Alternative synthesis of 9-{3-[(diisopropoxyphosphoryl)-methoxy]-2- hydroxypropyl}adenine and its free phosphonates substituted at the c-8 position of purine base

Article abstract of DOI:10.1135/cccc2009569

For its high therapeutic effect, (S)-9-[3-hydroxy-2-(phosphonomethoxy) propyl]adenine (HPMPA) is an important member of a class of acyclic nucleoside phosphonates (ANPs). Although its constitutional isomer, 9-[2-hydroxy-3- (phosphonomethoxy)propyl]adenine (iso-HPMPA), exhibits no antiviral activity, our general interest in C-8 substituted adenine ANPs led us to prepare certain iso-HPMPA derivatives modified at the C-8 position of adenine. Novel alkylating agent, diisopropyl {[2-(tetrahydro-2-pyranyl)oxy-3-tosyloxypropoxy]-methyl} phosphonate (9), was prepared by procedure starting from allyl alcohol (4). 9-{3-[(Diisopropoxyphosphoryl)methoxy]-2-hydroxypropyl}adenine (12) was prepared by alkylation of adenine with the alkylating agent 9 followed by acid hydrolysis, although elimination by-product 9-{3-[(diisopropoxyphosphoryl) methoxy]prop-1-enyl}adenine (11) predominated in the reaction mixture. Bromination of the compound 12 gave 8-bromoadenine derivative 13 quantitatively. Nucleophilic substitutions of the bromine atom of compound 13 with N-and O-nucleophiles, followed by phosphonate deprotection, afforded the free phoshonic acids 15-18.

Full text of DOI:10.1135/cccc2009569

Acyclic Nucleoside Phosphonates
371
ALTERNATIVE SYNTHESIS OF 9-{3-[(DIISOPROPOXYPHOSPHORYL)-
METHOXY]-2-HYDROXYPROPYL}ADENINE AND ITS FREE
PHOSPHONATES SUBSTITUTED AT THE C-8 POSITION OF PURINE
BASE
1,
2
3
Zlatko JANEBA *, Milena MASOJÍDKOVÁ and Antonín HOLÝ
Institute of Organic Chemistry and Biochemistry, Academy of Sciences of the Czech Republic, v.v.i.,
1
Flemingovo nám. 2, 166 10 Prague 6, Czech Republic; e-mail: janeba@uochb.cas.cz,
2
3
masojidkova@uochb.cas.cz, holy@uochb.cas.cz
Received December 17, 2009
Accepted January 7, 2010
Published online March 29, 2010
For its high therapeutic effect, (S)-9-[3-hydroxy-2-(phosphonomethoxy)propyl]adenine
(HPMPA) is an important member of a class of acyclic nucleoside phosphonates (ANPs).
Although its constitutional isomer, 9-[2-hydroxy-3-(phosphonomethoxy)propyl]adenine
(iso-HPMPA), exhibits no antiviral activity, our general interest in C-8 substituted adenine
ANPs led us to prepare certain iso-HPMPA derivatives modified at the C-8 position of ade-
nine. Novel alkylating agent, diisopropyl {[2-(tetrahydro-2-pyranyl)oxy-3-tosyloxypropoxy]-
methyl}phosphonate (9), was prepared by procedure starting from allyl alcohol (4).
9-{3-[(Diisopropoxyphosphoryl)methoxy]-2-hydroxypropyl}adenine (12) was prepared by
alkylation of adenine with the alkylating agent 9 followed by acid hydrolysis, although
elimination by-product 9-{3-[(diisopropoxyphosphoryl)methoxy]prop-1-enyl}adenine (11)
predominated in the reaction mixture. Bromination of the compound 12 gave 8-bromo-
adenine derivative 13 quantitatively. Nucleophilic substitutions of the bromine atom of
compound 13 with N- and O-nucleophiles, followed by phosphonate deprotection, afforded
the free phoshonic acids 15–18.
Keywords: Acyclic nucleoside phosphonates; Acyclic nucleotide analogues; HPMPA;
Alkylation; Nucleophilic substitution; Intramolecular cyclization.
Structural modifications of nucleosides and nucleotides have been for a
long time significant means of research in medicinal and bioorganic chem-
istry, biotechnology and other biological applications. Acyclic nucleoside
phosphonates (ANPs), or acyclic nucleotide analogues, have certainly occu-
pied very important position in the field1. Among other side chain modifi-
cations, N-[3-hydroxy-2-(phosphonomethoxy)propyl] (HPMP) derivatives2
of purine and pyrimidine bases have been studied in detail owing to their
antiviral properties and were found to be selectively active against DNA
Collect. Czech. Chem. Commun. 2010, Vol. 75, No. 3, pp. 371–381
© 2010 Institute of Organic Chemistry and Biochemistry
doi:10.1135/cccc2009569

372
Janeba, Masojídková, Holý:
viruses (herpes viruses, adenoviruses, poxviruses, iridoviruses, polyoma and
papilomaviruses)3. (S)-HPMPC (Cidofovir)2,4 has been approved for treat-
ment of CMV retinitis in AIDS patients (Vistide)5, although it also is effi-
cacious in other viral diseases6.
While the (S)-enantiomer is active in adenine series (HPMPA, 1, Chart 1)3,
its 3′-O-phosphonomethyl isomer (iso-HPMPA, 2)7 exhibits no antiviral ac-
tivity3a. Recently, certain ANPs were found to selectively inhibit hypoxanthine-
guanine-xanthine phosphoribosyltransferase of Plasmodium falciparum and
have been studied as potential anti-malarials8. These findings revived our
interest in the iso-HPMPA analogues, especially those modified at the C-8
position of the purines.
CHART 1
Some 10 years ago, as a part of the Ph.D. Thesis of the first author (Z.J.)9,
a synthesis of racemic 9-{3-[(diisopropoxyphosphoryl)methoxy]-2-hydroxy-
propyl}adenine was designed starting from allyl alcohol. Later, another syn-
thetic approach to iso-HPMP derivatives of purine bases using oxirane ring
opening was developed10 and immunobiological properties of these com-
pounds were studied11.
Within the extensive systematic studies of various types of acyclic
nucleoside and nucleotide analogues we have paid substantial attention to
a group of compounds modified at the C-8 position of purine bases12. These
compounds showed to be interesting both for their biological properties13
and for the synthetic and mechanistic studies (e.g. formation and ring
opening of 8,O-anhydro derivatives, e.g. compound 3, Chart 1)12a,12b,14. The
intramolecular cyclizations to form the corresponding anhydro derivatives
Collect. Czech. Chem. Commun. 2010, Vol. 75, No. 3, pp. 371–381

Acyclic Nucleoside Phosphonates
373
could be studied in the case of the HPMP analogues, where the free hydrox-
yl group in the acyclic moiety can participate in the cyclization process.
Due to possible important biological properties of the modified iso-HPMP
analogues we decided to describe the original procedure for the preparation
of racemic iso-HPMPA 9. An additional aim of this study was to prepare
8-substituted iso-HPMPA analogues for SAR studies, namely 8-hydroxy
(8-oxo) and N-substituted 8-aminoadenine derivatives. Also, compound of
special interest was 8,2′-O-anhydro analogue, a constitutional isomer of for-
merly described 8,3′-O-anhydro HPMPA derivative 3 (Chart 1)12b
.
RESULTS AND DISCUSSION
Allyl alcohol (4) was chosen as a convenient starting material for the syn-
thesis of suitable alkylating agent leading to iso-HPMPA analogue. Allyl
alcohol (4) was converted by chloromethylation to chloro derivative 5 15
which under Arbuzov reaction16 conditions with triisopropyl phosphite
afforded diisopropyl phosphonate 6 17 in 32% overall yield from the start-
ing alcohol 4 (Scheme 1). The double bond of the compound 6 was
hydroxylated under catalysis of OsO4 to afford 2,3-dihydroxy derivative 7
in moderate yield. The compound 7 was tosylated at the primary hydroxyl
to give derivative 8. Then the secondary hydroxyl of compound 8 was pro-
tected by the reaction with 3,4-dihydro-2H-pyran to afford compound 9 in
37% overall yield from 7.
SCHEME 1
Alkylation of adenine (10) with the tosylate 9 using DBU, followed by
treatment of the crude alkylation product with 1 M HCl at 60 °C for 2 h
Collect. Czech. Chem. Commun. 2010, Vol. 75, No. 3, pp. 371–381

374
Janeba, Masojídková, Holý:
gave a mixture of two products (Scheme 2): the desired product 12 (11%)
and compound 11 (31%) which was formed by elimination of the hydroxyl
group at C-2′ under the acid conditions. The value of vicinal coupling con-
stants18 of H-1′ and H-2′ protons (J(1′,2′) = J(2′,1′) = 14.5 Hz) confirmed the
trans isomer of 11 being the sole product of the elimination process.
SCHEME 2
Although, the yield of the desired iso-HPMPA derivative 12 was lower
compared to the procedure of Krečmerová et al.10 (31% of diethyl ester of
iso-HPMPA acetylated at C-2′ position), also their procedure suffers from
Collect. Czech. Chem. Commun. 2010, Vol. 75, No. 3, pp. 371–381

Acyclic Nucleoside Phosphonates
375
formation of a by-product (in this case a cyclic phosphonate) and acetylat-
ion of the reaction mixture was necessary to isolate the desired product. Re-
action conditions of our procedure (alkylation/deprotection) were not
further optimized.
Nucleophilic substitutions of bromine atom at the C-8 position of adenine
with corresponding O- and N-nucleophiles were used for modifications of
9-{3-[(diisopropoxyphosphoryl)methoxy]-2-hydroxypropyl}adenine (12).
Bromination of the adenine derivative 12 with bromine in a mixture of
dioxane and 10% aqueous solution of disodium hydrogen phosphate af-
forded 8-bromoadenine derivative 13 in 96% yield (Scheme 2). The attempt
to brominate compound 11 under the same reaction conditions failed,
probably because of the presence of the double bond at the aliphatic side
chain and the reaction led to decomposition of the starting material.
Similarly as in the synthesis of 8,3′-O-anhydro HPMPA derivative 3 with
annellated six-membered ring (Chart 1)12b, 8-bromoadenine derivative 13
gave on treatment with 2 equivalents of sodium hydride in DMF 8,2′-O-
anhydro derivative 14 with annellated five-membered cycle (Scheme 2).
Free phosphonic acid 15, the constitutional isomer of the compound 3, was
prepared in 26% yield by the standard procedure using TMSBr in acetoni-
trile followed by hydrolysis. The low yield of the compound 15 could be ex-
plained by susceptibility of the O-anhydro compounds to hydrolysis under
acidic conditions12b
.
Treatment of the 8-bromoadenine derivative 13 with sodium acetate in
acetic acid, followed by deprotection with TMSBr, gave 6-amino-7H-pu-
rine-8(9H)-one analogue 16 (Scheme 2). 8-(N-Methylamino)adenine deriva-
tive 17 and 8-(N,N-dimethylamino)adenine derivative 18 were obtained by
reactions of the compound 13 with ethanolic solutions of either methyl-
amine or dimethylamine, respectively, in an autoclave, followed by the
deprotection of the phosphonate group.
In conclusion, synthesis of alkylating agent 9 was developed starting
from allyl alcohol (4). Alkylation of adenine (10) with the tosylate 9, fol-
lowed by acid removal of the tetrahydro-2-pyranyl group, afforded
9-{3-[(diisopropoxyphosphoryl)methoxy]-2-hydroxypropyl}adenine (12)
and the unsaturated derivative 11. Bromination of the compound 12 gave
8-bromoadenine compound 13 quantitatively. Nucleophilic substitutions
of the bromine atom of the compound 13, followed by diester removal,
afforded the free phoshonic acids 15–18. Biological activities of the final
compounds will be examined.
Collect. Czech. Chem. Commun. 2010, Vol. 75, No. 3, pp. 371–381

376
Janeba, Masojídková, Holý:
EXPERIMENTAL
Melting points were determined on a Büchi Melting Point B-545 apparatus and are uncor-
rected. NMR spectra (δ, ppm; J, Hz) were recorded on a Bruker Avance 500 (1H at 500 MHz,
13C at 125.8 MHz) in DMSO-d6, or D2O (with addition of NaOD to improve solubility).
Chemical shifts were referenced to the solvent signal (DMSO-d6 for 1H NMR δ 2.5 ppm and
for 13C δ 39.7). Chemical shifts in D2O were referenced to 1,4-dioxane for 1H NMR δ 3.75
and for 13C NMR δ 67.19. UV spectra were measured on DU Spectrophotometer with solu-
tions in H2O. Mass spectra were measured on a ZAB-EQ (VG Analytical) spectrometer using
FAB (ionization by Xe, accelerating voltage 8 kV, glycerol matrix). The elemental analyses
were obtained on a Perkin–Elmer CHN Analyzer 2400, Series II Sys (Perkin–Elmer). Column
chromatography and thin-layer chromatography (TLC) were performed using Silica gel 60
(Fluka) and Silufol Silica gel 60 F254 foils (Merck), respectively. Solvents were evaporated at
2 kPa and bath temperature 30–60 °C. The compounds were dried at 2 kPa over P2O5.
Diisopropyl [(Prop-2-en-1-yloxy)methyl]phosphonate (6)17
Gaseous hydrogen chloride was introduced into the cooled (0 °C) mixture of allyl alcohol
(4; 116.2 g, 2.0 mol) and formaldehyde (180.2 g, 6.0 mol) for 3 h. Then the upper organic
layer was decanted off, dried (CaCl2), and the drying agent was filtered off. Distillation of
the residue afforded 3-(chloromethoxy)prop-1-ene15 (5; 145.0 g, 68%) as a colorless oil, b.p.
108–110 °C.
A mixture of compound 5 (145.0 g, 1.4 mol) and triisopropyl phosphite (291.5 g, 1.4 mol)
was refluxed at 120 °C for 7 h and distilled in vacuo affording compound 6 (155.9 g, 47%)
as an oil, b.p. 90–92 °C at 198 kPa. MS (FAB), m/z: 237 [M + H]+ (40), 153 [M – 2 (iPr) + H]+
(100).
Diisopropyl [(2-Hydroxy-3-tosyloxypropoxy)methyl]phosphonate (8)
A mixture of compound 6 (47.3 g, 0.2 mol), 50% aqueous ethanol (350 ml), NaClO3 (59.6 g,
0.6 mol) and OsO4 (0.20 g, 0.80 mmol) in CCl4 (20 ml) was heated at 50 °C for 6 h and
then stirred at room temperature overnight. Organic solvents were evaporated and the re-
sulting solution was saturated with NaCl. The solution was extracted with diethyl ether (3 ×
100 ml) and the organics were dried (MgSO4). Filtration, evaporation of organics and distil-
lation in vacuo gave compound 7 (19 g, 35%) as a colorless oil, b.p. 135–137 °C at 9.9 kPa.
MS (FAB), m/z: 271 [M + H]+ (100).
A solution of tosyl chloride (19 g, 0.10 mol) in pyridine (30 ml) was added dropwise with
stirring to ice-cool solution of compound 7 (30 g, 0.11 mol) and DMAP (0.3 g) in pyridine
(80 ml). The mixture was stirred at 0 °C for 3 h and set aside at room temperature over-
night. Methanol (25 ml) was added and the mixture was stirred at room temperature for
1 h. The mixture was diluted with EtOAc (500 ml), washed with water (2 × 100 ml), and the
organics were evaporated in vacuo. The residue was codistilled with toluene (3 × 50 ml) and
dried over phosphorus pentoxide to give compound 8 (28 g, 60%) as a thick oil. 1H NMR
(DMSO-d6): 1.19 d, 6 H, J(CH3,CH) = 6.1 (CH3); 1.22 d, 6 H, J(CH3,CH) = 6.1 (CH3); 2.42 s,
3 H (CH3-arom); 3.40 dd, 1 H, J(3b,2) = 6.1, J(gem) = 10.0 (H-3b); 3.46 dd, 1 H, J(3a,2) = 5.1,
J(gem) = 10.0 (H-3a); 3.69 d, 2 H, J(P,CH) = 7.9 (PCH2); 3.78 m, 1 H (H-2); 3.85 dd, 1 H,
J(1b,2) = 6.5, J(gem) = 9.9 (H-1b); 3.98 dd, 1 H, J(1a,2) = 3.4, J(gem) = 9.9 (H-1a); 4.56 m,
2 H (POCH); 5.34 d, 1 H, J(OH,2) = 5.4 (OH); 7.48 d, 2 H, J = 8.3 (arom); 7.77 d, 2 H, J = 8.3
Collect. Czech. Chem. Commun. 2010, Vol. 75, No. 3, pp. 371–381

Acyclic Nucleoside Phosphonates
377
(arom). 13C NMR (DMSO-d6): 21.26 (CH3-arom); 23.84 d, 2 C, J(P,C) = 4.9 (CH3); 23.96 d,
2 C, J(P,C) = 3.9 (CH3); 65.36 d, J(P,C) = 164.1 (PC); 67.01 (C-2); 70.29 d, 2 C, J(P,C) = 5.9
(POC); 72.05 (C-1); 73.19 d, J(P,C) = 10.7 (C-3); 127.79 (2 C, C-arom); 130.33 (2 C, C-arom),
132.43 (C-arom); 145.07 (C-arom).
Diisopropyl {[2-(Tetrahydro-2-pyranyl)oxy-3-tosyloxypropoxy]methyl}phosphonate (9)
A mixture of tosyl derivative 8 (28.0 g, 66.0 mmol), 3,4-dihydro-2H-pyran (12.3 g, 13.3 ml,
146.2 mmol) and HCl/DMF (4.5 mol/l, 2 ml) was stirred at room temperature for 4 h. The
mixture was diluted with diethyl ether (150 ml) and washed (3 × 50 ml of water, 50 ml of
1 M NaHCO3 aqueous solution, and 50 ml of water). The organic solvent was taken down in
vacuo, the residue was codistilled with ethanol (3 × 50 ml) and dried over phosphorus
pentoxide to give compound 9 (20.8 g, 62%) as a thick oil. 1H NMR (DMSO-d6): 1.19 d, 6 H,
J(CH3,CH) = 6.1 (CH3); 1.22 d, 6 H, J(CH3,CH) = 6.1 (CH3); 1.53–1.89 m, 6 H (pyr); 2.41 s,
3 H (CH3-arom); 3.53 dd, 1 H, J(3b,2) = 6.1, J(gem) = 10.0 (H-3b); 3.95 dd, 1 H, J(3a,2) = 5.1,
J(gem) = 10.0 (H-3a); 3.61 m, 1 H (pyr); 3.69 d, 2 H, J(P,CH) = 7.9 (PCH2); 3.74 m, 1 H (pyr);
4.35 dd, 1 H, J(1b,2) = 6.5, J(gem) = 9.9 (H-1b); 4.46 dd, 1 H, J(1a,2) = 3.4, J(gem) = 9.9
(H-1a); 4.68 m, 1 H (H-2); 4.60 m, 2 H (POCH); 4.76 m, 1 H (pyr); 7.46 d, 2 H, J = 8.3
(arom); 7.75 d, 2 H, J = 8.3 (arom).
Alkylation of Adenine with Tosylate 9
A mixture of adenine (10; 5.0 g, 37.0 mmol), DBU (6.2 g, 6.1 ml, 40.7 mmol) and com-
pound 9 (21 g, 41.3 mmol) in DMF (70 ml) was stirred at 110 °C for 22 h. The solvent was
evaporated in vacuo, the residue codistilled with toluene (3 × 30 ml), and dissolved in chlo-
roform (150 ml). The solution was washed with water (3 × 25 ml) and the organic solvent
was taken down. The residue was heated in 1 M HCl (120 ml) at 60 °C for 2 h. The mixture
was neutralized with 1 M NaOH and extracted with chloroform (3 × 50 ml). The extract was
chromatographed on silica gel (10% MeOH in CHCl3) and fractions were crystallized from
EtOH to give compounds 11 (4.2 g, 31%) and 12 (1.6 g, 11%).
9-{3-[(Diisopropoxyphosphoryl)methoxy]prop-1-enyl}adenine (11). White crystals, m.p. 123–
125 °C. MS (FAB), m/z: 370 [M + H]+ (100). 1H NMR (DMSO-d6): 1.25 d, 6 H, J(CH3,CH) =
6.1 (CH3); 1.255 d, 6 H, J(CH3,CH) = 6.1 (CH3); 3.78 d, 2 H, J(P,CH) = 8.6 (PCH2); 4.24 d,
2 H, J(3′,2′) = 6.4 (H-3′); 4.62 m, 2 H (POCH); 6.69 dt, 1 H, J(2′,3′) = 6.4, J(2′,1′) = 14.5
(H-2′); 7.31 d, 1 H, J(1′,2′) = 14.5 (H-1′); 7.38 brs, 2 H (NH2), 8.19 and 8.48 s, 2 × 1 H (H-2
and H-8). 13C NMR (DMSO-d6): 23.88 d, 2 C, J(P,C) = 4.9 (CH3); 23.99 d, 2 C, J(P,C) = 3.9
(CH3); 46.50 (C-1′); 65.50 d, J(P,C) = 163.5 (PC); 67.68 (C-2′); 70.42 d, 2 C, J(P,C) = 6.4
(POC); 74.85 d, J(P,C) = 10.3 (C-3′); 118.78 (C-5); 141.74 (C-8); 149.86 (C-4); 152.46 (C-2);
156.11 (C-6). For C15H26N5O5P (387.4) calculated: 46.51% C, 6.76% H, 18.08% N, 8.00% P;
found: 46.42% C, 6.78% H, 17.82% N, 8.10% P.
(R,S)-9-{3-[(Diisopropoxyphosphoryl)methoxy]-2-hydroxypropyl}adenine (12). White crystals,
m.p. 125–127 °C. MS (FAB), m/z: 388 [M + H]+ (100). 1H NMR (DMSO-d6): 1.24 d, 6 H,
J(CH3,CH) = 6.2 (CH3); 1.25 d, 6 H, J(CH3,CH) = 6.2 (CH3); 3.49 dd, 1 H, J(3′b,2′) = 5.2,
J(gem) = 12.4 (H-3′b); 3.52 dd, 1 H, J(3′a,2′) = 5.0, J(gem) = 12.4 (H-3′a); 3.80 d, 2 H,
J(P,CH) = 7.8 (PCH2); 4.01 m, 1 H (H-2′); 4.04 dd, 1 H, J(1′b,2′) = 8.1, J(gem) = 12.4 (H-1′b);
4.26 dd, 1 H, J(1′a,2′) = 2.2, J(gem) = 12.4 (H-1′a); 4.61 m, 2 H (POCH); 5.34 d, 1 H,
J(OH,2′) = 5.1 (OH); 7.21 brs, 2 H (NH2); 8.05 and 8.13 s, 2 × 1 H (H-2 and H-8). 13C NMR
Collect. Czech. Chem. Commun. 2010, Vol. 75, No. 3, pp. 371–381

378
Janeba, Masojídková, Holý:
(DMSO-d6): 23.88 d, 2 C, J(P,C) = 4.9 (CH3); 23.99 d, 2 C, J(P,C) = 3.9 (CH3); 46.50 (C-1′);
65.50 d, J(P,C) = 163.5 (PC); 67.68 (C-2′); 70.42 d, 2 C, J(P,C) = 6.4 (POC); 74.85 d, J(P,C) =
10.3 (C-3′); 118.78 (C-5); 141.74 (C-8); 149.86 (C-4); 152.46 (C-2); 156.11 (C-6). For
C
15H26N5O5P (387.4) calculated: 46.51% C, 6.76% H, 18.08% N, 8.00% P; found: 46.42% C,
6.78% H, 17.82% N, 8.10% P.
(R,S)-8-Bromo-9-{3-[(diisopropoxyphosphoryl)methoxy]-2-hydroxypropyl}adenine (13)
Bromine (0.10 ml, 0.31 g, 3.9 mmol) was added to a suspension of compound 12 (0.80 g,
2.1 mmol) in a mixture of dioxane (40 ml) and 10% aqueous solution of sodium hydrogen-
phosphate (25 ml) and the mixture was stirred at room temperature for 15 h. A concen-
trated solution of sodium hydrogensulfite was added until the mixture became colorless and
the product was taken up in chloroform (5 × 30 ml). The chloroform extract was dried over
anhydrous magnesium sulfate, filtered, and the solvent was evaporated. Purification on a
silica gel plate (15% MeOH in CHCl3) and crystallization (EtOAc) afforded compound 13
(0.92 g, 96%) as yellowish crystals, m.p. 138–140 °C. MS (FAB), m/z: 466/468 [M + H]+ (100).
1H NMR (DMSO-d6): 1.25 d, 12 H, J(CH3,CH) = 6.1 (CH3); 3.55 dd, 1 H, J(3′b,2′) = 5.1,
J(gem) = 10.1 (H-3′b); 3.59 dd, 1 H, J(3′a,2′) = 5.1, J(gem) = 10.1 (H-3′a); 3.78 dd, 1 H,
J(P,CH) = 7.7, J(gem) = 13.9 (PCH2); 3.82 dd, 1 H, J(P,CH) = 8.8, J(gem) = 13.9 (PCH2);
4.07 dd, 1 H, J(1′b,2′) = 8.1, J(gem) = 12.8 (H-1′b); 4.13 m, 1 H (H-2′); 4.17 dd, 1 H,
J(1′a,2′) = 3.4, J(gem) = 12.8 (H-1′a); 4.61 m, 2 H (POCH); 4.96 d, 1 H, J(OH,2′) = 5.2 (OH);
7.36 brs, 2 H (NH2); 8.12 s, 1 H (H-2). 13C NMR (DMSO-d6): 23.96 d, 2 C, J(P,C) = 4.5 (CH3);
24.05 d, 2 C, J(P,C) = 3.7 (CH3); 47.68 (C-1′); 65.50 d, J(P,C) = 163.6 (PC); 67.30 (C-2′);
70.38 d, J(P,C) = 6.3 (POC); 70.40 d, J(P,C) = 6.3 (POC); 75.00 d, J(P,C) = 10.2 (C-3′); 119.20
(C-5); 127.39 (C-8); 151.21 (C-4); 152.79 (C-2); 154.91 (C-6).
Diisopropyl (R,S)-{[(4-Amino-7,8-dihydro-[1,3]oxazolo[3,2-e]purin-7-yl)methoxy]methyl}-
phosphonate (14)
A mixture of compound 13 (0.6 g, 1.3 mmol) and 60% sodium hydride suspension (100 mg,
2.6 mmol) in DMF (20 ml) was stirred at room temperature for 1 h. The mixture was neu-
tralized (AcOH) and taken down in vacuo. The residue was codistilled with toluene (2 ×
10 ml) and purified on a silica gel plate (15% MeOH in CHCl3). Crystallization (EtOAc) af-
forded compound 14 (0.36 g, 72%) as white crystals, m.p. 150–152 °C. MS (FAB), m/z: 386
[M + H]+ (30), 302 [M – 2 (iPr) + H]+ (100). 1H NMR (DMSO-d6): 1.14 d, 6 H, J(CH3,CH) = 6.1
(CH3); 1.17 d, 3 H, J(CH3,CH) = 6.1 (CH3); 1.18 d, 3 H, J(CH3,CH) = 6.1 (CH3); 3.84 dd, 1 H,
J(P,CH) = 9.2, J(gem) = 13.9 (PCH2); 3.87 dd, 1 H, J(P,CH) = 9.2, J(gem) = 13.9 (PCH2);
3.88 dd, 1 H, J(3′b,2′) = 4.4, J(gem) = 11.6 (H-3′b); 3.98 dd, 1 H, J(3′a,2′) = 2.6, J(gem) = 11.6
(H-3′a); 4.07 dd, 1 H, J(1′b,2′) = 6.0, J(gem) = 9.4 (H-1′b); 4.37 dd, 1 H, J(1′a,2′) = 8.8,
J(gem) = 9.4 (H-1′a); 4.50 m, 2 H (POCH); 5.63 m, 1 H (H-2′); 6.74 brs, 2 H (NH2), 7.98 s,
1 H (H-2). 13C NMR (DMSO-d6): 23.76 d, 2 C, J(P,C) = 3.9 (CH3); 23.91 d, 2 C, J(P,C) = 3.9
(CH3); 42.13 (C-1′); 65.39 d, J(P,C) = 164.1 (PC); 70.44 d, J(P,C) = 6.8 (POC); 70.47 d,
J(P,C) = 6.8 (POC); 72.60 d, J(P,C) = 10.7 (C-3′); 86.70 (C-2′); 120.34 (C-5); 147.10 (C-4);
150.54 (C-2); 154.00 (C-6); 160.25 (C-8). For C15H24N5O5P (385.4) calculated: 46.75% C,
6.28% H, 18.17% N, 8.04% P; found: 46.46% C, 6.28% H, 17.82% N, 7.97% P.
Collect. Czech. Chem. Commun. 2010, Vol. 75, No. 3, pp. 371–381

Acyclic Nucleoside Phosphonates
379
(R,S)-{[(4-Amino-7,8-dihydro[1,3]oxazolo[3,2-e]purin-7-yl)methoxy]methyl}-
phosphonic Acid (15)
Method A 19: A mixture of compound 14 (386 mg, 1.0 mmol) and TMSBr (1.0 ml) in aceto-
nitrile (5 ml) was stirred overnight at ambient temperature, then evaporated and codistilled
with acetonitrile (10 ml). The residue was dissolved in water and alkalized with aqueous am-
monia. The mixture was evaporated in vacuo and the residue was deionized on a Dowex
50X8 (H+) column (50 ml) under standard conditions (washing with water and elution with
2.5% aqueous ammonia). The product was purified on a Dowex 1X2 (acetate) column by
elution with linear gradient of acetic acid (0–0.5 mol/l, 1 l each). The UV-absorbing fractions
were evaporated in vacuo and the residue was crystallized to afford compound 15 (79 mg,
26%) as white crystals (H2O), m.p. > 270 °C (dec.), EUp = 0.80. MS (FAB), m/z: 302 [M + H]+
(100). 1H NMR (D2O): 3.61 d, 2 H, J(P,CH) = 8.3 (PCH2); 4.01 dd, 1 H, J(3′b,2′) = 6.0,
J(gem) = 12.0 (H-3′b); 4.06 dd, 1 H, J(3′a,2′) = 3.3, J(gem) = 12.0 (H-3′a); 4.24 dd, 1 H,
J(1′b,2′) = 6.9, J(gem) = 9.5 (H-1′b); 4.47 dd, 1 H, J(1′a,2′) = 8.9, J(gem) = 9.5 (H-1′a); 5.81 m,
1 H (H-2′); 8.09 s, 1 H (H-2). 13C NMR (D2O): 45.43 (C-1′); 72.53 d, J(P,C) = 149.4 (PC);
74.86 d, J(P,C) = 8.8 (C-3′); 91.40 (C-2′); 122.51 (C-5); 149.01 (C-4); 153.13 (C-2); 155.82
(C-6); 163.91 (C-8). For C9H12N5O5P (301.2) calculated: 35.89% C, 4.02% H, 23.25% N,
10.28% P; found: 35.76% C, 4.15% H, 22.99% N, 10.17% P. UV, λmax (εmax): (pH 2) 262
(12 100); (pH 12) 262 (12 400).
(R,S)-6-Amino-9-[2-hydroxy-3-(phosphonomethoxy)propyl]-7H-purin-8(9H)-one (16)
A mixture of compound 13 (933 mg, 2 mmol), acetic acid (15 ml) and sodium acetate (1.31 g,
16 mmol) was refluxed for 8 h. Volatiles were evaporated in vacuo and the residue was
codistilled with water (3 × 10 ml). The residue was stirred with sodium methoxide (10 ml,
125 mmol) at 50 °C for 2 h, neutralized with HCl, and volatiles were evaporated in vacuo.
The residue was deionized on a Dowex 50X8 (H+) column (50 ml, elution with water fol-
lowed by 2.5% aqueous ammonia). The UV-absorbing ammonia eluate was collected and
evaporated in vacuo. The crude intermediate was dried over phosphorus pentoxide over-
night and then treated with TMSBr under standard conditions (method A) to give com-
pound 16 (237 mg, 37%) as yellowish crystals (H2O), m.p. 275 °C, EUp = 0.81. MS (FAB),
m/z: 320 [M + H]+ (100). 1H NMR (D2O): 3.47 dd, 1 H, J(P,CHb) = 8.9, J(gem) = 12.6 (PCHb);
3.52 dd, 1 H, J(P,CHa) = 8.7, J(gem) = 12.6 (PCHa); 3.54 dd, 1 H, J(3′b,2′) = 7.2, J(gem) =
10.8 (H-3′b); 3.60 dd, 1 H, J(3′a,2′) = 3.1, J(gem) = 10.8 (H-3′a); 3.89 dd, 1 H, J(1′b,2) = 5.9,
J(gem) = 14.4 (H-1′b); 3.95 dd, 1 H, J(1′a,2′) = 7.3, J(gem) = 14.4 (H-1′a); 4.24 m, 1 H (H-2′);
7.96 s, 1 H (H-2). 13C NMR (D2O): 46.57 (C-1′); 70.90 (C-2′); 72.32 d, J(P,C) = 150.3 (PC);
76.48 d, J(P,C) = 11.3 (C-3′); 103.35 (C-5); 146.59 and 146.60 (C-4 and C-6); 150.47 (C-2);
151.52 (C-8). HR MS (FAB) for C9H15N5O6P [M + H] calculated 320.0760, found 320.0761.
UV, λmax (εmax): (pH 2) 281 (10 500); (pH 7) 272 (12 300); (pH 12) 281 (13 900).
(R,S)-9-[2-Hydroxy-3-(phosphonomethoxy)propyl]-8-(N-methylamino)adenine (17)
Method B 12d: A mixture of compound 13 (933 mg, 2 mmol) in ethanolic methylamine so-
lution (33%, 30 ml) was heated in a steel autoclave at 100 °C for 15 h. Then the solvents
were evaporated, the residue was codistilled with ethanol (2 × 15 ml) and dried over phos-
phorus pentoxide overnight. The crude intermediate was treated with TMSBr under standard
conditions (method A) to give compound 17 (299 mg, 45%) as white crystals (H2O), m.p.
Collect. Czech. Chem. Commun. 2010, Vol. 75, No. 3, pp. 371–381

380
Janeba, Masojídková, Holý:
217–219 °C, EUp = 0.75. MS (FAB), m/z: 333 [M + H]+ (100). 1H NMR (D2O + NaOD): 2.98 s,
3 H (NCH3); 3.49 dd, 1 H, J(3′b,2′) = 5.7, J(gem) = 10.5 (H-3′b); 3.50 dd, 1 H, J(P,CHb) = 8.8,
J(gem) = 12.4 (PCHb); 3.53 dd, 1 H, J(P,CHa) = 8.9, J(gem) = 12.4 (PCHa); 3.59 dd, 1 H,
J(3′a,2′) = 3.8, J(gem) = 10.5 (H-3′a); 3.92 dd, 1 H, J(1′b,2′) = 7.1, J(gem) = 15.0 (H-1′b);
3.96 dd, 1 H, J(1′a,2′) = 5.1, J(gem) = 15.0 (H-1′a); 4.13 m, 1 H (H-2′); 7.92 s, 1 H (H-2).
13C NMR (D2O + NaOD): 31.43 (NCH3), 46.66 (C-1′); 70.85 (C-2′); 72.27 d, J(P,C) = 150.5
(PC); 76.43 d, J(P,C) = 11.3 (C-3′); 118.90 (C-5); 151.53 (C-2); 152.59 (C-4); 153.55 (C-6);
157.07 (C-8). For C10H17N6O5P (332.3) calculated: 36.15% C, 5.16% H, 25.29% N, 9.32% P;
found: 35.94% C, 5.24% H, 24.96% N, 9.05% P. UV, λmax (εmax): (pH 2) 275 (11 400);
(pH 12) 279 (13 900).
(R,S)-8-(N,N-Dimethylamino)-9-[2-hydroxy-3-(phosphonomethoxy)propyl]adenine (18)
Treatment of compound 13 (933 mg, 2 mmol) with ethanolic dimethylamine solution (33%,
30 ml) by method B, followed by deprotection (method A) gave compound 18 (270 mg,
39%) as white crystals (H2O), m.p. 250 °C, EUp = 0.77. MS (FAB), m/z: 347 [M + H]+ (100).
1H NMR (D2O + NaOD): 3.04 s, 6 H (NCH3); 3.51 dd, 1 H, J(P,CHb) = 8.9, J(gem) = 12.4
(PCHb); 3.54 dd, 1 H, J(P,CHa) = 8.9, J(gem) = 12.4 (PCHa); 3.56 dd, 1 H, J(3′b,2′) = 6.5,
J(gem) = 10.6 (H-3′b); 3.64 dd, 1 H, J(3′a,2′) = 3.3, J(gem) = 10.6 (H-3′a); 4.19 dd, 1 H,
J(1′b,2′) = 4.8, J(gem) = 15.0 (H-1′b); 4.24 dd, 1 H, J(1′a,2′) = 8.4, J(gem) = 15.0 (H-1′a);
4.36 m, 1 H (H-2′); 8.04 s, 1 H (H-2). 13C NMR (D2O + NaOD): 43.82 (2 C, NCH3), 48.90
(C-1′); 70.00 (C-2′); 72.21 d, J(P,C) = 150.4 (PC); 76.80 d, J(P,C) = 11.3 (C-3′); 118.75 (C-5);
152.80 (C-2); 153.38 (C-4); 155.04 (C-6); 160.01 (C-8). For C11H19N6O5P (346.3) calculated:
38.15% C, 5.53% H, 24.27% N, 8.94% P; found: 37.96% C, 5.67% H, 24.01% N, 8.92% P.
UV, λmax (εmax): (pH 2) 282 (15 800); (pH 12) 278 (16 200).
REFERENCES
1. Reviews: a) Naesens L., Snoeck R., Andrei G., Balzarini J., Neyts J., De Clercq E.:
Antiviral Chem. Chemother. 1997, 8, 1; b) De Clercq E.: J. Med. Microbiol. 1998, 47, 1;
c) De Clercq E.: Clin. Microbiol. Rev. 2003, 16, 569; d) De Clercq E.: Expert Rev. Anti-Infect.
2003, 1, 21; e) De Clercq E., Holý A.: Nat. Rev. Drug Discov. 2005, 4, 928; f) Holý A.:
Antiviral Res. 2006, 71, 248; g) Lee W. A., Martin J. C.: Antiviral Res. 2006, 71, 254.
2. Holý A.: Collect. Czech. Chem. Commun. 1993, 58, 649.
3. a) De Clercq E., Holý A., Rosenberg I., Sakuma T., Balzarini J., Maudgal P. C.: Nature
1986, 323, 464; b) Holý A., Votruba I., Merta A., Černý J., Veselý J., Vlach J., Šedivá K.,
Rosenberg I., Otmar M., Hřebabecký H., Trávníček M., Vonka V., Snoeck R., De Clercq E.:
Antiviral Res. 1990, 13, 295; c) De Clercq E., Holý A.: Antimicrob. Agents Chemother.
1991, 35, 701.
4. a) Neyts J., Snoeck R., Balzarini J., De Clercq E.: Antiviral Res. 1991, 16, 41; b) De Clercq E.:
Rev. Med. Virol. 1993, 3, 85; c) Flores-Aguilar M., Huang J.-S., Wiley C. A., De Clercq E.,
Vuong C., Bergeron-Lynn G. L., Chandler B., Munguia D., Freeman W. R.: J. Infect. Dis.
1994, 169, 642; d) Kirsch L. S., Arevalo J. F., Delapaz E. C., Munguia D., De Clercq E.,
Freeman W. R.: Ophthalmology 1995, 102, 533; e) Naesens L., De Clercq E.: Nucleosides
Nucleotides 1997, 16, 983.
5. a) Hitchcock M. J. M., Jaffe H. S., Martin J. C., Stagg R. J.: Antiviral Chem. Chemother.
1996, 7, 115; b) Lea A. P., Bryson H. M.: Drugs 1996, 52, 225; c) Rahhal F. M., Arevalo
Collect. Czech. Chem. Commun. 2010, Vol. 75, No. 3, pp. 371–381

Acyclic Nucleoside Phosphonates
381
J. F., Munguia D., Taskintuna I., Delapaz E. C., Azen S. P., Freeman W. R.: Ophthalmology
1996, 103, 1078; d) De Clercq E.: Collect. Czech. Chem. Commun. 1998, 63, 480.
6. a) Snoeck R., Van Ranst M., Andrei G., De Clercq E., Dewit S., Poncin M., Clumeck N.:
N. Engl. J. Med. 1995, 333, 943; b) Van Cutsem E., Snoeck R., Van Ranst M., Fiten P.,
Opdenakker G., Geboes K., Janssens J., Rutgeerts P., Van Trappen G., De Clercq E.:
J. Med. Virol. 1995, 45, 230; c) Meadows K. P., Tyring S. K., Pavia A. T., Rallis T. M.: Arch.
Dermatol. 1997, 133, 987; d) Blick G., Whiteside M., Griegor P., Hopkins U., Garton T.,
Lagravinese L.: Clin. Infect. Dis. 1998, 26, 191; e) Snoeck R., Wellens W., Desloovere C.,
Van Ranst M., Naesens L., De Clercq E., Feenstra L.: J. Med. Virol. 1998, 54, 291; f) Neyts J.,
Sadler R., De Clercq E., Raab-Traub N., Pagano J. S.: Cancer Res. 1998, 58, 384.
7. a) Holý A., Rosenberg I.: Collect. Czech. Chem. Commun. 1987, 52, 2775; b) Rosenberg I.,
Holý A.: Collect. Czech. Chem. Commun. 1987, 52, 2792.
8. a) Keough D. T., Hocková D., Holý A., Naesens L. M. J., Skinner-Adams T. S., de Jersey J.,
Guddat L. W.: J. Med. Chem. 2009, 52, 4391; b) Hocková D., Holý A., Masojídková M.,
Keough D. T., de Jersey J., Guddat L. W.: Bioorg. Med. Chem. 2009, 17, 6218.
9. Janeba Z.: Ph.D. Thesis. Institute of Organic Chemistry and Biochemistry, Academy of
Sciences of the Czech Republic, Prague 2001.
10. Krečmerová M., Masojídková M., Holý A.: Collect. Czech. Chem. Commun. 2004, 69,
1889.
11. Potměšil P., Krečmerová M., Kmoníčková E., Holý A., Zídek Z.: Eur. J. Pharmacol. 2006,
540, 191.
12. a) Janeba Z., Holý A., Votavová H., Masojídková M.: Collect. Czech. Chem. Commun. 1996,
61, 442; b) Janeba Z., Holý A., Masojídková M.: Collect. Czech. Chem. Commun. 2000, 65,
1126; c) Janeba Z., Holý A., Masojídková M.: Collect. Czech. Chem. Commun. 2000, 65,
1698; d) Janeba Z., Holý A., Masojídková M.: Collect. Czech. Chem. Commun. 2001, 66,
517; e) Janeba Z., Holý A., Masojídková M.: Collect. Czech. Chem. Commun. 2001, 66,
1393; f) Janeba Z., Holý A.: Nucleosides Nucleotides Nucleic Acids 2001, 20, 1103.
13. Janeba Z., Holý A., De Clercq E., Balzarini J.: Unpublished results.
14. Mieczkowski A., Roy V., Agrofoglio L. A.: Chem. Rev. 2009, doi:10.1021/cr900329y.
15. Hurd C. D., Ginger L. G., Gershbein L. L., Yarnall W. A.: J. Am. Chem. Soc. 1952, 74,
5128.
16. Arbuzov B. A., Bogonostseva N. P.: Zh. Obshch. Khim. 1959, 29, 2617.
17. Kolyamshin O. A., Kormachev V. V., Mitrasov Y. N.: Zh. Obshch. Khim. 1993, 63, 1075.
18. Williams D. H., Fleming I.: Spectroscopic Methods in Organic Chemistry, 4th ed., Table 3.27.
McGraw–Hill Book Company Ltd., London 1989.
19. Holý A., Günter J., Dvořáková H., Masojídková M., Andrei G., Snoeck R., Balzarini J.,
De Clercq E.: J. Med. Chem. 1999, 42 , 2064.
Collect. Czech. Chem. Commun. 2010, Vol. 75, No. 3, pp. 371–381