WO2021083438A1 - Inhibitors of purine nucleoside phosphorylase - synthesis and use thereof for treatment of t-cell acute lymphoblastic leukemia and lymphoma - Google Patents

Inhibitors of purine nucleoside phosphorylase - synthesis and use thereof for treatment of t-cell acute lymphoblastic leukemia and lymphoma Download PDF

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WO2021083438A1
WO2021083438A1 PCT/CZ2020/050085 CZ2020050085W WO2021083438A1 WO 2021083438 A1 WO2021083438 A1 WO 2021083438A1 CZ 2020050085 W CZ2020050085 W CZ 2020050085W WO 2021083438 A1 WO2021083438 A1 WO 2021083438A1
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mhz
nmr
pyrrolo
mmol
dihydro
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French (fr)
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Jan SKACEL
Zlatko Janeba
Helena MERTLIKOVA KAISEROVA
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Ustav Organicke Chemie A Biochemie Av Cr, V. V. I.
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Publication of WO2021083438A1 publication Critical patent/WO2021083438A1/en

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
    • A61K31/519—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim ortho- or peri-condensed with heterocyclic rings
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A61P35/02—Antineoplastic agents specific for leukemia
    • 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

Definitions

  • the present invention concerns new compounds for application in the treatment of T-cell acute lymphoblastic leukemia and lymphoma, their synthesis, and their use in vitro and in vivo.
  • T-cell acute lymphoblastic leukemia did not undergo such rapid development as other cancer treatments in past years. Because of this, new and innovative treatments and treatment procedures are needed to increase prognosis of T-ALL patients. Espetially prognosis of adult T-ALL patients are very poor and the overall five-year survival rate is only approximately 40%.
  • PNP purine nucleosid phosphorylase
  • forodesine hydrochloride (Mundesine), a PNP inhibitor, was approved for T-cell lymphoma treatment by PMDA.
  • Forodesine hydrochloride exhibits high inhibitory activity against PNP and T-cell cancer cell lines [Kicska, G., et al, Proc. Natl. Acad. Sci. U. S. A. 2001, 98 (8), 4593-4598],
  • forodesine hydrochloride exhibits poor oral bioavailability which is less than 10% in human and nonhuman primates [Kilpatrick, J. M., et al.. Int. Immunopharmacol. 2003, 3 (4), 541-548],
  • X is independently -NH- or -S- in combination with Ar, Z, G and R;
  • alkyl is a linear or branched hydrocarbon chain
  • aryl is an aromatic hydrocarbon group consists of 6 to 12 carbon atoms and at least one aromatic ring. It can be unsubstituted or substituted with one or more groups, including -OH, -O(aryl), - O(arylalkyl), -C(O)-, -CN and halogen; particularly preferably, aryl is phenyl, optionally substituted; heteroaryl is an aromatic hydrocarbon group consisting of 4 to 11 carbon atoms and at least one aromatic ring and at least one heteroatom included O, N, S.
  • arylalkyl is an alkyl group that contains one or more aryl groups which can be unsubstituted or substituted with one or more function groups, including -OH, -O(aryl), -O(arylalkyl), -C(O)-, - CN and halogen.
  • R 2 is independently linear or branched C 1 -C 10 alkyl chain and -CHR 3 COOR 4 , wherein R 3 is independently linear or branched C 1 -C 10 alkyl chain in which any -CH 2 - group can be replaced independently by -S-, -0-, -NH-, C 6 -C 12 aryl and C 6 -C 12 arylalkyl, and R 4 is independently linear or branched C 1 -C 10 alkyl chain;
  • R is independently -H, -OH, -O(aryl), -O(arylalkyl), -C(O)-, -CN and halogen in combination with Ar, Z and G;
  • Reaction conditions include: a protic or an aprotic solvent selected from methanol, ethanol, «-propanol, 2-propanol, «-butanol, 2- butanol, toluene, acetone, dimethylformamide, dioxane, tetrahydrofuran, ethyl acetate, butyl acetate, dimethylsulfoxide, diethylether, N-methylpyrrolidonc.
  • a protic or an aprotic solvent selected from methanol, ethanol, «-propanol, 2-propanol, «-butanol, 2- butanol, toluene, acetone, dimethylformamide, dioxane, tetrahydrofuran, ethyl acetate, butyl acetate, dimethylsulfoxide, diethylether, N-methylpyrrolidonc.
  • Reaction conditions include: a protic or an aprotic solvent selected from methanol, ethanol, «-propanol, 2-propanol, «-butanol, 2- butanol, toluene, acetone, dimethylformamide, dioxane, tetrahydrofuran, ethyl acetate, butyl acetate, dimethylsulfoxide, diethylether, N-methylpyrrolidone.
  • a protic or an aprotic solvent selected from methanol, ethanol, «-propanol, 2-propanol, «-butanol, 2- butanol, toluene, acetone, dimethylformamide, dioxane, tetrahydrofuran, ethyl acetate, butyl acetate, dimethylsulfoxide, diethylether, N-methylpyrrolidone.
  • chloroform dichlormethane, 1,2- dichloroethane, benzene, ere sole; temperature in the range 0 °C to 250 °C; an organic or an inorganic base selected from sodium carbonate, potassium carbonate, cesium carbonate, sodium bicarbonate, sodium phosphate, potassium phosphate, sodium hydroxide, potassium hydroxide, lithium hydroxide, triethylamine, trimethylamine, diisopropylamine, diisopropylethylamine, dimethylaminopyridine, 2,6-lutidine; catalysis with suitable palladium form selected from palladium(II) chloride, palladium(II) acetate, tris(dibenzylidenacetone) dipalladium, so-called precatalysts known as [4-(di-tert-butylphosphine)- N, N-dimethylaniline-2-(2'-aminobiphenyl)
  • di-tert-butylphenylphosphine 4,5- bis(diphenylphosphine)-9,9-dimethylxanthene, 4,6-bis(diphenylphosphine)- 10A-phcnoxazinc.
  • IC 50 The half maximal inhibitory concentration riPNP Human Purine Nucleoside Phosphorylase
  • Microwave experiments were performed in 10 mF or 30 ml vials with a CEM Discover (Explorer) microwave apparatus operating at a frequency of 2.45 GHz with continuous irradiation power from 0 W to 300 W. Large-scale reactions were carried-out in the Syrris Atlas Potassium system with 21, 1 1 and 0.5 1 jacket reactor coupled with Julabo FP50-HL Refrigerated/Heating Circulator.
  • Analytical TLC was performed on silica gel pre-coated aluminium plates with fluorescent indicator (Merck 60 F254). Flash column chromatography was carried out by Teledyne ISCO CombiFlash Rf200 with dual absorbance detector. Various types of columns were used: a) Teledyne ISCO columns RediSepRf HP Silica GOLD in sizes 12 g, 40 g, 80 g and 120 g; b) Teledyne ISCO columns RediSepRf HP C18 Aq GOLD in sizes 50 g and 100 g; c) column Chromabond Flash DL 40, DL 80, DL 120 and DL 200, fdled with FLUKA silica gel 60; d) Interchim puriFlash C18 Aq in sizes F0040 and F0080.
  • Preparative HPLC purifications were performed on Waters Delta 600 chromatography system with columns packed with Cl 8 reversed phase resin (Phenomenex Gemini 10 ⁇ m 21 x 250 mm, Phenomenex Gemini 5 ⁇ m 21 x 250 mm, Phenomenex Luna 10 ⁇ m 21 x 250 mm) using gradient H 2 0/MeOH as eluent.
  • Mass spectra, UV spectra and purity of compounds were measured on Waters UPLC-MS system consisted of Waters UPLC H-Class Core System (column Waters Acquity UPLC BEH C18 1.7 mm, 2.1 x 100 mm), Waters Acquity UPLC PDA detector and Mass spectrometer Waters SQD2.
  • NMR spectra were recorded on Bruker Avance 400 or 500 spectrometers referenced to the residual solvent signal or a specified additive.
  • Dowex® 50D resin was turned to Na + cycle by treatment of Dowex D50 resin in H + cycle with 1M NaOH aq. solution, followed by water wash to neutral pH.
  • the jacket reactor (21) was flushed with nitrogen, charged with benzyl alcohol (1 1) and the system was set to retain temperature 20 °C.
  • Sodium metal (22.5 g, 977 mmol, 1.5 eq.) was added in portions and the mixture was stirred 20 hours under small flow of nitrogen. The mixture was then heated at 80 °C for 1 hour, and then it was cooled back to 20 °C.
  • pyrimidinc (100 g, 651 mmol, 1 eq.) was charged and the mixture was stirred at 80 °C until complete conversion was achieved (ca. 4 hours).
  • the mixture was cooled to 5 °C, diluted with water (400 ml), pH was adjusted to 7 with 3M HC1 (aq.) (ca. 100 - 130 ml).
  • the mixture was heated to 20 °C, extracted with chloroform (3 x 400 ml) and the organic phase was washed with brine (500 ml).
  • the mixture was concentrated and benzyl alcohol was evaporated at high vacuum at ca. 90 °C.
  • the solid was filtered through a short pad of silica gel (600 g) with eluent (100% of chloroform, then chloroform with 5% methanol).
  • the jacket reactor (1 1) was flushed with nitrogen, 4-(benzyloxy)-5H-pyrrolo[3.2-d]
  • LI 4-(benzyloxy)-5H-pyrrolo[3.2-d]
  • the jacket reactor (2 1) was flushed with nitrogen and 4-(benzyloxy)-7-iodo-5H- pyrrolo[3.2-d]
  • the jacket reactor (1 1) was flushed with nitrogen and 4-(benzyloxy)-5H-pyrrolo[3.2-d]
  • the vial was sealed and inserted into the microwave reactor for 1 h at 120 °C.
  • the dark-brown reaction mixture was dissolved in chloroform and washed with a half- saturated aqueous solution of NaHCO 3 (2x), 1M HC1 (aq.) (2x), brine (1x), dried with MgSO 4 , filtered and evaporated.
  • the residue was adsorbed on silica gel in a mixture of cyclohexane/acetone and purified by flash chromatography on silica gel (chloroform to 5% of methanol), yielded coupling product as a brown oil.
  • the oil residue was dissolved in dry methanol and potassium carbonate (2.30 g, 16.6 mmol, 2 eq.) was added.
  • the vial was sealed and inserted into the microwave reactor for 2 h at 120 °C.
  • the dark-brown reaction mixture was dissolved in chloroform, washed with a half- saturated aqueous solution NaHCO 3 (2x), 1M HC1 (aq.) (2x), brine (1x), dried with MgSO 4 , filtered and solvents were evaporated. Purification by flash chromatography on silica gel (cyclohexane to 15% ethyl acetate modified with 10% methanol (v/vi)) afforded the title compound as a white -off oil.
  • Tetraethyl methylenediphosphonate (2.28 ml, 11.6 mmol, 1.2 eq.) was dissolved in dry tetrahydrofuran (60 ml) under an argon atmosphere and potassium tert-butoxide (1.3 g, 11.6 mmol, 1.2 eq.) was added. The mixture was stirred at room temperature for 1 hour and a solution of compound L19 (3 g, 9.65 mmol, 1 eq.) in dry tetrahydrofuran (20 ml) was added.
  • Diethyl (E )-(4-hydroxy-2-iodostyryl)phosphonate L23 Compound L22 (4 g, 16.1 mmol, 1 eq.) was dissolved in dry toluene (80 ml) and diethylphosphonoacetic acid (3.8 ml, 19.4 mmol, 1.2 eq.), piperidine (524 ⁇ l, 5.13 mmol, 0.33 eq.) and acetic acid (221 ⁇ l, 3.86 mmol, 0.24 eq.) were added subsequently.
  • the mixture was extracted with dichloromethane (3x), washed with brine (1x), dried with MgSO 4 , filtered and evaporated.
  • the solid was adsorbed on silica gel in a mixture of cyclohexane/acetone and it was purified by flash column chromatography on silica gel (cyclohexane to 10% ethyl acetate modified with 10% of methanol (v/v i )), yielded 4.1 g (87%) of the title compound as yellow needles.
  • the reaction was quenched with a half-saturated aqueous solution of NH 4 Cl, extracted with ethyl acetate (3x), washed with brine (1x) and dried with MgSO 4 .
  • the mixture was evaporated to dryness and it was adsorbed on silica gel in a mixture of cyclohexane/acetone . Purification by flash chromatography on silica gel (cyclohexane to 40% ethyl acetate modified with 10% methanol (v/vi)) afforded a pure product.
  • Tetraethyl methylenediphosphonate (597 ⁇ l, 2.4 mmol, 1.2 eq.) was dissolved in dry tetrahydrofuran (4 ml) under an argon atmosphere and potassium tert-butoxidc (269 mg, 2.4 mmol, 1.2 eq.) was added. The mixture was stirred at room temperature for 1 hour and a solution of 5-iodofuran-2-carbaldehyde (444 mg, 2.0 mmol, 1 eq.) in dry tetrahydrofuran (1 ml) was added.
  • Tetraethyl methylenediphosphonate (597 ⁇ l, 2.4 mmol, 1.2 eq.) was dissolved in dry tetrahydrofuran (4 ml) under an argon atmosphere and potassium tert-butoxidc (269 mg, 2.4 mmol, 1.2 eq.) was added. The mixture was stirred at room temperature for 1 hour and a solution of 2-iodothiophene-3- carbaldehyde (478 mg, 2.0 mmol, 1 eq.) in dry tetrahydrofuran (1 ml) was added.
  • Solvents were evaporated at 40 °C/900 mbar and the residue was dissolved in dry tetrahydrofuran (78 ml). The mixture was cooled to 0 °C and sodium hydride (3.9 g of 60% oil dispersion, 97.7 mmol, 1.25 eq.) was added portionwise. The mixture was stirred at 0 °C for 30 minutes and TfOCH 2 P(O)(Oi-Pr)2 (32 g, 97.7 mmol, 1.25 eq.) was added and the mixture was stirred 1 hour at 0 °C. The reaction was quenched with a half- saturated aqueous solution of NH 4 CI and extracted with ethyl acetate (3x).
  • the mixture was dissolved in chloroform, washed with a half-saturated aqueous solution of NaHCO 3 (2x), 1M HC1 (aq.) (2x), brine (1x), dried with MgSO 4 , filtered and solvents were evaporated.
  • the solid was adsorbed on silica gel in a mixture of cyclohexane/acetone and purified by flash chromatography on silica gel (cyclohexane to 15% ethyl acetate modified with 10% methanol (v/vi)), yielded 800 mg (99%) of the product as an oil.
  • the compound was used in the next step without further characterization.
  • the mixture was dissolved in chloroform, washed with a half-saturated aqueous solution of NaHCO 3 (2x), 1M HC1 (aq.) (2x), brine (1x), dried with MgSO 4 , filtered and solvents were evaporated.
  • the solid was adsorbed on silica gel in a mixture of cyclohexane/acetone and purified by flash chromatography on silica gel (cyclohexane to 15% ethyl acetate modified with 10% methanol (v/vi)), yielded 700 mg (81%) of the product as an oil.
  • the compound was used in the next step without further characterization.
  • the compound was dissolved in trifluoroacetic acid (2 ml/ 100 mg of the material) at room temperature and stirred for 15 minutes. Trifluoroacetic acid was evaporated, coevaporated with water (2x) and a cold solution of ammonia in ethanol (2 ml/100 mg of the material) was added. The mixture was evaporated, the flask was sealed with septum, flushed with argon, and then dry pyridine (10 ml/ 1 mmol of the starting compound) was added followed by trimethylsilyl bromide (1 ml/1 mmol of the starting compound). The mixture was stirred at room temperature overnight and then it was evaporated to dryness. The residue was dissolved in a 2M aq.
  • Ethyl (diethoxyphosphoryl)methanesulfonate (673 mg, 2.6 mmol, 1.2 eq.), prepared by published procedure [Otrubova, K.; Cravatt, B. F.; Boger, D. L. J. Med. Chem. 2014, 57 (3), 1079-1089], was dissolved in dry tetrahydrofiiran (20 ml) under an argon atmosphere and potassium tert-butoxidc (290 mg, 2.6 mmol, 1.2 eq.) was added.
  • Ethyl (E )-3-(2-iodophenyl)acrylate L76 Triethyl phosphonoacetate (1 g, 5.2 mmol, 1.2 eq.) was dissolved in dry tetrahydrofuran (20 ml) under an argon atmosphere and potassium tert-butoxidc (580 mg, 5.2 g, 1.2 eq.) was added. The mixture was stirred at room temperature for 1 hour and a solution of 2-iodobenzaldehyde (1 g, 4.3 mmol, 1 eq.) in dry tetrahydrofuran (10 ml) was added dropwise.
  • the solid was dissolved in trifluoroacetic acid (2 ml/100 mg of the material) at room temperature and stirred for 15 minutes. Trifluoroacetic acid was evaporated, coevaporated with water (2x) and a cold solution of ammonia in ethanol (2 ml/100 mg of the material) was added. The mixture was evaporated to dryness and the solid was purified by HPLC (Cl 8, gradient HaO/MeOH). Product-containing fractions were filtered through a short pad of Dowex® 50 in Na + -cycle and solvents were evaporated. Purified product was lyophilized from water, yielded 16 mg (21%) of the title compound as a white solid.
  • Trifluoroacetic acid was evaporated, coevaporated with water (2x) and a cold solution of ammonia in ethanol (2 ml/100 mg of the material) was added. The mixture was evaporated to dryness and the solid dissolved in small amount of water and purified by HPLC (C18, gradient H 2 O /MeOH). Product-containing fractions were filtered through a short pad of Dowex® 50 in Na + -cycle and solvents were evaporated. Purified product was lyophilized from water, yielded 26 mg (27%) of the title compound as a white solid.
  • Trifluoroacetic acid was evaporated, coevaporated with water (2x), a cold solution of ammonia in ethanol (2 ml/ 100 mg of the material) was added and the mixture was evaporated to dryness.
  • the solid was dissolved in a mixture of dioxane and water (1: 1), several drops of 10M NaOH (aq.) were added, and the mixture was stirred at room temperature for 2 hours. Solvents were evaporated and the solid dissolved in a small amount of water and purified by HPLC (Cl 8, gradient ffO/McOH).
  • Pd2(dba)3 (84 mg, 0.0922 mmol, 0.1 eq.) and 4,5-bis(diphenylphosphino)-9,9-dimethylxanthene (107 mg, 0.1843 mmol, 0.2 eq.) were dissolved in dry dioxane (5 ml) under an argon atmosphere, and then triethylamine (192 ⁇ l, 1.4 mmol, 1.5 eq.) was added.
  • Trifluoroacetic acid was evaporated, coevaporated with water (2x) and a cold solution of ammonia in ethanol (2 ml/100 mg of the material) was added. The mixture was evaporated to dryness and the solid was adsorbed on silica gel in a mixture of cyclohexane/acetone. Purification by flash chromatography on silica gel (chloroform to 10% methanol) afforded 334 mg (97%) of the diethyl phosphonate, which was used in the next step without further characterization. MS (ESI-QMS) m/z: [M + H] + calcd for C16H19N304PS, 380.1; found, 380.2.
  • 2-Iodobenzyl bromide (3 g, 10.1 mmol, 1 eq.) was charged into a microwave reactor tube and the tube was flushed with an argon. Dry toluene (20ml) and triisopropyl phosphite (3 ml, 11.1 mmol, 1.1 eq.) were added subsequently and the mixture was heated in the microwave reactor at 160 °C for 1 hour.
  • Trifluoroacetic acid was evaporated, coevaporated with water (2x) and a cold solution of ammonia in ethanol (2 ml/100 mg of the material) was added. The mixture was evaporated to dryness and the solid was adsorbed on silica gel in a mixture of cyclohexane/acetone. Purification by flash chromatography on silica gel (chloroform to 10% methanol) afforded 208 mg (42%) of the diisopropyl phosphonate, which was used in the next step without further characterization. MS (ESI-QMS) m/z: [M + H] + calcd for C19H25N304PS 422,1; found, 422.3.
  • the reaction was quenched with a half-saturated aqueous solution of NH 4 CI and extracted with ethyl acetate (3x).
  • the organic phase was washed with brine, dried with MgSO 4 , filtered and solvents were evaporated.
  • the solid was adsorbed on silica gel in a mixture of cyclohexane/acetone and purified by flash chromatography on silica gel (cyclohexane to 20% ethyl acetate modified with 10% methanol (v/vi)) afforded 82 mg (60%) of alkylated product, which was used in the next step without further characterization.
  • the coupling product (110 mg, 0.2023 mmol, 1 eq.) was dissolved in trifluoroacetic acid (2 ml/100 mg of the material) at room temperature and stirred for 15 minutes. Trifluoroacetic acid was evaporated, coevaporated with water (2x) and a cold solution of ammonia in ethanol (2 ml/100 mg of the material) was added. The mixture was evaporated to dryness and the solid was purified by HPLC (Cl 8, gradient H 2 O/McOH). Product-containing fractions were filtered through a short pad of Dowex® 50 in Na + -cycle and solvents were evaporated. Purified product was lyophilized from water, yielded 54 mg (84%) of the title compound as a white solid.
  • Bromomethanesulfonamide (2 g, 11.5 mmol, 1 eq.) was dissolved in dry tetrahydrofuran (20 ml) under an argon atmosphere and the solution was cooled to 0 °C.
  • Sodium hydride (460 mg of 60% oil dispersion, 11.5 mmol, 1 eq.) was added and the mixture was stirred at 0 °C for 15 minutes.
  • 2- (trimethylsilyl)ethoxymethyl chloride (2 ml, 11.5 mmol, 1 eq.) was added and the mixture was stirred at 0 °C for 15 minutes.
  • the addition of sodium hydride and 2-(trimethylsilyl)ethoxymethyl chloride was repeated once more.
  • the reaction was quenched with a half-saturated aqueous solution of NH4CI, extracted with ethyl acetate (3x), and the organic phase was washed with brine (1x) and it was dried with MgSO/i.
  • the whole suspension with desiccant was half-diluted with chloroform, and it was filtered through a short pad of neutral alumina and the pad was washed with a mixture of chloroform and ethyl acetate (2: 1, v/vi). Solvents were evaporated, yielded 4.6 g (92%) of the title compound as a clear oil.
  • Trifluoroacetic acid was evaporated, coevaporated with water (2x) and a cold solution of ammonia in ethanol (2 ml/100 mg of the material) was added. The mixture was evaporated to dryness and the solid was dissolved in a small amount of dimethylformamide. The sample was subjected to purification by C18 reverse phase flash chromatography (water to methanol) in a liquid injection mode. Purified product was lyophilized from dioxane, yielded 11 mg (22%) of title compound as a white solid.
  • the mixture was dissolved in chloroform, washed with a half-saturated aqueous solution of NaHCO 3 (2x), 1M HC1 (aq.) (2x), brine (1x), dried with MgSO 4 , filtered and solvents were evaporated.
  • the solid was adsorbed on silica gel in a mixture of cyclohexane/acetone and purified by flash chromatography on silica gel (cyclohexane to 15% ethyl acetate modified with 10% methanol (v/vi)), yielded 800 mg (99%) of the product as an oil.
  • the compound was used in the next step without further characterization.
  • a flask was charged with the diethyl phosphonate (100 mg, 0.2467 mmol, 1 eq.), it was sealed with septum, flushed with argon, and then dry pyridine (10 ml/1 mmol of the starting compound) was added, followed by trimethylsilyl bromide (1 ml/1 mmol of the starting compound). The mixture was stirred at room temperature overnight and then it was evaporated to dryness under an argon atmosphere. L-phenylalanine ethyl ester hydrochloride (227 mg, 0.9868 mmol, 4 eq.) was added and the flask was sealed with septum, flushed with argon.
  • the solid residue was adsorbed on silica gel in a mixture of cyclohexane/acetone and purified by flash chromatography on silica gel (chloroform to 20% methanol), followed by purification on HPLC (Cl 8, gradient H 2 O/MeOH as eluent). Purified product was lyophilized from dioxane, yielded 100 mg (58%) of the title compound as a white solid.
  • the mixture was warmed to 60 °C and TsOCH 2 P(O)(0/-Pr) 2 (21.8 g; 62.3 mmol; 1.5 eq.) and potassium tert-butoxide (7.00 g; 62.3 mmol; 1.5 eq.) were added.
  • the mixture was stirred at 60 °C for 1 hour and additional portion of TsOCH 2 P(O)(0/-Pr) 2 (21.8 g; 62.3 mmol; 1.5 eq.) and potassium tert-butoxide (7.00 g; 62.3 mmol; 1.5 eq.) was added.
  • the addition was repeated after another 1 hour.
  • Dry pyridine (10 ml/1 mmol of the starting compound) and triethylamine (191 ⁇ l, 1.37 mmol, 6 eq.) were added to the mixture.
  • the mixture was stirred 15 minutes at 70 °C and a solution of triphenylphosphine (359 mg, 1.37 mmol, 6 eq.) and AldrithiolTM-2 AldrithiolTM-2 (302 mg, 1.37 mmol, 6 eq.) in dry pyridine (10 ml/1 mmol of the starting compound) was added.
  • the reaction was quenched by evaporation of solvents and the residue was coevaporated with toluene.
  • the solid residue was adsorbed on silica gel in a mixture of cyclohexane/acetone and purified by flash chromatography on silica gel (chloroform to 20% methanol), followed by purification on HPLC (C18, gradient H 2 O/McOH as eluent). Purified product was lyophilized from dioxane, yielded 112 mg (70%) of the title compound as a white solid.
  • L-phenylalanine isopropyl ester hydrochloride (335 mg, 1.37 mmol, 2 eq.) was added and the flask was sealed with septum, flushed with argon and then dry pyridine (10 ml/1 mmol of the starting compound) and triethylamine (572 ⁇ l, 4.11 mmol, 6 eq.) were added. The mixture was stirred 15 minutes at 70 °C and a solution of triphenylphosphine (1.08 g, 4.11 mmol, 6 eq.) and AldrithiolTM-2 (905 mg, 4.11 mmol, 6 eq.) in dry pyridine (10 ml/1 mmol of the starting compound) was added.
  • step 1 The coupling product from Example 69 (step 1) (444 mg, 0.6570 mmol, 1 eq.) was dissolved in trifluoroacetic acid (2 ml/100 mg of the material) at room temperature and stirred for 15 minutes. Trifluoroacetic acid was evaporated, coevaporated with water (2x) and a cold solution of ammonia in ethanol (2 ml/100 mg of the material) was added. The mixture was evaporated to dryness and the solid was adsorbed on silica gel in a mixture of cyclohexane/acetone. Purification by flash chromatography on silica gel (chloroform to 15% methanol) afforded 243 mg (81%) of the diisopropyl phosphonate.
  • the solid residue was adsorbed on silica gel in a mixture of cyclohexane/acetone and purified by flash chromatography on silica gel (chloroform to 10% methanol), followed by purification on HPFC (Cellulose SC, heptane/ethanol (80:20) as the eluent). Purified product was lyophilized from dioxane, yielded 69 mg (47%) of the title compound as a white solid.
  • the coupling product from Example 70 (455 mg, 0.6615 mmol, 1 eq.) was dissolved in trifluoroacetic acid (2 ml/100 mg of the material) at room temperature and stirred for 15 minutes. Trifluoroacetic acid was evaporated, coevaporated with water (2x) and a cold solution of ammonia in ethanol (2 ml/100 mg of the material) was added.
  • the solid residue was adsorbed on silica gel in a mixture of cyclohexane/acetone and purified by flash chromatography on silica gel (chloroform to 10% methanol), followed by purification on HPLC (Cellulose SC, heptane/ethanol (80:20) as the eluent). Purified product was lyophilized from dioxane, yielded 31 mg (21%) of the racemic title compound as a white solid.
  • Tetraethyl methylene bisphosphonate (152 ⁇ l , 0.608 mmol, 1.2 eq.) and potassium tert-butoxidc (68 mg, 0.608 mmol, 1.2 eq.) were stirred in THF (8 mL) at 25 °C for 10 min. Then, compound Llll (180 mg, 0.405 mmol) was added dropwise in THF (7 mL) and the mixture was stirred at 25 °C for 30 min. The reaction mixture was diluted by saturated aqueous NH4CI (30 mL) and extracted by ethyl acetate (3x). Organic layer was dried by Na2S04 and evaporated.
  • hPNP recombinant human PNP protein
  • E. coli E. coli
  • purified by means of affinity chromatography NiNTA column, Thermo Fisher Scientific, Waltham, USA
  • 20 mM phosphate buffer pH 7.4 20 mM phosphate buffer pH 7.4 containing 0.3 M NaCl in aliquots at -80 °C.
  • All newly synthesized compounds were dissolved either in water or DMSO to yield 10 mM stock solutions.
  • the compounds then underwent basic screening at 10 mM concentration and in case that at least 50% inhibition was observed, dose-response curve was generated to calculate IC50 value (half -maximal inhibitory concentration).
  • Forodesine (MedChemExpress, Monmouth Junction, USA) was used as a reference compound.
  • Reaction mixture for PNP activity determination consisted of 1 mM P i , 200 pM [2,8- 3 H] inosine (ARC Inc., Saint Fouis, USA), variable concentrations of tested compound, 1 mM DTT, 0.2 pg/ml BSA and 1.75 pg of PNP protein.
  • Reaction was incubated for 10 min at 37°C and it was terminated by spotting a 2-pl aliquot of PEI-coated cellulose TEC plate (NO. 105579, Merck Darmstadt, Germany). The plate was developed in «-butanol-acetic acid-water at 10: 1 :3 ratio, let dry and analyzed by means of radio-TEC scanner RITA Star (Elysia-Raytest GmbH, Strauberhardt, Germany).
  • T-lymphoblastic cell line was employed (CCRF-CEM) representing the target tissue.
  • CCRF-CEM promyelocytic leukemia cell line
  • All cell lines were from ATCC (Manassas, VA, USA).
  • the cells were incubated in RPMI-1640 or DMEM culture medium containing 10% FBS and 1% GlutaMax at 37°C under the atmosphere containing 5% CO 2 .
  • Experiments were performed on the cells between passage No. 10 and 50.
  • cells were seeded in 384-well transparent plates (Brand GmbH, Wertheim, Germany) at a concentration 2,000 - 10,000 cells per well and left to rest in an incubator overnight.
  • PBMC Peripheral blood mononuclear cells
  • Table 2 Evaluation of kinetic solubility of selected PNP inhibitors in TRIS buffer at pH 7.4, in comparison with known compounds, in particular Peldesine. Peldesine failed in Phase III of clinical studies, likely due to its low solubility and low bioavailability (J. Am. Acad. Dermatol. 2001, 44 (6), 940).
  • the compounds of the present invention have a significantly higher solubility and bioavailability.
  • Structurally similar compounds exhibit similar physico-chemical properties (i.e. solubility).

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Abstract

The present invention relates to new compounds of general formula I, their synthesis, their pharmaceutically acceptable salts, and their use in treatment of T-cell acute lymphoblastic leukemia and lymphoma.

Description

Inhibitors of purine nucleoside phosphorylase - synthesis and use thereof for treatment of T-cell acute lymphoblastic leukemia and lymphoma
Field of art
The present invention concerns new compounds for application in the treatment of T-cell acute lymphoblastic leukemia and lymphoma, their synthesis, and their use in vitro and in vivo.
Background art
The treatment of T-cell acute lymphoblastic leukemia (T-ALL) did not undergo such rapid development as other cancer treatments in past years. Because of this, new and innovative treatments and treatment procedures are needed to increase prognosis of T-ALL patients. Espetially prognosis of adult T-ALL patients are very poor and the overall five-year survival rate is only approximately 40%.
Current therapies are based on application of high dosages of classic chemotherapeutics like hyper- CVAD (drug combination of cyclophosphamide, vincristine, adriamycine, dexamethasone in combination with high dosages of methotrextate and cytarabine), a combination of pegaspargase and methotrextate and combinations of these therapies with nelarabine [Litzow, M. R.; Ferrando, A. A. Blood. 2015, 126, 833-841] . These treatment procedures cause severe adverse effects (hair loss, edema, life-threathing infections) which affect patients’ quality of life and the success of the treatment. Potential of purine nucleosid phosphorylase (PNP) inhibitors for a treatment of blood cancers is known for a long time [Krenitsky, T. A., et al, J. Biol. Chem. 1968, 265 (6), 3066-3069], However, inhibitors published to this date failed in preclinical or clinical studies mostly due to poor pharmacodynamic or pharmacokinetic properties.
Peldesine failed in a phase III clinical study as it did not show better efficacy than placebo. This failure was blamed to poor solubility and permeability of the compound [Duvic, M., et ak, J. Am. Acad. Dermatol. 2001, 44 (6), 940-947],
Experimental compound CI-972 failed in clinical studies from unknown reasons. [Gilbertsen, R. B., et al. , Biochem. Biophys. Res. Commun. 1991, 178 (3), 1351-1358],
Other experimental peldesine-derivatives based on 9-deazapurines exhibited similar potency with very low aqueous solubility [Montgomery, J., et al., Med. Chem. 1993, 36 (1), 55-69],
Experimental compounds based on natural purine bases and benzylphosphonate moiety exhibited low inhibitory activity but better aqueous solubility. [Halazy, S., et al., Tetrahedron 1996, 52 (1), 177-184.]. Experimental compounds based on peldesine containing heteroatom between 9-deazapurine base and phenyl moiety exhibited higher inhibitory activity with extremely low aqueous solubility [Morris, P. E., et al., Nucleosides, Nucleotides and Nucleic Acids. 2000, 19 (1-2), 379-404.]. Most of the published compounds exhibited very poor water solubility, therefore, the compounds were not suititable for pharmaceutical application as therapeutic substances.
In 2017, forodesine hydrochloride (Mundesine), a PNP inhibitor, was approved for T-cell lymphoma treatment by PMDA. Forodesine hydrochloride exhibits high inhibitory activity against PNP and T-cell cancer cell lines [Kicska, G., et al, Proc. Natl. Acad. Sci. U. S. A. 2001, 98 (8), 4593-4598], However, forodesine hydrochloride exhibits poor oral bioavailability which is less than 10% in human and nonhuman primates [Kilpatrick, J. M., et al.. Int. Immunopharmacol. 2003, 3 (4), 541-548],
The reported experimental compounds were mostly prepared by ineffective linear synthetic sequences which were based on gradual constructions of the natural or modified purine bases. Such approach did not allow synthesis of complex structures containing sensitive and/or reactive functional groups which could improve properties of the PNP inhibitors.
From the above references it is apparent that in the field of PNP inhibitors, and their therapeutic applications for T-ALL treatment, there is ongoing search for novel compounds that would combine high efficacy in vitro and in vivo with good pharmacokinetic properties, especially high solubility in pharmaceutically relevant solvents and high bioavailability using various modes of medical administrations. Also, there is a need for innovative synthetic procedures that would allow synthesis of more complex inhibitors.
Disclosure of the invention
This invention describes novel purine nucleoside phosphorylase inhibitors of general formula I, including pharmaceutically acceptable salts, their preparation and use in the therapy of T-cell acute lymphoblastic leukemia (T-ALL) and lymphomas.
Although several compounds exhibiting high inhibitory activity againts PNP were published in the past, we were unable to reproduce most of those results. Previously published results could have been affected by wrongly chosen experimental methods. This could be the reason why these compounds were never further developed as potential therapeutics, or they failed in clinical studies. As example, we compare some of the reported compounds with our new compounds in our assays in the Table 1 (compounds L9, L114 and peldesine).
Due to innovative synthesis, we were able to combine structure features which were not possible to combine using previously published procedures. Newly developed compounds exhibit improved inhibition of the PNP enzyme and improved pharmacokinetic properties. One of skill in the art will recognize pharmacokinetic properties as a combination of physico-chemical properties that allow desired distribution of substances in biological systems. Such properties including, but not limited to, solubility, stability, permeability, bioavailability, etc. Combination of these properties allows various routes of administrations for compounds of the general formula I, including, but not limited to peroral, intravenous, topical, etc.
One aspect of the present invention is compounds of the following general formula I:
Figure imgf000004_0001
wherein:
X is independently -NH- or -S- in combination with Ar, Z, G and R;
Ar is independently aryl or heteroaryl in combination with X, Z, G and R;
Z is omitted or it is independently -CH2-, -CH2O-, -OCH2- -CH2OCH2- and -CH=CH- attached independently to the position 2- or 3- of the Ar relative to the X, in combination with X, Ar, G and
R;
G is a group selected from -COA1, -SO2A1, -P(O)(A1)(A2), -OH and -H in combination with X, Ar, Z and R, wherein:
A1 and A2 are identical or different groups and consist independently of -OH, -OR1 and -NHR2, wherein
R1 is independently linear or branched C1-C6 alkyl chain, C6-C12 aryl, arylalkyl and - CH2OC(O)R2 and
R2 is independently linear or branched C1-C10 alkyl chain and -CHR3COOR4, wherein R3 is independently linear or branched C1-C10 alkyl chain in which any -CH2- group can be replaced independently by -S-, -O-, -NH-, C6-C12 aryl and C6-C12 arylalkyl, and R4 is independently linear or branched C1-C10 alkyl chain;
R is -H or a group selected from -OH, -O(aryl), -O(arylalkyl), -C(O)-, -CN and halogen in combination with X, Ar, Z and G; with the proviso that when Ar is phenyl and Z is not present, then R and G are not both a hydrogen atom, and their pharmaceutically acceptable salts.
Preferably, G is a group selected from -COA1, -SO2A1, -P(O)(A1)(A2) and -OH; more preferably G is a group selected from -COA1, -SO2A1 and -P(O)(A1)(A2).
A person skilled in the art to which the invention pertains will understand the terms used herein as: alkyl is a linear or branched hydrocarbon chain; aryl is an aromatic hydrocarbon group consists of 6 to 12 carbon atoms and at least one aromatic ring. It can be unsubstituted or substituted with one or more groups, including -OH, -O(aryl), - O(arylalkyl), -C(O)-, -CN and halogen; particularly preferably, aryl is phenyl, optionally substituted; heteroaryl is an aromatic hydrocarbon group consisting of 4 to 11 carbon atoms and at least one aromatic ring and at least one heteroatom included O, N, S. It can be unsubstituted or substituted with one or more groups, including -OH, -O(aryl), -O(arylalkyl), -C(O)-, -CN and halogen; particularly preferably, heteroaryl is pyridyl, pyrimidinyl, furanyl, thiophenyl, optionally substituted; arylalkyl is an alkyl group that contains one or more aryl groups which can be unsubstituted or substituted with one or more function groups, including -OH, -O(aryl), -O(arylalkyl), -C(O)-, - CN and halogen.
Pharmaceutically acceptable salts include salts with alkali metals, salts with inorganic or organic anions and salts with inorganic or organic cations. Such salts exhibit similar biological efficacy and similar properties as the claimed compounds of general formula I and which can be used for application on human or mammal tissues without adverse effects, including, but not limited to irritation, allergic reaction, etc. A person skilled in the art to which the invention pertains will be able to identify which salts are pharmaceutically acceptable, especially, salts with one or more benefitial properties, including, but not limited to stability at various temperatures and humidity, solubility in water or oil, toxicity. List of suitable salts can be found in Remington's Pharmaceutical Sciences, 20th edition, Mack Publishing Company, Easton, Pa., (1985).
In a preferred embodiment, the present invention provides compounds of general formula I, including, but not limited to:
(E)-7-((2-(2-(phosphonato)vinyl)phenyl)thio)-3.5-dihydro-4H-pyrrolo|3.2-d]pyrimidin-4-onc. 7-((2-((phosphonato)methoxy)phenyl)thio)-3.5-dihydro-4H-pyrrolo [3,2-d]pyrimidin-4-one, (E )-7-((4-bromo-2-(2-(phosphonato)vinyl)phenyl)thio)-3,5-dihydro-4H-pyrrolo[3,2-d]pyrimidin-4- one,
(E )- 7 -((5 -bromo-2-(2-(phosphonato)vinyl)phenyl)thio)-3 ,5 -dihydro-4H-pyrrolo [3 ,2-d]pyrimidin-4- one,
(E )- 7 -((5 -hydroxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3 ,5 -dihydro-4H-pyrrolo [3 ,2-d]pyrimidin-4- one,
(E )-7-((4-hydroxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3 ,5 -dihydro-4H-pyrrolo [3 ,2-d]pyrimidin-4- one, (E)-7-((4-methoxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3.5-dihydro-4H-pyrrolo[3.2-d]|pyrimidin-4- one,
(E)-7-((4-isopropoxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3.5-dihydro-4H-pyrrolo[3.2-d]|pyrimidin-
4-one,
(E )-7-((4-(2,3,4,5,6-pentafluorophenyl)methoxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3,5-dihydro-4H-pyrrolo [3 ,2-d]pyrimidin-4-one,
(E )-7-((4-(2,3,4,5,6-pentafluorophenyl)oxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3,5-dihydro-4H- pyrrolo [3 ,2-d]pyrimidin-4-one,
(E )- 7 -((3 -hydroxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3 ,5 -dihydro-4H-pyrrolo [3 ,2-d]pyrimidin-4- one,
(E)-7-((3-methoxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3.5-dihydro-4H-pyrrolo[3.2-d]|pyrimidin-4- one,
(E)-7-((3-isopropoxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3.5-dihydro-4H-pyrrolo[3.2-d]|pyrimidin-
4-one,
(E )-7-((3-(2,3,4,5,6-pentafluorophenyl)methoxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3,5-dihydro- 4H-pyrrolo [3 ,2-d]pyrimidin-4-one,
(E )-7-((3-(2,3,4,5,6-pentafluorophenyl)oxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3,5-dihydro-4H- pyrrolo [3 ,2-d]pyrimidm-4-one,
(E)-7-((5-fluoro-2-((phosphonato)methoxy)phenyl)thio)-3.5-dihydro-4H-pyrrolo[3.2-d]|pyrimidin-4- one,
(E)-7-((3-methoxy-2-((phosphonato)methoxy)phenyl)thio)-3.5-dihydro-4H-pyrrolo[3.2-d]|pyrimidin-4- one,
(E )- 7 -((3 -(2-(phosphonato)vinyl)thiophene-2-yl)thio)-3.5-dihydro-4H-pyrrolo [3.2-d]pyrimidin-4-onc. 7-((3-((phosphonato)methoxy)thiophene-2-yl)thio)-3.5-dihydro-4H-pyrrolo[3.2-d]|pyrimidin-4-onc. 7-((2-((sulfo)methoxy)phenyl)thio)-3 ,5 -dihydro-4H-pyrrolo [3 ,2-d]pyrimidm-4-one,
(E )-7 -((2-(2-(sulfo)vinyl)phenyl)thio)-3 ,5 -dihydro-4H-pyrrolo [3 ,2-d]pyrimidin-4-one, 7-((2-(3-phosphonato-2-oxapropyl)phenyl)thio)-3.5-dihydro-4H-pyrrolo[3.2-d]|pyrimidin-4-onc. 7-((2-(((((S)-1-(isopropoxycarbonyl)cthyl)amino)(phcnoxy)phosphoiyl)methoxy)phenyl)thio)-3.5- dihydro-4H-pyrrolo[3.2-d]|pyrimidinc-4-on.
(E )- 7 -((2-(2-(phosphonato)vinyl)phenyl)amino)-3 ,5 -dihydro-4H-pyrrolo [3 ,2-d]pyrimidine-4-one .
The subject of the present invention are compounds of the general formula I and/or pharmaceutically acceptable salts for use as a medication.
The subject of the present invention are compounds of the general formula I and/or pharmaceutically acceptable salts for use as a treatment of diseases caused by abnormal growth of T-lymphocytes. The subject of the present invention are compounds of the general formula I and/or pharmaceutically acceptable salts for use in a treatment of T-cell acute lymphoblastic leukemia. The present invention also provides a pharmaceutical composition comprising therapeutically effective amount of at least one compound of formula I and optionally at least one pharmaceutically acceptable excipient as carrier, fdler, emulgator and/or diluent.
The subject of the present invention also provides a pharmaceutical composition for use in the treatment of diseases caused by abnormal growth of T-lymphocytes.
In particular, the invention provides a pharmaceutical composition for use in the treatment of T-acute lymphoblastic leukemia. Presented compounds exhibit significant and selective in vitro cytotoxic effects against leukemic T- lymphocytes. Presented compounds also exhibit good solubility in biologically relevant solvents and good permeability through biological membranes.
The presented invention also describes synthesis of compounds of the general formula II and III according to the scheme 1, scheme 2 and scheme 3,
Figure imgf000007_0001
Scheme 3 wherein:
Ar is independently aryl or heteroaryl in combination with X, Z, G and R;
Z is excluded or it is independently -CH2-, -CH2O-, -CH2OCH2- and -CHCH- attached independently to the position 2- or 3- of the Ar relative to -SH or I, in combination with Ar, G and
R;
G in independently -H, -COA1, -SO2A1, -P(O)(A1)(A2) and -OH in combination with Ar, Z and R, wherein:
A1 and A2 are identical or different and consist independently of -OH, -OR1 and -NHR2, wherein R1 is independently linear or branched C 1 -G, alkyl chain, C6-C12 aryl and -CH2OC(O)R2 and
R2 is independently linear or branched C1-C10 alkyl chain and -CHR3COOR4, wherein R3 is independently linear or branched C1-C10 alkyl chain in which any -CH2- group can be replaced independently by -S-, -0-, -NH-, C6-C12 aryl and C6-C12 arylalkyl, and R4 is independently linear or branched C1-C10 alkyl chain;
R is independently -H, -OH, -O(aryl), -O(arylalkyl), -C(O)-, -CN and halogen in combination with Ar, Z and G;
P1 is a protecting group selected from methyl, ethyl, tert- butyl, benzyl, halogenbenzyl, benzoyl, trityl, tosyl, acetyl;
P2 is a protecting group selected from terc- butyl, benzyl, halogenbenzyl, benzoyl, trityl, tosyl, acetyl, pivaloyl, benzyloxycarbonyl, benzyloxymethyl, p-methoxybenzylcarbonyl. tert- butyloxy carbonyl, 2-trimethylsilylethyloxymethyl .
Reaction conditions include: a protic or an aprotic solvent selected from methanol, ethanol, «-propanol, 2-propanol, «-butanol, 2- butanol, toluene, acetone, dimethylformamide, dioxane, tetrahydrofuran, ethyl acetate, butyl acetate, dimethylsulfoxide, diethylether, N-methylpyrrolidonc. chloroform, dichlormethane, 1,2- dichloroethane, benzene, ere sole; temperature in the range 0 °C to 250 °C; an organic or an inorganic base selected from sodium carbonate, potassium carbonate, cesium carbonate, sodium bicarbonate, sodium phosphate, potassium phosphate, sodium hydroxide, potassium hydroxide, lithium hydroxide, triethylamine, trimethylamine, diisopropylamine, diisopropylethylamine, dimethylaminopyridine, 2,6-lutidine; catalysis with organic or inorganic copper salt selected from copper(I) fluoride, copper(I) chloride, copper(I) bromide, copper(I) iodide, copper(I) acetate, copper(I) benzoate; if necessary, presence of a ligand selected from tetramethylethylendiamine, 1,10-phenantroline, 4,7- dimethyl- 1 , 10-phenantroline, 5,6-dimethyl- 1 , 10-phenantroline, 3,4,7,8-tetramethyl- 1,10- phenantroline, 2-isobutyrylcyclohexanone, 2-bipyridyl;
The presented invention also describes synthesis of compounds of the general formula IV according to scheme 4,
Figure imgf000009_0001
Scheme 4 wherein:
Z is not present or it is independently -CH2-, -CH2O-, -CH2OCH2- and -CH=CH- attached independently to the position 2- or 3- of the phenyl relative to -NH2, in combination with G and R; G in independently -H, -COA1, -SO2A1, -P(O)(A1)(A2) and -OH in combination with Z and R, wherein:
A1 and A2 are identical or different and consist independently of -OH, -OR1 and -NHR2, wherein R1 is independently linear or branched C1-C6 alkyl chain, C6-C12 aryl and -CH2OC(O)R2 and
R2 is independently linear or branched C1-C10 alkyl chain and -CHR3COOR4, wherein R3 is independently linear or branched C1-C10 alkyl chain in which any -CH2- group can be replaced independently by -S-, -0-, -NH-, C6-C12 aryl and C6-C12 arylalkyl, and R4 is independently linear or branched C1-C10 alkyl chain;
R is independently -H, -OH, -O(aryl), -O(arylalkyl), -C(O)-, -CN and halogen in combination with Z and G;
P1 is a protecting group selected from methyl, ethyl, tert- butyl, benzyl, halogenbenzyl, benzoyl, trityl, tosyl, acetyl; P3 is a protecting group selected from terc-butyl, benzyl, halogenbenzyl, benzoyl, trityl, tosyl, acetyl, pivaloyl, benzyloxycarbonyl, benzyloxymethyl, p-methoxybenzylcarbonyl, terc- butyloxycarbonyl, 2-trimethylsilylethyloxymethyl;
Reaction conditions include: a protic or an aprotic solvent selected from methanol, ethanol, «-propanol, 2-propanol, «-butanol, 2- butanol, toluene, acetone, dimethylformamide, dioxane, tetrahydrofuran, ethyl acetate, butyl acetate, dimethylsulfoxide, diethylether, N-methylpyrrolidone. chloroform, dichlormethane, 1,2- dichloroethane, benzene, ere sole; temperature in the range 0 °C to 250 °C; an organic or an inorganic base selected from sodium carbonate, potassium carbonate, cesium carbonate, sodium bicarbonate, sodium phosphate, potassium phosphate, sodium hydroxide, potassium hydroxide, lithium hydroxide, triethylamine, trimethylamine, diisopropylamine, diisopropylethylamine, dimethylaminopyridine, 2,6-lutidine; catalysis with suitable palladium form selected from palladium(II) chloride, palladium(II) acetate, tris(dibenzylidenacetone) dipalladium, so-called precatalysts known as [4-(di-tert-butylphosphine)- N, N-dimethylaniline-2-(2'-aminobiphenyl)|palladium(II). [(R)-(+)-2.2'-bis(diphenylphosphinc)- 1, 1'-binaphtyl]palladium(II) chloride, [(2-di-cyclohexylphosphine-3,6-dimethoxy-2',4',6'- triisopropyl- 1 , 1 '-biphenyl)-2-(2'-amino- 1 , 1 '-biphenyl)]palladium(II) [2-(dicyclohexylphosphine)- 2'-(N,N-dimethylamino)- 1 , 1 '-biphenyl](2'-amino- 1 , 1 '-biphenyl -2 -yl)palladium(II), bis(tri-tert- butylphosphine)-palladium(O), |(2-dicyclohexylphosphine-2'.6'-bis(N,N-dimethylamino)- 1. G- biphenyl)-2-(2'-amino-l,r-biphenyl)]palladium(II), (2-dicyclohexylphosphine-l,l'-biphenyl)[2-(2'- amino- 1 , 1 '-biphenyl)]palladium(II), 2 -dicyclohexyl phosphine-2-(N,N-dimethylamino)-biphenyl(2'- amino- 1 , 1 '-biphenyl -2 -yl)palladium(II), [1,1 '-bis(diphenylphosphine)- ferrocenejdichloro- palladium(II), (2-(di- 1 -adamantylphosphine)morpholinobenzene)[2-(2'-amino- 1,1'- biphenyl)]palladium(II), [(di-tert-butylneopentylphosphine)-2-(2-aminobiphenyl)]palladium(II), [(tricyclohexylphosphine)-2-(2'-aminobiphenyl)]-palladium(II), (2-dicyclohexylphosphine-2',6'- di/isopropoxy- 1 , 1 '-biphenyl)[2-(2'-amino- 1 , 1 '-biphenyl)]palladium(II), (2-dicyclohexylphosphine- 2',6'-dimethoxybiphenyl) [2-(2'-amino-l,r-biphenyl)]palladium(II), [(4,5-bis(diphenylphosphine)- 9,9-dimethylxanthene)-2-(2'-amino- 1 , 1 '-biphenyl)]palladium(II), (2-dicyclohexylphosphine-
2'.4'.6'-tri/.vopropyl- 1 , 1 '-biphenyl)|2-(2'-amino- 1. 1 '-biphenyl) |palladium(II). (di-tert- butylphosphinc-3.4.5.6-tctramethyl-2'.4'.6'-triisopropyl- 1 , 1 -biphenyl)(2'-amino- 1 , 1 '-biphenyl-2 - yl)-palladium(II), |2-(di- l-adamantylphosphinc)-2'.4'.6'-triisopropyl-3.6-dimethoxybiphenyl ][2-(2'- amino- l,r-biphenyl)]palladium(II), [(di-tert-butylphosphine-2',4',6'-triisopropyl-l,l'-biphenyl)- 2-(2'-amino- 1 , 1 '-biphenyl)]palladium(II), [(2- {bis [3 ,5 -bis(trifluoromethyl)phenyl] - phosphine } -3 ,6- dimethoxy-2',4',6'- triisopropyl- 1. 1 '-biphenyl )-2-(2'-amino-1, 1'-biphenyl)]palladium(II), (2-(di-1- adamantylphosphine)morpholinobenzene)[2-(2'-amino-l,l'-biphenyl)]palladium(II), 2-(di -tert- butylphosphine)-2', 4', 6'-triisopropyl-3,6-dimethoxy-l,l'-biphenyl)-2-(2'-amino-1,1'-biphenyl)]- palladium (II), [tri(o-tolyl)-phosphine] [2-(2'-amino- 1 , 1 '-biphenyl)]palladium(II); if necessary, presence of a ligand selected from [4-(dimethylamino)phenyl]bis(tert-butyl)phosphine, 2, 2'-bis(diphenylphosphine)- 1,1 '-binaphthalene, 2-(dicyclohexylphosphine)3,6-dimethoxy-2',4',6'- triisopropyl- 1, 1 '-biphenyl, 2'-(di-tert-butyl- phosphine)-A, A-dimethylbiphenyl-2-aminoe, tri -tert- butylphosphine, 2-dicyclohexyl- phosphine-2', 6'-bis(A,A-dimethylamino)biphenyl, 2-
(dicyclohexylphosphine)biphenyl, 2-dicyclohexylphosphine-2'-(A A-dimethylamino)biphenyl, 1,1'- ferrocenediyl-bis(diphenylphosphine), (di-tert-butylneopentylphosphine, tricyclohexylphosphine, 2- dicyclohexylphosphine-2'.6'-di/isopropoxybiphenyl. 2-dicyclohexylphosphine-2',6'- dimethoxybiphenyl, di-to'/-butylmethyl phosphine. di-tert-butylphenylphosphine, 4,5- bis(diphenylphosphine)-9,9-dimethylxanthene, 4,6-bis(diphenylphosphine)- 10A-phcnoxazinc. 2- dicyclohcxylphosphinc-2'.4'.6'-triisopropylbiphenyl. di-tert- butylphosphine-3.4.5.6-tetramethyl- 2', 4', 6'-triisopropyl- 1,1 -biphenyl, 2-(di- 1 -adamantylphosphinc)-2'.4'.6'-triisopropyl-3.6- dimethoxybiphenyl, 2-di-tert-butylphosphine-2',4',6'-triisopropylbiphenyl, 2-{bis[3,5- bis(trifluoromethyl)phenyl |-phosphinc}-3.6-dimethoxy-2'.4'.6'-triisopropyl- 1.1 '-biphenyl. 2-(di- tert-butylphosphine)-2',4',6'-triisopropyl-3,6-dimethoxy- 1 , 1 '-biphenyl, tris(o-tolyl)phosphine, tri- tert-butylphosphine, triphenylphosphine, s vyhodou 2-dicyclohexylphosphine-2',4',6'- triisopropylbiphenyl .
The range of the invention is not limited by the following examples.
Examples
List of Abbreviation:
TLC Thin Layer Chromatography
HPLC High Pressure Liquid Chromatography
C18 Dodecyl
HR-MS High Resolution Mass Spectrometry
MS Mass Spectrum
ESI Electrospray Ionization
Cl Chemical Ionization aq. Aqueous solution
THF Tetrahydrofuran
DMF A. A- d i m e t h y 1 fo rm am i de
DMSO Dimethylsulfoxide Me Methyl
Et Ethyl
Pr Propyl
7-Pr iso-Propyl
Ph Phenyl
Bn Benzyl
/-Bii terc-Butyl
SEM 2-(Trimethylsilyl)ethoxymethyl
Boc terc-Butoxy carbonyl
Tf Triflyl
Ts Tosyl
TMS Trimethylsilane dba Dibenzylidenacetone
Bn-F5 Pentafluorobenzyl; perfluorobenzyl
Ph-F5 2,3,4,5,6-Petrafluorophenyl; perfluorophenyl
IC50 The half maximal inhibitory concentration riPNP Human Purine Nucleoside Phosphorylase
ND Not Determine
Ni-NTA Nickel-charged affinity resin
RPMI Growth medium used in cell culture
DMEM Dulbecco's Modified Eagle Medium
FBS Fetal Bovine Serum
PBMC Peripheral blood mononuclear cel
Unless otherwise stated, solvents were evaporated at 40 °C/2 kPa and prepared compounds were dried at 30 °C at 2 kPa. Starting compounds and reagents were purchased from commercial suppliers (Sigma- Aldrich, Fluorochem, Acres Organics, Carbosynth, TCI) and used without further purification or were prepared according to the published procedures.
Diethyl ether, tetrahydrofuran, dioxane and acetonitrile were dried by activated neutral alumina (drysphere®). Dimethylformamide was dried by activated molecular sieves (3Å). Other dry solvents were purchased from commercial suppliers (Sigma- Aldrich, Acros Organics). Triethylamine was dried with potassium hydroxide under an argon atmosphere in a dark flask sealed with septum.
Microwave experiments were performed in 10 mF or 30 ml vials with a CEM Discover (Explorer) microwave apparatus operating at a frequency of 2.45 GHz with continuous irradiation power from 0 W to 300 W. Large-scale reactions were carried-out in the Syrris Atlas Potassium system with 21, 1 1 and 0.5 1 jacket reactor coupled with Julabo FP50-HL Refrigerated/Heating Circulator.
Analytical TLC was performed on silica gel pre-coated aluminium plates with fluorescent indicator (Merck 60 F254). Flash column chromatography was carried out by Teledyne ISCO CombiFlash Rf200 with dual absorbance detector. Various types of columns were used: a) Teledyne ISCO columns RediSepRf HP Silica GOLD in sizes 12 g, 40 g, 80 g and 120 g; b) Teledyne ISCO columns RediSepRf HP C18 Aq GOLD in sizes 50 g and 100 g; c) column Chromabond Flash DL 40, DL 80, DL 120 and DL 200, fdled with FLUKA silica gel 60; d) Interchim puriFlash C18 Aq in sizes F0040 and F0080. Preparative HPLC purifications were performed on Waters Delta 600 chromatography system with columns packed with Cl 8 reversed phase resin (Phenomenex Gemini 10 μm 21 x 250 mm, Phenomenex Gemini 5 μm 21 x 250 mm, Phenomenex Luna 10 μm 21 x 250 mm) using gradient H20/MeOH as eluent.
Mass spectra, UV spectra and purity of compounds were measured on Waters UPLC-MS system consisted of Waters UPLC H-Class Core System (column Waters Acquity UPLC BEH C18 1.7 mm, 2.1 x 100 mm), Waters Acquity UPLC PDA detector and Mass spectrometer Waters SQD2. The universal LC method was used (eluent water/acetonitrile, gradient 0% - 100%, run length 7 min) and MS method (ESI+ and/or ESI-, cone voltage = 30 V, mass detector range 100 - 1000 Da).
High-resolution mass spectra were measured on LTQ Orbitrap XL spectrometer (Thermo Fisher Scientific).
NMR spectra were recorded on Bruker Avance 400 or 500 spectrometers referenced to the residual solvent signal or a specified additive.
Dowex® 50D resin was turned to Na+ cycle by treatment of Dowex D50 resin in H+ cycle with 1M NaOH aq. solution, followed by water wash to neutral pH.
Example 1
4-(Benzyloxy)-5H-pyrrolo 13.2-d|py rimidinc L 1
Figure imgf000013_0001
The jacket reactor (21) was flushed with nitrogen, charged with benzyl alcohol (1 1) and the system was set to retain temperature 20 °C. Sodium metal (22.5 g, 977 mmol, 1.5 eq.) was added in portions and the mixture was stirred 20 hours under small flow of nitrogen. The mixture was then heated at 80 °C for 1 hour, and then it was cooled back to 20 °C. 4-Chloro-5H-pyrrolo[3.2-d]|pyrimidinc (100 g, 651 mmol, 1 eq.) was charged and the mixture was stirred at 80 °C until complete conversion was achieved (ca. 4 hours). The mixture was cooled to 5 °C, diluted with water (400 ml), pH was adjusted to 7 with 3M HC1 (aq.) (ca. 100 - 130 ml). The mixture was heated to 20 °C, extracted with chloroform (3 x 400 ml) and the organic phase was washed with brine (500 ml). The mixture was concentrated and benzyl alcohol was evaporated at high vacuum at ca. 90 °C. The solid was filtered through a short pad of silica gel (600 g) with eluent (100% of chloroform, then chloroform with 5% methanol). Solvents were evaporated and the residue was just dissolved in a refluxed mixture of ethyl acetate/methanol (1: 1) and the solution was cooled to 30 °C. Then an antisolvent pentane (same volume as the mixture) was slowly added and the mixture was cooled to -20 °C within 3 hours. Then, it was stirred overnight and during that time, the product crystallized. Crystals were collected, washed with pentane and dried, yielded 141 g (96%) of 4- (benzyloxy)-5H-pyrrolo[3.2-d]|pyrimidine as white crystals. 1H NMR (401 MHz, Chloroform-d) δ 9.33 (s, 1H), 8.58 (s, 1H), 7.49 - 7.43 (m, 2H), 7.41 (m, 1H), 7.38 - 7.32 (m, 2H), 7.35 (s, 1H), 6.67 (dd, J = 3.2, 2.1 Hz, 1H), 5.59 (s, 2H). 13C NMR (101 MHz, Chloroform -d) δ 155.46, 150.55, 150.11, 136.22, 128.76 - 128.28 (m), 115.14, 103.31, 67.94. MS (ESI+) m/z = 226.1 [M + H]+.
Example 2
4-(Benzyloxy)-7-iodo-5-((2-(trimethylsilyl)ethoxy)methyl)-5H-pyrrolo[3.2-d]|pyrimidine L2
Figure imgf000014_0001
The jacket reactor (1 1) was flushed with nitrogen, 4-(benzyloxy)-5H-pyrrolo[3.2-d]|pyrimidine (LI) (40 g, 178 mmol, 1 eq.) was dissolved in tetrahydrofuran (500 ml) and /v'-iodosuccinimide (44 g, 195 mmol, 1.1 eq.) was added. The mixture was stirred at room temperature for 1 hour, during which the product crystalized. Crystals were collected, washed with tetrahydrofuran and dried, yielded 56 g (90%) of the crude 9-iodo compound as a white solid, which was used in the next step without further purification and characterization. The jacket reactor (2 1) was flushed with nitrogen and 4-(benzyloxy)-7-iodo-5H- pyrrolo[3.2-d]|pyrimidinc (56 g, 159 mmol, 1 eq.) was dissolved in dry dimethylformamide (560 ml). The mixture was cooled to -5 °C and sodium hydride (8 g, 199 mmol, 1.25 eq.) was added portionwise under small flow of nitrogen, keeping the temperature under 5 °C. The mixture was stirred at 20 °C for 1 hour. 2-(Trimethylsilyl)ethoxymethyl chloride (35.3 ml, 199 mmol, 1.25 eq.) was added dropwise and the mixture was stirred at 20 °C for 1 hour. The reaction was quenched with a half-saturated aqueous solution of NH4CI (560 ml), extracted with ethyl acetate (3x 300 ml), washed with brine (lx 500 ml) and dried with MgSO4. Solvents were evaporated and the solid was filtered through a short pad of silica gel (300 g) with eluent (100% of chloroform, then chloroform with 1% methanol). Solvents were evaporated and the residue was just dissolved in a refluxed acetonitrile. The solution was slowly cooled to 20 °C within 3 hours, during which the product crystallized. Crystals were collected, washed with acetonitrile and dried, yielded 66 g (86%) of the title compound as white flakes. 1H NMR (401 MHz, Chloroform -d) δ 8.65 (s, 1H), 7.51 (s, 1H), 7.51 - 7.45 (m, 2H), 7.44 - 7.31 (m, 3H), 5.63 (s, 2H), 5.62 (s, 2H), 3.73 - 3.17 (m, 2H), 0.97 - 0.65 (m, 2H), -0.09 (s, 9H). 13C NMR (101 MHz, Chloroform-d) δ 155.90, 151.86, 151.05, 136.44, 136.11, 128.80, 128.52, 128.33, 115.87, 77.84, 68.55, 66.30, 59.18, 17.80, -1.34. MS (ESI+) m/z = 482.2 [M + H]+. MS (ESI-QMS) m/z: [M + H]+ ealed for C19H25IN302Si, 482.1; found, 482.2.
Example 3
4-(Bcnzyloxy)-7-bromo-5-((tertbutoxycarbonyl))-5H-pyrrolo[3.2-d]|pyrimidinc L3
OBn Boc
Figure imgf000015_0001
The jacket reactor (1 1) was flushed with nitrogen and 4-(benzyloxy)-5H-pyrrolo[3.2-d]|pyrimidinc (LI) (40 g, 178 mmol, 1 eq.) was dissolved in tetrahydrofuran (500 ml) and N-bomosuccinimide (34.7 g, 195 mmol, 1.1 eq.) was added. The mixture was stirred at room temperature for 1 hour, during which the product crystalized. The solvent was evaporated to dryness and the crude mixture was used in the next step. The jacket reactor (1 1) was flushed with nitrogen and the crude mixture was dissolved in dry acetonitrile (400 ml). Then, 4-(dimethylamino)pyridine (2.2 g, 18 mmol, 0.1 eq.) was added, followed by slow addition (30 minutes) of a solution of di-tert-butyl dicarbonate (58.2 g, 267 mmol, 1.5 eq.) in dry acetonitrile (100 ml). The reaction was stirred at room temperature until gas release ceased (ca. 2 hours). Solvents were evaporated and the solid was filtered through a short pad of neutral alumina (300 g) in a mixture of cyclohexane/ethylacetate (3: 1, v/v). Solvents were evaporated and the solid was lyophilized from dioxane, yielded 58 g (81%) of the title compound as a white solid. 1H NMR (401 MHz, Chloroform-d) δ 8.68 (s, 1H), 7.92 (s, 1H), 7.56 - 7.48 (m, 2H), 7.43 - 7.27 (m, 3H), 5.65 (s, 2H),
1.54 (s, 9H). 13C NMR (101 MHZ, Chloroform-d) δ 156.47, 152.90, 152.00, 147.23, 136.29, 131.34, 128.58, 128.22, 128.14, 114.26, 96.81, 85.90, 68.68, 27.87. MS (ESI+) m/z = 404.1 [M + H]+. HRMS (ESI-FTMS) m/z: [M + H]+ ealed for C18H19BrN303, 404.0604; found, 404.0603.
Example 4
4-(Benzyloxy)-5-((2-(trimethylsilyl)ethoxy)methyl)-7-(thio)-5H-pyrrolo|3.2-ri| pyrimidine:
1 ,2-Bis(4-(benzyloxy)-5-((2-(tri methyl si lyl)cthoxy)methyl)-5H-pyrrolo|3.2-ri|pyri midin-7- yl)disulfane L4
Figure imgf000015_0002
Compound L2 (4 g, 8.31 mmol, 1.0 eq.), copper(I) iodide (158 mg, 0.83 mmol, 0.1 eq.) and 1,10- phenantroline (299 mg, 1.63 mmol, 0.2 eq.) were charged into a microwave vial. The vial was flushed with argon, and then dry toluene (15 ml) was added, followed by a solution of thiobenzoic acid (1.18 ml, 9.97 mmol, 1.2 eq.) and triethylamine (1.73 ml, 12.5 mmol, 1.5 eq.) in dry toluene (20 ml). The solution turned into a dark-red color. The vial was sealed and inserted into the microwave reactor for 1 h at 120 °C. The dark-brown reaction mixture was dissolved in chloroform and washed with a half- saturated aqueous solution of NaHCO3 (2x), 1M HC1 (aq.) (2x), brine (1x), dried with MgSO4, filtered and evaporated. The residue was adsorbed on silica gel in a mixture of cyclohexane/acetone and purified by flash chromatography on silica gel (chloroform to 5% of methanol), yielded coupling product as a brown oil. The oil residue was dissolved in dry methanol and potassium carbonate (2.30 g, 16.6 mmol, 2 eq.) was added. The mixture was stirred at room temperature until full conversion was achieved (ca. 1 h). The reaction was quenched with water and the mixture was stirred 1 hour in an unsealed flask. The mixture was extracted with chloroform (3x), washed with brine (1x), dried with MgSO4, filtered and solvents were evaporated. Purification by flash chromatography on silica gel (chloroform to 5% of methanol), followed by C18 reverse phase chromatography (water to methanol) afforded 2.5 g (78%) of a yellow oil, which was lyophilized from dioxane to afford the yellow solid product. 1H NMR (400 MHz, Chloroform-d) δ 8.53 (s, 2H), 7.60 (s, 2H), 7.51 (dd, J= 8.1, 1.6 Hz, 4H), 7.44 - 7.29 (m, 8H), 5.64 (s, 5H), 5.61 (s, 4H), 3.45 (t, J= 8.0 Hz, 4H), 0.81 (t, J= 8.1 Hz, 4H), -0.10 (s, 18H). 13C NMR (101 MHz, Chloroform-d) δ 156.12, 151.46, 151.10, 138.34, 136.20, 128.80, 128.51, 128.34, 116.17, 110.52, 77.98, 68.42, 66.41, 17.83, -1.32. HRMS (ESI-FTMS) m/z: [M + H]+ calcd for C38H4904N6S2Si2, 773.2790; found, 773.2780.
General procedure for compounds L5 - L8 (SI)
Compound L2 (200 mg, 0.4154 mmol, 1.0 eq.), copper(I) iodide (8 mg, 0.0415 mmol, 0.1 eq.) and 1,10- phenantroline (15 mg, 0.0830 mmol, 0.2 eq.) were charged into a microwave vial. The vial was flushed with argon, dry toluene (2 ml) and triethylamine (87 μl, 0.6231 mmol, 1.5 eq.) were added. A thiophenol (0.4985 mmol, 1.2 eq.) was added as the last component and the solution turned into a dark-red color. The vial was sealed and inserted into the microwave reactor for 2 h at 120 °C. The dark-brown reaction mixture was dissolved in chloroform, washed with a half- saturated aqueous solution NaHCO3 (2x), 1M HC1 (aq.) (2x), brine (1x), dried with MgSO4, filtered and solvents were evaporated. Purification by flash chromatography on silica gel (cyclohexane to 15% ethyl acetate modified with 10% methanol (v/vi)) afforded the title compound as a white -off oil.
Example 5
4-(Benzyloxy)-7-(phenylthio)-5-((2-(trimethylsilyl)ethoxy)methyl)-5H-pyrrolo[3.2-d]|pyrimidine L5
Figure imgf000016_0001
General procedure SI afforded 150 mg (77%) of the title compound. 1H NMR (401 MHz, Chloroform- ,7) δ 8.68 (s, 1H), 7.68 (s, 1H), 7.55 - 7.47 (m, 2H), 7.44 - 7.33 (m, 3H), 7.24 - 7.14 (m, 4H), 7.12 - 7.03 (m, 1H), 5.67 (s, 4H), 3.47 (t, J= 8.0 Hz, 2H), 0.84 (t, J= 8.3, 7.8 Hz, 2H), -0.09 (s, 9H). 13C NMR (101 MHz, Chloroform -d) δ 156.56, 150.86, 150.75, 138.85, 137.65, 135.85, 128.99, 128.83, 128.64, 128.44, 127.45, 125.83, 116.31, 105.13, 78.07, 68.88, 66.52, 17.82, -1.35. MS (ESI-QMS) m/z: [M + H]+ calcd for C25H30N3O2 SSi, 464.2; found, 464.3.
Example 6
4-(Bcnzyloxy)-7-((pyrimidin-2-yl)thio)-5-((2-(tri methyl si lyl)cthoxy (methyl )-5H-pyrrolo 13.2- d]pyrimidinc L6
Figure imgf000017_0001
General procedure SI afforded 64 mg (33%) of the title compound. 1H NMR (401 MHz, Chloroform- d) δ 8.66 (s, 1H), 8.52 - 8.25 (m, 2H), 7.72 (s, 1H), 7.51 (d, J= 7.0 Hz, 2H), 7.45 - 7.31 (m, 3H), 7.03 - 6.84 (m, 1H), 5.70 (s, 2H), 5.66 (s, 2H), 3.49 (t, J= 8.0 Hz, 2H), 0.85 (t, J= 8.1 Hz, 2H), -0.08 (s, 9H). 13C NMR (101 MHZ, Chloroform-d) δ 172.10, 157.65, 156.59, 150.80, 138.97, 135.85, 128.79, 128.61, 128.42, 117.27, 116.43, 102.09, 78.16, 68.85, 66.51, 17.79, -1.35. MS (ESI-QMS) m/z: [M + H]+ calcd for C23H28N502SSi, 466.2; found, 466.2.
Example 7
4-(Benzyloxy)-7-((2-hydroxymethyl)phenylthio)-5 -((2-(trimethylsilyl)ethoxy)methyl)-5H-pyrrolo [3 ,2- d]pyrimidinc L7
Figure imgf000017_0002
General procedure SI afforded 130 mg (64%) of the title compound. 1H NMR (401 MHz, Chloroform- d) δ 8.51 (s, 1H), 7.70 (s, 1H), 7.47 (dd, J= 7.9, 1.7 Hz, 2H), 7.42 (dd, J= 7.6, 1.5 Hz, 1H), 7.40 - 7.32 (m, 3H), 7.27 (dd, J= 7.8, 1.4 Hz, 1H), 7.17 (ddd, J= 7.4, 1.4 Hz, 1H), 7.08 (ddd, J= 7.5, 1.6 Hz, 1H), 5.63 (s, 2H), 5.60 (s, 2H), 5.03 (s, 2H), 3.45 (t, J= 8.1 Hz, 2H), 1.42 (s, 1H), 0.83 (t, J= 8.1 Hz, 2H), - 0.08 (s, 9H). 13C NMR (101 MHZ, Chloroform-d) δ 156.18, 150.62, 150.50, 142.31, 137.38, 135.83, 134.76, 133.09, 130.01, 128.69, 128.57, 128.47, 128.27, 128.02, 116.16, 107.07, 77.82, 68.56, 66.31, 63.56, 17.72, -1.40. MS (ESI-QMS) m/z: [M + H]+ calcd for C26H32N303SSi, 494.2; found, 494.3. Example 8
4-(Bcnzyloxy)-7-((2-hydroxy)phenylthio)-5-((2-(trimethylsilyl (ethoxy (methyl )-5H-pyrrolo 13.2- d]pyrimidinc L8 General procedure SI afforded 132 mg (66%) of the title compound. 1H NMR (401 MHz, Chloroform - d) δ 8.66 (s, 1H), 7.60 (s, 1H), 7.59 (dd, J= 7.8, 1.6 Hz, 1H), 7.48 (dd, J= 7.9, 1.6 Hz, 2H), 7.44 - 7.31 (m, 3H), 7.21 (ddd, J= 8.2, 7.3, 1.7 Hz, 1H), 7.00 (dd, J= 8.1, 1.4 Hz, 1H), 6.79 (ddd, J= 7.5, 1.4 Hz, 1H), 5.63 (s, 2H), 5.58 (s, 2H), 3.43 (t, J= 8.0 Hz, 2H), 0.81 (t, J= 8.2 Hz, 2H), -0.10 (s, 9H). 13C NMR
(101 MHz, Chloroform -d) δ 158.52, 156.09, 150.89, 150.25, 136.69, 136.61, 135.81, 131.43, 128.73, 128.51, 128.26, 121.32, 120.64, 118.48, 115.83, 108.65, 77.74, 68.69, 66.36, 17.70, -1.41. MS (ESI- QMS) m/z: [M + H]+ calcd for C25H30N3O3SSi, 480.2; found, 480.2.
General procedure for compounds L9 - L12 (S2)
The starting compound (L5 - L8) was dissolved in TFA (2 ml/ 100 mg of the material) and stirred at room temperature for 15 minutes. Trifluoroacetic acid was evaporated, coevaporated with water (2x), and a cold solution of ammonia in ethanol (2 ml/100 mg of the material) was added. The mixture was evaporated to dryness and the solid was dissolved in dimethylformamide (5 ml). The sample was subjected to purification by Cl 8 reverse phase flash chromatography (water to methanol) in a liquid injection mode. Product was slowly washed from the column within the second half of the gradient. Fractions were analyzed by TEC and fractions with the pure product were collected and evaporated to obtain the product as a white solid.
Example 9
7-((Phenyl)thio)-3 ,5 -dihydro-4H-pyrrolo [3 ,2-d]pyrimidin-4-one L9
Figure imgf000018_0001
General procedure S2 afforded 44 mg (56%) of the title compound. 1H NMR (401 MHz, DMSO-d6) δ 7.84 (s, 1H), 7.72 (s, 1H), 7.21 (dd, J= 7.7 Hz, 2H), 7.13 - 6.97 (m, 3H). 13C NMR (101 MHz, DMSO- d6) δ 153.73, 145.66, 142.84, 138.98, 133.69, 128.84, 125.60, 124.91, 119.21, 102.05. HRMS (ESI- FTMS) m/z: [M + H]+ calcd for C12H10ON3S, 244.0539; found, 244.0537.
Example 10
7-((Pyrimidin-2-yl)thio)-3 ,5 -dihydro-4H-pyrrolo [3 ,2-d]pyrimidin-4-one L10 General procedure S2 afforded 15 mg (44%) of the title compound. 1H NMR (401 MHz, DMSO-d6) δ 8.29 (d,J= 5.0 Hz, 2H), 7.74 (s, lH), 7.39 (s, 1H), 7.02 (t, J= 5.1 Hz, 1H). 13CNMR(101 MHz, DMSO- d6) δ 174.81, 167.00, 159.33, 151.75 (d, J = 4.2 Hz), 150.43, 143.61, 127.96, 118.94, 93.55. HRMS (ESI-FTMS) m/z: [M + Na]+ calcd for C10H7N5NaOS, 268.0264; found, 268.0263.
Example 11
7-(((2-Hydroxy methyl )phenyl)thio)-3.5-dihydro-4H-pyrrolo 13.2-d]pyrimidin-4-one Lll
Figure imgf000019_0001
General procedure S2 afforded 51 mg (72%) of the title compound. 1H NMR (401 MHz, DMSO-d6) δ 7.84 (s, 1H), 7.70 (s, 1H), 7.42 (dd, J= 7.5, 1.5 Hz, 1H), 7.10 (ddd, J= 7.4, 1.4 Hz, 1H), 7.04 (ddd, J =
7.6, 1.6 Hz, 1H), 6.76 (dd,J= 7.8, 1.3 Hz, 1H), 4.67 (s, 2H). 13C NMR (101 MHz, DMSO-d6) δ 153.83, 145.65, 142.85, 138.75, 136.11, 133.66, 127.08, 126.34, 126.05, 124.71, 119.30, 101.85, 60.52. HRMS (ESI-FTMS) m/z: [M + H]+ calcd for C13H12N302S, 274.0645; found, 274.0644.
Example 12 7-(((2-Hydroxy)phenyl)thio)-3.5-dihydro-4H-pyrrolo[3.2-d]|pyrimidin-4-one L12
Figure imgf000019_0002
General procedure S2 afforded 44 mg (62%) of the title compound. 1H NMR (401 MHz, DMSO-d6) δ 12.62 (s, 1H), 12.09 (s, 1H), 9.87 (s, 1H), 7.83 (d, J= 2.1 Hz, 1H), 7.65 (d, J= 2.7 Hz, 1H), 6.89 (ddd, J = 8.0, 7.2, 1.7 Hz, 1H), 6.77 (dd, J= 7.9, 1.3 Hz, 1H), 6.58 (ddd, J= 7.6, 1.3 Hz, 1H), 6.48 (dd, J = 7.8, 1.6 Hz, 1H). 13C NMR (101 MHz, DMSO-d6) δ 153.63, 153.04, 145.88, 142.62, 133.59, 126.15,
125.50, 125.29, 119.49, 119.16, 114.50, 101.79. HRMS (ESI-FTMS) m/z: [M - H]- calcd for C12H8N302S, 258.0343; found, 258.03401.
Example 13
Diethyl (E )-(2-iodostyryl)phosphonate L13 A flask was charged with 2-iodobenzaldehyde (2 g, 8.62 mmol, 1 eq.), dichloromethane (20 ml) and NaOH (20 ml of 5M aqueous solution). Tetraethyl methylenediphosphonate (2.57 ml, 10.3 mmol, 1.2 eq.) was added and the mixture was vigorously stirred at room temperature overnight. The reaction mixture was extracted with dichloromethane (3x), washed with brine (1x), dried with MgSO4, filtered and evaporated. The oil was adsorbed on silica gel in cyclohexane and purified by flash chromatography on silica gel (chloroform to 10% methanol), yielded 3.1 g (98%) of diethyl (E )-( 2- iodostyryl)phosphonate as a clear oil. 1H NMR (401 MHz, Chloroform-d) δ 7.87 (dd, J= 7.9, 1.2 Hz, 1H), 7.63 (dd, J= 22.3, 17.3 Hz, 1H), 7.52 (dd, J= 7.9, 1.6 Hz, 1H), 7.35 (dddd, J= 7.9, 7.3, 1.3, 0.6 Hz, 1H), 7.04 (ddd, J= 7.7, 1.7 Hz, 1H), 6.15 (dd, J= 18.1, 17.3 Hz, 1H), 4.24 - 4.09 (m, 4H), 1.37 (t, J= 7.1 Hz, 6H). 13C NMR (101 MHZ, Chloroform-d) δ 151.33 (d, J= 7.5 Hz), 140.01, 138.38 (d, J = 23.7 Hz), 131.28, 128.68, 127.30, 117.86 (d, J= 190.6 Hz), 100.68, 62.26 (d, J= 5.7 Hz), 16.56 (d, J = 6.4 Hz). 31P NMR (162 MHz, Chloroform-d) δ 20.30. MS (ESI-QMS) m/z: [M + H]+ calcd for C12H17I03P, 367.0; found, 367.0.
Example 14
Diisopropyl ((2-iodophenoxy)methyl)phosphonate L14
Figure imgf000020_0001
2-Iodophenol (2 g, 9.09 mmol, 1.0 eq.) was dissolved in dry tetrahydrofuran (20 ml) under an argon atmosphere. The solution was cooled to 0 °C, sodium hydride (382 mg of 60% oil dispersion, 9.54 mmol, 1.05 eq.) was added, and the mixture was stirred at 0 °C for 15 minutes. Then, TfOCH2P(O)(0/- Pr)2 (3.58 g, 10.9 mmol, 1.2 eq.) was added and the mixture was stirred at 0 °C for additional 15 minutes. The reaction was quenched with a half-saturated aqueous solution of NH4CI and extracted with ethyl acetate (3x). The organic phase was washed with brine (1x), dried with MgSO4, filtered and solvents were evaporated. The oil was adsorbed on silica gel in a mixture of cyclohexane/acetone and purified by flash chromatography on silica gel (chloroform to 10% methanol), yielded 2.97 g (100%) of diisopropyl ((2-iodophenoxy)methyl)phosphonate as a clear oil. 1H NMR (401 MHz, Chloroform -d) δ 7.76 (dd, J= 7.8, 1.6 Hz, 1H), 7.30 (ddd, J= 8.3, 7.4, 1.6 Hz, 1H), 6.89 (dd, J= 8.3, 1.3 Hz, 1H), 6.74 (ddd, J = 7.6, 1.3 Hz, 1H), 4.96 - 4.82 (m, 2H), 4.27 (d, J= 10.1 Hz, 2H), 1.42 - 1.35 (m, 12H). 13C NMR (101 MHz, Chloroform-d) δ 157.47 (d, J= 13.8 Hz), 139.84, 129.68, 123.69, 112.29, 86.11, 72.42 (d,J= 6.6 Hz), 63.58 (d, J= 170.5 Hz), 26.51 - 21.78 (m). 33P NMR (162 MHz, Chloroform-d) δ 18.31. MS (ESI-QMS) m/z: [M + H]+ calcd for C13H2 1I04P, 399.0; found, 399.0.
Example 15 (5 -Bromo-2-iodophenyl)methanol L 15
Figure imgf000021_0001
The compound was synthesized according to the published procedure. [Zhou, N.; Wang, L.; Thompson, D. W.; Zhao, Y. Org. Lett. 2008, 10 (14), 3001-3004]
Methyl 5-bromo-2-iodobenzoate (4 g, 11.7 mmol, 1 eq.) was dissolved in dry dichloromethane (24 ml) and the solution was cooled to 0 °C. Diisobutylaluminum hydride (23.4 ml of 1M toluene solution, 23.4 mmol, 2 eq.) was added and the mixture was stirred overnight at room temperature. The reaction was quenched with an aqueous solution of citric acid (15%), extracted with dichloromethane (3x), washed with brine (1x), dried with MgSO4 and filtered. Evaporation of solvents afforded 3.52 g (96%) of the title compound as ayellowish solid. 1H NMR (401 MHz, Chloroform-d) δ 7.64 (d, ./= 8.3 Hz, 1H), 7.62 (d, J = 2.4 Hz, 1H), 7.13 (dd, J = 8.4, 2.4 Hz, 1H), 4.62 (s, 2H), 2.06 (s, 1H). 13C NMR (101 MHz, Chloroform -d) δ 144.70, 140.43, 132.31, 131.27, 123.13, 94.76, 68.82. MS (ESI-QMS) m/z: [M + H - H20 + CH3CN]+ calcd for C9H8BrIN, 335.9; found, 335.9.
Example 16
5-Bromo-2-iodobenzaldehyde L16
Figure imgf000021_0002
The compound was synthesized according to the published procedure. [Zhou, N.; Wang, L.; Thompson, D. W.; Zhao, Y. Org. Lett. 2008, 10 (14), 3001-3004]
Compound L15 and pyridinium dichromate (7.2 g, 19.2 mmol, 2 eq.) were dissolved in dry dichloromethane (40 ml) and the mixture was stirred at room temperature for 4 hours. The mixture was filtered through celite, washed with diethyl ether and solvents were evaporated. The solid was adsorbed on silica gel in a mixture of cyclohexane/acetone and it was purified by flash column chromatography on silica gel (cyclohexane to 20% ethyl acetate modified with 10% methanol (v/vi)), yielded 2.3 g (77%) of the title compound as a white solid (85% NMR purity). 1H NMR (401 MHz, Chloroform-d) δ 9.99 (s, 1H), 7.99 (d, J= 2.5 Hz, 1H), 7.81 (d, J= 8.4 Hz, 1H), 7.41 (dd, J= 8.4, 2.5 Hz, 1H). 13C NMR (101 MHz, Chloroform-d) δ 194.45, 141.95, 138.37, 136.47, 133.24, 123.63, 98.39. MS (CI-QMS) m/z: [M + H]+ calcd for C7H5BrIO, 310.9; found, 310.9.
Example 17
Diethyl (E )-(5-bromo-2-iodostyryl)phosphonate L17 Tetraethyl methylenediphosphonate (1.52 ml, 7.72 mmol, 1.2 eq.) was dissolved in dry tetrahydrofuran (40 ml) under an argon atmosphere and potassium tert-butoxidc (867 mg, 7.72 mmol, 1.2 eq.) was added. The mixture was stirred at room temperature for 1 hour and a solution of compound L16 (2 g, 6.43 mmol, 1 eq.) in dry tetrahydrofuran (15 ml) was added. The mixture was stirred at room temperature for 1 hour and the reaction was quenched with HC1 ( 1M (aq.)), extracted with dichloromethane (3x), washed with brine (1x), dried with MgSO4 and filtered. Solvents were evaporated, solid was adsorbed on silica gel in a mixture of cyclohexane/acetone and it was purified by flash column chromatography on silica gel (chloroform to 5% methanol), yielded 1.82 g (63%) of the title compound as a clear oil. 1H NMR (401 MHz, Chloroform-d) δ 7.72 (dd, J= 8.5, 1.5 Hz, 1H), 7.63 (d,J= 2.0 Hz, 1H), 7.55 (ddd, J= 22.2, 17.2, 1.5 Hz, 1H), 7.17 (d, J= 8.4 Hz, 1H), 6.17 (dd, J= 17.3 Hz, 1H), 4.17 (p, J= 7.3 Hz, 2H), 1.38 (t, J= 7.2 Hz, 6H). 13C NMR (101 MHz, Chloroform-d) δ 149.76 (d, J= 8.0 Hz), 141.09, 140.26 (d, J = 24.1 Hz), 133.98, 130.11, 122.92, 119.44 (d, J= 190.0 Hz), 98.21, 62.23 (d, J= 5.4 Hz), 16.48 (d, J = 6.5 Hz). MS (ESI-QMS) m/z: [M + H]+ calcd for C12H16BrI03P, 449.9; found, 444.9.
Example 18
4-Bromo-l-(bromomethyl)-2-iodobenzene L18
Figure imgf000022_0001
The compound was synthesized according to the published procedure. [Caruso, A.; Tovar, J. D. J. Org. Chem 2011, 76, 2227-2239]
4-Bromo-2-iodo-l-methylbenzene (3.94 g, 13.3 mmol, 1 eq.) was dissolved in dry 1,2-dichloroethane (19 ml) under an argon atmosphere and N-bromosuccinimide(2.63 g, 14.6 mmol, 1.1 eq.) was added, followed by dibenzoyl peroxide (164 mg, 0.6630 mmol, 0.05 eq.). The mixture was refluxed for 4 hours and the reaction was quenched with a half-saturated aqueous solution of NH4CI and extracted with chloroform (3x). The organic phase was washed with brine, dried with MgSO4, filtered and solvents were evaporated. The solid was adsorbed on silica gel in a mixture of cyclohexane/acetone and purified by flash chromatography on silica gel (cyclohexane), yielded 4.35 g (89%) of the title compound as a white solid (85% UV/LC purity). 1H NMR (401 MHz, Chloroform-d) δ 8.00 (d. .J = 2.1 Hz, 1H), 7.46 (dd, J= 8.3, 1.7 Hz, 1H), 7.33 (d, J= 8.3 Hz, 1H), 4.54 (s, 2H). 13C NMR (101 MHz, Chloroform-d) δ 141.98, 139.32, 132.04, 131.28, 122.85, 100.38, 37.65. MS (ESI-QMS) m/z: [M + H - HBr + H2O + CH3CN]+ calcd for C9H10BrINO, 353.9; found, 353.9. Example 19
4-Bromo-2-iodobenzaldehyde L19
Figure imgf000023_0001
The compound was synthesized according to the published procedure. [Caruso, A.; Tovar, J. D. J. Org. Chem 2011, 76, 2227-2239] N- Methylmorpholinc-N-oxidc (3.74 g, 31.9 mmol, 3 eq.) was dissolved in dry acetonitrile (80 ml) under an argon atmosphere, molecular sieves (4A, 26 g) were added, and the suspension was cooled to 0 °C. Compound L18 (4 g, 10.6 mmol, 1 eq.) was added and the mixture was stirred at 0°C for 2 hours. The mixture was filtered through a short pad of silica gel and washed with cyclohexane, yielded 3.3 g ( 100%) of the title compound as a white solid (65% UV/LC purity). 1H NMR (401 MHz, Chloroform-d) δ 9.98 (d, J= 0.8 Hz, 1H), 8.12 (d, J= 1.8 Hz, 1H), 7.72 (d, J= 8.3 Hz, 1H), 7.61 (ddd, J= 8.3, 1.8, 0.8 Hz, 1H). 13C NMR (101 MHZ, Chloroform-d) δ 194.56, 142.67, 133.95, 132.17, 131.00, 130.04, 100.81. MS (CI-QMS) m/z: [M + H]+ calcd for C7H5BrIO, 310.9; found, 310.9.
Example 20
Diethyl (E )-(4-bromo-2-iodostyryl)phosphonate L20
Figure imgf000023_0002
Tetraethyl methylenediphosphonate (2.28 ml, 11.6 mmol, 1.2 eq.) was dissolved in dry tetrahydrofuran (60 ml) under an argon atmosphere and potassium tert-butoxide (1.3 g, 11.6 mmol, 1.2 eq.) was added. The mixture was stirred at room temperature for 1 hour and a solution of compound L19 (3 g, 9.65 mmol, 1 eq.) in dry tetrahydrofuran (20 ml) was added. The mixture was stirred at room temperature for 1 hour and the reaction was quenched with HC1 (1M aq.), extracted with dichloromethane, washed with brine, dried with MgSO4 and filtered. The solvents were evaporated, solid was adsorbed on silica gel in a mixture of cyclohexane/acetone and it was purified by flash column chromatography on silica gel (chloroform to 5% methanol), yielded 2.41 g (56%) of the title compound as a clear oil (60% UV/LC purity). 1H NMR (401 MHz, Chloroform-d) δ 8.04 (d, J= 1.9 Hz, 1H), 7.56 (dd, J= 22.3, 17.4 Hz, 1H), 7.51 - 7.47 (m, 1H), 7.38 (d, J= 8.3 Hz, 1H), 6.16 (t, J= 17.4 Hz, 1H), 4.21 - 4.11 (m, 4H), 1.38 (t, J = 7.1 Hz, 6H). 13C NMR (101 MHz, Chloroform-d) δ 150.08 (d, J= 7.8 Hz), 141.99 (d, J = 1.4 Hz), 131.93, 128.10 (d, J= 1.6 Hz), 124.43, 118.61 (d, J= 190.7 Hz), 100.96 (d, J= 15.9 Hz), 62.37 (d, J = 5.7 Hz), 16.92 - 15.90 (m). MS (ESI-QMS) m/z: [M + H]+ calcd for C12H16BrI03P, 444.9; found, 445.0.
Example 21 4-(l,3-Dimethylimidazolidin-2-yl)phenol L21
Figure imgf000024_0001
A flask was charged with toluene (600 ml) and 4-hydroxybenzaldehyde (36 g, 295 mmol, 1 eq.) was added, followed by N,N'-dimethylethylenediamine (38 ml, 354 mmol, 1.2 eq.). The mixture was stirred at reflux for 15 minutes, and then 2/3 of the volume was distilled-off The mixture was cooled in an ice bath and crystals were collected, washed with cold toluene and dried, yielded 48.6 g (86%) of the title compound as a brownish solid. 3HNMR (401 MHz, DMSO-d6) δ 9.36 (s, 1H), 7.16 (d, J= 8.5 Hz, 2H), 6.71 (d, J= 8.5 Hz, 2H), 3.24 - 3.15 (m, 2H), 3.09 (s, 1H), 2.47 - 2.38 (m, 2H), 2.01 (s, 6H). 13C NMR (101 MHz, DMSO-d6) δ 157.42, 129.89, 129.85, 114.72, 91.41, 52.57, 39.10. MS (CI-QMS) m/z: [M + H]+ calcd for Cl 1H17N20, 193.1; found, 193.1.
Example 22
4-Hydroxy-2-iodobenzaldehyde L22
Figure imgf000024_0002
Compound L21 (10 g, 52.0 mmol, 1 eq.) was dissolved in dry diethyl ether (250 ml) under an argon atmosphere, the solution was cooled to -78 °C and a solution of tert- butyl lithium (92 ml of 1.7M in pentane, 156 mmol, 3 eq.) was added keeping the temperature of the solution under -70 °C. The mixture was stirred at room temperature overnight and then it was cooled to -78 °C again. A solution of 1,2- diiodoethane (44 g, 156 mmol, 3 eq.) and dry diethyl ether (200 ml) was added keeping the temperature under -60 °C and the mixture was stirred at room temperature for 30 minutes. The reaction was quenched with 1M HC1 (aq.), extracted with diethyl ether (3x), washed with brine (1x), dried with MgSO4 and filtered. Solvents were evaporated, the solid was adsorbed on silica gel in a mixture of cyclohexane/acetone and it was purified by flash column chromatography on silica gel (cyclohexane to 20% ethyl acetate modified with 10% of methanol (v/vi)), yielded 9.4 g (73%) of the title compound as a white solid (81% UV/LC purity). 1H NMR (401 MHz, DMSO-d6) δ 10.86 (s, 1H), 9.74 (d, J= 0.8 Hz, 1H), 7.65 (d, J= 8.6 Hz, 1H), 7.37 (d, J= 2.3 Hz, 1H), 6.91 (dd, J= 8.6, 2.3 Hz, 1H). 13C NMR (101 MHz, DMSO-d6) δ 193.53, 163.52, 132.00, 126.82, 126.79, 116.18, 103.01. MS (ESI-QMS) m/z: [M + H]+ calcd for C7H6I02, 248.9; found, 249.0.
Example 23
Diethyl (E )-(4-hydroxy-2-iodostyryl)phosphonate L23 Compound L22 (4 g, 16.1 mmol, 1 eq.) was dissolved in dry toluene (80 ml) and diethylphosphonoacetic acid (3.8 ml, 19.4 mmol, 1.2 eq.), piperidine (524 μl, 5.13 mmol, 0.33 eq.) and acetic acid (221 μl, 3.86 mmol, 0.24 eq.) were added subsequently. The mixture was refluxed overnight in open flask and the reaction was quenched with a half-saturated aqueous solution of NaHCO3, extracted with chloroform (3x), washed with brine (1x), dried with MgSO4 and filtered. Solvents were evaporated and the mixture was purified by Cl 8 reverse phase flash column chromatography (cyclohexane to 20% ethyl acetate modified with 10% of methanol (v/vi)) in the liquid injection mode, yielded 5.3 g (87%) of title compound as a yellowish oil. 1H NMR (401 MHz, Chloroform-d) δ 9.48 (s, 1H), 7.62 (dd, J= 23.0, 17.3 Hz, 1H), 7.50 (d, J= 2.4 Hz, 1H), 7.40 (d, J= 8.7 Hz, 1H), 6.92 (ddd, J= 8.7, 2.5, 0.6 Hz, 1H), 5.96 (dd, J = 19.0, 17.3 Hz, 1H), 4.21 - 4.09 (m, 4H), 1.37 (t, J = 7.1 Hz, 6H). 13C NMR (101 MHz, Chloroform -d) δ 160.04, 152.40 (d, J= 8.3 Hz), 128.63 (d, J= 24.2 Hz), 127.86 (d, J= 1.3 Hz), 126.92 (d, J= 1.4 Hz), 116.68, 111.99 (d, J= 194.2 Hz), 102.44, 62.63 (d, J= 5.7 Hz), 16.54 (d, J= 6.4 Hz). 31P NMR (162 MHz, Chloroform-d) δ 20.82. MS (ESI-QMS) m/z: [M + H]+ calcd for C12H17I04P, 383.0; found, 383.1.
Example 24
2-Iodo-5-methoxybenzaldehyde L24
Figure imgf000025_0001
A flask was charged with 3-methoxybenzaldehyde (20.0 g, 147 mmol, 1 eq.) and methanol (600 ml) followed by AgNO3 (24.8 g, 147 mmol, 1 eq.) and iodine (42.0 g, 162 mmol, 1.1 eq.). The flask was wrapped in an aluminum foil and the mixture was stirred at room temperature for 1 hour. The mixture was filtered through celite, excess of iodine was titrated by a saturated aqueous solution of Na2S2O3 and solvents were evaporated to dryness. The solid was dissolved in a mixture of chloroform and water and it was extracted with chloroform (2x), washed with brine (1x), dried with MgSO4 and filtered. Solvents were evaporated and the solid was dissolved in hot ethyl acetate and the product was precipitated with cyclohexane. Crystals were collected, yielded 24 g (62%) of the title compound as a yellow solid. 1H NMR (401 MHz, DMSO-d6) δ 9.89 (s, 1H), 7.89 (d, J= 8.7 Hz, 1H), 7.30 (d, J= 3.2 Hz, 1H), 7.05 (dd, J= 8.6, 3.2 Hz, 1H), 3.80 (s, 3H). 13C NMR (101 MHz, DMSO-d6) δ 195.16, 159.73, 141.25, 135.54, 122.87, 114.36, 89.57, 55.58. MS (ESI-QMS) m/z: [M + H]+ calcd for C8H8I02, 263.0; found, 263.0. Example 25
5 -Hydroxy-2 -iodobenzaldehyde L25 Compound L24 (5 g, 19.1 mmol, 1 eq.) was dissolved in dry dichloromethane (200 ml) under an argon atmosphere. The solution was cooled to -78 °C, a solution of boron tribromide (95 ml of 1M in dichloromethane, 95.5 mmol, 5 eq.) was added and the mixture was stirred at room temperature until full conversion was achieved (ca. 3 hours). The mixture was cooled to -15 °C and the reaction was slowly quenched by a saturated aqueous solution of NaHCO3. The mixture was extracted with dichloromethane (3x), washed with brine (1x), dried with MgSO4, filtered and evaporated. The solid was adsorbed on silica gel in a mixture of cyclohexane/acetone and it was purified by flash column chromatography on silica gel (cyclohexane to 10% ethyl acetate modified with 10% of methanol (v/vi)), yielded 4.1 g (87%) of the title compound as yellow needles. 1H NMR (401 MHz, DMSO-d6) δ 10.18 (s, 1H), 9.86 (s, 1H), 7.80 (d, J= 8.5 Hz, 1H), 7.21 (d, J= 3.1 Hz, 1H), 6.89 (dd, J= 8.5, 3.1 Hz, 1H). 13C NMR (101 MHZ, DMSO-d6) δ 195.45, 158.09, 141.27, 135.54, 123.90, 116.41, 87.33. MS (ESI- QMS) m/z: [M + H]+ calcd for C7H6I02, 248.9; found, 249.0.
Example 26
Diethyl (E )-(5-hydroxy-2-iodostyryl)phosphonate L26
Figure imgf000026_0001
A flask was charged with compound L25 (4.00 g, 16.1 mmol, 1 eq.) and absolute ethanol (80 ml). The mixture was stirred at room temperature and potassium carbonate (11.2 g, 80.6 mmol, 5 eq.) with tetraethyl methylenediphosphonate (12.0 mL, 48.4 mmol, 3 eq.) were added subsequently. The mixture was heated at reflux until full conversion was achieved (1 hour). The mixture was cooled to 0 °C and it was acidified with 1M hydrochloric acid, extracted by chloroform (3x), washed with brine (1x), dried with MgSO4 and filtered. Solvents were evaporated and the solid was adsorbed on silica gel in a mixture of acetone/cyclohexane and it was purified by flash column chromatography on silica gel (cyclohexane to 1% of ethyl acetate modified with 10% of methanol (v/vi)). This separation provided impure product containing tetraethyl methylenediphosphonate. This mixture was purified by flash chromatography on C18 silica gel (liquid injection in dimethylformamide, eluent water to methanol), yielded 5.39 g (88%) of the title product as a yellow solid. 1H NMR (401 MHz, Chloroform-d) δ 9.23 (s, 1H), 7.64 (d, J= 8.6 Hz, 1H), 7.54 (dd, J= 22.6, 17.3 Hz, 1H), 7.21 (d, J= 2.9 Hz, 1H), 6.68 (dd, J= 8.6, 2.9 Hz, 1H), 6.12 (dd, J= 19.6, 17.3 Hz, 1H), 4.29 - 4.06 (m, 4H), 1.37 (td, J= 7.1, 0.6 Hz, 6H). 13C NMR (101 MHz, Chloroform -d) δ 158.31, 152.31 (d, J= 6.9 Hz), 140.52, 138.11 (d, J= 23.5 Hz), 120.48, 115.74 (d, J= 192.6 Hz), 114.43, 87.79, 62.95 (d, J= 5.9 Hz), 16.49 (d, J= 6.5 Hz).31P NMR (162 MHz, Chloroform- d) δ 20.49. MS (ESI-QMS) m/z: [M + H]+ calcd for C12H17I04P, 383.0; found, 383.0. Example 27
2-(2-Methoxyphenyl)-l,3-dimethylimidazolidine L27
Figure imgf000027_0001
A flask was charged with toluene (600 ml) and 2-methoxybenzaldehyde (36.0 g, 295 mmol, 1 eq.) was added, followed by N,N'-dimethylethylenediamine (31.3 ml, 325 mmol, 1.1 eq.). The mixture was stirred at reflux for 15 minutes, and then toluene was evaporated. The solid was recrystallized from hexane yielded 36 g (67%) of the title compound as clear large crystals. 1H NMR (401 MHz, Chloroform-d) δ 7.67 (dd, J= 7.6, 1.9 Hz, 1H), 7.26 (td, J= 6.6, 1.9 Hz, 1H), 7.00 (td, J = 7.4, 1.3 Hz, 1H), 6.88 (dd, J = 8.2, 1.1 Hz, 1H), 4.06 (s, 1H), 3.82 (s, 3H), 3.37 - 3.31 (m, 2H), 2.65 - 2.57 (m, 2H), 2.19 (s, 6H). 13C NMR (101 MHz, Chloroform-d) δ 159.04, 129.40, 129.00, 127.75, 121.26, 110.52, 82.90, 55.63, 53.67, 39.83. MS (CI-QMS) m/z: [M + H]+ calcd for C12H19N20, 207.1; found, 207.1.
Example 28
2-Iodo-6-methoxybenzaldehyde L28
Figure imgf000027_0002
Compound L27 (10 g, 52.0 mmol, 1 eq.) was dissolved in dry diethyl ether (250 ml) under an argon atmosphere, the solution was cooled to -78 °C and a solution of tert- butyl lithium (61 ml of 1.7M in pentane, 104 mmol, 2 eq.) was added keeping the temperature of the solution under -70 °C. The mixture was stirred at room temperature overnight and then it was cooled to -78 °C again. A solution of 1,2- diiodoethane (29.3 g, 104 mmol, 2 eq.) in dry diethyl ether (200 ml) was added keeping the temperature under -60 °C. The mixture was stirred at room temperature for 30 minutes. The reaction was quenched with 1M HC1 (aq.), extracted with diethyl ether (3x), washed with brine (1x), dried with MgSO4 and filtered. Solvents were evaporated, the solid was adsorbed on silica gel in a mixture of cyclohexane/acetone and it was purified by flash column chromatography on silica gel (cyclohexane to 20% ethyl acetate modified with 10% methanol (v/vi)), yielded 5.8 g (46%) of the title compound as a white solid. 1H NMR (401 MHz, Chloroform-d) δ 10.24 (s, 1H), 7.59 (d, J= 7.8 Hz, OH), 7.13 (dd, J = 8.4, 7.8 Hz, 1H), 6.99 (dd, J= 8.3, 0.9 Hz, 1H), 3.91 (s, 3H). 13C NMR (101 MHz, Chloroform-d) δ 191.90, 161.98, 135.25, 133.92, 125.04, 112.08, 96.55, 56.16. MS (ESI-QMS) m/z: [M + H]+ calcd for C8H8I02, 263.0; found, 263.0.
Example 29
2-Hydroxy-6-iodobenzaldehyde L29 Compound L28 (5 g, 19.1 mmol, 1 eq.) was dissolved in dry dichloromethane (200 ml) under an argon atmosphere. The solution was cooled to -78 °C and a solution of boron tribromide (95 ml of 1M in dichloromethane, 95.5 mmol, 5 eq.) was added and the mixture was stirred at room temperature until full conversion was achieved (ca. 3 hours). The mixture was cooled to -15 °C and the reaction was slowly quenched by a saturated aqueous solution of NaHCO3. The mixture was extracted with chloroform (3x), washed with brine (1x), dried with MgSO4, filtered and evaporated. The solid was adsorbed on silica gel in a mixture of cyclohexane/acetone and it was purified by flash column chromatography on silica gel (cyclohexane to 10% ethyl acetate modified with 10% methanol (v/vi)), yielded 4.4 g (93%) of the title compound as a white-off solid. 1H NMR (401 MHz, DMSO-d6) δ 11.58 (s, 1H), 10.02 (s, 1H), 7.52 (dd, J= 7.7, 1.0 Hz, 1H), 7.22 (dd, J= 8.4, 7.7 Hz, 1H), 7.00 (dd, J= 8.4, 1.1 Hz, 1H). 13CNMR(101 MHZ, DMSO-d6) δ 198.92, 162.10, 137.58, 131.87, 120.06, 118.14, 101.00. MS (ESI-QMS) m/z: [M + H]+ calcd for C7H6I02, 248.9; found, 248.9.
Example 30
Diethyl (E )-(2-hydroxy-6-iodostyryl)phosphonate L30
Figure imgf000028_0001
Compound L29 (1.5 g, 6.05 mmol, 1 eq.) and tetraethyl methylenediphosphonate (3 ml, 12.1 mmol, 2 eq.) were dissolved in dry dimethylformamide (30 ml) under an argon atmosphere and the solution was cooled to 0 °C. Sodium hydride (484 mg of 60% oil dispersion, 12.1 mmol, 2 eq.) was added at once and the mixture was stirred at 40 °C overnight. The reaction was quenched with a half-saturated aqueous solution ofNH4Cl, extracted with ethyl acetate (3x), washed with brine (1x) and dried with MgSO4. The mixture was evaporated to dryness and it was adsorbed on silica gel in a mixture of cyclohexane/acetone. Purification by flash chromatography on silica gel (cyclohexane to 40% ethyl acetate modified with 10% methanol (v/vi)) afforded 1.2 g (52%) of the title compound as a clear oil. 1H NMR (401 MHz, DMSO-d6) δ 10.70 (s, 1H), 7.43 (dd, J= 6.9, 1.9 Hz, 1H), 7.42 (dd, J= 24.2, 16.7 Hz, 1H), 6.96 (dd, J = 8.3, 1.5 Hz, 1H), 6.91 (dd, J= 8.2, 7.5 Hz, 1H), 6.69 (dd, J= 21.7, 17.5 Hz, 1H), 4.02 (dq, J= 8.3, 7.1 Hz, 4H), 1.27 (t, J= 7.1 Hz, 6H). 13C NMR (101 MHz, DMSO-d6) δ 158.54 - 156.63 (m), 148.12
(d, J= 8.0 Hz), 131.89, 130.66, 123.24 (d, J= 22.1 Hz), 120.16 (d, J= 182.8 Hz), 116.74, 103.66, 61.31
(d, J= 5.6 Hz), 16.30 (d, J= 5.9 Hz).31P NMR (162 MHz, DMSO-d6) δ 21.53. MS (ESI-QMS) m/z: [M + H]+ calcd for C12H17I04P, 383.0; found, 383.1.
General procedure for compounds L31 - L42 (S3) The corresponding phenol (L23, L26, L30) (500 mg, 1.31 mmol, 1 eq.) was dissolved in dry dimethylformamide (5 ml) under an argon atmosphere and the mixture was cooled to 0 °C. Sodium hydride (79 mg of 60% oil dispersion, 1.96 mmol, 1.5 eq.) was added and the mixture was stirred at room temperature for 30 minutes. An alkylating agent was added (2.62 mmol, 2 eq.) and the mixture was stirred at room temperature for 1 hour. The reaction was quenched with a half-saturated aqueous solution of NH4Cl, extracted with ethyl acetate (3x), washed with brine (1x) and dried with MgSO4. The mixture was evaporated to dryness and it was adsorbed on silica gel in a mixture of cyclohexane/acetone . Purification by flash chromatography on silica gel (cyclohexane to 40% ethyl acetate modified with 10% methanol (v/vi)) afforded a pure product.
Example 31
Diethyl (E )-(2-iodo-4-methoxystyryl)phosphonate L31
Figure imgf000029_0001
General procedure S3 with methyl iodide (163 μl) afforded 462 mg (89%) of the title compound as a clear oil. 1H NMR (401 MHz, Chloroform-d) δ 7.53 (dd, J = 22.4, 17.3 Hz, 1H), 7.41 (d, J = 8.7 Hz, 1H), 7.33 (d, J = 2.6 Hz, 1H), 6.85 (ddd, J = 8.7, 2.6, 0.7 Hz, 1H), 5.98 (dd, J = 18.0, 17.3 Hz, 1H), 4.09 (dq, J = 7.9, 7.1 Hz, 4H), 3.74 (s, 3H), 1.31 (td, J = 7.0, 0.5 Hz, 6H). 13C NMR (101 MHz, Chloroform- d) δ 160.86, 150.62 (d,J= 8.1 Hz), 130.43 (d, J= 23.7 Hz), 127.54, 124.58, 114.98, 114.78 (d, J= 192.0 Hz), 101.57, 61.95 (d, J= 5.6 Hz), 55.60, 16.43 (d, J= 6.5 Hz). 31P NMR (162 MHz, Chloroform-d) δ 21.03. MS (ESI-QMS) m/z: [M + H]+ calcd for C13H19I04P, 397.0; found, 397.0.
Example 32
Diethyl (E )-(2-iodo-4-isopropoxystyryl)phosphonate L32
Figure imgf000029_0002
General procedure S3 with isopropyl iodide (261 μl) afforded 465 mg (84%) of the title compound as a yellowish oil. 1H NMR (401 MHz, Chloroform-d) δ 7.52 (dd, J= 22.4, 17.2 Hz, 1H), 7.39 (d, J= 8.8 Hz, 1H), 7.31 (d, J= 2.6 Hz, 1H), 6.80 (ddd, J= 8.8, 2.7, 0.7 Hz, 1H), 5.96 (dd, J= 18.0, 17.3 Hz, 1H), 4.47 (hept, J= 6.0 Hz, 1H), 4.08 (dq, J= 7.9, 7.1 Hz, 4H), 1.29 (td, J= 7.1, 0.6 Hz, 6H), 1.25 (d, J = 6.1 Hz, 6H). 13C NMR (101 MHZ, Chloroform-d) δ 159.35, 150.71 (d, J= 8.0 Hz), 130.04 (d, J= 24.2 Hz), 127.56, 126.36, 116.15, 114.49 (d,J= 192.2 Hz), 101.67, 70.43, 61.93 (d, J= 5.8 Hz), 21.83, 16.43 (d, J= 6.5 Hz). 31P NMR (162 MHz, Chloroform-d) δ 21.06. MS (ESI-QMS) m/z: [M + H]+ calcd for C15H23I04P, 425.0; found, 425.3.
Example 33
Diethyl (E )-(2-iodo-4-((2,3,4,5,6-pentafluorophenyl)methoxy)styryl)phosphonate L33 General procedure S3 with 2,3,4,5,6-pentafluorobenzyl bromide (395 μl) afforded 700 mg (95%) of the title compound as a clear oil. 1H NMR (401 MHz, Chloroform-d) δ 7.59 (dd, J = 22.4, 17.3 Hz, 1H),
7.50 (d, J= 5.1 Hz, 1H), 7.49 (d, J= 1.0 Hz, 1H), 6.97 (ddd, J= 8.8, 2.6, 0.7 Hz, 1H), 6.07 (dd, J= 17.5
Hz, 1H), 5.12 (s, 2H), 4.16 (dq, J= 7.9, 7.1 Hz, 4H), 1.37 (td, J= 7.1, 0.5 Hz, 6H). 13C NMR (101 MHz, Chloroform -d) δ 159.15, 150.44 (d, J= 8.0 Hz), 147.58 - 146.63 (m), 144.97 - 144.02 (m), 139.40 - 138.40 (m), 131.99 (d, J= 24.1 Hz), 127.91, 125.69, 116.01 (d, J= 191.7 Hz), 115.61, 109.86 - 108.61 (m), 101.49, 62.19 (d, J= 5.4 Hz), 57.77, 16.59 (d, J= 6.5 Hz). 19F NMR (377 MHz, Chloroform-d) δ - 138.39 - -143.87 (m), -147.08 - -153.58 (m), -158.12 - -164.61 (m). 31P NMR (162 MHz, Chloroform- d) δ 20.63. MS (ESI-QMS) m/z: [M + H]+ calcd for C19H18F5I04P, 563.0; found, 563.2.
Example 34
Diethyl (E )-(2-iodo-4-((2,3,4,5,6-pentafluorophenyl)oxy)styryl)phosphonate L34
Figure imgf000030_0001
General procedure S3 with hexafluorobenzene (151 μl), heating at 70 °C afforded 530 mg (74%) of the title compound as a clear oil. 1H NMR (401 MHz, Chloroform-d) δ 7.59 (dd, J = 22.3, 17.3 Hz, 1H), 7.50 (d, J= 8.7 Hz, 1H), 7.44 (d, J= 2.6 Hz, 1H), 6.97 (dd, J= 8.7, 2.6 Hz, 1H), 6.10 (dd, J= 17.4 Hz, 1H), 4.16 (dq, J = 7.9, 7.1 Hz, 4H), 1.37 (t, J = 7.1 Hz, 6H). 13C NMR (101 MHz, Chloroform-d) δ 157.73, 150.02 (d, J= 7.9 Hz), 134.36 (d,J= 23.7 Hz), 128.18, 126.43, 117.52 (d,J= 191.5 Hz), 115.90, 100.99, 62.34 (d, J= 5.7 Hz), 16.59 (d, J= 6.6 Hz). 19F NMR (377 MHz, Chloroform-d) δ -153.37 - - 153.56 (m), -157.93 - -158.31 (m), -160.86 - -161.12 (m).31P NMR (162 MHz, Chloroform-d) δ 20.20. MS (ESI-QMS) m/z: [M + H]+ calcd for C18H16F5I04P, 549.0; found, 549.2.
Example 35
Diethyl (E )-(2-iodo-5-methoxystyryl)phosphonate L35
Figure imgf000030_0002
General procedure S3 with methyl iodide (163 μl) afforded 326 mg (63%) of the title compound as a yellowish oil. 1H NMR (401 MHz, Chloroform-d) δ 7.63 (d, J= 8.7 Hz, 1H), 7.50 (dd, J= 22.2, 17.3 Hz, 1H), 6.98 (d, J= 3.0 Hz, 1H), 6.58 (dd, J= 8.7, 3.0 Hz, 1H), 6.08 (dd, J= 18.0, 17.3 Hz, 1H), 4.09 (dq, J= 8.0, 7.1 Hz, 4H), 3.72 (s, 3H), 1.30 (t, J= 7.0 Hz, 6H). 13C NMR (101 MHz, Chloroform-d) δ 159.97, 150.94 (d, J= 7.4 Hz), 140.22, 138.82 (d, J= 23.5 Hz), 117.75 (d,J= 190.1 Hz), 117.69, 112.53 (d, J = 1.6 Hz), 89.12, 62.04 (d, J = 5.7 Hz), 55.41, 16.39 (d, J = 6.6 Hz). 31P NMR (162 MHz, Chloroform -d) δ 20.12. MS (ESI-QMS) m/z: [M + H]+ calcd for C13H19I04P, 397.0; found, 397.1.
Example 36 Diethyl (E )-(2-iodo-5-isopropoxystyryl)phosphonate L36
Figure imgf000031_0001
General procedure S3 with isopropyl iodide (261 μl) afforded 402 mg (72%) of the title compound as a yellowish oil. 1H NMR (401 MHz, Chloroform-d) δ 7.56 (d, J= 8.7 Hz, 1H), 7.45 (dd, J= 22.2, 17.3 Hz, 1H), 6.93 (d, J= 3.0 Hz, 1H), 6.51 (dd, J= 8.7, 3.0 Hz, 1H), 6.01 (dd, J= 18.0, 17.3 Hz, 1H), 4.40 (hept, J= 6.0 Hz, 1H), 4.03 (dq, J= 8.0, 7.1 Hz, 4H), 1.24 (td, J= 7.1, 0.6 Hz, 6H), 1.18 (d, J= 6.0 Hz, 6H). 13C NMR (101 MHz, Chloroform-d) δ 158.12, 150.87 (d, J= 7.8 Hz), 140.09, 138.71 (d, J= 23.5 Hz), 119.05, 117.45 (d, J= 190.1 Hz), 114.57 (d, J= 1.4 Hz), 88.62, 70.02, 61.87 (d, J= 5.8 Hz), 21.66, 16.26 (d, J= 6.5 Hz).31P NMR (162 MHz, Chloroform-d) δ 20.13. MS (ESI-QMS) m/z: [M + H]+ calcd for C15H23I04P, 425.0; found, 425.2.
Example 37
Diethyl (E )-(2-iodo-5-((2,3,4,5,6-pentafluorophenyl)methoxy)styryl)phosphonate L37
Figure imgf000031_0002
General procedure S3 with 2,3,4,5,6-pentafluorobenzyl bromide (395 μl) afforded 675 mg (92%) of the title compound as a clear oil. 1H NMR (401 MHz, Chloroform-d) δ 7.63 (d, ./= 8.7 Hz, 1H), 7.46 (dd, J = 22.2, 17.3 Hz, 1H), 7.06 (d, J = 3.0 Hz, 1H), 6.64 (dd, J= 8.7, 3.0 Hz, 1H), 6.09 (dd, J = 17.4 Hz, 1H), 5.04 (s, 2H), 4.07 (dq, J = 8.1, 7.1 Hz, 4H), 1.27 (t, J = 7.1 Hz, 6H). 13C NMR (101 MHz, Chloroform -d) δ 158.27, 150.45 (d, J= 7.5 Hz), 147.19 - 146.31 (m), 144.87 - 143.92 (m), 143.47 - 142.27 (m), 140.42, 139.10 (d, J= 23.5 Hz), 118.25 (d, J= 189.9 Hz), 118.13, 113.35, 110.60 - 108.34 (m), 90.43, 62.02 (d, J= 5.5 Hz), 57.50, 16.23 (d, J= 6.5 Hz). 19F NMR (377 MHz, Chloroform-d) δ - 138.86 - -143.87 (m), -149.90 - -153.34 (m), -159.37 - -163.83 (m). 31P NMR (162 MHz, Chloroform- d) δ 19.75. MS (ESI-QMS) m/z: [M + H]+ calcd for C19H18F5I04P, 563.0; found, 563.0.
Example 38
Diethyl (E )-(2-iodo-5-((2,3,4,5,6-pentafluorophenyl)oxy)styryl)phosphonate L38
Figure imgf000031_0003
The general procedure with hexafluorobenzene (151 μl), heating at 70 °C afforded 300 mg (42%) of the title compound as a clear oil. 1H NMR (401 MHz, Chloroform-d) δ 7.75 (d, ./= 8.7 Hz, 1H), 7.50 (dd, J = 22.1, 17.3 Hz, 1H), 7.07 (d, J= 3.0 Hz, 1H), 6.65 (dd, J= 8.7, 3.0 Hz, 1H), 6.07 (dd, J= 34.8, 17.4 Hz, 1H), 4.12 (dq, J= 8.0, 7.1 Hz, 4H), 1.32 (td, J= 7.0, 0.6 Hz, 6H). 13C NMR (101 MHz, Chloroform- d) d 157.56, 149.94 (d, J = 7.7 Hz), 143.53 - 142.68 (m), 140.94, 139.94 (d, J = 23.7 Hz), 138.30 - 137.58 (m), 137.44 - 136.53 (m), 129.22 - 128.40 (m), 119.29 (d, J= 190.2 Hz), 118.29, 114.17 (d, J = 1.9 Hz), 93.06, 62.27 (d, J= 5.6 Hz), 16.41 (d, J= 6.4 Hz). 19F NMR (377 MHz, Chloroform-d) δ - 151.86 - -154.36 (m), -156.40 - -159.84 (m), -159.84 - -162.90 (m). 31P NMR (162 MHz, Chloroform- d) δ 19.40. MS (ESI-QMS) m/z: [M + H]+ calcd for C18H16F5I04P, 549.0; found, 549.1.
Example 39
Diethyl (E )-(2-iodo-6-methoxystyryl)phosphonate L39
Figure imgf000032_0001
General procedure S3 with methyl iodide (163 μl) afforded 483 mg (93%) of the title compound as a yellowish oil. 1H NMR (401 MHz, Chloroform-d) δ 7.49 (dd, J= 7.6, 1.3 Hz, 1H), 7.47 (dd, J= 25.0, 17.7 Hz, 1H), 6.92 (dd, J= 8.2, 7.8 Hz, 1H), 6.86 (dd, J= 8.4, 1.3 Hz, 1H), 6.54 (dd, J= 21.3, 17.6 Hz, 1H), 4.13 (dq, J = 7.9, 7.1 Hz, 4H), 3.80 (s, 3H), 1.34 (t, J = 7.1 Hz, 6H). 13C NMR (101 MHz, Chloroform -d) δ 158.54 (d,J= 2.1 Hz), 148.06 (d, J= 7.5 Hz), 132.32, 131.19, 126.54 (d, J= 23.0 Hz), 121.16 (d, J= 184.3 Hz), 111.36, 102.64, 61.96 (d, J= 5.3 Hz), 55.74, 16.50 (d, J= 6.5 Hz). 31P NMR (162 MHz, Chloroform-d) δ 20.51. MS (ESI-QMS) m/z: [M + H]+ calcd for C13H19I04P, 397.0; found, 369.9.
Example 40
Diethyl (E )-(2-iodo-6-isopropoxystyryl)phosphonate L40
Figure imgf000032_0002
The general procedure with isopropyl iodide (261 μl) afforded 488 mg (88%) of the title compound as a yellowish oil. 1H NMR (401 MHz, Chloroform-d) δ 7.46 (dd, J= 6.3, 2.6 Hz, 1H), 7.45 (dd, J= 24.9, 17.6 Hz, 1H), 6.92 - 6.84 (m, 2H), 6.53 (dd, J= 21.6, 17.6 Hz, 1H), 4.56 (hept, J= 6.1 Hz, 1H), 4.21 - 4.06 (m, 4H), 1.39 - 1.29 (m, 12H). 13CNMR(101 MHz, Chloroform-d) δ 156.79 (d,J= 2.1 Hz), 148.46
(d, J= 8.0 Hz), 132.03, 131.01, 127.50 (d, J= 22.6 Hz), 121.08 (d,J= 183.5 Hz), 113.75, 102.63, 71.33,
61.94 (d, J= 5.2 Hz), 22.05, 16.53 (d, J= 6.6 Hz). 31P NMR (162 MHz, Chloroform-d) δ 20.55. MS (ESI-QMS) m/z: [M + H]+ calcd for C15H23I04P, 425.0; found, 425.0.
Example 41
Diethyl (E )-(2-iodo-6-((2,3,4,5,6-pentafluorophenyl)methoxy)styryl)phosphonate L41
Figure imgf000032_0003
General procedure S3 with 2,3,4,5,6-pentafluorobenzyl bromide (395 μl) afforded 668 mg (91%) of the title compound as a clear oil. 1H NMR (401 MHz, Chloroform -d) δ 7.65 - 7.53 (m, 1H), 7.37 (ddd. ./ =
24.5, 17.6, 2.0 Hz, 1H), 7.03 (d, J= 8.4 Hz, 1H), 6.97 (dd, J= 8.0, 1.5 Hz, 1H), 6.29 (ddd, J= 20.4,
17.6, 1.7 Hz, 1H), 5.14 (d, J= 1.6 Hz, 2H), 4.21 - 3.95 (m, 4H), 1.29 (td,J= 7.1, 1.6 Hz, 6H).13C NMR (101 MHz, Chloroform-d) δ 156.23, 147.38 (d, J = 7.7 Hz), 147.25 - 146.51 (m), 145.30 - 144.00 (m), 136.77 - 135.83 (m), 133.59, 131.12, 128.10 (d, J= 24.2 Hz), 122.08 (d, J= 183.1 Hz), 112.82, 110.47 - 108.60 (m), 102.16, 61.95 (d, J = 5.2 Hz), 58.18, 16.38 (d, J = 6.5 Hz). 19F NMR (377 MHz, Chloroform -d) δ -138.82 - -144.30 (m), -147.51 - -154.01 (m), -158.55 - -165.05 (m).31P NMR (162 MHz, Chloroform-d) δ 20.15. MS (ESI-QMS) m/z: [M + H]+ calcd for C19H18F5I04P, 563.0; found, 563.1.
Example 42
Diethyl (E )-(2-iodo-6-((2,3,4,5,6-pentafluorophenyl)oxy)styryl)phosphonate L42
Figure imgf000033_0001
General procedure S3 with hexafluorobenzene (151 μl), heating overnight at 70 °C afforded 356 mg (50%) of the title compound as a clear oil. 1H NMR (401 MHz, Chloroform-d) δ 7.62 (dd, J = 7.9, 1.0 Hz, 1H), 7.44 (dd, J= 24.1, 17.7 Hz, 1H), 6.87 (dd, ./= 8.1 Hz, 1H), 6.61 (d, J= 7.9 Hz, 1H), 6.40 (dd, J = 19.6, 17.7 Hz, 1H), 4.11 (dq, J = 8.1, 7.1 Hz, 4H), 1.30 (td, J = 7.0, 0.6 Hz, 6H). 13C NMR (101 MHz, Chloroform -d) δ 154.61, 146.20 (d, J= 7.4 Hz), 135.43, 130.98, 128.21 (d, J= 23.5 Hz), 123.28 (d, J= 184.7 Hz), 113.99, 101.47 (d, J = 1.9 Hz), 62.15 (d, J= 5.3 Hz), 16.33 (d, J= 6.3 Hz). 19F NMR (377 MHz, Chloroform-d) δ -152.64 - -156.08 (m), -157.10 - -159.37 (m), -159.37 - -166.65 (m).31P NMR (162 MHz, Chloroform-d) δ 19.80.MS (ESI-QMS) m/z: [M + H]+ calcd for C18H16F5I04P, 549.0; found, 549.2.
Example 43
Diisopropyl ((4-fluoro-2-iodophenoxy)methyl)phosphonate L43
Figure imgf000033_0002
4-Fluoro-2-iodophenol (1 g, 4.20 mmol, 1 eq.) was dissolved in dry tetrahydrofuran (20 ml) under an argon atmosphere. The solution was cooled to 0 °C, sodium hydride (210 mg of 60% oil dispersion, 5.25 mmol, 1.25 eq.) was added, and the mixture was stirred at 0 °C for 15 minutes. Then, TfOCH2P(O)(0/-Pr)2 (1.72 g, 5.25 mmol, 1.25 eq.) was added and the mixture was stirred at 0 °C for additional 15 minutes. The reaction was quenched with a half-saturated aqueous solution of NH4CI and extracted with ethyl acetate (3x). The organic phase was washed with brine (1x), dried with MgSO4, filtered and solvents were evaporated. The solid was adsorbed on silica gel in a mixture of cyclohexane/acetone and purified by flash chromatography on silica gel (cyclohexane to 20% ethyl acetate modified with 10% methanol (v/vi)), yielded 1.39 g (79%) of the title compound as a clear oil. 1H NMR (401 MHz, Chloroform-d) δ 7.49 (dd, J = 7.5, 3.0 Hz, 1H), 7.02 (ddd, J= 9.0, 7.7, 3.0 Hz, 1H), 6.86 (dd, J= 9.1, 4.5 Hz, 1H), 4.87 (dhept, J= 7.5, 6.2 Hz, 2H), 4.24 (d, J = 9.9 Hz, 2H), 1.42 - 1.31 (m, 12H). 13C NMR (101 MHz, Chloroform-d) δ 157.48 (d, J= 245.0 Hz), 154.31 (dd, J= 13.2, 2.3 Hz), 126.47 (d, J= 25.0 Hz), 115.87 (d, J= 22.9 Hz), 112.95 (d, J= 8.2 Hz), 85.73 (d, J= 8.6 Hz), 72.26 (d, J= 6.6 Hz), 64.58 (d, J= 170.1 Hz), 26.53 - 22.24 (m). 19F NMR (377 MHz, Chloroform-d) d -116.31 - -124.37 (m). 31P NMR (162 MHz, Chloroform-d) δ 18.12. MS (ESI-QMS) m/z: [M + H]+ calcd for C13H20FIO4P, 417.0; found, 417.0.
Example 44
Diisopropyl ((2-iodo-6-methoxyphenoxy)methyl)phosphonate L44
Figure imgf000034_0001
4-Fluoro-2-iodophenol (1 g, 4.0 mmol, 1 eq.) was dissolved in dry tetrahydrofuran (20 ml) under an argon atmosphere. The solution was cooled to 0 °C, sodium hydride (200 mg of 60% oil dispersion, 5.0 mmol, 1.25 eq.) was added, and the mixture was stirred at 0 °C for 15 minutes. Then, TfOCH2P(O)(0/- Pr)2 (1.64 g, 5.0 mmol, 1.25 eq.) was added and the mixture was stirred at 0 °C for additional 15 minutes. The reaction was quenched with a half-saturated aqueous solution of NH4CI and extracted with ethyl acetate (3x). The organic phase was washed with brine (1x), dried with MgSO4, filtered and solvents were evaporated. The solid was adsorbed on silica gel in a mixture of cyclohexane/acetone and purified by flash chromatography on silica gel (cyclohexane to 20% ethyl acetate modified with 10% methanol (v/vi)), yielded 1.70 g (99%) of the title compound as a clear oil. 1H NMR (401 MHz, Chloroform -d) δ 7.33 (dd, J= 7.9, 1.5 Hz, 1H), 6.87 (dd, J= 8.2, 1.6 Hz, 1H), 6.80 (dd, ./= 8.0 Hz, 1H), 4.88 (dhept, J= 7.3, 6.2 Hz, 2H), 4.31 (d, J= 9.7 Hz, 2H), 3.83 (s, 3H), 1.41 (d, J= 6.2 Hz, 12H). 13C NMR (101 MHz, Chloroform -d) δ 152.56, 147.76 (d, J= 14.8 Hz), 130.87, 126.59, 112.90, 91.80, 72.17 (d, J= 6.6 Hz), 66.10 (d, J = 166.6 Hz), 55.97, 26.07 - 22.47 (m). 31P NMR (162 MHz, Chloroform-d) δ 19.60. MS (ESI-QMS) m/z: [M + H]+ calcd for C14H23I05P, 429.0; found, 429.2.
Example 45
Diisopropyl (((3-iodopyridin-4-yl)oxy)methyl)phosphonate L45
Figure imgf000034_0002
4-Hydroxy-2-iodopyridine (442 mg, 2.00 mmol, 1 eq.) and potassium tert-butoxide (247 mg, 2.2 mmol, 1.1 eq.) were dissolved in dry dimethylformamide (5 ml) under an argon atmosphere. Then, TfOCH2P(O)(0/-Pr)2 (841 mg, 2.4 mmol, 1.20 eq.) was added and the mixture was stirred overnight at 70 °C. The reaction was quenched with a half-saturated aqueous solution of NH4CI and extracted with ethyl acetate (3x). The organic phase was washed with brine (1x), dried with MgSO4, filtered and solvents were evaporated. The solid was adsorbed on silica gel in a mixture of cyclohexane/acetone and purified by flash chromatography on silica gel (cyclohexane to 20% ethyl acetate modified with 10% methanol (v/vi)), yielded 642 mg (80%) of the title compound as a clear oil. 1H NMR (401 MHz, DMSO- d6) δ 8.22 (d, J= 2.3 Hz, 1H), 7.63 (dd, J= 7.5, 2.2 Hz, 1H), 6.18 (d, J= 7.5 Hz, 1H), 4.68 - 4.54 (m, 2H), 4.52 (d, J= 11.6 Hz, 2H), 1.26 - 1.20 (m, 12H). 13C NMR (101 MHz, DMSO-d6) δ 173.01, 146.53, 141.38, 113.09, 92.32, 71.45 (d, J= 6.7 Hz), 50.17 (d, J= 151.7 Hz), 25.92 - 21.19 (m). 31P NMR (162 MHz, DMSO-d6) δ 19.42. MS (ESI-QMS) m/z: [M + H]+ calcd for C12H20INO4P, 400.0; found, 400.2. Example 46
Diethyl (E )-(2-(5-iodofuran-2-yl)vinyl)phosphonate L46
Figure imgf000035_0001
Tetraethyl methylenediphosphonate (597 μl, 2.4 mmol, 1.2 eq.) was dissolved in dry tetrahydrofuran (4 ml) under an argon atmosphere and potassium tert-butoxidc (269 mg, 2.4 mmol, 1.2 eq.) was added. The mixture was stirred at room temperature for 1 hour and a solution of 5-iodofuran-2-carbaldehyde (444 mg, 2.0 mmol, 1 eq.) in dry tetrahydrofuran (1 ml) was added. The mixture was stirred at room temperature for 1 hour and the reaction was quenched with 1M HC1 (aq.), extracted with dichloromethane (3x), washed with brine (1x), dried with MgSO4 and filtered. Solvents were evaporated, solid was adsorbed on silica gel in a mixture of cyclohexane/acetone and it was purified by flash column chromatography on silica gel (cyclohexane to 40% ethyl acetate modified with 10% methanol (v/vi)), yielded 624 mg (88%) of the title compound as a white solid. 1H NMR (401 MHz, DMSO-d6) δ 7.12 (dd, J= 22.4, 17.4 Hz, 1H), 6.84 (d, J = 3.4 Hz, 1H), 6.82 (d, J = 3.5 Hz, 1H), 6.08 (dd, J = 17.5 Hz, 1H), 3.99 (dq, J = 8.2, 7.0 Hz, 4H), 1.24 (t, J = 7.0 Hz, 6H). 13C NMR (101 MHz, DMSO-d6) δ 155.60 (d, J= 26.3 Hz), 133.28 (d, J= 7.6 Hz), 122.77, 117.03, 111.88 (d, J= 190.5 Hz), 96.77, 61.36 (d, J= 5.4 Hz), 16.22 (d, J= 6.0 Hz). 31P NMR (162 MHz, DMSO-d6) δ 20.99. MS (ESI- QMS) m/z: [M + H]+ calcd for C10H15IO4P, 357.0; found, 357.1.
Example 47
Diethyl (E )-(2-(2-iodothiophen-3-yl)vinyl)phosphonate L47
Figure imgf000035_0002
Tetraethyl methylenediphosphonate (597 μl, 2.4 mmol, 1.2 eq.) was dissolved in dry tetrahydrofuran (4 ml) under an argon atmosphere and potassium tert-butoxidc (269 mg, 2.4 mmol, 1.2 eq.) was added. The mixture was stirred at room temperature for 1 hour and a solution of 2-iodothiophene-3- carbaldehyde (478 mg, 2.0 mmol, 1 eq.) in dry tetrahydrofuran (1 ml) was added. The mixture was stirred at room temperature for 1 hour and the reaction was quenched with 1M HC1 (aq.), extracted with dichloromethane (3x), washed with brine (1x), dried with MgSO4 and filtered. Solvents were evaporated, solid was adsorbed on silica gel in a mixture of cyclohexane/acetone and it was purified by flash column chromatography on silica gel (cyclohexane to 40% ethyl acetate modified with 10% methanol (v/vi)), yielded 596 mg (80%) of the title compound as a white solid. 1H NMR (401 MHz, DMSO-d6) δ 7.82 (dd, J= 5.6, 0.8 Hz, 1H), 7.45 (d, J= 5.6 Hz, 1H), 7.15 (dd, J= 22.4, 17.1 Hz, 1H), 6.51 (dd, J= 18.1, 17.3 Hz, 1H), 4.02 (dq, J= 8.2, 7.1 Hz, 4H), 1.26 (t, J= 7.0 Hz, 6H). 13C NMR (101 MHz, DMSO-d6) δ 141.41 (d, J= 7.3 Hz), 140.87 (d, J= 24.7 Hz), 132.75, 125.90, 116.75 (d, J= 185.7 Hz), 86.01, 61.35 (d, J= 5.4 Hz), 16.25 (d, J= 6.0 Hz). 31P NMR (162 MHz, DMSO-d6) δ 21.59. MS (ESI-QMS) m/z: [M + H]+ calcd for C10H15IO3PS, 373.0; found, 373.1.
Example 48
Diisopropyl (((thiophen-3-yl)oxy)methyl)phosphonate L48
Figure imgf000036_0001
Thiophene-3-ylboronic acid (10 g, 78.2 mmol, 1 eq.) was dissolved in diethyl ether (190 ml). The solution was stirred at room temperature and an aqueous solution of hydrogen peroxide (54 ml, 10% solution) was added dropwise. The mixture was stirred 1 hour at 40 °C and then cooled to room temperature. The mixture was extracted with diethyl ether (3x), washed with an aqueous solution of Mohr’s salt (3x, 10% solution), washed with brine (1x), dried with MgSO4 and filtered. Solvents were evaporated at 40 °C/900 mbar and the residue was dissolved in dry tetrahydrofuran (78 ml). The mixture was cooled to 0 °C and sodium hydride (3.9 g of 60% oil dispersion, 97.7 mmol, 1.25 eq.) was added portionwise. The mixture was stirred at 0 °C for 30 minutes and TfOCH2P(O)(Oi-Pr)2 (32 g, 97.7 mmol, 1.25 eq.) was added and the mixture was stirred 1 hour at 0 °C. The reaction was quenched with a half- saturated aqueous solution of NH4CI and extracted with ethyl acetate (3x). The organic phase was washed with brine (1x), dried with MgSO4, filtered and solvents were evaporated. The solid was adsorbed on silica gel in a mixture of cyclohexane/acetone and purified by flash chromatography on silica gel (cyclohexane to 20% ethyl acetate modified with 10% methanol (v/vi)), yielded 10.5 g (48%) of the title compound as a black liquid. 1H NMR (401 MHz, Chloroform -d) δ 7.16 (dd. J = 5.3, 3.1 Hz, 1H), 6.78 (dd, J= 5.3, 1.6 Hz, 1H), 6.35 (dd, J= 3.2, 1.6 Hz, 1H), 4.81 (dhept, J= 7.6, 6.2 Hz, 2H), 4.22 (d, J = 9.9 Hz, 2H), 1.40 - 1.28 (m, 12H). 13C NMR (101 MHz, Chloroform-d) δ 157.77 (d, J = 15.5 Hz), 125.21, 119.39, 98.69, 71.93 (d, J= 6.7 Hz), 64.89 (d, J= 171.2 Hz), 28.00 - 22.97 (m). 31P NMR (162 MHz, Chloroform-d) δ 19.40. MS (ESI-QMS) m/z: [M + H]+ calcd for C11H120O4PS, 279.1; found, 279.1.
Example 49
Diisopropyl (((2-iodothiophen-3-yl)oxy)methyl)phosphonate L49
Figure imgf000036_0002
Diisopropyl ((thiophen-3-yloxy)methyl)phosphonate (557 mg, 2.0 mmol, 1 eq.) was dissolved in absolute ethanol (10 ml) and N-iodosuccinimide (450 mg, 2.0 mmol, 1 eq.) was added. The mixture was stirred for 1 hour at room temperature. The mixture was diluted with water and extracted with ethyl acetate (3x), washed with brine (1x), dried with MgSO4 and filtered. Solvents were evaporated, the solid was adsorbed on silica gel in a mixture of cyclohexane/acetone and it was purified by flash column chromatography on silica gel (cyclohexane to 40% ethyl acetate modified with 10% methanol (v/vi)), yielded 770 mg (95%) of the title compound as a white solid. 1H NMR (401 MHz, DMSO-d6) δ 7.73 (d, J= 5.8 Hz, 1H), 6.99 (d, J= 5.8 Hz, 1H), 4.70 (dhept, J= 7.6, 6.2 Hz, 2H), 4.42 (d, J= 9.1 Hz, 2H), 1.30 - 1.26 (m, 12H). 13C NMR (101 MHz, DMSO-d6) δ 159.16 (d, J= 13.0 Hz), 131.12, 117.99, 99.79, 71.28 (d, J = 6.5 Hz), 65.77 (d, J = 165.0 Hz), 27.37 - 20.46 (m). 31P NMR (162 MHz, DMSO-d6) δ 19.29. MS (ESI-QMS) m/z: [M + H]+ calcd for C11H19I04PS, 405.0; found, 405.1.
Example 50
Sodium (E )-7-((2-(2-(phosphonato)vinyl)phenyl)thio)-3,5-dihydro-4H-pyrrolo[3,2-ri]pyrimidin-4-one
L50
Figure imgf000037_0001
Compound L4 (500 mg, 1.29 mmol, 1 eq.), copper(I) iodide (25 mg, 0.1290 mmol, 0.1 eq.) and cesium carbonate (505 mg, 1.55 mmol, 1.2 eq.) were charged into a microwave reactor tube and the tube was flushed with argon. Dry toluene (5 ml), compound L13 (559 mg, 1.33 mmol, 1.1 eq.) and 2-isobutyrylcyclohexanone (43 μl, 0.2580 mmol, 0.2 eq.) were added subsequently and the mixture was heated in the microwave reactor at 120 °C for 2 hours. The mixture was dissolved in chloroform, washed with a half-saturated aqueous solution of NaHCO3 (2x), 1M HC1 (aq.) (2x), brine (1x), dried with MgSO4, filtered and solvents were evaporated. The solid was adsorbed on silica gel in a mixture of cyclohexane/acetone and purified by flash chromatography on silica gel (cyclohexane to 15% ethyl acetate modified with 10% methanol (v/vi)), yielded 800 mg (99%) of the product as an oil. The compound was used in the next step without further characterization. MS (ESI-QMS) m/z: [M + H]+ calcd for C31H41N305PSSi, 626.2; found, 626.2. The compound was dissolved in trifluoroacetic acid (2 ml/ 100 mg of the material) at room temperature and stirred for 15 minutes. Trifluoroacetic acid was evaporated, coevaporated with water (2x) and a cold solution of ammonia in ethanol (2 ml/ 100 mg of the material) was added. The mixture was evaporated, the solid was adsorbed on silica gel in a mixture of cyclohexane/acetone and purified by flash chromatography on silica gel (chloroform to 15% methanol), yielded 250 mg (48%) of the diethyl phosphonate as a white solid. MS (ESI-QMS) m/z: [M + H]+ calcd for C18H2 1N304PS, 406.1; found, 406.2. A flask charged with the diethyl phosphonate (100 mg) was sealed with septum, flushed with argon, and then dry pyridine (10 ml/1 mmol of the starting compound) was added followed by trimethylsilyl bromide (1 ml/1 mmol of the starting compound). The mixture was stirred at room temperature overnight and then it was evaporated to dryness. The residue was dissolved in 2M aq. solution of triethylammonium bicarbonate and a small amount of methanol, and it was evaporated to dryness. The solid residue was dissolved in a small amount of water (solubility can be enhanced by addition of several drops of aqueous ammonia) and purified by HPLC (C18, gradient H2O /McOH). Product-containing fractions were filtered through a short pad of Dowex® 50 in Na+-cycle and solvents were evaporated. Purified product was lyophilized from water, yielded 86 mg (89%) of the title compound as a white solid. 1H NMR (400 MHz, Deuterium Oxide) δ 8.04 (d, J= 3.0 Hz, 1H), 7.81 (d, J= 2.9 Hz, 1H), 7.81 - 7.70 (m, 1H), 7.62 (dd, J= 7.7, 2.7 Hz, 1H), 7.19 (ddd, J= 7.6, 2.7 Hz, 1H), 7.10 (ddd, J= 7.6, 2.7 Hz, 1H), 6.89 (dd, J= 7.9, 2.9 Hz, 1H), 6.51 (dd, J= 16.8, 3.0 Hz, 1H). 13C NMR (101 MHz, Deuterium Oxide) δ 156.17, 147.22, 144.45, 138.59 (d, J = 6.1 Hz), 138.22, 136.89, 135.91 (d, J= 21.3 Hz), 130.63, 128.77, 128.08, 127.66, 127.10 (d, J= 175.8 Hz), 119.69, 104.05. 31P NMR (162 MHz, Deuterium Oxide) δ 16.37. HRMS (ESI-FTMS) m/z for the acid: [M - H]- calcd for C14H1104N3PS, 348.0213; found, 348.0211.
Example 51
Sodium 7-((2-((phosphonato)methoxy)phenyl)thio)-3.5-dihydro-4H-pyrrolo[3.2-d]|pyrimidin-4-onc L51
Figure imgf000038_0001
Compound L4 (500 mg, 1.29 mmol, 1 eq.), copper(I) iodide (25 mg, 0.1290 mmol, 0.1 eq.) and cesium carbonate (505 mg, 1.55 mmol, 1.2 eq.) were charged into a microwave reactor tube and the tube was flushed with argon. Dry toluene (5 ml), compound L14 (565 mg, 1.33 mmol, 1.1 eq.) and 2- isobutyrylcyclohexanone (43m1, 0.2580 mmol, 0.2 eq.) were added subsequently and the mixture was heated in the microwave reactor at 120 °C for 2 hours. The mixture was dissolved in chloroform, washed with a half-saturated aqueous solution of NaHCO3 (2x), 1M HC1 (aq.) (2x), brine (1x), dried with MgSO4, filtered and solvents were evaporated. The solid was adsorbed on silica gel in a mixture of cyclohexane/acetone and purified by flash chromatography on silica gel (cyclohexane to 15% ethyl acetate modified with 10% methanol (v/vi)), yielded 700 mg (81%) of the product as an oil. The compound was used in the next step without further characterization. MS (ESI-QMS) m/z: [M + H]+ calcd for C32H45N306PSSi, 658.3; found, 658.2. The compound (680 mg) was dissolved in trifluoroacetic acid (2 ml/100 mg of the material) at room temperature and stirred for 15 minutes. Trifluoroacetic acid was evaporated, coevaporated with water (2x) and a cold solution of ammonia in ethanol (2 ml/100 mg of the material) was added. The mixture was evaporated, the solid was adsorbed on silica gel in a mixture of cyclohexane/acetone and purified by flash chromatography on silica gel (chloroform to 15% methanol), yielded 320 mg (71%) of diisopropyl phosphonate as a white solid. MS (ESI-QMS) m/z: [M + H]+ calcd for C19H25N305PS, 438.1; found, 438.2. A flask charged with the diethyl phosphonate (100 mg) was sealed with septum, flushed with argon, and then dry pyridine (10 ml/1 mmol of the starting compound) was added followed by trimethylsilyl bromide (1 ml/1 mmol of the starting compound). The mixture was stirred at room temperature overnight and then it was evaporated to dryness. The residue was dissolved in a 2M aq. solution of triethylammonium bicarbonate and a small amount of methanol, and it was evaporated to dryness. The solid residue was dissolved in a small amount of water (solubility can be enhanced by addition of several drops of aqueous ammonia) and purified by HPLC (C18, gradient ffO/McOH). Product-containing fractions were filtered through a short pad of Dowex® 50 in Na+-cycle and solvents were evaporated. Purified product was lyophilized from water, yielded 72 mg (79%) of the title compound as a white solid. 1H NMR (401 MHz, Deuterium Oxide) δ 8.03 (s, 1H), 7.82 (s, 1H), 7.17 (ddd, J= 8.7, 7.2, 1.6 Hz, 1H), 7.11 (dd, J= 8.4, 1.3 Hz, 1H), 6.76 (ddd, J= 7.5, 1.3 Hz, 1H), 6.65 (dd, J= 7.8, 1.6 Hz, 1H), 4.23 (d, J= 9.8 Hz, 2H). 13C NMR (101 MHz, Deuterium Oxide) δ 156.66 (d, J = 13.2 Hz), 156.53, 147.59, 144.48, 137.34, 128.30, 128.04, 127.66, 123.21, 119.81, 113.62, 102.97, 66.53 (d, J= 156.8 Hz).31P NMR (162 MHz, Deuterium Oxide) d 16.07. HRMS (ESI-FTMS) m/z for the acid: [M - H]- calcd for C13H1105N3PS, 352.0163; found, 352.0158.
General procedure for compounds L52 - L74 (S4)
Compound L4 (100 mg, 0.2580 mmol, 1 eq.) and copper(I) iodide (5 mg, 0.0258 mmol, 0.1 eq.) were charged into a microwave reactor tube and the tube was flushed with argon. Dry dimethylformamide (2 ml), an aryl iodide (0.3096 mmol, 1.2 eq.), triethylamine (54 μl, 0.3870 mmol, 1.5 eq.) and 2- isobutyrylcyclohexanone (9 μl, 0.0516 mmol, 0.2 eq.) were added subsequently and the mixture was heated in the microwave reactor at 120 °C for 2-3 hours. The mixture was dissolved in chloroform, washed with a half-saturated aqueous solution of NaHCO3 (2x), 1M HC1 (aq.) (2x), brine (1x) and it was dried with MgSO4, filtered and solvents were evaporated. The solid was adsorbed on silica gel in a mixture of cyclohexane/acetone and purified by flash chromatography on silica gel (cyclohexane to 30% of ethyl acetate modified with 10% of methanol (v/vi)), yielded a coupling product as an oil. The compound was used in the next step without further characterization.
The compound was dissolved in trifluoroacetic acid (2 ml/ 100 mg of the material) at room temperature and stirred for 15 minutes. Trifluoroacetic acid was evaporated, coevaporated with water (2x) and a cold solution of ammonia in ethanol (2 ml/100 mg of the material) was added. The mixture was evaporated, the flask was sealed with septum, flushed with argon, and then dry pyridine (10 ml/ 1 mmol of the starting compound) was added followed by trimethylsilyl bromide (1 ml/1 mmol of the starting compound). The mixture was stirred at room temperature overnight and then it was evaporated to dryness. The residue was dissolved in a 2M aq. solution of triethylammonium bicarbonate and a small amount of methanol, and it was evaporated to dryness. The solid residue was dissolved in a small amount of water (solubility can be enhanced by addition of several drops of aqueous ammonia) and purified by HPLC (Cl 8, gradient H2O/MeOH). Product-containing fractions were filtered through a short pad of Dowex® 50 in Na+-cycle and solvents were evaporated. Purified product was lyophilized from water.
Example 52
Sodium (E )-7-((4-bromo-2-(2-(phosphonato)vinyl)phenyl)thio)-3,5-dihydro-4H-pyrrolo[3,2- d]pyrimidin-4-one L52
Figure imgf000040_0001
General procedure S4, starting with 300 mg of the compound L4 and diethyl (E )-(5-bromo-2- iodostyryl)phosphonate L17 (414 mg), afforded 216 mg (40%) of the coupling product and 71 mg (49%) of the title compound as a white solid. NMR spectra were collected as a diastereomeric mixture of cis- and trans- isomers because of light induced isomerization in solution. 1H NMR (401 MHz, Deuterium Oxide) δ 8.14 (d, J= 2.2 Hz, 1H), 8.03 (d, J= 1.5 Hz, 1H), 7.81 (d, J= 2.2 Hz, 1H), 7.67 (d, J= 13.6 Hz, 1H), 7.51 (dd, J= 18.5, 17.2 Hz, 1H), 7.23 (ddd, J= 12.3, 8.5, 2.2 Hz, 1H), 7.10 (dd, J= 39.2, 14.2 Hz, 1H), 6.70 (dd, J= 31.4, 8.5 Hz, 1H), 6.55 (dd, J= 17.2, 13.4 Hz, 1H), 6.21 (dd, J= 14.2, 10.8 Hz, 1H). 13C NMR (101 MHz, Deuterium Oxide) δ 165.68, 152.68, 149.36 (d, J= 6.1 Hz), 139.42, 139.36, 139.17, 138.82, 138.26 (d,J= 19.6 Hz), 137.96 (d, J= 7.7 Hz), 132.11, 131.69, 129.34, 127.85, 124.47, 124.40, 119.70, 118.64, 98.08, 97.84. 31P NMR (162 MHz, Deuterium Oxide) δ 12.23, 9.47. HRMS (ESI-FTMS) m/z for the acid: [M - H]- calcd for C14H10O4N3BrPS, 425.9319; found, 425.9315. Example 53
Sodium (E )-7-((5-bromo-2-(2-(phosphonato)vinyl)phenyl)thio)-3,5-dihydro-4H-pyrrolo[3,2- d]pyrimidin-4-one L53
Figure imgf000040_0002
General procedure S4, starting with 300 mg of the compound L4 and diethyl (E )-( 4-bromo-2- iodostyryl)phosphonate L20 (414 mg), afforded 184 mg (34%) of the coupling product and 60 mg (49%) of the title compound as a white solid. 1H NMR (401 MHz, Deuterium Oxide) δ 8.05 (s, 1H), 7.70 (s, 1H), 7.54 (d, J= 8.1 Hz, 1H), 7.50 (dd, J= 18.0 Hz, 1H), 7.33 (dd, J= 8.5, 2.0 Hz, 1H), 6.89 (d, J= 2.0 Hz, 1H), 6.55 (dd , J= 17.2, 13.7 Hz, 1H). 13C NMR (101 MHz, Deuterium Oxide) δ 165.78, 152.87, 149.40, 142.40, 139.37, 135.28 (d,J= 20.1 Hz), 133.99 (d, J= 6.1 Hz), 132.38 (d,J= 167.3 Hz), 129.48, 129.16, 124.57, 123.18, 97.51. 31P NMR (162 MHz, Deuterium Oxide) δ 12.59. HRMS (ESI-FTMS) m/z forthe acid: [M - H]- calcd for C14H10O4N3BrPS, 425.9319; found, 425.9316.
Example 54
Sodium (E )-7-((5-hydroxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3,5-dihydro-4H-pyrrolo[3,2- d]pyrimidin-4-one L54
Figure imgf000041_0001
General procedure S4, starting with 200 mg of the compound L4 and diethyl (E )-(4-hydroxy-2- iodostyryl)phosphonate L23 (198 mg), afforded 100 mg (30%) of the coupling product and 15 mg (23%) of the title compound as a white solid. 1H NMR (401 MHz, Deuterium Oxide) δ 8.10 (s, 1H), 7.76 (s, 1H), 7.56 (dd, J= 19.0, 17.3 Hz, 1H), 7.44 (d, J= 8.6 Hz, 1H), 6.45 (ddd, J= 8.6, 2.6, 0.6 Hz, 1H), 6.25 (dd, J= 17.2, 14.3 Hz, 1H), 6.24 (d, J= 2.6 Hz, 1H). 13C NMR (101 MHz, Deuterium Oxide) δ 166.50, 163.91, 152.70, 148.30, 139.05, 135.09 (d, J= 4.3 Hz), 128.81, 125.70 (d, J= 171.5 Hz), 124.18 (d, J = 20.2 Hz), 120.97, 117.99, 117.66, 102.13. 31P NMR (162 MHz, Deuterium Oxide) δ 16.37. HRMS (ESI-FTMS) m/z forthe acid: [M - H]- calcd for C14H1105N3PS, 364.0163; found, 364.0157. Example 55
Sodium (E )-7-((5-methoxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3,5-dihydro-4H-pyrrolo[3,2- d]pyrimidin-4-one L55
Figure imgf000041_0002
General procedure S4 with diethyl (E )-(2-iodo-4-methoxystyryl)phosphonate L31 (123 mg) afforded 104 mg (62%) of the coupling product and 27 mg (59%) of the title compound as a white solid. 1H NMR (401 MHz, DMSO-d6) δ 7.84 (s, 1H), 7.54 (s, 1H), 7.42 (d, J= 8.6 Hz, 1H), 7.30 (dd, J= 18.9, 17.2 Hz, 1H), 6.63 (dd, J= 8.7, 2.7 Hz, 1H), 6.22 (dd, .J= 17.2, 13.6 Hz, 1H), 6.16 (d, J= 2.6 Hz, 1H), 3.48 (s, 3H). 13C NMR (101 MHz, DMSO-d6) δ 162.68, 159.96, 151.23, 147.81, 140.56, 134.39, 133.27, 130.06 (d, J = 167.9 Hz), 128.61, 121.22, 113.34, 111.21, 111.12, 100.56, 55.98. 31P NMR (162 MHz, Deuterium Oxide) δ 13.38. HRMS (ESI-FTMS) m/z for the acid: [M - H]- calcd for C15H1305N3PS, 378.0319; found, 378.0315.
Example 56
Sodium (E) -7 -((5 -isopropoxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3 ,5 -dihydro-4H-pyrrolo [3 ,2- ri|pyrimidin-4-onc L56
Figure imgf000042_0001
General procedure S4 with diethyl (E )-(2-iodo-4-isopropoxystyryl)phosphonate L32 (131 mg) afforded 170 mg (50%) of the coupling product and 32 mg (28%) of the title compound as a white solid. 1H NMR (401 MHz, DMSO-d6) δ 7.89 (s, 1H), 7.54 (s, 1H), 7.40 (d, J= 8.5 Hz, 1H), 7.28 (dd, J= 18.7, 17.3 Hz, 1H), 6.56 (dd, J= 8.6, 2.6 Hz, 1H), 6.23 (dd, J= 17.2, 13.9 Hz, 1H), 6.12 (d, J= 2.6 Hz, 1H), 4.12 (hept, J= 6.1 Hz, 1H), 0.93 (d, J= 6.0 Hz, 6H). 13C NMR (101 MHz, DMSO-d6) δ 164.88, 158.53, 153.77, 148.76, 140.82, 135.30, 134.17 (d,J= 5.8 Hz), 130.09 (d,J= 155.5 Hz), 129.63 (d, J= 20.2 Hz), 129.19, 121.65, 114.97, 114.38, 100.94, 72.40, 22.56. 31P NMR (162 MHz, Deuterium Oxide) δ 13.38. HRMS (ESI-FTMS) m/z for the acid: [M - H]- calcd for C17H1705N3PS, 406.0632; found, 406.0628. Example 57
Sodium (E )-7-((5-(2,3,4,5,6-pentafluorophenyl)methoxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3,5- dihydro-4H-pyrrolo|3.2-ri|pyrimidin-4-onc L57
Figure imgf000042_0002
General procedure S4 with diethyl (E )-(2-iodo-4-((2, 3,4,5, 6-pentafluorophenyl)methoxy)styryl)- phosphonate L33 (174 mg) afforded 153 mg (72%) of the coupling product and 9 mg (8%) of the title compound as a white solid. 1H NMR (500 MHz, Deuterium Oxide) δ 7.98 (s, 1H), 7.71 (s, 1H), 7.69 - 7.59 (m, 1H), 7.53 (d, J= 8.6 Hz, 1H), 6.76 (d, J= 8.7 Hz, 1H), 6.38 (dd, J= 16.7 Hz, 1H), 6.23 (s, 1H), 4.97 (s, 3H). 13C NMR (126 MHz, Deuterium Oxide) δ 158.96, 155.94, 148.25 - 146.09 (m), 146.99, 145.39 - 144.93 (m), 144.04, 140.31, 139.46 - 138.73 (m), 137.58 (d, J= 6.4 Hz), 136.63, 129.25 (d, J = 21.8 Hz), 129.11, 124.60 (d, J = 177.4 Hz), 119.64, 114.30, 113.50, 111.43 - 110.01 (m), 103.08, 59.01. 19F NMR (377 MHz, Deuterium Oxide) δ -139.07 - -145.81 (m), -153.48 (t, J= 20.9 Hz), -162.37 - -162.67 (m). 31P NMR (162 MHz, Deuterium Oxide) δ 13.41. HRMS (ESI-FTMS) m/z for the acid: [M - H]- calcd for C21H1205N3F5PS, 544.0161; found, 544.0154.
Example 58
Sodium (E )-7-((5-(2,3,4,5,6-pentafluorophenyl)oxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3,5- dihydro-4H-pyrrolo|3.2-ri|pyrimidin-4-onc L58
Figure imgf000043_0001
General procedure S4 with diethyl (E )-(2-iodo-4-((2,3,4,5,6-pentafluorophenyl)oxy)styryl)phosphonate L34 (170 mg) afforded 126 mg (78%) of the coupling product and 9 mg (12%) of the title compound as a white solid. 1H NMR (500 MHz, Deuterium Oxide) δ 7.95 (d, J= 0.7 Hz, 1H), 7.66 (d, J= 8.6 Hz, 1H), 7.59 - 7.52 (m, 1H), 7.42 (t, J= 17.9 Hz, 1H), 6.94 (dd, J= 8.6, 2.8 Hz, 1H), 6.50 (dd, J= 17.2, 14.3 Hz, 1H), 6.07 (d,J= 2.2 Hz, 1H). 13CNMR(126 MHz, Deuterium Oxide) δ 165.44, 158.15, 152.47, 149.17, 142.74, 138.11, 133.53 (d,J= 5.6 Hz), 131.44 (d, .J= 20.1 Hz), 131.22 (d,J= 168.0 Hz), 129.48, 124.10, 113.67, 112.05, 97.73. 19F NMR (377 MHz, Deuterium Oxide) δ -155.14 (d, J = 18.8 Hz), - 159.87 (t, J= 22.1 Hz), -162.17 - -162.35 (m).31P NMR (162 MHz, Deuterium Oxide) δ 13.42. HRMS (ESI-FTMS) m/z for the acid: [M - H]- calcd for C20H1005N3F5PS, 530.0004; found, 530.0000. Example 59
Sodium (E )-7-((4-hydroxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3,5-dihydro-4H-pyrrolo[3,2- ri|pyrimidin-4-onc L59
Figure imgf000043_0002
General procedure S4 with diethyl (E )-(5-hydroxy-2-iodostyryl)phosphonate L26 (99 mg) afforded 120 mg (73%) of the coupling product and 44 mg (58%) of the title compound as a white-off solid. 1H NMR (401 MHz, Deuterium Oxide) δ 8.06 (s, 1H), 7.83 (s, 1H), 7.71 (dd, J= 18.1 Hz, 1H), 7.10 (d, J = 2.7 Hz, 1H), 7.08 (d, J = 8.7 Hz, 1H), 6.65 (dd, J= 8.6, 2.7 Hz, 1H), 6.47 (dd, J= 17.3, 14.1 Hz, 1H). 13C NMR (101 MHz, Deuterium Oxide) δ 156.75, 156.69, 147.08, 144.66, 139.97 (d, J= 20.3 Hz), 136.55, 136.19 (d, J = 5.3 Hz), 133.44, 131.15 (d, J= 169.1 Hz), 127.81, 119.43, 117.82, 114.33, 106.50. 31P NMR (162 MHz, Deuterium Oxide) δ 12.90. HRMS (ESI-FTMS) m/z for the acid: [M - H]- calcd for C14H1105N3PS, 364.0163; found, 364.0159. Example 60
Sodium (E )-7-((4-methoxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3,5-dihydro-4H-pyrrolo[3,2- d]pyrimidin-4-onc L60
Figure imgf000044_0001
General procedure S4 with diethyl (E )-(2-iodo-5-methoxystyryl)phosphonate L35 (123 mg) afforded 160 mg (95%) of the coupling product and 51 mg (49%) of the title compound as a white solid. 1H NMR
(401 MHz, Deuterium Oxide) δ 8.05 (s, 1H), 7.83 (s, 1H), 7.69 (dd, J= 18.1 Hz, 1H), 7.20 (d, J = 2.9
Hz, 1H), 7.01 (d, J= 8.7 Hz, 1H), 6.67 (dd, J= 8.8, 2.8 Hz, 1H), 6.54 (dd, J= 17.3, 13.9 Hz, 1H), 3.80 (s, 3H). 13C NMR (101 MHz, Deuterium Oxide) δ 159.32, 156.86, 147.20, 144.87, 139.47 (d, J= 19.9 Hz), 136.70, 135.78 (d, J= 5.9 Hz), 132.38, 131.84 (d, J= 168.1 Hz), 129.16, 119.55, 116.30, 112.80, 105.80, 56.89. 31P NMR (162 MHz, Deuterium Oxide) δ 12.74. HRMS (ESI-FTMS) m/z for the acid:
[M - H]- calcd for C15H1305N3PS, 378.0319; found, 378.0315.
Example 61
Sodium (E )-7-((4-isopropoxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3,5-dihydro-4H-pyrrolo[3,2- d]pyrimidin-4-one (236) L61
Figure imgf000044_0002
General procedure S4 with diethyl (E )-(2-iodo-5-isopropoxystyryl)phosphonate L36 (131 mg) afforded 150 mg (85%) of the coupling product and 38 mg (38%) of the title compound as a white-off solid. 1H NMR (401 MHz, Deuterium Oxide) δ 8.04 (s, 1H), 7.81 (s, 1H), 7.80 (dd, J= 20.2, 17.3 Hz, 1H), 7.15 (d, J= 2.8 Hz, 1H), 7.06 (d, J= 8.7 Hz, 1H), 6.64 (dd, J= 8.8, 2.8 Hz, 1H), 6.48 (dd, J= 17.3, 15.4 Hz, 1H), 4.56 (hept, J= 6.1 Hz, 1H), 1.27 (d, J = 6.2 Hz, 6H). 13C NMR (101 MHz, Deuterium Oxide) δ 157.50, 156.34, 147.06, 144.42, 139.01 (d, J= 21.0 Hz), 138.17 (d, J= 6.4 Hz), 136.61, 133.03, 129.56, 128.57 (d, J= 173.6 Hz), 119.43, 118.67, 115.33, 106.07, 72.99, 22.62.31P NMR (162 MHz, Deuterium Oxide) δ 13.70. HRMS (ESI-FTMS) m/z for the acid: [M - H]- calcd for C17H1705N3PS, 406.0632; found, 406.0627.
Example 62 Sodium (E )-7-((4-(2,3,4,5,6-pentafluorophenyl)methoxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3,5- dihydro-4H-pyrrolo[3.2-d]|pyrimidin-4-onc L62
Figure imgf000045_0001
General procedure S4 with diethyl (E )-(2-iodo-5-((2, 3,4,5, 6-pentafluorophenyl)methoxy)- styryl)phosphonate L37 (174 mg) afforded 147 mg (69%) of the coupling product and 56 mg (54%) of the title compound as a white-off solid. 1H NMR (401 MHz, Deuterium Oxide) δ 8.05 (s, 1H), 7.83 (s, 1H), 7.67 (dd, J= 18.1 Hz, 1H), 7.30 (d, J= 2.8 Hz, 1H), 6.98 (d, J= 8.7 Hz, 1H), 6.69 (dd, J= 8.8, 2.8 Hz, 1H), 6.55 (dd, J= 17.3, 13.7 Hz, 1H), 5.23 (s, 2H). 13CNMR(101 MHz, Deuterium Oxide) δ 157.67, 157.18, 147.32, 145.30, 139.47 (d, J= 19.8 Hz), 136.76, 135.29 (d, J= 5.7 Hz), 132.40 (d, J= 167.4 Hz), 131.97, 130.71, 119.68, 117.38, 114.08, 105.26, 59.58. 19F NMR (377 MHz, Deuterium Oxide) δ - 143.07 (dd, J = 23.0, 8.3 Hz), -153.03 (t, J = 20.9 Hz), -161.92 - -162.22 (m).31P NMR (162 MHz, Deuterium Oxide) δ 12.51. HRMS (ESI-FTMS) m/z for the acid: [M - H]- calcd for C21H1205N3F5PS, 544.0161; found, 544.0155.
Example 63
Sodium (E )-7-((4-(2,3,4,5,6-pentafluorophenyl)oxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3,5- dihydro-4H-pyrrolo[3.2-d]|pyrimidin-4-one L63
Figure imgf000045_0002
General procedure S4 with diethyl (E )-(2-iodo-5-((2,3,4,5,6-pentafluorophenyl)oxy)styryl)phosphonate L38 (174 mg) afforded 169 mg (81%) of the coupling product and 70 mg (58%) of the title compound as a white-off solid. 1H NMR (401 MHz, Deuterium Oxide) δ 8.04 (s, 1H), 7.83 (s, 1H), 7.61 (dd, J = 17.9 Hz, 1H), 7.26 (d, J= 2.8 Hz, 1H), 6.89 (d, J= 8.8 Hz, 1H), 6.63 (dd, J= 8.8, 2.9 Hz, 1H), 6.48 (dd, J= 17.3, 13.6 Hz, 1H). 13C NMR (101 MHz, Deuterium Oxide) δ 157.01, 147.34, 145.17, 139.43 (d, J = 20.2 Hz), 136.97, 134.73 (d, J= 6.2 Hz), 133.14 (d, J= 167.1 Hz), 132.97, 131.24, 119.78, 116.67, 114.54, 104.52. 19F NMR (377 MHz, Deuterium Oxide) δ -155.26 (d, J= 18.8 Hz), -159.99 (t, J= 22.1 Hz), -162.29 - -162.47 (m). 31P NMR (162 MHz, Deuterium Oxide) δ 12.22. HRMS (ESI-FTMS) m/z for the acid: [M - H]- calcd for C20H1005N3F5PS, 530.0004; found, 529.9999.
Example 64 Sodium (E )-7-((3-hydroxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3,5-dihydro-4H-pyrrolo[3,2- d]pyrimidin-4-onc L64
Figure imgf000046_0001
General procedure S4 with diethyl (E )-(2-hydroxy-6-iodostyryl)phosphonate L30 (99 mg) (100 °C, 1 hour) afforded 100 mg (60%) of the coupling product and 10 mg (16%) of the title compound as a white solid. 1H NMR (401 MHz, Deuterium Oxide) δ 8.07 (s, 1H), 7.84 (s, 1H), 7.25 (dd, J= 20.1, 17.9 Hz, 1H), 6.96 (dd, J= 8.1 Hz, 1H), 6.73 (dd, J= 8.1, 1.1 Hz, 1H), 6.58 (dd, J= 17.9, 15.7 Hz, 1H), 6.36 (dd, J= 8.0, 1.1 Hz, 1H). 13C NMR (101 MHZ, Deuterium Oxide) δ 156.90, 155.60, 147.56, 145.09, 140.08, 137.08, 136.14 (d, J= 165.9 Hz), 132.47 (d, J= 5.2 Hz), 130.04, 124.52 (d,J= 19.6 Hz), 119.87, 119.31, 114.67, 104.19. 31P NMR (162 MHz, Deuterium Oxide) δ 12.72. HRMS (ESI-FTMS) m/z for the acid: [M - H]- calcd for C14H1105N3PS, 364.0163; found, 364.0159.
Example 65
Sodium (E )-7-((3-methoxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3,5-dihydro-4H-pyrrolo[3,2- d]pyrimidin-4-one L65
Figure imgf000046_0002
General procedure S4 with diethyl (E )-(2-iodo-6-methoxystyryl)phosphonate L39 (123 mg) afforded 118 mg (70%) of the coupling product and 39 mg (51%) of the title compound as a white solid. 1H NMR (401 MHz, Deuterium Oxide) δ 8.07 (s, 1H), 7.87 (s, 1H), 7.51 (dd, J= 22.4, 17.8 Hz, 1H), 7.13 (dd, J = 8.2 Hz, 1H), 6.92 (d, J= 8.3 Hz, 1H), 6.71 (dd, J= 17.9 Hz, 1H), 6.51 (d, J= 8.0 Hz, 1H), 3.90 (s, 3H). 13C NMR (101 MHz, Deuterium Oxide) δ 159.29, 156.21, 147.44, 144.51, 140.42, 137.02, 134.98 (d, J = 5.9 Hz), 131.39 (d, J = 173.0 Hz), 130.61, 124.35 (d, J = 20.9 Hz), 120.41, 119.79, 110.27, 104.33, 57.15. 31P NMR (162 MHz, Deuterium Oxide) δ 13.13. HRMS (ESI-FTMS) m/z for the acid: [M - H]- calcd for C15H1305N3PS, 378.0319; found, 378.0311.
Example 66
Sodium (E ) -7 -((3 -isopropoxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3 ,5 -dihydro-4H-pyrrolo [3 ,2- d]pyrimidin-4-one L66 General procedure S4 with diethyl (E )-(2-iodo-6-isopropoxystyryl)phosphonate L40 (131 mg) afforded 160 mg (91%) of the coupling product and 34 mg (32%) of the title compound as a white solid. 1H NMR (500 MHz, Deuterium Oxide) δ 8.04 (s, 1H), 7.79 (s, 1H), 7.48 (dd, J= 22.6, 17.8 Hz, 1H), 7.05 (dd, J = 8.2 Hz, 1H), 6.91 (d, J= 8.2 Hz, 1H), 6.70 (dd, J= 18.1 Hz, 1H), 6.49 (d, J= 8.1 Hz, 1H), 4.68 - 4.58 (m, 1H), 1.35 (d, J = 6.1 Hz, 6H). 13C NMR (126 MHz, Deuterium Oxide) δ 157.30, 156.62, 147.37, 144.61, 140.39, 137.05, 136.18 (d, J= 6.3 Hz), 130.71 (d, J= 173.7 Hz), 130.49, 129.71, 126.52 (d, J = 19.2 Hz), 121.04, 119.85, 115.00, 104.21, 74.90, 22.67. 31P NMR (162 MHz, Deuterium Oxide) δ 13.16. HRMS (ESI-FTMS) m/z for the acid: [M - H]- calcd for C17H1705N3PS, 406.0632; found, 406.0628.
Example 67
Sodium (E )-7-((3-(2,3,4,5,6-pentafluorophenyl)methoxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3,5- dihydro-4H-pyrrolo[3.2-d]|pyrimidin-4-one L67
Figure imgf000047_0001
General procedure S4 with diethyl (E )-(2-iodo-6-((2,3,4,5,6- pentafluorophenyl)methoxy)styryl)phosphonate L41 (174 mg) afforded 125 mg (59%) of the coupling product and 54 mg (60%) of the title compound as a white solid. 1H NMR (500 MHz, Deuterium Oxide) d 8.45 (s, 1H), 8.19 (s, 1H), 7.90 (dd, J= 22.4, 17.8 Hz, 1H), 7.48 (dd, ./= 8.1 Hz, 1H), 7.34 (d, J= 8.1 Hz, 1H), 7.03 (dd, J = 17.8 Hz, 1H), 6.99 (d, J = 7.9 Hz, 2H), 5.69 (s, 2H). 13C NMR (126 MHz, Deuterium Oxide) δ 157.65, 156.44, 148.38 - 147.69 (m), 147.53, 146.48 - 145.66 (m), 144.48, 141.04, 140.16 - 139.67 (m), 136.74, 135.20 (d, J= 6.2 Hz), 131.58 (d, J= 172.3 Hz), 130.14, 126.41 (d, J = 23.9 Hz), 122.24, 120.00, 112.92, 111.83 - 111.25 (m), 105.02, 60.24. 19F NMR (377 MHz, Deuterium Oxide) δ -138.45 - -145.19 (m), -152.86 (t, J= 20.9 Hz), -161.75 - -162.05 (m). 31P NMR (162 MHz, Deuterium Oxide) δ 14.84. HRMS (ESI-FTMS) m/z for the acid: [M - H]- calcd for C21H1205N3F5PS, 544.0161; found, 544.0155.
Example 68 Sodium (E )-7-((3-(2,3,4,5,6-pentafluorophenyl)oxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3,5- dihydro-4H-pyrrolo[3.2-d]|pyrimidin-4-onc L68
Figure imgf000048_0001
General procedure S4 with diethyl (E )-(2-iodo-6-((2,3,4,5,6-pentafluorophenyl)oxy)styryl)phosphonate L42 (174 mg) afforded 175 mg (84%) of the coupling product and 78 mg (62%) of the title compound as a white-off solid. 1H NMR (401 MHz, Deuterium Oxide) δ 8.07 (s, 1H), 7.83 (s, 1H), 7.37 (dd, J = 20.0, 17.8 Hz, 1H), 6.93 (dd, J= 8.2 Hz, 1H), 6.75 (dd, J= 17.8, 14.8 Hz, 1H), 6.59 (d, J= 8.2 Hz, 1H), 6.55 (d, J = 8.0 Hz, 1H). 13C NMR (101 MHz, Deuterium Oxide) δ 157.28, 156.29, 147.62, 145.55, 141.44, 137.79 (d,J= 165.0 Hz), 137.15, 130.47 (d, J= 5.0 Hz), 129.77, 126.13 (d,J= 19.3 Hz), 122.81, 120.04, 112.13, 103.44. 19F NMR (377 MHz, Deuterium Oxide) δ -155.40 (d, J= 21.0 Hz), -160.34 (t, J = 22.2 Hz), -162.44 - -162.72 (m). 31P NMR (162 MHz, Deuterium Oxide) δ 12.29. HRMS (ESI- FTMS) m/z for the acid: [M - H]- calcd for C20H1005N3F5PS, 530.0004; found, 530.0000.
Example 69
Sodium 7-((5-fluoro-2-((phosphonato)methoxy)phenyl)thio)-3.5-dihydro-4H-pyrrolo[3.2-d]|pyrimidin- 4-one L69
Figure imgf000048_0002
General procedure S4 with diisopropyl ((4-fluoro-2-iodophenoxy)methyl)phosphonate L43 (129 mg) afforded 105 mg (63%) of the coupling product and 21 mg (32%) of the title compound as a white solid. 1H NMR (401 MHz, Deuterium Oxide) δ 8.07 (s, 1H), 7.86 (s, 1H), 7.10 (dd, J= 9.1, 4.5 Hz, 1H), 6.89 (ddd, J= 8.7, 3.0 Hz, 1H), 6.31 (dd, J= 9.1, 3.0 Hz, 1H), 4.16 (d, J= 9.6 Hz, 2H). 13C NMR (101 MHz, Deuterium Oxide) δ 158.59 (d, J= 239.1 Hz), 156.30, 152.89 (d, J= 13.0 Hz), 147.44, 144.63, 137.23, 130.18 (d, J= 7.8 Hz), 119.96, 114.57 (d, J = 8.5 Hz), 113.87 (d, J= 26.4 Hz), 113.62 (d, J= 23.3 Hz), 102.18, 67.67 (d, J= 154.7 Hz). 19F NMR (377 MHz, Deuterium Oxide) δ -122.28 (d, J= 5.4 Hz). 31P NMR (162 MHz, Deuterium Oxide) δ 15.10. HRMS (ESI-FTMS) m/z for the acid: [M - H]- calcd for C13H10O5N3FPS, 370.0068; found, 370.0064.
Example 70 Sodium 7 -((3 -methoxy-2-((phosphonato)methoxy)phenyl)thio)-3 ,5 -dihydro-4H-pyrrolo [3,2- d]pyrimidin-4-onc L70
Figure imgf000049_0001
General procedure S4 with diisopropyl ((2-iodo-6-methoxyphenoxy)methyl)phosphonate L44 (133 mg) afforded 114 mg (64%) of the coupling product and 32 mg (46%) of the title compound as a white solid. 1H NMR (401 MHz, Deuterium Oxide) δ 7.99 (s, 1H), 7.63 (s, 1H), 6.92 (dd, J= 8.0 Hz, 1H), 6.87 (dd, J = 8.3, 1.7 Hz, 1H), 6.25 (dd, J= 7.8, 1.7 Hz, 1H), 4.11 (d, J= 9.5 Hz, 2H), 3.92 (s, 3H). 13C NMR (101 MHz, Deuterium Oxide) δ 166.33, 153.30, 152.04, 150.15, 145.13 (d,J= 13.6 Hz), 141.85, 136.73, 126.36, 126.14, 119.40, 110.71, 95.72, 72.23 (d, J= 145.9 Hz), 57.38.31P NMR (162 MHz, Deuterium Oxide) δ 14.84. HRMS (ESI-FTMS) m/z for the acid: [M - H]- calcd for C14H1306N3PS, 382.0268; found, 382.0265.
Example 71
Sodium 7-((4-((phosphonato)methoxy)pyridinc-3-yl)thio)-3.5-dihydro-4H-pyrrolo[3.2-d]|pyrimidin-4- one L71
Figure imgf000049_0002
General procedure S4 with diisopropyl (((3-iodopyridin-4-yl)oxy)methyl)phosphonate L45 (124 mg) afforded 57 mg (33%) of the coupling product and 11 mg (32%) of the title compound as a white solid. 1H NMR (401 MHz, Deuterium Oxide) δ 8.10 (s, OH), 7.89 (s, OH), 7.82 (d, J= 7.5 Hz, 1H), 7.35 (s, OH), 6.51 (d, J = 7.2 Hz, OH), 3.83 (d, J = 11.5 Hz, 2H). 13C NMR (101 MHz, Deuterium Oxide) δ 176.03, 156.08, 147.00, 145.16, 142.70, 139.98, 136.66, 129.11, 120.01, 115.33, 101.66, 58.02 (d, J = 128.5 Hz). 31P NMR (162 MHz, Deuterium Oxide) δ 11.50. HRMS (ESI-FTMS) m/z for the acid: [M - H]- calcd for C12H10N4O5PS, 353.0115; found, 353.0113.
Example 72
Sodium (E)-7-((5-(2-(phosphonato)\ inyl)furan-2-yl)thio)-3.5-dihydro-4H-pyrrolo[3.2-d]|pyrimidin-4- one L72 General procedure S4 with diethyl (E )-(2-(5-iodofuran-2-yl)vinyl)phosphonate L46 (110 mg) afforded 104 mg (65%) of the coupling product and 23 mg (43%) of the title compound as a white solid. 1H NMR (401 MHz, Deuterium Oxide) δ 8.15 (d, J= 1.3 Hz, 1H), 7.91 (d, J= 1.4 Hz, 1H), 6.75 (ddd, J= 17.7, 15.3, 1.3 Hz, 2H), 6.73 (d, J= 1.6 Hz, 1H), 6.51 (d, J= 3.3 Hz, 1H), 6.33 (ddd, J= 17.6, 13.9, 1.5 Hz,
1H). 13C NMR (101 MHZ, Deuterium Oxide) δ 156.61 (d, J= 23.1 Hz), 156.61, 146.41, 146.33, 144.95, 135.41, 127.49 (d, J = 178.2 Hz), 126.67, 119.23, 118.98, 112.94, 105.47. 31P NMR (162 MHz, Deuterium Oxide) δ 12.43. HRMS (ESI-FTMS) m/z for the acid: [M - H]- calcd for C12H9N305PS, 338.0006; found, 338.0001. Example 73
Sodium (E)-7 -((3 -(2-(phosphonato)vinyl)thiophene-2-yl)thio)-3 ,5 -dihydro-4H-pyrrolo [3,2- d]pyrimidin-4-one L73
Figure imgf000050_0001
General procedure S4 with diethyl (E )-(2-(2-iodothiophen-3-yl)vinyl)phosphonate L47 (115 mg) afforded 118 mg (72%) of the coupling product and 25 mg (33%) of the title compound as a white solid. 1H NMR (401 MHz, Deuterium Oxide) δ 8.13 (d, J= 0.9 Hz, 1H), 7.84 (d, J= 0.9 Hz, 1H), 7.46 (dd, J = 18.4, 17.4 Hz, 1H), 7.38 - 7.26 (m, 2H), 6.41 (ddd, J= 17.5, 13.8, 0.9 Hz, 1H). 13C NMR (101 MHz, Deuterium Oxide) δ 164.55, 153.49, 147.75, 141.49 (d, J = 20.9 Hz), 136.30, 134.27, 130.73 (d, J = 168.6 Hz), 130.33 (d, J = 6.0 Hz), 128.46, 127.45, 120.73, 104.90. 31P NMR (162 MHz, Deuterium Oxide) δ 12.92. HRMS (ESI-FTMS) m/z for the acid: [M - H]- calcd for C12H9N304PS2, 353.9778; found, 353.9775.
Example 74
Sodium 7-((3-((phosphonato)methoxy)thiophene-2-yl)thio)-3.5-dihydro-4H-pyrrolo[3.2-d]|pyrimidin- 4-one L74 General procedure S4 with diisopropyl (((2-iodothiophen-3-yl)oxy)methyl)phosphonate L49 (125 mg) afforded 133 mg (77%) of the coupling product and 28 mg (35%) of the title compound as a white solid. 1H NMR (401 MHz, Deuterium Oxide) δ 8.14 (s, 1H), 8.04 (d, J= 0.7 Hz, 1H), 7.40 (dd, J= 5.8, 0.7 Hz, 1H), 7.06 (d, J= 5.8 Hz, 1H), 4.13 (d, J= 9.5 Hz, 2H). 13C NMR (101 MHz, Deuterium Oxide) δ 160.87 (d, J = 13.1 Hz), 144.58, 136.19, 128.90, 119.01, 118.65, 110.74, 108.71, 70.84 (d, J= 151.8 Hz). 31P NMR (162 MHz, Deuterium Oxide) δ 14.30. HRMS (ESI-FTMS) m/z for the acid: [M - H| calcd for C11H9N305PS2, 357.9727; found, 357.9722.
Eample 75
Ethyl (E )-2-(2-iodophenyl)ethene- 1 -sulfonate L75
Figure imgf000051_0001
Ethyl (diethoxyphosphoryl)methanesulfonate (673 mg, 2.6 mmol, 1.2 eq.), prepared by published procedure [Otrubova, K.; Cravatt, B. F.; Boger, D. L. J. Med. Chem. 2014, 57 (3), 1079-1089], was dissolved in dry tetrahydrofiiran (20 ml) under an argon atmosphere and potassium tert-butoxidc (290 mg, 2.6 mmol, 1.2 eq.) was added. The mixture was stirred at room temperature for 1 hour and a solution of 2-iodobenzaldehyde (500 mg, 2.2 mmol, 1 eq.) in dry tetrahydrofiiran (10 ml) was added dropwise. The reaction was stirred at room temperature for 1 hour and it was quenched with a half-saturated aqueous solution of NH4CI, extracted with dichloromethane (3x), washed with brine (1x) and dried with MgSO4. Solvents were evaporated, the crude mixture was adsorbed on silica gel in a mixture of pentane/dichloromethane and purified by flash chromatography on silica gel (pentane to 50% dichloromethane), yielded 648 mg (89%) of the title compound as a clear oil. 1H NMR (401 MHz, Chloroform-d) δ 7.93 (dd, J= 8.0, 1.2 Hz, 1H), 7.84 (d, J= 15.4 Hz, 1H), 7.52 (dd, J= 7.8, 1.6 Hz, 1H), 7.41 (dddd, J= 7.9, 7.4, 1.2, 0.6 Hz, 1H), 7.13 (ddd, J= 7.9, 7.3, 1.7 Hz, 1H), 6.64 (d, J= 15.4 Hz, 1H), 4.28 (q, J= 7.1 Hz, 2H), 1.43 (t, J= 7.1 Hz, 3H). 13C NMR (101 MHz, Chloroform-d) δ 147.83, 140.36, 135.88, 132.33, 128.96, 127.91, 124.47, 101.07, 67.41, 15.09. MS (ESI-QMS) m/z: [M + H]+ calcd for C10H12IO3S, 339.0; found, 339.0.
Example 76
Ethyl (E )-3-(2-iodophenyl)acrylate L76 Triethyl phosphonoacetate (1 g, 5.2 mmol, 1.2 eq.) was dissolved in dry tetrahydrofuran (20 ml) under an argon atmosphere and potassium tert-butoxidc (580 mg, 5.2 g, 1.2 eq.) was added. The mixture was stirred at room temperature for 1 hour and a solution of 2-iodobenzaldehyde (1 g, 4.3 mmol, 1 eq.) in dry tetrahydrofuran (10 ml) was added dropwise. The reaction was stirred at room temperature for 1 hour and it was quenched with a half-saturated aqueous solution of NH4CI, extracted with dichloromethane (3x), washed with brine (1x) and dried with MgSCri. Solvents were evaporated, the crude mixture was adsorbed on silica gel in a mixture of pentane/dichloromethane and purified by flash chromatography on silica gel (pentane to 50% dichloromethane), yielded 648 mg (89%) of the title compound as a clear oil. 1H NMR (401 MHz, Chloroform-d) δ 7.90 (d, J = 15.8 Hz, 1H), 7.90 (dd, J = 8.0, 1.2 Hz, 1H), 7.56 (dd, J= 7.8, 1.7 Hz, 1H), 7.36 (dddd, J= 7.9, 7.4, 1.3, 0.6 Hz, 1H), 7.05 (ddd, J = 7.9, 7.3, 1.7 Hz, 1H), 6.31 (d, J= 15.8 Hz, 1H), 4.29 (q, J= 7.1 Hz, 2H), 1.35 (t, J= 7.1 Hz, 3H). 13C NMR (101 MHz, Chloroform-d) δ 166.30, 147.68, 140.02, 137.89, 131.18, 128.56, 127.37, 121.30, 101.15, 60.73, 14.33. MS (ESI-QMS) m/z: [M + H]+ calcd for Cl 1H12I02, 303.0; found, 303.0. Example 77
Sodium 7-((2-((sulfo)methoxy)phenyl)thio)-3.5-dihydro-4H-pyrrolo[3.2-d]|pyrimidin-4-onc L77
Figure imgf000052_0001
Compound L12 (100 mg, 0.2085, 1 eq.) was dissolved in dry dimethylformamide (2 ml) in a microwave reactor tube under an argon atmosphere. Sodium hydride (8 mg of 60% oil dispersion, 0.2085 mmol, 1 eq.) was added at 0 °C and the mixture was stirred at room temperature for 30 minutes. Sodium iodomethanesulfonate (102 mg, 0.4170 mmol, 2 eq.) was added and the mixture was heated in the microwave reactor at 150 °C for 15 minutes. A rich mixture of compounds was formed, which contained combinations of deprotected compounds. The mixture was dissolved in water, neutralized with 1M HC1 (aq.) and solvents were evaporated. The solid was dissolved in trifluoroacetic acid (2 ml/100 mg of the material) at room temperature and stirred for 15 minutes. Trifluoroacetic acid was evaporated, coevaporated with water (2x) and a cold solution of ammonia in ethanol (2 ml/100 mg of the material) was added. The mixture was evaporated to dryness and the solid was purified by HPLC (Cl 8, gradient HaO/MeOH). Product-containing fractions were filtered through a short pad of Dowex® 50 in Na+-cycle and solvents were evaporated. Purified product was lyophilized from water, yielded 16 mg (21%) of the title compound as a white solid. Ή NMR (401 MHz, DMSO-d6) δ 12.37 (s, 2H), 7.83 (s, 1H), 7.70 (s, 1H), 7.21 (dd, J= 8.3, 1.2 Hz, 1H), 7.04 - 6.91 (m, 1H), 6.71 (ddd, J= 7.6, 1.2 Hz, 1H), 6.47 (dd, J = 7.8, 1.6 Hz, 1H), 4.61 (s, 2H). 13C NMR (101 MHz, DMSO-d6) δ 154.13, 153.66, 146.05, 142.65, 133.88, 127.90, 125.29, 125.18, 121.42, 119.22, 113.55, 101.28, 79.90. HRMS (ESI-FTMS) m/z for the acid: [M - H]- calcd for C13H10O5N3S2, 352.0067; found, 352.0064.
Example 78
Sodium (E)-7-((2-(2-(sulfo)\ inyl)phenyl)thio)-3.5-dihydro-4H-pyrrolo[3.2-d]|pyrimidin-4-onc L78
Figure imgf000053_0001
Compound L4 (100 mg, 0.2580 mmol, 1 eq.) and copper(I) iodide (5 mg, 0.0258 mmol, 0.1 eq.) were charged into a microwave reactor tube and the tube was flushed with argon. Dry toluene (3 ml), ethyl (E )-2-(2-iodophenyl)ethene-l -sulfonate L75 (105 mg, 0.3096 mmol, 1.2 eq.), triethylamine (54 μl, 0.3870 mmol, 1.5 eq.) and 2-isobutyrylcyclohexanone (9 μl, 0.0516 mmol, 0.2 eq.) were added subsequently and the mixture was heated in the microwave reactor at 120 °C for 2 hours. The mixture was dissolved in chloroform, washed with a half-saturated aqueous solution of NaHCO3 (2x), 1M HC1 (aq.) (2x), brine (1x) and it was dried with MgSO4, filtered and solvents were evaporated. The solid was used in the next step without further characterization. MS (ESI-QMS) m/z: [M + H]+ calcd for C29H36N305S2Si, 598.2; found, 598.2. The solid was dissolved in trifluoroacetic acid (2 ml/100 mg of the material) at room temperature and stirred for 2 hours. Trifluoroacetic acid was evaporated, coevaporated with water (2x) and a cold solution of ammonia in ethanol (2 ml/100 mg of the material) was added. The mixture was evaporated to dryness and the solid dissolved in small amount of water and purified by HPLC (C18, gradient H2O /MeOH). Product-containing fractions were filtered through a short pad of Dowex® 50 in Na+-cycle and solvents were evaporated. Purified product was lyophilized from water, yielded 26 mg (27%) of the title compound as a white solid. 1H NMR (401 MHz, Deuterium Oxide) δ 8.04 (s, 1H), 7.92 (d, J= 15.5 Hz, 1H), 7.82 (s, 1H), 7.61 (dd, J= 7.6, 1.7 Hz, 1H), 7.26 (ddd, J = 7.5, 1.5 Hz, 1H), 7.21 (ddd, J= 7.6, 1.7 Hz, 1H), 7.07 (dd, J= 7.9, 1.4 Hz, 1H), 6.99 (d, J = 15.5 Hz, 1H). 13C NMR (101 MHZ, Deuterium Oxide) δ 156.31, 146.98, 144.57, 138.92, 136.77, 134.10, 133.48, 132.00, 131.63, 129.88, 128.78, 128.19, 119.74, 104.30. HRMS (ESI-FTMS) m/z for the acid: [M - H]- calcd for C15H10O3N3S, 312.0448; found, 312.0447.
Example 79
Sodium 7-((2-((carboxy)methoxy)phenyl)thio)-3.5-dihydro-4H-pyrrolo|3.2-ri|pyrimidin-4-onc L79 A vial was charged with compound L12 (100 mg, 0.2085, 1 eq.) and cesium carbonate (340 mg, 1.0 mmol, 1.5 eq.) and it was flushed with argon. Dry dimethylformamide (2 ml) and ethyl bromoacetate (204 mg, 0.8340 mmol, 4 eq.) were added and the mixture was stirred at 90 °C for 2 hours. The reaction was quenched with a half-saturated aqueous solution of NH4CI, extracted with ethyl acetate (3x), washed with brine (1x) and dried with MgSO4. Solvents were evaporated and the oil was used in the next step without further characterization. MS (ESI-QMS) m/z: [M + H]+ calcd for C29H36N305SSi, 566.2; found, 566.5. The solid was dissolved in a mixture of dioxane and water (1: 1), several drops of 1M NaOH (aq.) were added, and the mixture was stirred at room temperature for 1 hour. The mixture was neutralized with 1M HC1 (aq.), solvents were evaporated and the solid was used in the next step without further characterization. MS (ESI-QMS) m/z: [M + H]+ calcd for C27H32N305SSi, 538.2; found, 538.5. The solid was dissolved in trifluoroacetic acid (2 ml/ 100 mg of the material) at room temperature and stirred for 15 minutes. Trifluoroacetic acid was evaporated, coevaporated with water (2x) and a cold solution of ammonia in ethanol (2 ml/100 mg of the material) was added. The mixture was evaporated to dryness and the solid was purified by HPLC (C18, gradient H2O/MeOH). Product-containing fractions were filtered through a short pad of Dowex® 50 in Na+-cycle and solvents were evaporated. Purified product was lyophilized from water, yielded 24 mg (34%) of the title compound as a white solid. 1H NMR (401 MHz, DMSO-d6) δ 7.87 (s, 1H), 7.72 (s, 1H), 7.00 (ddd, J= 8.5, 7.3, 1.7 Hz, 1H), 6.71 (dd, J= 8.4, 1.2 Hz, 1H), 6.68 (ddd, J= 7.4, 0.8 Hz, 1H), 6.48 (dd, J= 7.9, 1.6 Hz, 1H), 4.37 (s, 2H) . 13C NMR (101 MHz, DMSO-d6) δ 173.21, 154.47, 154.41, 146.69, 143.17, 135.19, 126.59, 126.53, 126.24, 121.43, 118.79, 111.93, 101.98, 67.47. HRMS (ESI-FTMS) m/z for the acid: [M - H]- calcd for C14H10O4N3S, 316.0398; found, 316.0395.
Example 80
Sodium (E )-7-((2-(2-(carboxy)vinyl)phenyl)thio)-3,5-dihydro-4H-pyrrolo[3,2-ri]pyrimidin-4-one L80
Figure imgf000054_0001
Compound L4 (100 mg, 0.2580 mmol, 1 eq.) and copper(I) iodide (5 mg, 0.0258 mmol, 0.1 eq.) were charged into a microwave reactor tube and the tube was flushed with argon. Dry toluene (3 ml), (E )- 3- (2-iodophenyl)acrylate L76 (94 mg, 0.3096 mmol, 1.2 eq.), triethylamine (54 μl, 0.3870 mmol, 1.5 eq.) and 2-isobutyrylcyclohexanone (9 μl, 0.0516 mmol, 0.2 eq.) were added subsequently and the mixture was heated in the microwave reactor at 120 °C for 2 hours. The mixture was dissolved in chloroform, washed with a half-saturated aqueous solution of NaHCO3 (2x), 1M HC1 (aq.) (2x), brine (1x) and it was dried with MgSO4, filtered and solvents were evaporated. The solid was used in the next step without further characterization. MS (ESI-QMS) m/z: [M + H]+ calcd for C30H36N3O4SSi, 562.2; found, 562.2. The solid was dissolved in trifluoroacetic acid (2 ml/100 mg of the material) at room temperature and stirred for 2 hours. Trifluoroacetic acid was evaporated, coevaporated with water (2x), a cold solution of ammonia in ethanol (2 ml/ 100 mg of the material) was added and the mixture was evaporated to dryness. MS (ESI-QMS) m/z: [M + H]+ calcd for C17H16N303S, 342.1; found, 342.1. The solid was dissolved in a mixture of dioxane and water (1: 1), several drops of 10M NaOH (aq.) were added, and the mixture was stirred at room temperature for 2 hours. Solvents were evaporated and the solid dissolved in a small amount of water and purified by HPLC (Cl 8, gradient ffO/McOH). Product- containing fractions were filtered through a short pad of Dowex® 50 in Na+-cycle and solvents were evaporated. Purified product was lyophilized from water, yielded 3 mg (3.5%) of the title compound as a white solid. 1H NMR (401 MHz, DMSO-d6) δ 7.84 (s, 1H), 7.69 (s, 1H), 7.65 (d, J= 15.6 Hz, 1H), 7.52 - 7.43 (m, 1H), 7.13 - 6.91 (m, 2H), 6.79 - 6.60 (m, 1H), 6.34 (d, J= 15.6 Hz, 1H). 13C NMR (101 MHz, DMSO-d6) δ 170.88, 154.63, 146.27, 143.68, 138.38, 134.32, 133.98, 132.93, 131.72, 128.82, 126.60, 125.55, 119.60, 101.87. HRMS (ESI-FTMS) m/z for the acid: [M - H]- calcd for C15H10O3N3S, 312.0448; found, 312.0447.
Example 81
4-(Benzyloxy)-7-((3-bromo)phenylthio)-5-((2-(trimethylsilyl)ethoxy)methyl)-5i7-pyrrolo[3,2- ri|pyrimidinc L81
Figure imgf000055_0001
General procedure SI with s 3-bromothiophenol (103 μl; 1.0 mmol; 1.5 eq.) afforded 82 mg (89%) of the title compound. 1H NMR (401 MHz, Chloroform-d) δ 8.63 (s, 1H), 7.66 (s, 1H), 7.56 - 7.48 (m, 2H), 7.43 - 7.33 (m, 3H), 7.22 (dd, J= 1.8 Hz, 1H), 7.16 (ddd, J= 7.7, 1.9, 1.1 Hz, 1H), 7.05 (ddd, J= 7.9, 1.8, 1.2 Hz, 1H), 6.99 (dd, J= 7.8 Hz, 1H), 5.66 (s, 2H), 5.64 (s, 2H), 3.48 (t, J= 7.9 Hz, 2H), 0.84 (t, J= 8.1 Hz, 2H), -0.09 (s, 9H). 13C NMR (101 MHz, Chloroform-d) δ 156.22, 151.63, 151.36, 140.55, 138.64, 135.93, 130.08, 128.93, 128.69, 128.44, 128.40, 128.29, 125.09, 122.87, 116.34, 103.58, 77.95, 68.47, 66.36, 17.68, -1.40. MS (ESI-QMS) m/z: [M + H]+ calcd for C25H29BrN302SSi, 542.1; found, 542.2.
Example 82 Sodium 7 -((3 -(phosphonato)phenyl)thio)-3 ,5 -dihydro-4H-pyrrolo [3,2-ri]pyrimidin-4-one L82
Figure imgf000056_0001
Pd2(dba)3 (84 mg, 0.0922 mmol, 0.1 eq.) and 4,5-bis(diphenylphosphino)-9,9-dimethylxanthene (107 mg, 0.1843 mmol, 0.2 eq.) were dissolved in dry dioxane (5 ml) under an argon atmosphere, and then triethylamine (192 μl, 1.4 mmol, 1.5 eq.) was added. The mixture was stirred 15 minutes at room temperature and a solution of compound L81 (500 mg, 0.9215 mmol, 1 eq.) with diethyl phosphite (130 μl, 1.0 mmol, 1.1 eq.) in dry dioxane (2 ml) was added. The mixture was stirred 24 hours at 90 °C, then, it was cooled to 0 °C, quenched with 1M HC1 (aq.), extracted with ethyl acetate (3x), washed with brine (1x), dried with magnesium sulfate and evaporated. Purification by flash chromatography on silica gel (cyclohexane to 15% ethyl acetate modified with 10% methanol (v/vi)) afforded 514 mg (93%) of the coupling product as a white-off oil, which was used in the next step without further characterization. MS (ESI-QMS) m/z: [M + H]+ calcd for C29H39N305PSSi, 600.2; found, 600.2. The oil was dissolved in trifluoroacetic acid (2 ml/100 mg of the material) at room temperature and stirred for 15 minutes. Trifluoroacetic acid was evaporated, coevaporated with water (2x) and a cold solution of ammonia in ethanol (2 ml/100 mg of the material) was added. The mixture was evaporated to dryness and the solid was adsorbed on silica gel in a mixture of cyclohexane/acetone. Purification by flash chromatography on silica gel (chloroform to 10% methanol) afforded 334 mg (97%) of the diethyl phosphonate, which was used in the next step without further characterization. MS (ESI-QMS) m/z: [M + H]+ calcd for C16H19N304PS, 380.1; found, 380.2. A flask charged with the diethyl phosphonate was sealed with septum, flushed with argon, and then dry pyridine (10 ml/1 mmol of the starting compound) was added, followed by trimethylsilyl bromide (1 ml/1 mmol of the starting compound). The mixture was stirred at room temperature overnight and then it was evaporated to dryness. The residue was dissolved in a 2M aq. solution of triethylammonium bicarbonate and a small amount of methanol, and it was evaporated to dryness. The solid residue was dissolved in a small amount of water (solubility can be enhanced by addition of several drops of aqueous ammonia) and purified by HPLC (C18, gradient H2/OMeOH). Product-containing fractions were filtered through a short pad of Dowex® 50 in Na+-cycle and solvents were evaporated. Purified product was lyophilized from water, yielded 199 mg (70%) of the title compound as a white solid. 1H NMR (401 MHz, Deuterium Oxide) δ 8.07 (s, 1H), 7.73 (s, 1H), 7.71 (ddd, J= 11.9, 1.4 Hz, 1H), 7.48 (dddd, J= 11.2, 7.5, 1.2 Hz, 1H), 7.21 (ddd, J= 7.6, 3.0 Hz, 1H), 6.94 (dddd, J = 7.9, 2.1, 1.1 Hz, 1H). 13C NMR (101 MHz, Deuterium Oxide) δ 165.12, 153.38, 148.76, 143.49 (d, J = 165.0 Hz), 139.12 (d, J= 14.0 Hz), 136.15, 129.85 (d, J= 13.0 Hz), 129.46 (d, J= 9.7 Hz), 128.91 (d, J = 8.7 Hz), 127.34 (d, J = 2.4 Hz), 122.22, 101.04. 31P NMR (162 MHz, Deuterium Oxide) δ 12.86. HRMS (ESI-FTMS) m/z for the acid: [M - H]- calcd for C12H904N3PS, 322.0057; found, 322.0053.
Example 83
2-Iodobenzyl bromide L83
Figure imgf000057_0001
2-iodobenzyl alcohol (3 g, 12.8 mmol, 1 eq.) was dissolved in dry tetrahydrofuran (30 ml) under an argon atmosphere. Phosphorus(III) bromide (1.8 ml, 19.2 mmol, 1.5 eq.) was added and the mixture was stirred at room temperature overnight. The reaction was quenched with silica gel and solvents were evaporated. Purification by flash chromatography on silica gel (cyclohexane to 20% ethyl acetate modified with 10% methanol (v/vi)) afforded 3.7 g (97%) of 2-iodobenzyl bromide as a white solid. 1H NMR (401 MHz, Chloroform-d) δ 7.86 (dd, J= 7.9, 1.2 Hz, 1H), 7.48 (dd, J= 7.7, 1.7 Hz, 1H), 7.34 (ddd, J = 7.5, 1.3 Hz, 1H), 6.98 (ddd, J = 7.8, 1.7 Hz, 1H), 4.60 (s, 2H). 13C NMR (101 MHz, Chloroform -d) δ 140.33, 140.22, 130.63, 130.22, 129.02, 100.19, 38.91. MS (ESI-QMS) m/z: [M + H - HBr + ACN + H20]+ calcd for C9H1 UNO, 276.0; found, 275.9.
Example 84
Diisopropyl (2-iodobenzyl)phosphonate L84
Figure imgf000057_0002
2-Iodobenzyl bromide (3 g, 10.1 mmol, 1 eq.) was charged into a microwave reactor tube and the tube was flushed with an argon. Dry toluene (20ml) and triisopropyl phosphite (3 ml, 11.1 mmol, 1.1 eq.) were added subsequently and the mixture was heated in the microwave reactor at 160 °C for 1 hour. The mixture was evaporated and purified by flash chromatography on silica gel (cyclohexane to 40% ethyl acetate modified with 10% methanol (v/vi)) afforded 3.4 g (88%) of diisopropyl (2- iodobenzyl)phosphonate as a clear oil. 1H NMR (401 MHz, Chloroform -d) δ 7.83 (dd, J= 7.9, 1.2 Hz, 1H), 7.50 (ddt, J= 7.7, 2.8, 1.7 Hz, 1H), 7.29 (ddd, J= 7.6, 1.1 Hz, 1H), 6.91 (ddd, J= 7.9, 2.0 Hz, 1H), 4.82 - 4.33 (m, 2H), 3.36 (d, J= 22.0 Hz, 2H), 1.29 (d, J= 6.2 Hz, 6H), 1.18 (d, J= 6.2 Hz, 6H). 13C NMR (101 MHz, Chloroform-d) δ 139.75 (d, J= 2.9 Hz), 135.95 (d, J= 8.2 Hz), 130.79 (d, J= 5.1 Hz), 128.57 (d, J= 3.6 Hz), 128.37 (d, J= 3.1 Hz), 101.87 (d, J= 9.6 Hz), 70.97 (d, J= 6.7 Hz), 39.47 (d, J = 140.0 Hz), 24.83 - 23.34 (m). 31P NMR (162 MHz, Chloroform-d) δ 26.23. MS (ESI-QMS) m/z: [M + H]+ calcd for C13H2 1I03P, 383.0; found, 383.0.
Example 85
Sodium 7-((2-((phosphonato)methyl)phenyl)thio)-3.5-dihydro-4H-pyrrolo[3.2-d]|pyrimidin-4-onc L85 Compound L4 (500 mg, 1.3 mmol, 1 eq.), cesium carbonate (509 mg, 1.6 mmol, 1.2 eq.), copper(I) iodide (25 mg, 0.1290 mmol, 0.1 eq.) and 1,10-phenantroline (46 mg, 0.2580 mmol, 0.2 eq.) were charged into a microwave reactor tube and the tube was flushed with argon. Dry toluene (5 ml) and diisopropyl (2-iodobenzyl)phosphonate L84 (596 mg, 1.6 mmol, 1.2 eq.) were added subsequently and the mixture was heated in the microwave reactor at 120 °C for 2 hours. The mixture was dissolved in chloroform, washed with a half-saturated aqueous solution of NaHCO3 (2x), 1M HC1 (aq.) (2x), brine (1x) and it was dried with MgSO4, filtered and solvents were evaporated. The solid was adsorbed on silica gel in a mixture of cyclohexane/acetone and purified by flash chromatography on silica gel (cyclohexane to 20% ethyl acetate modified with 10% methanol (v/vi)), yielded 754 mg (91%) of the coupling product as a white-off oil. The product was used in the next step without further characterization. MS (ESI-QMS) m/z: [M + H]+ calcd for C32H45N305PSSi, 642.3; found, 642.2. The oil was dissolved in trifluoroacetic acid (2 ml/ 100 mg of the material) at room temperature and stirred for 15 minutes. Trifluoroacetic acid was evaporated, coevaporated with water (2x) and a cold solution of ammonia in ethanol (2 ml/100 mg of the material) was added. The mixture was evaporated to dryness and the solid was adsorbed on silica gel in a mixture of cyclohexane/acetone. Purification by flash chromatography on silica gel (chloroform to 10% methanol) afforded 208 mg (42%) of the diisopropyl phosphonate, which was used in the next step without further characterization. MS (ESI-QMS) m/z: [M + H]+ calcd for C19H25N304PS 422,1; found, 422.3. A flask charged with 100 mg of the diisopropyl phosphonate was sealed with septum, flushed with argon, then dry pyridine (10 ml/ 1 mmol of the starting compound) was added followed by trimethylsilyl bromide (1 ml/1 mmol of the starting compound). The mixture was stirred at room temperature overnight and then it was evaporated to dryness. The residue was dissolved in 2M aq. solution of triethylammonium bicarbonate and a small amount of methanol, then it was evaporated to dryness. The solid residue was dissolved in a small amount of water (solubility can be enhanced by addition of several drops of aqueous ammonia) and purified by HPLC (Cl 8, gradient H2O/MeOEl). Product-containing fractions were filtered through a short pad of Dowex® 50 in Na+-cycle and solvents were evaporated. Purified product was lyophilized from water, yielded 70 mg (78%) of the title compound as a white solid. 1H NMR (400 MHz, Deuterium Oxide) δ 8.01 (s, 1H), 7.76 (s, 1H), 7.37 (d, J= 7.7 Hz, 1H), 7.08 (dd, J= 7.6 Hz, 1H), 6.87 (dd, J= 7.7 Hz, 1H), 6.72 (d, J= 7.9 Hz, 1H), 3.36 (d, J= 20.8 Hz, 1H). 13C NMR (101 MHz, Deuterium Oxide) δ 156.27, 147.02, 144.37, 138.35 (d, J = 7.2 Hz), 136.79, 134.47 (d, J = 8.3 Hz), 132.15 (d, J = 5.1 Hz), 128.38, 127.89, 126.96, 119.57, 104.97, 35.03 (d, J= 128.6 Hz). 31P NMR (162 MHz, Deuterium Oxide) δ 17.54. HRMS (ESI-FTMS) m/z for the acid: [M - H]- calcd for C13H1104N3PS, 336.0213; found, 336.0209. Example 86
Sodium 7-((2-(3-phosphonato-2-oxapropyl)phenyl)thio)-3.5-dihydro-4H-pyrrolo[3.2-d]|pyrimidin-4- one L86
Figure imgf000059_0001
Compound Lll (100 mg, 0.2026 mmol, 1 eq.) was dissolved in dry tetrahydrofuran (50 ml) under an argon atmosphere. The solution was cooled to 0 °C, sodium hydride (12 mg of 60% oil dispersion, 0.3039 mmol, 1.5 eq.) was added and the mixture was stirred at 0 °C for 15 minutes. Then, TfOCH2P(O)(0/-Pr)2 (133 mg, 0.4052 mmol, 2 eq.) was added and the mixture was stirred at 0 °C for additional 15 minutes. The reaction was quenched with a half-saturated aqueous solution of NH4CI and extracted with ethyl acetate (3x). The organic phase was washed with brine, dried with MgSO4, filtered and solvents were evaporated. The solid was adsorbed on silica gel in a mixture of cyclohexane/acetone and purified by flash chromatography on silica gel (cyclohexane to 20% ethyl acetate modified with 10% methanol (v/vi)) afforded 82 mg (60%) of alkylated product, which was used in the next step without further characterization. MS (ESI-QMS) m/z: [M + H]+ calcd for C33H47N306PSSi, 672.3; found, 672.4. The oil was dissolved in trifluoroacetic acid (2 ml/100 mg of the material) at room temperature and stirred for 15 minutes. Trifluoroacetic acid was evaporated, coevaporated with water (2x) and a cold solution of ammonia in ethanol (2 ml/100 mg of the material) was added. The mixture was evaporated to dryness to afford the crude diisopropyl phosphonate, which was used in the next step without further characterization. MS (ESI-QMS) m/z: [M + H]+ calcd for C20H27N3O5PS, 452.1; found, 452.2. A flask charged with the diisopropyl phosphonate was sealed with septum, flushed with argon, and then dry pyridine (10 ml/1 mmol of the starting compound) was added followed by trimethylsilyl bromide (1 ml/1 mmol of the starting compound). The mixture was stirred at room temperature overnight and then it was evaporated to dryness. The residue was dissolved in a 2M aq. solution of triethylammonium bicarbonate and a small amount of methanol, and it was evaporated to dryness. The solid residue was dissolved in a small amount of water (solubility can be enhanced by addition of several drops of aqueous ammonia) and purified by HPLC (C18, gradient EhO/MeOEl). Product-containing fractions were filtered through a short pad of Dowex® 50 in Na+-cycle and solvents were evaporated. Purified product was lyophilized from water, yielded 6 mg (12%) ofthe title compound as a white solid. 1H NMR (500 MHz, Methanol-d4) δ 8.00 (s, 1H), 7.76 (s, 1H), 7.46 (dd, J= 7.6, 1.5 Hz, 1H), 7.16 (ddd, J= 7.5, 1.2 Hz, 1H), 7.07 (ddd, J= 7.7, 1.5 Hz, 1H), 6.78 (dd, J= 8.0, 1.0 Hz, 1H), 4.85 (s, 2H), 3.66 (d, J = 8.6 Hz, 2H). 13C NMR (126 MHz, Methanol-riQ d 156.19, 147.02, 144.04, 138.53, 136.54, 135.72, 130.54, 129.56, 127.54, 126.53, 119.48, 103.99, 73.42 (d, J= 11.1 Hz), 68.90 (d, J= 152.7 Hz).31P NMR (162 MHz, Deuterium Oxide) δ 16.23. HRMS (ESI-FTMS) m/z for the acid: [M - H]- calcd for C14H1305N3PS, 366.0319; found, 366.0317.
Example 87
Sodium 7-((2-(carboxy)phenyl)thio)-3.5-dihydro-4H-pyrrolo|3.2-ri|pyrimidin-4-onc L87
Figure imgf000060_0001
2-Mercaptobenzoic acid (77 mg, 0.4985 mmol, 1.2 eq.) was dissolved in dry dimethylformamide (2 ml) in a microwave reactor tube and the tube was flushed with argon. Sodium hydride (40 mg of 60% oil dispersion, 0.9970 mmol, 2.4 eq.) was added at 0 °C and the mixture was stirred at room temperature for 30 minutes. Compound L2 (200 mg, 0.4154 mmol, 1 eq.), copper(I) iodide (8 mg, 0.0420 mmol, 0.1 eq.) and 1,10-phenantroline (15 mg, 0.0840 mmol, 0.2 eq.) were added and the mixture was heated in the microwave reactor at 120 °C for 1 hour. The mixture was dissolved in 1M HC1 (aq.), extracted with ethyl acetate (3 x), washed with brine (1 x), dried with MgSO4, filtered, and solvents were evaporated. The solid was adsorbed on C18 silica gel in methanol and purified by reverse phase flash chromatography on Cl 8 silica gel (water to methanol), yielded 37 mg (18%) of the coupling product, which was used in the next step without further characterization. MS (ESI-QMS) m/z: [M + H]+ calcd for C26H30N3O4SSi, 508.2; found, 508.3. The coupling product was dissolved in trifluoroacetic acid ( 1 ml) at room temperature and stirred for 15 minutes. Trifluoroacetic acid was evaporated, coevaporated with water (2x) and a cold solution of ammonia in ethanol (2 ml/100 mg of the material) was added. The mixture was evaporated to dryness and the solid was purified by HPLC (Cl 8, gradient ITO/McOH). Product-containing fractions were filtered through a short pad of Dowex® 50 in Na+-cycle and solvents were evaporated. Purified product was lyophilized from water, yielded 8 mg (36%) of the title compound as a white solid. 1H NMR (500 MHz, DMSO-d6) δ 7.71 - 7.56 (m, 2H), 7.10 (s, 1H), 6.89 - 6.74 (m, 2H), 6.66 (d, J= 7.2 Hz, 1H). 13CNMR(126 MHz, DMSO-d6) δ 170.53, 162.93, 147.19, 141.76, 138.09, 128.79, 126.67, 125.05, 124.13, 121.48, 99.89. HRMS (ESI-FTMS) m/z for the acid: [M - H]- calcd for C13H803N3S, 286.0292; found, 286.0290.
Example 88
Sodium 2-iodobenzenesulfonate hydrate L88
Figure imgf000060_0002
The compound was synthesized according to the published procedure. [Tanaka, Y. et al., J. Fluor. Chem. 2012, 137, 98-103] A solution of NaNO2 (4.2 g, 60.6 mmol, 1.05 eq.) and water (40 ml) was added dropwise to a stirred mixture of 2-aminobenzenesulfonic acid (10 g, 57.8 mmol, 1 eq.), ice (40 g) and cone. HC1 (20 ml) at 0 °C. The mixture was stirred at 0 °C for 1 hour then at room temperature for 1 hour and then at 50 °C overnight. The mixture was filtered, evaporated and recrystallized from water. Crystals were collected, washed with cold ethanol (1x), cold diethyl ether (1x) and dried, yielded 9.8 g (52%) of the title compound as white crystals.
Example 89
Sodium 7 -((2-(sulfo)phenyl)thio)-3 ,5 -dihydro-4H-pyrrolo [3 ,2-ri]pyrimidin-4-one L89
Figure imgf000061_0001
Compound L4 (200 mg, 0.5160 mmol, 1 eq.), 2-iodobenzenesulfonate hydrate L88 (334 mg, 1.0 mmol, 2 eq.), copper(I) iodide (5 mg, 0.0516 mmol, 0.1 eq.) and 1,10-phenantroline (9 mg, 0.1532 mmol, 0.2 eq.) were charged into a microwave reactor tube and the tube was flushed with argon. Dry dimethylformamide (2 ml) and triethylamine (72 μl, 1.0 mmol, 2 eq.) were added subsequently and the mixture was heated in the microwave reactor at 120 °C for 1 hour. The mixture was diluted with water and pH was adjusted to 1 with 1M HC1 (aq.). pH was adjusted to 7 with triethylamine, mixture was extracted with ethyl acetate, washed with brine (1 x), it was dried with MgSO4, filtered, and solvents were evaporated. The solid was adsorbed on C18 silica gel in methanol and purified by reverse phase flash chromatography on Cl 8 silica gel (water to methanol), yielded 220 mg (66%) of the coupling product, which was used in the next step without further characterization. MS (ESI-QMS) m/z: [M + H]+ calcd for C25H30N3O5S2Si, 544.1; found, 544.3. The coupling product (110 mg, 0.2023 mmol, 1 eq.) was dissolved in trifluoroacetic acid (2 ml/100 mg of the material) at room temperature and stirred for 15 minutes. Trifluoroacetic acid was evaporated, coevaporated with water (2x) and a cold solution of ammonia in ethanol (2 ml/100 mg of the material) was added. The mixture was evaporated to dryness and the solid was purified by HPLC (Cl 8, gradient H2O/McOH). Product-containing fractions were filtered through a short pad of Dowex® 50 in Na+-cycle and solvents were evaporated. Purified product was lyophilized from water, yielded 54 mg (84%) of the title compound as a white solid. 1H NMR (401 MHz, DMSO-d6) δ 12.31 (s, 2H), 7.81 (s, 1H), 7.71 (dd, J= 7.4, 1.9 Hz, 1H), 7.60 (s, 1H), 7.08 - 6.86 (m, 2H), 6.56 (dd, J = 7.7, 1.6 Hz, 1H). 13C NMR (101 MHz, DMSO-d6) δ 154.03, 146.19, 143.43, 142.69, 137.77, 133.74, 128.49, 127.34, 125.07, 122.91, 119.40, 103.28. HRMS (ESI-FTMS) m/z for the acid: [M - H]- calcd for C12H804N3S2, 321.9962; found, 321.9959.
Example 90
Sodium bromomethanesulfonate L90 The compound was synthesized according to the published procedure. [Gao, F. et al., Bioorganic Med. Chem. Lett. 2008, 18 (20), 5518-5522]
Dibromomethane (24.5 ml, 350 mmol, 1 eq.), sodium sulfite (44 g, 350 mmol, 1 eq.) and tetrabutylammonium hydrogensulfate (2 g, 6 mmol, 0.02 eq.) were dissolved in a mixture of ethanol and water (200 ml, 1:2). The mixture was vigorously stirred overnight at reflux and then it was evaporated to dryness. The solid was suspended in a mixture of ethanol and water (240 ml, 5: 1) and the mixture was stirred 30 minutes at 45 °C. Then, the mixture was filtered and cooled to -20 °C for 7 days and after that, white crystals were collected and dried, yielded 48 g (70%) of the title compound.
Example 91
Bromomethanesulfonyl chloride L91
Figure imgf000062_0001
The compound was synthesized according to the published procedure. [Gao, F. et al., Bioorganic Med. Chem. Lett. 2008, 18 (20), 5518-5522]
Sodium bromomethanesulfonate (30 g, 152 mmol, 1 eq.) was vigorously stirred in a flask and phosphorus pentachloride (32 g, 152 mmol, 1 eq.) was added portionwise. The mixture was vigorously stirred without solvent 30 minutes at 130 °C and then 30 minutes at 70 °C. The solid mixture turned to red slurry within 5 minutes of the reaction. The reaction was cooled to room temperature, ice with water were added to the mixture, and the mixture was stirred for additional 30 minutes. The mixture was extracted with dichloromethane (3x) and the organic phase was washed with a cooled half-saturated aqueous solution of NaHCO3 (2x), water (1x), brine (1x) and it was dried with MgSO4. The desiccant was filtered and solvents were evaporated, yielded 15 g (51%) of the title compound as a yellow oil. The compound was used without further purification into next steps.
Example 92
Bromomethanesulfonamide L92
Figure imgf000062_0002
Bromomethanesulfonyl chloride (20 g, 103 mmol, 1 eq.) was dissolved in dry diethyl ether (200 ml) under an argon atmosphere. The solution was cooled to 0 °C and gaseous ammonia was slowly bubbled into the mixture for 15 minutes. The mixture was stirred for additional 30 minutes, and then the precipitate of ammonium chloride and ammonium bromomethanesulfonate was filtered. Evaporation of solvents afforded 12 g (67%) of the title compound as a white-off solid. 1H NMR (401 MHz, DMSO- d6) δ 7.32 (s, 2H), 4.67 (s, 2H). 13C NMR (101 MHz, DMSO-d6) δ 44.63. MS (ESI-QMS) m/z: [M + H - HBr + H2O + CH3CN]+ calcd for C3H9N203S, 153.0; found, 153.0.
Example 93
1 -Bromo- N,N'-bis((2-(trimethylsilyl)cthoxy)methyl)mcthancsulfonamidc L93
Figure imgf000063_0002
Bromomethanesulfonamide (2 g, 11.5 mmol, 1 eq.) was dissolved in dry tetrahydrofuran (20 ml) under an argon atmosphere and the solution was cooled to 0 °C. Sodium hydride (460 mg of 60% oil dispersion, 11.5 mmol, 1 eq.) was added and the mixture was stirred at 0 °C for 15 minutes. 2- (trimethylsilyl)ethoxymethyl chloride (2 ml, 11.5 mmol, 1 eq.) was added and the mixture was stirred at 0 °C for 15 minutes. The addition of sodium hydride and 2-(trimethylsilyl)ethoxymethyl chloride was repeated once more. The reaction was quenched with a half-saturated aqueous solution of NH4CI, extracted with ethyl acetate (3x), and the organic phase was washed with brine (1x) and it was dried with MgSO/i. The whole suspension with desiccant was half-diluted with chloroform, and it was filtered through a short pad of neutral alumina and the pad was washed with a mixture of chloroform and ethyl acetate (2: 1, v/vi). Solvents were evaporated, yielded 4.6 g (92%) of the title compound as a clear oil. 1H NMR (401 MHz, Chloroform-d) δ 4.82 (s, 4H), 4.52 (s, 2H), 3.64 - 3.51 (m, 4H), 0.98 - 0.88 (m, 4H), 0.02 (s, 18H). MS (ESI-QMS) m/z: [M + Na]+ calcd for C13H32BrNNa04SSi2, 456.01; found, 456.1.
Example 94
7-((2-((Sulfamoyl)methoxy)phenyl)thio)-3 ,5 -dihydro-4H-pyrrolo [3 ,2-d]pyrimidin-4-one L94
Figure imgf000063_0001
A flask was charged with compound L12 (200 mg, 0.4170 mmol, 1 eq.), compound L93 (724 mg, 1.7 mmol, 4 eq.), potassium carbonate (130 mg, 1.7 mmol, 4 eq.) and it was flushed with an argon atmosphere. Dry dimethylformamide (10 ml) was added and the mixture was stirred at 110 °C overnight. The reaction was quenched at ca. 50% conversion with a half-saturated aqueous solution of NH4CI, extracted with ethyl acetate (3x), and the organic phase was washed with brine (1x) and it was dried with MgSO4. The solid was adsorbed on C18 silica gel and purification by C18 reverse phase flash chromatography (water to methanol) afforded 120 mg (35%) of the title compound. The compound was used in the next step without characterization. MS (ESI-QMS) m/z: [M + H]+ calcd for C38H61N407S2Si3, 833.3; found, 833.5. The residue (100 mg, 0.1200 mmol, 1 eq.) was dissolved in trifluoroacetic acid (2 ml/100 mg of the material) and the mixture was stirred at room temperature for 15 minutes. Trifluoroacetic acid was evaporated, coevaporated with water (2x) and a cold solution of ammonia in ethanol (2 ml/100 mg of the material) was added. The mixture was evaporated to dryness and the solid was dissolved in a small amount of dimethylformamide. The sample was subjected to purification by C18 reverse phase flash chromatography (water to methanol) in a liquid injection mode. Purified product was lyophilized from dioxane, yielded 11 mg (22%) of title compound as a white solid. 1H NMR (500 MHz, DMSO-d6) δ 12.67 (s, 1H), 12.18 (s, 1H), 7.83 (s, 1H), 7.72 (s, 1H), 7.45 - 7.24 (m, 2H), 7.22 (dd, J= 8.2, 1.2 Hz, 1H), 7.05 (ddd, J= 7.8, 1.6 Hz, 1H), 6.83 (ddd, J= 7.7, 1.2 Hz, 1H), 6.53 (dd, J= 7.9, 1.5 Hz, 1H), 5.17 (s, 2H). 13C NMR (126 MHz, DMSO-d6) δ 153.70, 153.02, 145.97, 142.85, 134.04, 128.72, 125.94, 125.49, 122.98, 119.38, 114.27, 100.74, 81.65. HRMS (ESI-FTMS) m/z: [M - H]- calcd for C13H11N404S2, 351.0227; found, 351.0227.
Example 95
(E )-7-((2-(2-(bis(( S)-1-(ethoxycarbonyl)-2-(phenyl)ethyl)amino)phosphoryl)vinyl)phenyl)thio)-3,5- dihydro-4H-pyrrolo[3.2-d]|pyrimidin-4-onc L95
Figure imgf000064_0001
Compound L4 (500 mg, 1.29 mmol, 1 eq.), copper(I) iodide (25 mg, 0.1290 mmol, 0.1 eq.) and cesium carbonate (505 mg, 1.55 mmol, 1.2 eq.) were charged into a microwave reactor tube and the tube was flushed with argon. Dry toluene (5 ml), compound L13 (559 mg, 1.33 mmol, 1.1 eq.) and 2-isobutyrylcyclohexanone (43 μl, 0.2580 mmol, 0.2 eq.) were added subsequently and the mixture was heated in the microwave reactor at 120 °C for 2 hours. The mixture was dissolved in chloroform, washed with a half-saturated aqueous solution of NaHCO3 (2x), 1M HC1 (aq.) (2x), brine (1x), dried with MgSO4, filtered and solvents were evaporated. The solid was adsorbed on silica gel in a mixture of cyclohexane/acetone and purified by flash chromatography on silica gel (cyclohexane to 15% ethyl acetate modified with 10% methanol (v/vi)), yielded 800 mg (99%) of the product as an oil. The compound was used in the next step without further characterization. MS (ESI-QMS) m/z: [M + H]+ calcd for C31H41N305PSSi, 626.2; found, 626.2. The compound was dissolved in trifluoroacetic acid (2 ml/ 100 mg of the material) at room temperature and stirred for 15 minutes. Trifluoroacetic acid was evaporated, coevaporated with water (2x) and a cold solution of ammonia in ethanol (2 ml/ 100 mg of the material) was added. The mixture was evaporated, the solid was adsorbed on silica gel in a mixture of cyclohexane/acetone and purified by flash chromatography on silica gel (chloroform to 15% methanol), yielded 250 mg (48%) of the diethyl phosphonate as a white solid. MS (ESI-QMS) m/z: [M + H]+ calcd for C 18H2 1N304PS, 406.1 ; found, 406.2. A flask was charged with the diethyl phosphonate (100 mg, 0.2467 mmol, 1 eq.), it was sealed with septum, flushed with argon, and then dry pyridine (10 ml/1 mmol of the starting compound) was added, followed by trimethylsilyl bromide (1 ml/1 mmol of the starting compound). The mixture was stirred at room temperature overnight and then it was evaporated to dryness under an argon atmosphere. L-phenylalanine ethyl ester hydrochloride (227 mg, 0.9868 mmol, 4 eq.) was added and the flask was sealed with septum, flushed with argon. Dry pyridine (10 ml/1 mmol of the starting compound) and triethylamine (206 μl, 1.48 mmol, 6 eq.) were added to the mixture. The mixture was stirred 15 minutes at 70 °C and a solution of triphenylphosphine (388 mg, 1.48 mmol, 6 eq.) and Aldrithiol™-2 (326 mg, 1.48 mmol, 6 eq.) in dry pyridine (10 ml/1 mmol of the starting compound) was added. The reaction was quenched by evaporation of solvents and the residue was coevaporated with toluene. The solid residue was adsorbed on silica gel in a mixture of cyclohexane/acetone and purified by flash chromatography on silica gel (chloroform to 20% methanol), followed by purification on HPLC (Cl 8, gradient H2O/MeOH as eluent). Purified product was lyophilized from dioxane, yielded 100 mg (58%) of the title compound as a white solid. 1H NMR (401 MHz, DMSO-d6) δ 12.73 (d, J= 3.1 Hz, 1H), 12.12 (d, J= 3.5 Hz, 1H), 7.85 (d, J= 3.2 Hz, 1H), 7.74 (d, J= 3.0 Hz, 1H), 7.63 (dd, J= 21.0, 17.1 Hz, 1H), 7.38 - 7.31 (m, 1H), 7.29 - 7.04 (m, 12H), 6.87 - 6.72 (m, 1H), 6.02 (dd, J= 19.0, 17.1 Hz, 1H), 4.70 (t, J= 11.1 Hz, 1H), 4.39 (t, J= 10.8 Hz, 1H), 4.09 - 3.93 (m, 4H), 3.95 - 3.77 (m, 2H), 3.05 - 2.76 (m, 4H), 1.16 - 0.95 (m, 6H). 13C NMR (101 MHz, DMSO-d6) δ 176.02 - 171.21 (m), 153.60, 145.68, 142.90, 139.78 (d, J= 6.1 Hz), 138.51, 137.10 (d, J = 9.2 Hz), 133.83, 132.67 (d, J= 20.9 Hz), 129.57, 129.38 (d, J= 3.6 Hz), 128.11, 126.70, 126.48 (d, J = 2.0 Hz), 126.39, 125.14, 123.11 (d, J= 153.3 Hz), 119.28, 101.48, 61.98 - 58.15 (m), 56.65 - 50.71 (m), 39.94, 15.85 - 10.37 (m). 31P NMR (162 MHz, DMSO-d6) δ 22.01. HRMS (ESI-FTMS) m/z: [M + H]+ calcd for C36H39N506PS, 700.2353; found, 700.2350.
Example 96
7-((2-(((Diisopropoxy)phosphoryl)methoxy)phenyl)thio)-3 ,5 -dihydro-4H-pyrrolo [3.2-ri|pyrimidin-4- one L96
Figure imgf000065_0001
A flask was charged with compound L4 (20.0 g, 41.5 mmol, 1.0 eq.), copper(I) iodide (7.79 g, 4.15 mmol, 0.1 eq.) and 1,10-phenantroline (1.50 g, 8.31 mmol, 0.2 eq.). The flask was sealed with septum, flushed with argon, and dry toluene (200 ml), triethylamine (8.67 ml, 62.3 mmol, 1.5 eq.) and 2- mercaptophenol (5.16 ml, 49.9 mmol, 1.2 eq.) were added subsequently. The mixture was heated at 120 °C for 2 hours. Dark brown mixture was diluted with chloroform, washed with a half-saturated aqueous NaHCO3 solution (2x), 1M HC1 (aq.) (2x), brine (1x) and it was dried with MgSCri. Solvents were evaporated and the crude solid was dissolved in toluene and evaporated (2x). The solid was dissolved in dry dimethylformamide (100 ml) and the flask was flushed with argon. The mixture was warmed to 60 °C and TsOCH2P(O)(0/-Pr)2 (21.8 g; 62.3 mmol; 1.5 eq.) and potassium tert-butoxide (7.00 g; 62.3 mmol; 1.5 eq.) were added. The mixture was stirred at 60 °C for 1 hour and additional portion of TsOCH2P(O)(0/-Pr)2 (21.8 g; 62.3 mmol; 1.5 eq.) and potassium tert-butoxide (7.00 g; 62.3 mmol; 1.5 eq.) was added. The addition was repeated after another 1 hour. The mixture was cooled to 0 °C, neutralized with 1M HC1 (aq.), extracted with chloroform (3x), washed with water (2x), brine (2x), drided with MgSO4, filtered and evaporated. The mixture was dissolved in toluene and evaporated (2x), and then it was dissolved in trifluoroacetic acid (200 ml) and the solution was stirred at room temperature for 15 minutes. The trifluoroacetic acid was evaporated and codistilled with toluene (2x) and with a saturated solution of ammonia in ethanol (1 x 100 ml). The mixture was adsorbed on silica gel in a mixture cyclohexane/acetone and it was purified by flash chromatography on silica gel (chloroform to 10% methanol), yielded 12.3 g (68%) of white crystals. 1H NMR (401 MHz, DMSO-d6) δ 12.67 (d, J= 3.2 Hz, 1H), 12.09 (d, J= 3.5 Hz, 1H), 7.82 (d, J= 3.6 Hz, 1H), 7.69 (d, J= 3.2 Hz, 1H), 7.09 (dd, J =
8.2, 1.3 Hz, 1H), 7.03 (ddd, J= 8.2, 7.2, 1.6 Hz, 1H), 6.76 (td, J= 7.5, 1.3 Hz, 1H), 6.48 (dd, J= 7.8, 1.6 Hz, 1H), 4.86 - 4.72 (m, 2H), 4.43 (d, J= 9.9 Hz, 2H), 1.40 - 1.27 (m, 12H). 13C NMR (101 MHz, DMSO-d6) δ 153.77 (d, J= 14.1 Hz), 153.64, 145.97, 142.70, 133.90, 127.86, 125.46, 125.35, 121.79, 119.32, 111.73, 100.80, 70.94 (d, J= 6.4 Hz), 62.49 (d, J= 167.0 Hz), 24.73 - 21.20 (m). 31P NMR (162 MHz, DMSO-d6) δ 19.70. MS (ESI-QMS) m/z: [M + H]+ calcd for C19H25N305PS, 438.1; found,
438.2.
Example 97
7-((2-((Bis(((S)-1-(ethoxy carbonyl)-2-(phenyl)ethyl)amino)phosphoryl)methoxy)phenyl)thio)-3, 5- dihydro-4H-pyrrolo[3.2-d]|pyrimidin-4-one L97
Figure imgf000066_0001
A flask was charged with compound L96 (100 mg, 0.2286 mmol, 1 eq.), it was sealed with septum, flushed with argon, and then dry pyridine (10 ml/1 mmol of the starting compound) was added, followed by trimethylsilyl bromide (1 ml/1 mmol of the starting compound). The mixture was stirred at room temperature overnight and then it was evaporated to dryness under an argon atmosphere. L-phenylalanine ethyl ester hydrochloride (210 mg, 0.9144 mmol, 4 eq.) was added and the flask was sealed with septum, flushed with argon. Dry pyridine (10 ml/1 mmol of the starting compound) and triethylamine (191 μl, 1.37 mmol, 6 eq.) were added to the mixture. The mixture was stirred 15 minutes at 70 °C and a solution of triphenylphosphine (359 mg, 1.37 mmol, 6 eq.) and Aldrithiol™-2 Aldrithiol™-2 (302 mg, 1.37 mmol, 6 eq.) in dry pyridine (10 ml/1 mmol of the starting compound) was added. The reaction was quenched by evaporation of solvents and the residue was coevaporated with toluene. The solid residue was adsorbed on silica gel in a mixture of cyclohexane/acetone and purified by flash chromatography on silica gel (chloroform to 20% methanol), followed by purification on HPLC (C18, gradient H2O/McOH as eluent). Purified product was lyophilized from dioxane, yielded 112 mg (70%) of the title compound as a white solid. 1H NMR (401 MHz, DMSO-d6) δ 12.67 (s, 1H), 12.10 (s, 1H), 7.83 (s, 1H), 7.65 (s, 1H), 7.44 - 7.11 (m, 10H), 7.04 (ddd, J= 8.6, 7.4, 1.6 Hz, 1H), 6.79 (dd, J = 8.3, 1.2 Hz, 1H), 6.75 (ddd, J= 7.6, 1.1 Hz, 1H), 6.51 (dd, J= 7.8, 1.6 Hz, 1H), 4.70 (dd, J= 11.8, 10.8 Hz, 1H), 4.45 (dd, J= 12.2, 10.6 Hz, 1H), 4.28 - 4.05 (m, 2H), 4.05 - 3.92 (m, 4H), 3.80 (d, J= 9.0 Hz, 2H), 3.04 - 2.80 (m, 4H), 1.26 - 0.72 (m, 6H). 13C NMR (101 MHz, DMSO-d6) δ 174.82 - 171.21 (m), 154.16 (d,J= 12.5 Hz), 153.64, 145.99, 142.67, 137.03 (d, J= 10.2 Hz), 133.74, 129.53, 128.12, 127.91, 126.66 - 126.15 (m), 125.59, 125.34, 121.53, 119.30, 111.65, 101.17, 66.35, 65.22 (d, J= 137.0 Hz), 60.46, 54.99 - 50.19 (m), 15.40 - 10.15 (m). 31P NMR (162 MHz, DMSO-d6) δ 22.20. HRMS (ESI- FTMS) m/z: [M + H]+ calcd for C35H3907N5PS, 704.2302; found, 704.2298.
Example 98
7-((2-(((((S)-l-(Isopropoxycarbonyl)-2-
(phenyl)ethyl)amino)(phenoxy)phosphoryl)methoxy)phenyl)thio)-3 ,5 -dihydro-4H-pyrrolo [3 ,2- d]pyrimidin-4-one L98
Figure imgf000067_0001
A flask was charged with compound L96 (300 mg, 0.6858 mmol, 1 eq.) and it was sealed with septum, flushed with argon, and then dry pyridine (10 ml/1 mmol of the starting compound) was added followed by trimethylsilyl bromide (1 ml/1 mmol of the starting compound). The mixture was stirred at room temperature overnight and then it was evaporated to dryness under an argon atmosphere. L-phenylalanine isopropyl ester hydrochloride (335 mg, 1.37 mmol, 2 eq.) was added and the flask was sealed with septum, flushed with argon and then dry pyridine (10 ml/1 mmol of the starting compound) and triethylamine (572 μl, 4.11 mmol, 6 eq.) were added. The mixture was stirred 15 minutes at 70 °C and a solution of triphenylphosphine (1.08 g, 4.11 mmol, 6 eq.) and Aldrithiol™-2 (905 mg, 4.11 mmol, 6 eq.) in dry pyridine (10 ml/1 mmol of the starting compound) was added. The mixture was stirred overnight at 70 °C. The reaction was quenched by evaporation of solvents and the residue was coevaporated with toluene. The solid residue was adsorbed on silica gel in a mixture of cyclohexane/acetone and purified by flash chromatography on silica gel (chloroform to 20% methanol), followed by purification on HPLC (C18, gradient H2O/McOH as eluent). The purified product was lyophilized from dioxane, yielded 130 mg (31%) of the title compound as a white solid. 1H NMR (401 MHz, DMSO-d6) δ 12.71 - 12.62 (m, 1H), 12.14 - 12.03 (m, 1H), 7.85 - 7.78 (m, 1H), 7.71 - 7.61 (m, 1H), 7.37 - 7.11 (m, 10H), 7.09 - 6.99 (m, 1H), 6.97 - 6.84 (m, 1H), 6.82 - 6.73 (m, 1H), 6.53 - 6.46 (m, 1H), 4.87 - 4.65 (m, 1H), 4.28 - 4.23 (m, 1H), 4.36 - 4.12 (m, 1H), 3.04 - 2.80 (m, 2H), 1.12 - 0.94 (m, 6H). 13CNMR(101 MHZ, DMSO-d6) δ 171.91, 153.86 (d, J= 13.0 Hz), 153.68 - 153.59 (m), 150.24
- 149.91 (m), 146.16 - 145.82 (m), 142.71, 137.24 - 136.61 (m), 133.82, 129.82 - 129.21 (m), 128.38
- 128.04 (m), 128.04 - 127.70 (m), 126.76 - 126.38 (m), 125.71 - 125.49 (m), 125.45 - 125.32 (m), 124.55, 121.85, 120.92 - 120.45 (m), 119.43 - 119.15 (m), 111.75, 100.94, 68.29 - 67.83 (m), 64.03 (d, J= 158.1 Hz), 55.50 - 55.02 (m), 39.52, 21.60 - 21.01 (m). 31P NMR (162 MHz, DMSO-d6) δ 22.97, 22.91. HRMS (ESI-FTMS) m/z: [M + H]+ calcd for C31H3206N4PS, 619.1775; found, 619.1773. Example 99
7-((2-((Bis(((S)-1-(isopropoxycarbonyl)-2-(phenyl)ethyl)amino)phosphoryl)methoxy)phenyl)thio)- 3 ,5 -dihydro-4H-pyrrolo [3.2-ri|pyrimidin-4-onc L99
Figure imgf000068_0001
The title compound was recovered as a byproduct of the previous procedure. The purified product was lyophilized from dioxane, yielded 224 mg (45%) of the title compound as a white solid. 1H NMR (401 MHz, DMSO-d6) δ 7.82 (s, 1H), 7.64 (s, 1H), 7.30 - 7.10 (m, 1 OH), 7.04 (ddd, J= 8.3, 7.4, 1.6 Hz, 1H), 6.80 (dd, J= 8.3, 1.2 Hz, 1H), 6.75 (ddd, J= 7.6, 1.1 Hz, 1H), 6.51 (dd, J= 7.8, 1.6 Hz, 1H), 4.88 - 4.73 (m, 2H), 4.71 - 4.59 (m, 1H), 4.48 - 4.34 (m, 1H), 4.22 - 4.10 (m, 1H), 4.10 - 3.99 (m, 1H), 3.82 (d, J = 8.9 Hz, 2H), 3.04 - 2.81 (m, 4H), 1.19 - 0.98 (m, 12H). 13C NMR (101 MHz, DMSO-d6) δ 172.10, 154.18 (d, J= 12.7 Hz), 153.65, 146.01, 142.68, 137.00 (d, J= 6.7 Hz), 133.73, 129.58, 128.12, 127.96, 126.47 (d, J = 4.8 Hz), 125.63, 125.37, 121.58, 119.30, 111.70, 101.17, 68.02, 65.52 (d, J = 169.5 Hz), 54.09 (d, J= 14.7 Hz), 21.59 - 21.11 (m). 31P NMR (162 MHz, DMSO-d6) δ 21.33. HRMS (ESI-FTMS) m/z: [M + H]+ calcd for C37H4307N5PS, 732.2615; found, 732.2613.
Example 100
Mono(tetrabutylammonium) 7 -((2-((phosphonato)methoxy)phenyl)thio)-3.5-dihydro-4H-pyrrolo [3,2- ri]pyrimidin-4-one L100
Figure imgf000069_0001
A flask charged with the compound L96 (12.0 g, 27.4 mmol, 1 eq.) was sealed with septum, flushed with argon, and then dry pyridine (10 ml/1 mmol of the starting compound) was added followed by trimethylsilyl bromide (1 ml/1 mmol of the starting compound). The mixture was stirred at room temperature overnight and then it was evaporated to dryness and coevaporated with toluene (2x). The residue was dissolved in an aqueous solution of triethylammonium bicarbonate (2M) and tetrabutylammonium hydroxide (2 ml of -1.5M aq. solution/1 mmol of the starting compound) was added. The solution was filtered through P2 and P4 glass filter and it was evaporated to dryness. The resulting oil was dissolved in methanol and adsorbed on C18 silica gel, followed by Cl 8 reverse phase flash chromatography (water to 50% of methanol), yielded 14.5 g (89%) of the title mono(tetrabutylammonium) salt as a white solid. 1H NMR (401 MHz, DMSO-d6) δ 7.83 (s, 1H), 7.72 (s, 1H), 7.07 (dd, J= 8.3, 1.2 Hz, 1H), 6.94 (dd, J= 8.2, 7.8, 1.7 Hz, 1H), 6.61 (dd, J= 7.5, 1.2 Hz, 1H), 6.51 (dd, 1.7 Hz, 1H), 3.87 (d, J= 9.2 Hz, 2H), 3.24 - 3.09 (m, 8H), 1.63 - 1.46 (m, 8H), 1.37
- 1.20 (m, 8H), 0.92 (t, J = 13 Hz, 12H). 13C NMR (101 MHz, DMSO-d6) δ 155.84 (d, J = 9.4 Hz), 154.81, 146.22, 143.25, 134.38, 127.71, 125.51, 125.17, 120.05, 119.42, 111.78, 100.63, 67.56 (d, J = 151.6 Hz), 57.51, 23.08, 19.20, 13.50. 31P NMR (162 MHz, DMSO-d6) δ 10.93. MS (ESI-QMS) m/z for the acid: [M + H]+ calcd for C13H1305N3PS, 354.0; found, 354.0.
Example 101
7-((2-(((((S)-1-(Isopropoxycarbonyl)ethyl)amino)(phenoxy)phosphoryl)methoxy)phenyl)thio)-3,5- dihydro-4H-pyrrolo|3.2-ri|pyrimidin-4-onc L101 A flask charged with compound L100 (5.00 g, 8.40 mmol, 1 eq.), L-alanine isopropyl ester hydrochloride (2.50 g, 14.9 mmol, 1.77 eq.) and phenol (3.49 g, 37.1 mmol, 4.42 eq.) was sealed with septum and flushed with argon. Then, dry pyridine (40 ml/1 mmol of the starting compound) and triethylamine (12.4 ml, 89.0 mmol, 10.6 eq.) were added. The mixture was stirred 15 minutes at 60 °C and a solution of triphenylphosphine (11.5 g, 44.0 mmol, 5.24 eq.) and Aldrithiol™-2 (13.6 g, 61.7 mmol, 7.34 eq.) in dry pyridine (20 ml/1 mmol of the starting compound) was added. The mixture was stirred overnight at 60 °C. The reaction was quenched by evaporation of solvents and the residue was coevaporated with toluene (2x). The solid residue was adsorbed on silica gel in a mixture of cyclohexane/acetone and purified by flash chromatography on silica gel (chloroform to 10% methanol). This purification was repeated once more, yielded 3.52 g (77%) of the title compound as a white solid. 1H NMR (401 MHz, DMSO-d6) δ 12.66 (s, 1H), 12.09 (s, 1H), 7.83 (d, J= 1.6 Hz, 1H), 7.69 (d, J= 8.0 Hz, 1H), 7.44 - 7.34 (m, 2H), 7.34 - 7.22 (m, 2H), 7.22 - 7.13 (m, 1H), 7.13 - 7.07 (m, 1H), 7.08 - 7.00 (m, 1H), 6.82 - 6.71 (m, 1H), 6.57 - 6.47 (m, 1H), 5.97 - 5.76 (m, 1H), 4.93 - 4.72 (m, 1H), 4.59 - 4.37 (m, 2H), 4.21 - 4.04 (m, 1H), 1.37 - 1.19 (m, 3H), 1.16 - 1.04 (m, 6H). 13C NMR (101 MHz, DMSO-d6) δ 173.13 - 172.79 (m), 154.10 - 153.85 (m), 153.67, 150.29 - 149.99 (m), 146.00, 142.74, 134.08 - 133.74 (m), 129.68, 128.16 - 127.88 (m), 125.75 - 125.58 (m), 125.47, 124.76 - 124.53 (m), 122.02 - 121.78 (m), 120.95 - 120.73 (m), 119.35, 112.13 - 111.70 (m), 101.06 - 100.79 (m), 68.01, 65.72 - 63.04 (m), 49.93 - 48.85 (m), 21.73 - 21.04 (m), 20.56 - 19.98 (m).31P NMR (162 MHz, DMSO-d6) δ 23.09, 22.75. HRMS (ESI-FTMS) m/z: [M + H]+ calcd for C25H2806N4PS, 543.1462; found, 543.1462.
Example 102
7-((2-((Bis(((S)-1-(isopropoxycarbonyl)ethyl)amino)phosphoryl)methoxy)phenyl)thio)-3, 5 -dihydro- 4H-pyrrolo [3.2-ri|pyrimidin-4-onc L 102 The title compound was recovered as a byproduct of the procedure from Example 96 with L-alanine isopropyl ester hydrochloride (230 mg, 1.37 mmol, 2 eq.) as the amino acid. The purified product was lyophilized from dioxane, yielded 279 mg (75%) of the title compound as a white solid. 1H NMR (401 MHz, DMSO-d6) δ 12.67 (s, 1H), 12.09 (s, 1H), 7.82 (d, J= 2.8 Hz, 1H), 7.66 (d, J= 2.2 Hz, 1H), 7.11
- 6.98 (m, 2H), 6.75 (ddd, J= 7.8, 5.7, 2.8 Hz, 1H), 6.55 - 6.44 (m, 1H), 4.93 - 4.83 (m, 2H), 4.81 - 4.72 (m, 1H), 4.71 - 4.62 (m, 1H), 4.30 - 4.16 (m, 2H), 4.11 - 3.99 (m, 1H), 3.99 - 3.86 (m, 1H), 1.37
- 1.28 (m, 6H), 1.21 - 1.14 (m, 12H). 13C NMR (101 MHz, DMSO-d6) δ 173.47 (d, J= 5.1 Hz), 154.36 (d, J = 12.4 Hz), 153.65, 145.97, 142.67, 133.77, 127.99, 125.66, 125.46, 121.59, 119.29, 111.86, 101.18, 67.85, 65.71 (d, J= 135.0 Hz), 48.40 (d, J= 10.3 Hz), 21.43, 20.88 (dd, J= 13.6, 4.9 Hz).31P
NMR (162 MHz, DMSO-d6) δ 21.41. HRMS (ESI-FTMS) m/z: [M + H]+ calcd for C25H3507N5PS, 580.1989; found, 580.1985.
Example 103
Triethylammonium 7-((5-fluoro-2-((phosphonato)methoxy)phenyl)thio)-3.5-dihydro-4H-pyrrolo|3.2- ri]pyrimidin-4-one LI 03
Figure imgf000071_0001
The coupling product from Example 69 (step 1) (444 mg, 0.6570 mmol, 1 eq.) was dissolved in trifluoroacetic acid (2 ml/100 mg of the material) at room temperature and stirred for 15 minutes. Trifluoroacetic acid was evaporated, coevaporated with water (2x) and a cold solution of ammonia in ethanol (2 ml/100 mg of the material) was added. The mixture was evaporated to dryness and the solid was adsorbed on silica gel in a mixture of cyclohexane/acetone. Purification by flash chromatography on silica gel (chloroform to 15% methanol) afforded 243 mg (81%) of the diisopropyl phosphonate. A flask charged with the diisopropyl phosphonate was sealed with septum, flushed with argon, and then dry pyridine (10 ml/1 mmol of the starting compound) was added followed by trimethylsilyl bromide (1 ml/1 mmol of the starting compound). The mixture was stirred at room temperature overnight and then it was evaporated to dryness. The residue was dissolved in a 2M aq. solution of triethylammonium bicarbonate and a small amount of methanol and it was evaporated to dryness. The solid was dissolved in methanol and adsorbed on C 18 silica gel, followed by C 18 reverse phase flash chromatography (water to methanol). Solvents were evaporated and the solid was lyophilized from dioxane, yielded 293 mg (78% overall) of the title bis(triethylammonium) salt as a white solid which was used in the next step without further characterization. MS (ESI-QMS) m/z for the acid: [M + H]+ calcd for C13H1205N3FPS, 372.0; found, 372.2 .
Example 104
7-((5 -Fluoro-2-(((((S)-1 -
(isopropoxycarbonyl)ethyl)amino)(phenoxy)phosphoryl)methoxy)phenyl)thio)-3,5-dihydro-4H- pyrrolo[3.2-d]|pyrimidin-4-one L104
Figure imgf000072_0001
A flask charged with compound L103 (150 mg, 0.2615 mmol, 1 eq.), F-alanine isopropyl ester hydrochloride (78 mg, 0.4629 mmol, 1.77 eq.) and phenol (109 mg, 1.16 mmol, 4.42 eq.) was sealed with septum, flushed with argon and then dry pyridine (40 ml/1 mmol of the starting compound) and triethylamine (386 μl, 2.78 mmol, 10.62 eq.) were added. The mixture was stirred 15 minutes at 60°C and a solution of triphenylphosphine (359 mg, 1.37 mmol, 5.24 eq.) with Aldrithiol™-2 (422 mg, 1.92 mmol, 7.34 eq.) in dry pyridine (20 ml/1 mmol of the starting compound) was added. The mixture was stirred overnight at 60 °C. The reaction was quenched by evaporation of solvents and the residue was coevaporated with toluene. The solid residue was adsorbed on silica gel in a mixture of cyclohexane/acetone and purified by flash chromatography on silica gel (chloroform to 10% methanol), followed by purification on HPFC (Cellulose SC, heptane/ethanol (80:20) as the eluent). Purified product was lyophilized from dioxane, yielded 69 mg (47%) of the title compound as a white solid. 1H NMR (401 MHz, DMSO-d6) δ 12.76 (s, 1H), 12.15 (s, 1H), 7.85 (d, J= 1.5 Hz, 1H), 7.74 (d, J= 6.7 Hz, 1H), 7.42 - 7.33 (m, 2H), 7.33 - 7.22 (m, 2H), 7.22 - 7.15 (m, 1H), 7.15 - 7.09 (m, 1H), 6.87 (ddd, J= 8.5, 3.1 Hz, 1H), 6.26 - 6.15 (m, 1H), 5.96 - 5.80 (m, 1H), 4.92 - 4.70 (m, 1H), 4.61 - 4.35 (m, 2H), 4.23 - 3.99 (m, 1H), 1.31 - 1.19 (m, 3H), 1.17 - 1.03 (m, 6H). 13CNMR (101 MHz, DMSO-d6) δ 173.13 - 172.56 (m), 158.52 - 155.80 (m), 153.64, 150.45 - 149.80 (m), 145.76, 143.01, 134.14 - 133.89 (m), 130.91 - 130.56 (m), 129.70, 124.85 - 124.49 (m), 120.96 - 120.61 (m), 119.50, 113.68 - 112.93 (m),
111.92 (d, J= 30.3 Hz), 111.28 (d, J= 23.7 Hz), 100.15 - 99.84 (m), 68.26 - 67.87 (m), 66.05 - 63.61 (m), 49.30 (d, J = 7.0 Hz), 21.49 - 21.26 (m), 20.50 - 20.18 (m). 31P NMR (162 MHz, DMSO-d6) δ 22.92, 22.50. 19F NMR (377 MHz, DMSO-d6) δ -121.52. HRMS (ESI-FTMS) m/z: [M + H]+ calcd for C25H2706N4FPS, 561.1368; found, 561.1365.
Example 105
7-((5-Fluoro-2-((bis(((S)-1 -(isopropoxycarbonyl)cthyl)amino)phosphoryl)methoxy)phenyl)thio)-3.5- dihydro-4H-pyrrolo[3.2-d]|pyrimidin-4-onc L105
Figure imgf000073_0001
The title compound was recovered as a byproduct from the previous procedure. The purified product was lyophilized from dioxane, yielded 21 mg (13%) of the title compound as a white solid. 1H NMR (401 MHz, DMSO-d6) δ 12.75 (s, 1H), 12.14 (s, 1H), 7.85 (s, 1H), 7.70 (s, 1H), 7.06 (dd, J= 9.0, 4.6 Hz, 1H), 6.86 (ddd, J= 8.9, 8.2, 3.1 Hz, 1H), 6.19 (dd, J= 9.2, 3.1 Hz, 1H), 4.88 (hept, J= 6.3 Hz, 2H), 4.79 (dd, J= 12.1, 10.2 Hz, 1H), 4.67 (dd, J= 11.6, 10.1 Hz, 1H), 4.34 - 4.15 (m, 2H), 4.09 - 3.86 (m, 2H), 1.38 - 1.24 (m, 6H), 1.24 - 1.08 (m, 12H). 13C NMR (101 MHz, DMSO-d6) δ 173.45 (d, J= 5.3
Hz), 157.04 (d, J= 237.8 Hz), 153.63, 150.67 (d, J= 12.0 Hz), 145.76, 142.96, 133.89, 130.69 (d, J = 7.9 Hz), 119.47, 113.26 (d, J= 8.6 Hz), 111.89 (d, J= 27.1 Hz), 111.24 (d, J= 23.1 Hz), 100.21, 67.87, 66.34 (d, J= 135.2 Hz), 48.40 (d, J= 11.6 Hz), 21.44, 21.24 - 20.51 (m). 19F NMR (377 MHz, DMSO- d6) δ -121.96. 31P NMR (162 MHz, DMSO-d6) δ 21.27. HRMS (ESI-FTMS) m/z: [M + H]+ calcd for C25H3407N5FPS, 598.1895; found, 598.1892.
Example 106
Triethylammonium 7 -((3 -methoxy-2-((phosphonato)methoxy)phenyl)thio)-3,5-dihydro-4H- pyrrolo[3.2-d]|pyrimidin-4-one L106
Figure imgf000073_0002
The coupling product from Example 70 (455 mg, 0.6615 mmol, 1 eq.) was dissolved in trifluoroacetic acid (2 ml/100 mg of the material) at room temperature and stirred for 15 minutes. Trifluoroacetic acid was evaporated, coevaporated with water (2x) and a cold solution of ammonia in ethanol (2 ml/100 mg of the material) was added. The mixture was evaporated to dryness and the solid was adsorbed on silica gel in a mixture of cyclohexane/acetone. Purification by flash chromatography on silica gel (chloroform to 15% methanol) afforded 245 mg (79%) of the diisopropyl phosphonate. A flask charged with the diisopropyl phosphonate was sealed with septum, flushed with argon, and then dry pyridine (10 ml/1 mmol of the starting compound) was added followed by trimethylsilyl bromide (1 ml/1 mmol of the starting compound). The mixture was stirred at room temperature overnight and then it was evaporated to dryness. The residue was dissolved in a 2M aq. solution of triethylammonium bicarbonate and a small amount of methanol and it was evaporated to dryness. The solid was dissolved in methanol and adsorbed on C18 silica gel, followed by Cl 8 reverse phase flash chromatography (water to methanol). Solvents were evaporated and the solid was lyophilized from dioxane, yielded 300 mg (78% overall) of the title bis(triethylammonium) salt as a white solid. 1H NMR (401 MHz, DMSO-d6) δ 7.83 (s, 1H), 7.68 (s, 1H), 6.78 (dd, J= 8.0 Hz, 1H), 6.72 (dd, J= 8.3, 1.6 Hz, 1H), 6.09 (dd, J= 7.8, 1.6 Hz, 1H), 3.99 (d, J= 9.9 Hz, 2H), 3.75 (s, 3H), 3.01 (q, J= 7.3 Hz, 13H), 1.17 (t, J= 7.3 Hz, 19H). 13C NMR (101 MHz, DMSO- d6) δ 153.79, 152.27, 145.85, 144.12 (d, J= 14.0 Hz), 142.73, 134.00, 133.81, 124.03, 119.21, 117.73, 109.65, 101.61, 69.43 (d, J= 155.6 Hz), 55.93, 45.35, 8.70. 31P NMR (162 MHz, DMSO-d6) δ 13.28. MS (ESI-QMS) m/z for the acid: [M + H]+ calcd for C14H1506N3PS, 383.0; found, 383.0.
Example 107 7-((3-Methoxy-2-(((((S)-1 -
(isopropoxycarbonyl)ethyl)amino)(phenoxy)phosphoryl)methoxy)phenyl)thio)-3,5-dihydro-4H- pyrrolo|3.2-ri|pyrimidin-4-onc L107
Figure imgf000074_0001
A flask charged with compound L106 (150 mg, 0.2561 mmol, 1 eq.), L-alanine isopropyl ester hydrochloride (76 mg, 0.4533 mmol, 1.77 eq.) and phenol (106 mg, 1.13 mmol, 4.42 eq.) was sealed with septum, flushed with argon, and then dry pyridine (40 ml/1 mmol of the starting compound) and triethylamine (378 μl, 2.72 mmol, 10.62 eq.) were added. The mixture was stirred 15 minutes at 60°C and a solution of triphenylphosphine (352 mg, 1.34 mmol, 5.24 eq.) with Aldrithiol™-2 (414 mg, 1.88 mmol, 7.34 eq.) in dry pyridine (20 ml/1 mmol of the starting compound) was added. The mixture was stirred overnight at 60 °C. The reaction was quenched by evaporation of solvents and the residue was coevaporated with toluene. The solid residue was adsorbed on silica gel in a mixture of cyclohexane/acetone and purified by flash chromatography on silica gel (chloroform to 10% methanol), followed by purification on HPLC (Cellulose SC, heptane/ethanol (80:20) as the eluent). Purified product was lyophilized from dioxane, yielded 31 mg (21%) of the racemic title compound as a white solid. 1H NMR (401 MHz, DMSO-d6) δ 12.67 (s, 1H), 12.09 (s, 1H), 7.92 - 7.76 (m, 1H), 7.70 (s, 1H), 7.41 - 7.33 (m, 2H), 7.32 - 7.22 (m, 2H), 7.21 - 7.11 (m, 1H), 6.90 - 6.83 (m, 1H), 6.83 - 6.75 (m, 1H), 6.22 - 6.04 (m, 1H), 5.88 - 5.68 (m, 1H), 4.93 - 4.74 (m, 1H), 4.60 - 4.27 (m, 2H), 4.13 - 3.90 (m, 1H), 3.82 - 3.74 (m, 3H), 1.28 - 1.20 (m, 3H), 1.16 - 1.08 (m, 6H). 13CNMR (101 MHz, DMSO-d6) δ 173.19
- 172.78 (m), 153.63, 151.97 - 151.70 (m), 150.56 - 150.16 (m), 146.03 - 145.70 (m), 142.88, 142.75, 133.95, 133.56 - 133.25 (m), 129.57, 124.82 - 124.65 (m), 124.44, 121.03 - 120.54 (m), 119.27, 117.99
- 117.54 (m), 109.57, 101.34 - 100.89 (m), 68.07 - 67.79 (m), 68.23 - 66.19 (m), 55.94, 49.43 - 49.03 (m), 21.63 - 21.05 (m), 20.51 - 19.82 (m).31P NMR (162 MHz, DMSO-d6) 523.11. HRMS (ESI-FTMS) m/z: [M + H]+ calcd for C26H30O7N4PS, 573.1567; found, 573.1563.
Example 108
7-((3-Methoxy-2-((bis(((S)-1-(isopropoxycarbonyl)ethyl)amino)phosphoryl)methoxy)phenyl)thio)- 3 , 5 -dihydro-4H-pyrrolo [3 ,2-d]pyrimidin-4-one L 108
Figure imgf000075_0001
The title compound was recovered as a byproduct of the previous procedure. The purified product was lyophilized from dioxane, yielded 42 mg (27%) of the racemic title compound as a white solid. 1H NMR (401 MHz, DMSO-d6) δ 12.68 (s, 1H), 12.10 (s, 1H), 7.83 (s, 1H), 7.72 (s, 1H), 6.85 (dd, J= 8.0 Hz, 1H), 6.79 (dd, J= 8.3, 1.6 Hz, 1H), 6.15 (dd, J= 7.9, 1.5 Hz, 1H), 4.98 - 4.80 (m, 2H), 4.80 - 4.59 (m, 2H), 4.30 - 4.11 (m, 2H), 4.07 - 3.86 (m, 2H), 3.82 (s, 3H), 1.39 - 1.24 (m, 6H), 1.20 - 1.12 (m, 12H). 13C NMR (101 MHZ, DMSO-d6) δ 173.48, 153.62, 151.74, 145.84, 143.23 (d, J = 11.8 Hz), 142.78, 133.95, 133.32, 124.70, 119.24, 118.05, 109.54, 101.14, 68.91 (d, J= 132.5 Hz), 67.90, 55.89, 48.35 (d, J = 4.5 Hz), 21.44, 21.04 - 20.61 (m). 31P NMR (162 MHz, DMSO-d6) δ 21.31. HRMS (ESI-FTMS) m/z: [M + H]+ calcd for C26H3708N5PS, 610.2095; found, 610.2093.
Example 109 Tert- butyl 4-(benzyloxy)-7-((2-(((/er/-butyldimethylsilyl)oxy)methyl)phenyl)amino)-5H-pyrrolo[3,2- 1pyrimidine-5-carboxy1ate L109
Figure imgf000076_0001
All reactants were added into a secured Schlenk flask 2-(((tert-butyldimethylsilyl)oxy)methyl)aniline (1.22 g, 5.14 mmol, 1 eq.), compound L3 (1.04 g, 5.14 mmol, 1 eq.), cesium carbonate (2.5 g, 7.72 mmol, 1.5 eq.), chloro(2-dicyclohexylphosphino-2', 4', 6'-triisopropyl- 1,1 '-biphenyl) [2-(2'-amino- 1,1'- biphenyl)]palladium(II) (200 mg, 0.25 mmol, 0.05 eq.), 2-dicyclohexylphosphino-2',4',6'- tri/.vopropylbiphenyl (130 mg, 0.27 mmol, 0.05 eq.). The Schlenk flask was flushed with argon and toluene (10 mL) was added. The resulting suspension was heated at 110 °C for 16 hours. Solids were filtered through a celite pad and washed with ethyl acetate (50 mL). Solvents were evaporated and crude product was purified by Flash silica gel chromatography using gradient cyclohexane/ethyl acetate (0- 40%) to give the title compound as a white solid (1.14 g, 67%). 1H NMR (400 MHz, DMSO-d6) δ 8.55 (s, 1H), 7.71 (s, 1H), 7.66 (s, 1H), 7.52 (m, 2H), 7.38 (m, 2H), 7.32 (m, 1H), 7.27-7.23 (m, 2H), 7.20 (m, 1H), 6.84 (td, J= 7.3 Hz, 1.3, 1H), 5.63 (s, 2H), 4.79 (s, 2H), 1.46 (s, 9H), 0.90 (s, 9H), 0.10 (s, 6H). 13C NMR (100 MHz, DMSO-d6) δ 155.79, 151.26, 148.66, 148.23, 142.05, 136.62, 128.86, 128.51, 128.41, 128.09, 128.02, 127.48, 123.65, 119.58, 114.74, 114.67, 112.38, 84.54, 67.92, 63.98, 27.49, 25.93 , 18.10, -5.11. HRMS (ESI-FTMS) m/z : [M + H] + ealed for C3 lH40O4N4NaSi, 583.2711 ; found, 583.2713.
Example 110
Tert-butyl 4-(benzyloxy)-7-((2-(hydroxy methyl )phenyl)ami no)-5H-py rrolo| 3, 2-d]pyrimidine-5- carboxylate LI 10
Figure imgf000076_0002
Solution of TBAF.3H2O (1.9 mL, 1M, 1.9 mmol, 1 eq.) in THF (50 mL) was buffered with acetic acid (101 μl , 1.7 mmol, 1.03 eq.). Compound L109 (1.1 g, 1.96 mmol) was added in one portion and the solution was stirred at 25 °C for 16 hours. Solvent was evaporated and the crude product was purified by Flash chromatography using cyclohexane/EtOAc (0-80%). This procedure afforded the title compound as a solid (467 mg, 53%). 1H NMR (400 MHz, CDCl3) δ 8.56 (s, 1H), 7.72 (s, 1H), 7.57 (bs, 1H), 7.53 (m, 2H), 7.38 (m, 2H), 7.33 (m, 1H), 7.28-7.26 (m, 2H), 7.13 (bd, J= 7.4 Hz, 1H), 6.82 (m, 1H), 5.67 (s, 2H), 4.81 (s, 2H), 1.53 (s, 9H). 13C NMR (400 MHz, CDCl3) δ 156.23, 151.13, 148.52, 148.51, 142.77, 136.32, 129.35, 129.20, 128.43, 128.04, 128.02, 127.02, 123.45, 119.57, 115.02, 114.54, 113.03, 84.63, 68.39, 64.40, 27.85. HRMS (ESI-FTMS) m/z: [M - H]- calcd for C25H2 504N4, 445.1881; found, 445.1882.
Example 111
Tert-butyl 4-(benzyloxy)-7-((2-formyl phenyl )amino)-5H-pyrrolo| 3.2-d]pyrimidinc-5-carboxylatc Llll
OBn Boc
Figure imgf000077_0001
Compound L110 (400 mg, 0.90 mmol, 1 eq.) was dissolved in dichloromethane (20 mL) and MnO2 (780 mg, 8.9 mmol, 9.9 eq.) was added. The suspension was stirred at 35 °C for 6 hours. Solids were filtered through a celite pad and washed with dichloromethane (10 mL). Solvents were evaporated and the crude product was purified by Flash chromatography using gradient cyclohexane/ethyl acetate (0-30%) to give title compound as a white solid (142 mg, 36%, mixture of two rotamers). 1H NMR (400 MHz, CDCl3) δ 10.43 (bs, 1H), 9.97 (s, 1H), 8.67 (s, 1H), 7.89 (s, 1H), 7.62 (dd, J= 7.7 Hz, 1.6, 1H), 7.53 (m, 2H), 7.47 (m, 1H), 7.38 (m, 2H), 7.33 (m, 1H), 7.13 (d, J= 8.5 Hz, 1H), 6.91 (m, 1H), 5.67 (s, 2H), 1.54 (s, 9H). 13C NMR (400 MHz, CDC13) δ 194.28, 156.21, 156.20, 152.07, 152.04, 149.67, 149.62, 148.35, 146.65, 136.65, 136.36, 135.76, 128.44, 128.11, 128.06, 121.04, 121.02, 119.92, 119.90, 119.84, 117.68, 113.51, 113.14, 85.08, 85.07, 68.34, 68.32, 27.82. HRMS (ESI-FTMS) m/z: [M + H]+ calcd for C25H2 504N4, 445.1870; found, 445.1872.
Example 112
Diethyl (E )-(2-((4-oxo-4,5-dihydro-3i7-pyrrolo[3,2-d]pyrimidin-7-yl)amino)styryl)phosphonate L112
Figure imgf000077_0002
Tetraethyl methylene bisphosphonate (152 μl , 0.608 mmol, 1.2 eq.) and potassium tert-butoxidc (68 mg, 0.608 mmol, 1.2 eq.) were stirred in THF (8 mL) at 25 °C for 10 min. Then, compound Llll (180 mg, 0.405 mmol) was added dropwise in THF (7 mL) and the mixture was stirred at 25 °C for 30 min. The reaction mixture was diluted by saturated aqueous NH4CI (30 mL) and extracted by ethyl acetate (3x). Organic layer was dried by Na2S04 and evaporated. The crude residue was dissolved in trifluoroacetic acid (4 ml) and directly evaporated in vacuo. The crude product was purified by Flash chromatography using gradient chloroform/MeOH (0-20%) to give title compound as a white solid (91 mg, 58%). 1H NMR (400 MHz, DMSO-d6) δ 11.98 (bs, 1H), 11.92 (bs, 1H), 7.79 (dd, J= 22.8, 17.2 Hz, 1H), 7.73 (s, 1H), 7.56 (dd, J= 7.8, 1.5 Hz, 1H), 7.48 (bs, 1H), 7.22 (d, J= 1H), 7.07 (ddd, J= 8.4, 7.0, 1.5 Hz, 1H), 6.68 (m, 1H), 6.56 (dd, J= 8.4, 1.0 Hz, 1H), 6.34 (dd, J= 18.9, 17.2 Hz, 1H), 4.04 (m, 4H), 1.25 (t, 6H, J = 7.0 Hz). 31P NMR (400 MHz, DMSO-d6) δ 22.68. 13C NMR (100 MHz, DMSO- d6) d 153.88, 146.52, 143.86 (d, J = 7.8 Hz), 141.13, 139.20, 131.06, 127.11, 122.36, 120.85 (d, J = 22.8 Hz), 119.45, 117.96, 117.23, 115.24, 112.90 (d, J= 188.0 Hz), 61.26 (d, J= 5.3 Hz), 16.50 (d, J = 6.0 Hz). HRMS (ESI-FTMS) m/z: [M + H]+ calcd for C18H2 104N4NaP, 411.1193; found, 411.1194. Example 113 (E )-(2-((4-Oxo-4.5-dihydro-3H-pyrrolo|3.2-d|pyrimidin-7-yl)amino)styryl)phosphonic acid L113
Figure imgf000078_0001
Compound L112 (55mg, 0.14 mmol, 1 eq.) was dissolved in pyridine (4 mL) at 0 °C and bromotrimethylsilane (10 ml/1 mmol of the starting material) was added dropwise. The solution was stirred at 25 °C for 16 hours. Solvents were evaporated and the crude product was purified by reverse phase HPLC using gradient water (TEAB 0.1M as modifier)/methanol (5-95%, Xterra column, 5 μm, 10 x 100 mm) to give title compound as a white solid (10 mg, 21%). 1H NMR (400 MHz, D2O) <5 7.88 (s, 1H), 7.51-7.41 (m, 2H), 7.43 (s, 1H), 7.08 (m, 1H), 6.89 (m, 1H), 6.60 (dd, J= 8.2, 1.2 Hz, 1H), 6.42 (t, J= 16.8 Hz, 1H). 31P NMR (400 MHz, D20) δ 15.12. 13C NMR (100 MHz, D20) δ 156.04, 145.37, 142.18, 140.51, 136.93 (d, J= 5.9 Hz), 130.44, 127.92, 125.76, 125.54 (d, J= 175.0 Hz), 124.47 (d, J = 20.7 Hz), 120.09, 119.35, 116.9, 114.72. HRMS (ESI-FTMS) m/z: [M - H]- calcd for C14H1204N4P, 331.0601; found, 331.0601.
Example 114
7-(Phenylamino)-3 ,5-dihydro-4H-pyrrolo [3 ,2-d]pyrimidin-4-one L 114
Figure imgf000078_0002
All reactants were added into a secured vial, aniline (70 mg, 0.8 mmol, 3.2 eq.), compound L3 (100 mg, 0.25 mmol), cesium carbonate (170 mg, 0.3 mmol, 1.2 eq.), chloro(2-dicyclohexylphosphino-2',4',6'- tri/.vopropyl- 1. 1 '-biphenyl)|2-(2'-amino- 1. 1 '-biphenyl) |palladium(II) (30 mg, 0.04 mmol), 2- dicyclohexylphosphino-2'.4'.6'-tri/.vopropylbiphenyl (20 mg, 0.04 mmol). The vial was flushed with argon and toluene (3 mL) was added. The resulting suspension was heated at 110 °C for 16 hours. Solids were filtered through the celite pad and washed with ethyl acetate (50 mL). Solvents were evaporated and crude product was purified by Flash silica gel chromatography using gradient cyclohexane/ethyl acetate (0-100%) to give the coupling product as a white solid (54 mg, 52%). The coupling product (25 mg, 0.06 mmol) was dissolved in trifluoroacetic acid (2 mL) and the solution was immediately evaporated in vacuo. Crude compound was purified by reverse phase Flash silica gel chromatography using gradient water/methanol (with 20% DMF as modifier) (0-100%) to give title compound as a white solid (5 mg, 37%). 1H NMR (400 MHz, DMSO-d6) δ 11.90 (bs, 2H), 7.74 (s, 1H), 7.39 (s, 1H), 7.30 (s, 1H), 7.06 (m, 2H), 6.76 (m, 2H), 6.58 (m, 1H). 13C NMR (100 MHz, DMSO-d6) δ 153.90, 147.62, 140.87, 138.78, 128.93, 120.49, 119.75, 116.90, 116.85, 113.30. HRMS (ESI-FTMS) m/z: [M + H]+ calcd for C12H110N4, 227.0927; found, 227.0928.
Example 115
7-((4-Cyanophenyl)amino)-3 ,5 -dihydro-4H-pyrrolo [3 ,2-d]pyrimidin-4-one L 115
Figure imgf000079_0001
All reactants were added into a secured vial 4-aminobenzonitrile (90 mg, 0.8 mmol, 1.6 eq.), compound L3 (200 mg, 0.5 mmol, 1 eq.), cesium carbonate (430 mg, 1.3 mmol, 2.6 eq.), chloro(2- dicyclohexylphosphino-2'.4'.6'-triisopropyl- 1.1 '-biphenyl)|2-(2'-amino- 1. 1 '-biphenyl) |palladium(II)
(58 mg, 0.07 mmol, 0.14 eq.), 2 -dicyclohexyl phosph i no-2', 4', 6'-triisop ropy lbi phenyl (39 mg, 0.08 mmol, 0.16 eq.). The vial was flushed with argon and toluene (3 mL) was added. The resulting suspension was heated at 110 °C for 16 hours. Solids were filtered through the celite pad and washed with ethyl acetate (30 mL). Solvents were evaporated and crude product was purified by Flash silica gel chromatography using gradient cyclohexane/ethyl acetate (0-100%) to give the coupling product as a white solid (128 mg, 58%). The coupling product (37 mg, 0.08 mmol) was dissolved in trifluoroacetic acid (4 mL) and the solution was immediately evaporated in vacuo. Crude compound was purified by Flash silica gel chromatography using dichloromethane/methanol (with 10% DMF as modifier) (3/1, v/v) to give title compound as a poorly soluble white amorphous solid (10 mg, 47%). 1H NMR (400 MHz, DMSO-d6) δ 12.12 (d, J= 3.2 Hz, 1H), 11.95 (d, J= 3.5 Hz, 1H), 8.35 (bs, 1H), 7.77 (d, J= 3.5, 1H), 7.45 (m, 2H), 7.40 (dd, J = 3.2, 0.6 Hz, 1H), 6.77 (m, 2H). 13C NMR (100 MHz, DMSO-d6) δ 153.83, 151.72, 141.50, 139.19, 133.48, 122.32, 120.64, 117.22, 117.20, 113.19, 97.07. HRMS (ESI- FTMS) m/z: [M - H]- calcd for C13H80N5, 250.0734; found, 250.0732.
Biological activity evaluation PNP activity inhibition in vitro
To evaluate inhibitory activity of the compounds towards PNP, recombinant human PNP protein (hPNP) was expressed in E. coli, purified by means of affinity chromatography (NiNTA column, Thermo Fisher Scientific, Waltham, USA) and stored in 20 mM phosphate buffer pH 7.4 containing 0.3 M NaCl in aliquots at -80 °C. All newly synthesized compounds were dissolved either in water or DMSO to yield 10 mM stock solutions. The compounds then underwent basic screening at 10 mM concentration and in case that at least 50% inhibition was observed, dose-response curve was generated to calculate IC50 value (half -maximal inhibitory concentration). Forodesine (MedChemExpress, Monmouth Junction, USA) was used as a reference compound. Reaction mixture for PNP activity determination consisted of 1 mM Pi, 200 pM [2,8-3H] inosine (ARC Inc., Saint Fouis, USA), variable concentrations of tested compound, 1 mM DTT, 0.2 pg/ml BSA and 1.75 pg of PNP protein. Reaction was incubated for 10 min at 37°C and it was terminated by spotting a 2-pl aliquot of PEI-coated cellulose TEC plate (NO. 105579, Merck Darmstadt, Germany). The plate was developed in «-butanol-acetic acid-water at 10: 1 :3 ratio, let dry and analyzed by means of radio-TEC scanner RITA Star (Elysia-Raytest GmbH, Strauberhardt, Germany).
Cytotoxicity evaluation
To evaluate selectivity of the antitumor action of the compounds, T-lymphoblastic cell line was employed (CCRF-CEM) representing the target tissue. As a counterscreen, one promyelocytic leukemia cell line (HL-60) was used. All cell lines were from ATCC (Manassas, VA, USA). The cells were incubated in RPMI-1640 or DMEM culture medium containing 10% FBS and 1% GlutaMax at 37°C under the atmosphere containing 5% CO2. Experiments were performed on the cells between passage No. 10 and 50. For the experiments, cells were seeded in 384-well transparent plates (Brand GmbH, Wertheim, Germany) at a concentration 2,000 - 10,000 cells per well and left to rest in an incubator overnight. The next day, 10 pM deoxyguanosine (Sigma) and varying concentrations of the test compounds were added. The compounds were incubated with the cells in a CO2 incubator at 37°C for 72 h after which XTT dye Sigma-Aldrich, St. Louis, USA) was added. The absorbance signal at 450 nm was recorded after 1 - 4 h using a multimode plate reader. Signal of untreated cells was set to 100% cell viability. The effect of the compounds was expressed as IC50 values calculated by non-linear regression using the GraphPad Prism v. 5.04 (GraphPad Software, San Diego, USA).
Peripheral blood mononuclear cells (PBMC) were isolated from healthy donors (with their informed consent) by centrifugation of the buffy-coats through Ficoll gradient. Immediately after the isolation, cells were seeded in the 384-well white plates (Thermo Fisher Scientific, Waltham, USA) at a concentration 50,000 cells per well. The next day, cells were treated with the test compounds and deoxyguanosine at a concentration of 50 and 10 mM, respectively. The cells were then incubated at 37°C, 5% CO2 for 72 h after which CellTiter-Glo® detection reagent (Promega, Madison, USA) was added. The plate was left on a shaker (350 rμm) for 20 min at RT. Luminescence was measured by a multimode plate reader Cytation 3 (BioTek Instruments Inc., Winooski, USA). The signal of the compound-treated cells was compared to the value of untreated control which was set to 100% viability.
Table 1: Evaluation of in-vitro efficacy of the PNP inhibitors.
Figure imgf000081_0001
Figure imgf000082_0001
Figure imgf000083_0001
Figure imgf000084_0001
Figure imgf000085_0001
Figure imgf000086_0001
Figure imgf000087_0001
ND - not determined
Table 2: Evaluation of kinetic solubility of selected PNP inhibitors in TRIS buffer at pH 7.4, in comparison with known compounds, in particular Peldesine. Peldesine failed in Phase III of clinical studies, likely due to its low solubility and low bioavailability (J. Am. Acad. Dermatol. 2001, 44 (6), 940). The compounds of the present invention have a significantly higher solubility and bioavailability.
Figure imgf000087_0002
Figure imgf000088_0001
Note: Structurally similar compounds exhibit similar physico-chemical properties (i.e. solubility).
Industrial Applicability The invention is applicable in the pharmaceutical industry and clinical use, in preparation and manufacture of medicaments and medical therapy for treatment of serious diseases caused by abnormal growth of white blood cells.

Claims

Claims
1. A compound of general formula I:
Figure imgf000089_0001
wherein:
X is -NH- or -S-;
Ar is aryl or heteroaryl;
Z is not present or Z is selected from -CH2-, -CH2O-, -OCH2- -CH2OCH2- and -CH=CH- attached to the position 2- or 3- of the Ar relative to the X;
G is selected from -COA1, -SO2A1, -P(O)(A1)(A2), -OH and -H, wherein:
A1 and A2 are identical or different groups selected from of -OH, -OR1 and -NHR2, wherein R1 is linear or branched C1-C6 alkyl chain, C6-C12 aryl, arylalkyl and -CH2OC(O)R2 and R2 is linear or branched C1-C10 alkyl chain or -CHR3COOR4, wherein
R3 is linear or branched C1-C10 alkyl chain in which at least one -CH2- group can optionally be replaced by a group independently selected from -S-, -O-, -NH-, C6-C12 aryl and C6-C12 arylalkyl, and R4 is linear or branched C1-C10 alkyl chain;
R is -H or a group selected from -OH, -O(aryl), -O(arylalkyl), -C(O)-, -CN and halogen; with the proviso that when Ar is phenyl and Z is not present, then R and G are not both a hydrogen atom, and their pharmaceutically acceptable salts.
2. Compound of the general formula I according to claim 1, wherein G is selected from -COA1, -SO2A1, -P(O)(A1)(A2).
3. Compound of the general formula I according to claim 1, selected from the group comprising: (E )-7-((2-(2-(phosphonato)vinyl)phenyl)thio)-3,5-dihydro-4H-pyrrolo[3,2-ri]pyrimidin-4-one, 7-((2-((phosphonato)methoxy)phenyl)thio)-3.5-dihydro-4H-pyrrolo [3.2-ri]pyrimidin-4-onc. (E )-7-((4-bromo-2-(2-(phosphonato)vinyl)phenyl)thio)-3,5-dihydro-4H-pyrrolo[3,2-ri]pyrimidin-4- one, (E )- 7 -((5 -bromo-2-(2-(phosphonato)vinyl)phenyl)thio)-3.5-dihydro-4H-pyrrolo [3 ,2-d]pyrimidin-4- one,
(E )- 7 -((5-hydroxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3.5-dihydro-4H-pyrrolo [3 ,2-d]pyrimidin-4- one,
(E)-7-((4-hydroxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3.5-dihydro-4H-pyrrolo[3.2-d]|pyrimidin-4- one,
(E)-7-((4-methoxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3.5-dihydro-4H-pyrrolo[3.2-d]|pyrimidin-4- one,
(E)-7-((4-isopropoxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3.5-dihydro-4H-pyrrolo[3.2-d]|pyrimidin-
4-one,
(E )-7-((4-(2,3,4,5,6-pentafluorophenyl)methoxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3,5-dihydro- 4H-pyrrolo [3 ,2-d]pyrimidin-4-one,
(E )-7-((4-(2,3,4,5,6-pentafluorophenyl)oxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3,5-dihydro-4H- pyrrolo [3 ,2-d]pyrimidin-4-one,
(E )-7 -((3 -hydroxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3 ,5 -dihydro-4H-pyrrolo [3 ,2-d]pyrimidin-4- one,
(E)-7-((3-methoxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3.5-dihydro-4H-pyrrolo[3.2-d]|pyrimidin-4- one,
(E)-7-((3-isopropoxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3.5-dihydro-4H-pyrrolo[3.2-d]|pyrimidin-
4-one,
(E )-7-((3-(2,3,4,5,6-pentafluorophenyl)methoxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3,5-dihydro- 4H-pyrrolo [3 ,2-d]pyrimidin-4-one,
(E )-7-((3-(2,3,4,5,6-pentafluorophenyl)oxy-2-(2-(phosphonato)vinyl)phenyl)thio)-3,5-dihydro-4H- pyrrolo [3 ,2-d]pyrimidin-4-one,
(E )-7-((5 -fluoro-2-((phosphonato)methoxy)phenyl)thio)-3,5 -dihydro-4H-pyrrolo [3 ,2-d]pyrimidm-4- one,
(E)-7-((3-methoxy-2-((phosphonato)methoxy)phenyl)thio)-3.5-dihydro-4H-pyrrolo[3.2-d]|pyrimidin-4- one,
(E )- 7 -((3 -(2-(phosphonato)vinyl)thiophene-2-yl)thio)-3.5-dihydro-4H-pyrrolo [3.2-d|pvrimidin-4-onc.
7-((3-((phosphonato)methoxy)thiophene-2-yl)thio)-3.5-dihydro-4H-pyrrolo[3.2-d]|pyrimidin-4-onc.
7-((2-((sulfo)methoxy)phenyl)thio)-3.5-dihydro-4H-pyrrolo[3.2-d]|pyrimidin-4-onc.
(E )-7 -((2-(2-(sulfo)vinyl)phenyl)thio)-3 ,5 -dihydro-4H-pyrrolo [3 ,2-d]pyrimidin-4-one, 7-((2-(3-phosphonato-2-oxapropyl)phenyl)thio)-3.5-dihydro-4H-pyrrolo[3.2-d]|pyrimidin-4-onc. 7-((2-(((((S)- 1 -(isopropoxycarbonyl)cthyl)amino)(phcnoxy)phosphoryl)methoxy)phenyl)thio)-3.5- dihydro-4H-pyrrolo[3.2-d]|pyrimidine-4-on. (E )-7 -((2-(2-(phosphonato)vinyl)phenyl)amino)-3.5-dihydro-4H-pyrrolo [3 ,2-d]pyrimidine-4-one .
4. Compound of the general formula I according to any one of claims 1 to 3 for use as medicaments.
5. Compound of the general formula I according to any one of claims 1 to 3 for use as medicaments for the treatment of diseases caused by abnormal growth of T-lymphocytes.
6. Compound of the general formula I according to any one of claims 1 to 3 for use as medicaments for the treatment of T-cell acute lymphoblastic leukemia and/or lymphoma.
7. A pharmaceutical composition comprising at least one compound of the general formula I according to any one of claims 1 to 3 as an active pharmaceutical ingredient and optionally at least one pharmaceutically acceptable carrier and/or filler and/or diluent and/or adjuvant.
8. The pharmaceutical composition according to claim 7 for use in the treatment of diseases caused by abnormal growth of T-lymphocytes.
9. The pharmaceutical composition according to claim 7 for use in the treatment of T-cell acute lymphoblastic leukemia and/or lymphoma.
10. A method of treatment of T-cell acute lymphoblastic leukemia and/or lymphoma, comprising the step of administering a therapeutically effective amount of at least one compound of the general formula I according to any one of claims 1 to 3.
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