Synthesis and biological evaluation of some acyclic 4,6-disubstituted 1H-pyrazolo[3,4-d]pyrimidine nucleosides

Article abstract of DOI:10.1081/NCN-120022697

The chemical synthesis and biological evaluation of some acyclic α-[6-(1′-carbamoylalkylthio)-1H-pyrazolo[3,4-d]pyrimidin-4-yl] thioalkylamide nucleosides are described.

Full text of DOI:10.1081/NCN-120022697

NUCLEOSIDES, NUCLEOTIDES & NUCLEIC ACIDS
Vol. 22, Nos. 5–8, pp. 967–972, 2003
Synthesis and Biological Evaluation of Some Acyclic 4,6-
Disubstituted 1H-Pyrazolo[3,4-d]pyrimidine Nucleosides
1
O. Moukha-chafiq, M. L. Taha, A. Mouma, H. B. Lazrek,
1,*
1
2
3
J. J. Vasseur, and E. De Clercq
4
1
Laboratoire de Chimie Bio-Organique, Facult e´ des Sciences, Agadir, Morocco
2
Laboratoire de Chimie Bio-Organique, Facult e´ des Sciences I,
Marrakech, Morocco
3
Laboratoire de Chimie Organique Biomol e´ culaire de Synth e` se,
U. S. T. Montpellier II, France
4
Rega Institute for Medical Research, Katholieke Universiteit
Leuven, Leuven, Belgium
ABSTRACT
0
The chemical synthesis and biological evaluation of some acyclic a-[6-(1 -car-
bamoylalkylthio)-1H-pyrazolo[3,4-d]pyrimidin-4-yl]thioalkylamide nucleosides are
described.
Key Words: Acyclic nucleosides; Pyrazolo[3,4-d]pyrimidines; Biological
evaluation.
*
Correspondence: M. L. Taha, D e´ partement de Chin, Facult e´ des Sciences, Laboratoire de
Chimie Bio-Organique, BP 28=5, Agadir 80000, Morocco; Fax: +212 48 22 0100; E-mail:
labd1@caramail.com.
9
67
DOI: 10.1081/NCN-120022697
Copyright # 2003 by Marcel Dekker, Inc.
1525-7770 (Print); 1532-2335 (Online)
www.dekker.com

968
Moukha-chafiq et al.
INTRODUCTION
The pyrazolo[3,4-d]pyrimidines are of considerable chemical and pharmacolo-
[
1–3]
gical importance as purine analogues due to their anti-tumor effects
and their
[
4]
strong therapeutic activity against various diseases. However, only few acyclic pyr-
azolo[3,4-d]pyrimidine nucleosides have been reported in the literature. In this
respect, we recently reported the synthesis of some acyclic a-(1H-pyrazolo[3,4-d]pyr-
[
5,6]
imidin-4-yl) thioalkylamide nucleosides
[8]
equipped with the acyclic chains of the
[7]
[9]
acyclovir, HBG and iso-DHPG (Fig. 1). Some of these compounds showed
modest activity against some DNA viruses. Along these lines, we attempted at
synthesizing, through modification at the C position of pyrazolo[3,4-d]pyrimidine
ring, some acyclic a-[6-(1 -carbamoylalkylthio)-1H-pyrazolo[3,4-d]pyrimidin-4-
yl]thioalkylamide nucleosides (Schs. 1, 2 and 3) and determined their biological
activity.
6
0
RESULTS AND DISCUSSION
The 1H-pyrazolo[3,4-d]pyrimidin-4,6-dithione 6, depicted in Sch. 1, was pre-
[
10,11]
pared in five steps according to the literature procedure
and triethyl orthoformate as starting materials. The C and C sulfur atoms of the
from malononitrile
4
6
heterocycle 6 were alkylated with ethyl bromoacetate 7, (DL)-ethyl-2-bromopropio-
nate 8 or (DL) -ethyl-2-bromobutyrate 9 in a sodium hydroxide solution at room
0
temperature to give regioselectively ethyl a-[6-(1 -carboethoxyalkylthio) -1H-pyra-
zolo[3,4-d]pyrimidin-4-yl]thioalkylates 10a–c (Sch. 1) in good yield.
The preparation of the protected acyclic nucleosides (14–16)a–c (Sch. 2) was
achieved using the same conditions as previously described for the synthesis of some
[
12,13]
N -acyclic 4-substituted pyrazolo[3,4-d]pyrimidine nucleosides.
1
Thus, the alky-
[13,15]
[14]
lation of heterocycles 10a–c, separately with alkylating agents 11, 12
[16]
or 13,
using solid-liquid phase transfer catalysis method in which potassium tert-butoxide
Figure 1.

Acyclic Pyrazolo[3,4-d]pyrimidine Nucleosides
969
Scheme 1.
was used as alkali, tetrahydrofuran as solvent and 18-crown-6 as catalyst, afforded
regioselectively the N -regioisomers (14–16)a–c, respectively, in good yield.
1
It was reported that N -nucleoside (N -acyclonucleoside) formation occurred
2
2
[
17–23]
during glycosylation (alkylation) of the pyrazolo[3,4-d]pyrimidines.
case, the presumed N -regioisomers of (14–16)a–c were detected in only trace
. In our
2
amounts but not isolated.
Then, the treatment of N -protected acyclic nucleosides (14–16)a–c with a solu-
1
tion of methanolic ammonia at room temperature gave the deprotected acyclic
nucleosides (17–19) a–c (Sch. 3) in quantitative yield, through removing of the acetyl
and benzoyl groups and concomitant conversion of the esters into the amide
moieties.
The site of alkylation in compounds 10a–c was established to be at N by a
1
direct comparison of the UV spectra of the compounds (17–19)a–c with the UV spec-
[
24]
tra of the corresponding N -pyrazolo[3,4-d]pyrimidine nucleosides.
1
Scheme 2.

970
Moukha-chafiq et al.
Scheme 3.
1
All structures of the synthetic products were identified by H NMR, mass
a
spectra, UV and=or elemental analysis (for example, compound 18a).
BIOLOGICAL STUDIES
The acyclic nucleosides (17–19)a–c were evaluated against cytomegalovirus
þ
[
OKA strains) and TK VZV (07=1 and YS=R strains)] in human embryonic lung
CMV (AD-169 and Davis strains)] and varicella-zoster virus [TK VZV (YS and
ꢀ
(HEL) fibroblasts. No significant antiviral activity or cytotoxicity was noted at con-
centrations up to 50 mg=mL.
All above mentioned acyclic nucleosides were also evaluated for their inhibitory
activity against Mycobacterium tuberculosis [H Rv (ATCC 27294)] in BACTEC 12B
3
7
medium. No activity was noted against M.tuberculosis at concentrations up to
.25 mg=mL.
In conclusion, we have regioselectively synthesized some a-[6-(1 -carbamoylalk-
6
0
ylthio)()-1H-pyrazolo[3,4-d]pyrimidin-4-yl]thioalkylamide nucleosides with the alkyl
chains of acyclovir, HBG and iso-DHPG. No significant anti-CMV; anti-VZV or
anti-tuberculosis activity was witnessed. Further antiviral and anti-tumor evaluation
is in progress.
a
a-[1-(4-hydroxybutyl)-6-carbamoylmethylthio-1H-pyrazolo[3,4-d]pyrimidin-4-yl]thio-
f 3 3
acetamide 18a: Yield: 87%. R : 0.12 (CHCl :CH OH, 90 : 10, v:v). Appearence :
1
liquid. UV (methanol) lmax ¼ 250 nm (e ¼ 19 200). H-NMR (Me
2 6
SO-d , 250 MHz) d
:
1.35 (m, 2H, HOCH CH ), 1.84 (m, 2H, CH CH N), 3.42 (m, 2H, HOCH CH ),
2
2
2
2
2
2
3
4
.92 (s, 2H, 6-SCH ), 4.11 (s, 2H, 4-SCH
.44 (t, J ¼ 5.17 Hz, 1H, HO, D
2
2
), 4.33 (t, J ¼ 6.86 Hz, 2H, CH
2
CH
2
N),
O exchangeable), 7.23, 7.32, 7.66 et 7.74 (4sl, 4H, 2
2
þ
þ
NH , D O exchangeable), 8.28 (s, 1H, H ). MS (FAB , NBA) m=z: 371 [M þ H] . ele-
2
2
3
mental analysis calculated for C13
H
18
N
found : C 42.51%, H 5.00%, N 22.89%.
6
O
3 2
S (370.44): C 42.15%, H 4.89%, N 22.68%,

Acyclic Pyrazolo[3,4-d]pyrimidine Nucleosides
ACKNOWLEDGMENTS
971
This work was supported by ‘‘Recherches men e´ es dans le cadre du Programme
ꢁ
Th e´ matique d’Appui a` la Recherche Scientifique (PROTARS N P1T2=02) ’’ and by
the ‘‘Biomedical Research Programme of the European Commission’’. Cecil D.
Kwong, Ph.D. (Southern Research Institute, Birmingham, AL) is gratefully acknow-
ledged for the evaluation of the anti-tuberculosis activity.
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