trifluoromethoxy)phenoxy)-l H-purine-2,6(3H,7H)-dione
Figure imgf000517_0001
Step 1 3-ethyl-l -(3-hydroxypropyl)-8-(3-(trifluoromethoxy)phenoxy)-l H-purine-2,6(3H,7H)- dione
Figure imgf000517_0002
7-benzyl-3-ethyl-l-(3-hydroxypropyl)-8-(3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)- dione (0.62 g, 1.23 mmol, example 60) was dissolved in ethanol (20 mL); then the mixture was degassed and refilled with nitrogen three times. Ammonium formate (0.5 g, 7.94 mmol) and 10% Pd/C (30 mg) were added. The mixture was again degassed and refilled with nitrogen three times; then it was stirred at 80 °C overnight. The reaction was cooled and filtered. The filter cake was washed with methanol. The filtrate was concentrated to give 3-ethyl-l -(3- hydroxypropyl)-8-(3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (0.6 g, 100%) yield) as white solid. XH-NMR (DMSO-rf6) δ 7.61-7.57(t, IH), 7.50(s, IH), 7.40-7.38(dd, IH), 7.31 -7.29(d, IH), 4.50(s, IH), 3.95-3.85(m, 4H), 3.44-3.43(m, 2H), 1.73-1.66(t, 2H), 1.17-1.13(t, 3H).LCMS retention time 1.350 min; LCMS MH+ 415.
Step 2 3-ethyl-l -(3-hydroxypropyl)-7-((6-methylpyridin-3-yl)methyl)-8-(3- (trifluoromethox
Figure imgf000517_0003
To a solution of 3-ethyl-l -(3-hydroxypropyl)-8-(3-(trifluoromethoxy)phenoxy)- lH-purine- 2,6(3H,7H)-dione (1 10 mg, 0.22 mmol) in DMF (3 mL) was added 5-(chlorometbyl)-2- methylpyridine (50 mg, 0.4 mmol), potassium carbonate (91 mg, 0.66 mmol), and TBAI (2 mg, 0.02 mrnol). The reaction was heated at 50°C overnight. The mixture was cooled and partitioned between ethyl acetate and water. The combined organic layer was dried over sodium sulfate, filtered and concentrated to give a crude product, which was purified by preparative HPLC to give 3 -ethyl- 1 -(3 -hydroxypropy l)-7-((6-methylpyridin-3 -yl)methyl)-8-(3 - (trifluoromethoxy)phenoxy)-l H-purine-2,6(3H,7H)-dione (30 mg, 44.8% yield) as a white solid. 'H-NMR (DMSO-<¾ δ 8.53(d, 1H), 7.71 (dd, 1H), 7.55-7.63(m, 2H), 7.50(dd, 1H), 7.32(d, 1H), 7.25(d, 1H), 5.43(s, 2H), 4.48(t, 1 H), 3.85-3.96(m, 4H), 3.44(q, 2H), 2.44(s, 3H), 1.68- 1.73(m, 2H), 1.14(t, 3H). LCMS retention time 2.097min; LCMS MH+ 520.
The following examples 62a through 62d were prepared using the method of example 61. Example 62a 7-((5-chloropyridin-2-yl)methyl)-3-ethyl-l-(3-hydroxypropyl)-8-(3- (trifluoromethoxy)phenoxy)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000518_0001
White solid, 30 mg, 28.1% yield: 'H-NMR (DMSO-c¾ δ 8.47(s, 1Η), 7.84-7.87(dd, 1Η), 7.53- 7.57(t, 1Η), 7.46-7.48(d, 1Η), 7.34-7.39 (m, 2Η), 7.22-7.24 (d, 1Η), 5.63(s, 2Η), 4.03-4.08(m, 4Η), 3.56-3.59(t, 2Η), 1.82-1.85 (q, 2Η), 1.26-1.29(t, 3Η). LCMS retention time 2.701 min; LCMS MH+ 540.
Example 62b 7-(4-chlorobenzyl)-3-ethyl-l-(3-hydroxypropyl)-8-(3- (trifluoromethoxy)phenoxy)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000518_0002
White solid, 15 mg, 20.1% yield: 'H-NMR (DMSO-t¼) δ 7.60-7.56(t, IH), 7.49(s, IH), 7.43- 7.39(m, 5H), 7.30-7.28(d, I H), 5.40(s, 2H), 4.49(bs, IH), 3.92-3.82(m, 4H), 3.43-3.36(m, 2H), 1.7M .64(m, 2H), 1 .14-1 .10(t, 3H). LCMS retention time 1.906 min; LCMS MH+ 539. Example 62c 3,7-diethyl-l -(3-hydroxypropyl)-8-(3-(trifluoromethoxy)phenoxy)-l H-purine- 2,6(3H, -dione
Figure imgf000519_0001
White solid, 15 mg, 20.1 % yield: 'H-NMR (DMSO-<¾ δ 7.64-7.57(m, 2H), 7.47-7.45(dd, IH), 7.34-7.32(d, IH), 4.49-4.47(t, IH), 4.25-4.20(q, 2H), 3.96-3.85(m, 4H), 3.47-3.38(m, 2H), 1.74- 1.67(m, 2H), 1 .39-1.36(t, 3H), 1.16-1.13(t, 3H). LCMS retention time 2.667 min; LCMS MH+ 443.
Example 62d 3-ethyl-l-(3-hydroxypropyl)-7-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH- purine-2,6(3H,7H)-dione
Figure imgf000519_0002
White solid, 15 mg, 14.2% yield: 'H-NMR (DMSO- 6) δ 7.64-7.59(t, IH), 7.56(s, IH), 7.47- 7.44(dd, IH), 7.33-7.31 (d, IH), 4.50-4.48(t, IH), 3.95-3.84(m, 4H), 3.46-3.42(m, 2H), 1.73- 1.66(m, 2H), 1.15-1.12(t, 3H). LCMS retention time 2.504 min; LCMS MH+ 429.
Example 63 1 -(3-hydroxypropyl)-7-((6-methylpyridin-3-yl)methyl)-3-propyl-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000520_0001
Ste l l -(3-hydroxypropyl)-3-propyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)- dione
Figure imgf000520_0002
The title compound was prepared using the methods of examples 60 and 61 , step 1 from the product of example 59. White solid, 180 mg, 70.1% yield: LCMS retention time 1.452 min; LCMS MH+ 429.
Step 2 1 -(3-hydroxypropyl)-7-((6-methylpyridin-3-yl)methyl)-3-propyl-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000520_0003
The title compound was prepared using the method of example 61 , step 2. The product was purified via preparative HPLC to give l-(3-hydroxypropyl)-7-((6-methylpyridin-3-yl)methyl)-3- propyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (7.1 mg, 15.1% yield) as white solid. JH-NMR (DMSO-i¼) δ 8.53-8.52(d, IH), 7.72-7.68(dd, IH), 7.63-7.56(m, 2H), 7.45-7.42(dd, IH), 7.33-7.31 (Hd, IH), 7.26-7.24(d, IH), 5.42(s, 2H), 4.50-4.47(t, IH), 3.95- 3.91(t, 2H), 3.82-3.78(t, 2H), 3.46-3.42(q, 2H), 2.44(s, 3H), 1.73-1.58(m, 4H), 0.82-0.78(t,3H). LCMS retention time 2.276 min; LCMS MH+ 534. Example 64 7-(4-chlorobenzy])-8-(3-((dimethylamino)methyl)phenoxy)-l -(3- hydroxypropyl)-3-propyl-lH-purine-2,6(3H,7H)-dione
Figure imgf000521_0001
Step 1 8-chloro-7-(4-chlorobenzyl)-l -(3-hydroxypropyl)-3-propyl-lH-purine-2,6(3H,7H)- dione
Figure imgf000521_0002
The title compound was prepared using the method of example 60. White solid, 160 mg, 72.3% yield: LCMS retention time 1.574 min; LCMS MH+ 41 1. Step 3 3-(7-(4-chlorobenzyl)-l -(3-hydroxypropyl)-2,6-dioxo-3-propyl-2,3,6,7-tetrahydro-lH- purin-8-yloxy)benzaldehyde
Figure imgf000521_0003
The title compound was prepared using the method of example 59, step 1. Yellow oil, 190 mg, 70.9% yield: LCMS retention time 1.663 min; LCMS MH+ 497.
Step 4 7-(4-chlorobenzyl)-8-(3-((dimethylamino)methyl)phenoxy)-l-(3-hydroxypropyl)-3- propyl-lH-purine-2,6(3H,7H)-dione hydrochloride
Figure imgf000522_0001
The title product was prepared using the method of example 39, step 2 and the crude product was purified by preparative HPLC. White solid, 15 mg, 14.3%: JH-NMR (DMSO-c¼) δ 1 1 .04(bs, 1H), 7.60(bs, 1H), 7.57-7.49(m, 2H), 7.46-7.41 (m, 5H), 5.43(s, 2H), 4.29-4.28(d, 2H), 3.95- 3.91 (t, 2H), 3.81-3.77(t, 2H), 3.45-3.42(t, 2H), 2.68-2.67(d, 6H), 1.72-1.66(m, 2H), 1.63-1.54(m, 2H), 0.81-0.77(t, 3H). LCMS retention time 1.755 min; LCMS MH+ 526.
Example 65 7-(4-chlorobenz"yl)-8-(3-((dimethylamino)methyl)phenoxy)-3-ethyl-l-(3- hydroxypropyl)- 1 H-purine-2,6(3H,7H)-dione hydrochloride
Figure imgf000522_0002
The title compound was prepared using the method of example 64. White solid, 13 mg, 14.5% yield: !H-NMR (DMSO-^) δ 1 1.15(s, 1H), 7.63(s, 1H), 7.58-7.50(m, 2H), 7.47-7.42(m, 5H), 5.43(s, 2H), 4.31-4.29(d, 2H), 3.95-3.91 (t, 2H), 3.89-3.83(m, 2H), 3.46-3.42(t, 2H), 2.69-2.67(d, 6H), 1.73-1.66(m, 2H), 1.15-l. l l (t, 3H). LCMS retention time 1.668 min; LCMS MH+ 512 .
Example 66 7-(4-chlorobenzyl)-8-ethoxy-3-ethyl-l -(3-hydroxypropyl)-lH-purine- 2,6(3H,7H)-dione
Figure imgf000522_0003
To a solution of 8-ethoxy-3-ethyl-l -(3-hydroxypropyl)-lH-purine-2,6(3H,7H)-dione (70 mg,
0.248 mmol, intermediate 26) in DMF (3 mL) was added l-(bromomethyl)-4-chlorobenzene (80 mg, 0.390 mmol), followed by potassium carbonate (60 mg, 0.435 mmol). The mixture was stirred at 60 °C for 4 h, then diluted with ethyl acetate and water, and the phases were separated. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give a crude product which was purified by preparative HPLC to give 7-(4-chlorobenzyl)-8- ethoxy-3-ethyl-l -(3-hydroxypropyl)-lH-purine-2,6(3H,7H)-dione (15 mg, 14.9%yield) as white solid. 'H-NMR (DMSO-d6) δ 7.43-7.4 l (d, 2H), 7.34-7.32(d, 2H), 5.21 (s, 2H), 4.53-4.44(m, 3H), 3.99-3.88(m, 4H), 3.45-3.39(m, 2H), 1.71 -1.64(m, 2H), 1.37-1.33(t, 3H), 1.23-1.19(t, 3H). LCMS retention time 2.654 min; LCMS MH+ 407. Example 67 7-(4-chIorobenzyl)-8-ethoxy-l -(3-hydroxypropyl)-3-propyl-lH-purine- 2,6(3H,7H)-dione
Figure imgf000523_0001
The title compound was prepared using the methods of intermediate 26 and example 66. White solid, 5 mg, 17.0% yield: *H-NMR (DMSO-^) δ 7.43-7.42(d, 2H), 7.33-7.3 l(d, 2H), 5.21(s, 2H), 4.52-4.47(m, 3H), 3.92-3.87(m, 4H), 3.44-3.41(m, 2H), 1.72- 1.63(m, 4H), 1.36-1.33(t, 3H), 0.88-0.84(t, 3H). LCMS retention time 2.837 min; LCMS MH+ 421.
Example 68 7-(4-chlorobenzyl)-8-(2-ethoxyethoxy)-l -(3-hydroxypropyl)-3-methyl-lH-purine- 2,6(3H,7H)-dione
Figure imgf000523_0002
The title compound was prepared using the two-step method of example 36. White solid, 33 mg, 26.2% yield: 'H-NMR (DMSO-<¾ 5 7.34-7.41(m, 4 H), 5.21(s, 2 H), 4.58(t, 2 H), 4.47(t, 1 H), 3.90(t, 2 H), 3.69-3.72(m, 2 H), 3.41 -3.47(m, 7 H), 1.65-1.70(m, 2 H), 1.08(t, 3 H). LCMS retention time 2.423 min; LCMS MH+ 437.
Example 69 7-(4-chlorobenzyl)-8-(isopropylsulfonyl)-l-(isoxazol-5-ylmethyl)-3-methyl-lH- purine-2,6( -dione
Figure imgf000524_0001
Step 1 7-(4-chlorobenzyl)-8-(isopropylthio)-3-methyl-lH-purine-2,6(3H,7H)-dione
Figure imgf000524_0002
To a solution of 8-bromo-7-(4-chlorobenzyl)-3 -methyl- 1 H-purine-2,6(3H,7H)-dione (0.2 g, 0.54 mmol, intermediate 8) in DMF (5 mL) was added propane-2 -thiol (0.2 g, 2.7 mmol) and potassium carbonate (0.38 g, 2.7 mmol). The reaction was heated at 80 °C overnight. The mixture was cooled and partitioned between ethyl acetate and water. The organic phase was dried over sodium sulfate, filtered and concentrated to give a crude product (0.2 g) which was used without purification. LCMS retention time 1.015 min; LCMS MH+ 365. Step 2 7-(4-chlorobenzyl)-8-(isopropylsulfonyl)-3-methyl-lH-purine-2,6(3H,7H)-dione
Figure imgf000525_0001
To a solution of 7-(4-chlorobenzyl)-8-(isopropylthio)-3-methyl-l H-purine-2,6(3H,7H)-dione (0.2 g, 0.55mmol) in chloroform (10 mL) was added MCPBA (0.19 g, 1.37 mmol) portionwise at 0°C. The reaction was stirred at room temperature overnight. The mixture was partitioned between ethyl acetate and water. The combined organic layer was dried over sodium sulfate, filtered and concentrated to give a crude solid product which was triturated in ethanol and collected to afford 7-(4-chlorobenzyl)-8-(isopropylsulfonyl)-3-methyl-lH-purine-2,6(3H,7H)- dione (0.1 9 g, 84.5% yield) as a white solid. LCMS retention time 1.322"min; LCMS MH+ 397. Step 3 7-(4-chlorobenzyl)-8-(isopropylsulfonyl)-l-(isoxazol-5-ylmethyl)-3-methyl-l H-purine- 2,6(3H,7H)-dione
Figure imgf000525_0002
To a solution of 7-(4-chlorobenzyl)-8-(isopropylsulfonyl)-3-methyl-lH-purine-2,6(3H,7H)-dione (80 mg, 0.20 mmol) in DMF (5 mL) was added 5-(chloromethyI)isoxazole (55 mg, 0.5 mmol, intermediate 41 ), potassium carbonate (58 mg, 0.42 mmol), and TBAI (2 mg, 0.02 mmol). The reaction was heated at 50 °C overnight. The mixture was cooled and partitioned between ethyl acetate and water. The combined organic layer was dried over sodium sulfate, filtered and concentrated to give a crude product which was purified by preparative HPLC to give 7-(4- chlorobenzyl)-8-(isopropylsulfonyl)-l-(isoxazol-5-ylmethyl)-3-methyl-lH-purine-2,6(3H,7H)- dione (39 mg, 40.6% yield) as a white solid. ^-NMR (CD3OD) δ 8.49(d, 1H), 7.41(dd, 2 H), 7.28(d, 2H), 6.43(d, 1 H), 5.87(s, 2H), 5.2 l(s, 2H),3.60-3.73(m, 1 H), 3.47(s, 3H), 1.25(d, 6H). LCMS retention time 2.772 min; LCMS MH+ 478
Example 70 7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl-8-(methylsulfonyl)-lH-purine- 2,6(3H,7H)-dione
Figure imgf000526_0001
Step 1 7-(4-chlorobenzyl)-3-methyl-8-(methylthio)-l -(3-(tetrahydi -2M-pyran-2- yloxy)propyl)- -purine-2,6(3H,7 -dione
Figure imgf000526_0002
8-bromo-7-(4-chIorobenzyl)-3-methyI-l-(3-(tetrahydro-2H-pyran-2-y[oxy)propyl)-lH-purine- 2,6(3H,7H)-dione (0.15 g, 0.35 mmol, intermediate 14) was dissolved in sodium thiomethoxide (5 mL). The reaction was heated at 80 °C overnight. The mixture was cooled and partitioned between ethyl acetate and water. The combined organic layer was dried over sodium sulfate, filtered and concentrated to give a crude product which was used without purification. LCMS retention time 1.875 min; LCMS MH+-THP 395.
Step 2 7-(4-chlorobenzyl)-l -(3-hydroxypropyl)-3-methyl-8-(methylsulfonyl)-lH-purine- 2,6(3H,7H)-dione
Figure imgf000526_0003
To a solution of 7-(4-chlorobenzyl)-3-methyl-8-(methylthio)-l-(3-(tetrahydro-2H-pyran-2- yloxy)propyl)-lH-purine-2,6(3H,7H)-dione (0.14 g, 0.35mmol) in chloroform (10 mL) was added MCPBA (0.15 g, 0.87 mmol) portionwise at 0°C. The reaction was stirred at room temperature overnight. The mixture was partitioned between ethyl acetate and water. The combined organic layer was dried over sodium sulfate, filtered and concentrated to give a yellow oil. This oil was dissolved in ethanol (5 mL); then acetyl chloride (0.2 mL) was added dropwise at 0 °C. The reaction was stirred at this temperature for 1 h. The mixture was concentrated to give a crude product which was purified by preparative HPLC to give7-(4-chlorobenzyl)-l -(3- hydroxypropyl)-3-methyl-8-(methylsulfonyl)-lH-purine-2,6(3H,7H)-dione (15 mg, 10.0% yield) as a white solid. ^-NMR (CD3OD) δ 7.34-7.42 (m, 4 H), 5.97(s, 2 H), 4.1 l (t, 2H), 3.61 (t, 2 H), 3.57(s, 3 H), 3.38(s, 3 H),1.84-1.89(m, 2 H). LCMS retention time 2.052 min; LCMS MH+ 427
Example 71 7-(4-chlorobenzyl)-8-(ethylsulfonyl)-l -(3-hydroxypropyl)-3-methyl-l H-purine- -dione
Figure imgf000527_0001
Step 1 7-(4-chlorobenzyl)-8-(ethylthio)-3 -methyl- 1 -(3 -(tetrahydro-2H-pyran-2-yloxy)propy 1)- 1 H-purine-2 -dione
Figure imgf000527_0002
To a solution of 8-bromo-7-(4-chlorobenzyl)-3 -methyl- 1 -(3 -(tetrahydro-2H-pyran-2- yloxy)propyl)- lH-purine-2,6(3H,7H)-dione (0.15 g, 0.29 mmol, intermediate 14) in DMF (10 mL) was added ethanethiol (27 mg, 0.44 mmol) and potassium carbonate (81 mg, 0.58 mmol). The reaction was heated at 80 °C overnight. The mixture was cooled and partitioned between ethyl acetate and water. The combined organic layer was dried over sodium sulfate, filtered and concentrated to give a crude product which was used without purification. LCMS retention time 1.912 min; LCMS MH+-THP 409.
Step 2 7-(4-chlorobenzyl)-8-(ethylsulfonyl)-l -(3 -hydroxypropyl)-3 -methyl- lH-purine- 2,6(3H,7H)-dione
Figure imgf000527_0003
The title compound was prepared using the method of example 70, step 2 and purified by preparative HPLC to give 7-(4-chlorobenzyl)-8-(ethylsulfonyl)-l -(3-hydroxypropyl)-3-methyl- lH-purine-2,6(3H,7H)-dione (26 mg, 20.5% yield) as a white solid. 'H-NMR (CD3OD) δ= 7.34-7.41(m, 4H), 5.98(s, 2H), 4.1 l (t, 1H), 3.62(t, 2H), 3.57(s, 3H),3.47-3.53(m, 2H), 1.85- 1 .90(m, 2H), 1.31 (t, 3H). LCMS retention time 2.192 min; LCMS MH+ 441.
Example 72 7-(4-chIorobenzyl)- l -(3-hydroxypropyl)-8-(isopropylsulfonyl)-3-methyl-l H- purine- -dione
Figure imgf000528_0001
The title compound was prepared using the method of example 71 and purified by preparative HPLC to give 7-(4-chlorobenzyl)-8-(ethylsulfonyl)-l -(3-hydroxypropyl)-3-methyl-l H-purine- 2,6(3H,7H)-dione (26 mg, 20.5% yield) as white solid. 'H-NMR (CDC13) δ 7.43-7.45(d, 2Η), 7.31-7.34 (d, 2Η), 5.97(s, 2Η), 4.19-4.22(t, 2Η), 3.73-3.76 (m, 1Η), 3.59 (s, 3Η), 3.54-3.59 (m, 2Η), 3.04-3.06 (t, 1Η), 1 .90-1.93 (m, 2Η), 1.38-1.40 (d, 6Η). LCMS retention time 2.31 1 rn'in; LCMS ΜΗ+ 455.
Example 73 7-(4-chlorobenzyl)-l -(3-hydroxypropyl)-3-mei
(trifluoromethoxy)phenylthio)-lH-purine-2,6(3H,7H)-dione
Figure imgf000528_0002
Step 1 7-(4-chlorobenzyl)-3-methyl-l-(3-(tetrahydro-2H-pyran-2-yloxy)propy])-8-(3
(trifluoromethoxy)phenylthio)-lH-purine-2,6(3H,7H)-dione
Figure imgf000528_0003
The title compound was prepared using the method of example 71, step 1 and using 3-
(trifluoromethoxy)benzenethiol. Yellow oil, 100 mg, 81.9% yield. LCMS retention time 2.098 min; LCMS MH+-THP 541
Step 2 7-(4-chlorobenzyl)-l -(3-hydroxypropyl)-3-methyl-8-(3-(trifluoromethoxy)phenylthio)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000529_0001
The title compound was prepared using the method of example 37, step 2. Light yellow solid, 20 mg, 23.1 % yield: V -NMR (DMSO-< ) δ 7.47-7.42(t, IH), 7.33-7.30(m, 5H), 7.24-7.22(d, 2H), 5.63(s, 2H), 4.49-4.46(t, IH), 3.95-3.92(t, 2H), 3.47-3.42(m, 2H), 3.39(s, 3H), 1.72- 1.69(m, 2H). LCMS retention time 2.950 min; LCMS MH+ 541
Example 74 8-(butylsulfonyl)-7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl- lH-purine- 2,6(3H,7H)-dione
Figure imgf000529_0002
The title compound was prepared using the method of example 71. Light yellow oil, 30 mg, 39.3% yield: XH-NMR (CD3OD) δ 7.40-7.35(m, 4H), 5.99(s, 2H), 4.14-4.10(t, 2H), 3.64- 3.61 (t, 2H), 3.57(s, 3H), 3.44-3.40(m, 2H), 1.90-1 .86(m, 2H), 1.69-1.65(m, 2H), 1.44-1.38(m, 2H), 0.92-0.89(t, 3H). LCMS retention time 1.671 min; LCMS MIL" 469 Example 75 7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl-8-(propylsulfonyl)-l H-purine- 2,6(3H,7H)-dione
Figure imgf000529_0003
The title compound was prepared using the method of example 71. Light brown solid, 30 mg, 39.5% yield: *H-NMR (CD3OD) δ 7.40-7.34(m, 4H), 5.99(s, 2H), 4.13-4.09(t, 2H), 3.63- 3.60(t, 2H), 3.57(s, 3H), 3.43-3.39 (t, 2H), 1 .89-1.85(m, 2H), 1.80-1 .75(m, 2H), 1.04-0.99(t, 3H). LCMS retention time 2.353 min; LCMS MH+ 455 Example 76 7-(4-chlorobenzyl)-l -(3-hydroxypropyl)-3-methyl-8-(propylsulfonyl)-lH-purine- 2,6(3H,7H)-dione
Figure imgf000530_0001
Step 1 7-(4-chlorobenzyl)-3 -methyl- 1 -(3-(tetrahydro-2IT-pyran-2-yloxy)propyl)-8-(3- (trifluoromethoxy)benzylthio)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000530_0002
To a solution of 7-(4-chlorobenzyl)-8-mercapto-3-methyl-l-(3-(tetrahydro-2H-pyran-2- yloxy)propyl)-lH-purine-2,6(3H,7H)-dione (100 mg, 0.216 mmol) in DMF (5 mL) was added 1- (bromomethyl)-3-(trifluoromethoxy)benzene (66 mg, 0.259 mmol), followed by potassium carbonate (89 mg, 0.645 mmol). The reaction was stirred at 80 °C for 2 h. The mixture was cooled, diluted with ethyl acetate and water, and the phases were separated. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give 7-(4- chlorobenzyl)-3-methyl-l -(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-8-(3- (trifluoromethoxy)benzylthio)-lH-purine-2,6(3H,7H)-dione (120mg, 87.3%) as light yellow oil. LCMS retention time 2.106 min; LCMS MH+-THP 555.
Step 2 7-(4-chlorobenzy])- l-(3-hydroxypropyl)-3-methyl-8-(propylsulfonyl)-lH-purine- 2,6(3H,7H)-dione
Figure imgf000531_0001
The title compound was prepared using the method of example 70, step 2. White solid, 14.3% yield: J-NMR (CD3OD) δ 7.47-7.43(t, 1H), 7.32-7.30(d, 2H), 7.27-7.23(d, 2H), 7.18-7.16(d, 3H), 5.64(s, 2H), 4.91 (s, 2H), 4.10-4.07(t, 2H), 3.62-3.59(s, 5H), 1.88-1 .81 (m, 2H). LCMS retention time 2.922 min; LCMS MH+ 587.
Example 77 7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl-8-(3- (trifluoromethoxy)benzylsulfinyl)-l H-purine-2,6(3H,7H)-dione
Figure imgf000531_0002
Step 1 7-(4-chlorobenzyl)-3 -methyl- 1 -(3 -(tetrahydro-2H-pyran-2-y loxy)propyl)-8-(3 - (trifiuoromethoxy)benzylsulfinyl)-lH-purine-2,6(3H,7H)-dione
Figure imgf000531_0003
To a solution of 7-(4-chlorobenzyl)-3 -methyl- 1 -(3 -(tetrahydro-2H-pyran-2-y loxy)propyl)-8-(3 - (trifluoromethoxy)benzylthio)-lH-purine-2,6(3H,7H)-dione (60 mg, 0.108 mmol, example 129, step 2) in THF (4 mL) and water (2 mL) was added Oxone (73 mg, 0.1 19 mmol) at 0°C. The resulting mixture was stirred at room temperature for 16 h. The mixture was quenched with aqueous sodium thiosulfate, partitioned between DCM and water, and the phases were separated. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give crude product. This crude material was purified by silica gel chromatography eluting with petroleum/ethyl acetate (1 :0 to 1 : 1) to give 7-(4-chlorobenzyl)-3 -methyl- 1 -(3 -(tetrahydro-2H- pyran-2-yloxy)propyl)-8-(3-(trifluoromethoxy)benzylsulfinyl)-l H-purine-2,6(3H,7H)-dione (60 mg, 97.6%) as yellow oil. LCMS retention time 1.499 min; LCMS MH+-THP 571 . Step 2 7-(4-chlorobenzyl)-l -(3-hydroxypropyl)-3-methyl-8-(3- (trifluoromethoxy)benzylsulfinyl)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000532_0001
The title compound was prepared using the method of example 37, step 2. White solid, 30 mg, 57.4% yield: 'H-NMR (CD3OD) δ 7.43-7.39(t, 1 H), 7.27-7.23(m, 4H), 7.17-7.12(m, 3H), 5.59(s, 2H), 4.89-4.74(m, 2H), 4.12-4.08(t, 2H), 3.62-3.59(m, 5H), 1.88-1.84(m, 2H). LCMS retention time 2552 min; LCMS MH+ 571.
Example 78 7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-N-isopropyl-3-methyl-2,6-dioxo-2,3,6,7- tetrahydro-lH-purine-8-sulfonamide
Figure imgf000532_0002
Step 1 7-(4-chlorobenzyl)-N-isopropyl-3-methyl-2,6-dioxo-2,3,6,7-tetrahydro-l H- purine-8-sulfonamide
Figure imgf000532_0003
To a solution of 7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-2,3,6,7-tetrahydro-lH-purine-8-sulfonyl chloride (1 15 mg, 0.295 mmol, intermediate 64) in DCM (5 mL) was added isopropylamine (17.4 mg, 0.59 mmol) followed by TEA (60 mg, 0.59 mmol) at 0 °C and the mixture was stirred at room temperature for 1 h. The mixture was diluted with DCM and washed brine. The organic phase was dried and concentrated to give a crude product, which was purified by silica gel chromatography eluting with DCM /methanol (50: 1) to give 7-(4-chlorobenzyl)-N-isopropyl-3- methyl-2,6-dioxo-2,3,6,7-tetrahydro-lH-purine-8-sulfonamide (59 mg, 48.6% yield) as yellow syrup. LCMS retention time 1.436 min; LCMS MH+ 412.
Step 2 7-(4-chlorobenzyl)-N-isopropyl-3-methyl-2,6-dioxo-l-(3-(tetrahydro-2H-pyran-2- yloxy)propyl)-2,3,6,7-tetrahydro-l H-purine-8-sulfonamide
Figure imgf000533_0001
To a solution of 7-(4-chlorobenzyl)-N-isopropyl-3-methyl-2,6-dioxo-2,3,6,7-tetrahydro-lH- purine-8-sulfonamide (59 mg, 0.143 mmol) in DMF (3 mL) was added 2-(3- bromopropoxy)tetrahydro-2H-pyran (32 mg, 0.143 mmol) followed by potassium carbonate (30 mg, 0.214 mmol) and the mixture was stirred at 60 °C for 4 h. The reaction was diluted with ethyl acetate and washed with brine and saturated aqueous ammonium chloride. The organic phase was dried and concentrated to give 7-(4-chlorobenzyl)-N-isopropyl-3-methyl-2,6-dioxo-l - (3-(tetrahydro-2H-pyran-2-yloxy)propyl)-2,3,6,7-tetrahydro-lH-purine-8-sulfonamide (61 mg, 77% yield) as yellow solid. LCMS retention time 1.805 min; LCMS MH+-THP 470.
Step 3 7-(4-chlorobenzyl)-N-isopropyl-3-methyl-2,6-dioxo-l -(3-(tetrahydro-2H-pyran-2- yloxy)propyl)-2,3 6,7-tetrahydro-lH-purine-8-sulfonamide
Figure imgf000533_0002
A solution of 7-(4-chlorobenzyl)-N-isopropyl-3-methyl-2,6-dioxo-l-(3-(tetrahydro-2H-pyran-2- yloxy)propyl)-2,3,6,7-tetrahydro-l H-purine-8-sulfonamide (61 mg, 0.1 1 mmol) in 1 mM ethanolic HC1 (1 mL, lmmol/L) was stirred at room temperature for 30 min. The mixture was concentrated and purified by preparative HPLC to give 7-(4-chlorobenzyl)-l -(3-hydroxypropyl)- N-isopropyl-3-methyl-2,6-dioxo-2,3,6,7-tetrahydro-l H-purine-8-sulfonarnide (17.9 mg, 34.6% yield) as white solid. 'H-NMR (CD3OD) δ 7.31-7.38(m, 4Η), 5.89(s, 2Η), 4.06-4.10 (t, 2Η), 3.58-3.63 (m, 3Η), 3.56 (s, 3Η), 1 .83-1.87 (m, 2Η), 1 .19-1.21 (d, 6Η). LCMS retention time 2.315 min; LCMS MH+ 470.
Example 79 7-(4-chlorobenzyl)-N-cyclopropyl-N,l-bis(3-hydroxypropyl)-3-methyl-2,6-dioxo- 2,3,6 -tetrahydro-l H-purine-8-sulfonamide
Figure imgf000534_0001
The title compound was prepared using the method of example 78 except in step 2, 2 equivalent of 2-(3-bromopropoxy)tetrahydro-2H-pyran was added. The final product was purified via preparative HPLC to give 7-(4-chlorobenzyl)-N-cyclopropyl-N,l-bis(3-hydroxypropyl)-3- methyl-2,6-dioxo-2,3,6,7-tetrahydro-l H-purine-8-sulfonamide (21 mg, 38.9% yield) as white solid. 'H-NMR (CD3OD) δ 7.32-7.39(m, 4Η), 5.92(s, 2Η), 4.07-4.1 1 (t, 2Η), 3.57-3.62 (m, 6Η), 3.53 (s, 3Η), 2.71-2.75 (m, 1H),1.84-1.95 (m, 4H), 0.91-0.94 (m, 2H), 0.75-.078 (m, 2H). LCMS retention time 2.238 min; LCMS MH+ 526.
Example 80 N-(7-(4-chlorobenzyl)-l -(3-hydroxypropyl)-3-methyl-2,6-dioxo-2,3,6,7- tetrahydro- 1 H-purin-8-yl)propane-2-sulfonamide
Figure imgf000534_0002
Step 1 N-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-l-(3-(tetrahydro-2H-pyran-2-yloxy)propyl)- 2,3,6,7-tetrahydro- -purin-8-yl)propane-2-sulfonamide
Figure imgf000534_0003
To a solution of 8-amino-7-(4-ch'lorobenzyl)-3 -methyl- 1 -(3-(tetrahydro-2H-pyran-2- yloxy)propyl)-l H-purine-2,6(3H,7H)-dione (95 mg, 0.21 mmol, intermediate 53) in THF (5 mL) was added sodium hydride (10 mg, 0.23 mmol) at 0 °C. The reaction was stirred at room temperature for 1 h; then Isopropyl sulphonyl chloride (29 mg, 0.21 mmol) was added dropwise at 0°C. The mixture was stirred at room temperature 1 h. The reaction was quenched with aqueous ammonium chloride (2 mL) at 0 °C and then was partitioned between ethyl acetate and water. The combined organic layer was dried over sodium sulfate, filtered and concentrated to give the product as a yellow syrup(60 mg) which was used without purification. LCMS retention time 1.519 min; LCMS MH+ -ΊΉΡ 470.
Step 2 N-(7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl-2,6-dioxo-2,3,6,7-tetrahydro-lH- purin-8-yl)propane-2-sulfonamide
Figure imgf000535_0001
The title compound was prepared using the method of example 37, step 2 and purified by preparative HPLC. White solid, 19 mg, 37.2% yield: 'H-NMR (CD3OD) δ 7.44(d, 2H), 7.31 - 7.33(m, 2H), 5.37(s, 2H), 4.19(t, 2H), 3.56(s, 2H), 3.52(s, 3H), 3.39(brs, 1). 3.10(brs, 1H), 1 .88- 1.94 (m, 2H), 1.40(d, 6H). LCMS retention time 1.942 min; LCMS MH+ 470
Example 81 7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl
Figure imgf000535_0002
Stepl 7-(4-chlorobenzyl)-3-methyl-l-(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-8-(3- (trifluoromethoxy)phenylamino)-lH-purine-2,6(3H,7H)-dione (WYS-000356-026)
Figure imgf000536_0001
To a solution of 8-bromo-7-(4-chIorobenzyl)-3-methyl- l -(3-(tetrahydro-2H-pyran-2- yloxy)propyl)-lH-purine-2,6(3H,7H)-dione (100 mg, 0.196 mmol, Intermediate 14) in toluene(30 mL) was added 3-(trifluoromethoxy)aniline (45 mg, 0.254 mmol),
tris(dibenzylideneacetone)dipalladium(0) (10 mg, 0.01 1 mmol), X-phos (10 mg, 0.021 mmol) and potassium tert-butanolate (45 mg, 0.402 mmol). The mixture was degassed and refilled with nitrogen 3 times. The reaction was stirred at 100 °C overnight under nitrogen. The mixture was filtered and the. filter cake was washed with ethyl acetate. The filtrate was concentrated to give a crude product which was purified by silica gel chromatography eluting with petroleum/ethyl acetate (3: 1 to 3 :2). The product was collected, concentrated, and dried in vacuo to give 7-(4- chlorobenzyl)-3-methyl-l-(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-8-(3-
(trifluoromethoxy)phenylamino)-lH-purine-2,6(3H,7H)-dione (70 mg, 57.6%) as a yellow solid. LCMS MH+-THP 524. Step2 7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl-8-(3- (trifluoromethoxy)phenylamino)-lH-purine-2,6 -dione
Figure imgf000536_0002
The title compound was prepared as example 36, step 2 but with aqueous ethanolic HC1. White solid, 13 mg, 12.4% yield: 'H-NMR (DMSO-i¾ δ 9.63(s,lH), 7.91 (s,lH), 7.63-7.61 (d,l H), 7.45-7.38(m,3H), 7.24-7.22(d,2H), 6.97-6.94(d,lH), 5.57(s,2H), 4.44-4.42(t3 lH), 3.89-
3.86(t,2H), 3.42-3.38(m,4H), 3.31(s,3H), 1.69-1.6 l (m,2H). LCMS retention time 2.853 min; LCMS MH+ 524.
Example 82 7-(4-chlorobenzyl)- l-((l-(hydroxymethyl)cyclopropyl)methyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000537_0001
To a solution of 7-(4-chIorobenzyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)- lH-purine- 2,6(3H,7H)-dione (80 mg, 0.17 mmol, intermediate 9) in DMF (5 mL) was added (1 - (hydroxymethyl)cyclopropyl)methyl methanesulfonate (0.1 g, 0.5 mmol, intermediate 42), potassium carbonate (47 mg, 0.34 mmol) and TBAI (2 mg, 0.02 mmol). The reaction was stirred at 50 °C for 8 h. The mixture was cooled and partitioned between ethyl acetate and water. The combined organic layer was dried over sodium sulfate, filtered and concentrated to give a crude product which was purified by preparative HPLC to give 7-(4-chlorobenzyl)-l -((l - (hydroxymethyl)cyclopropyl)methyl)-3-methyl-8-(3(trifluoromethoxy)phenoxy)- lH-purine- 2,6(3H,7H)-dione (36 mg, 39.9% yield) as a white solid. 'H-NMR (CD3OD) δ 7.39(t, 1 H), 7.01(dd, 1 H), 6.88-6.90(m , 2H), 4.87-4.88(m, 2H), 4.44-4.46(m, 2H), 4.08-4.16(m, 4H), 3.61(t, 2H), 3.51 (s, 3H), 1.80-1.90 (m, 2H), 1.32(t, 3H). LCMS retention time 3.099 min; LCMS MH+ 533.
The following examples 83a and 83d were prepared using the method of example 82.
Example 83a 7-(4-chlorobenzyl)-l -(2-(l-hydroxycyclopentyl)ethyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000537_0002
The title compound was prepared from Intermediates 9 and 43. White solid, 18 mg, 17.9% yield 7J-NMR (CD3OD) S 7.54(t, 1H), 7.44(dd, 2H), 7.32-7.36(m, 4H), 7.22-7.25(m , 1H), 5.48(s, 2H), 4.14-4.18(m, 2H), 3.51 (s, 3H), 1.88-1.92(m, 2H), 1.62-1.82(m, 8H). LCMS retention time 3.304 min; LCMS MH+ 561 Example 83b 7-(4-chlorobenzyl)-l -(2-(l -hydroxycyclobutyl)ethyl)-3-methyl-8-(3- (trifluoromelhoxy)phenoxy)-l H-purine-2,6(3H,7H)-dione
Figure imgf000538_0001
The title compound was prepared from intermediates 9 and 44. White solid, 20 mg, 17.9% yield: 'H-NMR (CD3OD) δ 7.55(t5 IH), 7.54(d, 2H), 7.32-7.38(m, 4H), 7.23-7.26(m , IH), 5.51(s, 2H), 4.1 1-4.15(m, 2H), 3.50(s, 3H), 2.05-2.17(m, 4H), 1.93-1.97(m, 2H), 1.76-1.80(m, IH) 1.59- 1.63(m, IH). LCMS retention time 3.1 1 1 min; LCMS MI-1+ 547
Example 83c 7-(4-chlorobenzyl)-l -(3-hydroxy-3-methylbutyl)-3-methyl-8-(3- (trifluoromethox henox -lH- urine-2 6 3H 7H -dione
Figure imgf000538_0002
The title compound was prepared from intermediates 9 and 45. White solid, 23 mg, 39.6% yield: 1H-NMR (CD3OD) δ 7.55(t, IH), 7.44(d, 2H), 7.32-7.38(m, 4H), 7.24(d, IH), 5.50(s, 2H), 4.1 1-4.15(m, 2H), 3.42(s, 3H), 1.78-1.82(m, 2H), 1.29(s, 6H). LCMS retention time 3.235 min; LCMS MH+ 535
Example 83d 7-(4-chlorobenzyl)-l -(2-(l -hydroxycyclopropyl)ethyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000538_0003
The title compound was prepared from intermediates 9 and 46. White solid, 39 mg, 33.9% yield: 'H-NMR (CD3OD) δ 7.55(t, 1Η), 7.44(d, 2Η), 7.34-7.36(m, 4Η), 7.25(d, 1Η), 5.49(s, 2Η), 4.26(t, 2H), 3.42(s, 3H),1.84(t, 2H), 0.62(t, 2H), 0.38(t, 2H). LCMS retention time 3.301 min; LCMS MH+ 551
Example 84 7-(4-chlorobenzyl)-8-(2-(cyclohexyloxy)ethoxy)- 1 -(3-hydroxypropyl)-3-methyl- l H-purine-2,6(3H,7H)-dione
Figure imgf000539_0001
Step 1 7-(4-chlorobenzyl)-8-(2-(cyclohexyloxy)ethoxy)-3-methyl-l -(3-(tetrahydro-2H- pyran-2-ylox
Figure imgf000539_0002
To a solution of 2-(cyclohexyloxy)ethanol (63.4 mg, 0.439 mmol, intermediate 38) in THF (5 mL) was added sodium hydride (23.4 mg, 0.586 mmol, 60% dispersion in mineral oil) at 0 °C, and the mixture was stirred at room temperature for 30 min. Then a solution of 8-bromo-7-(4- chlorobenzyl)-3-methyl-l -(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-lH-purine-2,6(3H,7H)- dione (150 mg, 0.293 mmol, intermediate 14) in THF (1 mL) was added and the resulting mixture was stirred at reflux for 1 h. The reaction was cooled and quenched with ice-water. The mixture was extracted with ethyl acetate. The organic phase was dried and concentrated to give 7-(4-chlorobenzyl)-8-(2-(cyclohexyloxy)ethoxy)-3-methyl-l-(3-(tetrahydro-2H-pyran-2- yloxy)propyl)-l H-purine-2,6(3H,7H)-dione (123 mg, 73% yield) as a yellow solid. LCMS retention time 2.064 min; LCMS MHT-THP 491.
Step 2 7-(4-chlorobenzyl)-8-(2-(cyclohexyloxy)ethoxy)-l -(3-hydroxypropyl)-3-methyl- lH-purine-2,6(3H,7H)-dione
Figure imgf000539_0003
A solution of 7-(4-chlorobenzyl)-8-(2-(cyclohexyloxy)ethoxy)-3-methyl-l -(3-(tetrahydro-2H- pyran-2-yloxy)propyl)-lH-purine-2,6(3H,7H)-dione (123 mg, 0.214 mmol) in ImM ethanolic HC1 (2 mL) was stirred at room temperature for 30 min. The reaction was concentrated to dryness to give a crude product, which was purified by preparative HPLC to give 7-(4- chlorobenzyl)-8-(2-(cyclohexyloxy)ethoxy)-l -(3-hydroxypropyl)-3-methyl-lH-purine- 2,6(3H,7H)-dione (64 mg, 60.9% yield) as a white solid. 'H-NMR (CDC13) δ 7.41 -7.44 (d, 2H), 7.28-7.31 (d, 2H), 5.26 (s, 2H), 4.63-4.65 (m, 2H), 4.17-4.20 (t, 2H), 3.81 -3.84 (m, 2H), 3.68- 3.70 (br, 1H), 3.21 (m, 5H), 3.30-3.34 (m, 1H), 1.87-1.93 (m, 4H), 1.75-1.77 (t, 2H), 1.55-1.58 (m, 1H), 1.23-1.36 (m, 5H). LCMS retention time 2.893 min; LCMS MH+ 491.
Example 85 7-(4-chlorobenzyl)-l -(3-hydroxypropyl)-3-methyl-8-(2-propoxyethoxy)- l H- purine-2,6(3H,7H)-dione
Figure imgf000540_0001
The title compound was prepared using the method of example 84 and purified by preparative HPLC to give 7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl-8-(2-propoxyethoxy)-l H- purine-2,6(3H,7H)-dione (39 mg, 38.7% yield) as a white solid. yH-NMR (CD3OD) 8 7.39- 7.42(d, 2H), 7.31 -7.33 (d, 2H), 5.29 (s, 2H), 4.63-4.65 (m, 2H), 4.06-4.10 (t, 2H), 3.80-3.83 (m, , 2H), 3.58-3.61 (t, 2H), 3.49 (s, 3H), 3.44-3.47 (t, 2H), 1.84-1.87 (t, 2H), 1.55-1.60 (m, 2H), 0.90- 0.94 (t, 3H). LCMS retention time 2.561 min; LCMS MH+ 451.
Example 86 7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl-8-(6-methylpyridin-3-yloxy)- 1 H-purine-2,6(3H,7H)-dione hydrochloride
Figure imgf000540_0002
The title compound was prepared from intermediate 63 using the method of example 55 except with 6-methylpyridin-3-ol to give 35 mg (44.1% yield) as white solid. 'H-NMR (DMSO-<¾) δ 8.69(s, 1H), 8.03-8.01 (d, 1H), 7.59-7.57(d, 1H), 7.45(s, 4H), 5.45(s, 2H), 4.06-3.98(m, 2H), 3.45-3.42(t, 2H), 3.28(s, 3H), 2.59(s, 3H), 1 .71 - 1.67(1, 2H). LCMS retention time 2.207 min; LCMS MH+ 456.
Example 87 7-(4-chlorobenzyl)- 1 -(3 -hydroxypropyl)-3 -methyl-8-(2-methy lpyridin-4-yloxy)- 1 H-purine-2,6(3H,7H)-dione hydrochloride
Figure imgf000541_0001
HCI
The title compound was prepared using the method of example 55. White solid, 30 mg, 36.6% yield: 'H-NMR (DMSO-i¾ δ 8.48-8.46(d, 1H), 8.17(s, 1H), 7.43-7.33(m, 4H), 7.26-7.22(m, 2H), 5.42(s, 2H), 4.52(s, 1H), 3.95-3.92(t, 2H), 3.44-3.42(m, 2H), 3.37(s, 3H), 2.47(s, 3H), 1.72- 1.69(t, 2H). LCMS retention time 1.716 min; LCMS MH+ 456.
Example 88 7-(4-chlorobenzyl)-l -(3-hydroxypropyl)-3-methyl-8-(5-methylpyridin-3-yloxy)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000541_0002
The title compound was prepared using the method of example 55. White solid, 40 mg, 48.8% yield: 'H-NMR (DMSO-^) 5 8.45-8.44(d, 1H), 8.36(s, 1H), 7.65(s,lH), 7.46-7.41 (m, 4H), 5.44(s, 2H), 4.50-4.47(t, 1H), 3.94-3.91(t, 2H), 3.46-3.40(m, 2H), 3.28(s, 3H), 2.34(s, 3H), 1.73- 1.66(m, 2H). LCMS retention time 2.259 min; LCMS MH+ 456 Example 89 7-butyl-l-(3-hydroxypropyl)-3-methyl-8-(5-methylpyridin-3-yloxy)-lH-purine- 2,6(3H,7H)-dione
Figure imgf000542_0001
Step 1 3-methyl-8-(5-methylpyridin-3-yloxy)-l -(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-7- ((2-(tr
Figure imgf000542_0002
The title compound was prepared using the method of example 52, step 1. Yellow oil, 300 mg, 97.6% yield: LCMS retention time 1.838 min; LCMS MH+-THP 462
Step 2 7-(4-chlorobenzyl)-l -(3-hydroxybutyl)-3-methyl-8-(6-methylpyridin-3-yloxy)-lH- purine-2,6(3H,7H)-dione
Figure imgf000542_0003
To a solution of 3-methyl-8-(5-methylpyridin-3-yloxy)-l -(3-(tetrahydro-2H-pyran-2- yloxy)propyl)-7-((2-(trimethylsilyl)ethoxy)methyl)-lH-purine-2,6(3H,7H)-dione (300 mg, 0.554 mmol) in ethyl alcohol (10 ml) was added concentrated HCl (3 ml). The reaction was stirred for. 6 h at 90 °C. The reaction was cooled and concentrated to give 7-(4-chlorobenzyl)-l-(3- hydroxybutyl)-3-methyl-8-(6-methylpyridin-3-yloxy)-l H-purine-2,6(3H,7H)-dione (180 mg, 98.9%) as yellow oil. LCMS retention time 0.356 min; LCMS MH+ 332.
Step 3 7-butyl-l -(3-hydroxypropyl)-3-methyl-8-(5-methylpyridin-3-yloxy)-lH-purine- 2,6(3H,7H)-dione
Figure imgf000543_0001
The title compound was prepared using the method of example 61 , step 2. White solid, 30 mg, 42.8% yield: 'H-NMR (DMSO-i¾ δ 8.48-8.47(d, 1H), 8.37-8.36(d, 1H), 7.71 (s, 1H), 4.51 - 4.48(t, 1 H), 4.22-4.19(t, 2H), 3.94-3.91(t, 2H), 3.46-3.43(t, 2H), 3.28(s, 3H), 2.36(s, 3H), 1.81 - 1 .66(m, 4H), 1 .35-1.30(m, 2H), 0.93-0.89(t3 3H). LCMS retention time 1.854 min; LCMS MH+ 388.
Example 90 7-(4-chlorobenzyl)-8-(2-isopropoxyethoxy)-3-methyl-l -(3-(tetrahydro-2H-pyran- 2-yloxy)propyl)-lH-purine-2,6(3H,7H)-dione
Figure imgf000543_0002
Step 1 7-(4-chlorobenzyl)-8-(2-isopropoxyethoxy)-3-methyl- 1 -(3-(tetrahydro-2H-pyran-2- yloxy)propy -lH-purine-2,6(3H,7H)-dione
Figure imgf000543_0003
The title compound was prepared using the method of example 50, step 1. Yellow oil, 100 mg, 95.2%. LCMS retention time 1.864 min; LCMS MH+-THP 451
Step 2 7-(4-chlorobenzyl)-8-(2-isopropoxyethoxy)-3-methyl-l -(3-(tetrahydro-2H-pyran-2- yloxy)propyl)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000543_0004
The title compound was prepared using the method of example 37, step 2. White solid, 30 mg, 35.1% yield: 'H-NMR (CD3OD) δ 7.43-7.41(d, 2Η), 7.34-7.32(d, 2Η), 5.30(s, 2Η), 4.66- 4.64(m, 2Η), 4.1 1-4.07(t, 2Η), 3.83-3.81(m, 2Η), 3.68-3.58(m, 3Η), 3.50(s, 3Η), 1.88-1.85(m, 2Η), 1.17-1.56(d, 6H). LCMS retention time 2.464 min; LCMS MH+ 451
Example 91 7-(4-chlorobenzyl)-l -(3-hydroxypropyl)-3-methyl-8-(3- (trifluoromethoxy)benzoyl)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000544_0001
Step 1 7-(4-chlorobenzyl)-l -(4-methoxybenzyl)-3-methyl-8-(3-(trifluoromethoxy)benzoyl)- lH-purine-2,6(3H,7H)-dione
Figure imgf000544_0002
The title compound was prepared using the method of intermediate 16, step 3. White solid, 15 mg, 21.5% yield. LCMS retention time 2.159 min; LCMS MH+ 599. Step 2 7-(4-chlorobenzyl)-3-methyl-8-(3-(trifluoromethoxy)benzoyl)-lH-purine-2,6(3H,7H)- dione
Figure imgf000544_0003
To a solution of 7-(4-chlorobenzyl)-l-(4-methoxybenzyl)-3-methyl-8-(3- (trifluoromethoxy)benzoyl)-lH-purine-2,6(3H,7H)-dione (15 mg, 0.025mmol) in DCM (1 ml) was added TFA (1 ml) dropwise followed by trifluoromethanesulfonic acid (0.25 ml), both at 0 °C. The reaction was stirred at 0 °C for 10 min, then stirred at room temperature for 16 h. The reaction was quenched with aqueous saturated sodium bicarbonate and extracted with DCM. The organic phase was washed with water dried over sodium sulfate, filtered and concentrated to give 7-(4-chlorobenzyl)-3-methyl-8-(3-(trifluoromethoxy)benzoyl)-lH-purine-2,6(3H,7H)-dione (l Omg, 83.7%) as yellow oil. LCMS retention time 1.821 min; LCMS MIT" 479.
Step 3 7-(4-chlorobenzyl)-3-methyl- l-(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-8-(3- (trifluoromethoxy)benzoyl)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000545_0001
The title compound was prepared using the method of intermediate 14. Yellow oil, 10 mg, 76.8% yield: LCMS retention time 2.176 min; LCMS MH+ -THP 537.
Step 4 7-(4-chlorobenzyl)- l-(3-hydroxypropyl)-3-methyl-8-(3-(trifluoromethoxy)benzoyl)- purine-2,6(3H,7H)-dione
Figure imgf000545_0002
The title compound was prepared using the method of example 37, step 2. Light yellow solid, 8 mg, 92.6% yield: 'H-NMR (DMSO-c¾ δ 8.20-8.16(m, 2H), 7.74-7.72(m, 2H), 7.40-7.37(d, 2H), 7.30-7.28(d, 2H), 5.93(s, 2H), 4.51(s, 1H), 3.97-3.93(t, 2H), 3.47-3.39(m, 5H), 1.73- 1.69(m, 2H). LCMS retention time 2.996 min; LCMS MH+ 537
Example 92 7-(4-chlorobenzyl)-8-(hydroxy(3-(trifluoromethoxy)phenyl)methyl)-l-(3- hydroxypropyl)-3-methyl-l H-purine-2,6(3H,7H)-dione
Figure imgf000546_0001
To a solution of 7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl-8-(3- (trifluoromethoxy)benzoyl)-lH-purine-2,6(3H,7H)-dione (50 mg, 0.081 mmol, example 91) in methanol (3 mL) was added sodium borohydride (10 mg, 0.263 mmol) at 0 °C under nitrogen. The reaction was stirred at 0°C for 2 h. The reaction was quenched and partitioned with ethyl acetate and water. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give a crude product, which was purified by preparative HPLC to give 7-(4- chlorobenzyl)-8-(hydroxy(3-(trifluoromethoxy)phenyl)methyl)-l-(3-hydroxypropyl)-3-methyl- lH-purine-2,6(3H,7H)-dione (10 mg, 22.9%) as white solid. }H-NMR (CDC13) δ 7.36-7.3 l(t, 1H), 7.23-7.14(m, 5H), 6.94-6.92(d, 2H), 5.87-5.85(d, 1H), 5.60-5.56(d, 1H), 5.39-5.35(d, 1H), 4.20-4.17(t, 2H), 3.65-3.63(s, 4H), 3.54-3.51(m, 2H), 3.31(s, 1H), 1.91-1.88(t, 2H). LCMS retention time 2.605 min; LCMS MH+ 539.
Example 93 7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl-8-(3-(3
(trifluoromethoxy)phenoxy)propyl)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000546_0002
Step 1 7-(4-chlorobenzyl)-l-(4-methoxybenzyl)-3-methyl-8-(3-(3- (trifluoromethoxy)phenoxy)propyl)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000546_0003
To a solution of l -(4-rnethoxybenzyl)-3-methyl-8-(3-(3-(trifluorornethoxy)phenoxy)propyl)-l H- purine-2,6(3H,7H)-dione (130 mg, 0.261 mmol, intermediate 60) in DMF (3 mL) was added 1 - (bromomethyl)-4-chlorobenzene (70 mg, 0.345 mmol), followed by potassium carbonate (107 mg, 0.775 mmol) and TBAI (5 mg, 0.014 mmol). The mixture was stirred at 60 °C overnight. The reaction was cooled and partitioned between ethyl acetate and water. The combined organic phase was washed with brine, dried over sodium sulfate, and concentrated to give the crude product 7-(4-chlorobenzyl)-l -(4-methoxybenzyl)-3-methyl-8-(3-(3-
(trifluoromethoxy)phenoxy)propyl)-lH-purine-2,6(3H,7H)-dione (150 mg, 92.5% yield) as a yellow oil. LCMS retention time 2.075 min; LCMS MH+ 629.
Step 2 7-(4-chlorobenzyl)-3-methyl-8-(3-(3-(trifluoromethoxy)phenoxy)propyl)-l H- purine-2,6(3 -dione
Figure imgf000547_0001
To a solution of 7-(4-chlorobenzyl)-l-(4-methoxybenzyl)-3-methyl-8-(3-(3- (trifluoromethoxy)phenoxy)propyl)-lH-purine-2,6(3H,7H)-dione (105 mg, 0.167 mmol) in DCM (4 mL) was added TFA (1 mL) dropwise followed by trifluoromethanesulfonic acid (0.25 mL) at 0 °C. The reaction was stirred at 0 °C for 10 min, then at room temperature for 16 h. The reaction was quenched with saturated sodium bicarbonate and extracted with DCM. The organic phase was washed with water, dried over sodium sulfate, and concentrated to give 7-(4-chlorobenzyl)- 3-methyl-8-(3-(3-(trifluoromethoxy)phenoxy)propyl)-lH-purine-2,6(3H,7H)-dione (66 mg, 77.6% yield) as a yellow oil. LCMS retention time 2.868 min; LCMS MH+ 509.
Step 3 7-(4-chlorobenzyl)-3-methyl-l-(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-8-(3-(3- (trifluoromethoxy)phenoxy)propyl)-lH-purine-2,6(3H,7H)-dione
Figure imgf000547_0002
The title compound was prepared using the method of intermediate 14 and purified by silica gel chromatography eluting with petroleum/ ethyl acetate (10: 1 to 2: 1) to give 7-(4-chlorobenzyl)-3- methyl-l -(3-(tetrahydro-2H-pyran-2-yloxy)propyl)-8-(3-(3-(trifluoromethoxy)phenoxy)propyl)- lH-purine-2,6(3H,7H)-dione (39 mg, 38.5% yield)as yellow oil. LCMS retention time 2.048min; LCMS MH+-THP 567.
Step 4 7-(4-chlorobenzyl)-l -(3-hydroxypropyl)-3-methyl-8-(3-(3- (trifluorom
Figure imgf000548_0001
The title compound was prepared as example 37, step 2 and purified via preparative HPLC to give 7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl-8-(3-(3-
(trifluoromethoxy)phenoxy)propyl)-lH-purine-2,6(3H,7H)-dione (17 mg, 38.3% yield)as white solid. 'H-NMR (DMSO-i tf) 5 7.41 -7.36(m, 3H), 7.23-7.21 (d, 2H), 6.93-6.90(dd, 2H), 6.84(s, 1H), 5.56(s, 2H), 4.48-4.46(t, 1H), 4.05-4.02(t, 2H), 3.93-3.89(t, 2H), 3.45-3.33(m, 5H), 2.89- 2.85(t, 2H), 2.09-2.06(t, 2H), 1.70-1.66(t, 2H). LCMS retention time 2.925 min; LCMS MH+ 567.
Example 94 7-(4-chlorobenzyl)-8-ethyl-l-(3-hydroxypropyl)-3-methyl-lH-purine-2,6(3H,7H)- dione
Figure imgf000548_0002
The title compound was prepared using the method of intermediate 48 and example 91 and using propionic acid. White solid, 15 mg, 22.8% yield: 'H- MR (CD3OD) δ 7.35-7.33(d, 2Η), 7.20- 7.18(d, 2Η), 5.58(s, 2Η), 4.09-4.06(t, 2Η), 3.59-3.55(m, 5Η), 2.78-2.73(q, 2Η), 1.88-1.81(m, 2Η), 1.23- 1.19(t, 3Η). LCMS retention time 2.207 min; LCMS MH+ 377. Example 95 7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl-8-propyl-l H-purine- 2,6(3H,7H)-dione
Figure imgf000549_0001
The title compound was prepared using the methods of intermediate 48 and example 91 and using butyric acid. White solid, 10 mg, 30.4% yield: 'H-NMR (CD3OD) δ 7.33-7.3 l(d, 2H), 7.20-7.17(d, 2H), 5.58(s, 2H), 4.07-4.04(t, 2H), 3.59-3.56(t, 2H), 3.53(s, 3H), 2.72-2.69(t, 2H), 1 .87-1 .80(m, 2H), 1.70-1.61(m, 2H), 0.93-0.90(t, 3H). LCMS retention time 2.379 min; LCMS MH+ 391. Example 96 7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl-8-(2-(5-methylthiazol-2- yl)ethyl)-3,4,5,7-tetrahydro-lH-purine-2,6-dione
Figure imgf000549_0002
The title compound was prepared using the methods of intermediate 48 and example 91 and using intermediate 49. White solid, 15 mg, 45.4% yield: JH-NMR (CD3OD) δ= 7.32(d, 2H), 7.28(d, 1H), 7.18(d, 1H), 5.58(s, 2H), 4.10(t, 2H), 3.60(t, 2H), 3.57(s, 3H), 3.41(t, 2H),3.20(t, 2H), 2.40(d, 3H), 1 .85-1.89(m, 2H). LCMS retention time 2.334 min; LCMS MH+ 474.
Example 97 7-(4-chlorobenzyl)-l-(3-hydroxybutyl)-3-methyl-8-(6-methylpyridin-3-yloxy)-lH- purine-2,6(3H,7H)-dione
Figure imgf000549_0003
Step 1 8-bromo-7-(4-chlorobenzyl)-l -(3-hydroxybutyl)-3-methyl-lH-purine-2,6(3H,7H)-dione
Figure imgf000550_0001
The title compound was prepared using the method of example 7k from intermediates 8 and 33. White solid, 275 mg, 57.5% yield. LCMS retention time 1.465 min; LCMS MH+ 441 Step 2 7-(4-chlorobenzyl)-l -(3-hydroxybutyl)-3-methyl-8-(6-methylpyridin-3-yloxy)-lH- purine-2,6(3H,7H)
Figure imgf000550_0002
The title compound was prepared using the method of example 86. White solid, 35mg, 43.9% yield: 'H-NMR (DMSO-c¼) δ 8.46-8.45(d, 1 H), 7.78-7.75(dd, lH),7.47-7.45(d, 2H), 7.42- 7.37(m, 3H), 5.52(s, 2H), 4.17-4.05(m, 2H), 3.82-3.77(m, 1H), 3.40(s, 3H), 2.58(s, 3H), 1.79- 1 .73(m, 2H), 1.23-1.21(d, 3H). LCMS retention time 2.390 min; LCMS MH+ 470
Example 98 l-(3-hydroxybutyl)-3-methyl-7-((5-methylpyridin-2-yl)methyl)-8-(3- (trifluoromethoxy)benzyl)-lH-purine-2,6(3H,7H)-dione
Figure imgf000550_0003
To a solution of 3-methyl-7-((5-methylpyridin-2-yl)methyl)-8-(3-(trifluoromethoxy)benzyl)-lH- purine-2,6 (3H,7H)-dione (30m g, 0.067 mmol, intermediate 51) in DMF (3 mL) was added 3- hydroxybutyl-4-methylbenzenesulfonate (24.68 mg, 0.10 mmol), followed by potassium carbonate (27.93 mg, 0.20 mmol). The reaction was stirred at 70 °C for 4 h. The mixture was partitioned between ethyl acetate and water. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give crude product, which was purified by preparative HPLC to give l -(3-hydroxybutyl)-3-methyl-7-((5- mefhylpyridin-2-yl)methyl)-8-(3- (trifluoromethoxy)ben2yl)-lH-purine-2,6(3H,7H)-dione (9 mg, 25.8%) as white solid. 'H-NMR (CD3OD) δ 8.216(s,lH), 7.491 -7.467(d,l H), 7.312-7.272(m,l H), 7.154-6.995(m,4H),
5.688(s,2H), 4.305(s,2H), 4.143-4.01 l(m,2H), 3.772-3.440(m,l H), 3.585(s,3H) , 2.283(s,3H), 1.761 -1.703(m,2H), 1 .194-1.179(d,3H). LCMS retention time 2.569 min; LCMS MH+ 518.
Example 99 l -(2-(l-hydroxycyclopropyl)ethyl)-3-methyl-7-((5-methylpyridin-2-yl)methyl)-8- (3-(trifluoromethoxy)benzyl)- -purine-2,6(3H,7H)-dione
Figure imgf000551_0001
The title compound was prepared using the method of example 98 and using l-(2- bromoethyl)cyclopropanol, intermediate 46). White solid, 8.7 mg, 24.4%: *H-NMR (CD3OD) δ 8.220(s,lH), 7.487-7.463(d,lH), 7.319-7.279(m,lH), 7.169-6.984(m,4H), 5.690(s,2H),
4.315(s,2H), 4.256 - 4.221 (m, 2H), 3.589(s, 3H), 2.282(s,3H), 1.834-1.799(m,2H), 0.585- 0.569(m,2H), 0.386-0.356(m,2H). LCMS retention time 2.717 min; LCMS MH+ 530. Example 100 l -(3-hydroxybutyl)-3-methyl-7-((5-methylpyridin-2-yl)methyl)-8-(3- (trifl
Figure imgf000551_0002
To a solution of l -(3-hydroxybutyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH-pur ne- 2,6(3H,7H)- dione (50 mg, 0.121 mmol, intermediate 57) in DMF (3 mL) was added 2- (chloromethyl)-5-methylpyridine hydrochloride (30 mg, 0.182 mmol, intermediate 50), followed by potassium carbonate (50 mg, 0.363 mmol) and TBAI (10mg,0.027mmol). The reaction was stirred at 65 °C overnight. The reaction was cooled and partitioned between ethyl acetate and water. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give a crude product, which was purified by preparative HPLC to give l -(3- hydroxybutyl)-3-methyl-7-((5-methyl^
purine-2,6(3H,7H)-dione (20 mg, 31.9% yield) as white solid. 'H-NMR (CD3OD) δ
8.319(s,l H), 7.663-7.640(d,l H),7.553-7.510(t,l H), 7.348-7.316(m,3H), 7.233-7.208(m, lH), 5.609(s,2H), 4.1 12-4.004(m,2H), 3.763-3.731(m, l H), 3.457(s,3H), 2.339(s,3H), 1.758- 1 .698(m,2H), 1.197-1.182(d,2H). LCMS retention time 2.714 min; LCMS MH+ 520.
Example 101 l-(2-(l -hydroxycyclopropyl)ethyl)-3-methyl-7-((5-methylpyridin-2-yl)methyl)- 8-(3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7 -dione
Figure imgf000552_0001
To a solution of l -(2-(l-hydroxycyclopropyl)ethyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)- l H-purine- 2,6(3H,7H)-dione (50 mg, 0.1 17 mmol, intermediate 58) in DMF (3 mL) was added 2-(chloromethyl)-5-methylpyridine hydrochloride (24.9 mg, 0.176 mmol, intermediate 50), followed by potassium carbonate (48.6 mg, 0.351 mmol) and TBAI ( Omg, 0.027mmol). The reaction was stirred at 65 °C overnight. The reaction was cooled and partitioned between ethyl acetate and water. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give crude product, which was purified by preparative HPLC to give l-(2-(l - hydroxycyclopropyl)ethyl)-3-methyl-7- ((5-methylpyridin-2-yl)methyl)-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (21 mg, 33.7% yield) as white solid. 'H-NMR (CD3OD) 6 8.320-8.315(d, lH), 7.662-7.636(d,lH), 7.561-7.517(t,lH), 7.363- 7.315(m,3H), 7.240-7.215(m,lH), 5.608(s,2H), 4.247-4.212(t,2H), 3.462(s,3H), 2.338(s,3H), 1.829-1.793(t,2H), 0.582-0.554(m, 2H), 0.370-0.340(m, 2H). LCMS retention time 2.869 min; LCMS MH+ 532.
Example 102 l -(2-(l -hydroxycyclopropyl)ethyl)-3-methyl-7-(pyridin-2-ylmethyl)-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000553_0001
The title compound was prepared using the method of example 101 . White solid, 20 mg, 32.9% yield: -NMR (CD3OD) δ 8.488-8.475(d,lH), 7.83 l-7.627(t,lH), 7.562-7.521(t,lH), 7.446- 7.427(d, lH), 7.375-7.341 (m,3H), 7.240-7.220(d,lH), 5.656(s,2H), 4.242-4.207(t52H),
3.465(s,3H), 1.823-1.788(t,2H), 0.562-0.546(m,2H), 0.364-0.335(m,2H). LCMS retention time 2.759 min; LCMS MH+ 518.
Example 103 7-((5-chloropyridin-2-yl)methyI)-l-(2-(l-hydroxycyclopropyl)ethyl)-3-methyl-8- (3-
Figure imgf000553_0002
The title compound was prepared using the method of example 101. White solid, 30 mg, 49.4% yield: 'H-NMR (CD3OD) δ 8.469-8.464(d,lH), 7.868-7.841(d,lH), 7.557-7.535(t,lH), 7.484- 7.463(d,lH), 7.385-7.370(m,2H), 7.252-7.231 (d,lH), 5.638(s,2H), 4.237-4.201 (t,2H),
3.460(s,3H), 1.820-1.784(t,2H), 0.563-0.548(m,2H), 0.366-0.337(m,2H). LCMS retention time 3.058 min; LCMS MH+ 552.
Example 104 7-(4-chlorobenzyl)-l-(2-(l-hydroxycyclopropyl)ethyl)-3-methyl-8-((6-
Figure imgf000553_0003
To a solution of 8-bromo-7-(4-chlorobenzyl)-l -(2-(l -hydroxycyclopropyl)ethyl)-3-methyl-lH- purine- 2,6(3H,7H)-dione (128 mg, 0.282 mmol, intermediate 56) in DMF (5 mL) was added 6- methylpyridin-3-ol (37 mg, 0.338 mmol), followed by potassium carbonate (58 mg, 0.423 mmol). The reaction was stirred at 80 °C for 2 h. The reaction was partitioned between ethyl acetate and water. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give a crude product, which was purified by preparative HPLC to give 7-(4- chloi benzyl)-l -(2-(l -hydroxycyclopropyl)ethyl)-3-methyl-8-((6-methylpyridin- 3-yl)oxy)-lH- purine-2,6(3H,7H)-dione (1 10 mg, 80.9% yield) as white solid. Z-NMR (CD3OD) δ 8.457- 8.451(d,lH), 7.776-7.748(d,lH), 7.470-7.374(m,5H), 5.519(s,2H), 4.291 -4.256(t,2H),
3.41 l (s,3H), 2.584 (s,3H), 1.867-1 .832(t,2H), 0.618-0.610(m, 2H), 0.403-0.387(m, 2H). LCMS retention time 2.619 min; LCMS MH+ 482.
Example 105 7-(4-chlorobenzyl)-3 -methyl- 1 -(4,4,4-trifluorobutyl)-8-(3 - (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000554_0001
To a solution of 7-(4-chlorobenzyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine- 2,6(3H,7H)- dione (80 mg, 0.171 mmol, intermediate 9) in DMF (2 mL) was added 4,4,4- trifluorobutyl methanesulfonate (50 mg, 0.239 mmol, intermediate 52), followed by potassium carbonate (71 mg, 0.51 8 mmol). The reaction was stirred at 50 °C overnight. The mixture was cooled and partitioned between ethyl acetate and water. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give a crude product, which was purified by preparative HPLC to give 1 -(3 -hydroxybutyl)-3 -methyl- 7-((5-methylpyridin-2- yl)methyl)-8-(3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (20 mg, 31 .9% yield) as white solid. 'H-NMR (CD3OD) δ 7.571-7.529(t,lH), 7.458-7.43 l (m,2H), 7.377- 7.255(m,4H), 7.237-7.23 l (m,lH), 5.496(s,2H), 4.098-4.063(t,2H), 3.423(s,3H), 2.288- 2.219(m,2H), 1.958-1.901 (m,2H). LCMS retention time 3.653 min; LCMS MH+ 577.
Example 106 7-(4-chlorobenzyl)-l -(3-hydroxybutyl)-3-methyl-8-(propylsulfonyl)-lH-purine- 2,6(3H,7H)-dione
Figure imgf000555_0001
Step 1 7-(4-chlorobenzyl)-l -(3-hydroxybutyl)-3-methyl-8-(pi pylthio)-lH-purine-2,6(3H,7H)- dione
Figure imgf000555_0002
To a solution of 8-bromo-7-(4-chlorobenzyl)-l-(3-hydroxybutyl)-3-methyl- lH-purine- 2,6(3H,7H)-dione (100 mg, 0.23 mmol, example 97, step 1) in DMF (5 mL) was added propane^ 1 -thiol (20.7 mg, 0.27 mmol) followed by potassium carbonate (93.9 mg, 0.68 mmol). The reaction was stirred at 60 °C for 3 h. The reaction was partitioned between ethyl acetate and water. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give crude product (80 mg, 80.9% yield) as yellow oil. LCMS MH+ 437.
Step 2 7-(4-chlorobenzyl)-l -(3-hydroxybutyl)-3-methyl-8-(propylsulfonyl)-lH-purine- 2,6(3H,7H)-dione
Figure imgf000555_0003
To a solution of 7-(4-chlorobenzyl)-l -(3-hydroxybutyl)-3-methyl-8-(propylthio)-lH-purine- 2,6(3H,7H)-dione (80 mg, 0.18 mmol) in CHC13 (10 mL) was added MCPBA (126 mg, 0.73 mmol). The reaction was stirred at room temperature overnight. The reaction was quenched with aqueous Na2S203 and NaHC03 and extracted with DCM. The phases were separated. The organic phase was washed with aqueous NaHC03 and brine; then it was dried over sodium sulfate, filtered and concentrated to give a crude product, which was purified by preparative HPLC to give 7-(4-chlorobenzyl)-l-(3-hydroxybutyl)-3-methyl-8-(propylsulfonyl)-lH-purine- 2,6 (3H,7H)-dione (30 mg, 34.9%) as white solid. 'H-NMR (DMSO-fifc) 5 7.429-7.408(d,2H), 7.278-7.257(d,2H), 5.870(s,2H), 4.51 l-4.500(d,lH), 4.044-3.973(m, lH), 3.874-3.805(m,lH), 3.675-3.619(m,lH), 3.499-3.459(m,2H), 3.446(s,3H), 1.691 -1.540(m,4H), 1 .081 -1.065(m,3H) , 0.934-0.897(m,3H). LCMS retention time 2.599 min; LCMS MH+ 469.
Example 107 7-(4-chlorobenzyl)-l -(2-(l -hydroxycyclopropyl)ethyl)-3-methyl-8- (propylsulfonyl)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000556_0001
To a solution of 8-bromo-7-(4-chlorobenzyl)-3 -methyl- lH-purine-2,6(3H,7H)-di one (100 mg, 0.27 mmol, intermediate 8) in DMF (5 mL) was added propane- 1 -thiol (24.7 mg, 0.32 mmol) followed by potassium carbonate (112.2 mg, 0.81 mmol). The reaction was stirred at 60 °C for 4 h. The reaction was cooled and partitioned between ethyl acetate and water. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give a crude white solid product (124 mg, 99.9% yield). LCMS MH+ 365.
Step 2 7-(4-chlorobenzyl)-l -(2-(l -hydroxycyclopropyl)ethyl)-3-methyl-8-(propylthio)- lH- purine-2,6(3H,7H)-dione 2014/027920
Figure imgf000557_0001
To a solution of 7-(4-chlorobenzyl)-3-methyl-8-(propylthio)- lH-purine-2,6(3H,7H)-dione (124 mg, 0.34 mmol) in DMF (7 mL) was added l-(2-bromoethyl)cyclopropanol (67.3 mg, 0.41 mmol, intermediate 46), followed by potassium carbonate (141 mg, 1.02 mmol). The reaction was stirred at 70 °C for 5 h. The reaction was cooled and partitioned between ethyl acetate and water. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give a crude yellow oil product (100 mg, 66.9% yield). LCMS MH+ 449.
Step 3 7-(4-chlorobenzyl)- 1 -(2-( 1 -hydroxycyclopropyl)ethyl)-3-methyl-8-(propylsulfonyl)- 1 H- purine-2,6(3H,7H)-dione
Figure imgf000557_0002
To a solution of 7-(4-chlorobenzyl)-l-(2-(l-hydroxycyclopropyl)ethyl)-3-methyl-8-(propylthio)- lH-purine -2,6(3H,7H)-dione (100 mg, 0.22 mmol) in CHC13 (7 mL) was added MCPBA (153 mg, 0.89 mmol) in portions. The mixture was stirred at room temperature for 3 h. The reaction was quenched with aqueous Na2S203 and extracted with DCM. The phases were separated. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give a crude product, which was purified by preparative HPLC to give 7-(4-chlorobenzyl)-l -(2- (l-hydroxycyclopropyl)ethyl)-3-methyl-8-(propylsulfonyl)-lH-purine-2,6(3H,7H)-dione (20 mg, 18.7%) as white solid. JH-NMR (CD3OD) δ 7.392-7.336(m,4H), 5.982(s,2H), 4.285- 4.248(t,2H), 3.567(s,3H), 3.445-3.406(m,2H), 1.855-1.756(m,4H) , L042-1 .005(m,3H) , 0.633- 0.604(m,2H), 0.433-0.403(m,2H). LCMS retention time 2.734 min; LCMS MH+ 481.
Example 108 8-(3-chloro-5-(trifluoromethoxy)phenoxy)-7-(4-chlorobenzyl)-l-(3- hydroxypropyl)-3-methyl-lH-purine-2,6(3H,7H)-dione
Figure imgf000558_0001
To a solution of the 8-bi mo-7-(4-chloiObenzyl)-l-(3-hydroxypropyl)-3-methyl-lH-purine- 2,6(3H,7H)- dione (50 mg, 0.1 16 mmol, intermediate 63) in DMF (2 mL) was added 3-chloro-5- (trifluoromethoxy)phenol (29.82mg, 0.140 mmol) followed by potassium carbonate (24.24mg, 0.175mmol). The reaction was stirred at 85 °C for 2 h. The reaction was cooled and partitioned between ethyl acetate and water. The organic phase was washed with brine, dried over sodium sulfate, filtered and concentrated to give crude product, which was purified by preparative HPLC to give 8-(3-chloro-5-(trifluoromethoxy)phenoxy)-7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-3- methyl-lH-purine-2,6(3H,7H)-dione (35mg, 53.5% yield) as white solid. 'H-NMR (CD3OD) δ 7.458-7.435(m,3H), 7.381 -7.365(m,3H), 7.361 (s,lH), 5.507(s,2H), 4.138-4.102(t,2H), 3.640- 3.608(t,2H), 3.448(s,3H), 1 .905-1.871(m,2H). LCMS retention time 3.325 min; LCMS MH+ 559.
Example 109 7-(4-chlorobenzyl)-8-(2-fluoro-5-(trifluoromethoxy)phenoxy)-l -(3- hydroxypropyl)-3 -methyl- lH-purine-2,6(3H,7H)-dione
Figure imgf000558_0002
The title compound was prepared using the method of example 108. White solid, 30 mg, 47.27 yield: 'H-NMR (CD3OD) δ 7.588-7.566(m,lH), 7.491-7.425(m,3H), 7.392-7.371 (d,2H), 7.347-7.310(m,lH), 5.520(s,2H), 4.127-4.09 l(t,2H), 3.633-3.602(t,2H), 3.390(s,3H), 1.914- 1.847(m,2H). LCMS retention time 3.099 min; LCMS MH+ 543.
Example 110 l-(2-hydroxyethyl)-3-methyl-7-((6-methylpyridin-3-yl)methyl)-8-(2-(3- (trifluoromethoxy)phenoxy)ethoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000559_0001
The title compound was prepared using the method of example 42 and purified by preparative HPLC to give l -(2-hydroxyethyl)-3,7-dimethyl-8-(2-(3-(trifluoromethoxy)phenoxy)ethoxy)-l H- purine-2,6(3H,7H)-dione (13.1 mg, 29.9% yield) as white solid. 'H-NMR (DMSO-c¾ δ 8.45 (s, 2H), 7.57-7.59 (d, IH), 7.41-7.45(t, I H), 7.09-7.1 l (d, IH), 6.97-7.04(m, 3H), 5.20 (s, 2H), 4.79- 4.81(m, 4H), 4.73-4.75 (t, IH), 4.41-4.43(t, 2H), 3.92-3.96 (t, 2H), 3.49-3.51 (m, 2H), 3.47 (s, 3H), 2.39 (s, 3H). LCMS retention time 2.003 min; LCMS MH+ 536.
Example 111 7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl-8-(3- (trifluoromethoxy)phenyl)- lH-purine-2,6(3H,7H)-dione
Figure imgf000559_0002
To a solution of 8-bromo-7-(4-chlorobenzyl)-3-methyl-l -(3-(tetrahydro-2H-pyran-2- yloxy)propyl)-lH-purine-2,6(3H,7H)-dione (100 mg, 0.195 mmol, intermediate 14) in toluene and ethanol (6 mL/ 2mL) was added 3-(trifluoromethoxy)phenylboronic acid (60.2 mg, 0.293 mmol) followed by 1 mM aqueous sodium carbonate (1 mL). The mixture was degassed and refilled with a nitrogen atmosphere 3 times. Tetrakis(triphenylphosphine)palladium (12 mg, 0.01 mmol) was added and the reactin was stirred at 100 °C for 16 h. The reaction was cooled and partitioned between ethyl acetate and brine The organic phase was dried and concentrated to give a crude product (1 10 mg). This crude product was dissolved in 1 mM ethanolic HC1 (2 mL) and the mixture was stirred at room temperature for 30 min. The mixture was concentrated and purified by preparative HPLC to give 7-(4-chIorobenzyl)- l-(3-hydroxypropyl)-3-methyl-8-(3- (trifluoromethoxy)phenyl)-lH-purine-2,6(3H,7H)-dione (49 mg, 49.4% yield) as a white solid. 'H-NMR (CD3OD) δ 7.62-7.64(t, 1Η), 7.55(s, 1Η), 7.51 -7.53(m, 1Η), 7.29-7.31 (d, 2H)„7.22- 7.24 (d, 2H), 5.71(s, 2H), 4.1 1 -4.14(t, 2H), 3.63 (s, 3H), 3.61-3.63 (t, 2H), 1.87-1.90 (m, 2H). LCMS retention time 2.746 min; LCMS MH+ 509. Example 112 7-(4-chlorobenzyl)-8-(2-fluoro-5-(trifluoromethoxy)benzyl)- l -(3-hydroxypropyl)- 3-methyl-lH-purine-2,6(3H,7H)-dione
Figure imgf000560_0001
Step 1 7-(4-chlorobenzyl)-8-(2-fluoro-5-(trifluoromethoxy)benzyl)-3-methyl- l-(3- (tetrahydro-2H-pyran-2-yloxy)propyl)-l H-purine-2,6(3H,7H)-dione (JF-000357-069)
Figure imgf000560_0002
The title compound was prepared from intermediate 39 using the method of intermediate 14 to give 7-(4-chlorobenzyl)-8-(2-fluoro-5-(trifluoromethoxy)benzyl)-3-methyl- 1 -(3-(tetrahydro-2H- pyran-2-yloxy)propyl)-l H-purine-2,6(3H,7H)-dione (43.1 mg, 85.1% yield) as a yellow solid. LCMS retention time 2.046, LCMS MH+-THP 541.
Step 2 7-(4-chlorobenzyl)-8-(2-fluoro-5-(trifluoromethoxy)benzyl)-l-(3-hydroxypropyl)- 3-methyl-lH-purine-2,6(3H,7H)-dione
Figure imgf000560_0003
The title compound was prepared using the hydrolysis method of example 1 1 1 to give 7-(4- chlorobenzyl)-8-(2-fluoro-5-(trifluoromethoxy)benzyl)-l -(3-hydroxypropyl)-3-methyl-lH- purine-2,6(3H,7H)-dione (17 mg, 38.8% yield) as a white solid. 7H-NMR (CD3OD) δ 7.22- 7.25(m, 2H), 7.08-7.17(m, 4H), 7.01 -7.02(d, 1H), 5.67(s, 2H), 4.22 (s, 2H), 4.08-4.1 1 (t, 2H), 3.59-3.62(t, 2H), 3.54(s, 3H), 1.85-1.89(s, 2H). LCMS retention time 2.862 min; LCMS MH+ 541.
The following examples 1 13a to 1 13c were prepared using the methods of intermediate 39 and example 1 12. Example 113a 8-(4-chloro-3-(trifluoromethoxy)benzyl)-7-(4-chlorobenzyl)-l-(3- hydroxypropyl)-3-methyl-l H-purine- -dione
Figure imgf000561_0001
White solid, 10.7 mg, 35.2% yield: H-NMR (CD3OD) δ 7.35-7.37(d, 1H), 7.19-7.23(m, 3H), 7.02-7.04(m, 3H), 5.65(s, 2H), 4.24(s, 2H), 4.09-4.12(t, 2H), 3.58-3.62(1, 2H), 3.59(s, 3H), 1.86- 1.89(m, 2H). LCMS retention time 3.033 min; LCMS MH+ 557.
Example 113b 8-(3-chloro-5-(trifluoromethoxy)benzyl)-7-(4-chlorobenzyl)-l -(3- hydroxypropyl)-3-methyl-lH-purine-2,6(3H,7H)-dione
Figure imgf000561_0002
White solid, 24.7 mg, 35.5% yield: 7H-NMR (CD3OD) 8 7.19-7.21(m, 2H), 7.12"(s, 1H), 6.97- 7.04(m, 4H), 5.67(s, 2H), 4.24 (s, 2H), 4.09-4.13(t, 2H), 3.60-3.63(t, 2H), 3.59(s, 3H), 1.86- 1.90(m, 2H). LCMS retention time 3.044 min; LCMS MH+ 557. Example 113c7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-8-(2-methoxy-5- (trifluoromethoxy)benzyl)-3-methyl- -purine-2,6(3H,7H)-dione
Figure imgf000561_0003
White solid, 36.5 mg, 60.1% yield: 7H-NMR (CD3OD) δ 7.22-7.24 (m, 2H), 7.10-7.13 (d, 1H), 7.04-7.06 (d, 2H), 6.90-6.98 (m, 2H), 5.64 (s, 2H)3 4.15 (s, 2H), 4.07-4.1 1 (t, 2H), 3.78 (s, 3H), 3.58-3.61 (t, 2H), 3.55 (s, 3H), 1.85-1.88 (m, 2H). LCMS retention time 2.914 min; LCMS MH+ 553.
Example 114 1 -(3-hydroxybutyl)-3-methyl-7-(pyridin-2-ylmethyl)-8-(3- (trifluoromethoxy)benzyl)-lH-purine-2,6(3H,7H)-dione
Figure imgf000562_0001
Step 1 1 -(4-methoxybenzyl)-3 -methyl-8-(3 -(trifluoromethoxy)benzyl)- 1 H-purine-
2,6(3H,7H)-dione
Figure imgf000562_0002
The title compound was prepared using the method of intermediate 39 and purified by trituration with ethanol to give l -(4-methoxybenzyl)-3-methyl-8-(3-(trifluoromethoxy)benzyl)-lH-purine- 2,6(3H,7H)-dione (136 mg, 81.6% yield) as white solid. LCMS retention time 1.638 min; LCMS MH+ 461. Step 2 l-(4-methoxybenzyl)-3-methyl-7-(pyridin-2-ylmethyl)-8-(3- (trifluoromethoxy)benzyl)-l H-purine-2,6(3H,7H)-dione
Figure imgf000562_0003
To a solution of l -(4-methoxybenzyl)-3-methyl-8-(3-(trifluoromethoxy)benzyl)-lH-purine- 2,6(3H,7H)-dione (136 mg, 0.295 mmol) in DMF (2 mL) was added 2-(bromomethyl)pyridine hydrobromide (1 12 mg, 0.442 mmol) followed by potassium carbonate (102 mg, 0.738 mmol). The reaction was stirred at 65 °C for 2 h. The mixture partitioned between ethyl acetate and brine. The organic layer was dried and concentrated to give a crude product, which was purified by silica gel chromatography eluting with DCM /methanol (45: 1) to give 1 -(4-methoxybenzyl)- 3-methyl-7-(pyridin-2-ylmethyl)-8-(3-(trifluoromethoxy)benzyl)-lH-purine-2,6(3H,7H)-dione (96 mg, 59% yield) as a white solid. LCMS retention time 1.791 min; LCMS MH+ 552. 3-methyl-7-(pyridin-2-ylmethyl)-8-(3-(trifluoromethoxy)benzyl)-l H-purine- -dione
Figure imgf000563_0001
The title compound was prepared using the method of intermediate 39, step 3 to give 3 -methyl (pyridin-2-ylmethyl)-8-(3-(trifluoromethoxy)benzyl)-lH-purine-2,6(3H,7H)-dione (61 mg, 81.2% yield) as a light yellow solid. LCMS retention time 1.276 min; LCMS MH+ 432.
Step 4 l-(3-hydroxybutyl)-3-methyl-7-(pyridin-2-ylmethyl)-8-(3- (trifluoromethoxy)benzyl)-lH-purine-2,6(3H,7H)-dione
Figure imgf000563_0002
To a solution of 3-methyl-7-(pyridin-2-ylmethyl)-8-(3-(trifiuoromethoxy)benzyl)-lH-purine- 2,6(3H,7H)-dione (30 mg, 0.07 mmol) in DMF (1 mL) was added 3-hydroxybutyl 4- methylbenzenesulfonate (25.6 mg, 0.105 mmol, intermediate 33) followed by potassium carbonate (14.5 mg, 0.105 mmol) and a catalytic amount of TBAI. The reaction was stirred at 60 °C for 2 h. The reactin was partitioned between ethyl acetate and brine. The organic phase was dried and concentrated to give crude product, which was purified by preparative HPLC to give 1 - (3-hydroxybutyl)-3-methyl-7-(pyridin-2-ylmethyl)-8-(3-(trifluoromethoxy)benzyl)-lH-purine-
2,6(3H,7H)-dione (1 1.2 mg, 32% yield) as white solid. 'H-NMR (CD3OD) δ 8.38-8.39 (d, 1 H), 7.65-7.69 (t, 1H), 7.23-7.31 (m, 2H), 7.07-7.17 (m, 2H), 5.72 (s, 2H), 4.32 (s, 2H), 4.01-4.1 1 (m, 2H), 3.75-3.77 (m, 1H), 3.58 (s, 3H), 1.71-1.75 (m, 2H), 1.17-1.19 (d, 3H). LCMS retention time 2.480 min; LCMS MH+ 504.
Example 115 l-(3-hydroxybutyl)-3-methyl-7-(pyridin-2-ylmethyl)-8-(3- (trifluoromethoxy)benzyl)-lH-purine-2,6(3H,7H)-dione
Figure imgf000564_0001
The title compound was prepared using the method of example 1 14, step 4 and purified by preparative HPLC to give l -(2-(l-hydroxycyclopropyl)ethyl)-3-methyl-7-(pyridin-2-ylmethyl)- 8-(3-(trifluoromethoxy)benzyl)-lH-purine-2,6(3H,7H)-dione (14.1 mg, 33.8%) as white solid. H-NMR (CD3OD) S 8.39-8.40 (d, 1Η), 7.65-7.69 (t, 1Η), 7.28-7.32 (t, 1Η), 7.23-7,25 (m, 1Η), 7.16-7.19 (d, 1Η), 7.08-7.12 (m, 3Η), 5.72 (s, 2Η), 4.33 (s, 2Η), 4.21-4.24 (t, 2Η), 3.59 (s, 3Η), 1.79-1.82 (t, 2Η), 0.55-0.58 (t, 2Η), 0.34-0.37 (t, 2Η). LCMS retention time 2.609 min; LCMS MH+ 516. Example 116 7-(4-chlorobenzyl)-l-((2-hydroxycyclopentyl)methyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000564_0002
The title compound was prepared from intermediates 9 and 72 using the method of example 82 and purified by preparative HPLC to give 7-(4-chlorobenzyl)- 1 -((2-hydroxycyclopentyl)methyl)- 3-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (20 mg, 33.1% yield) as white solid. 'H-NMR (DMSO-^) δ 7.62-7.58(t, 1Η), 7.48(s, 1Η), 7.44-7.42(m, 5Η), 7.33- 7.31 (dd, 1Η), 5.44(s, 2Η), 4.43-4.42(d, 1Η), 3.92-3.87(m, 1Η), 3.81-3.71 (m, 2Η), 3.29(s, 3Η), 2.21-2.16(m, 1Η), 1.85-1.19(m, 6Η). LCMS retention time 3.307 min; LCMS MH+ 565.
Example 117 7-(4-chlorobenzyl)-l-(3-hydroxycyclobutyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000564_0003
Step 1 l-(3-(benzyloxy)cyclobutyl)-7-(4-chlorobenzyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000565_0001
The title compound was prepared using the method of example 52, step 1 and purified by silica gel chromatography eluting with DCM/ methanol (60: 1) to give l -(3-(benzyloxy)cyclobutyl)-7- (4-chlorobenzyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-l H-purine-2,6(3H,7H)-dione (1 10 mg, 50.1 % yield) as white solid. LCMS retention time 2.378 min; LCMS MH+ 627.
Step 2 7-(4-chlorobenzyl)-l -(3-hydroxycyclobutyl)-3-methyl-8-(3- (trifluoromethox henox -lH- urine-2 6 3H 7H -dione
Figure imgf000565_0002
To a solution of l-(3-(benzyloxy)cyclobutyl)-7-(4-chlorobenzyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (1 10 mg, 0.175 mmol) in methanol was added 5% Pd/C (15 mg). The mixture was was hydrogenated at 45 psi for 30 min. The reaction was filtered and concentrated to give a crude product which was purified via preparative HPLC to give 7-(4-chlorobenzyl)-l -(3-hydroxycyclobutyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (11 mg, 17.7% yield) as white solid. 'H-NMR (DMSO-<¾ δ 7.53-7.57 (t, IH), 7.43-7.45 (d, 2H), 7.36-7.38 (d, 2H), 7.31 -7.33 (m, IH), 7.23-7.25 (d, IH), 5.79-5.84 (m, 0.65H), 5.50 (s, 2H), 4.83-4.85 (m, 0.35H), 4.64-4.66 (m, 0.65H), 4.00-4.02 (m, 0.35H), 3.40(s, 3H), 3.16-3.19 (m, 1.3H), 2.88-2.91 (m, 0.7H), 2.66-2.70 (m, 0.7H), 2.25-2.30 (m, 1.3H). LCMS retention time 3.131 min; LCMS MH+ 537.
Example 118 7-(4-chlorobenzyl)-l -(3 -hydroxy cyclobutyl)-3 -methyl- 8 -(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000566_0001
The title compound was prepared from the product of example 1 17 using the method of example 1 17, step 2, but with a longer hydrogenation time. The product was purified by preparative HPLC to give 7-benzyl-l -(3-hydroxycyclobutyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH- purine-2,6(3H,7H)-dione (46 mg, 52.3% yield) as white solid. ]H-NMR (DMSO-t¼) δ 7.54-7.58 (t, 1H), 7.42-7.44 (d, 2H), 7.23-7.37 (m, 5H), 7.22-7.24 (d, 1H), 5.80-5.84 (m, 0.6H), 5.50 (s, 2H), 4.77-4.82 (m, 0.4H), 4.62-4.66 (m, 0.6H), 4.00-4.04 (m, 0.4H), 3.39(s, 3H), 3.16-3.23 (m, 1.2H), 2.83-2.91 (m, 0.8H), 2.68-2.73 (m, 0.8H), 2.25-2.31 (m, 1 .2H). LCMS retention time 2.961 min; LCMS MH+ 503.
Example 119 7-(4-chlorobenzyl)-l-(3-hydroxycyclobutyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000566_0002
The title compound was prepared using the method of example 82 with intermediates 9 and 74 and purified via preparative HPLC to give 7-(4-chlorobenzyl)-l-(3-hydroxycyclopentyl)-3- methyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (29 mg, 24.6% yield) as white solid. 'H-NMR (DMSO-i¾ δ 7.53-7.57 (t, 1H), 7.43-7.45 (d, 2H), 7.32-7.38 (m, 6H), 7.24-7.26 (dd, 1H), 5.52-5.56 (m, 1H), 5.50 (s, 2H), 4.21-4.24 (m, 0.4H), 3.42(s, 3H), 2.33-2.38 (m, 1H), 2.22-2.27 (m, 1H), 2.13-2.16 (m, 1H), 1.89-1.99 (m, 2H), 1.77-1.80 (m, 1H). LCMS retention time 3.168LCMS MH+ 551.
Example 120 7-(4-chlorobenzyl)-l -(3-(hydroxymethyl)cyclobutyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000567_0001
Step 1 benzyl 3-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-
(triiluoromethoxy)phenoxy)-2, -tetrahydro-lH-purin-l -yl)cyclobutanecarboxylate
Figure imgf000567_0002
The title compound was prepared using the method of example 82 with intermediates 9 and 75 to give 3-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-(trifluoromethoxy)phenoxy)-2,3,6,7- tetrahydro-lH-purin-l -yl)cyclobutanecarboxylate (59 mg, 26.1% yield) as a yellow solid. LCMS retention time 1.981 min; LCMS MH+ 655. Step 2 7-(4-chlorobenzyl)-l-(3-(hydroxymethyl)cyclobutyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3 -dione
Figure imgf000567_0003
To a solution of 3-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-(trifluoromethoxy)phenoxy)- 2,3,6,7-tetrahydro-lH-purin-l-yl)cyclobutanecarboxylate (59 mg, 0.09 mmol) in THF (3 mL) was added LAH (6.8 mg, 0.18 mmol) in one portion and the mixture was stirred at room temperature for 2 h. The reaction was carefully quenched with ethyl acetate and filtered. The filtrate was concentrated to dryness to give a crude product which was purified via preparative HPLC to give 7-(4-chlorobenzyl)-l-(3-(hydroxymethyl)cyclobutyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (17 mg, 34.3% yield) as white solid. 'H-NMR (CDC13)5 7.40-7.48 (m, 3H), 7.33-7.35 (m, 2H), 7.21 -7.26 (m, 2H), 7.15-7.17 (d, 1H), 5.52-5.63 (m, 1H), 5.44 (s, 2H), 3.73-3.80 (m, 2H), 3.1 1-3.17 (q, 1H), 2.96-2.98 (m, 1H), 2.60- 2.64 (m, 1H), 2.34-2.38 (m, 1H), 2.08-2.14 (m, 1H). LCMS retention time 3.054; LCMS MH+ 551. Example 121 l-(3-hydroxybutyl)-3-methyl-7-(pyridin-2-ylmethyl)-8-(3- (trifluoromethoxy)phenoxy)-l H-purine-2,6(3H,7H)-dione
Figure imgf000568_0001
The title compound was prepared from intermediate 57 using the method of example 100 and purified by preparative HPLC to give l -(3-hydroxybutyl)-3-methyl-7-(pyridin-2-ylmethyl)-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (40 mg, 21.9% yield) as a white solid. 'H-NMR (CD3OD) δ 8.49-8.48(d, I H), 7.88-7.81 (t, 1H), 7.53-7.51 (t, I H), 7.45-7.43(d, 1H), 7.36-7.33(m, 3H), 7.23-7.19(d, IH), 5.66(s, 2H), 4.14-3.98(m, 2H), 3.78-3.71(m, IH), 3.46(s, 3H), 1.75-1.71(m, 2H), 1.19-1.18(d, 3H). LCMS retention time 2.575 min; LCMS MH+ 506.
Example 122 7-((5-chloropyridin-2-yl)methyl)-l-(3-hydroxybutyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000568_0002
The title compound was prepared from intermediate 57 using the method of example 100 and purified by preparative HPLC to give 7-((5-chloropyridin-2-yl)methyl)-l -(3-hydroxybutyl)-3- methyI-8-(3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (40 mg, 21.9% yield) as a white solid. V -NMR (CD3OD) δ 8.47-8.46(d, IH), 7.87-7.84(d, IH), 7.57-7.46(t, IH), 7.37- 7.36(d, IH), 7.25-7.22(m, 2H), 7.14-7.12(d, IH), 5.64(s, 2H), 4.13-3.97(m, 2H), 3.78-3.72(m, IH), 3.45(s, 3H), 1.75-1.69(m, 2H), 1.19-1.18(d, 3H). LCMS retention time 2.944 min; LCMS ΜΗ' 540.
Example 123 7-((5-fluoropyridin-2-yl)methyl)-l -(3-hydroxybutyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000568_0003
The title compound was prepared from intermediate 57 using the method of example 100 and purified by preparative HPLC to give 7-((5-fluoropyridin-2-yl)methyl)-l -(3-hydroxybutyl)-3- methyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (40 mg, 21.1% yield) as a white solid. 'H-NMR (CD3OD) δ 8.38-8.37(d, 1H), 7.66-7.51(m, 3H), 7.37-7.35(m, 2H), 7.24- 7.22(d, 1 H), 5.64(s, 2H), 4.12-3.97(m, 2H), 3.77-3.72(m, IH), 3.45(s, 3H), 1.75-1 .68(m, 2H), 1.19-1.18(d, 3H). LCMS retention time 2.733 min; LCMS MH+ 524.
Example 124 7-((5-fluoropyridin-2-yl)methyl)-l-(2-(l-hydroxycyclopropyl)ethyl)-3-methyl-8- (3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000569_0001
The title compound was prepared from intermediate 58 using the method of example 100 and purified by preparative HPLC to give 7-((5-fluoropyridin-2-yl)methyl)-l -(2-(l - hydroxycyclopropyl)ethyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)- dione(25 mg, 33.7% yield) as white solid. 'H-NMR (CD3OD)£ 8.49(d, IH), 7.5 l-7.65(m, 3H), 7.36-7.39(m, 2H), 7.24(dd, I H), 5.64(s, 2H), 4.22(t, 2H), 3.44(s, 3H), 1.81(t, 2H), 0.56(t, 2H), 0.35 (t, 2H). LCMS retention time 2.894 min; LCMS MH+ 536.
Example 125 7-(4-chlorobenzyl)-3-ethyl-l -(3-hydroxypropyl)-8-((5-methylpyridin-3-yl)oxy)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000569_0002
To a solution of 8-chloro-7-(4-chlorobenzyl)-3-ethyl-l -(3-hydroxypropyl)-lH-purine- 2,6(3H,7H)-dione (30 mg, 0.08 mmol, intermediate 78) in DMF (2 mL) was added 5- methylpyridin-3-ol (12.36 mg, 0.12 mmol), followed by potassium carbonate (15.66 mg, 0.11 mmol). The reaction was stirred at 80 °C for 3 h. The reaction was cooled and partitione between ethyl acetate and water. The organic phase was washed with brine, dried over sodium sulfate, and concentrated to give a crude product, which was purified by preparative HPLC to give 7-(4- chlorobenzyl)-3-ethyl- l-(3-hydroxypropyl)-8-((5-methylpyridin-3-yl)oxy)- l H-purine- 2,6(3H,7H)-dione (8 mg, 22.5 %) as white solid. 'H-NMR (CD3OD) δ 8.35-8.42(d, 2Η), 7.68(s, 1Η), 7.38-7.48(m, 4Η), 5.52(s, 2Η), 4.01 -4.14(m, 4Η), 3.61 -3.64(m, 2Η), 2.43(s, 3Η), 1.87- 1 .90(m, 2Η), 1.22-1.26(m, 3Η). LCMS retention time 2.51 1 min; LCMS MH+ 470.
Example 126 7-(4-chlorobenzyl)-3-ethyl-l -(3-hydroxypropyl)-8-(propylsulfonyl)-lH-purine- 2,6(3H,7H)-dione
Figure imgf000570_0001
The title compound was prepared using the method of example 106 and purified by preparative HPLC to give 7-(4-chlorobenzyl)-3-ethyl-l-(3-hydroxypropyl)-8-(propylsulfonyl)-lH-purine- 2,6(3H,7H)-dione (7 mg, 16.3 %) as white solid. 'H-NMR (CD3OD) δ 7.35-7.41 (m, 4Η), 5.98 (s, 2Η), 4.10-4.18 (m, 4Η), 3.60-3.64(m, 2Η), 3.40-3.44 (m, 2Η), 1.77-1.89 (m, 4Η), 1.32- 1.36(m, 3Η), 1.00-1.04 (m, 3Η). LCMS retention time 2.521 min; LCMS MH+ 469.
Example 127 8-(4-chloro-3-(trifluoromethoxy)phenoxy)-7-(4-chlorobenzyl)-l -(3- hydroxypropyl)-3-methyl- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000570_0002
The title compound was prepared from intermediate 63 using the method of example 108 and purified by preparative HPLC to give 8-(4-chloro-3-(trifluoromethoxy)phenoxy)-7-(4- chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl-lH-purine-2,6(3H,7H)-dione (39.4 mg, 30.1% yield) as white solid. 'H-NMR (CD3OD) δ 7.66-7.68(d, 2Η), 7.58(s, 1Η), 7.44-7.46 (d, 2Η), 7.36-7.40(m, 2Η), 5.51 (s, 2Η), 4.10-4.13 (t, 2Η), 3.60-3.63(t, 2Η), 3.43(s, 3Η), 1.87-1.90(m, 2Η). LCMS retention time 3.268 min; LCMS MH+ 559. Example 128 7-(4-chlorobenzyl)-8-(4-fluoro-3-(trifluoromethoxy)phenoxy)- 1 -(3- hydroxypropyl)-3-methyl-lH-purine-2,6(3H,7H)-dione
Figure imgf000571_0001
The title compound was prepared from intermediate 63 using the method of example 108 and purified by preparative HPLC to give 7-(4-chlorobenzyl)-8-(4-fluoro-3- (trifluoromethoxy)phenoxy)-l-(3-hydroxypropyl)-3-methyl-lH-purine-2,6(3H,7H)-dione (52.9 mg, 55.6% yield) as white solid. 'H-NMR (CD3OD) 5=7.55-7.56(m, IH), 7.44-7.46(d, 2H), 7.39-7.42 (m, IH), 7.37-7.39(d, 2H), 5.50(s, 2H), 4.13-4.09 (t, 2H), 3.60-3.63(t, 2H), 3.42(s, 3H), 1.86-1.90(m, 2H). LCMS retention time 3.1 14 min; LCMS MH+ 543.
Example 129 7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-8-(2-methoxy-5- (trifluoromethoxy)phenoxy)-3-m -dione
Figure imgf000571_0002
The title compound was prepared from intermediate 63 using the method of example 108 and purified by preparative HPLC to give 7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-8-(2-methoxy-5- (trifluoromethoxy)phenoxy)-3-methyl-lH-purine-2,6(3H,7H)-dione (56.5 mg, 43.5% yield) as white solid. 'H-NMR (CD3OD) δ 7.50-7.52(d, 2H), 7.38-7.40(d, 2H), 7.37 (s, IH), 7.21 -7.26 (m, 2H), 5.49(s, 2H), 4.08-4.1 1 (t, 2H), 3.75(s, 3H), 3.59-3.62(t, 2H), 3.36(s, 3H), 1.85-1.89(m, 2H). LCMS retention time 3.066 min; LCMS MH+ 555.
Example 130 7-benzyl-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-l -(3,3,3-trifluoropropyl)- lH-purine-2,6(3H,7H)-dione
Figure imgf000571_0003
The title compound was prepared from intermediate 80 using the method example 100 and purified via preparative HPLC to give 7-benzyl-3-methyl-8-(3-(trifluoromethoxy)phenoxy)- l- (3,3,3-ΐπί1υοΓορΓοργ1)-1Η-ρυπηε-2,6(3Η,7Η)^ΐοη6 (35.6 mg, 59.1% yield) as white solid. XH- NMR (CD3OD) δ 7.52-7.56(t, 1 H), 7.44-7.46(d, 2H), 7.29-7.38 (m, 5H), 7.22-7.24 (d, 1 H), 5.52(s, 2H), 4.27-4.30 (t, 2H), 3.42(s, 3H), 2.54-2.60(m, 2H). LCMS retention time 3.382 min; LCMS MH+ 529.
Example 131 7-((5-chJoropyridin-2-yl)methyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-l- (3,3)3-trifluoropropyl)-l H-purine-2,6(3H,7H)-dione
Figure imgf000572_0001
The title compound was prepared using the method of example 100 and purified via preparative HPLC to give 7-benzyl-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-l-(3,3,3-trifluoropropyl)-lH- purine-2,6(3H,7H)-dione (18.7 mg, 29.1 % yield) as white solid. 'H-NMR (CD3OD) δ 8.52(s, 1H), 8.46(s, 1H), 7.84-7.87 (dd, 1 H), 7.53-7.57 (t, 1H), 7.47-7.49 (d, 1H), 7.36-7.37 (m, 1H), 7.22-7.24 (d, 1H), 5.63(s, 2H), 4.19-4.23 (t, 2H), 3.45(s, 3H), 2.50-2.55(m, 2H). LCMS retention time 3.369 min; LCMS MH+ 564.
Example 132 7-(4-fluorobenzyl)-8-(2-fluorophenoxy)-l-(3-hydroxypropyl)-3-methyl-lH- purine-2, -dione
Figure imgf000572_0002
To a solution of 8-bromo-7-(4-fluorobenzyl)-l-(3-hydroxypropyl)-3-methyl-lH-purine- 2,6(3H,7H)-dione (50 mg, 0.121 mmol, intermediate 62) in DMF (1 mL) was added 2- fluorophenol (16.4 mg, 0.146 mmol) followed by potassium carbonate (25.2 mg, 0.182 mmol). The reaction was stirred at 80 °C for 4 h. The mixture was partitioned between ethyl acetate and brine. The organic phase was washed with saturated aqueous ammonium chloride, dried and 4 027920
concentrated to give crude product, which was purified by preparative HPLC to give 7-(4- fluorobenzyl)-8-(2-fluorophenoxy)- 1 -(3-hydroxypropyl)-3-methyl- 1 H-purine-2,6(3H,7H)-dione (19.8 mg, 36.9% yield) as white solid.'H-NMR (CD3OD) δ 7.52-7.56 (m, 2Η), 7.43-7.45 (t, 1Η), 7.2-7.38 (m, 3Η), 7.08-7.13 (t, 1Η), 5.51 (s, 2Η), 4.09-4.13 (t, 2Η), 3.60-3.63 (t, 2Η), 3.37 (s, 3Η), 1 .86-1.90 (m, 2Η). LCMS retention time 2.604; LCMS ΜΙ- 443.
The compounds in Table 2 were prepared using the method of example 132.
Table 1
Figure imgf000573_0001
Figure imgf000574_0001
F F 1 H-purine-2,6(3H,7H)-dione
Figure imgf000575_0001
27920
Figure imgf000576_0001
7-(4-fluorobenzyl)-l-(3- 2.572 455 hydroxypropyl)-8-(4- methoxyphenoxy)-3-methyl-
Figure imgf000577_0001
1 H-purine-2,6(3H,7H)-dione
7-(4-fiuorobenzyl)-l -(3- 2.889 493 hydroxypropyl)-3 -methyl -8- (4-(trifluoromethyl)phenoxy)- lH-purine-2,6(3H,7H)-dione
7-(4-fluorobenzyl)-l -(3- 2.957 509 hydroxypropyl)-3-methyl-8-
(4-
(trifluoromethoxy)phenoxy)- 1 H-purine-2,6(3H,7H)-dione
4-((7-(4-fluorobenzyl)-l -(3- 2.431 450 hydroxypropyl)-3-methyl- 2,6-dioxo-2,3 ,6,7-tetrahydro-
Figure imgf000577_0002
lH-purin-8- yl)oxy)benzonitrile
8-(4-ethylphenoxy)-7-(4- 2.936 453 fluorobenzyl)- l-(3- ? N H hydroxypropyl)-3-methyl- lH-purine-2,6(3H,7H)-dione
7-(4-fluorobenzyl)-l-(3- 3.096 467 hydroxypropyl)-8-(4- isopropylphenoxy)-3-methyl- lH-purine-2,6(3H,7H)-dione
7-(4-chlorobenzyl)-8-(2- 3.692 475 chlorophenoxy)-l-(3- hydroxypropyl)-3-methyl- 1 H-purine-2,6(3H,7H)-dione 165 7-(4-chlorobenzyl)-l-(3- 2.871 455 hydroxypropyl)-3-methyl-8- (o-tolyloxy)- 1 H-purine- 2,6(3H,7H)-dione
166 7-(4-chlorobenzyl)-l-(3- 3.535 471 hydroxypropyl)-8-(2- methoxyphenoxy)-3-methyl- 1 H-purine-2,6(3H,7H)-dione
167 7-(4-chlorobenzyl)-l-(3- 3.695 509 hydroxypropyl)-3-methyl-8- (2-(trifluoromethyl)phenoxy)- 1 H-purine-2,6(3H,7H)-dione
68 7-(4-chlorobenzyl)- 1 -(3 - 3.003 525
1 rrQr° hydroxypropyl)-3-methyl-8-
(2-
(tri fl uoromethoxy)phenoxy) - 1 H-purine-2,6(3H,7H)-dione
69 2-((7-(4-chlorobenzyl)-l -(3- 2.601 466 hydroxypropyl)-3-methyl-
2,6-dioxo-2,3,6,7-tetrahydro- l H-purin-8- yl)oxy)benzonitrile
70 7-(4-chlorobenzyl)- 1 -(3 - 3.100 483 hydroxypropyl)-8-(2- isopropylphenoxy)-3 -methyl- 1 H-purine-2,6(3H,7H)-dione
71 7-(4-chlorobenzyl)-8-(2- 3.035 469 ethylphenoxy)-l -(3- hydroxypropyl)-3-methyl- lH-purine-2,6(3H,7H)-dione
Figure imgf000579_0001
1 H-purine-2,6(3H,7H)-dione
Figure imgf000580_0001
phenoxy- 1 H-purine- 27920
Figure imgf000581_0001
2-((7-((5-chloropyridin-2- 2.273 467
V0* yl)methyl)-l -(3-
1 NC hydroxypropyl)-3-methyl- 2,6-dioxo-2,3 ,6,7-tetrahydro- 11-1 -pur in- 8- yl)oxy)benzonitrile
7-((5-chloropyridin-2- 2.810 484 yl)methyl)-l -(3- hydroxypropyl)-8-(2- i sopropy lphenoxy ) -3 -methy 1- lH-purine-2,6(3H,7H)-dione
7-((5-chloropyridin-2- 2.682 470 yl)methyl)-8-(2- ethy lphenoxy)- 1 -(3 - hydroxypropyl)-3-methyl- 1 H-purine-2,6(3H,7H)-dione
7-((5 -chloropyridin-2- 2.760 510 yl)methyl)-8-(2,3-
A 1X}- t®l CI dichlorophenoxy)-l -(3- hydroxypropyl)-3 -methy 1 - lH-purme-2,6(3H,7H)-dione
7-((5-chloropyridin-2- 1.535 478 yl)methyl)-8-(3,4-
( F difluorophenoxy)- 1 -(3 - hydroxypropyl)-3-methyl- 1 H-purine-2,6(3H,7H)-dione
7-((5-chloropyridin-2- 1.507 478 yl)methyl)-8-(2,3- difluorophenoxy)-l-(3- hydroxypropyl)-3 -methyl- 1 H-purine-2,6(3H,7H)-dione 198 7-((5-chloropyridin-2- 1.560 478 yl)methyl)-8-(3,5-
1 V difluorophenoxy)-l -(3- hydroxypropyl)-3 -methyl- 1 H-purine-2,6(3H,7H)-dione
199 8-(3-chlorophenoxy)-7-((5- 2.648 476 chloropyridin-2-yl)methyl)- 1 - (3-hydroxypropyl)-3-methyl- lH-purine-2,6(3H,7H)-dione
200 7-((5 -chloropyridin-2- 2.476 460 yl)methyl)-8-(3- fluorophenoxy)- 1 -(3 -
.1
hydroxypropyl)-3 -methy 1- 1 H-purine-2,6(3H,7H)-dione
201 7-((5-chloropyridin-2- 2.584 456 yl)methyl)-l-(3- hydroxypropyl)-3-methyl-8- (m-tolyloxy)- 1 H-purine- 2,6(3H,7H)-dione
202 7-((5 -chloropyri din-2- 2.432 472 yl)methyl)-l -(3- hydroxypropyl)- 8 -(3 - methoxyphenoxy)-3-methyl- lH-purine-2,6(3H,7H)-dione
203 7-((5-chloropyridin-2- 2.735 510 yl)methyl)-l -(3-
1 hydroxypropylmethyl)phenox
y)-l H-purine-2,6(3H,7H)- dione 204 7-((5-chloropyridin-2- 1 .528 508 yl)methyl)-8-(3-
1 (difluoromethoxy)phenoxy)- 1 -(3-hydroxypropyl)-3- methyl- 1 H-purine- 2,6(3H,7H)-dione
205 3-((7-((5-chloropyridin-2- 2.296 467 yl)methyl)-l-(3-
1 hydroxypropyl)-3-methyl-
2,6-dioxo-2,3,6,7-tetrahydro- lH-purin-8- yl)oxy)benzonitrile
206 7-((5-chloropyridin-2- 2.748 470 yl)methyl)-8-(3- ethylphenoxy)- 1 -(3 - hy droxypropyl ) -3 -methyl -
1 H-purine-2,6(3 H,7H)-dione
207 7-((5 -chloropyridin-2- 1.753 484 yl)methyl)-l-(3- hydroxypropyl)-8-(3- isopropylphenoxy)-3-methyl- lH-purine-2,6(3H,7H)-dione
208 8-(4-chlorophenoxy)-7-((5- 2.676 476 chloropyridin-2-yl)methyl)- 1 -
1 (3-hydroxypropyl)-3-methyl- lH-purine-2,6(3H,7H)-dione
209 7-((5-chloropyridin-2- 2.477 460 yl)methyl)-8-(4-
1 fluorophenoxy)- 1 -(3 - hydroxypropyl)-3-methyl- lH-purine-2,6(3H,7H)-dione 7-((5-chloropyridin-2- 2.598 456 yl)methyl)-l -(3-
1 hydroxypropyl)-3-methyl-8- (p-tolyloxy)- 1 H-purine- 2,6(3H,7H)-dione
7-((5-chloropyridin-2- 2.433 472 yl)methyl)-l-(3- hydroxypropyl)-8-(4- methoxyphenoxy)-3 -methy l- 1 H-purine-2,6(3 H, 7H)-dione
7-((5-chloropyridin-2- 2.746 510 yl)methyl)-l -(3-
1 ' hydroxypropyl)-3-methyl-8- (4-(trifluoromethyl)phenoxy)- 1 H-purine-2,6(3H,7H)-dione
7-((5-chloropyridin-2- 2.802 526 yl)methyl)-l-(3- hydroxypropyl)-3-methyl-8- (4-
(trifluorornethoxy)phenoxy)- 1 H-purine-2 ,6(3 H,7H)-dione
4-((7-((5 -chloropyridin-2 - 2.270 467 yl)methyl)-l -(3-
1 hydroxypropyl)-3-methyl-
2,6-dioxo-2,3,6,7-tetrahydro- lH-purin-8- yl)oxy)benzonitrile
? 7-((5 -chloropyridin-2- 1.667 470 yl)methyl)-8-(4-
1 ethylphenoxy)-l -(3- hydroxypropyl)-3 -methy 1-
Figure imgf000586_0001
1 H-purine-2,6(3H,7H)-dione 2014/027920
Figure imgf000587_0001
230 7-benzyl-l -(3- 3.541 421
I rQ hydroxypropyl)-3-methyl-8- (m-tolyloxy)-l H-purine- 2,6(3H,7H)-dione
231 7-benzyl-l-(3- 3.399 437 hydroxypropyl)-8-(3- methoxyphenoxy)-3-methyl- 1 H-purine-2,6(3H,7H)-dione
232 7-benzyl-l-(3- 2.603 491
1 rQ' hydroxypropyl)-3-methyl-8-
(3-
(trifluoromethoxy)phenoxy)- 1 H-purine-2,6(3H,7H)-dione
33 3-((7-benzyl-l -(3- 3.160 432 hydroxypropyl)-3 -methyl- 2,6-dioxo-2,3,6,7-tetrahydro- 1 H-purin-8- yl)oxy)benzonitrile
34 7-benzyl-8-(3-ethylphenoxy)- 3.669 435
1 -(3-hydroxypropyl)-3- methyl- 1 H-purine- 2,6(3H,7H)-dione
35 7-benzyl-l-(3- . 3.015 449
I rO hydroxypropyl)-8-(3- isopropylphenoxy)-3-methyl- 1 H-purine-2,6(3H,7H)-dione
36 4-((7-benzyl-l -(3- 3.140 432
5 rO hydroxypropyl)-3-methyl-
2,6-dioxo-2,3,6,7-tetrahydro- lH-purin-8- yl)oxy)benzonitrile
Figure imgf000589_0001
2,6(3H,7H)-dione 7-benzyl-l -(3- 3.054 449 hydroxypropyl)-8-(4- isopropylphenoxy)-3-methyl- 1 H-purine-2,6(3H,7H)-dione
7-(4-chlorobenzyl)-3 -ethyl- 1 - 2.936 455 (3-hydroxypropyl)-8-
Figure imgf000590_0001
phenoxy- 1 H-purine- 2,6(3H,7H)-dione
7-(4-chlorobenzyl)-8-(2- 3.034 489 chlorophenoxy)-3-ethyl-l-(3-
Figure imgf000590_0002
hydroxypropyl)- 1 H-purine- 2,6(3H,7H)-dione
7-(4-chlorobenzyl)-3-ethyl-8- 2.835 473 (2-fluorophenoxy)- 1 -(3 - hydroxypropyl)- 1 H-purine- 2,6(3H,7H)-dione
7-(4-chlorobenzyl)-3-ethyl-l - 2.021 469 (3-hydroxypropyl)-8-(o- -Q tolyloxy)-lH-purine- 2,6(3H,7H)-dione
7-(4-chlorobenzyl)-3-ethyl-l- 2.884 485 (3-hydroxypropyl)-8-(2-
Figure imgf000590_0003
methoxyphenoxy)- 1 H-purine- 2,6(3H,7H)-dione
7-(4-chlorobenzyl)-3-ethyl-l- 3.063 523 l rO-CI (3-hydroxypropyl)-8-(2- (trifluoromethyl)phenoxy)- 1 H-purin e-2, 6 (3 H, 7H) -d i one
7-(4-chlorobenzyl)-3-ethyl-l- 3.120 539
(3-hydroxypropyl)-8-(2-
(trifluoromethoxy)phenoxy)-
Figure imgf000591_0001
purine-2,6(3H,7H)-dione 259 7-(4-chlorobenzyl)-8-(3- 3.148 489 chlorophenoxy)-3-ethyl-l -(3- hydroxypropyl)- 1 H-purine- 2,6(3H,7H)-dione
260 7-(4-chlorobenzyl)-3-ethyl-8- 3.901 473 l rOra (3-fluorophenoxy)-l -(3- hydxoxypropyl)- 1 H-purine- 2,6(3H,7H)-dione
261 7-(4-chlorobenzyl)-3-ethyl-l - 3.001 469
(3-hydroxypropyl)-8-(m-
° / N tolyloxy)- 1 H-purine- 2,6(3H,7H)-dione
262 7-(4-chlorobenzyl)-3 -ethyl- 1- 2.904 485
(3-hydroxypropyl)-8-(3- methoxyphenoxy)- 1 H-purine- 2,6(3H,7H)-dione
263 7-(4-chlorobenzyl)-3 -ethyl- 1 - 3.138 523
(3 -hy droxypropy 1) - 8 -(3 - (trifluoromethyl)phenoxy)- lH-purine-2,6(3H,7H)-dione
264 7-(4-chlorobenzyl)-8-(3 - 2.976 521
(difluoromethoxy)phenoxy)- 3 -ethyl- 1 -(3-hydroxypropyl)- lH-purine-2,6(3H,7H)-dione
265 3-((7-(4-chlorobenzyl)-3- 2.690 480
1 €ra ethyl- 1 -(3 -hydroxypropyl)-
.J <L -CN 2,6-dioxo-2,3,6,7-tetrahydro- lH-purin-8- yl)oxy)benzonitrile
Figure imgf000593_0001
1 H-purine-2,6(3H,7H)-dione
Figure imgf000594_0001
Example 277 7-(4-fluorobenzyl)-l-(3-hydroxypropyl)-3-methyl-8-p-tolyl-lH-purine- 2,6(3H,7H)-dione
Figure imgf000594_0002
To a solution of 8-bromo-7-(4-fluorobenzyl)-l-(3-hydroxypropyl)-3-methyl-lH-purine- 2,6(3H,7H)-dione (50 mg, 0.121 mmol, intermediate 62) in toluene (2 mL) and ethanol (0.5 mL) was added p-tolylboronic acid (19.7 mg, 0.145 mmol) followed by aqueous sodium carbonate solution (0.5 mL), and the mixture was degassed under nitrogen atmosphere three times.
Tetrakis(triphenylphosphine)platinum (7.5 mg, 0.006 mmol) was added to the reaction under nitrogen and the resulting mixture was stirred at 100 °C for 16 h. The mixture was partitioned between ethyl acetate and brine. The organic phase was dried and concentrated to give crude product, which was purified via preparative HPLC to give 7-(4-fluorobenzyl)- l -(3- hydroxypropyl)-3-methyl-8-p-tolyl-lH-purine-2,6(3H,7H)-dione (20.9 mg, 40.9% yield) as white solid. 'H-NMR ( CD3OD) δ 7.51 -7.53 (d, 2Η), 7.34-7.36 (d, 2Η), 6.97-7.07 (m, 4Η), 5.66 (s, 2Η), 4.09-4.12 (t, 2Η), 3.59-3.62 (t, 2Η), 3.59 (s, 3Η), 2.44 (s, 3Η), 1 .85-1.89 (m, 2Η). LCMS retention time 2.306; LCMS MH+ 423.
The examples in Table 3 were prepared using the method of example 277.
Table 2
Figure imgf000595_0001
Figure imgf000596_0001
lH-purine-2,6(3H,7H)- dione
288 7-(4-fluorobenzyl)-l -(3- 2.367 439 hydroxypropyl)-8-(4- methoxyphenyl)-3-methyl- lH-purine-2,6(3H,7H)- dione
289 7- (4-fluorobenzyl)-l -(3- 2.453 477 hydroxypropyl)-3 -methyl -
1 8- (4-
(trifluoromethyl)phenyl)- lH-purine-2,6(3H,7H)- dione
290 8-(2-chlorophenyl)-7-(4- 2.219 443 fluorobenzyl)- 1 -(3 - hydroxypropyl)-3-methyl- lH-purine-2,6(3H,7H)- dione
91 7- (4-fluorobenzyl)-l -(3- 2.432 423 hydroxypropyl)-3-methyl-
8- (o-tolyl)- 1 H-purine- 2,6(3H,7H)-dione
92 7-(4-fluorobenzyl)-8-(2- 2.307 427 fluorophenyl)- 1 -(3 -
Figure imgf000597_0001
hydroxypropyl)-3-methyl- lH-purine-2,6(3H,7H)- dione
93 7-(4-fluorobenzyl)-l -(3- 2.303 439 hydroxypropyl)-8-(2-
1 0 methoxyphenyl)-3-methyl-
\
lH-purine-2,6(3H,7H)- dione
Figure imgf000598_0001
hydroxypropyl)-3 -methy 1 - l H-purine-2,6(3H,7H)- dione
300 7-((5-chloropyridin-2- 2.3 13 444 o yl)methyl)-8-(3- fluorophenyl)-l -(3- hydroxypropyl)-3-methyl- lH-purine-2,6(3H,7H)- dione
301 7- ((5-chloropyridin-2- 2.432 440 yl)methyl)-l -(3- hydroxypropyl)-3 -methyl-
8- (m-tolyl)- 1 H-purine- 2,6(3H,7H)-dione
302 7-((5-chloropyridin-2- 2.276 456 yl)methyl)-l -(3- hydroxypropyl)-8-(3- methoxyphenyl)-3-methyl- l H-purine-2,6(3H,7H)- dione
303 7-((5-chloropyridin-2- 2.791 468 yl)methyl)-l -(3- hydroxypropyl)-8-(3- i sopropylphenyl) -3 - methyl- 1 H-purine- 2,6(3H,7H)-dione
304 7- ((5-chloropyridin-2- 2.614 494 yl)methyl)-l -(3- hydroxypropyl)-3-methyl-
8- (3-
(trifluoromethyl)phenyl)- lH-purine-2,6(3H,7H)- dione
305 7- ((5-chloropyridin-2- 2.735 510 yl)methyl)-l -(3- hydroxypropyl)-3-methyl-
8- (3-
(trifluoromethoxy)phenyl)- lH-purine-2,6(3H,7H)- dione
306 8-([l ,l'-biphenyl]-3-yl)-7- 2.750 502
((5-chloropyridin-2- yl)methyl)-l-(3- hydroxypropyl)-3-methyl- lH-purine-2,6(3H,7H)- dione
307 8-(4-chlorophenyl)-7-((5- 2.501 461 chloropyridin-2- yl)methyl)-l -(3- hydroxypropyl)-3 -m ethyl - lH-purine-2,6(3H,7H)- dione
308 7-((5-chloropyridin-2- 2.281 444 yl)methyl)-8-(4- fluorophenyl)- 1 -(3- hydroxypropyl)-3-methyl- lH-purine-2,6(3H,7H)- dione
309 7- ((5 -chloropyridin-2- 2.413 440 yl)methyl)-l -(3- hydroxypropyl)-3-methyl-
8- (p-tolyl)-l H-purine- 2,6(3H,7H)-dione
310 7-((5-chloropyridin-2- 2.360 456 yl)methyl)-l -(3- hydroxypropyl)-8-(4- methoxyphenyl)-3-methyl- lH-purine-2,6(3H,7H)- dione
311 7- ((5-chloropyridin-2- 2.666 494 yl)methyl)-l -(3- hydroxypropyl)-3-methyl-
8- (4-
(trifluoromethyl)phenyl)- lH-purine-2,6(3H,7H)- dione
312 8-(2-chlorophenyl)-7-((5- 2.100 461 chloropyridin-2- yl)methyl)-l -(3-
Figure imgf000601_0001
hy droxypropyl)-3 -methyl - lH-purine-2,6(3H,7H)- dione
313 7- ((5-chloropyridin-2- 2.306 440 yl)methyl)-l -(3- hydroxypropyl)-3 -methyl -
8- (o-tolyl)- 1 H-purine- 2,6(3H,7H)-dione
314 7-((5-chloropyridin-2- 2.175 444 yl)methyl)-8-(2- fluorophenyl)- 1 -(3 - hydroxypropyl)-3 -methyl - lH-purine-2,6(3H,7H)- dione
Figure imgf000602_0001
dione
Example 319 7-((5-fluoropyridin-3-yl)methyl)-l-(3-hydroxypropyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000602_0002
To a solution of l -(3-hydroxypropyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)- l H-purine- 2,6(3H,7H)-dione (60 mg, 0.15 mmol, intermediate 15) in DMF (5 mL) was added 3- (chloi methyl)-5-fluoropyridine (80 mg, 0.55 mmol), potassium carbonate (0.55 g, 0.6 mmol), and TBAI (2 mg, 0.02 mmol). The reaction was heated at 60 °C for 2h. The mixture was cooled and partitioned between ethyl acetate and water. The organic phase was dried over sodium sulfate, and concentrated. The residue was purified by preparative HPLC to give 7-((5- fluoropyridin-3-yl)methyl)-l-(3-hydroxypropyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)- lH- purine-2,6(3H,7H)-dione(35 mg, 46 % yield) as white solid. LCMS retention time 2.577 min; LCMS. MH+ 510.
Example 320 1 -(3-hydroxypropyl)-3-methyl-7-((6-methylpyridin-2-yl)methyl)-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000603_0001
The title compound was prepared using the method of example 319 and purified via preparative HPLC to give l-(3-hydroxypropyl)-3-methyl-7-((6-methylpyridin-2-yl)methyl)-8-(3-
(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (20 mg, 36.2% yield) as white solid. 'H-NMR (CD3OD) 6 7.60-7.56(t, 1H), 7.42-7.40(m, 2H), 7.27-7.25(d, 1H), 4.30-4.26(t, 2H), 4.13-4.10(t, 2H), 3.64-3.61(1, 2H), 3.44(s, 3H), 1.94-1.87(m, 4H), 1.02-0.980(t, 3H). LCMS retention time 2.803 min; LCMS MH+ 443.
The examples in Table 3 were prepared using the methods of example 319.
Table 3
Figure imgf000603_0002
Figure imgf000604_0001
dione
Figure imgf000605_0001
-7-(3 ,3 ,3 -trifluoropropy 1)-
Figure imgf000606_0001
dione
Example 335 7-(4-chlorobenzyl)-3-ethyl-l-(3-hydroxypropyl)-8-propoxy- 2,6(3H,7H)-dione
Figure imgf000607_0001
To a solution of 8-chloro-3-ethyl-7-(4-fluorobenzyl)-l -(3-hydroxypropyl)-lH-purine- 2,6(3H,7H)-dione (50 mg, 0.1 17 mmol, intermediate 78) in propanol (5 mL) was added sodium (10 mg, 0.43 mmol). The reaction was stirred at room temperature for 1 h. The reaction was partitioned between ethyl acetate and brine. The organic phase was dried and concentrated to give a crude product, which was purified via preparative HPLC to give 7-(4-chlorobenzyl)-3- ethyl-l -(3-hydroxypiOpyl)-8-propoxy- lH-purine-2,6(3H,7H)-dione (1 1.6 mg, 24.4% yield) as white solid. 'H-NMR (CD3OD) δ 7.24-7.29(s, 4H), 5.21 (s, 2H), 4.39-4.42(t, 2H), 3.98-4.05(m, 4H), 3.49-3.53(m, 2H), 1 .73-1.81 (m, 4H), 1.20-1.23(m, 3H), 0.90-0.94(m, 3H). LCMS retention time 2.881 ; LCMS MH+ 421.
The examples in Table 4 were prepared using the method of example 335.
Table 4
Figure imgf000607_0002
Figure imgf000608_0001
dione
Figure imgf000609_0001
purine-2,6(3H,7H)-dione
Example 348 7-(4-Chlorobenzyl)-3-methyl-l-(2-oxopropyl)-8-(3-(trifluoromethoxy)phenoxy)- lH- urine-2,6(3H,7H)-dione
Figure imgf000609_0002
7-(4-Chlorobenzyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (0.25 g, 0.54 mmol, intermediate 9), potassium carbonate (0.1 1 g, 0.80 mmol), TBAI (0.10 g) and 1 -chloropropan-2-one (0.040 mL, 0.59 mmol) were combined in DMF (3 mL) and heated at 50°C for 1.5 h. The reaction was cooled to room temperature, diluted with water (100 mL) and extracted with ethyl acetate (3 x 75 mL). The combined organic extracts were washed with IN lithium chloride (2 x 75 mL) dried with magnesium sulfate, filtered and evaporated under reduced pressure to an off-white solid. The solid was purified using a 25 g silica gel flash column eluted with 1% methanol / DCM to give 7-(4-chlorobenzyl)-3 -methyl- 1 -(2-oxopropyl)-8-(3 - (trifluoromethoxy)phenoxy)-l H-purine-2,6(3H,7H)-dione (0.22 g, 79% yield) as a white solid. LCMS retention time 4.433 minutes and 99% purity, LCMS MH"" 523. 'HNMR (DMSO-d6) δ 7.60 (t, 1Η, J = 8 Hz), 7.50-7.53 (m, 1H), 7.38-7.47 (m, 5H), 7.32 (d, 1H, J = 8 Hz), 5.40 (s, 2H), 4.73 (s, 2H) 3.28 (s, 3H), 2.19 (s, 3H).
Example 349 7-(4-Chlorobenzyl)-3-methyl-l-((3-methylisoxazol-5-yl)methyl)-8-(3- trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000610_0001
The title compound was prepared using the method of example 348. White solid. LCMS retention time = 4.601 minutes and 99% purity, LCMS MH+ 562. 'HNMR (DMSO-d6) δ (DMSO-d6) δ 7.60 (t, 1Η, J = 8 Hz), 7.47-7.51 (m, 1H), 7.40-7.45 (m, 5H), 7.32 (d, 1 H, J - 8 Hz), 6.20 (s, 1H), 5.43 (s, 2H), 5.13 (s, 2H), 3.30 (s, 3H), 2.17 (s, 3H).
Example 350 Ethyl-2(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-(trifluoromethoxy)phenoxy)- 2 -tetrahydro- 1 H-purin- 1 -yl)acetate
Figure imgf000610_0002
The title compound was prepared using the method of example 348. White solid. LCMS retention time = 4.697 minutes and 99% purity, LCMS MH+ 553. 'HNMR (DMSO-d6) δ 7.58 (t, 1Η, J = 8 Hz), 7.51-7.54 (m, 1H), 7.38-7.47 (m, 5H), 7.32 (d, 1H, J = 12 Hz), 5.41 (s, 2H), 4.60 (s, 2H), 4.12 (dd, 2H, J = 8 Hz and 16 Hz), 3.30 (s, 3H). 1.18 (t, 3H, J = 8 Hz). Example 351 7-(4-Chlorobenzyl)-l -(2-hydroxypropyl)-3-methyl-8-(3- purine-2,6(3H,7H)-dione
Figure imgf000611_0001
7-(4-Chlorobenzyl)-3-methyl-l-(2-oxopropyl)-8-(3-(trifluoromethoxy)phenoxy)-l H-purine- 2,6(3H,7H)-dione (0.18 g, 0.35 mmol, example 348) was dissolved in methanol (3 mL) and cooled to 0°C. Then sodium borohydride (0.040 g, 1.05 mmol) was added portion wise over 20 minutes. The reaction was stirred in the cold for 2 h. The reaction solvent was removed under reduced pressure, then diluted with water (100 mL) and extracted with ethyl acetate (3 x 75 mL). The combined extracts were dried with magnesium sulfate, filtered and evaporated under reduced pressure to a light golden solid. Solid was purified using a 12 g silica gel flash column eluted with 1% methanol / DCM to give 7-(4-chlorobenzyl)-l -(2-hydroxypropyl)-3-methyl-8- (3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (0.15 g, 79% yield) as a white solid. LCMS retention time = 4.280 minutes and 97% purity, LCMS MH+ 525. 'HNMR
(DMSO-d6) δ 7.59 (t, 1H, J = 8 Hz), 7.46-7.48 (m, 1H), 7.38-7.43 (m, 5H), 7.31 (d, 1H, J - 8 Hz), 5.43 (s, 2H), 4.65 (d, 1H, J = 8 Hz), 3.90-3.95 (m, 2H), 3.68-3.74 (m,lH), 3.28 (s, 3H), 1.02 (d, 3H, J = 8 Hz).
Example 352 (S*)-l-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-trifluoromethoxy)phenoxy)- 2 3, ,7-tetrahydro-lH-purin-l -yl)propan-2yl acetate
Figure imgf000611_0002
and (R*)-7-(4-chlorobenzyl)-l-(2-hydroxypropyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-2,6(3H,7i )-dione
Figure imgf000612_0001
7-(4-Chlorobenzyl)- 1 -(2-hydroxypropyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)- 1 H-purine- 2,6(3H,7H)-dione (0.25g, 0.48 mmol, example 351), vinyl acetate (0.15 mL, 1.67 mmol) and Candida antartica acrylic resin ("Novozym") (0.14 g) were combined in ethyl acetate (5 iriL) and stirred at room temperature for 24 h. The reaction was filtered and the filtrate was evaporated under reduced pressure to a clear oil. The oil was purified using a 25 g silica gel flash column eluted with a gradient of 20% to 30% ethyl acetate / hexanes to yield both (S*)-l -(7-(4- chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-trifluoromethoxy)phenoxy)-2,3,6,7-tetrahydro-lH-purin- l-yl)propan-2yl acetate (0.12 g, 91% yield). LCMS retention time = 4.448 minutes and 94% purity, LCMS MH+ 567. 'HNMR (DMSO-d6) δ 7.60 (t, 1H, J = 8 Hz), 7.48-7.54 (m, 1H), 7.38-7.45 (m, 5H), 7.32 (d, 1H, J = 8 Hz), 5.42 (s, 2H), 5.10-5.19 (m, 1H), 4.13-4.22 (m, 1H), 3.85 (dd, 1H, J = 4 Hz and 12 Hz), 3.27 (s, 3H), 1.79 (s, 3H), 1.17 (d, 3H, J =? 4 Hz) and (R*)-7- (4-chlorobenzyl)-l -(2-hydroxypropyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-2,6(3H,7H)- dione (0.1 1 g, 88% yield) as a clear oil. LCMS retention time = 4.019 minutes and 95% purity, LCMS MH+ = 525. 'HNMR (DMSO-d6) δ 7.59 (t, 1Η, J = 8 Hz), 7.45-7.48 (m, 1H), 7.39-7.44 (m, 5H), 7.31 (d, 1H, J = 8 Hz), 5.43 (s, 2H), 4.65 (d, 1H, J = 4 Hz), 3.89-4.00 (m, 2H), 3.67- 3.74 (m, 1H), 3.28 (s , 3H), 1.02 (d, 3H, J = 8 Hz).
Example 353 2-(7-(4-Chlorobenzyl)-3-methyl-3-methyl-2,6-dioxo-8-(3- trifluoromethoxy)phenoxy)-2,3,6,7-tetrahydro-lH-purin-l -yl)-N-methylacetamide
Figure imgf000612_0002
The title compound was prepared using the method of example 348 but with heating at 50 °C for 15 h. White solid: 0.18g, 63% yield: LCMS retention time = 4.084 minutes and 99% purity, LCMS MH+ 538. 'HNMR (DMSO-d6) δ 7.92-7.98 (m, ΝΗ), 7.60 (t, 1 Η, J = 8 Hz), 7.47-7.51 (m, 1 H), 7.38-7.45 (m, 5H), 7.29-7.34 (d, 1 H, J = 8 Hz), 5.42 (s, 2H), 4.41 (s, 2H), 3.28 (s, 3H), 2.57 (d, 3H, J = 4 Hz).
Example 354 7-(4-Chlorobenzyl)-3-methyl- 1 -(thiazol-5-ylmethyl)-8-(3- trifluoromethoxy)phenoxy)- 1H-purine-2,6(3H,7//)-dione
Figure imgf000613_0001
The title compound was prepared using the method of example 348 but with heating at 50 °C for 3 h. White solid, 0.088 g, 29% yield: LCMS retention time = 4.552 minutes and 99% purity, LCMS MH+ 564. 'HNMR (DMSO-d6) δ 8.97 (s, lH), 7.85 (s, 1H), 7.58 (t, 1H, J = 8 Hz), 7.46-7.48 (m, 1H), 7.39-7.43 (m, 5H), 7.30 (d, 1H, J = 12 Hz), 5.43 (s, 2H), 5.24 (s, 2H), 3.29 (s, 3H).
Example 355 7-(4-Chlorobenzyl)-l -(2-hydroxy-2-methylpentyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000613_0002
7- (4-Chlorobenzyl)-3-methyl-l -(2-oxopropyl)-8-(3-(trifluoromethoxy)phenoxy)- lH-purine- 2,6(3//,7H)-dione (0.25 g, 0.48 mmol, example 348) was dissolved in THF (6 mL), cooled to 0°C and 2.0 M propyl magnesium bromide in THF (0.36 mL, 0.72 mmol) was added drop wise. The reaction was stirred in the cold for 15 min then warmed to room temperature and stirred 1 h. The reaction was diluted with water (100 mL) and extracted with ethyl acetate (3 x 75 mL). The combined extracts were dried with magnesium sulfate, filtered and evaporated under reduced pressure to a golden oil. The oil was purified using a 25 g silica gel flash column eluted with 20% ethyl acetate / hexanes to give 7-(4-chlorobenzyl)-l-(2-hydroxy-2-methylpentyl)-3-methyl-
8- (3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7 /)-dione (0.88 g, 32% yield) as a clear oil. LCMS retention time = 4.792 minutes and 98% purity, LCMS MH+ = 567. 'HNMR (DMSO-d6) δ 7.59 (t, l H, J = 8 Hz), 7.47-7.50 (m, 1H), 7.38-7.44 (m, 5H), 7.31 (d, 1 H, J = 8 Hz), 5.42 (s, 2H), 4.23 (s, 1H) 3.92-4.02 (dd, 2H, J = 4 Hz and 16 Hz), 3.29 (s, 3H), 1.30-1.41 (m, 4H), 0.99 (s, 3H), 0.80 (t, 3H, J = 8 Hz).
Example 356 7-(4-Chlorobenzyl)-l -(3-hydroxypropyl)-3-methyl-8-(3- 2,6(3H,7H)-dione
Figure imgf000614_0001
Step 1 7-(4-chlorobenzyl)-3-methyl-8-(3-(triiluoromethoxy)phenoxy)-l H-2,6(3H,7H)-dione (3.73 g, 7.99 mmol, intermediate 9) and potassium carbonate (1.66 g, 1 1 .99 mmol) were combined in DMF (56 mL) and (3-bromopropoxy)(tert-butyl)dimethylsilane (2.43 g, 9.59 mmol) was added. The reaction was heated at 100°C. After heating for 3 h the reaction was cooled, diluted with water (200 mL) and extracted with ethyl acetate (3 x 200 ml). The combined extracts were washed with IN lithium chloride (2 x 100 mL), dried with magnesium sulfate and evaporated under reduced pressure to provide l-(3-((tert-butyldimethylsilyl)oxy)propyl)-7-(4- chlorobenzyl)-3-methyl-8-(3-trifluoromethoxy)phenoxy)-l /-purine-2,6(3H,7H)-dione (5.7 g crude) as a light golden oil. LCMS retention time = 5.646 and 98% purity, LCMS MH+ 639.
Step 2 l-(3-((tert-Butyldimethylsilyl)oxy)propyl)-7-(4-chlorobenzyl)-3-methyl-8-(3- trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (5.7 g crude, 7.99 mmol) was dissolved in ethanol (40 mL) and 6N aqueous HC1 (4 mL) was added. The clear solution was stirred at room temperature. After 1 h the reaction was evaporated under reduce pressure, diluted with water (200 mL) and extracted with ethyl acetate (3 x 100 mL). The combined extracts were dried with magnesium sulfate, filtered and evaporated under reduced pressure to a clear oil. The oil was purified using an 80 g silica gel flash column eluted with 1 : 1 ethyl acetate / hexanes to provide 7-(4-chlorobenzyl)-l -(3-hydroxypropyl)-3-methyl-8-(3-trifluoromethoxy)phenoxy)-lH- purine-2,6(3H,7H)-dione (3.8 g, 90.9% yield) as a white crystalline solid. LCMS retention time = 3.955 and 100% purity, LCMS MH+ 525. 'HNMR (DMSO-d6) δ 7.59 (t, 1 H, J = 2 Hz), 7.47 (s, 1H), 7.41 (s, 4H), 7.41 (m, 1H), 7.31 (d, 1 H, J = 2 Hz), 5.42 (s, 2H), 4.44 (t, OH, J = 1 Hz), 3.92 (t, 2H, J = 1 Hz), 3.41 (m, 2H), 3.28 (s, 3H), 1.68 (m, 2H). Example 357 l -(3-Aminopropyl)-7-(4-chlorobenzyl)-3-methyl-8-(3- rifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione hydrochloride
Figure imgf000615_0001
Step 1 7-(4-Ch]orobenzyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)- dione (1.0 g, 2.14 mmol, intermediate 9), 2-(3-bromopropyl)isoindoline-l ,3-dione (0.63 g, 2.36 mmol), potassium carbonate (0.44 g, 3.21 mmol) and TBAI (0.040 g) were combined in DMF (20 mL) and heated at 100°C for 3 h. The reaction was cooled to room temperature, diluted with water (200 mL) and extracted with ethyl acetate (3 x 100 mL). The combined organic extracts were washed with IN lithium chloride (2 x 100 mL), dried with magnesium sulfate, filtered and the solvent removed under reduced pressure to give a clear oil. The oil was purified using an 80 g silica gel flash column eluted with a gradient of 25% to 50% ethyl acetate / hexanes to give 7- (4-chlorobenzyl)-l -(3-(l ,3-dioxoisoindolin-2-yl)propyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-l /-purine-2,6(3 ,7H)-dione (1.4 g, 100% yield) as a white solid. LCMS retention time = 4.660 minutes and 100% purity, LCMS MH+ 654. 'HNMR (CDC13) δ 7.78-7.83 (m, 2H), 7.65-7.70 (m, 2H), 7.41 (t, 1H, J = 8 Hz), 7.32 (dd, 4H, J = 12 Hz and 28 Hz), 7.10-7.23 (m, 4H), 5.35 (s, 2H), 4.05-4.14 (m, 2H), 3.76 (t, 2H, J = 8 Hz), 3.38 (s, 3H), 2.01-2.1 1 (m, 2H).
Step 2 7-(4-Chlorobenzyl)-l-(3-(l,3-dioxoisoindolin-2-yl)propyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)-dione (1.4 g, 2.14 mmol) was slurried in ethanol (20 mL) and hydrazine hydrate (0.42 mL, 8.56 mmol) was added and the reaction was heated at reflux. The reaction at reflux changed to a clear solution then after 30 min setup as a white mass. The reaction was cooled to room temperature, diluted with water (100 mL) and extracted with DCM (3 x 75 mL). The combined organic extracts were dried with magnesium sulfate, filtered and the solvent removed under reduced pressure to give an oil. The oil was purified using a 40 g silica gel flash column eluted with 10% methanol / DCM to give a light tan solid. Solid was dissolved in DCM (5 mL) and excess IN HQ / diethyl ether was added. The solvent was removed under reduced pressure to give l -(3-aminopropyl)-7-(4-chlorobenzyl)-3- methyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3 /,7H)-dione hydrochloride (0.87 g, 49% yield) as a tan solid. LCMS retention time = 2.802 minutes and 97% purity, LCMS MH+ 524. 'HNMR (DMSO-d6) δ 7.91-8.1 1 (brd s, NH2 and HC1), 7.60 (t, 1H, J = 8 Hz), 7.38-7.48 (m, 6H), 7.31 (d, 1H, J - 12 Hz), 5.43 (s, 2H), 3.93 (t, 2H, J = 8 Hz), 3.29 (s, 3H), 2.73-2.85 (m, 2H), 1.83-1 .94 (m, 2H).
Example 358 l-(2-Aminoethyl)-7-(4-chlorobenzyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-.
- urine-2,6(3H,7H)-dione hydrochloride
Figure imgf000616_0001
The title compound was prepared using the method of example 357. White solid, 0.062 g, 33% yield: LCMS retention time = 2.802 minutes and 99% purity, LCMS•MHf 510. 'HNMR
(DMSO-d6) δ 7.82-7.99 (brd s, N¾ and HC1), 7.62 (t, 1H, J = 8 Hz), 7.38-7.48 (m, 6H), 7.31 (d, 1H, J = 12 Hz), 5.43 (s, 2H), 4.1 1 (m, 2H) 3.29 (s, 3H), 2.99-2.15 (m, 2H).
Example 359 N-(3-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-(trifluoromethoxy)phenoxy)- 2,3 ,6,7-tetrahydro- 1 H-purin- 1 -yl)propyl)acetamide
Notebo : C5-0249-047
Figure imgf000616_0002
l -(3-Aminopropyl)-7-(4-chlorobenzyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine- 2,6(3H,7H)-dione (0.10 g, 0.19 mmol, example 357) and TEA (0.80 mL, 0.57 mmol) were combined in THF (1 mL). Acetyl chloride (0.28 mL, 0.42 mmol) was added and the reaction was stirred at room temperature for 4 h. The reaction was diluted with water (100 mL) and extracted with ethyl acetate (3 x 75 mL). The combined extracts were dried with magnesium sulfate, filtered and evaporated under reduced pressure to an oil. The oil was purified using a 2000 micron preparative TLC plate eluted with 5 % methanol / DCM. Target band was scraped off plate and eluted off silica gel with eluent to give N-(3-(7-(4-chlorobenzyl)-3-methyl-2,6- dioxo-8-(3-(trifluoromethoxy)phenoxy)-2,3,6,7-tetrahydro-l H-purin-l -yl)propyl)acetamide (0.053 g, 49% yield) as a light tan solid. LCMS retention time = 3.789 minutes and 99% purity, LCMS MH+ 566. ' NMR (DMSO-d6) δ 7.78-7.82 (m, 1H), 7.59 (t, 1H, J = 8 Hz), 7.44-7.48 (m, 1H), 7.38-7.42 (m, 5H), 7.31 (d, 1H, J = 8 Hz), 5.42 (s, 2H), 3.87 (t, 2H, J = 8 Hz), 3.28 (s, 3H), 2.04 (dd, 2H, J = 4 Hz and 12 Hz), 1.78 (s, 3H), 1.61-1.70 (m, 2H).
Example 360 N-(3-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-(trifluoromethoxy)phenoxy)- 2 3 ,6,7-tetrahydro- 1 H-purin- 1 -yl)propyl)methanesulfonamide
Figure imgf000617_0001
l -(3-Aminopropyl)-7-(4-chlorobenzyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine- 2,6(3H,7H)-dione (0.10 g, 0.19 mmol, example 357) and TEA (0.80 mL, 0.57 mmol) were combined in THF (1 mL). Methanesulfonyl chloride (0.30 mL, 0.38 mmol) was added and the reaction was stin-ed at room temperature for 3 h. The reaction was diluted with water (100 mL) and extracted with ethyl acetate (3 x 75 mL). The combined extracts were dried with magnesium sulfate, filtered and evaporated under reduced pressure to an oil. The oil was purified using a 2000 micron preparative TLC plate eluted with 5 % methanol / DCM. Target band was scraped off plate and eluted off silica gel with eluent to give N-(3-(7-(4-chlorobenzyl)-3-methyl-2,6- dioxo-8-(3-(trifluoromethoxy)phenoxy)-2,3,6,7-tetrahydro-l H-purin-l - yl)propyl)methanesulfonamide (0.045 g, 39% yield) as a tan solid. LCMS retention time = 4.048 minutes and 99% purity, LCMS MH+ 602. '//NMR (DMSO-d6) δ 7.59 (t, 1H, J - 8 Hz), 7.45- 7.48 (m, 1 H), 7.39-7.43 (m, 5H), 7.31 (d, 1H, J = 8 Hz), 6.96 (t, 1H), 5.43 (s, 2H), 3.91 (t, 2H, J = 8 Hz), 3.29 (s, 3H), 2.96 (dd, 2H, J = 4 Hz and 12 Hz), 2.87 (s, 3H), 1.69- 1.79 (m, 2H). Example 361 2-(7-(4-Chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-(trifluoromethoxy)phenoxy)- 6,7-tetrahydro- 1 H-purin- 1 -yl)acetic acid
Figure imgf000618_0001
Step 1 7-(4-Chlorobenzyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine-2,6(3H,7H)- dione (1.0 g, 2.14 mmol, intermediate 9), /er/-butyl 2-bromoacetate (0.34 mL, 2.36 mmol), and potassium carbonate (0.45 g, 3.21 mmol) were combined in DMF (20 mL) and heated at 50°C for 2 h. The reaction was cooled to room temperature diluted with water (200 mL) and extracted with ethyl acetate (3 x 100 mL). The combined extracts were washed with IN lithium chloride (2 x 100 mL), dried with magnesium sulfate, filtered and evaporated under reduced pressure to give tert-butyl-2-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-(tri fluoromethoxy)phenoxy)-2, 3,6,7- tetrahydro-1 H-purin- l -yl)acetate (1.24 g, 100% yield) as a clear oil. LCMS retention time = 4.817 minutes and 97% purity, LCMS MH+ 581. Step 2 teri-Butyl-2-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-(trifluoromethoxy)phenoxy)- 2,3,6,7-tetrahydro-lH-purin-l-yl)acetate (0.20 g, 0.34 mmol) was dissolved in DCM (4 mL) and TFA (0.26 ml, 3.4 mmol) was added. The reaction was stirred at room temperature for 15 h then evaporated under reduced pressure to give 2-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3- (trifluoromethoxy)phenoxy)-2,3, 6,7-tetrahydro-l H-purin- l -yl)acetic acid (0.18 g , 100% yield). LCMS retention time = 3.915 minutes and 97% purity, LCMS MH+ 525. !HNMR (DMSO-d6) δ 7.59 (t, 1Η, J = 8 Hz), 7.50-7.54 (m, 1H), 7.45 (d, 1 H, J = 8 Hz), 7.41 (s, 4H), 7.31 (d, 1 H, J = 8 Hz), 5.41 (s, 2H), 4.52 (s, 2H), 3.30 (s, 3H)
Example 362 2-(7-(4-Chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-(trifluoromethoxy)phenoxy)- 2,3,6,7-tetrahydro- 1 H-purin- 1 -yl)-N-propylacetamide
Figure imgf000619_0001
2-(7-(4-Chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-(trifluoromethoxy)phenoxy)-2,3,6,7-tetrahydro- lH-purin-l -yl)acetic acid (0.25g, 0.48 mmol, Example 361) and CDI (0.93 g, 0.57 mmol) were combined in DMF (4 mL) and stirred at room temperature for 20 min. Propan- 1 -amine was added and the reaction was stirred at room temperature for 3 h. The reaction was diluted with water (100 mL) and extracted with ethyl acetate (3 x 75 mL). The combined extracts were washed with IN lithium chloride (2 x 75 mL), dried with magnesium sulfate, filtered and evaporated under reduced pressure to a white solid. Solid was purified using a 25 g silica gel flash column eluted with 1% methanol / DCM to give 2-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo- 8-(3-(trifluoromethoxy)phenoxy)-2,3,6,7-tetrahydro-lH-purin-l-yl)-N-propylacetamide (0.13 g, 47 % yield) as a white solid. LCMS retention time = 4.070 min and 99% purity, LCMS MH+ 566. 'HNMR (DMSO-d6) δ 7.99 (t, NH), 7.60 (t, 1H, J - 8 Hz), 7.48-7.52 (m, lH), 7.39-7.46 (m, 5H), 7.32 (d, 1H, J = 8 Hz), 5.42 (s, 2H), 4.42 (s, 2H), 3.31 (s, 3H), 3.00 (dd, 2H, J = 8 Hz and 16 Hz), 1.34-1.45 (m, 2H), 0.83 (t, 3H, J = 8 Hz).
Example 363 7-(4-Chlorobenzyl)-3-methyl-l -(2-oxobutyl)-8-(3-(trifluoromethoxy)phenoxy)- lH- urine-2,6(3H,7H)-dione
Figure imgf000619_0002
The title compound was prepared using the method of example 348. White solid, 0.49 g, 85 % yield: LCMS retention time = 4.408 min and 99% purity, LCMS MH+ 537. 'HNMR (DMSO- d6) δ 7.60 (t, 1Η, J = 8 Hz), 7.51-7.54 (m, 1H), 7.37-7.47 (m, 5H), 7.30-7.33 (m, 1H), 5.40 (s, 2H), 4.72 (s, 2H), 3.28 (s, 3H), 2.55 (dd, 2H, J = 8 Hz and 16 Hz), 0.96 (t, 3H, J = 8 Hz). Example 364 7-(4-Chlorobenzyl)-8-ethoxy-l -(3 -hydroxypropyl)-3 -methyl- lH-purine- 2 6(3H,7H)-dione
Figure imgf000620_0001
Step 1 Sodium (0.35 g, 15.03 mmol) was dissolved in ethanol (30 mL) and 8-bromo-7-(4- chlorobenzyl)-3 -methyl- lH-purine-2,6(3H,7H)-dione (1.63 g, 3.01 mmol, intermediate 77) was added and the reaction was stirred at room temperature for 24 h. The reaction was evaporated to dryness under reduced pressure, diluted with water (100 mL) and extracted with ethyl acetate (3 x 75 mL). The combined extracts were dried with magnesium sulfate, filtered and the solvent was removed under reduced pressure to give l-(3-(( ^butyldimethylsilyl)oxy)propyl)-7-(4- cl lorobenzyl)-8-ethoxy-3-methyl-lH-purine-2,6(3H,7H)-dione (1.5 g, 100 % yield) as a light golden oil. LCMS retention time = 5.317 min and 95% purity, LCMS MH+ 507.
Step 2 7-(4-Chlorobenzyl)-8-ethoxy-l-(3-hydroxypropyl)-3-methyl-lH-purine-2,6(3H,7H)- dione
(1.5 g, 3.01 mmol) was dissolved in ethanol (20 mL) and 6N aqueous HC1 (4 mL) was added. The clear solution was stirred at room temperature for 1 h. The reaction was diluted with water (150 mL) and extracted with DCM (3 x 100 mL). The combined extracts were dried with magnesium sulfate, filtered and the solvent was removed under reduced pressure to an off-white solid. The solid was purified using an 80 g silica gel flash column eluted with 2% methanol / DCM which gave 7-(4-chlorobenzyl)-8-etlioxy-l-(3-hydroxypropyl)-3-methyl-lH-purine- 2,6(3 H,7H)-dione (1 .0 g, 84% yield) as a white solid. LCMS retention time = 3.173 min and 98% purity, LCMS MH+ 393. 'HNMR (DMSO-d6) δ 7.35 (dd, 4H, J = 4 Hz and 12 Hz), 5.20 (s, 2H), 4.47 ( dd, 2H, J = 8 Hz and 16 Hz), 4.43 (t, 1H, J = 4 Hz)), 3.88 (t, 2H, J = 8 Hz), 3.40 (dd, 2H, J = 4 Hz and 12 Hz), 3.36 (s, 3H), 1.62- 1.71 (m, 2H), 1.33 (t, 3H, J = 8 Hz).
Example 365 7-(4-Chlorobenzyl)-l-(2-hydroxybutyl)-3-methyl-8-(3- (trifluoromethoxy)phenoxy)- 1 H-purine-2,6(3H,7H)-dione
Figure imgf000621_0001
7-(4-Chlorobenzyl)-3-methyl-l -(2-oxobutyl)-8-(3-(tn:fluoromethoxy)phenoxy)-l / -purine- 2,6(3H,7H)-dione (0.44 g, 0.82 mmol, example 363) was dissolved in methanol (8 mL) and DCM (1 mL), then cooled to 0°C. Sodium borohydride (0.93 g, 2.46 mmol) was added portion wise over 20 min and reaction was stirred in the cold for 2 h. The reaction solvent was removed under reduced pressure then the mixture was diluted with water (100 mL) and extracted with ethyl acetate (3 x 75 mL). The combined organic extracts were dried with magnesium sulfate, filtered and the solvent removed under reduced pressure to give a clear oil. The oil was purified using an 40 g silica gel flash column eluted with 1% methanol / DCM to give 7-(4- chlorobenzyl)-l -(2-hydroxybutyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-lH-purine-
2,6(3H,7H)-dione (0.32 g, 73% yield) as a white solid. LCMS retention time = 4.265 min and 97% purity, LCMS MH+ 539. 'HNMR (DMSO-d6) 6 7.59 (t, 1H, J - 8 Hz), 7.45-7.48 (m, 1 H), 7.38-7.45 (m, 5H), 7.30 (d, 1H, J = 12 Hz), 5.43 (s, 2H), 4.56 (d, 1H, J = 4 Hz), 3.92-4.00 (m, 1H), 3.65-3.76 (m, 2H), 3.28 (s, 3H), 1.23-1 .41 (m, 2H).
Example 366 7-(4-Chlorobenzyl)-l -(3-hydroxypropyl)-3-methyl-8-(propylamino)-lH-purine- 2 6(3H,7H)-dione hydrochloride
Figure imgf000621_0002
Step 1 8-Bromo-l-(3-((tert-butyldimethylsilyl)oxy)propyl)-7-(4-chlorobenzyl)-3-methyl-lH- purine-2,6(3H,7H)-dione (0.25 g, 0.46 mmol, intermediate 77) and propan- 1 -amine (1.44 mL, 9.2 mmol) were combined in ethanol (5 mL) and heated at reflux for 24 h. The reaction was - cooled to room temperature, diluted with water (75 mL) and extracted with ethyl acetate (3 x 75 mL). The combined organic extracts were dried with magnesium sulfate, filtered and the solvent evaporated under reduced pressure to yield a light golden oil. The oil was purified using a 25 g silica gel column eluted with 20% ethyl acetate / hexanes to give l -(3-((tert- butyldimethylsilyl)oxy)propyl-7-(4-chlorobenzyl)-3-methyl-8-(propylamino)-l H-purine- 2,6(3H,7H)-dione (0.15 g, 63% yield) as a white solid. LCMS retention time = 5.013 min and 97% purity, LCMS MH+ 520.
Step 2 l -(3-((½rt-Butyldimethylsilyl)oxy)propyl-7-(4-chlorobenzyl)-3-methyl-8-(propylamino)- lH-purine-2,6(3H,7H)-dione (0.15 g, 0.29 mmol) was dissolved in ethanol (3 mL) and 6N aqueous HCl (0.5 mL) was added. The clear solution was stirred at room temperature for 1 h. The reaction solvent was removed under reduced pressure to give 7-(4-chlorobenzyl)-l-(3- hydroxypropyl)-3-methyl-8-(propylamino)-lH-purine-2,6(3H,7H)-dione hydrochloride (0.13 g, 100% yield) as a white foam. LCMS retention time = 3.021 min and 99% purity, LCMS MH+ 406. 'HNMR (DMSO-d6) δ 7.31 (dd, 4H, J = 8 Hz and 42 Hz), 6.15-6.48 (brd s, NH2 and HCl), 5.30 (s, 2H), 3.84 (t, 2H, J = 8 Hz), 3.38 (t, 2H, J = 8 Hz), 3.34 (s, 3H), 3.26 (t, 2H, J = 8 Hz), 1.59-1.68 (m, 2H), 1.46-1.57 (m, 2H), 0.81 (s, 3H, J = 8 Hz).
Example 367 7-(4-Chlorobenzyl)-l -(3-hydroxypropyl)-3-methyl-8-propoxy-lH-purine- 2 6(3H,7H)-dione
Figure imgf000622_0001
The title compound was prepared using the method of example 364. White solid, 0.14 g, 78%) yield: LCMS retention time = 2.914 min and 98% purity, LCMS MH+ 379. 'HNMR (DMSO- d6) δ 7.35 (dd, 4Η, J - 12 Hz and 40 Hz), 5.21 (s, 2H), 4.36-4.44 (m, 3H), 3.89 (t, 2H, J = 4 Hz), 3.41 (dd, 2H, J = 4 Hz and 12 Hz), 3.35 (s, 3H), 1.62-1.78 (m, 4H), 0.88 (t, 3H, J = 8 Hz).
Example 368 7-(4-Chlorobenzyl)-l-(3-hydroxypropyl)-8-methoxy-3-methyl-lH-purine- 2,6(3H,7H)-dione
Figure imgf000623_0001
The title compound was prepared using the method of example 364. White solid, 0.14 g, 78%: LCMS retention time = 2.914 min and 98% purity, LCMS MH+ 379. 'HNMR (DMSO-d6) δ 7.34 (dd, 4H, J = 8 Hz and 48 Hz), 5.21 (s, 2H), 4.42 (t, 1H, J = 4 Hz), 4.08 (s, 3H), 3.88 (t, 2H, J - 8 Hz)), 3.38-3.43 (m, 2H), 3.37 (s, 3H), 1 .62-1.70 (m, 2H).
Example 369 8-Butoxy-7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl-lH-purine- 2,6(3H,7H)-dione
Figure imgf000623_0002
The title compound was prepared using the method of example 364. White solid, 0.15 g, 77% yield: LCMS retention time = 3.690 min and 97% purity, LCMS MH+ 421. 'HNMR (DMSO-d6) δ 7.34 (dd, 4H, J = 4 Hz and 12 Hz), 5.21 (s, 2H), 4.43 ( m, 3H), 3.89 (t, 1 H, J = 8 Hz)), 3.41 (dd, 2H, J = 4 Hz and 12 Hz), 1.62-1.72 (m, 2H), 1.25-1.36 (m, 2H), 0.86 (t, 3H, J = 8 Hz).
Example 370 7-(4-Chlorobenzyl)-l -(3-hydroxypropyl)-8-isopropoxy-3-methyl- lH-purine- 2 6(3H,7H)-dione
Figure imgf000623_0003
The title compound was prepared using the method of example 364. White solid, 0.056 g, 30% yield: LCMS retention time = 3.355 min and 94% purity, LCMS MH+ 407. 'HNMR (DMSO-d6) δ 7.36 (dd, 4H, J = 8 Hz and 48 Hz), 5.19 (s, 2H), 5.12-5.18 (m, 1H), 4.44 (t, 1H, J = 4 Hz), 3.80 (t, 2H, J = 8 Hz), 3.38-3.45 (m, 2H), 3.37 (s, 3H), 1.63-1.72 (m, 2H), 1.34 (d, 6H, J = 8 Hz).
Example 371 7-(4-Chlorobenzyl)- l -(3-hydroxypropyl)-3-methyl-8-(2-mo holinoethoxy)-l H- urine-2,6(3H,7H)-dione hydrochloride
Figure imgf000624_0001
Step 1 Sodium (0.053 g, 2.31 mmol) was dissolved in 2-morpholinoethanol (5 mL) and 8- bromo-l -(3-((tert-butyldimethylsilyl)oxy)propyl)-7-(4-chlorobenzyl)-3-methyl-lH-purine- 2,6(3H,7H)-dione (0.25 g, 0.46 mmol, intermediate 77) was added. The reaction was stirred at room temperature for 24 h. The reaction was diluted with water (75 mL) and extracted with ethyl acetate (3 x 75 mL). The combined extracts were dried with magnesium sulfate, filtered and the solvent was removed under reduced pressure to give l-(3-((tert-butyldimethylsilyl)oxy)propyl)- 7-(4-chlorobenzyl)-3-methyl-8-(2-morpholinoethoxy)-lH-purine-2,6(3H,7H)-dione (0 27 g, 100% yield) as a clear oil. LCMS retention time = 2.990 min and 96% purity, LCMS MH+ 592.
Step 2 1 -(3-((ter/-Butyldimethylsilyl)oxy)propyl)-7-(4-chlorobenzyl)-3-methyl-8-(2- morpholinoethoxy)-lH-purine-2,6(3//,7H)-dione (0.27g, 0.46 mmol) was dissolved in ethanol (5 mL) and 6N aqueous HC1 (1 mL) and stirred at room temperature for 1 h. The reaction was evaporated and azeotroped with methanol (3 x 10 mL). The solid residue was triturated with methanol (5 mL) and filtered. The white solid was washed with diethyl ether (2 x 10 mL)and high vacuum dried to give 7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl-8-(2- morpholinoethoxy)-lH-purine-2,6(3H,7H)-dione hydrochloride (0.14, 57% yield) as a white solid. LCMS retention time = 1.908 min and 95% purity, LCMS MH+ 478. ]HNMR (DMSO-d6) δ 1 1.54 (s, HC1), 7.38 (dd, 4H, J = 12 and 28 Hz), 5.33 (2, 2H), 4.82-4.89 (m, 2H), 3.75-3.95 (m, 2H), 3.58-3.64 (m, 2H), 3.33-3.44 (m, 5H), 3.06 (m, 2H), 1 .62-1.72 (m, 2H). Example 372 7-(4-Chlorobenzyl)-8-(2-(dimethylamino)ethoxy)-l -(3-hydroxypropyl)-3-methyl- lH- urine-2,6(3H,7H)-dione hydrochloride
Figure imgf000625_0001
The title compound was prepared using the method of example 371 . White solid, 0.153 g, 95% yield: LCMS retention time = 1.837 min and 94% purity, LCMS MH+ 436. 'HNMR (DMSO-d6) δ 10.70 (s, HC1), 7.37-7.42 (m, 4H), 5.35 (s, 2H), 4.77-4.83 (m, 2H), 3.87-3.93 (m, 3H), 3.52- 3.60 (m, 2H), 3.40-3.44 (t, 2H, J = 8 Hz), 3.39 (s, 3H), 2.50 (s, 6H), 1.62-1.72 (m, 2H).
Example 373 7-(4-chlorobenzyl)-8-(cyclopentyloxy)-l -(3-hydroxypropyl)-3-methyl-lH-purine- 2 6(3H,7H)-dione
Figure imgf000625_0002
Step 1 Cyclopentanol (0.21 mL, 2.31 mmol) was dissolved in THF (5 mL) and sodium hydride (60% in oil, 0.092 g, 2.31 mmol) was added. The reaction was stirred for 1 h. 8-Bromo-l -(3- ((tert-butyldimethylsilyl)oxy)propyl)-7-(4-chlorobenzyl)-3 -methyl- lH-purine-2,6(3H,7/J)-dione (0.25 g, 0.46 mmol, intermediate 77) was added and the reaction was stirred at room temperature for 15 h. The reaction was diluted with water (100 mL) and extracted with ethyl acetate (3 x 75 mL). The combined extracts were dried with magnesium sulfate, filtered and the solvent was removed under reduced pressure to give l-(3-((tert-butyldimethylsilyl)oxy)propyl)-7-(4- chlorobenzyl)-8-(cyclopentyloxy)-3-methyl-l H-purine-2,6(3H,7H)-dione (0.25 g, 100% yield) as a golden oil. LCMS retention time = 5.843 min and 71% purity, LCMS MH+ 547.
Step 2 1 -(3-((tert-Butyldimethylsilyl)oxy)propyl)-7-(4-chlorobenzyl)-8-(cyclopentyloxy)-3- methyl-lH-purine-2,6(3H,7H)-dione (0.25 g, 0.46 mmol) was dissolved in ethanol (5 mL) and 6N aqueous HC1 (1 mL) was added. The clear solution was stirred at room temperature for 1 h. The reaction was diluted with water (100 mL) and extracted with ethyl acetate (3 x 100 mL). The combined extracts were dried with magnesium sulfate, filtered and the solvent was removed under reduced pressure to a golden oil. The oil was purified using a 2000 micron preparative TLC plate eluted with ethyl acetate. Target band was scraped off plate, eluted off silica gel with ethyl acetate and evaporated under reduced pressure to give 7-(4-chlorobenzyl)-8- (cyclopentyloxy)-l -(3-hydroxypropyl)-3-methyl-lH-purine-2,6(3H,7H)-dione (0.028 g, 14% yield) as a white solid. LCMS retention time = 3.656 min and 98% purity, LC S MH+ 433. H
NMR (DMSO-d6) δ 7.34 (dd, 4H, J = 12 and 48 Hz), 5.34-5.40 (m, 1H), 5.1 8 (s, 2H), 4.42 (t, 1H, J = 4 Hz), 3.89 (t, 2H, J = 8 Hz), 3.38-3.45 (m, 2H), 3.36 (s, 3H), 1.55- 1.93 (m, 8H).
Example 374 7-(4-Chlorobenzyl)-8-(cyclohexyloxy)-l -(3-hydroxypropyl)-3-methyl-l -purine- -dione
Figure imgf000626_0001
The title compound was prepared using the method of example 373. White solid, 0.86 g, 42% yield: LCMS retention time = 3.929 min and 97% purity, LCMS MH+ 447. 'HNMR (DMSO-d6) δ 7.35 (dd, 4H, J = 12 Hz and 36 Hz), 5.20 (s, 2H), 4.92-4.99 (m, 1H), 4.42 (t, 1 H, J = 4 Hz), 3.99 (t, 2H, J = 8 Hz), 3.38-3.44 (m, 2H), 3.35 (s, 3H), 1.83-1.94 (m, 2H), 1 .50-1.72 (m, 6H), 1.20-1.45 (m, 4H).
Example 375 7-(4-Chlorobenzyl)-l -(3-hydroxypropyl)-3-methyl-8-(pentyloxy)-lH-purine- 2,6(3H,7H)-dione
Figure imgf000626_0002
The title compound was prepared using the method of example 373. White solid, 0.13 g, 63% yield: LCMS retention time = 3.886 min and 98% purity, LCMS MH+ 435. 'HNMR (DMSO-d6) δ 7.34 (dd, 4H, J - 8 Hz and 44 Hz), 5.21 (s, 2H), 4.36-4.49 (m, 3H), 3.99 (t, 3H, J = 8 Hz), 3.37-3.45 (m, 2H), 3.35 (s, 3H), 1 .62-1.72 (m, 4H), 1.16-1.30 (m, 4H), 0.82 (t, 3H, J = 8 Hz).
Example 376 7-(4-Chlorobenzyl)-8-(cyclopentylmethoxy)-l -(3-hydroxypropyl)-3-methyl-l/ - purine-2,6(3H,7H)-dione
using the method of example 373. White solid, 0.13 g, 63% yield: LCMS retention time = 3.970 min and 95% purity, LCMS MH+ 447 and. !HNMR (DMSO-d6) δ 7.34 (dd, 4H, J = 8 Hz and 44 Hz), 5.21 (s, 2H), 4.42 (brd s, 1H), 4.31 (d, 2H, J = 4 Hz), 3.89 (t, 3H, J = 8 Hz), 3.41 (t, 2H, J = 8 Hz), 3.35 (s, 3H), 2.23-2.35 (m, 2H), 1.62-1.72 (m, 4H), 1.44-1.59 (m, 4H), 1.16-1.28 (m, 2H).
Example 377 7-(4-Chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl-8-((6-methylpyridin-3- l)methoxy)-lH-purine-2,6(3H,7H)-dione
Figure imgf000627_0001
The title compound was prepared using the method of example 373. White solid, 0.0.87 g, 44% yield: LCMS retention time = 2.255 min and 97% purity, LCMS MH+ 470. 'HNMR (DMSO-d6) δ 7.34 (dd, 4H, J = 8 Hz and 44 Hz), 5.21 (s, 2H), 4.36-4.49 (m, 3H), 3.99 (t, 3H, J = 8 Hz), 3.37-3.45 (m, 2H), 3.35 (s, 3H), 1.62-1.72 (m, 4H), 1.16-1.30 (m, 4H), 0.82 (t, 3H, J = 8 Hz).
Example 378 Ethyl 2-(7-(4-chlorobenzyl)- l -(3-hydroxypropyl)-3-methyl-2,6-dioxo-2,3,6,7- tetrahydro- 1 H-purin-8-yl)acetate
Figure imgf000627_0002
Step 1 Diethyl malonate (0.35 mL, 2.31 mmol) was dissolved in DMF ( 5 mL) and sodium hydride (60% in oil, 0.74 g, 1.85 mmol) was added portion wise. The mixture was stirred for 15 min and 8-bromo- l -(3-((ter/-butyldimethylsilyl)oxy)propyl)-7-(4-chlorobenzyl)-3-methyl-l H- purine-2,6(3H,7H)-dione (0.50 g, 0.92 mmol, intermediate 77) was added. The reaction was heated at 100 °C for 15 h. The reaction was cooled, diluted with water (100 mL) and extracted with ethyl acetate (3 x 75 mL). The combined extracts were washed with IN lithium chloride (2 x 100 mL), dried with magnesium sulfate, filtered and the solvent was removed under reduced pressure to a golden oil. The oil was purified using a 25 g silica gel flash column eluted with 20% ethyl acetate / hexanes to give diethyl-2-(l -(3-((/er/-butyldimethylsilyl)oxy)propyl)-7-(4- chlorobenzyl)-3-methyl-2,6-dioxo-253,6,7-tetrahydro-lH-purin-8-yl)malonate (0.23 g, 40% yield) as a clear oil. LCMS retention time = 5.015 min and 95% purity, LCMS MH+ 621. ]H NMR (DMSO-d6) δ 7.27 (dd, 4H, J = 8 Hz and 72 Hz), 5.62 (d, 2H, J = 12 Hz), 5.54 (s, 2H), 5.61 (s, IH), 3.90-4.03 (m, 6H), 3.62 (t, 2H, J = 8 Hz), 3.37 (s, 3H), 1 .70-1.78 (m, 2H), 1.08 (t, 6H, J = 8 Hz), 0.80 (s, 9H).
Step 2 Diethyl-2-(l -(3-((/er/-butyldimethylsilyl)oxy)propyl)-7-(4-chlorobenzyl)-3-methyl-2,6- dioxo-2,3,6,7-tetrahydro-lH-purin-8-yl)-2-(3-trifiuoromethyl)phenyl)malonate (0.23 g, 0.36 mmol) was slurried in 18% aqueous HC1 (3 mL) and heated at reflux for 3 h, yielding a clear solution. The solvent was removed under reduced pressure and the residue was high vacuum dried for 15 h to give 2-(7-(4-chlorobenzyl)-l -(3-hydroxypropyl)-3-methyl-2,6-dioxo-2,3,6,7- tetrahydro-l H-purin-8-yl)acetic acid (0.14 g, 100% yield) as a tan solid. LCMS retention time = 2.318 min and 70% purity, LCMS MH+ 407.
Step 3 Crude 2-(7-(4-chlorobenzyl)-l -(3-hydroxypropyl)-3-methyl-2,6-dioxo-2,3,6,7- tetrahydro-lH-purin-8-yl)acetic acid (0.14 g, 0.36 mmol) was dissolved in ethanol (5 mL) and concentrated sulfuric acid (1 drop). The reaction was heated at reflux for 1 h. The reaction was cooled, evaporated under reduced pressure to remove the ethanol then diluted with water (50 mL) and extracted with ethyl acetate (3 x 40 mL). The combined extracts were dried with magnesium sulfate, filtered and the solvent was removed under reduced pressure to a light golden oil. The oil was purified using a 12 g silica gel flash column and eluted with 20% ethyl acetate / hexanes to give ethyl 2-(7-(4-chlorobenzyl)-l -(3-hydroxypropyl)-3-methyl-2,6-dioxo- 2,3,6,7-tetrahydro-lH-purin-8-yl)acetate (0.062, 40% yield) as a light tan solid. LCMS retention time = 2.804 min and 99% purity, LCMS MH+ 435. NMR (CDC13) δ 7.21 (dd, 4H, J = 8 Hz and 84 Hz), 5.57 (s, 2H), 4.1 1 -4.23 (m, 4H), 3.77 (s, 2H), 3.59 (s, 3H), 3.50-3.56 (m, 2H), 3.32- 3.39 (m, lH), 1.86-1.94 (m, 2H), 1.25 (t, 3H, J = 8 Hz).
Example 379 7-(4-Chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl-8-(3- -2,6(3/y, 7H)-dione
Figure imgf000629_0001
Step 1 Sodium hydride (60% in oil, 0.15 g, 3.69 mmol) was added portion wise to ethyl-2-(3- (trifluoromethoxy)phenyl)acetate (1.03 g, 4.15 mmol) in DMF (5 mL) and the dark golden solution was stirred at room temperature for 20 min. 8-Bromo-l-(3-((tert- butyldimethylsilyl)oxy)propyl)-7-(4-chlorobenzyl)-3-methyl-lH-purine-2,6(3H,7H)-dione (0.50 g, 0.92 mmol, intermediate 77) was added and the reaction was heated at 100°C for 2 h. The reaction was cooled, diluted with water (100 mL) and extracted with ethyl acetate (3 x 100 mL). The combined extracts were washed with IN lithium chloride (2 x 100 mL), dried with magnesium sulfate, filtered and the solvent was removed under reduced pressure to a golden oil. The oil was purified using a 40 g silica gel flash column eluted with 10% ethyl acetate / hexanes to give ethyl-2-(l -(3-((½rt-butyldimethylsilyl)oxy)propyl)-7-(4-chloi benzyl)-3-methyl-2,6- dioxo-2,3,6,7-tetrahydro-lH-purin-8-yl)-2-(3-trifluoromethoxy)phenyl)acetate (0.54 g, 82% yield) as a light golden oil. LCMS retention time = 5.407 min and 99% purity, LCMS MH+ 709.
Step 2 Ethyl-2-(l -(3-((tert-butyldimethylsilyl)oxy)propyl)-7-(4-chlorobenzyl)-3-methyl-2,6- dioxo-2,3,6,7-tetrahydro-lH-purin-8-yl)-2-(3-trifluoromethoxy)phenyl)acetate was slurried in 6N HCl (6 mL) and heated at reflux for 1 h. The reaction solution was cooled, diluted with water (50 mL) and extracted with DCM (3 x 50 mL). The combined extracts were dried with magnesium sulfate, filtered and the solvent was removed under reduced pressure to a light golden oil. The oil was purified using a 12 g silica gel flash column eluted with a gradient of 20% ethyl acetate / hexanes to 100% ethyl acetate to give 7-(4-chlorobenzyl)-l -(3- hydiOxypropyl)-3-methyl-8-(3-(trifluorornethoxy)benzyl)-lH-purine-2,6.(3H, 7H)-dione (0.088 g, 60% yield) as a clear oil. LCMS retention time = 3.623 min and 98% purity, LCMS MH+ 523. 'HNMR (DMSO-de) δ 7.33 (t, 1H, J = 8 Hz), 7.25-7.29 (m, 2H), 7.05-7.18 (m, 5H), 5.61 (s, 2H), 4.41 (t, 11-1, J = 4 Hz), 4.24 (s, 2H), 3.89 (t, 2H, J = 12 Hz), 3.40 (s, 3H), 3.38-3.43 (m, 2H), 1.62-1.72 (m, 2H).
Example 380 Ethyl-2-(7-(4-chlorobenzyl)-l -(3-hydroxypropyl)-3-methyl-2,6-dioxo-2,3,6,7- tetrahydro-lH-purin-8-yl)-2-(3-trifluoromethoxy)phenyl)propanoate
Figure imgf000630_0001
Sodium (0.025 g, 1.07 mmol) was dissolved in ethanol (3 mL) and ethyl-2-(l -(3-((ter/- butyldimethylsilyl)oxy)propyl)-7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-2,3,6,7-tetrahydro-lH- purin-8-yl)-2-(3-trifluoromethoxy)phenyl)acetate (0.33 g , 0.47 mmol, example 379, step 1) was added. The reaction was stirred for 10 min and methyl iodide (0.29 mL, 4.7 mmol) and the reaction was heated at reflux for 1 h. The reaction was cooled, evaporated under reduced pressure, diluted with water (50 mL) and extracted with ethyl acetate (3 x 50 mL). The combined extracts were dried with magnesium sulfate, filtered and the solvent was removed under reduced pressure to a golden oil. The oil was purified using a 25 g silica gel flash column eluted with 20% ethyl acetate / hexanes to give ethyl-2-(7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl- 2,6-dioxo-2,3,6,7-tetrahydro-lH-purin-8-yl)-2-(3-trifluoromethoxy)phenyl)propanoate (0.17 g, 67% yield) as a white foam. LCMS retention time = 3.263 min and 99% purity, LCMS MH+ 609. 'HNMR (CDC13) 5 7.06-7.24 (m, 5H), 6.73 (d, 2H, J = 8 Hz), 5.14 (s, 2H), 4.39 (t, 1H, J = 8 Hz), 3.95-4.05 (m, 2H), 3.81-3.89 (m, 2H), 3.35-3.42 (m, 2H), 1 .95 (s, 3H), 1.59-1.67 (m, 2H), 1.09 (t, 3H, J = 8 Hz). Example 381 7-(4-Chlorobenzyl)-l -(3-hydroxypropyl)-3-methyl-8-(3-(trifluoromethyl)benzyl)- lH-purine-2,6(3H,7H)-dione
Figure imgf000631_0001
Step 1 Ethyl-2-( l -(3-((t<?r/-butyldimethylsilyl)oxy)propyl)-7-(4-chlorobenzyl)-3-methyl-2,6- dioxo-2,3,6,7-tetrahydro-17-/-purin-8-yl)-2-(3-trifluoromethyl)phenyl)acetate was prepared from intermediate 77 and ethyl 3-trifluoromethylphenylacetate using the method of example 378, step 1. (0.45 g, 70 % yield) as a yellow foam. LCMS retention time = 5.334 min and 87% purity, LCMS MH" 693.
Step 2 7-(4-Chlorobenzy])-l-(3-hydiOxypropyl)-3-methyl-8-(3-(trifluoromethyl)benzyl)-lH- purine-2,6(3H,7//)-dione was prepared using the method of example 379, step 2. Clear oil, 0.032 g, 44% yield: LCMS retention time = 3.523 min and 99% pure, LCMS MLT 507. 'HNMR (DMSO-d6) δ 7.65-7.75 (m, 1H), 7.47-7.52 (m, lH), 7.43 (d, 1 H, J = 4 Hz), 7.38 (s, 1H), 7.17 (dd, 4H, J = 8 Hz and 68 Hz), 5.53 (s, 2H), 4.29 (s, 2H), 4.1 1 -4.15 (m, 1H), 3.90 (t, 1H, J = 4 Hz), 3.40 (s, 3H), 3.37-3.45 (m, 2H), 1.60-1.71 (m, 2H). Example 382 Ethyl 2-(7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl-2,6-dioxo-2,3,6,7- tetrah dro-lH-purin-8-yl)-2-(3-(trifluoromethyl)phenyl)acetate
Figure imgf000631_0002
Ethyl-2-(l -(3-((tert-butyldimethylsilyl)oxy)propyl)-7-(4-chlorobenzyl)-3-methyl-2,6-dioxo- 2J3,6,7-tetrahydro-lH-purin-8-yl)-2-(3-trifluoromethyl)phenyl)acetate (0.050 g, 0.72 mmol, example 381 , step 1 ) was dissolved in ethanol (3 mL) and 6N aqueous HC1 (0.3 mL) was added. The clear solution was stirred at room temperature for 1 h. The reaction solvent was removed under reduced pressure, added DCM (3 mL) and directly spotted on a 2000 micron preparative TLC plate which was eluted with 50% ethyl acetate / hexanes. Target band was scraped off plate, eluted off silica gel with ethyl acetate and the solvent removed under reduced pressure to give ethyl 2-(7-(4-chlorobenzyl)-l -(3-hydroxypropyl)-3-methyl-2,6-dioxo-2,3,6,7-tetrahydro-lH- purin-8-yl)-2-(3-(trifluoromethyl)phenyl)acetate ( 0.025 g, 50 % yield) as a white foam. LCMS retention time = 3.73 1 min and 96% purity, LCMS MH+ 579. 'HNMR (DMSO-d6) δ 7.55-7.65 (m, 3H), 7.48 (t, 1 H, J = 8 Hz), 7.12 (dd, 4H, J = 8 Hz and 72 Hz), 5.69 (dd, 2H, J = 16 Hz and 32 Hz), 4.40 (brd s, 1 H), 4.05 (dd, 2H, J = 8 Hz and 16 Hz), 3.88 (t, 2H, J = 8 Hz), 3.41 ( s, 3H), 1 .61 -1 .69 (m, 2H).
Example 383 Ethyl 2-(7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl-2,6-dioxo-2,3,6,7- tetrahydro-lH-purin-8-yl)-2-(3-(trifluoromethyl)phenyl)propanoate
Figure imgf000632_0001
Ethyl 2-(7-(4-chlorobenzyl)-l -(3-hydroxypropyl)-3-methyl-2,6-dioxo-2,3,6,7-tetrahydro-lH- purin-8-yl)-2-(3-(trifluoromethyl)phenyl)propanoate was prepared from example 381 , step 1 using the method of example 380. Pale yellow foam, 0.13 g, 61 % yield: LCMS retention time = 3.999 min and 96% purity, LCMS MH+ 593. 'HNMR (DMSO-d6) δ 7.51 (s, 2H), 7.40 (t, 1H, J - 8 Hz), 7.32 (d, 1H, J = 8 Hz), 6.90 (dd, 4H, J = 8 Hz and 92 Hz), 5.15 (s, 2H), 4.39 (t, 1H), 3.98-4.07 (m, 2H), 3.84-3.95 (m, 3H), 3.48 (s, 3H), 3.35-3.42 (m, 2H), 3.61 (s, 3H), 1.59-1.78 (m, 2H), 1.12 (t, 3H, J = 8 Hz).
Example 384 2-(7-(4-Chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl-2,6-dioxo-2,3,6,7- tetrah dro-lH-purin-8-yl)-2-(3-(trifluoromethoxy)phenyl)acetonitrile
Figure imgf000632_0002
Step 1 Sodium hydride (60% in oil, 0.15g, 3.69 mmol) was added portion wise to 2-(3- (trifluoromethoxy)phenyl)acetonitrile (0.84 g, 4.15 mmol) and stirred at room temperature over 15 min. 8-Bromo-l -(3-((ier/-butyldimethylsilyl)oxy)propyl)-7-(4-chlorobenzyl)-3-methyl-lH- purine-2,6(3H,7/-/)-dione (0.50 g, 0.92 mmol, intermediate 77) was added and the reaction was stirred at room temperature for 1 h. The reaction was diluted with water (100 mL) and extracted with ethyl acetate (3 x 100 mL). The combined extracts were washed with IN lithium chloride (2 x 75 mL), dried with magnesium sulfate, filtered and the solvent was removed under reduced pressure to a golden oil. The oil was purified using a 25 g silica gel flash column eluted with 10% ethyl acetate / hexanes to give 2-(l-(3-((tert-butyldimethylsilyl)oxy)propyl)-7-(4- chlorobenzyl)-3-methyl-2,6-dioxo-2,3,6,7-tetrahydro-lH-purin-8-yl)-2-(3- trifluoromethoxy)phenyl)acetonitrile (0.58 g, 95% yield) as a pale yellow foam. LCMS retention time = 5.265 min and 90% purity, LCMS MH+ 662.
Step 2 2-(l -(3-((tert-Butyldimethylsilyl)oxy)propyl)-7-(4-chlorobenzyl)-3-methyl-2,6-dioxo- 2,3,6,7-tetrahydro-lH-purin-8-yl)-2-(3-trifluoromethoxy)phenyl)acetonitrile (0.20 g, 0.30 mmol) was dissolved in ethanol (5 mL) and 6 N aqueous HC1 (1 mL) was added. The clear solution was stirred at room temperature for 1 h. The reaction was diluted with water (50 mL) and extracted with DCM (3 x 50 ml). The combined extracts were dried with magnesium sulfate, filtered and the solvent was removed under reduced pressure to yield a golden oil. The oil was purified using a 25 g silica gel flash column eluted with a gradient of 20% to 50% ethyl acetate / hexanes to give 2-(7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl-2,6-dioxo-2,3,6,7-tetrahydro-l H- purin-8-yl)-2-(3-(trifluoromethoxy)phenyl)acetonitrile (0.082g, 50% yield) as a white foam. LCMS retention time = 3.709 min and 97% purity, LCMS MH+ 548. 'HNMR (CDC13) δ 7.34 (t, 1 H, J = 8 Hz), 7.05-7.25 (m, 5H), 6.88 (d, 2H, J = 8 Hz), 5.54 (dd, 2H, J = 12 Hz and 80 Hz), 4.18 (t, 2H, J = 8 Hz). 3.64 (s, 3H), 3.50-3.58 (m, 2H), 3.13 (t, 1 H, J = 8 Hz), 1.85-1.93 (m, 2H).
Example 385 7-(4-Chlorobenzyl)- l-(3-hydroxypropyl)-3-methyl-8-(l -(3- trifluoi methoxy)phenyl)ethyl-l H-purine- 2,6(3H,7H)-dione
Figure imgf000634_0001
Ethyl-2-(7-(4-chlorobenzyl)-l -(3-hydroxypropyl)-3-methyl-2,6-dioxo-2,3,6,7-tetrahydro-lH- purin-8-yl)-2-(3-trifluoromethoxy)phenyl)propanoate (0.14 g, 0.23 mmol, example 380) was slurried in 6N HC1 (6 mL) and heated at reflux for 15 h. The clear solution was cooled, diluted with water (75 mL) and extracted with DCM (3 x 50 mL). The combined extracts were dried with magnesium sulfate, filtered and the solvent was removed under reduced pressure to a clear oil. The oil was purified using a 2000 micron preparative TLC plate eluted with 50 % ethyl acetate / hexanes. Target band was collected and extracted with ethyl acetate. The extracts were evaporated under reduced pressure to give 7-(4-chlorobenzyl)-l -(3-hydroxypropyl)-3-methyl-8- (l-(3-trifluoromethoxy)phenyl)ethyl-lH-puri e-2,6(3H,7H)-dione (0.026 g, 22% yield) as a white foam. LCMS retention time = 4.029 min and 96% purity, LCMS MH+ 537. 'HNMR (CDCI3) δ 7.20-7.30 (m, 3H), 7.09 (d, 1H, J - 8 Hz), 7.00-7.05 (m, 2H), 6.91 (d, 2H, J = 12 Hz), 5.38 (dd, 2H, J = 16 Hz and 100 Hz), 4.18 ( t, 2H, J = 8 Hz), 4.00-4.09 (m, 1H), 3.67 (s, 3H), 3.48-3.53 (m, 2H), 3.39 (t, 1 H, J = 4 Hz), 1.85-1.93 (m, 2H), 1.66 (d, 3H, J = 8 Hz).
Example 386 7-(4-Chlorobenzyl)-8-(2-hydroxy-l -(3-(trifluoromethoxy)phenyl)ethyl)-l-(3- h droxypropyl)-3 -methyl -lH-purine-2,6(3H,7H)-dione
Figure imgf000634_0002
Ethyl 2-)l -(3-((tert-butyldimethylsilyl)oxy)propyl)-7-(4-chlorobenzyl)-3-methyl-2,6-dioxo- 2,3,6,7-tetrahydro-lH-purin-8-yl)-2-(3-trifluoromethoxy)phenyl)acetate (0.25 g, 0.352 mmol, example 379, step 1) was dissolved in THF (4 mL) and sodium borohydride (0.080 g, 2.1 1 mmol) was added. The reaction was refluxed for 15 min then methanol (2 mL) was added dropwise through condenser. After refluxing another 15 min the reaction was cooled and acidified with 6N HC1 until pH 1 was achieved. The mixture was stirred at room temperature for 1 h. The reaction was diluted with water (100 mL) and extracted with DCM (3 x 75 ml). The combined extracts were dried with magnesium sulfate, filtered and the solvent was removed under reduced pressure to yield an oil. The oil was purified using a 25 g silica gel column eluted with a gradient of 5% to 10% methanol / DCM to give 7-(4-chlorobenzyl)-8-(2-hydroxy-l -(3- (trifluorometboxy)phenyl)ethyl)-] -(3-hy
(0.030 g, 13% yield) as a white foam. LC S retention time = 2.665 min and 98% purity, LCMS MH+ 553. 'HNMR (CDC13) δ 7.25-7.30 (m, 2H), 7.03 (dd, 4H, J = 8 Hz and 91 Hz), 7.1 l(d, lH, J = 8 Hz), 6.99-7.02 (m, 1H), 5.26-5.57 (dd, 2H, J = 16 Hz and 91 Hz), 5.30 (s, 1H), 4.17 (t, 2H, J = 8 Hz), 4.04-4.10 (m, OH), 3.67 (s, 3H), 3.46-3.50 (m, 3H), 3.20-3.27 (m, 1H), 1 .85- 1.93
(m, 2H).
Example 387 7-(4-Chlorobenzyl)-8-(l-hydroxyethyl)-l -(3-hydroxypropyl)-3-methyl-lH- purine-2,6(3H,7H)-dione
Figure imgf000635_0001
Step 1 8-Bromo-l-(3-((rer/-butyldimethylsilyl)oxy)propyl)-7-(4-chlorobenzyl)-3-methyl-lH- purine-2,6(3H,7H)-dione (0.25 g, 0.46 mmol, intermediate 77) was dissolved in THF (5 mL) and cooled to
-78°C. To the clear solution was added 2.5 M n-butyllithium in hexanes (0.20 mL, 0.51 mmol) dropwise. The reaction was stirred in the cold for 5 min and acetaldehyde (0.13 mL, 2.31 mmol) was added. The reaction was warmed to room temperature and stirred for 1 h. The reaction was diluted with water (100 mL) and extracted with ethyl acetate (3 x 75 mL). The combined extracts were dried with magnesium sulfate, filtered and the solvent was removed under reduced pressure to leave a golden oil. The oil was purified using two 1000 microns preparative TLC plates eluted with 50% ethyl acetate / hexanes. Target band was extracted with ethyl acetate and the solvent was removed under reduced pressure to give l -(3-((tert-butyldimethysilyl)oxy)propyl)-7-(4- chlorobenzyl)-8-(l -hydroxyethyl)-3-methyl-lH-purine-2,6(3/ ,7H)-dione (0.052 g, 30% yield) as a clear oil. LCMS retention time = 4.490 min and 97% purity, LCMS MH+ 507. Step 2 l-(3-((tert-butyldimethysilyl)oxy)propyl)-7-(4-chlorobenzyl)-8-(l -hydroxyethyl)-3- methyl- lH-purine-2,6(3H,7H)-dione (0.052 g, 0.10 mmoL) was dissolved in ethanol (2 mL) and 6N aqueous HC1 (0.5 mL) was added. The clear solution was stirred at room temperature for 1 h. The reaction was diluted with water (25 mL) and extracted with DCM (3 x 25 mL). The combined organic extracts were dried with magnesium sulfate, filtered and the solvent removed under reduced pressure to a golden oil. The oil was purified using a 2000 micron preparative TLC plate eluted with 50% ethyl acetate / hexanes. Target band was extracted with ethyl acetate and the solvent was removed under reduced pressure to give 7-(4-chlorobenzyl)-8-(l- hydiOxyethyl)-l -(3-hydroxypropyl)-3-methyl-l /-purine-2,6(3H,7H)-dione (0.012 g, 29% yield) as a clear oil. LCMS retention time = 2.429 min and 96% purity, LCMS MH+ 393. 'HNMR
(CDC13) δ 7.25 (dd, 4H, J = 8 Hz and 64 Hz), 5.65 (dd, J = 12 Hz and 40 Hz), 4.91 -4.97 (m, 1H), 4.19 (t, 2H, J = 4 Hz), 3.61 (s, 3H), 3.51-3.55 (m, 2H), 2.52 (d, 1H, J = 4 Hz), 1.87-1.93 (m, 2H), 1.57 (d, 3H, J = 4 Hz). Example 388 7-(4-Chlorobenzyl)-8-(l-hydroxy-2-methylpropyl)-l -(3-hydroxypropyl)-3- methyl-lH-purine-2,6(3H,7H)-dione
Figure imgf000636_0001
Step 1 8-Bromo-l -(3-((tert-butyldimethylsilyl)oxy)propyl-7-(4-chlorobenzyl)-3-methyl-lH- purine-2,6(3H,7H)-dione (0.50 g , 0.923 mmol, intermediate 77) was dissolved in THF (5 mL) and cooled to -78°C. To the clear solution was slowly added drop wise 2.5 M n-butyllithium in THF (0.35 mL, 0.877 mmol) followed by the immediate addition of isobutyraldehyde (0.42 mL, 4.61 mmol). The reaction was warmed to room temperature and stirred for I h. The reaction was diluted with water (75 mL) and extracted with ethyl acetate (3 x 75 mL). The combined extracts were dried with magnesium sulfate, filtered and the solvent was removed under reduced pressure to yield an oil. The oil was purified using a 25 g silica column eluted with a gradient of 10% to 30% ethyl acetate / hexanes yielding l-(3-((tert-butyldimethylsiIyl)oxy)propyl)-7-(4- chlorobenzyl)-8-(l -hydroxy-2-methylpropyl)-3-methyl-lH-purine-2,6(3H,7H)-dione (0.20 g, 40% yield) as a light golden oil. LCMS retention time = 4.923 min and 98% purity, LCMS MH+ 535.
Step 2 1 -(3-((tert-Butyldimethylsilyl)oxy)propyl)-7-(4-chlorobenzyl)-8-(l -hydroxy-2- methylpropyl)-3 -methyl- lH-purine-2,6(3H,7H)-dione (0.19 g, 0.355 mmol) was dissolved in ethanol (5mL) and 6 N HC1 (0.5 mL) was added. The clear solution was stirred at room temperature for 1 h. The reaction was diluted with water (75 mL) and extracted with DCM (3 x 50 ml). The combined extracts were dried with magnesium sulfate, filtered and the solvent was removed under reduced pressure to yield a white solid. Solid was slurried in diethyl ether (4 mL) and collected to give 7-(4-chlorobenzyl)-8-(l -hydroxy-2-methylpropyl)-l -(3-hydroxypropyl)-3- methyl-lH-purine-2,6(3H,7 /)-dione (0.085g, 56% yield) as a white solid. LCMS retention time - 3.263 min and 94% purity, LCMS MH+ 421. 'HNMR (CDC13) δ 7.23 (dd, 4H, J = 8 Hz and 68 Hz), 5.59 (dd, 2H, J = 16 Hz and 40 Hz), 4.43 (t, 1H, J = 8 Hz). 4.17 (t, 2H, J = 8 Hz), 3.59 (s, 3H), 3.48-3.54 (m, 2H), 3.35-3.40 (m, 1H), 2.48 (d, 1 H, J = 8 Hz), 2.08-2.16 (m, 1H), 1 .85-1.92 (m, 2H), 1.03 (d, 3H, J = 8 Hz), 0.82 (d, 3H, J = 8 Hz).
Example 389 7-(4-Chlorobenzyl)-8-(3-chlorophenoxy)-l -(3-hydroxypropyl)-3-methyl-lH- purine-2,6(3H,7H)-dione
Figure imgf000637_0001
Step 1 8-Bromo-l -(3-((tert-butyldimethylsilyl)oxy)propyl)-7-(4-chlorobenzyl)-3-methyl-l H- purine-2,6(3H,7H)-dione (0.50 g, 0.92 mmol, intermediate 77), potassium carbonate (0.255 g, 1.85 mmol) and 3-chlorophenol (0.104 mL, 0.97 mmol) were combined in DMF (5 mL) and heated at 90°C for 3 h. The reaction was cooled to room temperature, diluted with water (100 mL) and extracted with ethyl acetate (3 x 75 mL). The combined organic extracts were washed with IN lithium chloride (2 x 100 mL), dried with magnesium sulfate, and concentrated under reduced pressure to yield a light golden oil (0.65 g). The oil was purified using a 24 g silica gel column eluted with a gradient of 10% to 20% ethyl acetate / hexanes to yield l-(3-(((/er/- butyldimethylsilyl)oxy)propyl)-7-(4-chlorobenzyl)-8-(3-chlorophenoxy)-3-methyl-l H-purine- 2,6(3H,7H)-dione as an off-white solid (0.413 g, 76% yield). LCMS retention time = 5.598 min and 98% purity, LCMS MH+ 589.
Syep 2 l -(3-(((tert-Butyldimethylsilyl)oxy)propyl)-7-(4-chlorobenzyl)-8-(3-chloiOphenoxy)-3- methyl-lH-purine-2,6(3H,7H)-dione (0.413 g, 0.70 mmol) was dissolved in ethanol (8 mL) and 6N HC1 (1 mL) was added. The clear solution was stirred at room temperature for 1 h. The reaction was diluted with water (75 mL) and extracted with DCM (3 x 75 mL). The combined organic extracts were dried with magnesium sulfate, and concentrated under reduced pressure to leave an off-white solid. The solid was slurried in diethyl ether (3 mL) and filtered to yield 7-(4- chloi benzyl)-8-(3-chlorophenoxy)- l-(3-hydroxypropyl)-3-methyl- lH-purine-2,6(3H,7H)-dione (0.30 g, 90% yield) as an off-white solid. LCMS retention time = 3.810 min and 99% purity, LCMS MH+ 475. 'HNMR (DMSO-d6) δ 7.15-7.41 (m, 8H), 5.41 (s, 2H), 4.20 (t, 2H, J = 8Hz), 3.49-3.51 (m, 2H), 3.45 (s, 3H), 1.87-1.93 (m, 2H).
The following examples 390a through 390r were prepared using the two step method of example 389 Example 390a 7-(4-Chlorobenzyl)-8-(3,4-dichlorophenoxy)l -(3-hydroxypropyl)-3-
Figure imgf000638_0001
White solid, 0.34 g, 92% yield: LCMS retention time = 4.080 min and 99% purity, LCMS MI J+ = 509. 'H NMR (CDCl3) δ 7.50 (d, 1 H, J = 12 Hz), 7.31 -7.44 (m, 5H), 7.15 (dd, 2H, J = 4 Hz and 8 Hz), 5.41 (s, 2H), 3.48-3.51 (m, 2H), 3.45 (s, 3H), 1.87-1.95 (m, 2H). Example 390b 7-(4-Chlorobenzy])-l -(3-hydroxypropyl)-3-methyl-8-(m-tolyloxy)-lH- urine-2,6(3H,7H)-dione
Figure imgf000639_0001
White solid, 0.29 g, 76% yield: LCMS retention time = 3.724 min and 96% purity, LCMS ΜΙ-Γ = 455. 'HNMR (CDC13) δ (CDC13) δ 7.27-7.45 (m, 5Η), 7.01 -7.13 (m, 3Η), 5.41 (s, 2Η), 4.20 (t, 2Η, J = 8 Hz), 3.49-3.57 (m, 2H), 3.44 (s, 3H), 2.39 (s, 3H), 1.86-1.94 (m, 2H).
Example 390c 7-(4-Chlorobenzyl)-l-(3-hydroxypropy])-8-(3-methoxyphenoxy)-3-methy]-lH- purine-2,6(3H,7H)-dione
Figure imgf000639_0002
White solid, 0.14 g, 41% yield: LCMS retention time = 3.550 min and 97% purity, LCMS MI1+ = 471. 'HNMR (CDC13) δ (CDC13) δ 7.29-7.44 (m, 5Η), 6.79-6.86 (m, 3Η), 5.41 (s, 2Η), 4.20 (t, 2Η, J = 8 Hz), 3.82 (s, 3H), 3.50-3.55 (m, 2H), 3.44 (s, 3H), 1.87-1.94 (m, 2H). Example 390d 7-(4-Chlorobenzyl)-8-(3-fluorophenoxy)-l -(3-hydroxypropyl)-3-methyl- -purine-2,6(3H,7H)-dione
Figure imgf000639_0003
White solid, 0.23 g, 74% yield: LCMS retention time = 3.584 min and 99% purity, LCMS MH+ = 459. 'HNMR (CDCI3) δ (CDC13) δ 7.30-7.43 (m, 5Η), 6.98-7.10 (m, 3Η), 5.42 (s, 2Η), 4.20 (t, 2Η, J = 8 Hz), 3.49-3.55 (m, 2H), 3.45 (s, 3H), 1 .88-1.93 (m, 2H). Example 390e7-(4-Chlorobenzyl)-8-(4-chlorophenoxy)-l -(3-hydroxypropyl)-3-methyl-lH- urine-2,6(3H,7H)-dione
Figure imgf000640_0001
White solid, 0.28 g, 76% yield: LCMS retention time = 3.814 min and 99% purity, LCMS MH+ = 475. lHNMR (CDC13) 6 7.37-7.43 (m, 5H), 7.30-7.35 (m, 2H), 7.19-7.23 (m, 2H), 5.41 (s, 2H), 4.20 (t, 2H, J = 8Hz), 3.47-3.56 (m, 2H), 3.43 (s, 3H), 1.86-1.94 (m, 2H).
Example 390f7-(4-Chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl-8-(p-tolyloxy)-lH-purine- 2,6(3H,7H)-dione
Figure imgf000640_0002
White solid, 0.14 g, 41 % yield: LCMS retention time = 3.732 min and 99% purity, LCMS MH+ = 455. 'HNMR (CDCI3) δ (CDC13) δ 7.46 (dd, 4H, J = 8 Hz and J = 44 Hz), 7.24 (dd, 4H, J = 8 Hz and J = 36 Hz), 5.41 (s, 2H), 4.19 (t, 2H, J = 8 Hz), 3.50-3.59 (m, 2H), 3.42 (s, 3H), 2.38 (s, 3H), 1.88-1.92 (m, 2H).
Example 390g 7-Benzyl-8-(3-chlorophenoxy)-l-(3-hydroxypropyl)-3-methyl-lH-purine- -dione
Figure imgf000640_0003
The title compound was prepared from intermediate 83 using the method of example 389.
solid, 0.1 1 g, 61 % yield: LCMS retention time = 3.503 min and 98% purity, LCMS MH+ 'HNMR (CDCI3) δ 7.42-7.47 (m, I H), 7.31 -7.38 (m, 5H), 7.24-7.30 (m, 2H), 7.14-7.18 (m, 1 H), 5.45 (s, 2H), 4.20 (t, 2H, J= 8Hz), 3.50-3.56 (m, 2H), 3.45 (s, 3H), 1 .85-1.95 (m, 2H).
Example 390h 7-Benzyl-8-(3-fluorophenoxy)-l -(3-hydroxypropyl)-3-methyl- l H-purine- 2,6(3H,7H)-dione
Figure imgf000641_0001
The title compound was prepared from intermediate 83 using the method of example 389. White solid, 0.17 g, 88% yield: LCMS retention time = 3.282 min and 98% purity, LCMS MH+ = 425. lHNMR (CDCI3) δ 7.43-7.47 (m, 2H), 7.30-7.41 (m, 4H), 7.04-7.08 (m, 2H), 6.96-7.02 (m, IH), 5.45 (s, 2H), 4.20 (t, 2H, J = 8Hz), 3.50-3.55 (m, 2H), 3.45 (s, 3H), 1.87-1.94 (m, 2H).
Example 390i 7-BenzyI-8-(4-chloro-3-fluorophenoxy)-l -(3-hydroxypropyl)-3-methyl-lH- purine-2,6(3H,7H)-dione
Figure imgf000641_0002
The title compound was prepared from intermediate 83 using the method of example 389. White solid, 0.25 g, 56% yield: LCMS retention time = 3.860 min and 99% purity, LCMS MH+ = 459. 'HNMR (CDCI3) δ 7.15-7.48 (m, 8H), 5.46 (s, 2H), 4.20 (t, 2H, J = 8 HZ), 3.50-3.61 (m, 3H), 3.43 (s, 3H), 1.87 -1.94 (m, 2H).
Example 390j 7-Benzyl-l -(3-hydroxypropyl)-3-methyl-8-phenoxy- lH-purine-2,6(3H,7H)-dione
Figure imgf000642_0001
The title compound was prepared from intermediate 83 using the method of example 389. White solid, 0.18 g, 46% yield: LCMS retention time = 3.435 min and 99% purity, LCMS MH+ = 407. 'HNMR (CDC13) δ 7.15-7.48 (m, 10Η), 5.46 (s, 2Η), 4.20 (t, 2Η, J = 8 Hz), 3.50-3.61 (m, 3H), 3.43 (s, 3H), 1.87-1.94 (m, 2H).
Example 390k 7-Benzyl-8-(3-difluoromethoxy)phenoxy)-l-(3-hydroxypropyl)-3-methyl- lH-purine-2,6(3H,7H)-dione
Figure imgf000642_0002
The title compound was prepared from intermediate 83 using the method of example 389. White solid, 0.18 g, 39% yield: LCMS retention time = 3.581 min and 97% purity, LCMS MH+ = 473. 'HNMR (CDCI3) 6 7.31-7.47 (m, 6Η), 7.10-7.14 (m, 2Η), 7.02-7.06 (m, 2Η), 6.34-6.72 (t, 1Η, J = 18Hz), 5.46 (s, 2H), 4.20 (t, 2H, J = 2Hz). 3.51-3.55 (m, 2H), 3.45 (s, 3H), 1.87-1.94 (m, 2H)
Example 3901 7-(4-Chlorobenzyl)-8-(3-difluoromethoxy)phenoxy)-l-(3-hydroxypropyl)-3- methyl-lH-purine-2,6(3H,7H)-dione
Figure imgf000642_0003
White solid, 0.14 g, 27% yield: LCMS retention time = 3.820 min and 99% purity, LCMS ΜΗ4 507. 'HNMR (CDCI3) δ 7.44 (s, 1 Η), 7.36 (dd, 4Η, J = 12 Hz and 32 Hz), 7.1 1 -7.14 (m, 2H), 7.04-7.07 (m, 1H), 6.54 (t, 1H, J = 72 Hz), 5.42 (s, 2H), 4.20 (t, 2H, J = 2 Hz), 3.48-3.56 (m, 2H), 3.45 (s, 3H), 1 .87-1.94 (m, 2H).
Example 390m 7-(4-Chlorobenzyl)-l -(3-hydroxypropyl)-3-methyl-8-phenoxy- lH- urine-2,6(3H,7H)-dione
Figure imgf000643_0001
White solid, 0.24 g, 59% yield: LCMS retention time = 3.471 min and 99% purity, LCMS MH+ = 441. 'HNMR (CDC13) δ 7.41 -7.45 (m, 4H), 7.24-7.33 (m, 4H), 5.42 (s, 3H), 4.20 (t, 2H, J = 8 Hz), 3.51 -3.54 (m, 2H), 3.43 (s, 3H), 1.87-1.94 (m, 2H).
Example 390n 3-((7-Benzyl-l-(3-hydroxypropyl)-3-methyl-2,6-dioxo-2,3,6,7-tetrahydro- -purin-8-yl)oxy)benzonitrile
Figure imgf000643_0002
The title compound was prepared from intermediate 83 using the method of example 389. White solid, 0.32 g, 74% yield: LCMS retention time = 3.292 min and 99% purity, LCMS ΜΗ'" = 466. 'HNMR (CDCI3) 6 7.52-7.65 (m, 4Η), 7.36 (dd, 4Η, J = 8 Hz and 24 Hz), 5.43 (s, 2H), 4.20 (t, 2H, J= 8 Hz), 3.51 -3.56 (m, 2H), 3.45 (s, 3H), 1.88-1.93 (m, 2H).
Example 390o 3-((7-Benzyl-l -(3-hydroxypropyl)-3-methyl-2,6-dioxo-2,3,6,7-tetrahydro- 1 H-purin-8 -yl)oxy)benzonitri 1 e
Figure imgf000644_0001
The title compound was prepared from intermediate 83 using the method of example 389. White solid, 0.24 g, 74%: LCMS retention time = 3.581 min and 99% purity, LCMS MH+ = 432. Ή NMR (CDC ) δ 7.54 (s, 3H), 7.46 (d, 3H, J = 8 Hz), 7.30-7.40 (m, 3H), 5.48 (s, 2H), 4.21 (t, 2H, J = 8 Hz), 3.50-3.56 (m, 2H), 3.45 (s, 3H), 1.88-1.96 (m, 2H)
Example 390p 7-(4-Chlorobenzyl)-l-(3-hydroxypropyl)-8-(4-isopropylphenoxy)-3- meth l-lH-purine-2,6(3H,7H)-dione
Figure imgf000644_0002
White solid, 0.33 g, 88% yield: LCMS retention time = 4.160 min and 99% purity, LCMS MH+ = 483. 'HNMR (CDCI3) δ 7.38 (dd, 4H, J = 8 Hz and 44 Hz), 7.22 (dd, 4H, J = 8 Hz and 40 Hz), 5.41 (s, 2H), 4.20 (t, 2H, J = 8 Hz), 3.50-3.59 (m, 2H), 3.45 (s, 3H), 2.91-2.98 (m, 2H), 1.85-1.95 (m, 2H), 1.28 (d, 2H, J = 2 Hz). Example 390q 7-Benzyl-l -(3-hydroxypropyl)-8-(4-isopropylphenoxy)-3-methyl-lH- purine-2,6(3H,7H)-dione
Figure imgf000644_0003
The title compound was prepared from intermediate 83 using the method of example 389. White solid, 0.33 g, 91% yield: LCMS retention time = 3.894 min and 99% purity, LCMS MH+ = 449. •HNMR (CDCI3) δ 7.48 (d, 2H, J = 8 Hz), 7.31 -7.37 (m, 3H), 7.21 (dd, 4H, J = 8 Hz and 32 Hz), 5.45 (s, 2H), 4.20 (t, 2H, J = 8 Hz), 3.62 (brd s, 1 H), 3.50-3.56 (m, 2H), 3.44 (s, 3H), 2.90- 2.93 ( m, 1H), 1.91 (m, 2H), 1.27 (d, 6H, J = 2 Hz).
Example 390r7-Benzyl-l-(3-hydroxypropyl)-3-methyl-8-(3-(trifluoromethyl)phenoxy)-l H- purine-2,6(3 ,7H)-dione
Figure imgf000645_0001
The title compound was prepared from intermediate 83 using the method of example 389. White solid, 0.20 g, 79% yield: LCMS retention time = 3.600 min and 99% purity, LCMS MH+ = 475. 'H NMR (CDC13) 6 7.34-7.59 (m, 8H), 5.48 (s, 2H), 4.21 (t, 2H, J = 8 Hz), 3.49-3.55 (m, 2H), 3.45 (s, 3H), 1.88-1.95 (m, 2H)
Example 391 7-Benzyl)-8-(4-fluoro-3-(trifluoromethyl)phenoxy)-l-(3-hydroxypropyl)- -methyl-lH-purine-2,6(3H,7H)-dione
Figure imgf000645_0002
Step 1 7-Benzyl-8-bromo-l-(3-((tert-butyldimethylsilyl)oxy)propyl)-3-methyl-lH-purine- 2,6(3H57H)-dione (0.50 g , 0.985 mmol, intermediate 83), potassium carbonate (0.272 g, 1.97 mmol) and 4-fluoro-3-(trifluoromethyl)phenol (0.19 g, 1.03 mmol) were combined in DMF (5 mL). The reaction was heated at 90°C. After 6 h the reaction was cooled, diluted with water (100 mL) and extracted with ethyl acetate (3 x 75 mL). The combined extracts were dried with magnesium sulfate, filtered and the solvent was removed under reduced pressure to yield a white solid. This solid was purified using a 25 g silica column eluted with 20% ethyl acetate / hexanes to give 7-benzyl-l -(3-((tert-butyldimethylsilyl)oxy)propyl)-8-(4-fluoro-3- (trifluoromethyl)phenoxy)-3-methyl-lH-purine-2,6(3H,7H)-dione (0.54 g, 90% yield) as a white solid. LCMS retention time = 5.342 min and 97% purity, LCMS MH+ = 607. Step 2 7-Benzyl-l -(3-((/er -butyldimethylsilyl)oxy)propyl)-8-(4-fluoro-3- (trifluoromethyl)phenoxy)-3 -methyl- lH-purine-2,6(3H,7/J)-dione (0.44 g, 0.686 mmol) was dissolved in ethanol (15mL) and DCM (1 mL) then 6 N aqueous HCl (2.0 mL) were added. The clear solution was stirred at room temperature. After 1 h reaction was diluted with water (100 mL) and extracted with DCM (3 x 75 ml). The combined extracts were dried with magnesium sulfate, filtered and the solvent was removed under reduced pressure to yield a white solid. Solid was slurried in diethyl ether (6 mL) and filtered to give 7-benzyl)-8-(4-fluoro-3- (trifluoromethyl)phenoxy)-l -(3 -hydroxypropyl)-3 -methyl- lH-purine-2,6(3H,7H)-dione (0.35 g, 79% yield) as a white solid. LCMS retention time = 3.641 min and 99% purity, LCMS MH+ = 493. 'HNMR (CDC13) 5 7.23-7.52 (m, 8H), 5.47 (s, 2H), 4.20 (t, 2H, J■= 8 Hz), 3.47-3.54 (m, 2H), 3.43 (s, 3H), 1.88-1.92 (m, 2H).
Example 392 7-(4-Chlorobenzyl)-8-(4-fluoro-3-(trifluoromethyl)phenoxy)-l -(3- hydroxypropyl)-3 -methyl- lH-purine-2,6(3H,7H)-dione
Figure imgf000646_0001
The title compound was prepared using the two step method of example 391 . White solid, 0.33 g, 95% yield: LCMS retention time = 3.870 min and 99% purity, LCMS MH+ = 527. 'HNMR (CDCI3) δ 7.25-7.56 (m, 7Η), 5.43 (s, 2Η), 4.20 (t, 2Η, J = 8 Hz), 3.55-3.51 (m, 2H), 3.45 (t, 2H, J = 8 Hz), 3.43 (s, 3H), 1.89-1.92 (m, 2H).
Example 393 8-(4-Chloro-3-fluorophenoxy)-7-(4-chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl- lH-purine-2,6(3H,7H)-dione
Figure imgf000647_0001
The title compound was prepared using the two step method of example 391 except the crude product was purified by slurrying in ether and filtration. White solid, 0.33 g, 95% yield: LCMS retention time = 3.840 min and 99% purity, LCMS MH+ = 493. 'HNMR (CDC13) δ 7.45 (t, 1H, J = 2 Hz), 7.36 (dd, 4H, J = 8 Hz and 24 Hz), 7.20 (dd, 1H, J = 4 Hz and 12 Hz), 7.04 (d, 1 H, J = 8 Hz), 5.41 (s, 2H), 4.20 (t, 2H, J= 8 Hz), 3.51 - 3.55 (m, 2H), 3.43-3.47 (m, 5H), 1.89-1 .92 (m, 2H)
Example 394 3-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-(trifluoromethoxy)phenoxy)- 2 3,6,7-tetrahydro- lH-purin-1 -yl)propanol
Figure imgf000647_0002
7-(4-Chlorobenzyl)-l-(3-hydroxypropyl)-3-methyl-8-(3-(trifluoromethoxy)phenoxy)-l H-purine- 2,6(3H,7H)-dione (2.0 g , 3.81 mmol, example 356) and pyridinium chlorochromate (1.64 g, 7.62 mmol) were dissolved in DCM (40 mL). The orange solution was stirred at room
temperature for 15 h. Solvent was decanted away from salts, absorbed onto silica gel and purified using a 40 g silica gel flash column eluted with 40% ethyl acetate / hexanes to give 3-(7- (4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-(trifluoromethoxy)phenoxy)-2,3,6,7-tetrahydro-lH- purin-l-yl)propanol (1.25 g , 63% yield) as a white solid. 'HNMR (CDCI3) δ 9.84 (s, 1H), 7.12-7.47 (m, 8H), 5.40 (s, 2H), 4.38 (t, 2H, J= 8 Hz), 3.45 (s, 3H), 2.75-2.83 (m, 2H).
Example 395 7-Benzyl-8-ethoxy-l -(3-hydroxypropyl)-3-methyl-lH-purine-2,6(3H,7H)-dione
Figure imgf000648_0001
Step 1 Sodium metal (0.1 l g, 4.93 mm'ol) was dissolved in ethanol (10 mL) and 7-benzyl-8- bromo-l -(3-((tert-butyldimethylsilyl)oxy)propyl)-3-methyl-l H-purine-2,6(3H,7H)-dione (0.50 g, 0.985 mmol, intermediate 83) was added. The clear solution was stirred at room temperature 15 h. The reaction was diluted with water (100 mL) and extracted with ethyl acetate (3 x 75 mL). The combined extracts were dried with magnesium sulfate, filtered and the solvent was removed under reduced pressure to give 7-benzyl-l -(3-((tert-butyldimethylsilyl)oxy)propyl)-3-ethoxy-3- methyl-lH-purine-2,6(3H,7H)-dione (0.47 g, 100% yield) as a white solid. LCMS retention time = 5.074 min and 97% purity, LCMS MH+ = 473.
Step 2 7-Benzyl-l-(3-((tert-butyldimethylsilyl)oxy)propyl)-3-ethoxy-3-methyl-lH-purine- 2,6(3H,7H)-dione (0.47 g, 0.985 mmol) was dissolved in ethanol (8 mL) and 6 N HC1 (2.0 mL) was added. The clear solution was stirred at room temperature. After 1 h the reaction was diluted with water (75 mL) and extracted with DCM (3 x 50 ml). The combined extracts were dried with magnesium sulfate, filtered and the solvent was removed under reduced pressure to yield a white solid. Solid was slurried in hexanes (5 mL) and filtered to give 7-benzyl-8-ethoxy-l -(3- hydroxypropyl)-3-methyl-lH-purine-2,6(3H,7H)-dione (0.26 g, 73% yield) as a white solid. LCMS retention time = 2.848 min and 97% purity, LCMS MH+ = 359. ]HNMR (CDC13) δ 7.26-7.40 (m, 5H), 5.26 (s, 2H), 4.53-4.59 (m, 2H), 4.17 (t, 2H, J = 8Hz), 3.49-3.51 (m, 5H), 1.85-1.91 (m, 2H), 1 .45 (t, 3H, J = 8 Hz).
Exam e 396 7-Benzyl- 1 -(3 -hydroxypropyl)-3 -methyl- 8-propoxy- 1 /-r-purine-2,6(3H,7H)-dione
Figure imgf000648_0002
The title product was prepared using the two step method of example 395. White solid, 0.32 g, 87% yield: LCMS retention time = 3.1 19 min and 99% purity, LC S MPT = 373.
Example 397 tert-Butyl 3-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3- '
trifluoromethoxy)phenoxy)-2,3,6,7-tetrahydro-l H-purin-l -yl)propanoate
Figure imgf000649_0001
7-(4-Chlorobenzyl)-3-methyl-8-(3-(trifluoromethoxy)-lH-purine-2,6(3H,7H)-dione (0.50 g, 1 .07 mmol, intermediate 9), tert-butyl 3-bromopropanoate (0.22 mL, 1.29 mmol) and potassium carbonate (0.22 g, 1.61 mmol) were combined in DMF (5 mL) and heated at 100°C for 24 h. The reaction was cooled, diluted with water (100 mL) and extracted with ethyl acetate (3 x 75 mL). The combined extracts were washed with IN lithium chloride (2 x 50 mL), dried with magnesium sulfate, filtered and the solvent was removed under reduced pressure to leave a white solid. This solid was purified using a 25 g silica gel flash column eluted with 10% ethyl acetate / hexanes to give tert-butyl 3-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3- (trifluoromethoxy)phenoxy)-2,3,6,7-tetrahydro-lH-purin-l -yl)propanoate (0.26 g, 41 % yield) as a white solid. LCMS retention time = 4.841 min and 99% purity, LCMS MH+ = 595. 'HNMR (CDC13) δ 7.12-7.46 (m, 8H), 5.41 (s, 2H), 4.29 (t, 2H, J = 8 Hz), 3.53 (s, 3H), 2.61 (t, 2H, J = 8 Hz), 1.43 (s, 9H). Example 398 3-(7-(4-Chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-(trifluoromethoxy)phenoxy)- 2 3,6,7-tetrahydro- lH-purin-l-yl)propanoic acid
Figure imgf000649_0002
tert-Butyl 3-(7-(4-chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-(trifluoromethoxy)phenoxy)-2,3,6,7- tetrahydro-l H-purin-l -yl)propanoate (0.23 g, 0.383 mmol, Example 397) was dissolved in DCM (4 mL) and TFA (0.57 mL, 7.66 mmol) was added. The reaction was stirred at room temperature for 1 h; then the solvent was removed under reduced pressure to leave a golden oil (0.21 g). The oil was purified using a 12 g silica gel column eluted with 5% methanol / DCM to give 3-(7-(4- chlorobenzyl)-3-methyl-2,6-dioxo-8-(3-(trifluoromethoxy)phenoxy)-2,3,6,7-tetrahydro-lH- purin-l -yl)propanoic acid (0.13 g, 63% yield) as an white solid. LCMS retention time = 3.903 min and 99% purity, LCMS MH+ = 539. 'H NMR (CDC13) δ 7.14-7.47 (m, 8H), 5.41 (s, 2H), 4.35 (t, 2H, J = 8 HZ), 3.44 (s, 3H), 2.77 (t, 2H, J = 8 HZ).
Example 399 8-(3-Chlorophenoxy)-l-(3-hydroxypropyl)-7-isopentyl-3-methyl— lH-purine- -dione
Figure imgf000650_0001
Step 1 8-Bromo-l -(3-((ter/-butyldimethylsilyl)oxy)propyl)-7-isopentyl-3-methyl-lH-purine- 2,6(3H,7H)-dione (0.50 g, 1.03 mmol, intermediate 82), 3-chlorophenol (0.14 g, 1.08 mmol), potassium carbonate (0.29 g, 2.06 mmol) were combined in DMF (5 mL) and heated at 90°C for 15 h. The reaction was cooled, diluted with water (100 mL) and extracted with ethyl acetate (3 x 75 mL). The combined extracts were washed with 1 N LiCl (2 x 75 mL), dried with magnesium sulfate, filtered and evaporated under reduced pressure to give l-(3-((tert- butyldimethylsilyl)oxy)propyl-8-(3-chlorophenoxy)-7-isopentyl-3-methyl-lH-purine- 2,6(3H,7H)-dione (0.55 g, 100 % yield) as a yellow oil. LCMS retention time = 6.238 min and 85% purity, LCMS MH+ = 535.
Step 2 l -(3-((tert-Butyldimethylsilyl)oxy)propyl-8-(3-chlorophenoxy)-7-isopentyl-3-methyl-lH- purine-2,6(3H,7//)-dione (0.55 g, 1.03 mmol) was dissolved in ethanol (l o mL) and 6N HC1 (2 mL) was added and the reaction was stirred at room temperature for 1 h. The reaction was diluted with water (100 mL) and extracted with DCM (3 x 75 mL). The combined extracted were dried with magnesium sulfate, filtered and evaporated under reduced pressure to leave a golden oil. The oil was purified using a 24 g silica gel flash column eluted with 50% ethyl acetate / hexanes to give 8-(3-chlorophenoxy)-l -(3-hydroxypropyl)-7-isopentyl-3-methyl— l /-purine- 2,6(3H,7H)-dione (0.21 g, 49% yield) as a white solid. LCMS retention time = 3.859 min and 98% purity, LCMS MH+ - 421. 'HNMR (CDC13) δ 7.35-7.38 (m, 2H), 7.24-7.28 (m, 1H), 7.18-7.25 (m, 1H), 4.27 (t, 2H, J - 8 Hz), 4.20 (t, 2H, J = 8 Hz), 3.52-3.58 (m, 3H), 3.47 (s, 3H), 1.86-1.94 (m, 2H), 1.70-1.78 (m, 2H), .1.59-1.69 (m, 1 H), 0.98 (d, 6H, J = 8 Hz).
Example 400 1 -(3-Hydroxypropyl)-7-isopentyl-8-isopropoxy-3-methyl-lH-purine-2,6(3H,7H)- dione
Figure imgf000651_0001
Sodium (0.12 g, 5.15 mmol) was dissolved in isopropanol (8 mL) and 8-bromo-l-(3-((/ert- butyldimethylsilyl)oxy)propyl-7-isopentyl-3 -methyl- lH-purine-2,6(3H,7H)-dione (0.50 g, 1.03 mmol, intermediate 82) was added. The reaction was stirred at room temperature for 15 h. The reaction was treated with 6N HC1 to achieve pH = 1, and stirred at room temperature for 1 h. The reaction was diluted with water (100 mL) and extracted with DCM (3 x 75 mL). The combined extracts were dried with magnesium sulfate, filtered and evaporated under reduced pressure to leave a golden oil. The oil was purified using two 2000 micron preparative TLC plate eluted with 50% ethyl acetate / hexanes to give l -(3-hydroxypropyl)-7-isopentyl-8-isopropoxy3-methyl-lH- purine-2,6(3H,7H)-dione (0.27 g, 75 yield) as a white solid. LCMS retention time = 3.325 min and 97% purity, LCMS MH"1" = 353. HNMR (CDCI3) δ 5.20-5.30 (m, 1Η), 4.17 (t, 2Η, J = 8 Hz), 4.07 (t, 2H, J = 8 Hz), 3.73 (t, 1H, J = 8 Hz), 3.45-3.54 (m, 5H), 1.85-1.93 (m, 2H), 1.58- 1.66 (m, 2H), 1.48-1.57 (m, 1H), 1.42 (d, 6H, J = 8 Hz), 0.94 (d, 6H, J = 8 Hz).
Example 401 8-Ethoxy-l-(3-hydroxypropyl)-7-isopentyl-3-methyl-lH-purine-2,6(3H,7H)- dione
Figure imgf000652_0001
The title compound was prepared using the method of example 400 as a white solid. LCMS retention time - 3.050 min and 96% purity, LCMS MH+ 339. JHNMR (CDC13) δ 4.53 (dd, 2H, J - 8 Hz and 16 Hz), 4.17 (t, 2H, J = 8 Hz), 4.08 (t, 2H, J = 8 Hz), 3.71 (t, 1H, J = 4 Hz), 3.47- 3.53 (m, 5H), 1.84-1.92 (m, 2H), 1.60-1.67 (m, 2H), 1.49-1.58 (m, 1H), 1.45 (t, 2H, J - 4 Hz), 0.94 (d, 6H, J = 4 Hz).
Example 402 1 -(3-Hydroxypropyl)-7-isopentyl-3-methyl-8-propoxy-lH-purine-2,6(3H,7H)- dione
Figure imgf000652_0002
The title compound was prepared using the method of example 400 as a white solid. LCMS retention time = 3.383 min and 95% purity, LCMS MH+ = 352. '//NMR (CDC13) δ 4.42 (t, 2H, J = 8 Hz), 4.17 (t, 2H, J - 8 Hz), 4.09 (t, 2H, J = 8 Hz), 3.70 (t, 1H, J = 4 Hz), 3.45-3.56 (m, 5H), 1.78-1.94 (m, 4H), 1.51-1.72 (m, 3H), 1.04 (t, 3H, J = 8 Hz), 0.94 (d, 6H, J = 8 Hz).
Example 403 8-Butoxy l-(3-hydroxypropyl)-7-isopentyl-3-methyl-lH-purine-2,6(3H,7H)- dione
Figure imgf000652_0003
The title compound was prepared using the method of example 400 as a white solid. LCMS retention time = 3.681 min and 99% purity, LCMS MH+ = 367. ' /NMR (CDC13) δ 4.47 (t, 2H, J = 8 Hz), 4.16 (t, 2H, J = 8 Hz), 4.08 (t, 2H, J = 8 Hz), 3.70 (t, 1H, J = 8 Hz), 3.45-3.59 (m, 5H), 1.73- 1 .93 (m, 4H), 0.99 (t, 3H, J = 8 Hz), 0.94 (d, 6H, J = 8 Hz).
Example 404 7-Benzyl-8-(3,4-dichlorophenyl)- l -(3-hydroxypropyl)-3-methyl-l/ -purine- 2,6(3 /,7H)-dione
Figure imgf000653_0001
Step 1 7^Benzyl-8-bromo-l-(3-((tert-butyldimethylsilyl)oxy)propyl)-3-methyl-l H-purine- 2,6(3//, 7H)-dione (0.50 g , 0.985 mmol, intermediate 83), potassium carbonate (0.326g, 2.36 mmol), tetrakis(triphenylphosphine) palladium(O) (0.032 g, 0.028 mmol) and 3,4-dichlorophenyl boronic acid (0.206 g, 1.08 mmol) were combined in ethanol (12.0 mL), toluene (2.0 mL) and water (2.0 mL) in a sealed vial. The reaction was heated at 85°C for 18 h. The reaction was cooled and filtered through Celite. The filtrate was diluted with water (100 mL) and extracted with ethyl acetate (3 x 75 mL). The combined extracts were dried with magnesium sulfate, filtered and the solvent was removed under reduced pressure to yield a golden solid. The solid was purified using a 25 g silica gel CombiFlash column eluted with 20% ethyl acetate / hexanes to give 7-benzyl-l-(3-((tert-butyldimethylsilyl)oxy)propyl)-8-(3,4-dichlorophenyl)-3-methyl-l//- purine-2,6(3H,7H)-dione (0.49 g, 86% yield) as a golden oil. LCMS retention time = 5.489 min and 97% purity, LCMS MH+ = 573.
Step 2 7-Benzyl-l-(3-((tert-butyldimethylsilyl)oxy)propyl)-8-(3,4-dichlorophenyl)-3-methyl- lH-purine-2,6(3H,7 )-dione (0.49 g, 0.854 mmol) was dissolved in ethanol (15 mL) and 6 N HC1 (2.0 mL) was added. The clear solution was stirred at room temperature for 1 h. The reaction was diluted with water (100 mL) and extracted with DCM (3 x 75 ml). The combined extracts were dried with magnesium sulfate, filtered and the solvent was removed under reduced pressure to give a white solid. The solid was slurried in diethyl ether (5 mL) and filtered to give 7-benzyl-8-(3,4-dichlorophenyl)-l -(3-hydroxypiOpy])-3-methyl-lH-purine-2,6(3H,7H)-dione (0..28 g, 71 % yield) as a white solid. LCMS retention time = 3.567 min and 96% purity, LCMS MH+ = 459. 'HNMR (CDC13) δ 7.71 (s, 1 H), 7.53 (d, 1 H, J = 8 Hz), 7.38-7.41 (d, 1H, J = 4 Hz), 7.26-7.29 (m, 3H), 7.04 (d, 2H), 5.64 (s, 2H), 4.20 (t, 2H, J = 8 Hz), 3.65 (s, 3H), 3.47-3.54 (m, 2H), 3.30 (t, 2H, J = 8 Hz).
Example 405 8-(3-Chlorophenyl)-l-(3-hydroxypropyl)-7-isopentyl-3-methyl- lH-purine- -dione
Figure imgf000654_0001
The title compound was prepared from intermediate 82 using the method of example 404. Golden yellow solid, 0.31 g, 74% yield: LCMS retention time = 3.516 min and 85% purity, LCMS MH+ = 405. 'HNMR (CDC13) δ 7.65-7.70 (m, 1Η), 7.45-7.54 (m, 3Η), 4.33-4.39 (m, 2Η), 4.23 (t, 2Η, J = 4 Hz), 3.63 (s, 3H), 3.52-3.58 (m, 2H), 3.43-3.48 (m, 2H), 1.89-1 .96 (m, 2H), 1.69-1.76 (m, 2H), 1 .53-1.64 (m, 2H), 0.89 (d, 6H, J = 4 Hz).
Example 406 1 -(3-Hydroxypropyl)-7-isopentyl-3-methyl-8-(3-(trifluoromethyl)phenyl)-lH- purine-2,6(3H,7H)-dione
Figure imgf000654_0002
The title compound was prepared from intermediate 82 using the method of example 404. solid, 0.089 g, 40% yield: LCMS retention time = 3.598 min and 97% purity, LCMS ΜΗ
438. Example 407 l -(3-hydroxypropyl)-7-isopentyl-8-(2-isopropylphenyl)-3-methyl-lH-purine- -dione
Figure imgf000655_0001
The title compound was prepared from intermediate 82 using the method of example 404.
solid, 0.089 g, 40% yield: LCMS retention time = 3.749 min and 99% purity, LCMS MH+ 413.
Example 408 8-(3-Fluorophenoxy)-l-(3-hydroxypropyl)-7-isopentyl-3-methyl-lH-purine- -dione
Figure imgf000655_0002
8-Bromo-l -(3-((tert-butyldimethylsilyl)oxy)propyl-7-isopentyl-3-methyl-lH-purine-2,6(3H,7H)- dione (0.50 g, 1.03 mmol, intermediate 82), 3-fluorophenol (0.098 mL, 1.08 mmol) and potassium carbonate (0.29 g, 2.06 mmol) were combined in DMF (8 mL) and heated at 90°C for 15 h. The reaction was cooled to room temperature and acidified with 6N HC1 to pH = 1. The reaction was stirred for 1 h then diluted with water (100 mL) and extracted with ethyl acetate (3 x 75 mL). The combined extracts were washed with IN LiCl (2 x 100 mL), dried with magnesium sulfate, filtered and the solvent was removed under reduced pressure to yield a golden oil. The oil was purified using a 24 g silica gel flash column eluted with a gradient of 20% ethyl acetate / hexanes to 100%» ethyl acetate to give 8-(3-fiuorophenoxy)-l-(3- hydroxypropyl)-7-isopentyl-3-methyl-lH-purine-2,6(3H,7H)-dione (0.20 g, 48% yield) as a white solid. LCMS retention time = 3.524 min and 96% purity, LCMS MH+ = 405.
Example 409 l -(3-Hydroxypropyl)-7-isopentyl-3-methyl-8-(3-(trifluoromethyl)phenoxy)- lH- purine-2,6(37J,7H)-dione
Figure imgf000656_0001
The title compound was prepared using the method of example 408. White solid, 0.26 g, 55% yield: LCMS retention time = 3.838 min and 99% purity, LCMS MH+ = 455.
The following examples 410a through 410w were prepared using the method of example 404.
Example 410a 7-Benzyl-8-(4-chloro-3-(trifluoromethyl)phenyl)-l-(3-hydroxypropyl)-3-
Figure imgf000656_0002
White solid, LCMS retention time = 3.625 min and 97% purity, LCMS MH+ = 493. 'HNMR (CDC13) δ 7.91 (s, 1H), 7.63 (d, 2H, J = 8 Hz), 7.59 (d, 2H, J = 8 Hz), 7.30-7.35 (m, 2H), 7.01 (d, 2H, J = 8 Hz), 5.55 (s, 2H), 4.21 (t, 2H, J = 8 Hz), 3.66 (s, 3H), 3.47-3.55 (m, 2H), 3.28 (t, 2H, J = 8 Hz), 1.89-1 .93 (m, 2H).
7-Benzyl-8-(3-fluoi phenyl)-l -(3-hydroxypropyl)-3-methyl-l H-purine-
Figure imgf000656_0003
White solid, 0.051 g, 80% yield: LCMS retention time = 3.004 min and 99% purity, LCMS MH+ = 409. Exam le 410c7-Benzyl-l -(3-hydroxypropyl)-3-methyl-8-(m-tolyl)-lH-purine-2,6(3/ ,7H)-dione
Figure imgf000657_0001
White solid, 0.054 g, 85% yield: LCMS retention time = 3.133 min and 99% purity, LCMS MH+ = 405.
7-benzyl- l-(3-hydroxypropyl)-8-(3-methoxyphenyl)-3-methyl-lH-purine-
Figure imgf000657_0002
White solid, 0.0 1 g, 84% yield: LCMS retention time = 2.950 min and 99% purity, LCMS MH+ = 421.
Example 410e7-Benzyl-l-(3-hydroxypropyl)-3-methyl-8-(3-(trifluoromethoxy)phenyl)-l H- purine-2,6(3H,7 )-dione
Figure imgf000657_0003
White solid, 0.052g, 80% yield: LCMS retention time = 3.312 min and 99% purity, LCMS MH = 459.
Example 410f 7-Benzyl-8-(4-chlorophenyl)-l-(3-hydroxypropyl)-3-methyl-lH-purine- 2,6(3/J,7H)-dione
Figure imgf000658_0001
White solid, 0.050 g, 91% yield: . LCMS retention time = 3.227 min and 98% purity, LCMS MH+ = 425. Example 410g 7-Benzyl-8-(4-fluorophenyl)-l-(3-hydroxypropyl)-3-methyl-l H-purine- -dione
Figure imgf000658_0002
White solid, 0.052 g, 60% yield: LCMS retention time = 2.977 min and 99% purity, LCMS MH+ = 409.
Example 410h 7-Benzyl-l -(3-hydroxypropyl)-3-methyl-8-(p-tolyl)-lH-purine- -dione
Figure imgf000658_0003
Clear oil, 0.028 g, 55% yield: LCMS retention time = 3.122 min and 98% purity, LCMS MH+ = 405.
Example 410i 7-Benzyl-l -(3-hydroxypropyl)-8-(4-methoxyphenyl)-3-methyl-lH-purine- 2,6(3H,7H)-dione
Figure imgf000659_0001
White solid, 0.049 g, 91 % yield: LCMS retention time = 2.944 min and 96% purity, LCMS MH+ = 421.
Example 410j 7-Benzyl-l-(3-hydroxypropyl)-3-methyl-8-(4-(trifluoromethyl)phi
purine-2,6(3H,7H)-dione
Figure imgf000659_0002
White solid, 0.052g, 76% yield: LCMS retention time = 3.348 min and 99% purity, LCMS MH+ = 459.
Example 410k 7-Benzyl-8-(2-chlorophenyl)-l-(3-hydroxypropyl)-3-methyl-lH-purine- -dione
Figure imgf000659_0003
Clear oil, 0.057 g, 93% yield: LCMS retention time = 2.979 min and 99% purity, LCMS MH = 425.
Example 4101 7-Benzyl-8-(2-fluorophenyl)-l-(3-hydroxypropyl)-3-methyl-lH-purine- 2,6(3H,7H)-dione
Figure imgf000660_0001
Golden oil, 0.030 g, 91% yield: LCMS retention time = 2.858 min and 99% purity, LCMS MH+ = 409. Example 410m 7-Benzyl- 1 -(3 -hydroxypropyl)-3 -methyl-8-(o-tolyl)-lH-purine- 2 /J,7H)-dione
Figure imgf000660_0002
White solid, 0.027 g, 77% yield: LCMS retention time = 2.995 min and 98% purity, LCMS MH+ = 405.
Example 410n 7-Benzyl-l-(3-hydroxypropyl)-8-(2-methoxyphenyl)-3-methyl-lH-purine- -dione
Figure imgf000660_0003
White solid, 0.061 g, 86% yield: LCMS retention time = 2.831 min and 98% purity, LCMS MH+ = 421.
Example 410o 7-Benzyl- 1 -(3-hydroxypropyl)-3-methyl-8-(2-(trifluoromethyl)phenyl)- lH-purine-2,6(3/J,7H)-dione
Figure imgf000661_0001
White solid, 0.017 g, 57% yield: LCMS retention time = 3.055 min and 97% purity, LCMS MH+ = 459. Example 410p 7-Benzyl-l -(3-hydroxypropyl)-8-(4-(isopropylsulfonyl)phenyl)-3-methyl- -purine-2,6(3H,7H)-dione
Figure imgf000661_0002
Colorless oil, 0.057g, 88% yield: LCMS retention time = 2.886 min and 98% purity, LCMS MH+ = 498.
Example 410q 7-Benzyl-l-(3-hydroxypropyl)-8-(2-isopropylphenyl)-3-methyl-lH- purine-2,6(3H,7H)-dione
Figure imgf000661_0003
Colorless oil, 0.038g, 79% yield LCMS retention time - 3.520 min, LCMS MH+ = 433 and 99% pure.
Example 410r7-Benzyl-l-(3-hydroxypropyl)-8-(3-isopropylphenyl)-3-methyl-lH-purine- 2,6(3 /,7H)-dione
Figure imgf000662_0001
Colorless oil, 0.026 g, 81 % yield: LCMS retention time = 3.609 min and 98% purity, LCMS MH+ = 433. Example 410s 8-([l , -Biphenyl]-3-yl)-7-benzyl-l -(3-hydroxypropyl)-3-methyl- lH-purine- 2 6(3H,7H)-dione
Figure imgf000662_0002
Colorless oil, 0.045 g, 80% yield: LCMS retention time = 3.609 min, LCMS MH+ - 467 and 99% pure.
Example 410t 7-Benzyl-l-(3-hydroxypropyl)-3-methyl-8-(2-(trifluoromethoxy)phenyl)-l H- rine-2, 6(3/7, 7H)-dione
Figure imgf000662_0003
Colorless oil, 0.048 g, 83% yield: LCMS retention time = 3.231 min and 99% purity, LCMS MH+ = 475.
Example 410u 7-Benzyl-8-(3-(difluoromethoxy)phenyl)-l -(3-hydroxypropyl)-3-methyl- 1 H-purine-2,6(3H,7 /)-dione
Figure imgf000663_0001
Colorless oil, 0.048 g, 83%: LCMS retention time = 3.150 min and 99% purity, LCMS MH = 457.
Example 410v 7-Benzyl-8-(4-(difluoromethoxy)phenyl)-l -(3-hydroxypropyl)-3-methyl- -purine-2,6(3H,7H)-dione
Figure imgf000663_0002
Colorless oil,0.050 g, 78% yield: LCMS retention time = 3.161 min and 99% purity, LCMS MH+ = 457.
Example 410 7-Benzyl-8-(4-(difluoromethyl)phenyl)-l -(3-hydroxypropyl)-3-methyl- -purine-2,6(3H,7H)-dione
Figure imgf000663_0003
Colorless oil, 0.048 g, 75% yield: LCMS retention time = 3.098 min and 99% purity, LCMS yhexyl)-3;7-dimethyl-lH-purine-2,6(3H,7H)-dione
Figure imgf000663_0004
Pentoxifylline (1 .0 g, 3.59 mmol) was dissolved in methanol (20 mL) and DCM (3 mL) then cooled to 0°C. To the reaction was added sodium borohydride (0.41 g, 10.8 mmol) portionwise over 30 min. The reaction was stirred in the cold for 2 h; then it was evaporated to dryness under reduced pressure, diluted with water (100 mL) and extracted with ethyl acetate (3 x 75 mL). The combined extracts were dried with magnesium sulfate, filtered and the solvent was removed under reduced pressure to give l -(5-hydroxyhexyl)-3,7-dimethyl-l H-purine-2,6(3H,7H)-dione (0.72 g, 71 % yield) as a white solid. LCMS retention time = 1.849 min and 98% purity, LCMS MH+ 281. 'HNMR (DMSO-d6) δ 8.00 (s, 1H), 4.31 (d, 1H, J = 4 Hz), 3.88 (s, 3H), 3.84 (t, 2H, J = 8 Hz), 3.51 -3.59 (m, 1H), 1.45-1.56 (m, 2H), 1.25-1.37 (m, 4H), 1.02 (d, 3H, J = 4 Hz).
Example 434. Anti-deprcssive Effect of Compounds of the Invention in the Forced Swim Test Model of Depression
The forced swim test (FST) is one of the most widely used in vivo models of
antidepressant activity. The immobility observed in this test represents a depressed stress- evoked behavior that can be reversed by all major classes of antidepressants. (Porsolt et al., Arch Int Pharmacodvn Ther. 229(2):327-36, 1977; Porsolt et al., Nature 266: 730 - 732, 1977) (Lucki, et al., Psvchopharmacology (BerO. 155(3):315-22, 2001). Briefly, groups of male CD-I mice (7 weeks old, Charles River Labs) were dosed orally with 0.5% methylcellulose (MC) or compounds of the invention at various doses. The positive control, imipramine, was
administered via intraperitoneal injection (IP) at 20mg/kg. Immediately following dosing, mice were returned to their home cage. At 60 minutes post methylcellulose administration or administration of compounds of the invention, and 45 minutes post imipramine administration, mice were placed individually into Polycarbonate cylinders (22.2 cm in diameter, 25.3 cm high; Fisher Scientific, Pittsburgh, PA) filled with 15 cm of water, maintained at a temperature of 24+1 °C. In the examples below, immobility or swimming behavior was recorded for 6 minutes after exposure to compounds of the invention. After the 6-min swim session, mice were towel- dried and returned to the home cage.
Digital video output of swim sessions was analyzed by a computer running Noldus software (Ethovision XT Version 8 Video Tracking, Noldus Information Technology,
Wageningen, Netherlands). Individual mouse immobility time was acquired and the last 4 minutes of the 6-min swim session analyzed. The software traced each animal's displacement in a defined arena; images of these displacements were acquired five times per second. In multiple examples, compounds of the invention were able to decrease the time the mice spent immobile. As immobility correlates with a depressed stress-evoked behavior, the compounds ability to reverse immobility demonstrates an antidepressant treatment effect.
Compound 31
Mice were administered 10 mL/kg 0.5% methylcellulose (MC) orally, or 0.1 , 0.3 or 1 mg/kg Compound 31 orally, 60 minutes prior to chamber placement. Imipramine. was administered by intraperitoneal injection at 20 mg/kg 45 minutes prior to testing.
Figure imgf000665_0001
Compound 260
Mice were administered 10 mL/kg 0.5% methylcellulose (MC) orally, or 0.3, 1 or 3 mg/kg Compound 260, 60 minutes prior to chamber placement. The positive control, imipramine, was administered by intraperitoneal injection at 20 mg/kg 45 minutes prior to testing.
Figure imgf000665_0002
Compound 587
Mice received lOmL/kg of orally administered 0.5% methyl cellulose (MC), or
Compound 587 at doses of 3, 10, 30 or 100 mg/kg, one hour before testing; or they received mg/kg imipramine, by intraperitoneal injection 45 minutes before testing.
Figure imgf000665_0003
immobile
MC l OmL/kg oral 59.1 24
Compound 587 3 mg/kg oral 31.0 12
Compound 587 10 mg/kg oral 38.9 24
Compound 587 30mg/kg oral 33.5 24
Compound 587 1 OOmg/kg oral 20.3 12 imipramine 20mg/kg IP 26.4 24
Compound 458
Mice received l OmL/kg of orally administered 0.5% methyl cellulose (MC) or Compound 458 at 3, 10 or 30 mg/kg one hour before testing; or they received 20 mg/kg imipramine, by intraperitoneal injection 45 minutes before testing. Compound 458 decreased the time mice spent immobile demonstrating an anxiolytic effect.
Figure imgf000666_0001
Compound 556
Mice received l OmL/kg of orally administered 0.5% methyl cellulose (MC) or Compound 556 at 0.3, 1 , or 3 mg/kg one hour before testing; or they received 20 mg/kg imipramine, by intraperitoneal injection 45 minutes before testing.
Figure imgf000666_0002
Example 435. Anti-depressive Effect of Compound 260 in the Tail Suspension Model of Depression The Tail Suspension Test as described by Steru et al. (L. Stem, R. Chermat, B. Thierry, and P. Simon, Psvchopharmacology 85(3): 367-370, 1985) has become one of the most widely used animal models of depression along with the forced swim test. (Cryan JF, Mombereau C, Vassout A., Neurosci. Biobehav. Rev. 29(4-5): 571 -625, 2005).
Briefly, NMRI mice were treated with 0.5% Methylcellulose (MC) (4000 CPS) orally or Compound 260 at oral doses of 0.3, 1 , 3 and 10 mg/kg one hour prior to testing, or desipramine by intraperitoneal injection of 30 mg/kg, 30 minutes prior to testing. After which, mice were suspended by their tail, upside down, for a total of six minutes. A period of one minute was allowed to lapse allowing the mice to acclimate themselves and then recorded the amount of time mice attempted to upright themselves within five minutes. Historically untreated mice become immobile, a reaction to the unavoidable and inescapable stress of their situation. This is thought to model a despair-type behavior seen in depressed patients. Clinically effective anti-depressants historically reverse such immobility. (Steru) Compound 260 at all dose levels was also able to reverse immobility as shown in FIG. 1.
Example 436. Anxiolytic Effect of Compounds Using the Marble Burying Test Model
Rodents have been shown to use bedding material to bury noxious materials, however they also bury harmless objects such as rat chow pellets and glass marbles. This behavior is prevented by administration of serotonin uptake inhibitors. It has been proposed that marble burying is a compulsive behavior as it can be reduced by serotonin uptake inhibitors which have been found to be effective in human obsessive-compulsive disorder (Njung'e, K., et al.
Pharmacol. Biochem. Behav. 38(Π: 63-7, 1991).
In the examples below, groups of female C57BL/6 mice (7 weeks old, Charles River Laboratories) were dosed orally with 0.5% methylcellulose (MC) or compounds of the invention at various doses. The positive control, 10 mg/kg zimelidine, was administered by intraperitoneal injection (IP) 45 minutes prior to testing. Immediately following dosing, mice were returned to their home cage. At 60 minutes post vehicle or compound administration, or 45 minutes post zimelidine administration, mice were placed into clean rat cages (45cm X 24.5cm X 13cm) filled to 5cm with BetaChip bedding and containing 20 black glass marbles evenly spaced over the BetaChip bedding for a 30 minute test session. Upon completion, the animals were carefully removed and each cage was photographed for later review. Photos were manually scored for number of marbles >25% visible by a scorer blinded to treatment. The examples below show that compounds of the invention were able to reduce the burying behavior in the mice, where treated mice left more unburied marbles than the control groups.
Compound 31
Mice were administered 10 mL/kg 0.5% methylcellulose (MC) orally, or 0.1, 0.3 or 1 mg/kg of Compound 31 orally, 60 minutes prior to testing. The positive control, 10 mg/kg zimelidine, was administered by intraperitoneal injection 45 minutes prior to testing. At 0.3 & 1 mg/kg, Compound 31 was able to decrease the number of buried marbles compared to the oral control.
Figure imgf000668_0001
.
Compound 260
Mice were administered 10 mL/kg 0.5% methylcellulose (MC) orally, or 1 , 3 or 10 mg/kg of Compound 260 orally, 60 minutes prior to testing. The positive control, 10 mg/kg zimelidine, was administered by intraperitoneal injection 45 minutes prior to testing.
Figure imgf000668_0002
Compound 587
Mice were administered 10 mL/kg 0.5% methylcellulose (MC) orally, or 3, 10 or 30 mg/kg of Compound 587 orally, 60 minutes prior to testing. The positive control, 10 mg/kg zimelidine, was administered by intraperitoneal injection 45 minutes prior to testing. Unburicd Marbles
Mice
Dose (n)
(n)
Average
MC l OmL/kg 12.7 10
Compound 587 3mg/kg 14.2 10
Compound 587 l Omg/kg 15.5 10
Compound 587 30mg/kg 17.1 10
zimelidinc 1 Omg/kg 16.5 10
Compound 458
Mice were administered 10 mL/kg 0.5% methylcellulose (MC) orally, or 3, 10 30 or 60 mg/kg of Compound 458 orally, 15 minutes prior to testing. The positive control, 10 mg/kg zimelidine, was administered by intraperitoneal injection 45 minutes prior to testing.
Figure imgf000669_0001
Example 437. Anxiolytic Effect of Compounds Using the Vogel Conflict Test of Anxiety
For over 30 years, the Vogel Conflict Test has been used as an in vivo animal model to measure the ability of drugs to overcome anxiety in rodents. (Millan MJ, Brocco M, Eur J Pharmacol. 463(1-3): 67-96, 2003). Briefly, water is withheld from animals for a certain period of time such as 48 hours. Then after the time period lapses, animal are given free access to water, but they receive mild electrical shocks from the water source at intermittent periods occurring after a defined number of licks such as on shock every 20 licks. Untreated rats develop an avoidance behavior manifested by a refusal to drink. This avoidance-type behavior or internal-conflict behavior in the rats is similar to the avoidance conflicts in man. (Cryan JF, Sweeney FF, British Journal of Pharmacology, 201 1 ; 164: 1 129-1 161).
However, Riccio et al (Riccio A. et al., Cell 137: 761-772, 2009), indicated that these types of conflict-avoidance tests may not be appropriate models for all mechanisms of anxiolytic activity. Riccio showed that TrpC5 Knock-out mice, in a similar conflict avoidance test to the Vogel test above, the novelty suppressed feeding test using food rather than water, did not behave differently than the wild-type mice in the conflict-avoidance model, yet knock-out mice showed behavioral differences in other anxiety behavior models.
Compound 260 was tested for activity in the Vogel Conflict Test Model. Briefly, three month old male Wistar-Kyoto rats, weighing 225-250g received 0.5% Methylcellulose (4000 CPS), or Compound 260 at doses of 0.3, 1, 3 and 10 mg/kg orally 1 hour prior to testing, or Midazolam 1 mg/kg by intraperitoneal injection, 30 minutes prior to testing. Results are shown in FIG. 2A and FIG. 2B. As expected, Compound 260, a TrpC5 antagonist, did not demonstrate positive activity when compared to the control in this conflict avoidance model.
Example 438. Anxiolytic Effects of Compounds of the Invention
The elevated plus maze test is an in vivo model to assess anxiolytic activity. The test is based upon the observation by Montgomery (Montgomery, KC, T Comp Physiol Psychol. 48: 254-260, 1958) that rodents exposed to an open or closed alley spent more time in the closed alley, characterized as an approach-avoidance conflict. Based upon this, Handley and Mithani (Handley SL, Mithani S., Naunyn-Schmiedeberg's Arch Pharmacol. 327: 1 -5, 1984.) devised an adaptation using an elevated maze in the shape of a plus with 2 closed and 2 open arms. In the elevated plus maze test, a motivational conflict is established in which the animals innate tendency to explore a novel environment is opposed by their fear of open spaces. Anxiolytic manipulations reduce the drive to avoid the open spaces, thus increasing entry into, and time spent on the open arms. Furthermore, Riccio et all demonstrated through the use of C57/BL6 TrpC5 knock-out mice that activation of TrpC5 leads to an increased fear response in the elevated plus maze (Riccio A el al., Cell 137: 761-772, 2009) but not the conflict-avoidance novelty suppressed feeding test.
In the following examples, compounds of the invention were tested to determine whether these compounds could reproduce the results seen with the knock-out mice when tested in C57/BL6 wild-type mice in the elevated plus maze of anxiety. An inactivation of TrpC5 in the wild-type mice with compounds of the invention would correlate to an anxiolytic treatment benefit. Briefly, using the model described in Riccio, C57/BL6 wild-type mice were dosed with vehicle, positive control, or compounds of the invention. Under reverse light, groups of male C57/BL6 mice (10 weeks old, Jackson Labs) were dosed orally with 0.5% methyl cellulose or compounds of the invention. The positive control, 1 .5 mg/kg diazepam, was administered via intraperitoneal injection 30 minutes prior to testing. Immediately following dosing, mice were returned to their home cage. At 60 minutes post vehicle or compound administration, and 30 5 minutes post diazepam administration, mice were placed onto the elevated plus maze, (0547M, Mouse Elevated Plus Maze, Columbus Instruments, Columbus, OH) one at a time, and their session recorded for 5 minutes. A distal open arm entry occurred when mice travelled to the farthest point of the open arm in the plus-maze configuration and an open arm result occurred when mice progress into the open arm portion of the plus maze configuration but failed to 10 progress to the farthest point of such arm. Videos were manually scored for number of open arm entries by a scorer blinded to treatment. In some studies, mice falling off the maze were included. As an additional way to analyze the data, fallen mice were omitted from the analysis of some examples as noted. As both methods are employed in reporting the elevated plus maze assay, where available, both results are included in the table below. The number of open arm
15 entries was recorded for 5 minutes. Results of the studies are summarized in the table below.
Figure imgf000671_0001
Compound
1 oral 1 1 21.3 2.6 10.0 8 1 8.0 1.3 9.6 31
5 mL
0.5% MC oral 12 12.1 1.1 4.9
(volume)
Compound
1 oral 12 26.7 3.3 1 1.3
31
c Compound
30 oral 12 14.5 1.3 6.0
77
10% 5 mL
IP 12 19.7 2.1 7.3
HPbCD (volume)
diazepam 1.5 IP 12 32.4 3.8 13.1
5 mL
0.5% MC oral 8 17.8 2.4 7.1 8 17.8 2.4 7.1
(volume)
Compound
0.1 oral 8 25.3 3.9 9.8 7 26.0 3.9 10.1 31
D Compound
0.3 oral 8 34.3 6.5 10.0 5 39.0 8.4 1 1.4 31
Compound
1 oral 8 30.2 5.5 1 1.4 5 34.3 6.4 1 1.4 31
diazepam 1.5 IP 8 33.5 6.8 12.9 6 33.9 6.5 12.2
5 mL
0.5% MC oral 8 5.7 0.3 2.9
(volume)
Compound
1 oral 9 23.7 3.0 10.7
31
E
Compound
3 oral 7 14.3 0.9 6.7
175
Compound
1 oral 8 30.7 4.6 10.5
260
5 mL
0.5% MC oral 8 5.9 0.4 3.5
(volume)
Compound
1 oral 8 23.7 4.3 9.9
31
F
Compound
3 oral 8 14.5 1.5 3.3
205
Compound
3 oral 8 16.4 2.4 5.1
256
5 mL
G 0.5% MC oral 10 14.9 1.8 14.9 7 13.8 1.8 6.0
(volume) Compound
0.3 oral 10 36.9 7.3 36.9 8 20.1 3.8 7.4 260
Compound
1 oral 10 23.5 3.1 23.5 9 23.1 3.1 9.9 260
Compound
3 oral 10 23.6 3.8 23.6 5 36.1 7.3 13.0 260
diazepam 1.5 IP 10 33.1 3.2 33.1 3 29.2 3.2 12.3
Example 439. Persistence of Treatment Effect with Compound 31
Using the methods described above, the duration of the effect seen with mice treated with Compound 31 was tested. Mice received either an oral dose of 0.5% methylcellulose (MC) 30 minutes before testing; or an oral dose of Compound 31 , at a dose of lmg/kg, 30 minutes, 60 minutes, or 120 minutes prior to testing. Videos were manually scored for number of open arm entries by a scorer blinded to treatment. Compound 31 treated mice spent more often and spent■ more time in the open arm areas of the maze than MC treated mice, indicating that the effect seen with Compound 31 has a duration of at least 2 hours. Longer durations were not tested.
Figure imgf000673_0001
Example 440. Anxiolytic effect of Compounds of the Invention in the Wistar -Kyoto Rat Elevated Plus Maze of Anxiety
Wistar-Kyoto rats were employed in an elevated-plus maze assay, using Midazolam l mg/kg dose and 0.5% methylcellulose as positive and negative controls, respectively, as well as Compound 260 at doses of 0.3, 1 , 3 and 10 mg/kg. Results showed that rats did not spend significant time in the open arm portions of the maze (FIGs. 3A and 3B); however, they were more active in the closed arm portions than rats receiving vehicle or midazolam (FIG. 3C). Rats travelled a greater distance (measured in centimeters) as the dose of Compound 260 (FIG. 3D) was increased.
Example 441. Stimulant Effects of Compounds of the Invention in NMRI Mice
Compound 260 was tested to ensure rats were not responding to a stimulant effect of compounds of the invention. In this example, phencyclidine (PCP) was tested at 5mg/kg dose delivered subcutaneously and with 0.5% methylcellulose (MC) (vehicle) (FIG. 4A), or
Compound 260 at various doses in NMRI mice (FIG. 4B). PCP treated mice had a significantly higher level of locomotor activity compared to methylcellulose demonstrating a marked stimulatory effect whereas Compound 260 at multiple doses did not demonstrate any stimulatory effect different than the control. This example demonstrates that the increased exploration of Compound 260-treated rats in the elevated plus maze was not due to a PCP-like effect of Compound 260.
Incorporation by Reference
All publications and patents mentioned herein, are hereby incorporated by reference in their entirety as if each individual publication or patent was specifically and individually indicated to be incorporated by reference.
Equivalents
Those skilled in the art will recognize, or be able to ascertain using no more than routine experimentation, many equivalents to the specific embodiments of the invention described herein. Such equivalents are intended to be encompassed by the following claims.

Claims

ciaim:
1. A method of treating a TRPC5 mediated d sorder in a subject, the method comprising administering to the subject a compound of Formula I:
Figure imgf000675_0001
Formula or a pharmaceutically acceptable salt thereof, wherein:
R3 is C;-C6 alkyl, C2-C alkenyl or C C^ alkynyl, each of which is optionally substituted with 1-4 R5;
R is C{-Cg alkyl, Cj-C6 lieteroalkyl, C2-C6 alkenyl, Cj-Ce alkynyl, Ci-C6 lialoaikyl, halo. Ci-Ce haloalkoxy, hydroxy!, Cj-Ce alkoxy, C3-C7 cyeloalkyloxy, C6-CJ O and, Ce-Cjo aryloxy, C7~ Cjfj arylalkoxy, amino, Ci-Cg akylamino, Cj-Ciz dialkyiamino, -S-, -S-Ci-Ce alkyl, -S(0)-, - 8(0)2-, heterocycloalk i, heieroaryl, heteroaryloxy, sulfonamidyl, amido, urea, sulfonylurea, acyl, nitro, cyano, wherein each of Ci-C6 alkyl, CrC6 heteroalkyl, C2-C6 alkenyl, C2-C6 alkynyl, C\~C haloalkyl, Q-Ce haloalkoxy, hydroxyl, Q-Ce alkoxy, C3-C7 cyeloalkyloxy, Ce-Cio aryl, C6-C|o aryloxy. C7-C16 arylalkoxy, amino, C C6 akylamino, C2-C12 dialkyiamino, -S-, -S-Ci-Ce alkyl, -S(0)-, ~S(0)2-, heterocycloalkyi. heieroaryl, heteroaryloxy. sulfonamidyl, amido, urea, sulfonylurea, acyl, is optionally substituted with 1 -3 R6;
R3 is Ci-Ce alkyl, Cj-Q lieteroalkyl, C3-C7 cycloalkyl, C2-C alkenyl, C2-C6 alkynyl, C2- C6 hydroxyalkyl, or Q-Ce alkoxy, each of which is optionally substituted with 1 -4 R 7;
R* is C j-Cfi alkyl, Ct~C6 heteroalkyl, C2-C6 alkenyl or C2-C6 alkynyl, each of which is optionally substituted with 1 -4 RS;
R\ R°, R7, and R8 are each independently Ci-Cg alkyl, Q-Ce heteroalkyl, halo, Ci-C6 haloalkyl. Ci-C6 haloalkoxy, hydroxyl, C C6 alkoxy, amino, ( C6 alkyiamino, CVC12 dialkyiamino, cyano, nitro, amido, Cj-C6 alkyiamido, C2-C12 diaikylamido, -S-, -8(0)2-,
-C(0)O, -C(0)-, -C(0)()-Ci-C6 alkyl, C3-C7 cycloalkyl, C6-Ci0 aryl, heterocycloalkyi, or heieroaryl, wherein each of d-Ce alkyl, Q-Ce heteroalkyl, C1-C haloalkyl, Ci-Ce haloalkoxy, hydroxy], Q-Cg alkoxy, amino, Q-Ce alkylamino, C2-C12 dialkylamino, amido, Q-Qs alkylamido, C2-C12 dialkylamido, -S-, -S(0)2-, -C(0)0-, -C(O)-, -C(0)0-Q-C6 alkyl, C3-C7 cycloalkyl, C6-Qo aryl, heterocycloalkyl, or heteroaryl is optionally substituted with 1-3 R : and each R is independently Q-Q alkyl, Q-C-6 heteroalkyl, Q-C6 ha.loa3k.yL Q-Cg haloalkoxy, heterocycloalkyl, C6-Cio aryl, heteroaryl, C4-Cio cycloaikylalkyl, heterocycloalkyl- Ci-Ce alkyl, C7-C16 arylaikyl, heteroaryl-Q-Ce alkyl, halo, hydroxy!, Q-Ce alkoxy, C6-Qo aryloxy, C7-C16 arylalkoxy, Q-Cg alkoxyaikoxyi, amino, Q-Q akylamino, C2-C12 dialkylamino Ci-C6 akyl-amino-Cj-Ce akyl C|-C6 akykaniino-tVCS2 dialkyl, -S-, ~S-Q-C6 alkyl -S(0)2-Q- C-6 alkyl, sulfonamidyf, amido, urea, sulfonylurea, acyl, -C(0)-Cg-Cio aryl -NHC(0)-C6-Qo ary -C(0)NH-C6-Qo aryl -C(0)OHs -C(0)G-CrC6 alkyl -C(0)-Cj-C6 alkyl acyl, nitro, or cyano; to thereby treat the subject.
2. The method of claim 1 , wherein the TRPC5 media ted disorder is selected from the group consisting of: 'a neuropsychiatrie disorder, a neurodegenerative disorder, nephropathy, and seizure disorder,
3. The method of claim 1, or a pharmaceutically acceptable salt thereof, wherein R1 is Ci-C alkyl,
4. The method of claim 1, or a pharmaceutically acceptable salt thereof wherein R1 is Ci-C alkyl and R5 is independently Q-Qo aryl or heteroaryl
5. The method of claim 1, or a pharmaceiiticaily acceptable salt thereof, wherein R2 is C6- C10 aryl, C6-Cio aryloxy or heteroaryloxy.
6. The method of claim 1, or a pharmaceutically acceptable salt thereoi", wherein R is C6- CJO aryloxy,
7. The method of claim 1, or a pharmaceutically acceptable salt thereof, wherein R" is Q-C alkoxy.
8. The method of claim 1 , or a pharmaceutically acceptable salt thereof, wherein R' is Q-C alkyl
9. The method of claim 1 , or a pharmaceutically acceptable salt thereof, wherein R" is or Cj-C6 akylamino.
10. The method of claim 1 , or a pharmaceutically acceptable salt thereof, wherein R' is - S(O)- or -S(0)2-.
1 1. The method of claim 1 , or a pharmaceutically acceptable salt thereof, wherein RJ is Cj-C6 alkyl, ,2~ e hydroxyalkyl, or Cj-Ca alkoxy.
12. The method of claim 1 , or a pharmaceutically acceptable salt thereof, wherein RJ is hydroxypropyl.
13. The method of claim I , or a pharmaceutically acceptable salt thereof, wherein is Ci-Q; a kyl
14. The method of claim 1 , or a pharmaceutically acceptable salt thereof, wherein R: is independently Ce-Qo aryl, heteroaryl, C3-C7 cyeloalkyL or heterocycloalkyl.
15. The method of claim 1 , or a pharmaceutically acceptable salt thereof, wherein R5 is phenyl, pyridyl, thiazolyl, pyrimidinyl, or oxazolyl.
16. The method of claim L or a pharmaceutically acceptable salt thereof, wherein R5 is phenyl.
17. The method of claim 1 , or a pharmaceutically acceptable salt thereof, wherein Rs is pyridyl,
1 8. The method of claim 1 , or a pharmaceutically acceptable salt thereof, wherein R5 is thiazolyl.
19. The method of claim 1 , or a pharmaceutically acceptable salt thereof, wherein R9 is independently Ci -Cg alkyl, Q-Cg alkoxy, halo, Cj-Cg haloalkyl, C C6 haloalkoxy, or
heterocycloalkyl,
20. The method of claim L or a pharmaceutically acceptable salt thereof, wherein is C(,- CJO aryl or C^-Cio aryloxy, and R6 is independently C\-C& alkyl, Ci-C6 haloalkyi, C; ~C6 or haloalkoxy.
21. The method of claim 1 , or a pharmaceutical ly acceptable salt thereof, wherein is C&- Cio aryloxy and R'J is independently Cj-Cg haloalkyi or \~C or haloalkoxy.
22. The method of claim 1 , or a pharmaceutically acceptable salt thereof, wherein RX is C6~ C;o aryloxy and R6 is -CF3 or -OCF3,
23. A compound of Formul a 1(a) :
Figure imgf000678_0001
Formula If a) or a pharmaceutically acceptable salt thereof, wherein:
R1 is Ci-Cg alkyl. Cj-Ce aikenyl or C2-C6 aikynyl, each of which is optionally substituted with 1 -4 R5;
R is CJ -C , alkyl, Ci-Cg heteroalkyl, C2-C6 aikenyl, Ca-Ce aikynyl, C . -Cg haloalkyi, halo, C1-C6 haloalkoxy, hydroxyl, Cj-Cg alkoxy, C3-C7 cycloalkyloxy, Cg-Cio aryl, Q-Qo aryloxy, C7- Ci6 arylalkoxy, amino, Ci-Ce akyiammo, C2-C12 dialkyiamino, -S-, -S-C1-C6 alkyl, -S(O)-, - S(0)2-, heterocycloaikyh heteroarvl, heteroaryloxy, sulfonamidyl, arnido, urea, sulfonylurea, acyl, niiro, cyano, wherein each of C-rC alkyl, Q-Ce heteroalkyl, Cj-Ce aikenyl, Ca-Cg aikynyl, C1-C6 haloalkyi, Ci-C6 haloalkoxy, hydroxyl, C -C alkoxy, C3-C7 cycloalkyloxy, Q-Cio aryl, Cg-Cj o aryloxy, C7-C16 arylalkoxy, amino, CrQ akylamino, C2-CS2 dialkyiamino, -S-, -S-Cj-Ce alkyl, -S(O)-. -8(0)2-, heterocycloaikyh heteroaryl, heteroaryloxy, sulfonamidyl, amido, urea, sulfonylurea, acyl, is optionally substituted with 1 -3 R&;
RJ is C2-C6 hydroxy alkyl or C-rCf. heteroalkyl;
R4 is Ci-C6 alkyl, Cj-C-6 heteroalkyl, C2-C6 aikenyl or C2-C6 aikynyl, each of which is optionally substituted with 1 -4 R8; R" , R°, and R' are each independently CrC6 alkyl, Q-Cg heteroalky , halo, CpC 6 haloalkyi, Ci-C6 haioalkoxy, hydroxy!, Cj-Cg alkoxy, amino, Ci-Cg aikylammo, C2-C12 dialkylamino, cyano, nitro, amido, C Cs alk lamido, C2-Cj2 dialkylamido, -S-, -8(0)2-,
-C(0)0-5 -C(O)-, -C(0)0-CrC6 alkyl, C3-C7 cycloalkyl, C6-C10 aryl, heterocycloalkyl, or heteroaryl, each of CpCe alkyl, Cj-C6 heteroalkyi, Cj-Q haloalkyi, -Ce haioalkoxy, hyd Ci-Ce alkoxy, amino, CpCe alkylamino, C2-C12 dialkylamino, amido, Ci-Cg alkylamido, C ?-C;2 dialkylamido, -S-, -S(0)2-, -C(0)0-, -C(O)-, -C(0)0-CrC6 alkyl, C3-C7 cycloalkyl, C6-C10 aryl, heterocycloalkyl, or heteroaryl is optional ly substituted with 1 -3 R9; and
each R'J is independently Ci-C6 alkyl, Ci-Q, heteroalkyi, Ci-C6 haloalkyi, Ci -C6 haioalkoxy, heterocycloalkyl, C6-Cio aryl, heteroaryl, Gt-Cjo cycloalkylalkyl, heterocycloalkyl - Ci-Ce alkyl, C7-Ci6 arylalkyl, heteroaryl-Q-Ce alkyl, halo, hydroxyl, Ci-C6 alkoxy, Ce-Cso aryioxy, C7-Cj6 arylalkoxy, C2-Cg alkoxyalkoxyl, amino, -Ce akylamino, C2-C12 dialkylamino, Ci-f 6 akyl-arnino-Ci-Cs akyl, C{-C6 akyl-amino-C -C-^ dialkyl, -S-, -S-Ci-Ce alkyl, -SiO)rCj- C alkyl, sulfonamidyl, amido, urea, sulfonylurea, acyl, -C(0)-C6-Cio aryi, -NHC(0)-C6-CJO aryl, -C(O)NH-C6-C10 aryl -C(0)OH, -C(0)0-C C6 alkyl, -C(0)-d-C6 alkyl acyl, nitro, or cyano.
24. The compound of claim 23, or a pharmaceutically acceptable salt thereof, wherein R1 is Ci-Q alkyl and R5 is phenyl, pyridyl, thiazolyl, pyrimidinyl, or oxaz.ol.yl.
25. The compound of claim 23, or a pharmaceutically acceptable salt thereof, wherein R2 is C( -C;o aryl or C6-Cio aryioxy, and R6 is independently Ci-C6 alkyl, Q-Ce haloalkyi, ]-C or haioalkoxy.
26. The compound of claim 23, or a pharmaceutically acceptable salt thereof, wherein R"' is C Cf, hydroxyalkyh
27. A compound of Formula IT
Figure imgf000679_0001
Formula II
or a pharmaceutically acceptable salt thereof, wherein:
Ring A is phenyl, thiazoiyl, pyriniidinyl, or oxazoly!;
R2 is Cj-Ce alkyl, C] -C6 heteroalkyl, C2-C6 alkenyl, C i-Cf, aikynyl, Cj-C6 haloalkyL halo, Ci-C6 haloalkoxy. hydroxvl, Ci-Ce alkoxy, C3-C7 cycloalkyloxy, C6-CJ O aryl, Ce-Cio aryloxy, C7- Cs6 arylalkoxy, amino, CrC6 akylamino, C2-Ci2 dialkylamino, -S-, -S-Ci-Ce alkyl, -S(OV, - 8(0)2-, heterocycloaikyl, heteroaryl, heteroaryloxv, sulfonamidyL amido, urea, sulfonylurea, acyl, nitro, cyano, wherein each of C¾ -C6 alkyl, Ct- , heteroalkyl, C2-C.6 alkenyl, C2-C6 aikynyl, Ct-C6 haloalkyL C Ce haloalkoxy, hydroxy], Q-Q alkoxy, C3-C7 cycloalkyloxy, Cg-Cjc aryl, Ce-Cio aryloxy, C7-C16 arylalkoxy, amino, CrC6 akylamino, C2-Ci2 dialkylamino, -S-, -S-Cj-Ce alkyl, -S(O)-, -S(0)2-, heterocycloaikyl, heteroaryl, heteroaryloxv, sulionamidyl, amido, urea, sulfonylurea, acyl, is optionally substituted with 1 -3 R6:
R:' is CrC-c alkyl, C · -( :<-. heteroalkyl, C2-Cg alkenyl, C2-C6 aikynyl, C2-C;, hydroxyalkyl, or Cj -Cf, alkoxy, each of which is optionally substituted with 1 -4 R7;
R is Ci-Ce alkyl, Ci-Cg heteroalkyl, C2-C6 alkenyl or C2-Ce aikynyl, each of which is optionally substituted with 1-4 Rx;
R , R , and R are each independently Ci-Ce alkyl, Ci-C heteroalkyl, halo, Cj-Ce haloaikyl, Cj-Q haloalkoxy, hydroxvl, Ct-Q alkoxy, amino, Ci -Q alkylarnino, C2-C12 dialkylamino, cyano, nitro, amido, Cj-Q alkyi amido, C2-Q2 d alkylamido, -S-, -S(0)2-,
-C(0)0-, -C(O)-, -C(0)0-Ci-C6 alkyl, C3-C7 cycloalkyl, C6~C10 aryl, heterocycloaikyl, or heteroaryl. wherein each of Q-Q alkyl, Q-Q heteroalkyl, Q-Q haloaikyl, Cj-Q haloalkoxy, hydroxy 1, -Q alkoxy, amino, Q-Q alkylamino, C2-Ci2 dialkylamino, amido, Q-Q
aikylarnido, C2-Cl2 dialkylamido, -S-, -S(0)2-, -C(0)0-, -C(O)-, -C(0)0-Ci-C6 alkyl, C3-C7 cycloalkyl, Q-Qo aryl, heterocycloaikyl, or heteroaryl is optionally substituted with 1 -3 R9; each R9 is independently Q-Q alkyl, Q-Q heteroalkyl, Q-Q haloaikyl, Q-Q haloalkoxy, heterocycloaikyl, Q-Qo a^y heteroaryl, Q~Qo cycloalkylalkyl, heterocycloaikyl- Q-Q alkyl, C7-C16 arylalkyL heteroaryl-Q-Q alkyl, halo, hydroxy), Q -Q alkoxy, C6-C10 aryloxy, C7-Q6 arylalkoxy, Q-Q alkoxy alkoxyl, amino, Q-Q akylamino, C2-Ci2 dialkylamino, Q -Q, akyhamino-Q-Q akyl, Q-Q akyl-amino-Q-C 12 dialkyl, -S-, -S-Q-Q alkyl, -S(0)2-Q- Q alkyl, sulionamidyl, amido, urea, sulfonylurea, acyl, -C(0)-Q-Qo aryl ~NHC(0)-Q~Qo ar l, -C(0)NH-C6-C,o aryl, -C(0)OH, -C(0)Q-Q-Q alkyl, -C(0)-Ci-C6 alkyl acyl, nitro, or cyano; each Ra is C\~C(, alkyl, Cj-Cf, haioalkyl, halo;
n is 1 or 2: and
m is I , 2, or 3.
28. The compound of claim 27. or a pharmaceutically acceptable salt thereof, wherein Ring A is phenyl or thiazolyl,
29. The compound of claim 27, wherein RJ is hydroxypropyl.
30. A compound of Formula III:
Figure imgf000681_0001
Formula III
or a pharmaceutically acceptable salt thereof, wherein:
R2 is Cj -·(.";. alkoxy or aryloxy substituted with 1-3 R6;
RJ is Cj -C6 heteroalkyl or C2-C6 hydroxyalkyl:
R4 is C]-C6 aikyi;
R6 is independently Ci-Ce alkyl, halo, Ci-C6 haioalkyl, Ci-Ce haloalkoxy, or Cj-Ce alkoxy;
each Ra is Ci-C6 alkyl, Q-Cg haioalkyl, halo;
n is 1 or 2; and
m is I , 2, or 3.
31. The compound of claim 30, or a pharmaceutically acceptable salt thereof, wherein RJ is hydroxypropyl.
32. The compound of claim 30, or a pharmaceutically acceptable salt thereof, wherein Ra is independently chioro, fluoro, or methyl.
33. A composition comprising a compound of any one of claims 1 to 32, or a pharmaceutically acceptable salt thereof.
34. A method of treating a. TRPC5 mediated disorder in a subject, the method comprising administering to the subject a compound or composition of any one of claims 23 to 33, to thereby treat the subject.
35. The method of claim 34, wherein the TRPC5 mediated disorder is selected from the group consisting of: a neuropsychiairic disorder, a neurodegenerative disorder, nephropathy, and seizure disorder,
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WO2016159746A1 (en) 2015-03-27 2016-10-06 Latvian Institute Of Organic Synthesis Novel ethynylxanthines, their preparation and use as inhibitors of matrix metalloproteinases and angiogenesis
WO2016159745A1 (en) 2015-03-27 2016-10-06 Latvian Institute Of Organic Synthesis Ethynylxanthines, preparation and use for cancer treatment
CN107530349B (en) * 2015-04-08 2021-01-08 路易斯和克拉克制药公司 Xanthine substituted alkynyl carbamates/trans carbamates as A2B antagonists
WO2018146485A1 (en) * 2017-02-09 2018-08-16 University Of Leeds Trpc ion channel inhibitors for use in therapy
WO2018157800A1 (en) * 2017-02-28 2018-09-07 National Institute Of Biological Sciences, Beijing Mlkl inhibitors
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AU2018299824B2 (en) * 2017-07-11 2022-06-16 Boehringer Ingelheim International Gmbh Novel substituted xanthine derivatives
WO2019173327A1 (en) 2018-03-05 2019-09-12 Goldfinch Bio, Inc. Imidazodiazepinediones and methods of use thereof
CN112351781A (en) * 2018-03-05 2021-02-09 金翅雀生物公司 Imidazodiazepine diones and methods of use thereof
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WO2020120449A1 (en) 2018-12-12 2020-06-18 Boehringer Ingelheim International Gmbh Substituted xanthine derivatives
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