EP3515923A1

EP3515923A1 - Prodrugs of fluorinated acyclic nucleoside phosphonates

Info

Publication number: EP3515923A1
Application number: EP17791943.8A
Authority: EP
Inventors: Graciela Andrei; Steven De Jonghe; Elisabetta GROAZ; Piet Herdewijn; Min Luo; Dominique Schols; Robert Snoeck
Original assignee: Katholieke Universiteit Leuven
Current assignee: Katholieke Universiteit Leuven
Priority date: 2016-09-23
Filing date: 2017-09-22
Publication date: 2019-07-31
Also published as: CN108137630A; WO2018055071A1

Abstract

The present invention relates to novel phosphonoamidate prodrugs of acyclic nucleoside phosphonates with a 3-fluoro-2-(phosphonomethoxy)propyl side chain. The invention also relates to the use of these novel phosphonate-modified nucleosides to treat or prevent viral infections and their use to manufacture a medicine to treat or prevent viral infections, particularly infections with viruses such as the hepatitis B virus, the human immunodeficiency virus, the human cytomegalovirus and the varicella zoster virus.

Description

Prodrugs of fluorinated acyclic nucleoside phosphonates

FIELD OF THE INVENTION

BACKGROUND OF THE INVENTION

Acyclic nucleoside phosphonates (ANPs) are a key class of antiviral drugs. Their main structural features are : (1 ) an aliphatic side chain instead of the cyclic sugar moiety and (2) the presence of a phosphonate group, attached to this acyclic nucleoside moiety.

The phosphonomethoxy functionality (P-C-O), as present in the ANPs, can be considered as an isoster of the naturally occurring phospho-oxymethyl (P-O-C) moiety in nucleosidemonophosphate. In contrast to the phosphate group, a phosphonate is not susceptible to phosphodiesterase and phosphatase hydrolysis and hence is enzymatically stable. The success of this motif is due to the fact that it is isopolar and isosteric with the phosphate group. Hence, they can undergo enzymatic phosphorylation that converts them into the corresponding phosphonate diphosphates, which act as analogues of the natural nucleoside triphosphates. As this phosphonate moiety is a phosphate mimicking group, ANPs only need two (instead of three for regular nucleoside analogues) phosphorylation steps to reach their biologically active stage. This is advantageous as the first phosphorylation is very often inefficient and rate-limiting in the formation of nucleoside-triphosphate. Flexibility in the acyclic sugar side chain allows these compounds, once dephosphorylated to adopt a conformation appropriate for interaction with the active sites of DNA polymerase or reverse transcriptase.

Extensive research on the synthesis and antiviral evaluation of ANPs led to marketing of three ANPs (Figure 1 ). Cidofovir (HPMPC, (S)-1 -(3-hydroxy-2-phosphonylmethoxypropyl)cytosine) received marketing approval for the treatment of CMV retinitis in AIDS patients. Adefovir (PMEA, 9-(2-phosphonylmethoxyethyl)adenine) has been licensed for the treatment of hepatitis B infected patients, and tenofovir (PMPA, (R)-9-{2- phosphonylmethoxypropyl)adenine) is being used as anti-HIV and anti-HBV agent. The latter two are marketed as an orally bioavailable prodrug form.

l An introduction of the fluorine atom into biologically active molecules often leads to remarkable changes in their physical, biological, and pharmacokinetic properties. ANPs with a fluorinated side chain have been synthesized and tested for antiviral activity. The best studied class of fluorinated ANPs are the 3-fluoro-2-(phosphonomethoxy)propyl (FPMP) derivatives. Both enantiomers of FPMPA and FPMPG (Figure 2) are completely devoid of antiviral activity against a broad range of DNA viruses (HSV-1 , HSV-2, CMV, VZV, VV). On the other hand, (S)-FPMPA shows potent and selective antiviral activity against HIV-1 and HIV-2, with EC50 values in the 8 μΜ range. The corresponding (R)-FPMPA completely lacks anti-HIV activity (Antimicrob. Agents Chemother. 1993, 37, 332-338). For the guanosine containing congeners (FPMPG), both enantiomers are equally active against HIV-1 and HIV-2, with EC50 values for both isomers in the 5 μΜ range. Also, nucleobase modified FPMP derivatives have been synthesized and evaluated for antiviral activity. The diaminopurine congener (FPMPDAP) shows no activity against a panel of DNA viruses, whereas (R)-FPMPDAP displays an EC50 value of 4.3 μΜ and 4.6 μΜ, against HIV-1 and HIV-2, respectively.

Phosphonates are negatively charged at physiological pH and hence, are not able to penetrate the lipid-rich cell membrane, which hampers their antiviral activity. Various prodrug or 'pronucleotide' approaches have been investigated to promote passive diffusion through the lipophilic cell membranes and to liberate the parent nucleotide intracellular^, where it can be further phosphorylated to the pharmacologically active species.

The present invention is based on the unexpected finding that phosphoramidate prodrugs of fluorinated ANPs with a 3-fluoro-2-(phosphonomethoxy)propyl (FPMP) side chain show unexpected biological properties, in particular have significant antiviral activity against the hepatitis B virus, the human immunodeficiency virus, the human cytomegalovirus and the varicella zoster virus.

SUMMARY OF THE INVENTION

The present invention relates to novel phosphoramidate and phosphorodiamidate prodrugs of acyclic nucleoside phosphonates (ANPs), and their use as agents for treating viral diseases. It is based on the unexpected finding that certain nucleoside prodrugs show unexpected biological properties, in particular have significant activity against the hepatitis B virus, the human immunodeficiency virus, the human cytomegalovirus and the varicella zoster virus.

Numbered statements of the invention are:

1 . A compound of formula I:

I

B is any natural or modified nucleobase

R¹ has the general formula I I

II

wherein

- R³ is selected from the group consisting of aryl, heteroaryl, C1-C10 alkyl, Cs-Cs-cycloalkyl, C3- Cs-cycloalkyl-alkyl, aryl(Ci-C6)alkyl, C2-C10 alkenyl, C2-C10 alkynyl, hydroxyl C1-C10 alkyl, halo C1-C10 alkyl, and alkoxyalkyl;

- R⁴ is selected from the group consisting of aryl, heteroaryl, C1-C10 alkyl, Cs-Cs-cycloalkyl, C3- Cscycloalkyl-alkyl, aryl(Ci-C6)alkyl, C2-C10 alkenyl, C2-C10 alkynyl, hydroxyl C1-C10 alkyl, halo C1-C10 alkyl, alkoxyalkyl, X-COOR⁵, X-0(C=0)-R⁵;

wherein X is aryl, heteroaryl, C1-C10 alkyl, C2-C10 alkenyl, C2-C10 alkynyl, or Cs-Cs-cycloalkyl, and wherein said aryl, heteroaryl, C1-C10 alkyl, C2-C10 alkenyl, C2-C10 alkynyl, Cs-Cs-cycloalkyl optionally contains one or more functions, atoms or radicals independently selected from the group consisting of halogen, halo-alkyl, cyano, C1-C7 alkoxy; and

wherein R⁵ is selected from the group consisting of aryl, heteroaryl, C1-C10 alkyl, Cs-Cs- cycloalkyl, C3-Cscycloalkyl-alkyl, aryl(Ci-C6)alkyl, C2-C10 alkenyl, C2-C10 alkynyl, hydroxyl Ci- C10 alkyl, halo C1-C10 alkyl, and alkoxyalkyl;

R² is O-Ar, wherein Ar is a fused bicyclic aryl moiety or a monocyclic aryl moiety, either of which aryl moieties is carbocyclic or heterocyclic and is optionally substituted with a halogen, C1-C6 alkyl, C1-C6 alkoxy;

or R² has the general formula I I wherein R¹ and R² can be identical or different; and/or a pharmaceutical acceptable addition salt thereof and/or a stereoisomer thereof and/or a solvate thereof.

2. The compound according to statement 1 , wherein B is selected from the group of adenine, thymine, cytosine and guanine.

3. The compound according to statement 1 or 2, wherein R² is O-Ph.

4. The compound according to any one of statements 1 to 3, wherein R³ is selected from C1-C10 alkyl.

5. The compound according to any one of statements 1 to 4, wherein R⁴ is X-COOR⁵ and wherein X is aryl, heteroaryl, C1-C10 alkyl, C2-C10 alkenyl, C2-C10 alkynyl, or Cs-Cs-cycloalkyl, and wherein said aryl, heteroaryl, C1-C10 alkyl, C2-C10 alkenyl, C2-C10 alkynyl, Cs-Cs-cycloalkyl optionally contains one or more functions, atoms or radicals independently selected from the group consisting of halogen, halo-alkyl, cyano, C1-C7 alkoxy; and wherein R⁵ is selected from the group consisting of aryl, heteroaryl, C1-C10 alkyl, Cs-Cs-cycloalkyl, C3-Cscycloalkyl-alkyl, aryl(Ci-C6)alkyl, C2-C10 alkenyl, C2-C10 alkynyl, hydroxyl C1-C10 alkyl, halo C1-C10 alkyl, and alkoxyalkyl.

6. The compound according to any one of statements 1 to 5, wherein X is C1-C10 alkyl and R⁵ is C1-C10 alkyl.

7. The compound according to any of statements 1 to 6, wherein R² is O-Ph, and wherei ¹ is

8. A compound selected from the group consisting of : (S)-1 -{3-Fluoro-2-[phenyloxy- bis(amyl-L-aspartyl)phosphoryl]methoxy]propyl}thymine; (R)-1 -{3-Fluoro-2-[phenyloxy- bis(amyl-L-aspartyl)phosphoryl]methoxy]propyl}thymine; (S)-9-{3-Fluoro-2-[phenyloxy- bis(amyl-L-aspartyl)phosphoryl]methoxy]propyl}adenine; (R)-9-{3-Fluoro-2-[phenyloxy- bis(amyl-L-aspartyl)phosphoryl]methoxy]propyl}adenine; (S)-1 -{3-Fluoro-2-[phenyloxy- bis(amyl-L-aspartyl)phosphoryl]methoxy]propyl}cytosine; (R)-1 -{3-Fluoro-2-[phenyloxy- bis(amyl-L-aspartyl)phosphoryl]methoxy]propyl}cytosine; (S)-9-{3-Fluoro-2-[phenyloxy- bis(amyl-L-aspartyl)phosphoryl]methoxy]propyl}guanine; (R)-9-{3-Fluoro-2-[phenyloxy- bis(amyl-L-aspartyl)phosphoryl]methoxy]propyl}guanine; (S)-0²-{3-Fluoro-2-[phenyloxy- bis(amyl-L-aspartyl)phosphoryl]methoxy]propyl}cytosine; (R)-0²-{3-Fluoro-2-[phenyloxy- bis(amyl-L-aspartyl)phosphoryl]methoxy]propyl}cytosine; Diamyl (((((R)-1 -(2-amino-4-oxo- 3,4-dihydro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-3-fluoropropan-2- yl)oxy)methyl)(phenoxy)phosphoryl)-L-aspartate; Tetraamyl 2,2'-((((((R)-1 -(2-amino-4-oxo-

3,4-dihydro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-3-fluoropropan-2- yl)oxy)methyl)phosphoryl)bis(azanediyl))(2S,2'S)-disuccinate.

9. A compound according to any one of statements 1 to 8 for use as a medicine.

10. A compound according to any one of statements 1 to 8 for use as a medicine for the prevention or treatment of a viral infection in an animal, mammal or human.

1 1 . The compound according to statement 10, wherein said viral infection is an infection with the hepatitis B virus (HBV), the human immunodeficiency virus (HIV), varicella-zoster virus (VZV), cytomegalovirus (CMV), vaccinia virus (VV), herpes simplex virus (HSV), BK virus, Epstein-barr virus (EBV), papillomavirus, Monkeypox virus, Cowpox virus, hepatitis C virus (HCV), respiratory syncytial virus (RSV), dengue virus, influenza virus, adenovirus, parainfluenza virus and/or rhinovirus.

12. A compound according to any one of statements 1 to 8 for use as a medicine for the prevention or treatment of a proliferative disorder such as cancer in an animal, mammal or human.

13. A pharmaceutical composition comprising a therapeutically effective amount of a compound according to any one of statements 1 to 8 and one or more pharmaceutically acceptable excipients.

14. The pharmaceutical composition according to statement 13, further comprising one or more biologically active drugs being selected from the group consisting of antiviral drugs and/or anti-proliferative drugs.

15. A method of prevention or treatment of a viral infection in an animal, mammal or human, comprising the administration of a therapeutically effective amount of a compound according to any one of statements 1 to 8, optionally in combination with one or more pharmaceutically acceptable excipients.

16. A method of prevention or treatment of a proliferative disorder in an animal, mammal or human, comprising the administration of a therapeutically effective amount of a compound according to any one of statements 1 to 8, optionally in combination with one or more pharmaceutically acceptable excipients.

The present invention will now be further described. In the following passages, different aspects of the invention are defined in more detail. Each aspect so defined may be combined with any other aspect or aspects unless clearly indicated to the contrary. In particular, any feature indicated as being preferred or advantageous may be combined with any other feature or features indicated as being preferred or advantageous.

DETAILED DESCRIPTION OF THE INVENTION

Scheme 1 schematically shows a method for the synthesis of the acyclic fluorinated phosphonate esters 4a/b that are the key building blocks for the preparation of ANPs bearing a 3-fluoro-2-(phosphonomethoxy)propyl side chain. They were prepared according to a modified literature report (Baszczynski, O.; Jansa, P.; Dracinsky, M.; Klepetarova, B.; Holy, A.; Votruba, I.; Clerck, E, D.; Balzarini, J.; Janeba, Z. Bioorg. Med. Chem. 2011 , 19, 21 14-2124), starting from commercially available enantiomeric O-tritylated glycidols 1 a/b, and using diethyl tosyloxymethylphosphonate in place of diisopropyl tosyloxymethylphosphonate 1.

( ) 1a (S) 2a (S) 3a (42% over two steps) (S) 4a (60%)

(S) 1b (R) 2b (R) 3b (46% over two steps) (R) 4b (62%)

Reagents and conditions: (a) KHF₂, PhCI, cat. BU4NH2F3, 135 °C, 15 h; (b) diethyl tosyloxymethylphosphonate, NaH, THF, 0 °C to rt, 5 h; (c) 80% AcOH, 90 °C, 1 h.

Scheme 1 : Synthesis of (S)/(R)-fluorinated phosphonate esters 4a/b.

Scheme 2 shows a method for the synthesis of 3-fluoro-2-(phosphonomethoxy)propyl ANP with thymidine as nucleobase. Condensation of 4a/b with A^-Bom-thymine delivered compounds 6a/b in high yields. Removal of the A^-Bom group by catalytic hydrogenation using 10% palladium on carbon provided compounds 7a/b, which was followed by hydrolysis of the diester groups with TMSBr in dry acetonitrile overnight to form the corresponding phosphonate acids 8a/b.

(S) 4a (S) 6a (80%)

(R) (R) 6b (70%)

(S) 7a (90%) (S) 8a (70%)

(R) 7b (90%) (R) 8b (70%)

Reagents and conditions: (a) Ph₃P, DIAD, THF, rt, 12 h; (b) Pd/C, H₂, EtOH, rt, 24 h; (c) TMSBr, 2,6-lutidine, CH₃CN, rt, 12 h.

Scheme 2 : Synthesis of (S)/(R)-acyclic fluorinated thymine phosphonate acids 8a/8b

Scheme 3 shows a method for the synthesis of 3-fluoro-2-(phosphonomethoxy)propyl ANPs with adenine as nucleobase. Mitsunobu reaction in the presence of 6-chloropurine proceeded smoothly affording compounds 10a/b in 60% and 65% yield, respectively. The chloropurine moiety was then converted into adenine by treatment with NH3 in EtOH under pressure, and finally the phosphonate esters were cleaved with TMSBr to provide compounds 12a/b in 65 and 70% yields, respectively.

(S) 11a (80%) (S) 12a (65%)

(R) 11b (90%) (R) 12b (70%)

Reagents and conditions: (a) Ph₃P, DIAD, THF, rt, 12 h; (b) NH₃/EtOH, 50 °C, 24 h; (c) TMSBr, 2,6-lutidine, CH₃CN, rt, 12 h. Scheme 3 : Synthesis of (S)/(R)-acyclic fluorinated adenine phosphonate acids 12a/12b.

Scheme 4 shows a method for the synthesis of 3-fluoro-2-(phosphonomethoxy)propyl ANP with cytosine as nucleobase. Moderate yields were achieved with /V sobutyrylcytosine along with the unexpected formation of O-2-alkylated products 15a/b (ratio 1 :1 , 0-2:/V-1 alkylation). After the removal of the /V sobutyryl protecting groups in ΝΗβ/ΜβΟΗ and cleavage of the phosp

(S) 16a (90%) (S) 17a (70%)

(R) 16b (82%) (R) 17b (70%)

(S) 15a (S) 18a (82%) (S) 19a (70%) (R) 15b (R) 18b (90%) (R) 19b (70%)

Reagents and conditions: (a) Ph₃P, DIAD, THF, rt, 12 h; (b) NH₃/MeOH, 45 °C, 15 h; (c) TMSBr, 2,6-lutidine, CH₃CN, rt, 12 h.

Scheme 4 : Synthesis of (S)/(R)-acyclic fluorinated cytosine and 0²-cytosine phosphonate acids 17a/b and 19a/b.

Scheme 5 shows a method for the synthesis of 3-fluoro-2-(phosphonomethoxy)propyl ANPs with guanosine as nucleobase. The reaction between 6-O-benzylguanine 20 and compounds 4a/b gave a mixture of compounds 21 a/b and the corresponding triphenylphosphine adducts (not shown). This crude mixture residue was then refluxed for several hours in THF/H2O to remove the undesired triphenylphosphine adducts. Sequential removal of the 6-O-benzyl protecting group using 10% palladium on carbon and treatment with TMSBr provided the desired products.

(S) 4a (S) 21a (50%)

20

(R) 4b (R) 21 b (44%)

(S) 22a (70%) (S) 23a (60%)

(R) 22b (80%) (R) 23b (60%)

Reagents and conditions: (a) Ph₃P, DIAD, THF, rt, 12 h; (b) Pd/C, H₂, EtOAc, 10 h.; (c) TMSBr, 2,6-lutidine, CH₃CN, rt, 12 h.

Scheme 5: Synthesis of (S)/(R)-acyclic fluorinated guanine phosphonate acids 23a/b.

Schemes 6 and 7 show methods to convert the free phosphonic acids into the corresponding aryloxyphosphonamidates. Phosphonamidates 24a/b-27a/b were successfully obtained in the presence of triethylamine, 2,2'-dithiodipyridine, and triphenylphosphine in pyridine overnight. However, for the synthesis of guanine phosphonamidates 29a/b, only the unexpected triphenylphosphine adducts 28a/b were formed (Scheme 7). In the case of adenine and cytosine, only minor quantities of triphenylphosphine adducts were formed. These triphenylphosphine adducts could be successfully cleaved by treatment with 1 M HCI at room temperature.

Reagents and conditions: (a) L-Aspartic acid amyl diester HCI salt, PhOH, 2,2'-dithiodipyridine, PPh₃, Et₃N, Pyr, 60 °C, 12 h.

Scheme 6: Synthesis of (S)/(R)-acyclic fluorinated nucleoside phosphonamidates 24a/b- 27a/b.

(S) 29a (5% over two steps)

(R) 29b (4% over two steps)

Reagents and conditions: (a) L-Aspartic acid amyl diester HCI salt, PhOH, 2,2'-dithiodipyridine, PPh₃, Et₃N, Pyr, 60 °C, 12 h; (b) 1 M HCI, 0 °C to rt, 1 h.

Scheme 7 : Synthesis of (S)/(R)-acyclic fluorinated guanine phosphonamidates 29a/b.

Scheme 8 shows a method for the synthesis of 3-fluoro-2-(phosphonomethoxy)propyl ANPs with 2-amino-6-(4-methoxy-thiophenol)purine as nucleobase. Under Mitsunobu reaction conditions (PhsP, DIAD), 2-amino-6-chloropurine 30 condensed with precursors 4a/b to provide compounds 31 a/b with the concomitant formation of the corresponding triphenylphosphine adducts. By refluxing the crude reaction mixtures for 24 h, these adducts were decomposed successfully. Heating of compounds 31 a/b with 4-methoxythiophenol in presence of triethylamine in DMF furnished 32a/b in 87% and 67% yield, respectively. Lewis- acid mediated deprotection of the phosphonate esters furnished compounds 33a/b.

(R) 32a (87%) (R) 33a (70%)

(S) 32b (67%) (S) 33b (65%)

Reagents and conditions: (a) (i) Ph₃P, DIAD, THF, rt, 24 h; (ii) THF/H₂0, reflux, 24 h; (b) Et₃N, DMF, 100 °C, 4 h; (c) TMSBr, 2,6-lutidine, CH₃CN, rt, 12 h.

Scheme 8 : Synthesis of (R)/(S)-fluorinated acyclic phosphonates 33a/b.

Scheme 9 shows a method for the synthesis of 3-fluoro-2-(phosphonomethoxy)propyl ANPs with 7-deaza-guanine and 7-deaza-7-fluoro-guanine as nucleobase. The preparation of 4- chloro-5-fluoro-2-pivaloylamino-7H-pyrrolo[2,3-d]pyrimidine 35 started from commercially available 30. Selectfluor was chosen as a fluorinating reagent following a literature protocol (Seela, F.; Xu, K.; Chittepu, P. Fluorinated pyrrolo[2,3-d]pyrimidine nucleosides: 7-fluoro-7- deazapurine 2'-deoxyribofuranosides and 2'-deoxy-2'-fluoroarabinofuranosyl derivatives. Synthesis 2006, 12, 2005-2012). The phosphonate diethyl esters 36a/b and 37a/b were prepared using the standard Mitsunobu conditions (Ph₃P, DIAD). Sequential replacement of chloro with hydroxyl and deprotection of pivaloyl group was achieved by DABCO/K2C0₃ and NH₃/MeOH, respectively, affording compounds 38a/b and 39a/b. Hydrolysis of the phosphonate esters groups yielded compounds 40a/b and 41a/b.

30 34 (87%) 35 (30%)

(S) 39b (35% over 2 steps) R = F

(R) 40a (70%)

H

(S) 40b (70%)

(R) 41a (50%) _R

(S) 41b (60%)

Reagents and conditions: (a) PivCI, Pyr, rt, 3 h; (b) Selectfluor, MeCN, 50 °C, 30 min.

(c) Ph₃P, DIAD, THF, rt, 24 h; (d) (i) DABCO, K₂C0₃, dioxane/H₂0, 90 °C, 3 h; (ii) NH₃/MeOH, rt, 12 h; (e) TMSBr, 2,6-lutidine, CH₃CN, rt, 12 h.

Scheme 9 : Synthesis of (R)/(S)-fluorinated acyclic phosphonates 40a/b and 41a/b.

Scheme 10 shows a method for the synthesis of 3-fluoro-2-(phosphonomethoxy)propyl ANPs with 7-deaza-7-cyano-guanine as nucleobase. Synthesis of the heterocyclic building block 46 (Llona-Minguez, S.; Mackay, S. P. Stereoselective synthesis of carbocyclic analogues of the nucleoside Q precursor (PreQO). Beilstein J. Org. Chem. 2014, 10, 1333-1338) started by formylation of chloroacetonitrile 42 with methyl formate. The resulting unstable chloroaldehyde 43 was used for the next step immediately without further purification to afford cyclocondensation compound 44. Then, the exocyclic amine was protected as a pivaloyl group, and the subsequent chlorination step was accomplished using POC in the presence of a phase transfer catalyst to furnish the desired chloro-intermediate 46. By treatment with Ph₃P and DIAD, alkylation of 4a/b with 46 were successfully performed, affording compounds 47a/b. Since only low yields were obtained for 48a using DABCO/K2C0₃ for converting the 4-chloro to the 4-oxo, an alternative method by treatment with DABCO/NaOAc was applied on 47b (Brijckl, T; Klepper, R; Gutsmiedl, K.; Carell, T. A short and efficient synthesis of the tRNA nucleosides PreQO and archaeosine. Org. Biomol. Chem. 2007, 5, 3821 -3825). As such, the desired product 48b was isolated in good yield (60% over 2 steps). Finally, cleavage of the phosphonate esters afforded compounds 49a/b.

46 (65%) 45 (60%)

(R) 49a (70%) (R) 48a (30% over 2 steps)

(S) 49b (75%) (S) 48b (60% over 2 steps)

Reagents and conditions: (a) Methyl formate, NaH, THF, 0 °C, 5 h; (b) 2,6-diamino-6- hydroxyprimidine, NaOAc, H₂0, 100 °C, 24 h; (c) PivCI, Pyr, 85 °C, 2 h; (d) POCI₃, DMA, BnEtsNCI, MeCN, 90 °C, 1 h. (e) Ph₃P, DIAD, THF, rt, 24 h; (f) (i) DABCO, K₂C0₃, dioxane/H₂0, 90 °C, 3 h or DABCO, NaOAc, DMF, rt, 48 h; (ii) NH₃/MeOH, rt, 12 h; (g) TMSBr, 2,6-lutidine, CHsCN, rt, 12 h.

Scheme 10 : Synthesis of (R)/(S)-fluorinated acyclic phosphonates 49a/b.

Scheme 11 shows a method for synthesizing an aryloxyphosphonoamidate prodrug 50 and a phosphonobisamidate prodrug 51 , from the corresponding free phosphonic acid 40a. Compound 51 was prepared from the parent nucleoside phosphonate 40a using 2,2'- dithiodipyridine and triphenylphosphine as activating agents and utilizing a mixture of phenol and the appropriate amino acid ester. The phosphonobisamidate prodrug 51 was prepared in a similar way, using the desired amino acid ester.

(^R) ^0a 50: R† = Asp-amyl, R₂ = OPh (40%)

51 : R-i = R₂ = Asp-amyl (20%)

Reagents and conditions: (a) L-Aspartic acid amyl diester HCI salt, PhOH, 2,2'-dithiodipyridine, PPh₃, Et₃N, Pyr, 60 °C, 12 h for 50; L-Aspartic acid amyl diester HCI salt, 2,2'-dithiodipyridine, PPh₃, Et₃N, Pyr, 60 °C, 12 h for 51.

Scheme 11 : Synthesis of (R)-fluorinated acyclic phosphonamidates 50 and 51

According to one embodiment, the present invention encompasses compounds of the general formula I :

B is any natural or modified nucleobase

R¹ has the general formula II

II

wherein

wherein X is aryl, heteroaryl, C1-C10 alkyl, C2-C10 alkenyl, C2-C10 alkynyl, or Cs-Cs-cycloalkyl, and wherein said aryl, heteroaryl, C1-C10 alkyl, C2-C10 alkenyl, C2-C10 alkynyl, Cs-Cs-cycloalkyl optionally contains one or more functions, atoms or radicals independently selected from the group consisting of halogen, halo-alkyl, cyano, C1-C7 alkoxy; and wherein R⁵ is selected from the group consisting of aryl, heteroaryl, C1-C10 alkyl, C3-C8- cycloalkyl, Cs-Cscycloalkyl-alkyl, aryl(Ci-C6)alkyl, C2-C10 alkenyl, C2-C10 alkynyl, hydroxyl Ci- C10 alkyl, halo C1-C10 alkyl, and alkoxyalkyl;

or R² has the general formula II

wherein R¹ and R² can be identical or different;

and/or a pharmaceutical acceptable addition salt thereof and/or a stereoisomer thereof and/or a solvate thereof.

Said base (B) is selected from the group of the pyrimidine and purine bases. Such bases include natural bases, such as adenine, thymine, cytosine, uracyl, guanine and modified bases or modifications of said natural bases. In certain embodiments of the present invention said base is a guanine, cytosine, adenine, thymine, cytosine, or uracyl. In a more specific embodiment of the present invention, said base is a adenine or guanine. In another specific embodiment of the present invention said base is a cytosine. In another specific embodiment of the present invention said base is a thymine. In another specific embodiment of the present invention said base is uracil. In certain embodiments of the present invention said base is attached to the rest of the compound via its nitrogen atom, such as in examples 31 , 32, 33, 34, 35, 36, 39, 40, 67 and 68. In other embodiments of the present invention said base is attached to the rest of the compound via an oxygen atom, such as in examples 37 and 38. In more specific embodiments thereof, said base (attached to the rest of formula I via its oxygen atom) is selected from the list consisting of:

wherein R is selected from the group consisting of H , halogen, and methyl.

In a more specific embodiment thereof, said base is

id embodiment encompasses compounds of formula lb :

wherein R¹ and R² can have any values as described herein.

In another embodiment, the present invention concerns a compound according to the invention, including the compound of formula I, lb, or any subgroup thereof, wherein Ar is a fused bicyclic aryl moiety or a monocyclic aryl moiety, either of which aryl moieties is carbocyclic or heterocyclic and is optionally substituted with a halogen, C1-C6 alkyl, C1-C6 alkoxy. In a more specific embodiment said Ar is phenyl. In a specific embodiment of the present invention, the compound of formula I, lb, or any subgroup thereof, can have any value for R¹ as described herein, wherein Ar is phenyl.

In a more specific embodiment said R³ is C1-C10 alkyl. In another specific embodiment said R³ is C3-C10 alkyl. In another specific embodiment said R³ is C1-C5 alkyl. In yet another specific embodiment said R³ is C3-C5 alkyl. In yet another specific embodiment said R³ is C5 alkyl.

In another specific embodiment, said R⁴ is selected from the group consisting of C1-C10 alkyl or X-COOR⁵, wherein R⁵ can have any values as described herein. In a more specific embodiment, said R⁵ is selected from the group consisting of aryl, heteroaryl, C1-C10 alkyl, C3- Cs-cycloalkyl, Cs-Cscycloalkyl-alkyl, aryl(Ci-C6)alkyl, C2-C10 alkenyl, C2-C10 alkynyl, hydroxyl C1-C10 alkyl, halo C1-C10 alkyl, and alkoxyalkyl. In a more specific embodiment R⁵ is C1-C7 alkyl or C3-C8 cycloalkyi; in a more specific embodiment R⁵ is C1-C5 alkyl, and in another more specific embodiment R⁵ is C3-C7 alkyl, in an even more specific embodiment R⁵ is C3-C5 alkyl. In a yet more specific embodiment R⁵ is C5 alkyl. In another specific embodiment, R⁵ is aryl- (Ci-C2)alkyl; in another more specific embodiment, R⁵ is benzyl or phenyl-methyl.

In another specific embodiment, X is selected from the group consisting of aryl, heteroaryl, C1-C10 alkyl, C2-C10 alkenyl, C2-C10 alkynyl or Cs-Cs-cycloalkyl, and wherein said aryl, heteroaryl, C1-C10 alkyl, and Cs-Cs-cycloalkyl optionally contains one or more functions, atoms or radicals independently selected from the group consisting of halogen, carbonyl, thiocarbonyl, hydroxyl, thiol, ether, thio-ether, acetal, thio-acetal, amino, imino, oximino, alkyloximino, aminoacid, cyano, acylamino, thioacylamino, carbamoyl, thiocarbamoyl, ureido, thio-ureido, carboxylic acid ester or halide or anhydride or amide, thiocarboxylic acid or ester or thioester or halide or anhydride or amide, nitro, thio C1-7 alkyl, thio C3-C10 cycloalkyl, hydroxylamino, mercaptoamino, alkyl-amino, cycloalkylamino, alkenylamino, cycloalkenylamino, alkynylamino, arylamino, arylalkylamino, hydroxyalkylamino, mercaptoalkylamino, heterocyclic-substituted alkylamino, hetero-cyclic amino, heterocyclic- substituted arylamino, hydrazine, alkylhydrazino, phenylhydrazino, sulfonyl, sulfinyl and sulfonamide. In a more specific embodiment, X is selected from the group consisting of aryl, heteroaryl, C1-C10 alkyl, or Cs-Cs-cycloalkyl, more specifically said X is a C1-C6 alkyl, even more specifically said X is a C1-C3 alkyl or C1-C2 alkyl or -CH2-.

Special novel compounds in accordance with the present invention include each of the compounds whose preparation is described in the accompanying Examples, and pharmaceutically acceptable salts and solvates thereof.

The present invention also concerns a compound having formula I , lb, or any subgroup thereof, or stereoisomeric forms thereof, for use as a medicine.

The present invention also concerns a compound having formula I, lb, or any subgroup thereof, or stereoisomeric forms thereof, for use as a medicine for the prevention or treatment of viral disorders and oncological disorders in an animal, preferably in a mammal. In an embodiment, said disorder is a viral disorder, including a disease caused by a viral infection, for example an infection with HBV, HIV, HCV, RSV, dengue virus, influenza virus, VZV, CMV, adenovirus, parainfluenza, rhinovirus, BK virus, and/or HSV; in another embodiment said disorder is an oncological disorder, which may be acute or chronic, including a proliferative disorder, especially cancer. In an embodiment, said mammal is a human being. In a specific embodiment, said compounds for use as a medicine, in particular for the prevention or treatment of viral disorders, are the compounds of formula I and lb, including any subgroups thereof. In a more specific embodiment thereof, said compounds of formula I have as a base B a pyrimidine base, more specifically a pyrimidine base represented by the structural formula

(IV):

IV or a purine base, more specifically a purine base represented by the structural formula (V):

wherein:

R⁷ and R⁹ are independently selected from the group consisting of H, -OH, -SH, -IMH2, and - NH-Me;

R⁸ and R¹⁰ are independently selected from the group consisting of H, methyl, ethyl, isopropyl, hydroxyl, amino, ethylamino, trifluoromethyl, cyano and halogen; and

X¹ and Y¹ are independently selected from CR¹¹ and N, wherein R¹¹ is selected from the group consisting of H, halogen and cyano.

The present invention also concerns the use of the compounds of formula I, lb, or any subgroup thereof, or stereoisomeric forms thereof, for the manufacture of a medicament for the prevention or treatment of a viral disorder and/or an oncological disorder in an animal. In an embodiment, said animal is a mammal, preferably said mammal is a human being.

In a further specific embodiment, said viral disorder is a disease caused by a viral infection, for example an infection with HBV, HIV, HCV, RSV, dengue virus, influenza virus, CMV, VZV, adenovirus, parainfluenza, rhinovirus, BK virus, and/or HSV. In an even more specific embodiment said viral disorder is a disease caused by a viral infection with HBV and/or HIV. In a specific embodiment, said viral disorder is a disease caused by a viral infection with HBV. In another specific embodiment, said viral disorder is a disease caused by a viral infection with HIV1 .

The present invention also concerns a pharmaceutical composition comprising a therapeutically effective amount of a compound having formula I, lb, or any subgroup thereof, or stereoisomeric forms thereof and one or more pharmaceutically acceptable excipients. Said composition may further comprise one or more biologically active drugs being selected from the group consisting of antiviral drugs, and antineoplastic drugs.

The present invention also concerns a method of prevention or treatment of a viral disorder in an animal, comprising the administration of a therapeutically effective amount of a compound having formula I, lb, or any subgroup thereof, or stereoisomeric forms thereof, optionally in combination with one or more pharmaceutically acceptable excipients.

The present invention also concerns a method of prevention or treatment of an oncological disorder in an animal, comprising the administration of a therapeutically effective amount of a compound having formula I, lb, or any subgroup thereof, or stereoisomeric forms thereof, optionally in combination with one or more pharmaceutically acceptable excipients.

For use in medicine, the salts of the compounds of formula I or lb, will be pharmaceutically acceptable salts. Other salts may, however, be useful in the preparation of the compounds of the invention or of their pharmaceutically acceptable salts. Suitable pharmaceutically acceptable salts of the compounds of this invention include acid addition salts which may, for example, be formed by mixing a solution of the compound of the invention with a solution of a pharmaceutically acceptable acid such as hydrochloric acid, sulphuric acid, methanesulphonic acid, fumaric acid, maleic acid, succinic acid, acetic acid, benzoic acid, citric acid, tartaric acid or phosphoric acid. Furthermore, where the compounds of the invention carry an acidic moiety, e.g. carboxy, suitable pharmaceutically acceptable salts thereof may include alkali metal salts, e.g. sodium or potassium salts; alkaline earth metal salts, e.g. calcium or magnesium salts; and salts formed with suitable organic ligands, e.g. quaternary ammonium salts.

The present invention includes within its scope solvates of the compounds of formula I and lb above. Such solvates may be formed with common organic solvents, e.g. hydrocarbon solvents such as benzene or toluene; chlorinated solvents such as chloroform or dichloromethane; alcoholic solvents such as methanol, ethanol or isopropanol; ethereal solvents such as diethyl ether or tetrahydrofuran; or ester solvents such as ethyl acetate. Alternatively, the solvates of the compounds of formula I and lb may be formed with water, in which case they will be hydrates.

The compounds in accordance with the present invention are beneficial in the treatment and/or prevention of various animal, mammal or human ailments or diseases. These include viral diseases, such as diseases caused by a viral infection, for example an infection with HBV, HIV, HCV, RSV, dengue virus, influenza virus, herpes simplex viruses 1 and 2 (HSV-1 and HSV-2), varicella-zoster virus (VZV), Epstein-Barr virus (EBV or HHV-4), human cytomegalovirus (HCMV or HHV-5), human herpesvirus 6A and 6B (HHV-6A and HHV-6B), human herpesvirus 7 (HHV-7), and Kaposi's sarcoma-associated herpesvirus (KSHV, also known as HHV-8), adenovirus, parainfluenza, rhinovirus, and/or BK virus; and oncological disorders such as proliferative disorders (eg. cancer).

Viral diseases include infections caused by various families of virus, including the Hepadnaviridae, Retroviridae, Herpesviridae, Papovaviridae, Papillomaviridae or pappilomaviruses, Flaviviridae, Picornaviridae. Various genera within the Hepadnaviridae include the genera Orthohepadnavirus and the Avihepadnavirus; Members of the Orthohepadnavirus genus include Hepatitis B virus (HBV) and the Woodchuck hepatitis virus. Members of the Avihepadnavirus genus include the Duck hepatitis B virus. Various genera within the Retroviridae family include Alpharetrovirus, Betaretrovirus, Gammaretrovirus, Deltaretrovirus, Epsilonretrovirus, Lentivirus and Spumavirus. Members of the Lentivirus genus include human immunodeficiency virus 1 (HIV-1 ) and human immunodeficiency virus 2 (HIV-2). Various genera within the Herpesviridae family include (i) within the subfamily of the Alphaherpesvirinae: Varicellovirus, Scutavirus, lltovirus, Mardivirus, Simplexvirus; (ii) within the subfamily of the Betaherpesvirinae: Cytomegalovirus, Muromegalovirus, Proboscivirus, Roseolovirus; and (iii) within the subfamily of the Gammaherpesvirinae: Lymphocryptovirus, Macavirus, Percavirus, Rhadinovirus. Members of the Varicellovirus genus include Varicella zoster virus (VZV); Simian varicella virus; Phocine herpesvirus 1 ; Suid herpesvirus 1 ; Feline herpesvirus 1 ; Equine herpesvirus 1 , 3, 4, 8 and 9; Cervine herpesvirus 1 , and 2; Cercopithecine herpesvirus 9; Caprine herpesvirus 1 ; Bovine herpesvirus 1 and 5; Bubaline herpesvirus 1 ; Canine herpesvirus 1 . Members of the Simplexvirus genus include Human herpesvirus 1 and 2. Members of the Scutavirus genus include Chelonid herpesvirus 5. Members of the lltovirus genus include Gallid herpesvirus 1 . Members of the Mardivirus genus include Gallid herpesvirus 2. Members of the Cytomegalovirus genus include Human cytomegalovirus (CMV). Members of the Proboscivirus genus include Elephantid herpesvirus 1 . Members of the Muromegalovirus genus include Murid herpesvirus 1 . Members of the Roseolovirus genus include Human herpesvirus 6A, 6B and 7. Members of the Lymphocryptovirus genus include Human herpesvirus 4. Members of the Macavirus genus include Alcelaphine herpesvirus 1 . Members of the Percavirus genus include Equid herpesvirus 2. Members of the Rhadinovirus genus include Saimiriine herpesvirus 2, Kaposi's sarcoma-associated virus. Various genera within the Flaviviridae family include Flavivirus, Pestivirus, Hepacivirus and Hepatitis G Virus. The Papovaviridae family include the genus Polyomavirus (e.g. JC virus; BK virus; Merkel cell polyomavirus; Trichodysplasia spinulosa polyomavirus; Human polyomavirus 6, 7, 9 and 12; New Jersey polyomavirus; Kl polyomavirus; WU polyomavirus; MW polyomavirus; STL polyomavirus). The Pappilomaviridae or pappilomaviruses include the human pappilomaviruses (HPV). Members of the Flavivirus genus include Dengue fever virus, yellow fever virus, West Nile encephalitis virus and Japanese encephalitis virus. Members of the Pestivirus genus include bovine viral diarrhoea virus (BVDV), classical swine fever virus and border disease virus 2 (BDV-2). Members of the Hepacivirus genus include hepatitis C virus (HCV). Members of the Hepatitis G Virus genus include hepatitis G virus. Various genera within the Picornaviridae family include Aphthovirus, Avihepatovirus, Cardiovirus, Enterovirus, Erbovirus, Hepatovirus, Kobuvirus, Parechovirus, Sapelovirus, Senecavirus, Teschovirus and Tremovirus. Members of the Enterovirus genus include poliovirus, coxsackie A virus, coxsackie B virus and rhinovirus.

Oncological disorders, which may be acute or chronic, include proliferative disorders, especially cancer, in animals, including mammals, especially humans. Particular categories of cancer include haematological malignancy (including leukaemia and lymphoma) and non- haematological malignancy (including solid tumour cancer, sarcoma, meningioma, glioblastoma multiforme, neuroblastoma, melanoma, gastric carcinoma and renal cell carcinoma). Chronic leukaemia may be myeloid or lymphoid. Varieties of leukaemia include lymphoblastic T cell leukaemia, chronic myelogenous leukaemia (CML), chronic lymphocytic/lymphoid leukaemia (CLL), hairy-cell leukaemia, acute lymphoblastic leukaemia (ALL), acute myelogenous leukaemia (AML), myelodysplastic syndrome, chronic neutrophilic leukaemia, acute lymphoblastic T cell leukaemia, plasmacytoma, immunoblastic large cell leukaemia, mantle cell leukaemia, multiple myeloma, acute megakaryoblastic leukaemia, acute megakaryocytic leukaemia, promyelocytic leukaemia and erythroleukaemia. Varieties of lymphoma include malignant lymphoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, lymphoblastic T cell lymphoma, Burkitt's lymphoma, follicular lymphoma, MALT1 lymphoma and marginal zone lymphoma. Varieties of non-haematological malignancy include cancer of the prostate, lung, breast, rectum, colon, lymph node, bladder, kidney, pancreas, liver, ovary, uterus, cervix, brain, skin, bone, stomach and muscle.

The present invention also provides a pharmaceutical composition which comprises a compound in accordance with the invention as described above, or a pharmaceutically acceptable salt or solvate thereof, in association with one or more pharmaceutically acceptable carriers.

Pharmaceutical compositions according to the invention may take a form suitable for oral, buccal, parenteral, nasal, topical, ophthalmic or rectal administration, or a form suitable for administration by inhalation or insufflation.

The quantity of a compound of use in the invention required for the prophylaxis or treatment of a particular condition or disease will vary depending on the compound chosen and the condition of the animal, mammal or human patient to be treated. In general, however, daily dosages may range from around 10 ng/kg to 1000 mg/kg, typically from 100 ng/kg to 100 mg/kg, e.g. around 0.01 mg/kg to 40 mg/kg body weight, for oral or buccal administration, from around 10 ng/kg to 50 mg/kg body weight for parenteral administration, and from around 0.05 mg to around 1000 mg, e.g. from around 0.5 mg to around 1000 mg, for nasal administration or administration by inhalation or insufflation. DEFINITIONS

The term "pyrimidine and purine bases" as used herein includes, but is not limited to, adenine, thymine, cytosine, uracyl, guanine, 2,6-diaminopurine, 5-fluorocytosine, 5- fluorouracil, 7-deazaguanosine, 7-deazaadenine and analogues thereof. A purine or pyrimidine base as used herein includes a purine or pyrimidine base found in naturally occurring nucleosides as mentioned above. An analogue thereof is a base which mimics such naturally occurring bases in such a way that their structures (the kinds of atoms and their arrangement) are similar to the naturally occurring bases but may either possess additional or lack certain of the functional properties of the naturally occurring bases. Such analogues include those derived by replacement of a CH moiety by a nitrogen atom (e.g. 5-azapyrimidines such as 5-azacytosine) or vice versa (e.g. 7-deazapurines, such as 7-deazaadenine or 7- deazaguanine) or both (e.g., 7-deaza, 8-azapurines). By derivatives of such bases or analogues are meant those bases wherein ring substituents are either incorporated, removed, or modified by conventional substituents known in the art, e.g. halogen, hydroxyl, amino, (Ci- Ce)alkyl and others. Such purine or pyrimidine bases, and analogues thereof, are well known to those skilled in the art, e.g. as shown at pages 20-38 of WO 03/093290.

In particular purine and pyrimidine analogues B for the purpose of the present invention may be selected from the group comprising pyrimidine bases represented by the structural formula (IV):

IV and purine bases represented by the structural formula (V):

wherein:

R⁷ and R⁹ are independently selected from the group consisting of H, -OH, -SH, -Nhb, and - NH-Me; R⁸ and R¹⁰ are independently selected from the group consisting of H, methyl, ethyl, isopropyl, hydroxyl, amino, ethylamino, trifluoromethyl, cyano and halogen; and

Just as a few non-limiting examples of pyrimidine analogues, can be named substituted uracils with the formula (IV) wherein X¹ is CH, R⁷ is hydroxyl, and R⁸ is selected from the group consisting of methyl, ethyl, isopropyl, amino, ethylamino, trifluoromethyl, cyano, fluoro, chloro, bromo and iodo.

The term "alkyl" as used herein refers to a straight (normal) or branched (eg. secondary, or tertiary) hydrocarbon chains having the number of carbon atoms as indicated (or where not indicated, preferably having 1 -20, more preferably 1 -6 carbon atoms). The term "C1-C6 alkyl" refers to such hydrocarbon chains having from 1 to 6 carbon atoms. Examples thereof are methyl, ethyl, 1 -propyl, 2-propyl, 1 -butyl, 2-methyl-1 -propyl(i-Bu), 2-butyl (s-Bu) 2-methyl-2- propyl (t-Bu), 1 -pentyl (n-pentyl), 2-pentyl, 3-pentyl, 2-methyl-2-butyl, 3-methyl-2-butyl, 3- methyl-1 -butyl, 2-methyl-1 -butyl, 1 -hexyl, 2-hexyl, 3-hexyl, 2-methyl-2-pentyl, 3-methyl-2- pentyl, 4-methyl-2-pentyl, 3-methyl-3-pentyl, 2-methyl-3-pentyl, 2,3-dimethyl-2-butyl, 3,3- dimethyl-2-butyl, n-pentyl, n-hexyl.

As used herein with respect to a substituting radical, and unless otherwise stated, the term "C2-C10 alkenyl " designate a straight and branched acyclic hydrocarbon monovalent radical having one or more ethylenic unsaturations and having from 2 to 10 carbon atoms such as, for example, vinyl, 1 -propenyl, 2-propenyl (allyl), 1 -butenyl, 2-butenyl, 2-pentenyl, 3- pentenyl, 3-methyl-2-butenyl, 3-hexenyl, 2-hexenyl, 2-heptenyl, 1 ,3-butadienyl, pentadienyl, hexadienyl, heptadienyl, heptatrienyl, 2-octenyl and the like, including all possible isomers thereof.

As used herein with respect to a substituting radical, and unless otherwise stated, the term "C2-C10 alkynyl " defines straight and branched chain hydrocarbon radicals containing one or more triple bonds and optionally at least one double bond and having from 2 to 10 carbon atoms such as, for example, acetylenyl, 1 -propynyl, 2- propynyl, 1 -butynyl, 2-butynyl, 2- pentynyl, 1 -pentynyl, 3-methyl-2-butynyl, 3-hexynyl, 2-hexynyl, 1 -penten-4-ynyl, 3-penten-1 - ynyl, 1 ,3-hexadien-1 -ynyl and the like.

As used herein and unless otherwise stated, the term "cycloalkyl" means a monocyclic saturated hydrocarbon monovalent radical having the number of carbon atoms as indicated (or where not indicated, preferably having 3-20, more preferably 3-10 carbon atoms, more preferably 3-8 or 3-6 carbon atoms). "C3-C8 cycloalkyl" refers to such monocyclic saturated hydrocarbon monovalent radical having from 3 to 8 carbon atoms, such as for instance cyclo- propyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl. The term "alkoxy" refers to the group alkyl-O-, where alkyl is as defined above. "(C1-C6) alkoxy" as used herein includes but is not limited to methoxy, ethoxy, propoxy, isopropoxy, butoxy, iso-butoxy, sec-butoxy, pentoxy, 3-pentoxy, or hexyloxy.

As used herein and unless otherwise stated, the term "halogen" or "halo" means any atom selected from the group consisting of fluorine (F), chlorine (CI), bromine (Br) and iodine

(I)-

As used herein and unless otherwise stated, the term "Ar" or "aryl" means a monovalent unsaturated aromatic carbocyclic radical having one, two, three, four, five or six rings, preferably one, two or three rings, which may be fused or bicyclic. An aryl group may optionally be substituted by one, two, three or more substituents as set out in this invention with respect to optional substituents that may be present on the group Ar or aryl. Preferred aryl groups are: an aromatic monocyclic ring containing 6 carbon atoms; an aromatic bicyclic or fused ring system containing 7, 8, 9 or 10 carbon atoms; or an aromatic tricyclic ring system containing 10, 1 1 , 12, 13 or 14 carbon atoms. Non- limiting examples of aryl include phenyl and naphthyl. Preferred substituent groups of Ar are independently selected from halogen, C1-C6 alkyl, Ci- C₆ alkoxy , hydroxy (-OH), acyl (R'-C(=0)-, acyloxy (R'-C(=0)-0-), nitro (-NO2), amino (-NH₂), -SO3H, -SH, -SR', wherein R' is an alkyl. Preferred Ar are phenyl, bromophenyl and naphthyl.

As used herein and unless otherwise stated, the term " stereoisomer " refers to all possible different isomeric as well as conformational forms which the compounds of formula I and lb may possess, in particular all possible stereochemical and conformationally isomeric forms, all diastereomers, enantiomers and/or conformers of the basic molecular structure. Some compounds of the present invention may exist in different tautomeric forms, all of the latter being included within the scope of the present invention.

As used herein and unless otherwise stated, the term " enantiomer " means each individual optically active form of a compound of the invention, having an optical purity or enantiomeric excess (as determined by methods standard in the art) of at least 80% (i.e. at least 90% of one enantiomer and at most 10% of the other enantiomer), preferably at least 90% and more preferably at least 98%.

Examples

EXPERIMENTAL SECTION

For reactions, all reagents and solvents were purchased from commercial sources and were used as obtained. Moisture sensitive reactions were carried out in oven-dried glassware under a nitrogen or argon atmosphere. ¹H, ¹³C, and ³¹P NMR spectra were recorded on Bruker Avance 300, 500, or 600 MHz spectrometers with tetramethylsilane as internal standard or referenced to the residual solvent signal, and 85% Η₃Ρ0₄ ίθΓ ³¹P NMR experiments. The intermediates and final compounds were characterized by using 2D NMR (H-COSY, HSQC, and HMBC) spectroscopic techniques. High-resolution mass spectra (HRMS) were obtained on a quadruple orthogonal acceleration time-of-flight mass spectrometer (Synapt G2 HDMS, Waters, Milford, MA). Samples were infused at 3 μΙ_/η"ΐίη, and spectra were obtained in positive (or in negative) ionization mode with a resolution of 15 000 (fwhm) using leucine enkephalin as the lock mass. Pre-coated aluminum sheets (254 nm) were used for TLC. Products were purified by column chromatography on silica gel (60 A, 0.035-0.070 mm, Acros Organics). Preparative RP-HPLC purifications were performed on a Phenomenex Gemini 1 1 OA column (C18, 10 m, 21 .2 mm ^χ 250 mm) using H₂0/CH₃CN with 50 mmol TEAB or H₂0/CH₃CN as eluent gradient.

Example 1 : Synthesis of (SJ-l -iS-Fluoro^-^diethylphosphorylJmethoxylpropy^-yV³- (benzyloxymethyl)thymine (6a)

A solution of DIAD (0.32 mL, 1.64 mmol) in anhydrous THF (1 mL) was added dropwise to a mixture of compound 4a¹ (200 mg, 0.82 mmol), compound 5 (240 mg, 0.98 mmol), and Ph₃P (430 mg, 1.64 mmol) in anhydrous THF (5 mL) at room temperature. The reaction mixture was stirred for 12 h, and it was then concentrated under reduced pressure. The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 80:1 , v/v; DCM/MeOH, 50:1 , v/v) to give 6a (310 mg, 80%) as a colorless oil. ¹H NMR (300 MHz, CDCI₃): δ 7.39-7.25 (m, 5H, ArH), 7.14-7.12 (m, 1 H, H-6), 5.50 (s, 2H, OCH₂N), 4.73-4.33 (m, 4H, CH r, H-3'), 4.17-3.92 (m, 7H, H-1 ', H-2', 2 x CH₂CH₃), 3.84-3.66 (m, 2H, PCH₂), 1 .93 (d, J = 1 .2 Hz, 3H, CH₃-5), 1 .34-1.28 (m, 6H, 2 x CH₂CH₃); ¹³C NMR (75 MHz, CDCI₃): δ 163.8 (C-4), 151 .8 (C- 2), 140.5 (C-6), 138.1 (Ar-C), 128.3 (Ar-C), 127.7 (Ar-C), 109.7 (C-5), 82.3 (d, ¹J_C,F = 173.4 Hz, C-3'), 78.6 (dd, ²J_C,F = 18.5 Hz, ³J_C,P = 9.8 Hz, C-2'), 72.3 (OCH₂N), 70.8 (CH₂-Ar), 64.6 (d, ¹Jc,p = 167.4 Hz, CH₂P), 62.6, 62.5 (2 x d, ²J_c,p = 3.8 Hz, CH₂CH₃), 62.5 (d, ²J_c,p = 3.7 Hz, CH₂CH₃), 49.3 (d, ³J_C,F = 8.6 Hz, C-1 '), 16.6, 16.5 (CH₂CH₃), 13.0 (CH₃-5); ³¹P NMR (121 MHz, CDCIs): δ 20.8; HRMS for C₂iH₃₀FN₂O₇P [M+H]⁺ calcd.: 473.1847, found: 473.1848.

Example 2 : Synthesis of (R)-1 -{3-Fluoro-2-[(diethylphosphoryl)methoxy]propyl}-N3- (benzyloxymethyl)thymine (6b)

Compound 6b was obtained as a colorless oil (460 mg, 70%) according to the procedure used for the preparation of 6a, starting from compound 4b¹ (300 mg, 1.23 mmol), compound 5 (360 mg, 1.47 mmol), Ph₃P (640 mg, 2.46 mmol), and DIAD (0.48 mL, 2.46 mmol) in anhydrous THF (8 mL). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 80:1 , v/v; DCM/MeOH, 50:1 , v/v). ¹H NMR (300 MHz, CDCI₃): δ 7.39-7.25 (m, 5H, ArH), 7.13 (t, J = 1.3 Hz, 1 H, H-6), 5.50 (s, 2H, OCH₂N), 4.73-4.33 (m, 4H, CH r, H-3'), 4.17-3.91 (m, 7H, H-1 ', H-2', 2 x CH₂CH₃), 3.84-3.66 (m, 2H, PCH₂), 1 .93 (d, J = 1 .1 Hz, 3H, CH₃-5), 1 .34-1.28 (m, 6H, 2 x CH₂CH₃); ¹³C NMR (75 MHz, CDCI₃): δ 163.8 (C-4), 151 .8 (C- 2), 140.5 (C-6), 138.0 (Ar-C), 128.3 (Ar-C), 127.7 (Ar-C), 109.7 (C-5), 82.3 (d, ¹J_C,F = 173.5 Hz, C-3'), 78.6 (dd, ²J_C,F = 18.5 Hz, ³J_C,P = 9.7 Hz, C-2'), 72.3 (OCH₂N), 70.7 (CH₂-Ar), 64.6 (d, ¹Jc,p = 167.3 Hz, CH₂P), 62.6, 62.5 (2 x d, ²J_c,p = 3.5 Hz, CH₂CH₃), 49.3 (d, ³J_C,F = 8.8 Hz, C- 1 '), 16.6, 16.5 (CH₂CH₃), 13.0 (CH₃-5); ³¹P NMR (121 MHz, CDCI₃): δ 20.8; HRMS for C₂i H₃₀FN₂O₇P [M+H]⁺ calcd.: 473.1847, found: 473.1841 .

Example 3: (S)-1 -{3-Fluoro-2-[(diethylphosphoryl)methoxy]propyl}thymine (7a)

A suspension of compound 6a (240 mg, 0.51 mmol) and Pd/C on charcoal (10%w/w, 120 mg) in EtOH (10 mL) was purged with nitrogen, followed by hydrogen, and then allowed to stir under a hydrogen atmosphere. After 24 h, the mixture was filtered through a pad of Celite and the filtrate was evaporated under reduced pressure to give a crude product. The residue was then redissolved in MeOH (10 mL) and Et₃N (1 mL) and the solution was stirred for 3 h at room temperature. The reaction mixture was concentrated under reduced pressure and the resulting crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 30:1 , v/v; DCM/MeOH, 20:1 , v/v) to give 7a (160 mg, 90%) as a colorless oil. ¹H NMR (300 MHz, CDCI₃): δ 9.63 (s, 1 H, NHCO), 7.16 (t, J = 1 .3 Hz, 1 H, H-6), 4.72-4.33 (m, 2H, H-3'), 4.17-3.90 (m, 7H, H-1 ', H-2', 2 x CH₂CH₃), 3.83-3.63 (m, 2H, PCH₂), 1 .89 (d, J = 1 .2 Hz, 3H, CH₃-5), 1 .33-1.28 (m, 6H, 2 x CH₂CH₃); ¹³C NMR (75 MHz, CDCI₃): δ 164.5 (C-4), 151 .3 (C-

2), 141 .9 (C-6), 1 10.4 (C-5), 82.3 (d, ¹J_C,F = 173.4 Hz, C-3'), 78.7 (dd, ²J_C,F = 1 8.5 Hz, ³J_C,P = 9.5 Hz, C-2'), 64.6 (d, ¹J_c,p = 167.3 Hz, CH₂P), 62.7, 62.6 (CH₂CH₃), 48.6 (d, ³J_C,F = 8.8 Hz, C- 1 '), 16.6, 16.5 (CH2CH3), 12.3 (CH₃-5); ³¹P NMR (121 MHz, CDC ): δ 21 .0; HRMS for C9H14FN2O6P [M+H]⁺ calcd.: 353.1272, found: 353.1262.

Example 4 Synthesis

[(diethylphosphoryl)methoxy]propyl}thymine (7b).

Compound 7b was obtained as a colorless oil (270 mg, 90%) according to the procedure used for the preparation of 7a, starting from compound 6b (400 mg, 0.85 mmol), Pd/C on charcoal (10%w/w, 200 mg) in EtOH (20 ml_), and Et₃N (1 mL) in MeOH (20 ml_). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 30:1 , v/v; DCM/MeOH, 20:1 , v/v). ¹H NMR (300 MHz, CDCb): δ 9.67 (s, 1 H, NHCO), 7.18 (d, J = 1.4 Hz, 1 H, H-6), 4.75-4.35 (m, 2H, H-3'), 4.19-3.93 (m, 7H, H-1 ', H-2', 2 x CH2CH3), 3.86-3.66 (m, 2H, PCH₂), 1 .92 (d, J = 1 .2 Hz, 3H, CH₃-5), 1.35-1.31 (m, 6H, 2 x CH2CH3); ¹³C NMR (75 MHz, CDCb): δ 164.5 (C-4), 151 .3 (C-2), 141.9 (C-6), 110.4 (C-5), 82.3 (d, ¹J_C,F = 173.4 Hz, C-3'), 78.7 (dd, ²JC,F = 18.5 Hz, ³J_c,p = 9.5 Hz, C-2'), 64.6 (d, ¹J_c,p = 167.4 Hz, CH₂P), 62.7, 62.6 (2 x d, ²Jc,p = 5.5 Hz, CH2CH3), 48.5 (d, ³J_C,F = 8.8 Hz, C-1 '), 16.6, 16.5 (CH₂CH₃), 12.3 (CH₃-5); ³¹P NMR (121 MHz, CDCb): δ 21 .0; HRMS for C9H14FN2O6P [M+H]⁺ calcd.: 353.1272, found: 353.1278.

Example 5 Synthesis of (S)-1 -[3-Fluoro-2-(phosphonomethoxy)propyl]thymine triethylammonium salt (8a)

Bromotrimethylsilane (0.15 mL, 1.14 mmol) was added dropwise to a solution of diethyl phosphonate ester 7a (100 mg, 0.28 mmol) and 2,6-lutidine (0.26 mL, 2.27 mmol) in anhydrous acetonitrile (5 mL) at 0 °C. After the addition was completed, the mixture was slowly warmed to room temperature and set aside in the dark for 12 h. The reaction was quenched with 0.1 M TEAB and was then concentrated under reduced pressure. The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH/Et3N, 10:5:1 , v/v/v; 7.5:5:1 , v/v/v). The collected eluate was freeze-dried repeatedly until constant mass to give the desired phosphonate acid triethylammonium salt 8a (59 mg, 70%) as a white foam. ¹H NMR (300 MHz, D₂0): δ 7.52 (s, 1 H, H-6), 4.77-4.48 (m, 2H, H-3', overlapped with H₂0), 3.93-3.79 (m, 3H, H- 1 ', H-2'), 3.58-3.46 (m, 2H, PCH₂), 1 .80 (s, 3H, CH₃-5); ¹³C NMR (75 MHz, D₂0): δ 166.8 (C- 4), 152.2 (C-2), 143.5 (C-6), 110.3 (C-5), 82.1 (d, ¹J_C,F = 167.3 Hz, C-3'), 77.1 (dd, ²J_C,F = 18.1 Hz, ³JC,P = 10.4 Hz, C-2'), 67.7 (d, ¹J_c,p = 150.9 Hz, CH₂P), 47.3 (d, ³J_C,F = 6.9 Hz, C-1 '), 11 .0 (CHs-5); ³¹P NMR (121 MHz, D₂0): δ 12.9; HRMS for C₉Hi₄FN₂0₆P [M-H]^" calcd.: 295.0501 , found: 295.0499.

Spectral data are in accordance with literature data (Pomeisl, K.; Pohl, R.; Holy, A.; Votruba, I. Collect. Czech. Chem. Commun. 2005, 70, 1465-1481 ).

Example 6 Synthesis of ( ?)-1 -[3-Fluoro-2-(phosphonomethoxy)propyl]thymine triethylammonium salt (8b)

Compound 8b was obtained as a white foam (176 mg, 70%) according to the procedure used for the preparation of 8a, starting from compound 7b (300 mg, 0.85 mmol), bromotrimethylsilane (0.45 mL, 3.41 mmol), and 2,6-lutidine (0.79 mL, 6.81 mmol) in anhydrous acetonitrile (10 mL). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH/Et₃N, 10:5:1 , v/v/v; 7.5:5:1 , v/v/v). ¹H NMR (300 MHz, D₂0): δ 7.55-7.54 (m, 1 H, H-6), 4.80-4.43 (m, 2H, H-3', overlapped with H₂0), 4.09-3.88 (m, 3H, H-1 ', H-2'), 3.79-3.60 (m, 2H, PCH₂), 1.87 (d, J = 1.1 Hz, 3H, CH₃); ¹³C NMR (75 MHz, D₂0): δ 166.7 (C-4), 152.0 (C-2), 143.5 (C-6), 110.2 (C-5), 82.0 (d, ¹J_C,F = 168.0 Hz, C-3'), 77.5 (dd, ²J_C,F = 18.2 Hz, ³JC,P = 11 .5 Hz, C-2'), 66.4 (d, ¹J_c,p = 156.3 Hz, CH₂P), 47.6 (d, ³J_C,F = 7.6 Hz, C-1 '), 1 1.0 (CH₃); ³¹P NMR (121 MHz, D₂0): δ 14.7; HRMS for C₉Hi₄FN₂0₆P [M-H]^" calcd.: 295.0501 , found: 295.0498.

Example 7 Synthesis of (S)-6-Chloro-9-{3-fluoro-2-

[(diethylphosphoryl)methoxy]propyl}purine (10a)

Compound 10a was obtained as a colorless oil (190 mg, 60%) according to the procedure used for the preparation of 6a, starting from compound 4a (200 mg, 0.82 mmol), compound 9 (150 mg, 0.98 mmol), Ph₃P (430 mg, 1 .64 mmol), and DIAD (0.32 mL, 1 .64 mmol) in anhydrous THF (6 mL). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 50:1 , v/v; DCM/MeOH, 30:1 , v/v). ¹H NMR (300 MHz, CDCI₃): δ 8.75 (s, 1 H, H- 2), 8.32 (s, 1 H, H-8), 4.74-4.40 (m, 4H, H-3', H-1 '), 4.22-3.94 (m, 6H, H-2', 2 x CH₂CH₃, PCH₂a), 3.78 (dd, J = 14.0, 8.5 Hz, 1 H, PCH₂b), 1.34-1.24 (m, 6H, 2 x CH₂CH₃); ¹³C NMR (75 MHz, CDCI₃): δ 152.0 (C-2, C-4), 151.1 (C-6), 146.5 (C-8), 131 .4 (C-5), 81.6 (d, ¹J_C,F = 174.3 Hz, C- 3'), 78.0 (dd, ²JC,F = 19.6 Hz, ³J_C,P = 9.0 Hz, C-2'), 64.4 (d, ¹ J_c,p = 167.1 Hz, CH₂P), 62.7, 62.5 (2 x d, ²Jc,p = 6.7 Hz, CH₂CH₃), 44.2 (d, ³J_C,F = 8.0 Hz, C-1 '), 16.5, 16.4 (2 x d, ³J_c,p = 3.8 Hz, CH₂CH₃); ³¹P NMR (121 MHz, CDCI₃): δ 20.3; HRMS for Ci₃Hi₉CIFN₄0₄P [M+H]⁺ calcd.: 381 .0889, found: 381.0884.

Example 8 Synthesis of ( ?)-6-Chloro-9-{3-fluoro-2-

[(diethylphosphoryl)methoxy]propyl}purine (10b)

Compound 10b was obtained as a colorless oil (300 mg, 65%) according to the procedure used for the preparation of 6a, starting from compound 4b (300 mg, 1.23 mmol), compound 9 (230 mg, 1.50 mmol), Ph₃P (640 mg, 2.46 mmol), and DIAD (0.48 mL, 2.46 mmol) in anhydrous THF (10 mL). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 50:1 , v/v; DCM/MeOH, 30:1 , v/v). ¹H NMR (300 MHz, CDCI₃): δ 8.71 (s, 1 H, H- 2), 8.26 (s, 1 H, H-8), 4.69-4.35 (m, 4H, H-3', H-1 '), 4.17-3.89 (m, 6H, H-2', 2 x CH₂CH₃, PCH₂a), 3.73 (dd, J = 14.0, 8.4 Hz, 1 H, PCH₂b), 1.30-1.19 (m, 6H, 2 x CH₂CH₃); ¹³C NMR (75 MHz, CDCI₃): δ 152.0 (C-2, C-4), 151.1 (C-6), 146.5 (C-8), 131 .5 (C-5), 81.6 (d, ¹J_C,F = 174.3 Hz, C- 3'), 78.1 (dd, ²JC,F = 19.6 Hz, ³J_C,P = 8.9 Hz, C-2'), 64.5 (d, ¹ J_c,p = 167.0 Hz, CH₂P), 62.7, 62.6 (2 x d, ²Jc,p = 6.6 Hz, CH₂CH₃), 44.2 (d, ³J_C,F = 8.2 Hz, C-1 '), 16.5, 16.4 (2 x d, ³J_c,p = 4.5 Hz, CH₂CH₃); ³¹P NMR (121 MHz, CDCI₃): δ 20.6; HRMS for Ci₃Hi₉CIFN₄0₄P [M+H]⁺ calcd.: 381 .0889, found: 381.0883. Example 9 : Synthesis of (S)-9-{3-Fluoro-2-[(diethylphosphoryl)methoxy]propyl}adenine

(11 a)

A solution of 10a (150 mg, 0.40 mmol) in 10% ethanolic ammonia (20 mL) was stirred at 50 °C for 24 h. After removal of all the volatiles, the residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 20:1 , v/v; 15:1 , v/v) to give 11 a (110 mg, 80%) as a colorless oil. ¹H NMR (300 MHz, CDCI₃): δ 8.30 (s, 1 H, H-2), 8.00 (s, 1 H, H-8), 6.75 (s, 2H, NH₂), 4.71 -4.27 (m, 4H, H-3', H-1 '), 4.14-3.90 (m, 6H, H-2', 2 x CH₂CH₃, PCH₂a), 3.76 (dd, J = 13.9, 8.9 Hz, 1 H, PCH₂b), 1 .30-1.22 (m, 6H, 2 x CH₂CH₃); ¹³C NMR (75 MHz, CDC ): δ 155.0 (C-6), 151 .6 (C-2), 159.9 (C-4), 142.1 (C-8), 119.2 (C-5), 82.0 (d, ¹J_C,F = 173.8 Hz, C-3'), 78.5 (dd, ²JC,F = 19.3 Hz, ³JC,P = 10.0 Hz, C-2'), 64.6 (d, ¹J_c,p = 167.3 Hz, CH₂P), 62.7, 62.6 (CH₂CH₃), 43.6 (d, ³J_C,F = 8.3 Hz, C-1 '), 16.5, 16.4 (CH₂CH₃); ³¹P NMR (121 MHz, CDCI₃): δ 20.5; HRMS for Ci₃H₂iFN₅0₄P [M+H]⁺ calcd.: 362.1389, found: 362.1386.

Example 10 Synthesis of (R)- 9-{3-Fluoro-2-

[(diethylphosphoryl)methoxy]propyl}adenine (11 b)

Compound 11 b was obtained as a colorless oil (260 mg, 90%) according to the procedure used for the preparation of 11a, starting from compound 10b (300 mg, 0.80 mmol) in 10% ethanolic ammonia (30 mL). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 20:1 , v/v; 15:1 , v/v). ¹H NMR (300 MHz, CDCI₃): δ 8.35 (s, 1 H, H-2), 7.95 (s, 1 H, H-8), 5.86 (s, 2H, NH₂), 4.74-4.28 (m, 4H, H-3', H-1 '), 4.18-3.91 (m, 6H, H- 2', 2 x CH₂CH₃, PCH₂a), 3.78 (dd, J = 13.9, 8.9 Hz, 1 H, PCH₂b), 1 .34-1.25 (m, 6H, 2 x CH₂CH₃); ¹³C NMR (150 MHz, CDCI3): δ 155.4 (C-6), 153.1 (C-2), 150.2 (C-4), 141 .6 (C-8), 119.4 (C-5), 82.0 (d, ¹JC,F = 173.8 Hz, C-3'), 78.5 (dd, ²J_C,F = 19.4 Hz, ³J_C,P = 9.4 Hz, C-2'), 64.6 (d, ¹J_c,p = 167.3 Hz, CH₂P), 62.5, 62.4 (d, ²J_c,p = 6.5 Hz, CH₂CH₃), 43.6 (d, ³J_C,F = 8.1 Hz, C-1 '), 16.4, 16.3 (CH₂CH₃); ³¹P NMR (121 MHz, CDCb): δ 20.6; HRMS for Ci₃H₂iFN₅0₄P [M+H]⁺ calcd.: 362.1389, found: 362.1390.

Example 11 : Synthesis of (S)-9-[3-Fluoro-2-(phosphonomethoxy)propyl]adenine (12a)

Compound 12a was obtained as a white foam (38 mg, 65%) according to the procedure used for the preparation of 8a, starting from compound 11 a (70 mg, 0.19 mmol), bromotrimethylsilane (0.10 mL, 0.77 mmol), and 2,6-lutidine (0.18 mL, 1.59 mmol) in anhydrous acetonitrile (5 mL). The crude residue was purified by column chromatography on silica gel (gradient acetone/H₂0/Et₃N, 6:1 :1 , v/v/v). ¹H NMR (300 MHz, D₂0): δ 8.23 (s, 1 H, H- 2), 8.14 (s, 1 H, H-8), 4.97-4.27 (m, 4H, H-3', H-1 '), 4.05-3.95 (m, 1 H, H-2'), 3.55-3.40 (m, 2H, PCH₂); ¹³C NMR (75 MHz, D₂0): δ 154.2 (C-6), 150.7 (C-2), 148.7 (C-4), 143.4 (C-8), 1 17.8 (C-5), 81.7 (d, ¹J_C,F = 168.0 Hz, C-3'), 77.6 (dd, ²J_C,F = 18.9 Hz, ³J_C,P = 11 .4 Hz, C-2'), 66.1 (d, ¹Jc,p = 157.0 Hz, CH₂P), 43.3 (d, ³J_C,F = 7.5 Hz, C-1 '); ³¹P NMR (121 MHz, D₂0): δ 12.7; HRMS for C9H13FN5O4P [M-H]- calcd.: 304.0616, found: 304.0603.

Spectral data are in accordance with literature data (Jindrich, J.; Holy, A.; Dvorakova, H. Collect. Czech. Chem. Commun. 1993, 58, 1645-1667).

Example 12 : Synthesis of ( ?)-9-[3-Fluoro-2-(phosphonomethoxy)propyl]adenine (12b)

Compound 12b was obtained as a white foam (148 mg, 70%) according to the procedure used for the preparation of 8a, starting from compound 11 b (250 mg, 0.67 mmol), bromotrimethylsilane (0.37 mL, 2.77 mmol), and 2,6-lutidine (0.64 mL, 5.54 mmol) in anhydrous acetonitrile (10 mL). The crude residue was purified by column chromatography on silica gel (gradient acetone/H₂0/Et₃N, 6:1 :1 , v/v/v). ¹H NMR (300 MHz, D₂0): δ 8.01 (d, J = 1.0 Hz, 1 H, H-2), 7.89 (d, J = 1 .0 Hz, 1 H, H-8), 4.71 -4.18 (m, 4H, H-3', H-1 '), 3.96-3.85 (m, 1 H, H- 2'), 3.62-3.40 (m, 2H, PCH₂); ¹³C NMR (75 MHz, D₂0): δ 154.6 (C-6), 151.6 (C-2), 148.2 (C- 4), 142.6 (C-8), 117.3 (C-5), 81.7 (d, ¹J_C,F = 168.1 Hz, C-3'), 77.3 (dd, ²J_C,F = 18.9 Hz, ³J_C,P = 1 1.4 Hz, C-2'), 66.4 (d, ¹J_c,p = 155.5 Hz, CH₂P), 43.0 (d, ³J_C,F = 7.4 Hz, C-1 '); ³¹P NMR (121 MHz, D₂0): δ 14.2; HRMS for C9H13FN5O4P [M-H]^" calcd.: 304.0616, found: 304.0609. Spectral data are in accordance with literature data (Jindrich, J.; Holy, A.; Dvorakova, H. Collect. Czech. Chem. Commun. 1993, 58, 1645-1667).

Example 13 : Synthesis of (SJ-l -iS-Fluoro^-^diethylphosphorylJmethoxylpropyl}-/^- isobutyrylcytosine (14a)

Compound 14a was obtained as a colorless oil (130 mg, 40%) according to the procedure used for the preparation of 6a, starting from compound 4a (200 mg, 0.82 mmol), compound 13 (180 mg, 0.98 mmol), Ph₃P (430 mg, 1 .63 mmol), and DIAD (0.33 mL, 1 .63 mmol) in anhydrous THF (6 mL). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 30:1 , v/v; DCM/MeOH, 25:1 , v/v). ¹H NMR (300 MHz, CDCI₃): δ 9.14 (s, 1 H, NHCO), 7.71 (d, J = 7.3 Hz, 1 H, H-6), 7.41 (d, J = 7.3 Hz, 1 H, H-5), 4.82-4.31 (m, 3H, H-3', H- 1 'a), 4.22-3.95 (m, 7H, H-1 'b, H-2', 2 x CH₂CH₃, PCH₂a), 3.70 (dd, J = 13.6, 8.3 Hz, 1 H, PCH₂b), 2.72-2.63 (m, 1 H, CH(CH₃)₂), 1 .36-1.20 (m, 12H, 2 x CH₂CH₃, CH(CH₃)₂); ¹³C NMR (75 MHz, CDCI₃): δ 177.4 (CONH), 163.0 (C-4), 156.1 (C-2), 150.4 (C-6), 96.3 (C-5), 82.3 (d, ¹J_C,F = 173.4 Hz, C-3'), 78.3 (dd, ²J_C,F = 18.3 Hz, ³J_C,P = 11 .9 Hz, C-2'), 64.4 (d, ¹J_c,p = 166.9 Hz, CH₂P), 62.7, 62.5 (2 x d, ²J_c,p = 6.6 Hz, CH₂CH₃), 51 .0 (d, ³J_C,F = 8.7 Hz, C-1 '), 36.7 (CH(CH₃)₂), 19.3, 19.1 (CH(CH₃)₂), 16.6, 16.5 (CH₂CH₃); ³¹P NMR (121 MHz, CDCI₃): δ 20.7; HRMS for Ci₆H₂₇FN₃06P [M+H]⁺ calcd.: 408.1694, found: 408.1690.

Example 14 : Synthesis of (R)-1 -{3-Fluoro-2-[(diethylphosphoryl)methoxy]propyl}-N4- isobutyrylcytosine (14b)

Compound 14b was obtained as a colorless oil (260 mg, 40%) according to the procedure used for the preparation of 6a, starting from compound 4a (400 mg, 1 .64 mmol), compound 9 (360 mg, 1.96 mmol), Ph₃P (860 mg, 3.26 mmol), and DIAD (0.66 mL, 3.26 mmol) in anhydrous THF (10 mL). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 30:1 , v/v; DCM/MeOH, 25:1 , v/v). Ή NMR (300 MHz, CDCI₃): δ 8.89 (s, 1 H, NHCO), 7.71 (d, J = 7.3 Hz, 1 H, H-6), 7.40 (d, J = 7.4 Hz, 1 H, H-5), 4.82-4.31 (m, 3H, H-3', H- 1 'a), 4.24-3.95 (m, 7H, H-1 'b, H-2', 2 x CH₂CH₃, PCH₂a), 3.70 (dd, J = 13.4, 8.2 Hz, 1 H, PCH₂b), 2.69-2.60 (m, 1 H, CH(CH₃)₂), 1 .35-1.20 (m, 12H, 2 x CH₂CH₃, CH(CH₃)₂); ¹³C NMR (75 MHz, CDCI₃): δ 177.2 (CONH), 162.9 (C-4), 156.1 (C-2), 150.4 (C-6), 96.3 (C-5), 82.3 (d, ¹J_C,F = 173.3 Hz, C-3'), 78.3 (dd, ²J_C,F = 18.4 Hz, ³J_c,p = 11 .8 Hz, C-2'), 64.4 (d, ¹J_c,p = 166.9 Hz, CH₂P), 62.7, 62.5 (2*d, ²J_c,p = 6.6 Hz, CH₂CH₃), 51 .0 (d, ³J_C,F = 9.1 Hz, C-1 '), 36.7 (CH(CH₃)₂), 19.3, 19.1 (CH(CH₃)₂), 16.6, 16.5 (CH₂CH₃); ³¹P NMR (121 MHz, CDCI₃): δ 21 .4; HRMS for Ci₆H₂₇FN₃06P [M+H]⁺ calcd.: 408.1694, found: 408.1691 .

Example 15 : Synthesis of (S)-0²-{3-Fluoro-2-[(diethylphosphoryl)methoxy]propyl}-Ai - isobutyrylcytosine (15a)

Compound 15a was obtained as a colorless oil (150 mg, 45%) according to the procedure used for the preparation of 6a, starting from compound 4a (200 mg, 0.82 mmol), compound 13 (180 mg, 0.98 mmol), Ph₃P (430 mg, 1 .63 mmol), and DIAD (0.33 mL, 1 .63 mmol) in anhydrous THF (6 mL). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 80:1 , v/v; DCM/MeOH, 50:1 , v/v). Ή NMR (600 MHz, CDCI₃): δ 8.83 (s, 1 H, NHCO), 8.38 (dd, J = 5.6, 0.6 Hz, 1 H, H-6), 7.84 (d, J = 5.7 Hz, 1 H, H-5), 4.72-4.50 (m, 3H, H- 3', H-1 'a), 4.38-4.35 (m, 1 H, H-1 'b), 4.23-4.15 (m, 5H, H-2', 2 x CH₂CH₃), 4.08 (d, J = 8.1 Hz, 2H, PCH₂), 2.68-2.61 (m, 1 H, CH(CH₃)₂), 1 .36-1.33 (m, 6H, 2 x CH₂CH₃), 1 .26-1.25 (m, 6H, CH(CH₃)₂); ¹³C NMR (150 MHz, CDCI₃): δ 176.8 (CONH), 164.1 (C-2), 160.6 (C-6), 159.5 (C- 4), 104.4 (C-5), 82.9 (d, ¹J_C,F = 172.0 Hz, C-3'), 78.1 (dd, ²J_C,F = 19.1 Hz, ³J_c,p = 8.9 Hz, C-2'), 65.0 (d, ³JC,F = 8.1 Hz, C-1 '), 64.9 (d, ¹J_c,p = 195 Hz, CH₂P), 62.8, 62.6 (2 x d, ²J_c,p = 6.5 Hz, CH₂CH₃), 36.6 (CH(CH₃)₂), 19.2, 19.1 (CH(CH₃)₂), 16.5, 16.4 (CH₂CH₃); ³¹P NMR (121 MHz, CDCI₃): δ 20.7; HRMS for Ci₆H₂₇FN₃06P [M+H]⁺ calcd.: 408.1694, found: 408.1699.

Example 16 : Synthesis of ( ?)-0²-{3-Fluoro-2-[(diethylphosphoryl)methoxy]propyl}-Ai - isobutyrylcytosine (15b)

Compound 15b was obtained as a colorless oil (300 mg, 45%) according to the procedure used for the preparation of 6a, starting from compound 4b (400 mg, 1.64 mmol), compound 13 (360 mg, 1 .96 mmol), Ph₃P (860 mg, 3.26 mmol), and DIAD (0.66 mL, 3.26 mmol) in anhydrous THF (10 mL). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 80:1 , v/v; DCM/MeOH, 50:1 , v/v). ¹H NMR (300 MHz, CDCI₃): δ 8.94 (s, 1 H, NHCO), 8.38 (d, J = 5.6 Hz, 1 H, H-6), 7.84 (d, J = 5.6 Hz, 1 H, H-5), 4.77-4.48 (m, 3H, H-3', H-1 'a), 4.39-4.33 (m, 1 H, H-1 'b), 4.25-4.07 (m, 7H, H-2', 2 x CH₂CH₃, PCH₂), 2.73- 2.63 (m, 1 H, CH(CH₃)₂), 1 .37-1.24 (m, 12H, 2 x CH₂CH₃, CH(CH₃)₂); ¹³C NMR (75 MHz, CDCI₃): δ 176.9 (CONH), 164.1 (C-2), 160.5 (C-6), 159.5 (C-4), 104.4 (C-5), 82.8 (d, ¹J_C,F = 172.3 Hz, C-3'), 78.1 (dd, ²J_C,F = 18.8 Hz, ³J_C,P = 9.1 Hz, C-2'), 64.9 (d, ³J_C,F = 8.2 Hz, C-1 '), 64.8 (d, ¹Jc,p = 165 Hz, CH₂P), 62.8, 62.5 (d, ²J_c,p = 6.6 Hz, CH₂CH₃), 36.4 (CH(CH₃)₂), 19.2, 19.1 (CH(CH₃)₂), 16.5, 16.4 (CH₂CH₃); ³¹P NMR (121 MHz, CDCI₃): δ 20.9; HRMS for Ci₆H₂₇FN₃06P [M+H]⁺ calcd.: 408.1694, found: 408.1693.

Example 17 Synthesis of (S)-1 -{3-Fluoro-2-

[(diethylphosphoryl)methoxy]propyl}cytosine (16a)

A solution of 14a (120 mg, 0.31 mmol) in 30% methanolic ammonia (20 mL) was stirred at 45 °C for 15 h. After removal of all the volatiles, the residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 10:1 , v/v; DCM/MeOH, 8:1 , v/v) to give 16a (90 mg, 90%) as a colorless foam. ¹H NMR (300 MHz, CDCI₃): δ 7.40 (d, J = 7.2 Hz, 1 H, H-6), 5.92 (d, J = 7.2 Hz, 1 H, H-5), 4.77-4.35 (m, 2H, H-3'), 4.18-3.95 (m, 7H, H-1 ', H-2', 2 x CH₂CH₃), 3.83 (dd, J = 13.8, 9.0 Hz, 1 H, PCH₂a), 3.70 (dd, J = 13.4, 7.2 Hz, 1 H, PCH₂b), 1 .36- 1 .30 (m, 6H, 2 x CH₂CH₃); ¹³C NMR (75 MHz, CDCI₃): δ 165.8 (C-4), 156.3 (C-2), 147.1 (C-6), 94.7 (C-5), 82.6 (d, ¹J_C,F = 172.9 Hz, C-3'), 78.7 (dd, ²J_C,F = 18.4 Hz, ³J_C,P = 10.8 Hz, C-2'), 64.6 (d, ¹Jc,p = 167.6 Hz, CH₂P), 62.8, 62.6 (2 x d, ²J_c,p = 6.6 Hz, CH₂CH₃), 50.1 (d, ³J_C,F = 8.7 Hz, C-1 '), 16.6, 16.5 (CH₂CH₃); ³¹P NMR (121 MHz, CDCI₃): δ 20.4; HRMS for C12H21 FN3O5P [M- H]⁺ calcd.: 338.1275, found: 338.1280.

Example 18 Synthesis of (/?)-1 -{3-Fluoro-2-

[(diethylphosphoryl)methoxy]propyl}cytosine (16b)

Compound 16b was obtained as a white foam (140 mg, 82%) according to the procedure used for the preparation of 16a, starting from compound 14b (200 mg, 0.49 mmol) in 30% methanolic ammonia (20 mL). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 10:1 , v/v; DCM/MeOH, 8:1 , v/v). ¹H NMR (300 MHz, CDCI₃): δ 7.35 (d, J = 1.2 Hz, 1 H, H-6), 5.81 (m, d, J = 7.2 Hz, 1 H, H-5), 4.76-4.34 (m, 2H, H-3'), 4.18-3.93 (m, 7H, H-r, H-2', 2 x CH2CH3), 3.87-3.66 (m, 2H, PCH₂), 1 .36-1.30 (m, 6H, 2 x CH2CH3); ¹³C NMR (75 MHz, CDCI3): δ 166.5 (C-4), 156.8 (C-2), 146.8 (C-6), 94.5 (C-5), 82.7 (d, ¹J_C,F = 172.9 Hz, C-3'), 78.8 (dd, ²J_C,F = 18.6 Hz, ³J_C,P = 10.6 Hz, C-2'), 64.7 (d, ¹J_c,p = 167.1 Hz, CH₂P), 62.7, 62.6 (2 x d, ²J_c,p = 6.4 Hz, CH2CH3), 50.1 (d, ³J_C,F = 8.4 Hz, C-1 '), 16.6, 16.5 (CH2CH3); ³¹P NMR (121 MHz, CDCI3): δ 20.9; HRMS for C12H21 FN3O5P [M+H]⁺ calcd.: 338.1275, found: 338.1278.

Example 19 : Synthesis of (S)-1 -[3-Fluoro-2-(phosphonomethoxy)propyl]cytosine (17a)

Compound 17a was obtained as a white foam (58 mg, 70%) according to the procedure used for the preparation of 8a, starting from compound 16a (100 mg, 0.30 mmol), bromotrimethylsilane (0.16 mL, 1.19 mmol), and 2,6-lutidine (0.27 mL, 2.37 mmol) in anhydrous acetonitrile (5 mL). The crude residue was purified by column chromatography on silica gel (gradient acetone/H₂0/Et₃N, 6:1 :1 , v/v/v; acetone/H₂0/Et₃N, 5:1 :1 , v/v/v). ¹H NMR (600 MHz, D₂0): δ 7.75-7.74 (m, 1 H, H-6), 6.07-6.06 (m, 1 H, H-5), 4.80-4.48 (m, 1 H, H-3', overlapped with H₂0), 4.14 (dd, J = 14.1 , 3.4 Hz, 1 H, H-1 'a), 3.98-3.88 (m, 2H, H-2', H-1 'b), 3.77 (dd, J = 13.1 , 9.1 Hz, 1 H, PCH₂a), 3.59 (dd, J = 13.1 , 9.6 Hz, 1 H, PCH₂b); ¹³C NMR (75 MHz, D₂0): δ 1 62.4 (C-4), 1 53.0 (C-2), 149.0 (C-6), 94.5 (C-5), 82.0 (d, ¹J_C,F = 1 67.9 Hz, C- 3'), 77.4 (dd, ²JC,F = 18.4 Hz, ³J_c,p = 11 .5 Hz, C-2'), 66.1 (d, ¹J_c,p = 157.2 Hz, CH₂P), 49.1 (d, ³JC,F = 7.9 Hz, C-1 '); ³¹P NMR (121 MHz, D₂0): δ 14.9; HRMS for CsHisFNsOsP [M-H]^" calcd.: 280.0504, found: 280.0501 .

Spectral data are in accordance with literature data (Yu, K. L; Bronson, J. J.; Yang, H.; Patick, A.; Alam, M.; Brankovan, V.; Datema, R.; Hitchcock, M. J. M.; Martin, J.C. J. Med. C em. 1993, 36, 2726-2738).

Example 20 : Synthesis of ( ?)-1 -[3-Fluoro-2-(phosphonomethoxy)propyl]cytosine (17b)

Compound 17b was obtained as a white foam (82 mg, 70%) according to the procedure used for the preparation of 8a, starting from compound 16b (140 mg, 0.42 mmol), bromotrimethylsilane (0.22 mL, 1.66 mmol), and 2,6-lutidine (0.38 mL, 3.32 mmol) in anhydrous acetonitrile (5 mL). The crude residue was purified by column chromatography on silica gel (gradient acetone/H₂0/Et₃N, 6:1 :1 , v/v/v; acetone/H₂0/Et₃N, 5:1 :1 , v/v/v). ¹H NMR (300 MHz, D₂0): δ 7.72 (d, J = 7.6 Hz, 1 H, H-6), 6.05 (d, J = 7.5 Hz, 1 H, H-5), 4.82-4.45 (m, 1 H, H-3', overlapped with H₂0), 4.18-4.12 (m, 1 H, H-1 'a), 4.02-3.75 (m, 3H, H-2', H-1 'b, PCH₂a), 3.64-3.57 (m, 1 H, PCH₂b); ¹³C NMR (75 MHz, D₂0): δ 163.9 (C-4), 155.0 (C-2), 148.4 (C-6), 94.7 (C-5), 82.1 (d, ¹J_C,F = 167.7 Hz, C-3'), 77.6 (dd, ²J_C,F = 18.3 Hz, ³J_c,p = 11 .6 Hz, C- 2'), 66.2 (d, ¹Jc,p = 157.1 Hz, CH₂P), 49.2 (d, ³J_C,F = 7.9 Hz, C-1 '); ³¹P NMR (121 MHz, D₂0): δ 14.9; HRMS for CsHisFNsOsP [M-H]^" calcd.: 280.0504, found: 280.0507.

Spectral data are in accordance with literature data (Yu, K. L.; Bronson, J. J.; Yang, H.; Patick, A.; Alam, M.; Brankovan, V.; Datema, R.; Hitchcock, M. J. M.; Martin, J.C. J. Med. Chem. 1993, 36, 2726-2738).

Example 21 Synthesis of (S)-0²-{3-Fluoro-2-

[(diethylphosphoryl)methoxy]propyl}cytosine (18a)

Compound 18a was obtained as a white foam (100 mg, 82%) according to the procedure used for the preparation of 16a, starting from compound 15a (150 mg, 0.37 mmol) in 30% methanolic ammonia (15 mL). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 25: 1 , v/v; DCM/MeOH, 20: 1 , v/v). ¹ H NMR (300 MHz, CDCI₃): δ 7.92 (dd, J = 5.7 Hz, 1 H, H-6), 6.11 (d, J = 5.7 Hz, 1 H, H-5), 5.71 (s, 2H, NH₂), 4.71 -4.60 (m, 1 H, H- 3'a), 4.56-4.44 (m, 1 H, H-3'b), 4.41 -4.31 (m, 2H, H-1 '), 4.19-3.96 (m, 7H, 2 x CH₂CH₃, H-2', PCH₂), 1 .32-1 .27 (m, 6H, 2 x CH₂CH₃); ¹³C NMR (75 MHz, CDCI₃): δ 165.3 (C-4), 164.7 (C-2), 157.1 (C-6), 100.1 (C-5), 83.0 (d, ¹J_C,F = 172.2 Hz, C-3'), 78.8 (dd, ²J_C,F = 19.0 Hz, ³J_C,P = 11 .3 Hz, C-2'), 64.8 (d, ¹J_c,p = 165.0 Hz, CH₂P), 64.6 (d, ³J_C,F = 8.3 Hz, C-1 '), 62.9, 62.7 (2 x d, ²J_c,p = 6.4 Hz, CH₂CH₃), 16.5, 16.4 (CH₂CH₃); ³¹ P NMR (121 MHz, CDCI₃): δ 20.5; HRMS for Ci₂H₂iFN₃0₅P [M+H]⁺ calcd.: 338.1276, found: 338.1292.

Example 22 Synthesis of ( ?)-0²-{3-Fluoro-2-

[(diethylphosphoryl)methoxy]propyl}cytosine (18b)

Compound 18b was obtained as a white foam (190 mg, 90%) according to the procedure used for the preparation of 16a, starting from compound 15b (250 mg, 0.60 mmol) in 30% methanolic ammonia (20 mL). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 25: 1 , v/v; DCM/MeOH, 20: 1 , v/v). ¹ H NMR (500 MHz, CDCI₃): δ 7.92 (dd, J = 5.7 Hz, 1 H, H-6), 6.11 (d, J = 5.7 Hz, 1 H, H-5), 5.71 (s, 2H, NH₂), 4.67-4.48 (m, 2H, H- 3'), 4.39-4.32 (m, 1 H, H-1 '), 4.17-4.11 (m, 4H, 2 x CH₂CH₃), 4.06-3.98 (m, 3H, H-2', PCH₂), 1 .31 -1 .28 (m, 6H, 2 x CH₂CH₃); ¹³C NMR (125 MHz, CDCI3): δ 165.3 (C-4), 164.7 (C-2), 157.0 (C-6), 100.1 (C-5), 83.0 (d, ¹J_C,F = 172.2 Hz, C-3'), 78.8 (dd, ²J_C,F = 18.9 Hz, ³J_C,P = 11 .3 Hz, C- 2'), 64.8 (d, ¹Jc,p = 162.5 Hz, CH₂P), 64.6 (d, ³J_C,F = 8.5 Hz, C-1 '), 62.9, 62.7 (2 x d, ²J_c,p = 6.6 Hz, CH₂CH₃), 16.5, 16.4 (CH₂CH₃); ³¹ P NMR (202 MHz, CDCI₃): δ 20.5; HRMS for C12H21FN3O5P [M+H]⁺ calcd. : 338.1276, found: 338.1271.

Example 23 : Synthesis of (S)-0²-[3-Fluoro-2-(phosphonomethoxy)propyl]cytosine (19a)

Compound 19a was obtained as a white foam (82 mg, 70%) according to the procedure used for the preparation of 8a, starting from compound 18a (100 mg, 0.30 mmol), bromotrimethylsilane (0.16 mL, 1.20 mmol), and 2,6-lutidine (0.27 mL, 2.37 mmol) in anhydrous acetonitrile (5 mL). The crude residue was purified by column chromatography on silica gel (gradient acetone/H₂0/Et₃N, 5:1 :1 , v/v/v; acetone/H₂0/Et₃N, 4:1 :1 , v/v/v). ¹H NMR (300 MHz, D₂0): 5 7.75 (d, J = 6.0 Hz, 1 H, H-6), 6.16 (d, J = 6.0 Hz, 1 H, H-5), 4.75-4.46 (m, 2H, H-3'), 4.27 (d, J = 5.0 Hz, 2H, H-1 '), 3.98-3.87 (m, 1 H, H-2'), 3.53 (d, J = 9.1 Hz, 2H, PCH₂); ¹³C NMR (75 MHz, D₂0): δ 165.3 (C-4), 163.7 (C-2), 155.6 (C-6), 100.2 (C-5), 82.3 (d, ¹J_C,F = 165.9 Hz, C-3'), 78.9 (dd, ²J_C,F = 18.6 Hz, ³J_C,P = 10.9 Hz, C-2'), 67.8 (d, ¹J_c,p = 149.9 Hz, CH₂P), 64.2 (d, ³JC,F = 7.6 Hz, C-1 '); ³¹P NMR (121 MHz, D₂0): δ 12.7; HRMS for C₈Hi3FN₃0₅P [M-H]- calcd.: 280.0504, found: 280.0505.

Example 24 : Synthesis of ( ?)-0²-[3-Fluoro-2-(phosphonomethoxy)propyl]cytosine

(19b)

Compound 19b was obtained as a white foam (160 mg, 70%) according to the procedure used for the preparation of 8a, starting from compound 18b (200 mg, 0.60 mmol), bromotrimethylsilane (0.32 mL, 2.20 mmol), and 2,6-lutidine (0.54 mL, 4.80 mmol) in anhydrous acetonitrile (10 mL). The crude residue was purified by column chromatography on silica gel (gradient acetone/H₂0/Et₃N, 5:1 :1 , v/v/v; acetone/H₂0/Et₃N, 4:1 :1 , v/v/v). ¹H NMR (300 MHz, D₂0): δ 7.74 (d, J = 6.1 Hz, 1 H, H-6), 6.15 (dd, J = 6.0, 1.0 Hz, 1 H, H-5), 4.75-4.46 (m, 2H, H-3'), 4.33-4.23 (m, 2H, H-1 '), 4.03-3.91 (m, 1 H, H-2'), 3.65 (d, J = 9.2 Hz, 2H, PCH₂); ¹³C NMR (75 MHz, D₂0): δ 165.2 (C-4), 163.4 (C-2), 155.1 (C-6), 100.2 (C-5), 82.3 (d, ¹J_C,F = 166.5 Hz, C-3'), 77.2 (dd, ²J_C,F = 18.6 Hz, ³J_C,P = 10.9 Hz, C-2'), 66.7 (d, ¹J_c,p = 154.2 Hz, CH₂P), 64.3 (d, ³JC,F = 7.7 Hz, C-1 '); ³¹P NMR (121 MHz, D₂0): δ 14.2; HRMS for C₈Hi₃FN₃0₅P [M-H]^" calcd.: 280.0504, found: 280.0513.

Example 25 : Synthesis of (S)-9-{3-Fluoro-2-[(diethylphosphoryl)methoxy]propyl}-0⁶- benzylguanine (21a) O

P-OEt

OEt

Compound 21 a was obtained as a colorless oil (290 mg, 50%) according to the procedure used for the preparation of 6a, starting from compound 4a (300 mg, 1 .23 mmol), compound 20 (360 mg, 1 .47 mmol), Ph₃P (650 mg, 2.46 mmol), and DIAD (0.48 mL, 2.46 mmol) in anhydrous THF (8 mL). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 40: 1 , v/v; DCM/MeOH, 30: 1 , v/v). ¹ H NMR (300 MHz, CDCI₃): δ 7.70 (s, 1 H , H- 8), 7.51 -7.29 (m, 5H, ArH), 5.56 (s, 2H, CH r), 5.01 (s, 2H, NH₂), 4.69-4.28 (m, 3H, H-3', H- 1 'a), 4.20-4.01 (m, 6H, H-1 'b, H-2', 2 x CH₂CH₃), 3.95-3.77 (m, 2H, PCH₂), 1 .34-1 .23 (m, 6H, 2 x CH2CH3); ¹³C NMR (75 MHz, CDCI3): δ 161 .1 (C-6), 159.4 (C-2), 154.2 (C-4), 140.3 (C-8), 136.5 (Ar-C), 128.4 (Ar-C), 128.3 (Ar-C), 128.0 (Ar-C), 115.4 (C-5), 82.1 (d, ¹J_C,F = 173.5 Hz, C-3'), 78.6 (dd, ²J_C,F = 19.3 Hz, ³J_C,P = 10.3 Hz, C-2'), 68.1 (CH₂-Ar), 64.6 (d, ¹J_c,p = 166.9 Hz, CH₂P), 62.7, 62.6 (2 x d, ²J_c,p = 7.0 Hz, CH₂CH₃), 44.3 (d, ³J_C,F = 7.9 Hz, C-1 '), 16.5, 16.4 (CH₂CH₃); ³¹ P N MR (121 MHz, CDCI₃): δ 19.9; HRMS for C₂oH₂7FN₅0₅P [M+H]⁺ calcd.: 468.1806, found: 468.1801 .

Example 26 : Synthesis of ( ?)-9-{3-Fluoro-2-[(diethylphosphoryl)methoxy]propyl}-0⁶- benzylguanine (21 b)

Compound 21 b was obtained as a colorless oil (250 mg, 44%) according to the procedure used for the preparation of 6a, starting from compound 4b (300 mg, 1 .23 mmol), compound 20 (360 mg, 1 .47 mmol), Ph₃P (650 mg, 2.46 mmol), and DIAD (0.48 mL, 2.46 mmol) in anhydrous THF (8 mL). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 40: 1 , v/v; DCM/MeOH, 30: 1 , v/v). ¹ H NMR (300 MHz, CDCI₃): δ 7.70 (s, 1 H, H-8), 7.52-7.28 (m, 5H, ArH), 5.57 (s, 2H, CH r), 4.94 (s, 2H, NH₂), 4.64-4.28 (m, 3H, Η-3', H-1 'a), 4.21 -4.00 (m, 6H, H-1 'b, Η-2', 2 x CH₂CH₃), 3.95-3.77 (m, 2H, PCH₂), 1 .34-1 .24 (m, 6H , 2 x CH₂CH₃); ¹³C NMR (75 MHz, CDCI₃): δ 161 .2 (C-6), 159.4 (C-2), 154.3 (C-4), 140.4 (C-8), 136.5 (Ar-C), 128.4 (d, J = 8.2 Hz, Ar-C), 128.1 (Ar-C), 115.4 (C-5), 82.2 (d, ¹JC,F = 173.5 Hz, C-3'), 78.6 (dd, ²J_C,F = 19.3 Hz, ³J_c,p = 10.1 Hz, C-2'), 68.2 (CH₂-Ar), 64.7 (d, ¹Jc,p = 167.0 Hz, CH₂P), 62.7, 62.6 (2 x d, ²J_c,p = 7.0 Hz, CH₂CH₃), 44.3 (d, ³J_C,F = 8.1 Hz, C- 1 '), 16.5, 16.4 (2 x d, ³Jc,p = 2.9 Hz, CH₂CH₃); ³¹ P N MR (121 MHz, CDCI₃): δ 20.5; HRMS for C₂oH₂₇FN₅0₅P [M+H]⁺ calcd.: 468.1806, found: 468.181 1 .

Example 27: Synthesis

[(diethylphosphoryl)methoxy]propyl}guanine (22a)

Compound 21 a (300 mg, 0.64 mmol) and Pd/C on charcoal (10% w/w, 150 mg) were suspended in EtOAc (20 ml_), and the solution was purged first with nitrogen, then hydrogen, and allowed to stir under a hydrogen atmosphere. After 10 h, the mixture was filtered through a pad of Celite, and the filtrate was evaporated under reduced pressure to give a crude product. The residue was then purified by column chromatography on silica gel (gradient DCM/MeOH, 10:1 , v/v; 7: 1 , v/v) to give 22a (170 mg, 70%) as a colorless foam. ¹ H NMR (300 MHz, MeOD): δ 7.74 (s, 1 H, H-8), 4.77-4.37 (m, 2H, H-3'), 4.33-3.85 (m, 9H, H-1 ', H-2', 2 x CH₂CH₃, PCH₂), 1 .31 -1 .23 (m, 6H, 2 x CH₂CH₃); ¹³C NMR (75 MHz, MeOD): δ 159.5 (C-6), 155.3 (C-2), 153.4 (C-4), 140.4 (C-8), 117.3 (C-5), 83.2 (d, ¹J_C,F = 171 .7 Hz, C-3'), 79.9 (dd, ²J_C,F = 19.1 Hz, ³J_c,p = 11 .9 Hz, C-2'), 64.5 (d, ¹J_c,p = 165.0 Hz, CH₂P), 64.3, 64.1 (2 x d, ²J_c,p = 6.6 Hz, CH₂CH₃), 44.3 (d, ³JC,F = 8.6 Hz, C-1 '), 16.7, 16.6 (CH₂CH₃); ³¹ P NMR (121 MHz, MeOD): δ 21 .1 ; HRMS for Ci₃H₂i FN₅0₅P [M+H]⁺ calcd.: 378.1337, found: 378.1329.

Example 28 Synthesis of ( ?)-9-{3-Fluoro-2-

[(diethylphosphoryl)methoxy]propyl}guanine (22b)

Compound 23b was obtained as a colorless foam (160 mg, 80%) according to the general procedure used for the preparation of 22a, starting from compound 21 b (250 mg, 0.54 mmol) and Pd/C on charcoal (10% w/w, 120 mg) in EtOAc (30 mL). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 10:1 , v/v; 7:1 , v/v). ¹H NMR (300 MHz, MeOD): δ 7.74 (s, 1 H, H-8), 4.76-4.37 (m, 2H, H-3'), 4.33-3.85 (m, 9H, H-1 ', H-2', 2 x CH2CH3, PCH₂), 1 .31 -1.23 (m, 6H, 2 x CH2CH3); ¹³C NMR (75 MHz, MeOD): δ 159.5 (C-6), 155.4 (C-2), 153.4 (C-4), 140.4 (C-8), 117.3 (C-5), 83.2 (d, ¹J_C,F = 171 .6 Hz, C-3'), 79.9 (dd, ²JC,F = 19.0 Hz, ³JC,P = 11 .9 Hz, C-2'), 64.5 (d, ¹J_c,p = 165.0 Hz, CH₂P), 64.3, 64.1 (2 x d, ²J_c,p = 6.6 Hz, CH2CH3), 44.3 (d, ³J_C,F = 8.6 Hz, C-1 '), 16.7, 16.6 (CH2CH3); ³¹P NMR (121 MHz, MeOD): δ 21 .1 ; HRMS for C13H21 FN5O5P [M+H]⁺ calcd.: 378.1337, found: 378.1329.

Example 29 : Synthesis of (S)-9-[3-Fluoro-2-(phosphonomethoxy)propyl]guanine (23a)

Compound 23a was obtained as a white foam (87 mg, 60%) according to the procedure used for the preparation of 8a, starting from compound 22a (170 mg, 0.45 mmol), bromotrimethylsilane (0.24 mL, 1.80 mmol), and 2,6-lutidine (0.42 mL, 3.60 mmol) in anhydrous acetonitrile (5 mL). The crude residue was purified by column chromatography on silica gel (gradient acetone/H₂0/Et₃N, 5:1 :1 , v/v/v; acetone/H₂0/Et₃N, 4:1 :1 , v/v/v). ¹H NMR (300 MHz, D₂0): δ 7.74 (s, 1 H, H-8), 4.79-4.59 (m, 4H, H-3', H-1 '), 3.97-3.88 (m, 1 H, H-2'), 3.51 -3.39 (m, 2H, PCH₂); ¹³C NMR (75 MHz, D₂0): δ 166.8 (C-6), 160.0 (C-2), 151 .2 (C-4), 138.8 (C-8), 116.7 (C-5), 81 .9 (d, ¹J_C,F = 167.4 Hz, C-3'), 77.2 (dd, ²J_C,F = 18.7 Hz, ³J_C,P = 10.6 Hz, C-2'), 68.0 (d, ¹Jc,p = 150.0 Hz, CH₂P), 42.2 (d, ³J_C,F = 7.1 Hz, C-1 '); ³¹P NMR (121 MHz, D₂0): δ 13.6; HRMS for C₉Hi3FN₅0₅P [M-H]^" calcd.: 320.0565, found: 320.0565.

Spectral data are in accordance with literature data (Baszczynski, O.; Hockova, D.; Janeba, Z.; Holy, A.; Jansa, P.; Dracinsky, M.; Keough, D. T; Guddat, L. W. Eur. J. Med. C em. 2013, 67, 81 -89).

Example 30 : Synthesis of ( ?)-9-[3-Fluoro-2-(phosphonomethoxy)propyl]guanine (23b)

Compound 23b was obtained as a white foam (87 mg, 60%) according to the procedure used for the preparation of 8a, starting from compound 22b (160 mg, 0.42 mmol), bromotrimethylsilane (0.22 ml_, 1.70 mmol), and 2,6-lutidine (0.39 ml_, 3.40 mmol) in anhydrous acetonitrile (5 ml_). The crude residue was purified by column chromatography on silica gel (gradient acetone/H₂0/Et₃N, 5:1 :1 , v/v/v; acetone/H₂0/Et₃N, 4:1 :1 , v/v/v). ¹H NMR (300 MHz, D₂0): δ 7.76 (s, 1 H, H-8), 4.70-4.29 (m, 2H, H-3'), 4.25-4.09 (m, 2H, H-1 '), 4.02- 3.88 (m, 1 H, H-2'), 3.65-3.47 (m, 2H, PCH₂); ¹³C NMR (75 MHz, D₂0): δ 158.4 (C-6), 153.3 (C-2), 151 .2 (C-4), 140.3 (C-8), 115.2 (C-5), 81.9 (d, ¹J_C,F = 167.9 Hz, C-3'), 77.5 (dd, ²J_C,F = 18.7 Hz, ³JC,P = 11 .3 Hz, C-2'), 66.4 (d, ¹J_c,p = 156.0 Hz, CH₂P), 42.7 (d, ¹J_C,F = 7.6 Hz, C-1 '); ³¹P NMR (121 MHz, D₂0): δ 14.5; HRMS for C9H13FN5O5P [M-H]^" calcd.: 320.0565, found: 320.0571 .

Examples 31 - 40 : Synthesis of aryloxyphosphonoamidate prodrugs of the free phosphonates 8a-b, 12a-b, 17a-b, 19a-b and 23a-b.

General procedure

The relevant phosphonic acid (1 eq.) was mixed with L-aspartic acid ester HCI salt (1.7 eq.) and phenol (4.4 eq.) in anhydrous pyridine. Then EtsN (10 eq.) was added and the mixture was stirred at 60 °C under a nitrogen atmosphere for 15-20 min. 2,2'-Dithiodipyridine (7 eq.) was mixed in a separate flask with PP i3 (7 eq.) in anhydrous pyridine and the resultant mixture was stirred for 10-15 min to give a clear light yellow solution. This solution was then added to the above solution and the combined mixture was stirred at 60 °C for 12 h. The mixture was then concentrated under reduced pressure to give a residue that was redissolved in EtOAc. This solution was washed with saturated aq. NaHCC and brine, the organic layer was separated, dried over Na₂S0₄, filtered, and concentrated under reduced pressure. The crude residue was purified by silica gel chromatography to give the desired phosphonamidate.

The following compounds were made according to this procedure :

Example 31 : (S)-1 -{3-Fluoro-2-[phenyloxy-bis(amyl-L- aspartvDphosphoryllmethoxylpropyDthymine (24a)

Compound 24a was obtained as a colorless oil (45 mg, 45%) according to the general procedure, starting from compound 8a (50 mg, 0.17 mmol), aspartic acid ester HCI salt (90 mg, 0.29 mmol), PhOH (70 mg, 0.75 mmol), Et₃N (0.24 mL, 1.7 mmol), 2,2'-dithiodipyridine (260 mg, 1.20 mmol), and PPh₃ (310 mg, 1.20 mmol) in anhydrous pyridine (5 mL). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 60:1 , v/v; 50:1 , v/v). ¹H NMR (300 MHz, CD₃CN): δ 9.35 (s, 1 H, NHCO), 7.36-7.13 (m, 6H, H-6, ArH), 4.72-3.65 (m, 13H, H-3', H-2', H-1 ', 2 x OCH₂(CH₂)₃CH₃, PCH₂, H-a-Asp, NHPO), 2.77-2.49 (m, 2H, Η-β-Asp), 1 .74 (d, J = 1 .2 Hz, 3H, CH₃-5), 1.71 (d, J = 1 .2 Hz, 3H, CH₃-5), 1 .60-1.50 (m, 4H, 2 x OCH₂CH2(CH2)2CH₃), 1 .35-1.26 (m, 8H, 2 x 0(CH₂)₂(CH2)2CH₃), 0.91 -0.85 (m, 6H, 2 x 0(CH₂)₄CH₃); ¹³C NMR (75 MHz, CD₃CN): δ 172.9 (d, ³J_c,p = 4.7 Hz, CO-a), 172.6 (d, ³J_c,p = 4.6 Hz, CO-a), 171 .5 (CO-β), 165.3 (C-4), 152.3, 152.2 (C-2), 151 .3 (Ar-C), 143.0 (C-6), 130.7 (Ar-C), 125.9, 125.8 (Ar-C), 121.9 (d, ³J_c,p = 4.3 Hz, Ar-C), 121.7 (d, ³J_c,p = 4.5 Hz, Ar- C), 1 10.4, 110.3 (C-5), 83.5 (d, ¹J_C,F = 172.5 Hz, C-3'), 83.2 (d, ¹J_C,F = 165.0 Hz, C-3'), 79.9, 79.8, 79.7, 79.6, 79.5, 79.4, 79.2 (C-2'), 66.9 (d, ¹J_C,P = 157.5 Hz, CH₂P), 66.5, 66.4, 65.8, 65.7 (OCH₂(CH₂)₃CH₃), 51 .5, 51 .4 (C-1 '), 49.0, 48.8 (C-a-Asp), 39.8 (d, ³J_c,p = 4.0 Hz, C-3-Asp), 39.6 (d, ³Jc,p = 4.1 Hz, C-3-Asp), 29.0, 28.9, 28.7, 28.6 (OCH2(CH₂)2CH₂CH₃), 23.0, 22.9 (0(CH₂)₃CH₂CH₃), 14.3 (0(CH₂)₄CH₃), 12.3 (CH₃-5); ³¹P NMR (121 MHz, CD₃CN): 5 22.3, 21.3; HRMS for C₂9H₄₃FN₃0₉P [M+Na]⁺ calcd.: 650.2613, found: 650.2629.

Example 32 : (/?)-1 -{3-Fluoro-2-[phenyloxy-bis(amyl-L- aspartvDphosphoryllmethoxylpropyDthymine (24b)

Compound 24b was obtained as a colorless oil (80 mg, 50%) according to the general procedure, starting from compound 8b (80 mg, 0.27 mmol), aspartic acid ester HCI salt (142 mg, 0.46 mmol), PhOH (112 mg, 1 .20 mmol), Et₃N (0.38 mL, 2.70 mmol), 2,2'-dithiodipyridine (420 mg, 1.90 mmol), and PPh₃ (500 mg, 1.90 mmol) in anhydrous pyridine (5 mL). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 60:1 , v/v; 50:1 , v/v). ¹H NMR (300 MHz, CD₃CN): δ 9.59 (s, 1 H, NHCO), 7.39-7.13 (m, 6H, H-6, ArH), 4.77-3.71 (m, 13H, H-3', H-2', H-1 ', 2 x OCH₂(CH₂)₃CH₃, PCH₂, H-a-Asp, NHPO), 2.82-2.53 (m, 2H, Η-β-Asp), 1 .79 (s, 3H, CH₃-5), 1.73 (s, 3H, CH₃-5), 1 .64-1 .55 (m, 4H, 2 x OCH₂CH2(CH2)2CH₃), 1 .41 -1 .28 (m, 8H, 2 x 0(CH₂)₂(CH2)2CH₃), 0.97-0.88 (m, 6H, 2 x 0(CH₂)₄CH₃); ¹³C NMR (75 MHz, CD₃CN): δ 172.8 (d, ³J_c,p = 5.0 Hz, CO-a), 172.6 (d, ³J_c,p = 4.5 Hz, CO-a), 171 .6, 171 .5 (CO-β), 165.4 (C-4), 152.3 (C-2), 151 .4, 151 .3 (Ar-C), 143.1 , 143.0 (C-6), 130.7, 130.6 (Ar-C), 125.8 (Ar-C), 121 .8 (d, ³J_c,p = 4.0 Hz, Ar-C), 121 .7 (d, ³J_c,p = 4.4 Hz, Ar-C), 1 10.4, 1 10.3 (C-5), 83.4 (d, J = 172.5 Hz, C-3'), 82.2 (d, ¹J_C,F = 165.0 Hz, C-3'), 79.8, 79.6, 79.5, 79.4, 79.2 (C-2'), 66.9 (d, ¹J_c,p = 157.5 Hz, CH₂P), 66.5, 66.4, 65.7 (OCH₂(CH₂)₃CH₃), 51.5 (C-1 '), 49.0 (d, ²J_c,p = 8.3 Hz, C-a-Asp), 48.8 (d, ²J_c,p = 8.0 Hz, C-a- Asp), 39.7 (d, ³Jc,p = 4.0 Hz, C-3-Asp), 39.6 (d, ³J_c,p = 4.6 Hz, C-3-Asp), 29.0, 28.9, 28.7, 28.6 (OCH2(CH₂)2CH₂CH₃), 23.0, 22.9 (0(CH₂)₃CH₂CH₃), 14.3 (0(CH₂)₄CH₃), 12.3 (CH₃-5); ³¹P NMR (121 MHz, CD₃CN): δ 22.2, 21.5; HRMS for C29H₄3FN₃0₉P [M-H]^" calcd.: 626.2648, found: 626.2648.

Example 33 : (S)-9-{3-Fluoro-2-[phenyloxy-bis(amyl-L- aspartvDphosphoryllmethoxylpropyDadenine (25a)

Compound 25a was obtained as a colorless oil (26 mg, 40%) according to the general procedure, starting from compound 12a (30 mg, 0.10 mmol), aspartic acid ester HCI salt (52 mg, 0.17 mmol), PhOH (40 mg, 0.43 mmol), Et₃N (0.14 mL, 1.00 mmol), 2,2'-dithiodipyridine (150 mg, 0.69 mmol), and PPh₃ (180 mg, 0.69 mmol) in anhydrous pyridine (3 mL). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 25:1 , v/v; 20:1 , v/v). ¹H NMR (300 MHz, CD₃CN): δ 8.25 (s, 1 H, H-2), 8.22 (s, 1 H, H-2), 7.99 (s, 1 H, H- 8), 7.97 (s, 1 H, H-8), 7.37-7.05 (m, 6H, ArH), 6.02 (s, 2H, NH₂), 4.75-3.83 (m, 13H, H-3', H-2', H-1 ', 2 x OCH₂(CH₂)₃CH₃, PCH₂, H-a-Asp, NHPO), 2.79-2.53 (m, 2H, Η-β-Asp), 1 .61 -1.53 (m, 4H, 2 x OCH₂CH2(CH₂)₂CH₃), 1 .36-1 .28 (m, 8H, 2 x 0(CH₂)₂(CH2)2CH₃), 0.93-0.87 (m, 6H, 0(CH₂)₄CH₃); ¹³C NMR (75 MHz, CD₃CN): δ 172.9 (d, ³J_c,p = 4.4 Hz, CO-a), 172.6 (d, ³J_c,p = 4.7 Hz, CO-a), 171 .5, 171 .4 (CO-β), 156.9 (C-6), 153.8, 153.7 (C-2), 151.4, 151 .2, 151.1 (Ar- C, C-4), 142.9, 142.8 (C-8), 130.6 (Ar-C), 125.8, 125.7 (Ar-C), 121 .9 (d, ³J_c,p = 4.4 Hz, Ar-C), 121 .7 (d, ³Jc,p = 4.4 Hz, Ar-C), 120.1 , 120.0 (C-5), 83.3 (d, ¹J_C,F = 165.0 Hz, C-3'), 83.1 (d, ¹J_C,F = 172.5 Hz, C-3'), 79.9, 79.8, 79.7, 79.5, 79.4, 79.3 (C-2'), 67.0 (d, ¹J_c,p = 1 57.5 Hz, CH₂P), 66.7 (d, ¹Jc,p = 1 57.5 Hz, CH₂P), 66.5, 66.4, 65.8, 65.7 (OCH₂(CH₂)3CH₃), 51 .6, 51 .5 (C-1 '), 44.3 (d, ²Jc,p = 2.4 Hz, C-a-Asp), 44.2 (d, ²J_c,p = 2.4 Hz, C-a-Asp), 39.8 (d, ³J_c,p = 3.9 Hz, C- β-Asp), 39.6 (d, ³Jc,p = 4.2 Hz, C-3-Asp), 29.0, 28.9, 28.7, 28.6 (OCH₂(CH₂)₂CH₂CH₃), 23.0, 22.9 (0(CH₂)₃CH₂CH₃), 14.3 (0(CH₂)₄CH₃); ³¹P NMR (121 MHz, CD₃CN): δ 22.2, 21 .2; HRMS for C₂9H₄₂FN₆0₇P [M+H]⁺ calcd.: 637.2909, found: 637.2913.

Example 34 : (/?)-9-{3-Fluoro-2-[phenyloxy-bis(amyl-L- aspartvDphosphoryllmethoxylpropyDadenine (25b)

Compound 25b was obtained as a colorless oil (110 mg, 55%) according to the general procedure, starting from compound 12b (80 mg, 0.26 mmol), aspartic acid ester HCI salt (140 mg, 0.45 mmol), PhOH (110 mg, 1 .15 mmol), Et₃N (0.36 mL, 2.60 mmol), 2,2'-dithiodipyridine (400 mg, 1.83 mmol), and PPh₃ (480 mg, 1.83 mmol) in anhydrous pyridine (5 mL). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 25:1 , v/v; 20:1 , v/v). ¹H NMR (300 MHz, CD₃CN): δ 8.26 (s, 1 H, H-2), 8.05 (s, 1 H, H-8), 8.03 (s, 1 H, H- 8), 7.35-7.01 (m, 5H, ArH), 6.02 (s, 2H, NH₂), 4.87-3.90 (m, 13H, H-3', H-2', H-1 ', 2 x OCH₂(CH₂)₃CH₃, PCH₂, H-a-Asp, NHPO), 2.82-2.56 (m, 2H, H-3-Asp),1.57-1.48 (m, 4H, OCH₂CH₂(CH₂)₂CH₃), 1 .32-1 .24 (m, 8H, 2 x 0(CH₂)₂(CH₂)₂CH₃), 0.90-0.81 (m, 6H, 2 x 0(CH₂)₄CH₃); ¹³C NMR (75 MHz, CD₃CN): δ 172.8 (d, ³J_c,p = 5.0 Hz, CO-a), 172.7 (d, ³J_c,p = 4.4 Hz, CO-a), 171 .5, 171.4 (CO-β), 157.0 (C-6), 153.8 (C-2), 151.3, 151 .2, 151 .1 , 151 .0 (Ar- C, C-4), 142.8 (C-8), 130.6, 130.5 (Ar-C), 125.7 (Ar-C), 121 .7 (d, ³J_c,p = 4.6 Hz, Ar-C), 119.9 (C-5), 83.3 (d, ¹JC,F = 172.5 Hz, C-3'), 83.1 (d, ¹J_C,F = 172.5 Hz, C-3'), 79.7, 79.6, 79.5, 79.4, 79.3 (C-2'), 66.8 (d, ¹J_c,p = 150.0 Hz, CH₂P), 66.6 (d, ¹J_c,p = 157.5 Hz, CH₂P), 66.4, 65.7 (OCH₂(CH₂)₃CH₃), 51.5 (C-1 '), 44.2 (d, ²J_c,p = 8.2 Hz, C-a-Asp), 44.0 (d, ²J_c,p = 7.8 Hz, C-a- Asp), 39.7 (d, ³Jc,p = 3.9 Hz, C-3-Asp), 39.6 (d, ³J_c,p = 3.8 Hz, C-3-Asp), 28.9, 28.8, 28.7, 28.6 (OCH₂(CH₂)₂CH₂CH₃), 23.0, 22.9 (0(CH₂)₃CH₂CH₃), 14.2 (0(CH₂)₄CH₃); ³¹P NMR (121 MHz, CD₃CN): δ 22.1 , 21 .5; HRMS for C₂₉H₄₂FN₆0₇P [M-H]^" calcd.: 635.2764, found: 635.2775.

Example 35_. : (S)-1 -{3-Fluoro-2-[phenyloxy-bis(amyl-L- aspartvDphosphoryllmethoxylpropyDcytosine (26a)

Compound 26a was obtained as a colorless oil (23 mg, 26%) according to the general procedure, starting from compound 17a (40 mg, 0.14 mmol), aspartic acid ester HCI salt (75 mg, 0.24 mmol), PhOH (59 mg, 0.63 mmol), Et₃N (0.20 mL, 1.40 mmol), 2,2'-dithiodipyridine (220 mg, 1.00 mmol), and PPh₃ (260 mg, 1.00 mmol) in anhydrous pyridine (3 mL). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 20:1 , v/v; 10:1 , v/v). ¹H NMR (300 MHz, CD₃CN): δ 7.45-7.16 (m, 6H, ArH, H-6), 5.95 (s, 2H, NH₂), 5.70 (s, 1 H, H-5), 5.68 (s, 1 H, H-5), 4.75-3.66 (m, 13H, H-3', H-2', H-1 ', 2 x OCH₂(CH₂)₃CH₃, PCH₂, H-a-Asp, NHPO), 2.82-2.56 (m, 2H, Η-β-Asp), 1 .63-1.55 (m, 4H, 2 x OCH₂CH₂(CH₂)₂CH₃), 1 .35-1.27 (m, 8H, 2 x 0(CH₂)₂(CH₂)₂CH₃), 0.94-0.88 (m, 6H, 0(CH₂)₄CH₃); ¹³C NMR (75 MHz, CD₃CN): δ 172.9 (d, ³J_c,p = 4.7 Hz, CO-a), 172.7 (d, ³J_c,p = 4.5 Hz, CO-a), 171.5 (CO-β), 167.5 (C-4), 157.5, 157.4 (C-2), 151 .5, 151.4, 151 .3, 151.2 (Ar-C), 148.2 (C-6), 130.7, 130.6 (Ar-C), 125.8, 125.7 (Ar-C), 122.0 (d, ³J_c,p = 4.4 Hz, Ar-C), 121 .8 (d, ³J_c,p = 4.4 Hz, Ar-C), 94.2, 94.1 (C-5), 83.8 (d, ¹JC,F = 172.5 Hz, C-3'), 83.4 (d, ¹J_C,F = 165.0 Hz, C-3'), 80.0, 79.8, 79.7, 79.6, 79.5, 79.4, 79.3 (C-2'), 66.9 (d, J = 150.0 Hz, CH₂P), 66.8 (d, ¹J_c,p = 150.0 Hz, CH₂P), 66.5, 66.4, 65.8, 65.7 (OCH₂(CH₂)₃CH₃), 51.6, 51 .5 (C-1 '), 50.6, 50.5, 50.4 (C-a-Asp), 39.8 (d, ³J_c,p = 4.0 Hz, C-3-Asp), 39.5 (d, ³J_c,p = 4.2 Hz, C-3-Asp), 29.0, 28.9, 28.8, 28.7 (OCH₂(CH₂)₂CH₂CH₃), 23.0, 22.9 (0(CH₂)₃CH₂CH₃), 14.3 (0(CH₂)₄CH₃); ³¹P NMR (121 MHz, CD₃CN): δ 22.4, 21 .4; HRMS for C₂₈H₄₂FN₄0₈P [M+H]⁺ calcd.: 613.2797, found: 613.2802.

Example 36 : (/?)-1 -{3-Fluoro-2-[phenyloxy-bis(amyl-L- aspartvDphosphoryllmethoxylpropyDcytosine (26b)

Compound 26b was obtained as a colorless oil (20 mg, 20%) according to the general procedure, starting from compound 17b (40 mg, 0.14 mmol), aspartic acid ester HCI salt (75 mg, 0.24 mmol), PhOH (59 mg, 0.63 mmol), Et₃N (0.20 mL, 1.40 mmol), 2,2'-dithiodipyridine (220 mg, 1.00 mmol), and PPh₃ (260 mg, 1.00 mmol) in anhydrous pyridine (3 mL). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 20:1 , v/v; 10:1 , v/v). ¹H NMR (300 MHz, CD₃CN): δ 7.45-7.13 (m, 6H, ArH, H-6), 5.74-5.69 (m, 1 H, H-5), 4.78-3.70 (m, 13H, H-3', H-2', H-1 ', 2 x OCH₂(CH₂)₃CH₃, PCH₂, H-a-Asp, NHPO), 2.82-2.56 (m, 2H, Η-β-Asp), 1 .64-1.52 (m, 4H, 2 x OCH₂CH₂(CH₂)₂CH₃), 1 .40-1.24 (m, 8H, 2 x 0(CH₂)₂(CH₂)₂CH₃), 0.93-0.87 (m, 6H, 2 x 0(CH₂)₄CH₃); ¹³C NMR (75 MHz, CD₃CN): δ 172.8 (d, ³Jc,p = 5.1 Hz, CO-a), 172.7 (d, ³J_c,p = 4.4 Hz, CO-a), 171 .6, 171 .5 (CO-β), 167.5 (C-4), 157.5, 157.4 (C-2), 151.5, 151 .3, 151 .2 (Ar-C), 148.2, 148.1 (C-6), 130.6 (Ar-C), 125.8, 125.7 (Ar-C), 121.9 (d, ³J_c,p = 4.4 Hz, Ar-C), 121 .8 (d, ³J_c,p = 4.4 Hz, Ar-C), 94.3, 94.2 (C-5), 88.7 (d, ¹JC,F = 172.5 Hz, C-3'), 83.5 (d, ¹J_C,F = 165 Hz, C-3'), 79.8, 79.7, 79.6, 79.4, 79.3 (C-2'), 66.9 (d, ¹Jc,p = 157.5 Hz, CH₂P), 66.7 (d, ¹J_c,p = 150.0 Hz, CH₂P), 66.5, 66.4, 65.7 (OCH₂(CH₂)3CH₃), 51.5 (C-1 '), 50.5 (d, ²J_c,p = 8.3 Hz, C-a-Asp), 50.4 (d, ²J_c,p = 8.3 Hz, C-a- Asp), 39.8 (d, ³Jc,p = 4.2 Hz, C-3-Asp), 39.6 (d, ³J_c,p = 3.8 Hz, C-3-Asp), 29.0, 28.9, 28.8, 28.7 (OCH₂(CH₂)₂CH₂CH₃), 23.0, 22.9 (0(CH₂)₃CH₂CH₃), 14.3 (0(CH₂)₄CH₃); ³¹P NMR (121 MHz, CD₃CN): δ 22.4, 21 .7; HRMS for C₂₈H₄₂FN₄0₈P [M-H]^" calcd.: 61 1 .2651 , found: 61 1 .2655.

Example 37 : (S)-0²-{3-Fluoro-2-[phenyloxy-bis(amyl-L- aspartvDphosphoryllmethoxylpropyDcytosine (27a)

Compound 27a was obtained as a colorless oil (12 mg, 28%) according to the general procedure, starting from compound 19a (20 mg, 0.07 mmol), aspartic acid ester HCI salt (38 mg, 0.12 mmol), PhOH (30 mg, 0.32 mmol), Et₃N (0.10 mL, 0.70 mmol), 2,2'-dithiodipyridine (110 mg, 0.50 mmol), and PPh₃ (130 mg, 0.50 mmol) in anhydrous pyridine (3 mL). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 40:1 , v/v; 30:1 , v/v). ¹H NMR (300 MHz, CD₃CN): δ 7.95-7.93 (m, 1 H, H-6), 7.38-7.16 (m, 5H, ArH), 6.19 (s, 1 H, H-5), 6.17 (s, 1 H, H-5), 5.61 (s, 2H, NH₂), 4.75-3.93 (m, 13H, H-3', H-2', H-1 ', 2 x OCH₂(CH₂)₃CH₃, PCH₂, H-a-Asp, NHPO), 2.86-2.63 (m, 2H, Η-β-Asp), 1 .64-1.52 (m, 4H, 2 x OCH₂CH₂(CH₂)₂CH₃), 1 .34-1 .27 (m, 8H, 2 x 0(CH₂)₂(CH₂)₂CH₃), 0.93-0.87 (m, 6H, 2 x 0(CH₂)₄CH₃); ¹³C NMR (75 MHz, CD₃CN): δ 173.0 (d, ³J_c,p = 4.4 Hz, CO-a), 172.8 (d, ³J_c,p = 4.5 Hz, CO-a), 171 .5 (CO-β), 166.6 (C-4), 165.9 (C-2), 158.0 (C-6), 151 .3 (Ar-C), 130.6 (Ar- C), 125.8 (Ar-C), 122.0 (d, ³J_c,p = 4.0 Hz, Ar-C), 121 .9 (d, ³J_c,p = 4.3 Hz, Ar-C), 100.6 (C-5), 83.9 (d, ¹JC,F = 168.0 Hz, C-3'), 82.7 (d, ¹J_C,F = 168.0 Hz, C-3'), 80.1 , 80.0, 79.9, 79.7, 79.6, 79.5, 79.3 (C-2'), 66.9 (d, ¹J_c,p = 152.3 Hz, CH₂P), 66.7 (d, ¹J_c,p = 150.0 Hz, CH₂P), 66.5, 66.4, 65.7, 65.4, 65.3, 65.2 (OCH₂(CH₂)3CH₃, C-1 '), 51.7, 51 .5 (C-a-Asp), 39.8 (d, ³J_c,p = 3.9 Hz, C- β-Asp), 39.6 (d, ³Jc,p = 4.2 Hz, Ο-β-Asp), 29.0, 28.9, 28.8, 28.7 (OCH₂(CH₂)₂CH₂CH₃), 23.0, 22.9 (0(CH₂)₃CH₂CH₃), 14.3 (0(CH₂)₄CH₃); ³¹P NMR (121 MHz, CD₃CN): δ 22.5, 21.9; HRMS for C₂₈H₄₂FN₄0₈P [M-H]^" calcd.: 611.2651 , found: 611.2642.

Example 38 : (/?)-0²-{3-Fluoro-2-[phenyloxy-bis(amyl-L- aspartvDphosphoryllmethoxylpropyDcytosine (27b)

Compound 27b was obtained as a colorless oil (31 mg, 24%) according to the general procedure, starting from compound 19b (60 mg, 0.21 mmol), aspartic acid ester HCI salt (114 mg, 0.36 mmol), PhOH (90 mg, 0.96 mmol), Et₃N (0.30 mL, 2.10 mmol), 2,2'-dithiodipyridine (330 mg, 1.50 mmol), and PPh₃ (390 mg, 1.50 mmol) in anhydrous pyridine (5 mL). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 40:1 , v/v; 30:1 , v/v). ¹H NMR (300 MHz, CD₃CN): δ 7.94 (s, 1 H, H-6), 7.92 (s, 1 H, H-6), 7.39-7.19 (m, 5H, ArH), 6.18 (s, 1 H, H-5), 6.16 (s, 1 H, H-5), 5.73 (s, 2H, NH₂), 4.75-3.93 (m, 13H, H-3', H-2', H-r, 2 x OCH₂(CH₂)₃CH₃, PCH₂, H-a-Asp, NHPO), 2.86-2.59 (m, 2H, Η-β-Asp), 1 .63-1.51 (m, 4H, 2 x OCH₂CH₂(CH₂)₂CH₃), 1 .39-1.24 (m, 8H, 2 x 0(CH₂)₂(CH₂)₂CH₃), 0.95-0.87 (m, 6H, 2 x 0(CH₂)₄CH₃); ¹³C NMR (75 MHz, CD₃CN): δ 172.8, 172.7 (CO-a), 171 .6, 171 .5 (CO-β), 166.6 (C-4), 165.9 (C-2), 157.9 (C-6), 151.4 (Ar-C), 130.6 (Ar-C), 125.8, 125.7 (Ar-C), 122.0 (d, ³J_c,p = 4.1 Hz, Ar-C), 121.9 (d, ³J_c,p = 4.4 Hz, Ar-C), 100.7, 100.6 (C-5), 83.8 (d, ¹J_C,F = 165.0 Hz, C-3'), 80.2, 80.0, 79.9, 79.7, 79.6, 79.5, 79.3 (C-2'), 67.0 (d, ¹J_c,p = 157.5 Hz, CH₂P), 66.6 (d, ¹Jc,p = 150.0 Hz, CH₂P), 66.5, 66.4, 65.7, 65.6 (OCH₂(CH₂)₃CH₃), 65.3 (d, ³J_C,F = 7.5 Hz, C-1 '), 51.5 (C-a-Asp), 39.8 (d, ³J_c,p = 4.0 Hz, ΰ-β-Αβρ), 39.6 (d, ³J_c,p = 3.9 Hz, ΰ-β-Αβρ), 29.0, 28.9, 28.8, 28.7 (OCH₂(CH₂)₂CH₂CH₃), 23.0, 22.9 (0(CH₂)₃CH₂CH₃), 14.3 (0(CH₂)₄CH₃); ³¹P NMR (121 MHz, CD₃CN): δ 22.8, 21.8; HRMS for C₂₈H₄₂FN₄0₈P [M-H]^" calcd.: 61 1 .2651 , found: 611.2650.

Example 39 : (S)-9-{3-Fluoro-2-[phenyloxy-bis(amyl-L- aspartvDphosphoryllmethoxylpropyDguanine (29a)

According to the general procedure, starting from compound 23a (30 mg, 0.09 mmol), aspartic acid ester HCI salt (50 mg, 0.16 mmol), PhOH (40 mg, 0.41 mmol), Et₃N (0.13 mL, 0.90 mmol), 2,2'-dithiodipyridine (145 mg, 0.65 mmol), and PPh₃ (170 mg, 0.65 mmol) in anhydrous pyridine (3 mL). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 40:1, v/v; 30:1, v/v) to give the triphosphine adduct of compound 29a. The collected eluate was evaporated and redissolved in CH3CN/H2O (1 :1 , v/v, 10 mL). One drop of a 1 M HCI solution was added at 0 °C and then the mixture was stirred for another 1 h at room temperature. The solution was neutralized with 2 M TEAB. After removal of all the volatiles, the resulting residue was purified by RP-HPLC (CH3CN, H2O) to give compound 29a (3 mg, 5%) as a colorless oil.¹H NMR(300 MHz, CD₃CN): 57.64 (s, 1H, H-8), 7.61 (s, 1H, H-8), 7.38-7.13 (m, 5H, ArH), 4.74-3.92 (m, 13H, H-3', H-2', H-1', 2 x OCH₂(CH₂)₃CH₃, PCH₂, H-a-Asp, NHPO), 2.82-2.54 (m, 2H, Η-β-Asp), 1.61-1.52 (m, 4H, 2 x OCH₂CH2(CH₂)2CH₃), 1.33-1.26 (m, 8H, 2 x 0(CH₂)2(CH2)2CH₃), 0.92-0.85 (m, 6H, 0(CH₂)₄CH₃); ¹³C NMR (75 MHz, CD₃CN): δ 172.8, 172.7 (CO-a), 171.6, 171.5 (CO-β), 154.7, 154.6 (C-2), 152.8 (C-4), 151.2 (Ar-C), 139.3 (C-8), 130.6 (Ar-C), 125.9, 125.8 (Ar-C), 121.9 (d, ³J_c,p = 4.3 Hz, Ar-C), 121.7 (d, ³J_c,p = 4.4 Hz, Ar- C), 118.3 (C-5, overlapped with CD₃CN), 83.5 (d, ¹J_C,F = 172.5 Hz, C-3'), 83.2 (d, ¹J_C,F = 165.0 Hz, C-3'), 79.7, 79.6, 79.5 (C-2'), 66.7 (d, ¹J_c,p = 157.5 Hz, CH₂P), 66.6, 66.5, 65.8, 65.7 (OCH₂(CH₂)₃CH₃), 51.6, 51.5 (C-1'), 43.8 (C-a-Asp), 39.8, 39.6 (C-3-Asp), 29.0, 28.9, 28.8, 28.7 (OCH2(CH₂)2CH₂CH₃), 23.0, 22.9 (0(CH₂)₃CH₂CH₃), 14.3 (0(CH₂)₄CH₃); ³¹P NMR (121 MHz, CD₃CN): δ 22.3, 21.5; HRMS for C29H₄2FN₆0₈P [M-H]^" calcd.: 651.2713, found: 651.2733.

Example 40: (/?)-9-{3-Fluoro-2-[phenyloxy-bis(amyl-L- aspartvDphosphoryllmethoxylpropyDguanine (29b)

According to the general procedure, starting from compound 23b (60 mg, 0.18 mmol), aspartic acid ester HCI salt (100 mg, 0.32 mmol), PhOH (80 mg, 0.82 mmol), Et₃N (0.26 mL, 1.80 mmol), 2,2'-dithiodipyridine (290 mg, 1.30 mmol), and PPh₃ (340 mg, 1.30 mmol) in anhydrous pyridine (5 mL). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 40:1 , v/v; 30:1 , v/v) to give the triphosphine adduct of compound 28b. The collected eluate was evaporated and redissolved in CH3CN/H2O (1 :1 , v/v, 10 mL). One drop of 1 M HCI solution was added at 0 °C and then the mixture was stirred for another 1 h at room temperature. The solution was neutralized with 2 M TEAB. After removal of all the volatiles, the resulting residue was purified by RP-HPLC (CH3CN, H2O) to give compound 29b (5 mg, 4%) as a colorless oil. ¹H NMR (300 MHz, CD₃CN): δ 7.67 (s, 1 H, H-8), 7.65 (s, 1 H, H- 8), 7.38-7.09 (m, 5H, ArH), 4.76-3.94 (m, 13H, H-3', H-2', H-1 ', 2 x OCH₂(CH₂)3CH₃, PCH₂, H- a-Asp, NHPO), 2.81 -2.56 (m, 2H, Η-β-Asp), 1 .59-1.52 (m, 4H, 2 x OCHzC ^CHz^CHs), 1 .32- 1 .28 (m, 8H, 2 x 0(CH₂)2(CH2)2CH₃), 0.92-0.86 (m, 6H, 2 x 0(CH₂)₄CH₃); ¹³C NMR (75 MHz, CD3CN): δ 172.8 (CO-a), 171.6 (CO-β), 158.6 (C-6), 154.6 (C-2), 152.7 (C-4), 151 .4 (Ar-C), 139.3 (C-8), 130.7, 130.6 (Ar-C), 125.9 (Ar-C), 121 .8 (t, ³J_C,P = 4.7 Hz, Ar-C), 1 18.3 (C-5, overlapped with CD₃CN), 83.4 (d, ¹J_C,F = 172.5 Hz, C-3'), 83.2 (d, ¹J_C,F = 165.0 Hz, C-3'), 79.8, 79.6, 79.5, 79.4 (C-2'), 66.9 (d, ¹J_c,p = 150.0 Hz, CH₂P), 66.6 (d, ¹J_c,p = 157.5 Hz, CH₂P), 66.5, 65.8 (OCH₂(CH₂)3CH₃), 51 .5 (C-1 '), 43.8 (C-a-Asp), 39.8, 39.6 (C-3-Asp), 29.0, 28.9, 28.8, 28.7 (OCH₂(CH2)2CH₂CH3), 23.0, 22.9 (0(CH2)3CH₂CH₃), 14.3 (0(CH₂)₄CH₃); ³¹P NMR (121 MHz, CD3CN): δ 22.3, 21.7; HRMS for C29H₄2FN₆0₈P [M-H]^" calcd.: 651 .2726, found: 651 .2733.

( ?)-2-Amino-6-chloro-9-{2-[(diethylphosphoryl)-methoxyl]-3-

A solution of DIAD (0.24 mL, 1.20 mmol) in anhydrous THF (1 mL) was added dropwise to a mixture of 30 (120 mg, 0.74 mmol), 4a (150 mg, 0.61 mmol), and Ph₃P (300 mg, 1 .20 mmol) in anhydrous THF (5 mL) at room temperature. The reaction mixture was stirred for 24 h, then H2O (5 mL) was added. The resulting mixture was refluxed for 24 h (to remove triphenylphosphine adducts). After removal of all the volatiles, the crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 50:1 , v/v; 30:1 , v/v; 25:1 , v/v) to give 31 a (200 mg, 83%) as a colorless oil. ¹H NMR (300 MHz, CDCI₃): δ 7.91 (s, 1 H, H-8), 5.50 (s, 1 H, NH₂), 4.71 -4.35 (m, 3H, H-3 ' , H-1 'a), 4.26-3.92 (m, 7H, H-1 'b, H-2 ' , 2 x CH₂CH₃, PCH₂a), 3.83 (dd, J = 13.9, 8.8 Hz, 1 H, PCH₂b), 1 .35-1.26 (m, 6H, 2 x CH2CH3); ¹³C NMR (75 MHz, CDCI3): δ 159.5 (C-2), 154.1 (C-4), 151.5 (C-6), 143.5 (C-8), 125.1 (C-5), 82.0 (d, ¹J_c,p = 173.9 Hz, C-3 ' ), 78.4 (dd, ²J_C,F = 19.5 Hz, ³J_C,P = 9.6 Hz, C-2 ' ), 64.7 (d, ¹J_c,p = 167.1 Hz, CH₂P), 62.9, 62.8 (CH2CH3), 43.8 (d, ³J_C,F = 8.0 Hz, C-1 ' ), 16.7,16.6 (CH2CH3); ³¹P NMR (121 MHz, CDCIs): δ 20.5; HRMS for Ci3H₂oCIFN₅0₄P [M+H]⁺ calcd.: 396.0998, found: 396.1002.

Example 42 3.2.2 (S)-2-Amino-6-chloro-9-{2-[(diethylphosphoryl)-methoxyl]-3- fluoropropyl}purine (31 b)

Compound 31 b was obtained as a colorless oil (300 mg, 70%) according to the procedure used for the preparation of 31 a, starting from 30 (250 mg, 1 .47 mmol), 4b (300 mg, 1 .23 mmol), Ph₃P (640 mg, 2.46 mmol), and DIAD (0.48 mL, 2.46 mmol) in anhydrous THF (10 mL). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 50:1 , v/v; 30:1 , v/v; 25:1 , v/v). ¹H NMR (300 MHz, CDCI₃): δ 7.90 (s, 1 H, H-8), 5.40 (s, 1 H, NH₂), 4.71 -4.34 (m, 3H, H-3 ' , H-1 'a), 4.25-3.79 (m, 8H, H-1 'b, H-2 ' , 2 x CH2CH3, PCH₂), 1 .35-1.24 (m, 6H, 2 x CH2CH3); ¹³C NMR (75 MHz, CDCI3): δ 159.5 (C-2), 154.2 (C-4), 151 .5 (C-6), 143.6 (C-8), 125.1 (C-5), 82.0 (d, ¹J_c,p = 173.7 Hz, C-3 ' ), 78.4 (dd, ²J_C,F = 19.7 Hz, ³Jc,p = 9.5 Hz, C-2 ' ), 64.8 (d, ¹J_c,p = 167.0 Hz, CH₂P), 62.9, 62.8 (CH2CH3), 43.8 (d, ³J_C,F = 7.8 Hz, C-1 ' ), 16.7,16.6 (CH2CH3); ³¹P NMR (121 MHz, CDCI3): δ 20.5; HRMS for Ci₃H₂oCIFN₅0₄P [M+H]⁺ calcd.: 396.0998, found: 396.0998.

Example 43 : ( ?)-2-Amino-6-(4-methoxyphenylthio)-9-{2-[(diethylphosphoryl)methoxyl]- 3-fluoropropyl}purine (32a)

4-Methoxythiophenol (0.12 mL, 0.96 mmol) was added to a mixture of 31a (190 mg, 0.48 mmol) and triethylamine (0.07 mL, 0.48 mmol) in anhydrous DMF (10 mL) at room temperature and the mixture was stirred at 100 °C for further 4 h. The solution was diluted with EtOAc (100 mL) and washed with saturated aq. NaHCC (50 mL) and brine (50 mL). The organic layer was dried with Na2S0₄, filtered and concentrated under reduced pressure. The residue was then purified by column chromatography on silica gel (gradient DCM/MeOH, 60:1 , v/v; 50:1 , v/v; 40:1 , v/v) to give 32a (210 mg, 87%) as a colorless oil. ¹H NMR (600 MHz, CDCI₃): δ 7.76 (s, 1 H, H-8), 7.52 (d, J = 8.9 Hz, 2H, ArH), 6.94 (d, J = 8.9 Hz, 2H, ArH), 4.88 (s, 2H, NH₂), 4.64- 4.38 (m, 2H, H-3 ' ), 4.33-4.28 (m, 1 H, H-1 'a), 4.19-4.04 (m, 6H, H-1 'b, H-2 ' , 2 x CH₂CH₃), 3.91 (dd, J = 13.9, 8.7 Hz, 1 H, PCH₂a), 3.85-3.77 (m, 4H, OCH₃, PCH₂b), 1 .34-1 .27 (m, 6H, 2 x CH2CH3); ¹³C NMR (150 MHz, CDC ): δ 161 .7 (C-2), 160.4 (Ar-C), 159.1 (C-6), 150.9 (C-4), 141.0 (C-8), 137.1 (Ar-C), 124.6 (C-5), 1 18.0 (Ar-C), 114.5 (Ar-C), 82.0 (d, ¹J_c,p = 173.6 Hz, C- 3 ' ), 78.3 (dd, ²JC,F = 19.2 Hz, ³J_C,P = 9.9 Hz, C-2 ' ), 64.5 (d, ¹J_c,p = 166.9 Hz, CH₂P), 62.5, 62.4 (d, ²Jc,p = 6.4 Hz, CH₂CH₃), 55.3 (OCH₃), 43.1 (d, ³J_C,F = 7.9 Hz, C-1 ' ), 16.4,16.3 (CH₂CH₃); ³¹ P N MR (121 MHz, CDCI₃): δ 19.9; HRMS for C₂oH₂7FN₅0₅PS [M+H]⁺ calcd.: 500.1527, found: 500.1526.

Example 44 : (S)-2-Amino-6-(4-methoxyphenylthio)-9-{2-[(diethylphosphoryl)methoxyl]- 3-fluoropropyl}purine (32b)

Compound 32b was obtained as a colorless oil (160 mg, 67%) according to the procedure used for the preparation of 32a, starting from 31 b (190 mg, 0.48 mmol), 4-methoxythiophenol (0.12 mL, 0.96 mmol) and triethylamine (0.07 mL, 0.48 mmol) in anhydrous DMF (10 mL). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 60:1 , v/v; 50:1 , v/v; 40:1 , v/v). ¹H NMR (300 MHz, CDCI3): δ 7.77 (s, 1 H, H-8), 7.53 (d, J = 8.8 Hz, 2H, ArH), 6.95 (d, J = 8.8 Hz, 2H, ArH), 5.03 (s, 1 H, NH₂), 4.70-4.30 (m, 3H, H-3 ' , H-1 'a), 4.22-3.98 (m, 6H, H-1 'b, H-2 ' , 2 x CH₂CH₃), 3.93-3.77 (m, 5H, PCH₂, OCH₃), 1 .35-1.26 (m, 6H, 2 x CH₂CH₃); ¹³C N MR (75 MHz, CDCI3): δ 162.1 (C-2), 160.9 (Ar-C), 159.0 (C-6), 151.0 (C-4), 141 .7 (C-8), 137.5 (Ar-C), 124.9 (C-5), 1 18.0 (Ar-C), 115.0 (Ar-C), 82.3 (d, ¹J_c,p = 173.7 Hz, C-3 ' ), 78.7 (dd, ²J_C,F = 19.5 Hz, ³J_C,P = 9.9 Hz, C-2 ' ), 64.9 (d, ¹J_c,p = 166.9 Hz, CH₂P), 62.9, 62.8 (CH₂CH₃), 55.6 (OCH₃), 43.6 (d, ³J_C,F = 8.1 Hz, C-1 ' ), 16.8,16.7 (CH₂CH₃); ³¹ P NMR (121 MHz, CDCb): δ 20.5; HRMS for C₂oH₂7FN₅0₅PS [M+H]⁺ calcd.: 500.1527, found: 500.1518.

Example 45 : 4-Chloro-5-fluoro-2-pivaloylamino-7H-pyrrolo[2,3-rflpyrimidine (35)

4-Chloro-2-pivaloylamino-7H-pyrrolo[2,3-d]pyrimidine 34 (150 mg, 0.59 mmol) and selectfluor (320 mg, 0.90 mmol) were placed in a round-bottom flask, followed by the addition of anhydrous MeCN (6 mL) and AcOH (0.6 ml_). The mixture was then heated at 50 °C for 30 min under N2. The mixture was then cooled by ice-water and diluted with CH2CI2 (50 mL). The organic layer was washed with sat. NaHCC>3 (50 mL). The inorganic layer was separated and extracted with CH2CI2 (2 ^χ 20 ml). The combined organic layers were dried with Na2S0₄, filtered and concentrated under reduce pressure. The residue was then purified by column chromatography on silica gel (gradient DCM/MeOH, 200:1 ) to give 35 (48 mg, 30%) as a colorless foam. ¹H NMR (300 MHz, CDCI₃): δ 11.79 (s, 1 H, NHCO), 8.25 (s, 1 H, H-6), 1.39 (s, 9H, (CH₃)₃C); ¹³C NMR (75 MHz, CDCI3): 176.5 (NHCO), 151.3 (C-2), 150.8 (C-4), 147.8 (C- 7a), 141 .4 (d, ¹J_C,F = 248.8 Hz, C-5), 11 1.2 (d, ²J_C,F = 26.0 Hz, C-6), 103.6 (d, ²J_C,F = 14.9 Hz, C-4a), 40.5 (C(CH₃)₃), 27.7 (C(CH₃)₃).

Example 46 ( ?)-4-Chloro-7-{2-[(diethylphosphoryl)-methoxyl]-3-fluoropropyl}-2- pivaloylamino-7H-pyrrolo[2,3-rflpyrimidin (36a)

A solution of DIAD (0.24 mL, 1.20 mmol) in anhydrous THF (1 mL) was added dropwise to a mixture of 34 (180 mg, 0.74 mmol), 4a (150 mg, 0.61 mmol), and Ph₃P (300 mg, 1 .20 mmol) in anhydrous THF (10 mL) at room temperature. The reaction mixture was stirred for 24 h, and it was then concentrated under reduced pressure. The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 60:1 , v/v; 40:1 , v/v; 30:1 , v/v) to give 36a (220 mg, 78%) as a colorless oil. ¹H NMR (300 MHz, CDCI₃): δ 8.16 (s, 1 H, NHCO), 7.29(dd, J = 0.9 Hz, 1 H, H-6), 6.53 (dd, J = 3.6, 0.9 Hz, 1 H, H-6), 4.74-4.31 (m, 4H, H-3 ' , H-1 ' ), 4.18- 3.86 (m, 7H, H-2', 2 x CH₂CH₃, PCH₂), 1 .36-1.27 (m, 15H, (CH₃)₃C, 2 x CH2CH3); ¹³C NMR (150 MHz, CDCI₃): δ 175.7 (NHCO), 152.3 (C-4, C-7a), 151 .6 (C-2), 130.1 (C-6), 114.2 (C-4a), 100.1 (C-5), 82.9 (d, ¹J_c,p = 173.2 Hz, C-3 ' ), 79.4 (dd, ²J_C,F = 19.1 Hz, ³J_C,P = 10.9 Hz, C-2 ' ), 64.9 (d, ¹Jc,p = 166.9 Hz, CH₂P), 62.7, 62.6 (CH₂CH₃), 44.9 (d, ³J_C,F = 8.5 Hz, C-1 ' ), 40.5 (C(CH₃)₃), 27.7 (C(CH₃)₃), 16.7, 16.6 (CH₂CH₃); ³¹ P NMR (121 MHz, CDCI₃): δ 20.9; HRMS for Ci₉H₂9CIFN₄0₅P [M+H]⁺ calcd.: 479.1621 , found: 479.1617.

Example 47 (S)-4-Chloro-7-{2-[(diethylphosphoryl)-methoxyl]-3-fluoropropyl}-2- pivaloylamino-7H-pyrrolo[2,3-rflpyrimidin (36b)

Compound 36b was obtained as a colorless oil (230 mg, 79%) according to the procedure used for the preparation of 36a, starting from 34 (180 mg, 0.74 mmol), 4b (150 mg, 0.61 mmol), Ph₃P (300 mg, 1 .20 mmol), and DIAD (0.24 mL, 1.20 mmol) in anhydrous THF (10 mL). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 60:1 , v/v; 40:1 , v/v; 30:1 , v/v). ¹H NMR (600 MHz, CDCI₃): δ 8.13 (s, 1 H, NHCO), 7.30 (dd, J = 3.6, 2.0 Hz, 1 H, H-6), 6.53 (dd, J = 3.6, 2.5 Hz, 1 H, H-6), 4.69-4.59 (m, 1 H, H-3 ' a), 4.55-4.51 (m, H-1 ' a), 4.46-4.33 (m, 2H, H-3 ' b, H-1 ' b), 4.15-4.08 (m, 5H, H-2', 2 x CH₂CH₃), 4.00- 3.96 (m, 1 H, PCH₂a), 3.92-3.88 (m, 1 H, PCH₂b), 1 .35-1 .29 (m, 15H, (CH₃)₃C, 2 x CH₂CH₃); ¹³C NMR (150 MHz, CDCI₃): δ 175.5 (NHCO), 152.1 (C-4, C-7a), 151.3 (C-2), 129.8 (C-6), 1 13.9 (C-4a), 99.8 (C-5), 82.5 (d, ¹J_c,p = 173.0 Hz, C-3 ' ), 79.1 (dd, ²J_C,F = 19.0 Hz, ³J_C,P = 1 1.1 Hz, C-2 ' ), 64.5 (d, ¹J_c,p = 167.2 Hz, CH₂P), 62.5, 62.4 (CH₂CH₃), 44.5 (d, ³J_C,F = 8.5 Hz, C-1 ' ), 40.2 (C(CH₃)₃), 27.4 (C(CH₃)₃), 16.4 (CH₂CH₃); ³¹P NMR (121 MHz, CDCI₃): δ 20.9; HRMS for Ci9H₂₉CIFN40₅P [M+H]⁺ calcd.: 479.1621 , found: 479.1624.

Example 48 : ( ?)-4-Chloro-5-fluoro-7-{2-[(diethylphosphoryl)-methoxyl]-3-fluoropropyl}- 2-pivaloylamino-7H-pyrrolo[2,3-d]pyrimidin (37a)

Compound 37a was obtained as a colorless oil (70 mg, 25%) according to the procedure used for the preparation of 36a, starting from 35 (200 mg, 0.74 mmol), 4a (150 mg, 0.61 mmol), Ph₃P (300 mg, 1 .20 mmol), and DIAD (0.24 mL, 1.20 mmol) in anhydrous THF (10 mL). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 80:1 , v/v; 60:1 , v/v; 40:1 , v/v). ¹H NMR (300 MHz, CDCI₃): 5 8.13 (s, 1 H, NHCO), 7.10 (d, J = 2.6 Hz, 1 H, H-6), 4.72-3.96 (m, 9H, H-3 ' , H-1 ' , H-2', 2 x CH₂CH₃), 4.00 (dd, J = 13.8, 8.7 Hz, 1 H, PCH₂a), 3.88 (dd, J = 13.8, 9.2 Hz, 1 H, PCH₂b), 1.34-1.28 (m, 15H, (CH₃)₃C, 2 x CH₂CH₃); ¹³C NMR (125 MHz, CDCI₃): δ 175.6 (NHCO), 152.0 (C-2), 150.9 (d, ³J_C,F = 4.2 Hz, C-4), 147.9 (C-7a), 141.6 (d, ¹J_C,F = 252.5 Hz, C-5), 112.5 (d, ²J_C,F = 26.5 Hz, C-6), 103.6 (d, ²JC,F = 14.9 Hz, C-4a), 82.6 (d, ¹J_c,p = 173.3 Hz, C-3 ' ), 79.5, 79.4, 79.2, 79.1 (C-2 ' ), 64.8 (d, ¹ Jc,p = 166.9 Hz, CH₂P), 62.7, 62.6 (CH₂CH₃), 44.4 (d, ³J_C,F = 8.5 Hz, C-1 ' ), 40.5 (C(CH₃)₃), 27.6 (C(CH₃)₃), 16.6, 16.5 (CH₂CH₃); ³¹P NMR (121 MHz, CDCI₃): δ 20.9; HRMS for Ci9H28CIF₂N₄0₅P [M+H]⁺ calcd.: 497.1527, found: 497.1523.

Example 49 : (S)-4-Chloro-5-fluoro-7-{2-[(diethylphosphoryl)-methoxyl]-3-fluoropropyl}- 2-pivaloylamino-7H-pyrrolo[2,3-rflpyrimidin (37b)

Compound 37b was obtained as a colorless oil (100 mg, 33%) according to the procedure used for the preparation of 36a, starting from 35 (200 mg, 0.74 mmol), 4b (150 mg, 0.61 mmol), Ph₃P (300 mg, 1 .20 mmol), and DIAD (0.24 mL, 1.20 mmol) in anhydrous THF (10 mL). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 80:1 , v/v; 60:1 , v/v; 40:1 , v/v). ¹H NMR (600 MHz, CDCI₃): 5 8.13 (s, 1 H, NHCO), 7.10 (d, J = 2.5 Hz, 1 H, H-6), 4.67-4.25 (m, 4H, H-3 ' , H-1 ' ), 4.18-4.02 (m, 5H, H-2', 2 x CH₂CH₃), 4.00 (dd, J = 13.8, 8.7 Hz, 1 H, PCH₂a), 3.89 (dd, J = 13.8, 9.2 Hz, 1 H, PCH₂b), 1 .34-1.30 (m, 15H, (CH₃)₃C, 2 x CH₂CH₃); ¹³C NMR (125 MHz, CDCI₃): δ 175.5 (NHCO), 151 .8 (C-2), 150.7 (C- 4), 147.6 (C-7a), 141.3 (d, ¹J_C,F = 252.3 Hz, C-5), 112.4 (d, ²J_C,F = 26.3 Hz, C-6), 103.4 (d, ²J_C,F = 14.9 Hz, C-4a), 82.4 (d, ¹J_c,p = 173.3 Hz, C-3 ' ), 79.2, 79.0 (C-2 ' ), 64.5 (d, ¹J_c,p = 167.0 Hz, CH₂P), 62.5 (CH₂CH₃), 44.1 (d, ³J_C,F = 8.5 Hz, C-1 ' ), 40.3 (C(CH₃)₃), 27.4 (C(CH₃)₃), 16.4 (CH₂CH₃); ³¹P NMR (121 MHz, CDCI₃): δ 20.8; HRMS for Ci₉H₂₈CIF₂N₄0₅P [M+H]⁺ calcd.: 497.1527, found: 497.1521 .

Example 50 ( ?)-2-amino-3,4-dihydro-7-{2-[(diethylphosphoryl)-methoxyl]-3- fluoropropyl}-7H-pyrrolo[2,3-d]pyrimidin-4-one (38a)

The mixture of 36a (200 mg, 0.42 mmol), DABCO (46 mg, 0.42 mmol) and K₂C0₃ (58 mg, 0.42 mmol) in mixture solvent of dioxane and H₂0 (6 mL, 5:1 , v/v) was stirred at 90 °C for 3 h. The reaction was then concentrated under reduced pressure to give a residue that was redissolved in 30% methanolic ammonia (20 mL) and stirred at room temperature for 12 h. After removal of all the volatiles, the residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 20:1 , v/v; 15:1 , v/v; 10:1 , v/v) to give 38a (100 mg, 64%) as a colorless oil. ¹H NMR (300 MHz, CDCI₃): δ 11.01 (s, 1 H, NHCO), 6.59 (dd, J = 3.5, 0.9 Hz, 1 H, H-6), 6.50 (dd, J = 3.5, 0.9 Hz, 1 H, H-5), 6.23 (s, 2H, NH₂), 4.67-4.28 (m, 2H, H-3 ' ), 4.23-4.10 (m, 6H, H-1 ', 2 x CH₂CH₃), 4.05-3.91 (m, 2H, H-2', PCH₂a), 3.85-3.77 (m, 1 H, PCH₂b), 1 .37-1.31 (m, 6H, 2 x CH2CH3); ¹³C NMR (75 MHz, CDC ): δ 160.7 (C-4), 152.8 (C-2), 151.1 (C-7a), 121 .6 (C-6), 102.6 (C-4a), 101.3 (C-5), 82.2 (d, ¹J_c,p = 173.4 Hz, C-3 ' ), 79.6 (dd, ²J_C,F = 18.7 Hz, ³J_c,p = 1 1.5 Hz, C-2 ' ), 64.7 (d, ¹J_c,p = 168.3 Hz, CH₂P), 62.3, 62.2 (CH₂CH₃), 44.5 (d, ³J_C,F = 8.0 Hz, C-1 ' ), 16.7,16.6 (CH₂CH₃); ³¹P NMR (121 MHz, CDCI₃): δ 21 .0; HRMS for Ci₄H₂₂FN₄0₅P [M+H]⁺ calcd.: 377.1384, found: 377.1387.

Example 51 (S)-2-amino-3,4-dihydro-7-{2-[(diethylphosphoryl)-methoxyl]-3- fluoropropyl}-7H-pyrrolo[2,3-d]pyrimidin-4-one (38b)

Compound 38b was obtained as a colorless oil (50 mg, 64%) according to the procedure used for the preparation of 38a, starting from 36b (100 mg, 0.21 mmol), DABCO (23 mg, 0.21 mmol) and K₂C03 (29 mg, 0.21 mmol) in mixture solvent of dioxane and H₂0 (6 ml_, 5:1 , v/v), and ΝΗβ/ΜβΟΗ (20 ml_). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 20:1 , v/v; 15:1 , v/v; 10:1 , v/v). ¹H NMR (300 MHz, CDCI₃): δ 11 .04 (s, 1 H, NHCO), 6.60 (d, J = 3.4 Hz, 1 H, H-6), 6.49 (d, J = 3.5 Hz, 1 H, H-5), 6.21 (s, 2H, NH₂), 4.67-4.29 (m, 2H, H-3 ' ), 4.22-4.11 (m, 6H, H-1 ', 2 x CH₂CH₃), 4.04-3.93 (m, 2H, H-2', PCH₂a), 3.86-3.78 (m, 1 H, PCH₂b), 1 .36-1.30 (m, 6H, 2 x CH₂CH₃); ¹³C NMR (75 MHz, CDCb): δ 160.6 (C-4), 152.7 (C-2), 150.7 (C-7a), 121 .6 (C-6), 102.6 (C-4a), 101.2 (C-5), 82.2 (d, ¹J_c,p = 173.6 Hz, C-3 ' ), 79.6 (C-2 ' ), 64.7 (d, ¹J_c,p = 168.3 Hz, CH₂P), 62.3, 62.2 (CH₂CH₃), 44.6 (d, ³J_C,F = 8.0 Hz, C-1 ' ), 16.7,16.6 (CH₂CH₃); ³¹P NMR (121 MHz, CDCb): δ 21.0; HRMS for Ci₄H₂₂FN₄0₅P [M+H]⁺ calcd.: 377.1384, found: 377.1384.

Example 52 : ( ?)-2-amino-5-fluoro-3,4-dihydro-7-{2-[(diethylphosphoryl)-methoxyl]-3- fluoropropyl}-7H-pyrrolo[2,3-d]py

Compound 39a was obtained as a colorless oil (20 mg, 30%) according to the procedure used for the preparation of 38a, starting from 37a (60 mg, 0.12 mmol), DABCO (14 mg, 0.12 mmol) and K₂C03 (17 mg, 0.12 mmol) in mixture solvent of dioxane and H₂0 (6 ml_, 5:1 , v/v), and ΝΗβ/ΜβΟΗ (20 ml_). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 18:1 , v/v; 15: 1 , v/v; 10: 1 , v/v). ¹ H NMR (300 MHz, CDCI₃): δ 6.42 (d, J = 2.3 Hz, 1 H, H-6), 5.99 (s, 2H, NH₂), 4.66-3.83 (m, 11 H, H-3 ' , H-1 ' , H-2 ' , 2 x CH₂CH₃, PCH₂), 1 .37-1 .32 (m, 6H, 2 x CH₂CH₃); ¹³C NMR (75 MHz, CDCI₃): δ 158.4 (d, ²J_C,F = 2.6 Hz, C-4), 152.3 (C-2), 146.7 (C-7a), 145.4 (d, ¹J_C,F = 248.6 Hz, C-5), 103.6 (d, ²J_C,F = 26.7 Hz, C- 6), 90.6 (d, ²JC,F = 13.9 Hz, C-4a), 81 .9 (d, ¹J_c,p = 173.2 Hz, C-3 ' ), 79.0 (dd, ²J_C,F = 19.0 Hz, ³Jc,p = 10.8 Hz, C-2 ' ), 64.2 (d, ¹J_c,p = 167.5 Hz, CH₂P), 62.4, 62.3 (CH₂CH₃), 43.5 (d, ³J_C,F = 7.9 Hz, C-1 ' ), 16.1 ,16.0 (CH₂CH₃); ³¹ P NMR (121 MHz, CDCI₃): δ 20.3; HRMS for Ci₄H₂i F₂N₄0₅P [M+H]⁺ calcd.: 395.1290, found: 395.1284.

Example 53 : (S)-2-amino-5-fluoro-3,4-dihydro-7-{2-[(diethylphosphoryl)-methoxyl]-3- fluoropropyl}-7H-pyrrolo[2,3-d]pyrimidin-4-one (39b)

Compound 39b was obtained as a colorless oil (25 mg, 35%) according to the procedure used for the preparation of 38a, starting from 37b (90 mg, 0.18 mmol), DABCO (20 mg, 0.18 mmol) and K₂C03 (25 mg, 0.18 mmol) in mixture solvent of dioxane and H₂0 (6 ml_, 5:1 , v/v), and ΝΗβ/ΜβΟΗ (20 ml_). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH , 18: 1 , v/v; 15: 1 , v/v; 10: 1 , v/v). ¹ H NMR (300 MHz, CDCI3): δ 6.41 (d, J = 2.4 Hz, 1 H, H-6), 6.00 (s, 2H, NH₂), 4.66-3.82 (m, 11 H, H-3 ' , H-1 ' , H-2 ' , 2 x CH₂CH₃, PCH₂), 1 .36-1 .31 (m, 6H, 2 x CH₂CH₃); ¹³C NMR (75 MHz, CDCI3): δ 159.0 (C-4), 152.9 (C-2), 147.3 (C-7a), 146.1 (d, ¹J_C,F = 248.6 Hz, C-5), 104.2 (d, ²J_C,F = 26.5 Hz, C-6), 91 .2 (d, ²J_C,F = 13.7 Hz, C-4a), 82.5 (d, ¹J_c,p = 173.1 Hz, C-3 ' ), 79.6 (dd, ²J_C,F = 19.2 Hz, ³J_c,p = 11 .1 Hz, C- 2 ' ), 64.8 (d, ¹Jc,p = 167.6 Hz, CH₂P), 63.0, 62.9 (CH₂CH₃), 44.2 (d, ³J_C,F = 7.9 Hz, C-1 ' ), 16.7,16.6 (CH₂CH₃); ³¹ P NMR (121 MHz, CDCI₃): δ 20.9; HRMS for Ci₄H₂i F₂N₄0₅P [M+H]⁺ calcd.: 395.1290, found: 395.1286.

Example 54 : 4-Chloro-5-cyano-2-pivaloylamino-7H-pyrrolo[2,3-d]pyrimidine (46)

5-Cyano-3,4-dihydro-2-pivaloylamino-7H-pyrrolo[2,3-c/]pyrimidine-4-one 45 (500 mg, 1 .93 mmol), N,N-dimethylaniline (980 mg, 8.10 mmol) and triethylbenzylammonium chloride (220 mg, 0.96 mmol) in dry MeCN (3 mL) was treated dropwise with POCI3 (2.95 g, 19.30 mmol). The reaction mixture was refluxed for 1 h, allowed to cool down and concentrated in vacuo. The resulting dark oil was cautiously treated with ice and was set to pH = 5 using 33% ammonia in H2O. The aqueous layer was extracted with EtOAc (2 50 mL). The combined organic layers were dried with Na2SC>4, filtered and concentrated under reduce pressure. The residue was then purified by column chromatography on silica gel (gradient Hexane:EtOAc, 3:1 , v/v; 1 :1 , v/v) to give 46 (350 mg, 65%) as a white powder. ¹H NMR (300 MHz, CDCI₃): δ 10.30 (s, 1 H, NHCO), 8.52 (s, 1 H, H-6), 1 .24 (s,9H, (CH₃)₃C); ¹³C NMR (75 MHz, CDC ): δ 176.0 (NHCO), 153.2, 152.9, 150.8 (C-4, C-7a, C-2), 137.7 (C-6), 114.6 (C-5), 111 .3 (C-4a), 83.5 (CN), 40.5- 38.8 (C(CH₃)₃, overlapped with DMSO), 26.9 (C(CH₃)₃).

Example 55 : ( ?)-4-Chloro-5-cyano-7-{2-[(diethylphosphoryl)-methoxyl]-3-fluoropropyl}- 2-pivaloylamino-7H-pyrrolo[2,3-rflpyrimidin (47a)

Compound 47a was obtained as a colorless oil (220 mg, 80%) according to the procedure used for the preparation of 36a, starting from 46 (150 mg, 0.54 mmol), 4a (120 mg, 0.49 mmol), Ph₃P (260 mg, 0.98 mmol), and DIAD (0.19 mL, 0.98 mmol) in anhydrous THF (10 mL). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 50:1 , v/v; 40:1 , v/v; 30:1 , v/v). ¹H NMR (300 MHz, CDCb): δ 8.24 (s, 1 H, NHCO), 7.91 (s, 1 H, H-6), 4.74-4.29 (m, 4H, H-3 ' , H-1 ' ), 4.17-3.96 (m, 6H, H-2', 2 x CH₂CH₃, PCH₂a), 3.81 (dd, J = 13.9, 9.0 Hz, 1 H, PCH₂b), 1 .34-1.25 (m, 15H, (CH₃)₃C, 2 x CH2CH3); ¹³C NMR (75 MHz, CDCb): δ 175.8 (NHCO), 153.4, 153.2, 152.3 (C-4, C-7a, C-2), 138.2 (C-6), 113.5 (C-5), 112.1 (C-4a), 86.0 (CN), 82.3 (d, ¹J_c,p = 174.0 Hz, C-3 ' ), 78.7 (dd, ²J_C,F = 19.2 Hz, ³J_C,P = 9.8 Hz, C-2 ' ), 64.7 (d, ¹Jc,p = 167.1 Hz, CH₂P), 62.9, 62.8 (CH2CH3), 45.6 (d, ³J_C,F = 8.4 Hz, C-1 ' ), 40.6 (C(CH₃)₃), 27.6 (C(CH₃)₃), 16.7 (CH₂CH₃); ³¹P NMR (121 MHz, CDCb): δ 20.7; HRMS for C2oH₂8CIFN₅0₅P [M+H]⁺ calcd.: 504.1573, found: 504.1567.

Example 56 : (S)-4-Chloro-5-cyano-7-{2-[(diethylphosphoryl)-methoxyl]-3-fluoropropyl}- 2-pivaloylamino-7H-pyrrolo[2,3-d]pyrimidin (47b)

Compound 47b was obtained as a colorless oil (150 mg, 60%) according to the procedure used for the preparation of 36a, starting from 46 (150 mg, 0.54 mmol), 4b (120 mg, 0.49 mmol), Ph₃P (260 mg, 0.98 mmol), and DIAD (0.19 mL, 0.98 mmol) in anhydrous THF (10 mL). The crude residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 50:1 , v/v; 40:1 , v/v; 30:1 , v/v). ¹H N MR (300 MHz, CDCI₃): δ 8.24 (s, 1 H, NHCO), 7.92 (s, 1 H, H-6), 4.74-4.30 (m, 4H, H-3 ' , H-1 ' ), 4.15-3.96 (m, 6H, H-2\ 2 x CH₂CH₃, PCH₂a), 3.81 (dd, J = 13.9, 9.0 Hz, 1 H, PCH₂b), 1 .35-1.26 (m, 15H, (CH₃)₃C, 2 x CH₂CH₃); ¹³C NMR (75 MHz, CDCI₃): δ 175.7 (NHCO), 153.2, 153.0, 152.1 (C-4, C-7a, C-2), 138.1 (C-6), 1 13.4 (C-5), 11 1.9 (C-4a), 85.8 (CN), 82.1 (d, ¹J_c,p = 174.0 Hz, C-3 ' ), 78.5 (dd, ²J_C,F = 19.2 Hz, ³J_c,p = 9.9 Hz, C-2 ' ), 64.5 (d, ¹Jc,p = 167.0 Hz, CH₂P), 62.8-62.6 (CH₂CH₃), 45.4 (d, ³J_C,F = 8.6 Hz, C-1 ' ), 40.5 (C(CH₃)₃), 27.5 (C(CH₃)₃), 16.6, 16.5 (CH₂CH₃); ³¹P NMR (121 MHz, CDCI₃): δ 20.7; HRMS for C₂oH₂8CIFN₅0₅P [M+H]⁺ calcd.: 504.1573, found: 504.1578.

Example 57 : ( ?)-2-amino-5-cyano-3,4-dihydro-7-{2-[(diethylphosphoryl)-methoxyl]-3- fluoropropyl}-7H-pyrrolo[2,3-d]py

Compound 48a was obtained as a colorless oil (50 mg, 30%) according to the procedure used for the preparation of 38a, starting from 47a (220 mg, 0.44 mmol), DABCO (33 mg, 0.30 mmol) and K₂C0₃ (61 mg, 0.44 mmol) in mixture solvent of dioxane and H₂0 (6 mL, 5:1 , v/v), and NH₃/MeOH (20 mL). The residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 20:1 , v/v; 15:1 , v/v; 10:1 , v/v). ¹H NMR (300 MHz, CDCI₃): δ 10.89 (s, 1 H, NHCO), 7.30 (d, J = 1 .1 Hz, 1 H, H-6), 6.16 (s, 2H, NH₂), 4.71 -4.29 (m, 3H, H-3 ' , H-1 'a), 4.21 -3.93 (m, 7H, H-1 ' b, H-2 ' , 2 x CH₂CH₃, PCH₂a), 3.83 (dd, J = 13.8, 9.1 Hz, 1 H, PCH₂b), 1 .38-1.32 (m, 6H, 2 x CH₂CH₃); ¹³C NMR (75 MHz, CDCI₃): δ 159.2 (C-4), 154.0 (C-2), 152.2 (C-7a), 130.9 (C-6), 115.5 (C-5), 99.6 (C-4a), 86.7 (CN), 82.4 (d, ¹J_c,p = 173.6 Hz, C-3 ' ), 79.1 (dd, ²JC,F = 19.1 Hz, ³Jc,p = 10.4 Hz, C-2 ' ), 64.8 (d, ¹J_c,p = 167.7 Hz, CH₂P), 62.9, 62.8 (CH₂CH₃), 45.2 (d, ³J_C,F = 8.1 Hz, C-1 ' ), 16.8,16.7 (CH₂CH₃); ³¹P NMR (121 MHz, CDCI₃): δ 20.7; HRMS for Ci₅H₂iFN₅0₅P [M+H]⁺ calcd.: 402.1336, found: 402.1324.

Example 58 : (S)-2-amino-5-cyano-3,4-dihydro-7-{2-[(diethylphosphoryl)-methoxyl]-3- fluoropropyl}-7H-pyrrolo[2,3-d]py

The mixture of 47b (150 mg, 0.30 mmol), Et₃N (0.12 mL, 0.89 mmol), NaOAc (73 mg, 0.89 mmol) and DABCO (33 mg, 0.30 mmol) in DMF (5 mL) was stirred at room temperature for 40 h. Then H2O (5 mL) was added into the reaction and stirred for another 1 h. The mixture was extracted with EtOAc (2 x 25 mL). The combined organic phases dried with Na2S0₄, filtered and concentrated under reduced pressure to give the crude product. The residue was then resolved in 30% methanolic ammonia (20 mL) and stirred at room temperature for 12 h. After removal of all the volatiles, the residue was purified by column chromatography on silica gel (gradient DCM/MeOH, 20:1 , v/v; 15:1 , v/v; 10:1 , v/v) to give 48b (80 mg, 67%) as a colorless oil. ¹H NMR (300 MHz, CDCI3): δ 10.89 (s, 1 H, NHCO), 7.30 (d, J = 1 .1 Hz, 1 H, H-6), 4.71 - 4.28 (m, 3H, H-3 ' , H-1 'a), 4.21 -3.93 (m, 7H, H-1 ' b, H-2 ' , 2 x CH2CH3, PCH₂a), 3.87-3.80 (m, 1 H, PCH₂b), 1 .38-1.32 (m, 6H, 2 x CH2CH3); ¹³C NMR (75 MHz, CDCI3): δ 159.2 (C-4), 154.0 (C-2), 152.2 (C-7a), 130.9 (C-6), 1 15.5 (C-5), 99.5 (C-4a), 86.6 (CN), 82.4 (d, ¹J_c,p = 173.5 Hz, C-3 ' ), 79.1 (dd, ²J_C,F = 19.2 Hz, ³J_C,P = 10.4 Hz, C-2 ' ), 64.8 (d, ¹J_c,p = 167.2 Hz, CH₂P), 63.0, 62.9 (CH2CH3), 45.2 (d, ³J_C,F = 8.2 Hz, C-1 ' ), 16.8,16.7 (CH2CH3); ³¹P NMR (121 MHz, CDCI3): δ 20.7; HRMS for C15H21 FN5O5P [M+H]⁺ calcd.: 402.1336, found: 402.1329.

Example 59 ( ?)-2-Amino-6-(4-methoxyphenylthio)-9-[2-(phosphonomethoxyl)-3- fluoropropyl]purine (33a)

Bromotrimethylsilane (0.43 mL, 3.20 mmol) was added dropwise to a solution of diethyl phosphonate ester 32a (200 mg, 0.40 mmol) and 2,6-lutidine (0.74 mL, 6.40 mmol) in anhydrous acetonitrile (10 mL) at 0 °C. After the addition was completed, the mixture was slowly warmed to room temperature and set aside in the dark for 12 h. The reaction was quenched with 0.1 M TEAB and was then concentrated under reduced pressure. The crude residue was purified by column chromatography on silica gel (gradient acetone/Et.3N/H20, 6:1 :1 , v/v/v), followed by RP-HPLC (linear gradient, 2-98% CH₃CN in 0.05 M TEAB solution) to give the desired phosphonate acid triethylammonium salt 33a (120 mg, 70%) as a white foam. ¹H NMR (300 MHz, D₂0): δ 7.93 (s, 1 H, H-8), 7.19 (d, J = 8.8 Hz, 2H, ArH), 6.73-6.70 (m, 2H, ArH), 4.60-4.21 (m, 2H, H-3 ' ), 4.18-4.08 (m, 2H, H-1 ' ), 3.89-3.81 (m, 1 H, H-2 ' ), 3.61 (s, 3H, OCH3), 3.59-3.41 (m, 2H, PCH₂); ¹³C NMR (75 MHz, D₂0): δ 161.1 (C-2), 160.0 (Ar-C), 159.3 (C-6), 150.1 (C-4), 142.8 (C-8), 136.8 (Ar-C), 123.0 (C-5), 116.8 (Ar-C), 1 14.8 (Ar-C), 82.1 (d, ¹J_c,p = 168.0 Hz, C-3 ' ), 77.4 (dd, ²J_C,F = 18.9 Hz, ³J_C,P = 11.2 Hz, C-2 ' ), 66.9 (d, ¹Jc,p = 154.8 Hz, CH₂P), 55.3 (OCH₃), 42.7 (d, ³J_C,F = 7.1 Hz, C-1 ' ); ³¹P NMR (121 MHz, D₂0): δ 14.1 ; HRMS for C16H19FN5O5PS [M-H]^" calcd.: 442.0756, found: 442.0752.

Example 60 (S)-2-Amino-6-(4-methoxyphenylthio)-9-[2-(phosphonomethoxyl)-3- fluoropropyl]purine (33b)

Compound 33b was obtained as a white foam (86 mg, 65%) according to the procedure used for the preparation of 33a, starting from compound 32b (150 mg, 0.30 mmol), bromotrimethylsilane (0.32 ml_, 2.40 mmol), and 2,6-lutidine (0.56 ml_, 4.80 mmol) in anhydrous acetonitrile (10 ml_). The crude residue was purified by column chromatography on silica gel (gradient acetone/Et₃N/H₂0, 6:1 :1 , v/v/v). ¹H NMR (300 MHz, D₂0): δ 7.95 (s, 1 H, H- 8), 7.22 (d, J = 8.4 Hz, 2H, ArH), 6.82-6.65 (m, 2H, ArH), 4.63-4.24 (m, 2H, H-3 ' ), 4.20-4.10 (m, 2H, H-1 ' ), 3.93-3.85 (m, 1 H, H-2 ' ), 3.64 (s, 3H, OCH₃), 3.61 -3.45 (m, 2H, PCH₂); ¹³C NMR (75 MHz, D₂0): δ 161.1 (C-2), 160.0 (Ar-C), 159.3 (C-6), 150.1 (C-4), 142.8 (C-8), 136.8 (Ar-C), 123.0 (C-5), 116.8 (Ar-C), 1 14.8 (Ar-C), 82.1 (d, ¹J_c,p = 168.1 Hz, C-3 ' ), 77.5 (dd, ²J_C,F = 18.9 Hz, ³Jc,p = 1 1.1 Hz, C-2 ' ), 66.7 (d, ¹J_c,p = 155.5 Hz, CH₂P), 55.3 (OCH₃), 42.8 (d, ³J_C,F = 7.1 Hz, C-1 ' ); ³¹P NMR (121 MHz, D₂0): δ 14.3; HRMS for C16H19FN5O5PS [M-H]^" calcd.: 442.0756, found: 442.0734.

Example 61 ( ?)-2-amino-3,4-dihydro-7-(2-phosphonomethoxyl-3-fluoropropyl)-7H- pyrrolo[2,3-d]pyrimidin-4-one (40a)

Compound 40a was obtained as a white foam (59 mg, 70%) according to the procedure used for the preparation of 33a, starting from compound 38a (100 mg, 0.26 mmol), bromotrimethylsilane (0.28 ml_, 2.12 mmol), and 2,6-lutidine (0.50 ml_, 4.24 mmol) in anhydrous acetonitrile (10 ml_). The crude residue was purified by column chromatography on silica gel (gradient acetone/Et₃N/H₂0, 4:1 :1 , v/v/v). ¹H NMR (300 MHz, D₂0): δ 6.65 (d, J = 3.7 Hz, 1 H, H-6), 6.17 (d, J = 3.6 Hz, 1 H, H-5), 4.50-4.04 (m, 2H, H-3 ' ), 4.95-3.93 (m, 2H, H- 1 ' ), 3.83-3.70 (m, 1 H, H-2 ' ), 3.41 (d, J = 9.2 Hz, 2H, PCH₂); ¹³C NMR (75 MHz, D₂0): δ 160.8 (C-4), 152.0 (C-2), 150.3 (C-7a), 123.1 (C-6), 100.9 (C-5), 99.5 (C-4a), 82.3 (d, ¹J_c,p = 167.3 Hz, C-3 ' ), 78.0 (dd, ²JC,F = 18.4 Hz, ³J_c,p = 10.7 Hz, C-2 ' ), 67.6 (d, ¹J_c,p = 151 .9 Hz, CH₂P), 43.4 (d, ³JC,F = 7.3 Hz, C-1 ' ); ³¹P NMR (121 MHz, D₂0): δ 12.3; HRMS for CioHi₄FN₄0₅P [M-H]- calcd.: 319.0612, found: 319.0611 .

Example 62 (S)-2-amino-3,4-dihydro-7-(2-phosphonomethoxyl-3-fluoropropyl)-7H- pyrrolo[2,3-d]pyrimidin-4-one (40b)

Compound 40b was obtained as a white foam (30 mg, 70%) according to the procedure used for the preparation of 33a, starting from compound 38b (50 mg, 0.13 mmol), bromotrimethylsilane (0.14 mL, 1.06 mmol), and 2,6-lutidine (0.25 mL, 2.12 mmol) in anhydrous acetonitrile (5 mL). The crude residue was purified by column chromatography on silica gel (gradient acetone/Et₃N/H₂0, 4:1 :1 , v/v/v). ¹H NMR (300 MHz, D₂0): δ 6.73 (d, J = 3.5 Hz, 1 H, H-6), 6.25 (d, J = 3.5 Hz, 1 H, H-5), 4.56-4.11 (m, 2H, H-3 ' ), 4.09-3.98 (m, 2H, H- 1 ' ), 3.90-3.78 (m, 1 H, H-2 ' ), 3.42 (d, J = 9.2 Hz, 2H, PCH₂); ¹³C NMR (75 MHz, D₂0): δ 166.0 (C-4), 156.5 (C-2), 151.1 (C-7a), 122.3 (C-6), 100.6 (C-4a, C-5), 82.4 (d, ¹J_c,p = 167.0 Hz, C-3 ' ), 78.0 (dd, ²J_C,F = 18.2 Hz, ³J_c,p = 10.4 Hz, C-2 ' ), 68.1 (d, ¹J_c,p = 150.0 Hz, CH₂P), 43.2 (d, ³JC,F = 7.2 Hz, C-1 ' ); ³¹P NMR (121 MHz, D₂0): δ 12.6; HRMS for Ci₀Hi₄FN₄O₅P [M- H]- calcd.: 319.0612, found: 319.0598.

Example 63 ( ?)-2-amino-5-fluoro-3,4-dihydro-7-{2-phosphonomethoxyl -3- fluoropropyl}-7H-pyrrolo[2,3-d]py

Compound 41a was obtained as a white foam (9 mg, 50%) according to the procedure used for the preparation of 33a, starting from compound 39a (20 mg, 0.05 mmol), bromotrimethylsilane (0.05 mL, 0.40 mmol), and 2,6-lutidine (0.09 mL, 0.81 mmol) in anhydrous acetonitrile (5 mL). The crude residue was purified by column chromatography on silica gel (gradient acetone/Et₃N/H₂0, 4:1 :1 , v/v/v). ¹H NMR (300 MHz, D₂0): δ 6.56 (d, J = 2.1 Hz, 1 H, H-6), 4.59-4.16 (m, 2H, H-3 ' ), 4.00-3.98 (m, 2H, H-1 ' ), 3.90-3.81 (m, 1 H, H-2 ' ), 3.49 (d, J = 9.1 Hz, 2H, PCH₂); ¹³C NMR (75 MHz, D₂0): δ 159.5 (d, ³J_C,F = 2.8 Hz, C-4), 1 52.3(C-2), 146.0 (d, ³JC,F = 3.0Hz, C-7a), 144.6 (d, ¹J_C,F = 246.3 Hz, C-5), 105.1 (d, ¹J_C,F = 26.6 Hz, C-6), 90.0 (d, ¹J_C,F = 14.4 Hz, C-4a), 82.4 (d, ¹J_c,p = 167.2 Hz, C-3 ' ), 78.1 (dd, ²J_C,F = 18.3 Hz, ³Jc,p = 10.4 Hz, C-2 ' ), 67.2 (d, ¹J_c,p = 153.2 Hz, CH₂P), 43.2 (d, ³J_C,F = 7.4 Hz, C- 1 ' ); ³¹P NMR (121 MHz, D₂0): δ 13.8; HRMS for C10H13F2N4O5P [M-H]^" calcd.: 337.0519, found: 337.0503.

Example 64 (S)-2-amino-5-fluoro-3,4-dihydro-7-{2-phosphonomethoxyl -3- fluoropropyl}-7H-pyrrolo[2,3-d]py

Compound 41 b was obtained as a white foam (10 mg, 60%) according to the procedure used for the preparation of 33a, starting from compound 39b (20 mg, 0.05 mmol), bromotrimethylsilane (0.05 mL, 0.40 mmol), and 2,6-lutidine (0.09 mL, 0.81 mmol) in anhydrous acetonitrile (5 mL). The crude residue was purified by column chromatography on silica gel (gradient acetone/Et₃N/H₂0, 4:1 :1 , v/v/v). ¹H NMR (300 MHz, D₂0): δ 6.52 (d, J = 2.2 Hz, 1 H, H-6), 4.57-4.09 (m, 2H, H-3 ' ), 4.07-3.92 (m, 2H, H-1 ' ), 3.89-3.77 (m, 1 H, H-2 ' ), 3.43 (d, J = 9.1 Hz, 2H, PCH₂); ¹³C NMR (75 MHz, D₂0): δ 167.1 (C-4), 159.1 (C-2), 147.4 (C- 7a), 144.6 (d, ¹J_C,F = 244.3 Hz, C-5), 103.5 (d, ²J_C,F = 27.2 Hz, C-6), 91.2 (d, ²J_C,F = 14.4 Hz, C-4a), 82.4 (d, ¹J_c,p = 166.9 Hz, C-3 ' ), 77.9 (dd, ²J_C,F = 18.2 Hz, ³J_c,p = 10.3 Hz, C-2 ' ), 68.1 (d, ¹Jc,p = 150.1 Hz, CH₂P), 42.7 (d, ³J_C,F = 7.2 Hz, C-1 ' ); ³¹P NMR (121 MHz, D₂0): δ 12.7; HRMS for CioHi₃F₂N₄0₅P [M-H]^" calcd.: 337.0519, found: 337.0522.

Example 65 ( ?)-2-amino-5-cyano-3,4-dihydro-7-{2-phosphonomethoxyl -3- fluoropropyl}-7H-pyrrolo[2,3-d]pyrimidin-4-one (49a)

Compound 49a was obtained as a white foam (30 mg, 70%) according to the procedure used for the preparation of 33a, starting from compound 48a (50 mg, 0.13 mmol), bromotrimethylsilane (0.13 mL, 1.00 mmol), and 2,6-lutidine (0.23 mL, 2.00 mmol) in anhydrous acetonitrile (5 mL). The crude residue was purified by column chromatography on silica gel (gradient acetone/Et₃N/H₂0, 5:1 :1 , v/v/v). ¹H NMR (300 MHz, D₂0): δ 7.51 (s, 1 H, H- 6), 4.60-4.21 (m, 2H, H-3 ' ), 4.10-4.08 (m, 2H, H-1 ' ), 3.90-3.83 (m, 1 H, H-2 ' ), 3.44 (d, J = 9.1 Hz, 2H, PCH₂); ¹³C NMR (75 MHz, D₂0): δ 162.0 (C-4), 155.6 (C-2), 151.1 (C-7a), 132.6 (C-6), 116.2 (C-5), 99.3 (C-4a), 84.3 (CN), 82.2 (d, ¹J_c,p = 167.1 Hz, C-3 ' ), 77.3 (dd, ²J_C,F = 18.6 Hz, ³Jc,p = 10.5 Hz, C-2 ' ), 68.0 (d, ¹J_c,p = 149.9 Hz, CH₂P), 44.1 (d, ³J_C,F = 7.0 Hz, C- 1 ' ); ³¹P NMR (121 MHz, D₂0): δ 12.5; HRMS for C11 H13FN5O5P [M-H]^" calcd.: 344.0565, found: 344.0573.

Example 66 (S)-2-amino-5-cyano-3,4-dihydro-7-{2-phosphonomethoxyl -3- fluoropropyl}-7H-pyrrolo[2,3-d]pyrimidin-4-one 49b

Compound 49b was obtained as a white foam (51 mg, 75%) according to the procedure used for the preparation of 33a, starting from compound 48b (80 mg, 0.20 mmol), bromotrimethylsilane (0.21 mL, 1.60 mmol), and 2,6-lutidine (0.37 mL, 3.20 mmol) in anhydrous acetonitrile (5 mL). The crude residue was purified by column chromatography on silica gel (gradient acetone/Et₃N/H₂0, 5:1 :1 , v/v/v). ¹H NMR (300 MHz, D₂0): δ 7.40 (d, J = 1.1 Hz, 1 H, H-6), 4.70-4.20 (m, 2H, H-3 ' ), 4.08-3.93 (m, 2H, H-1 ' ), 3.90-3.78 (m, 1 H, H-2 ' ), 3.58-3.42 (m, 2H, PCH₂); ¹³C NMR (75 MHz, D₂0): δ 159.0 (C-4), 153.3 (C-2), 150.7 (C-7a), 132.7 (C-6), 115.7 (C-5), 98.7 (C-4a), 84.6 (CN), 82.2 (d, ¹J_c,p = 167.7 Hz, C-3 ' ), 77.7 (dd, ²JC,F = 18.5 Hz, ³Jc,p = 11 .3 Hz, C-2 ' ), 66.7 (d, ¹J_c,p = 155.2 Hz, CH₂P), 44.3 (d, ³J_C,F = 7.6 Hz, C-1 ' ); ³¹P NMR (121 MHz, D₂0): δ 14.3; HRMS for C11 H13FN5O5P [M-H]^" calcd.: 344.0565, found: 344.0557.

Example 67 : Diamyl (((((R)-1 -(2-amino-4-oxo-3,4-dihydro-7H-pyrrolo[2,3-d]pyrimidin-7- yl)-3-fluoropropan-2-yl)oxy)methyl)(phenoxy)phosphoryl)-L-aspartate (50)

Compound 40a (40 mg, 0.09 mmol, EtsN salt) was mixed with L-aspartic acid amyl diester HCI salt (Maiti, M.; Maiti, M.; Rozenski, J.; De Jonghe, S.; Herdewijn, P. Aspartic acid based nucleoside phosphoramidate prodrugs as potent inhibitors of hepatitis C virus replication. Org. Biomol. Chem. 2015, 13, 5158-5174) (50 mg, 0.16 mmol) and phenol (40 mg, 0.42 mmol) in anhydrous pyridine (53 mL). Then, EtsN (0.13 mL, 0.95 mmol) was added and the mixture was stirred at 60 °C under a nitrogen atmosphere for 15-20 min. 2,2'-Dithiodipyridine (150 mg, 0.66 mmol) was mixed in a separate flask with PPh₃ (180 mg, 0.66 mmol) in anhydrous pyridine (2 mL) and the resultant mixture was stirred for 10-15 min to give a clear light yellow solution. This solution was then added to the above solution and the combined mixture was stirred at 60 °C for 12 h. The mixture was then concentrated under reduced pressure to give a residue that was redissolved in EtOAc (50 mL). This solution was washed with saturated aq. NaHCC>3 (25 mL) and brine (25 mL), the organic layer was separated, dried over Na₂S0₄, filtered, and concentrated under reduced pressure. The crude residue was purified by silica gel chromatography (gradient DCM/MeOH, 30:1 , v/v; 20:1 , v/v) and then purified by RP-HPLC (linear gradient, 5-95% CH₃CN in water) to compound 50 (60 mg, 40%) as a white powder. ¹ H NMR (300 MHz, CD₃CN): δ 10.08 (s, 1 H, NHCO), 7.37-7.08 (m, 5H, ArH), 6.72-6.69 (m, 1 H, H-6), 6.40-6.38 (m, 1 H, H-5), 5.80, 5.78 (s, 2H, NH₂), 4.70-4.25 (m, 4H, H-3 ' , NHPO, H-a- Asp), 4.16-3.89 (m, 9H, H-1 ' , H-2 ' , 2 x OCH₂(CH₂)₃CH₃, PCH₂), 2.84-2.57 (m, 2H, Η-β- Asp), 1 .58-1 .52 (m, 4H , 2 x OCH₂CH₂(CH₂)₂CH₃), 1 .32-1 .24 (m, 8H, 2 x 0(CH₂)₂(CH₂)₂CH₃), 0.91 -0.84 (m, 6H, 2 x 0(CH₂)₄CH₃); ¹³C N MR (75 MHz, CD₃CN): δ 172.9, 172.8, 172.7 (CO- a), 171 .6 (CO-β), 160.5 (C-4), 153.5, 153.4 (C-2), 152.0, 151 .9 (C-7a), 130.7 (Ar-C), 125.9 (Ar- C), 122.4 (C-6), 121 .9, 121 .8 (Ar-C), 102.7 (C-5), 101 .7 (C-4a), 83.6 (d, ¹J_C,F = 170.3 Hz, C- 3 ' ), 83.5 (d, ¹JC,F = 170.0 Hz, C-3 ' ), 80.7, 80.5, 80.4, 80.3, 80.1 (C-2 ' ), 66.8 (d, ¹J_c,p = 156.2 Hz, CH₂P), 66.6 (d, ¹J_c,p = 154.2 Hz, CH₂P), 66.6, 65.9, 65.8 (OCH₂(CH₂)₃CH₃), 51 .7, 51 .6 (C-a-Asp), 45.1 , 45.0, 44.9 (C-1 ' ), 39.9 (d, ³J_c,p = 3.9 Hz, C-3-Asp), 39.7 (d, ³J_c,p = 3.9 Hz, C-3-Asp), 29.1 , 29.0, 28.8, 28.7 (OCH₂(CH₂)₂CH₂CH₃), 23.1 , 23.0 (0(CH₂)₃CH₂CH₃), 14.3 (0(CH₂)₄CH₃); ³¹ P NMR (121 MHz, CD₃CN): 5 23.0, 22.2; HRMS for C₃₀H₄₃FN₅O₈P [M+H]⁺ calcd.: 652.2905, found: 652.2923.

Example 68 : Tetraamyl 2,2'-((((((R)-1 -(2-amino-4-oxo-3,4-dihydro-7H-pyrrolo[2,3- d]pyrimidin-7-yl)-3-fluoropropan-2-yl)oxy)methyl)phosphoryl)bis(azanediyl))(2S,2'S)- disuccinate (51 )

Compound 40a (15 mg, 0.04 mmol, Et₃N salt) was mixed with L-aspartic acid amyl diester HCI salt (Maiti, M.; Maiti, M.; Rozenski, J.; De Jonghe, S.; Herdewijn, P. Aspartic acid based nucleoside phosphoramidate prodrugs as potent inhibitors of hepatitis C virus replication. Org. Biomol. Chem. 2015, 13, 5158-5174) (44 mg, 0.14 mmol) in anhydrous pyridine (3 mL). Then, ΕίβΝ (0.04 mL, 0.40 mmol) was added and the mixture was stirred at 60 °C under a nitrogen atmosphere for 15-20 min. 2,2 ' -Dithiodipyridine (55 mg, 0.25 mmol) was mixed in a separate flask with PP i3 (65 mg, 0.25 mmol) in anhydrous pyridine (2 mL) and the resultant mixture was stirred for 10-15 min to give a clear light yellow solution. This solution was then added to the above solution and the combined mixture was stirred at 60 °C for 12 h. The mixture was then concentrated under reduced pressure to give a residue that was redissolved in EtOAc (50 mL). This solution was washed with saturated aq. NaHCC>3 (25 mL) and brine (25 mL), the organic layer was separated, dried over Na₂S0₄, filtered, and concentrated under reduced pressure. The crude residue was purified by silica gel chromatography (gradient DCM/MeOH, 30:1 , v/v; 20:1 , v/v) and then purified by RP-HPLC (linear gradient, 5-95% CH3CN in water) to compound 51 (5 mg, 20%) as a white powder. ¹H NMR (300 MHz, CD₃CN): δ 9.87 (s, 1 H, NHCO), 6.63 (d, J = 3.5 Hz, 1 H, H-6), 6.32 (d, J = 3.5 Hz, 1 H, H-5),6.06 (s, 2H, NH₂), 4.68, 3.74 (m, 19H, H-3 ' , 2 x NHPO, 2 x H-a-Asp, H-1 ' , H-2 ' , 4 x OCH₂(CH₂)₃CH₃, PCH₂), 2.89-2.73 (m, 4H, 2 x Η-β-Asp), 1.61 -1.53 (m, 8H, 4 x OCH₂CH₂(CH₂)₂CH₃), 1 .33-1.24 (m, 16H, 4 x 0(CH₂)₂(CH₂)₂CH₃), 0.91 -0.84 (m, 12H, 4 x 0(CH₂)₄CH₃); ¹³C NMR (75 MHz, CD₃CN): δ 173.3, 173.2 (CO-a), 171.9, 171.8 (CO-β), 160.0 (C-4), 153.7 (C-2), 151.6 (C-7a), 122.1 (C-6), 102.8 (C-5), 101 .8 (C-4a), 83.1 (d, ¹J_C,F = 170.2 Hz, C-3 ' ), 80.13 (dd, ²J_C,F = 18.7 Hz, ³J_C,P = 11 .5 Hz, C-2 ' ), 68.1 (d, ¹Jc,p = 136.9 Hz, CH₂P), 66.6, 66.5, 65.8 (OCH₂(CH₂)₃CH₃), 51.1 , 50.5 (C- a-Asp), 44.86 (d, J = 8.1 Hz, C-1 ' ), 40.0, 39.9, 39.8 (d, ³J_c,p = 3.9 Hz, Ο-β-Asp), 29.1 , 29.0, 28.8 (OCH₂(CH₂)₂CH₂CH₃), 23.1 , 23.0 (0(CH₂)₃CH₂CH₃), 14.3 (0(CH₂)₄CH₃); ³¹P NMR (121 MHz, CD₃CN): δ 21 .5; HRMS for C38H₆₄FN₆0iiP [M-H]^" calcd.: 829.4282, found: 829.4298.

Example 69 : HBV Antiviral Assay in HepG2 2.2.15 cells

The primary anti-HBV assay is performed as previously described (Korba, BF and Milman, G. A cell culture assay for compound which inhibit hepatitis B virus replication. Antiviral Res. 1991 , 15, 217-228; and Korba, BF and Gerin, JL. Use of a standardized cell culture assay to assess activities of nucleoside analogs again hepatitis B virus replication. Antiviral Res. 1992, 19, 55-70) with modifications to use real-time qPCR (TaqMan) to measure extracellular HBV DNA copy number associated with virions released from HepG2 2.2.15 cells. The HepG2 2.2.15 cell line is a stable human hepatoblastoma cell line that contains two copies of the HBV wild-type strain aywl genome and constitutively produces high levels of HBV. Antiviral compounds blocking any late step of viral replication such as transcription, translation, pregenome encapsidation, reverse transcription, particle assembly and release can be identified and characterized using this cell line. Briefly, HepG2 2.2.15 cells are plated in 96-well microtiter plates at 1 .5x10⁴ cells/well in Dulbecco's Modified Eagle's Medium supplemented with 2% FBS, 380 μg mL G418, 2.0 mM L-Glutamine, 100 units/mL Penicillin, 100 μg mL Streptomycin, and 0.1 mM non-essential amino acids. Only the interior wells are utilized to reduce "edge effects" observed during cell culture; the exterior wells are filled with complete medium to help minimize sample evaporation. After 16-24 hours the confluent monolayer of HepG2 2.2.15 cells is washed and the medium is replaced with complete medium containing various concentrations of a test compound in triplicate. Lamivudine (3TC) and entecavir were used as the positive control, while media alone is added to cells as a negative control (virus control, VC). Three days later the culture medium is replaced with fresh medium containing the appropriately diluted test compounds. Six days following the initial administration of the test compound, the cell culture supernatant is collected, treated with pronase and then used in a real-time quantitative TaqMan qPCR assay. The PCR-amplified HBV DNA is detected in real-time by monitoring increases in fluorescent signal that result from the exonucleolytic degradation of a quenched fluorescent probe molecule that hybridizes to the amplified HBV DNA. For each PCR amplification, a standard curve is simultaneously generated using dilutions of purified HBV DNA. Antiviral activity is calculated from the reduction in HBV DNA levels (EC50 & EC90 values determined). A tetrazolium dye (MTS; 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxy-methoxyphenyl)-2-(4- sulfophenyl)-2H-tetrazolium; CellTiter®96 Reagent, Promega) uptake assay is then employed to measure cell viability, which is used to calculate toxicity (CC50). The Selectivity Index (SI50) is calculated as CC50/IC50. The data are shown in Table 1 .

Table 1

Example 70 : Antiviral activity against the HIV-1 X4 strain NL4.3 and HIV-1 R5 strain BaL

TZM-bl cells (Montefiori, D.C. Methods Mol. Biol. 2009, 485, 395-405) were seeded in transparent 96-well plates at 1 x 104 cells per well in DMEM (Dulbecco's Modified Eagle Medium; Life Technologies, Waltham, MA, USA) with 10% Fetal Bovine Serum (FBS) and 10 mM HEPES. Subsequently, compounds were added and the cell/compound mixture was incubated at 37°C. After 30 min, virus was added at 100 pg p-24 per well. After 48 h of incubation, the assay plates were analyzed. For the analysis, steadylite plus substrate solution (PerkinElmer, Waltham, MA, USA) was added to the assay plates. The luminescent signal of the lysed cell suspension was analyzed in white 96-well plates on a SpectraMax L luminescence microplate reader (Molecular Devices, Sunnyvale, CA, USA) after a 10 min incubation period in the dark. Luciferase activity induced by HIV-1 Tat protein expression was measured as an assessment of the amount of HIV replication. The data are shown in Table 2.

Table 2

Example 71 Antiviral activity against the varicella-zoster virus and human cytomegalovirus

The compounds of the invention were evaluated against varicella-zoster virus (VZV) strain Oka, and thymidine kinase deficient (TK^") VZV strain 07-1 and human cytomegalovirus (HCMV) strains AD-169 and Davis. The antiviral assays were based on inhibition of virus- induced cytopathicity or plaque formation in human embryonic lung (HEL) fibroblasts. Confluent cell cultures in microtiter 96-well plates were inoculated with 100 CCID₅o of HCMV (1 CCID₅o being the virus dose to infect 50% of the cell cultures) or with 20 plaque forming units (PFU) (VZV). After 1 -2 h adsorption period, residual virus was removed, and the cell cultures were incubated in the presence of varying concentrations of the test compounds. Viral cytopathicity (HCMV) or plaque formation (VZV) was recorded as soon as it reached completion in the control virus-infected cell cultures that were not treated with the test compounds. Antiviral activity was expressed as the EC50 or concentration (expressed in micromolar) required for reducing virus-induced cytopathogenicity or viral plaque formation by 50%. Cytostatic measurements were based on the inhibition of cell growth. HEL cells were seeded at a rate of 5 ^χ 10³ cells/well into 96-well microtiter plates and allowed to proliferate for 24 h. Then, medium containing different concentrations of the test compounds was added. After 3 days of incubation at 37 °C, the cell number was determined with a Coulter counter. The cytostatic concentration (expressed in micromolar) was calculated as the CC50 or the compound concentration required for reducing cell proliferation by 50% relative to the number of cells in the untreated controls. CC50 values were estimated from graphic plots of the number of cells (percentage of control) as a function of the concentration of the test compounds. Alternatively, cytotoxicity of the test compounds was expressed as the minimum cytotoxic concentration (MCC) or the compound concentration that caused a microscopically detectable alteration of cell morphology. Ganciclovir and cidofovir are included as positive controls for HCMV testing, whereas acyclovir and brivudin are used as reference drugs in the VZV assays. The data are shown in Table 3.

a EC50 = effective concentration (expressed in μΜ) required to reduce virus-induced cytopathicity by 50%. ^b MCC = minimum concentration (expressed in μΜ) required to cause a microscopically detectable alteration of cell morphology.

c CC50 = cytotoxic concentration (expressed in μΜ) required to reduce cell growth by 50%. ^d ND = not determined

Table 3

References

1 . Pertusati, R; Hinsinger, K.; Flynn, A. S.; Powell, N.; Tristram, A.; Balzarini, J.; McGuigan, C. Eur. J. Med. Chem. 2014, 78, 259-268.

2. Pomeisl, K.; Pohl, R.; Holy, A.; Votruba, I. Collect. Czech. Chem. Commun. 2005, 70, 1465-1481 .

3. Jindrich, J.; Holy, A.; Dvorakova, H. Collect. Czech. Chem. Commun. 1993, 58, 1645- 1667.

4. Baszczynski, O.; Hockova, D.; Janeba, Z.; Holy, A.; Jansa, P.; Dracinsky, M.; Keough, D. T; Guddat, L. W. Eur. J. Med. Chem. 2013, 67, 81-89.

Claims

1. A compound of formula I:

wherein

B is any natural or modified nucleobase

R¹ has the general formula I I

II

wherein

or R² has the general formula I I wherein R¹ and R² can be identical or different;

2. The compound according to claim 1 , wherein B is selected from the group of adenine, thymine, cytosine and guanine.

3. The compound according to claim 1 or 2, wherein R² is O-Ph.

4. The compound according to any one of claims 1 to 3, wherein R³ is selected from Ci- Cio alkyl.

6. The compound according to any one of claims 1 to 5, wherein X is C1-C10 alkyl and R⁵ is C1-C10 alkyl.

7. The compound according to any of claims 1 to 6, wherein R² is O-Ph, and wherein R¹ is

9. A compound according to any one of claims 1 to 8 for use as a medicine.

10. A compound according to any one of claims 1 to 8 for use as a medicine for the prevention or treatment of a viral infection in an animal, mammal or human.

1 1 . The compound according to claim 10, wherein said viral infection is an infection with the hepatitis B virus (HBV), the human immunodeficiency virus (HIV), varicella-zoster virus (VZV), cytomegalovirus (CMV), vaccinia virus (VV), herpes simplex virus (HSV), BK virus, Epstein-barr virus (EBV), papillomavirus, Monkeypox virus, Cowpox virus, hepatitis C virus (HCV), respiratory syncytial virus (RSV), dengue virus, influenza virus, adenovirus, parainfluenza virus and/or rhinovirus.

12. A compound according to any one of claims 1 to 8 for use as a medicine for the prevention or treatment of a proliferative disorder such as cancer in an animal, mammal or human.

13. A pharmaceutical composition comprising a therapeutically effective amount of a compound according to any one of claims 1 to 8 and one or more pharmaceutically acceptable excipients.

14. The pharmaceutical composition according to claim 13, further comprising one or more biologically active drugs being selected from the group consisting of antiviral drugs and/or antiproliferative drugs.

15. A method of prevention or treatment of a viral infection in an animal, mammal or human, comprising the administration of a therapeutically effective amount of a compound according to any one of claims 1 to 8, optionally in combination with one or more pharmaceutically acceptable excipients.

16. A method of prevention or treatment of a proliferative disorder in an animal, mammal or human, comprising the administration of a therapeutically effective amount of a compound according to any one of claims 1 to 8, optionally in combination with one or more pharmaceutically acceptable excipients.