Synthesis and evaluation of oligo-1,3-thiazolecarboxamide derivatives as HIV-1 reverse transcriptase inhibitors

Article abstract of DOI:10.1016/S0968-0896(00)00029-8

A set of oligo-1,3-thiazolecarboxamide derivatives able to interact with the minor groove of nucleic acids was synthesized. These oligopeptides contained different numbers of thiazole units presenting dimethylaminopropyl or EDTA moieties on the C-terminus, and aminohexanoyl or EDTA moieties on the N-terminus. The inhibition of such compounds on HIV-1 reverse transcriptase activity was evaluated using different model template-primer duplexes: DNA·DNA, RNA·DNA, DNA·RNA and RNA·RNA. The biological properties of the thiazolecarboxamide derivatives were compared to those of distamycin, another minor groove binder which contains three pyrrole rings. Similar to distamycin, the thiazole containing oligopeptides were good inhibitors of the reverse transcription reaction in the presence of DNA·DNA. But in contrast to distamycin, the oligothiazolide derivatives were able to inhibit reverse transcription in the presence of RNA·DNA or DNA·RNA template-primers. Both distamycin and oligothiazolecarboxamides had low affinity for RNA·RNA duplexes. The inhibition obtained with the newly synthesized thiazolecarboxamides showed that these compounds were more powerful and versatile inhibitors of the RT-dependent polymerization than the natural minor groove binder distamycin. (C) 2000 Elsevier Science Ltd.

Full text of DOI:10.1016/S0968-0896(00)00029-8

Bioorganic & Medicinal Chemistry 8 (2000) 985±993
Synthesis and Evaluation of Oligo-1,3-thiazolecarboxamide
Derivatives as HIV-1 Reverse Transcriptase Inhibitors
Vladimir A. Ryabinin, a Olga D. Zakharova, b Ekaterina Y. Yurchenko, b
Olga A. Timofeeva, b Igor V. Martyanov, b Andrei A. Tokarev, a Eugeny F. Belanov, a
Nikolai I. Bormotov, a Laura Tarrago-Litvak, c Marie Line Andreola, c Simon Litvak, c,*
Georgy A. Nevinsky b and Alexander N. Sinyakov a
aInstitute of Molecular Biology, Koltsovo, Novosibirsk Region, 633159, Russia
bNovosibirsk Institute of Bioorganic Chemistry, Siberian Division of Russian Academy of Sciences, Novosibirsk 630090, Russia
c
 Â
UMR 5097 CNRS-Universite Victor Segalen Bordeaux 2, IFR 66 ``Pathologies Infectieuses''. 146, rue Leo Saignat,
33076 Bordeaux cedex, France
Received 20 September 1999; accepted 8 December 1999
AbstractÐA set of oligo-1,3-thiazolecarboxamide derivatives able to interact with the minor groove of nucleic acids was synthe-
sized. These oligopeptides contained di€erent numbers of thiazole units presenting dimethylaminopropyl or EDTA moieties on the
C-terminus, and aminohexanoyl or EDTA moieties on the N-terminus. The inhibition of such compounds on HIV-1 reverse tran-
.
.
.
scriptase activity was evaluated using di€erent model template±primer duplexes: DNA DNA, RNA DNA, DNA RNA and
.
RNA RNA. The biological properties of the thiazolecarboxamide derivatives were compared to those of distamycin, another minor
groove binder which contains three pyrrole rings. Similar to distamycin, the thiazole containing oligopeptides were good inhibitors
.
of the reverse transcription reaction in the presence of DNA DNA. But in contrast to distamycin, the oligothiazolide derivatives
.
were able to inhibit reverse transcription in the presence of RNA DNA or DNA RNA template±primers. Both distamycin and
.
.
oligothiazolecarboxamides had low anity for RNA RNA duplexes. The inhibition obtained with the newly synthesized thiazole-
carboxamides showed that these compounds were more powerful and versatile inhibitors of the RT-dependent polymerization than
the natural minor groove binder distamycin. # 2000 Elsevier Science Ltd. All rights reserved.
Introduction
The prototype minor groove binders are netropsin (1)
and distamycin (2) (Fig. 1). Distamycin was ®rst iso-
lated from Streptomyces distallicus4 and later
synthesized.5 7 It inhibits both Gram-positive and
Gram-negative bacteria and replication of viral DNA8
(vaccinia, herpes simplex virus). The synthesis of certain
analogues of distamycin, containing either additional
pyrrole units or derivatives in which the pyrrole rings
were replaced by benzene, pyridine, thiophene, thiazole,
imidazole, pyrazole, or triazole has been reported.8 18
These compounds have been used as antitumor or anti-
viral drugs8 or as tools to study the modulation of
enzymes.19 23 Several distamycin analogues possessing
antitumor activity are undergoing clinical trials.24,25
Most of the therapeutic agents used against HIV-1
induce the emergence of resistance strains.1,2 That is
why one of the most important lines in the pharmacol-
ogy concerning HIV-1 is focused on the development of
new, or on the modi®cation of already existing drugs.
Among the new and promising antivirals tools are
compounds which interact reversibly with nucleic
acids.3 Minor groove binding molecules are capable of
binding to the minor groove of a double-stranded
DNA. Such molecules contain several small aromatic
rings such as pyrrole, furan or benzene connected by
bonds with torsional freedom. The DNA minor groove
.
may ``®t'' aromatic molecules better at A T-rich relative
.
to G C-rich regions.
The development of methods to synthesize minor
groove molecules capable of binding with virtually any
DNA sequence increased the interest for such com-
pounds.23,26 For example, lexitropsins have been
Abbreviations: MGB, minor groove binders; HIV-1, human immuno-
de®ciency virus type 1; RT, reverse transcriptase.
*Corresponding author. Tel.: +33-557-57-1760; fax: +33-557-57-
1766; e-mail: simon.litvak@reger.u- bordeaux2.fr
.
.
designed to recognize A T and also G C base-pairs.
Sulfonated and phosphonated distamycin derivatives
0968-0896/00/$ - see front matter # 2000 Elsevier Science Ltd. All rights reserved.
PII: S0968-0896(00)00029-8

986
V. A. Ryabinin et al. / Bioorg. Med. Chem. 8 (2000) 985±993
.
the enzyme is ®rst complexed with an RNA RNA tem-
.
plate±primer; then, with an RNA DNA, and ®nally, at
.
the second step of replication, with a DNA DNA duplex.
Here we describe the synthesis of a new type of minor
groove binder. They are thiazole-containing oligo-
peptides of various lengths (up to 4 thiazole rings),
containing di€erent groups on their C- and N-termini.
The e€ect of these oligocarboxamides on the activity of
HIV-1 reverse transcriptase was determined.
Results and Discussion
Synthesis of oligo-1,3-thiazolecarboxamide
The basic scheme of the synthesis of oligo-1,3-thiazole-
carboxamide derivatives is shown in Figure 2. The ethyl-
2-amino-1,3-thiazole-4-carboxylate (4) was selected as
the starting material to obtain the target products. The
amino group-protected 1-hydroxy-1,2,3-benzotriazole
ester of 2-amino-1,3-thiazole-4-carboxylate was used
as a synthon. Attempts to obtain the (tert-butyloxy)-
carbonyl (Boc)-derivative were unsuccessful, possibly
due to the extremely low nucleophilicity of the hetero-
cyclic amino group. Hence, we used trityl group for the
protection of the amino group. The 1-hydroxy-1,2,3-
benzotriazole ester 6 was readily synthesized from equi-
molar amounts of acid 5 and 1-hydroxy-1,2,3-benzo-
triazole by employing dicyclohexylcarbodiimide (DCC)
as a coupling reagent. Compound 6 is suciently stable
and can be stored for several months at 0±4 ꢀC without
any alteration.
Figure 1. Structural formulas of netropsin (1), distamycin (2) and
thiazolecarboxamides (3).
capable of suppressing HIV-1 replication are of poten-
tial therapeutic importance.27 Of obvious interest are
the analogues of distamycin and netropsin in which the
N-methylpyrrole fragment is substituted with a thiazole
cycle, since these compounds are likely to possess dif-
ferent site-speci®city compared to oligopyrrolecarbox-
amides. Due to the low reactivity of the key oligomer
2-amino-4-carbonic acid-1,3-thiazole, studies concern-
ing the minor groove binders which contain the thiazole
fragment in their structure are not numerous.28 31
The cycle of oligocarboxamide chain lengthening com-
prised detritylation under the e€ect of tri¯uoroacetic
acid (TFA) in chloroform with consequent combination
of the amine formed hydroxy-1,2,3-benzotriazole ester 6
in CH2Cl2±DMF, since the amine solubility decreased
with the growth of the chain. The compounds synthe-
sized were puri®ed by chromatography on silica gel.
First, the residue of N-tritylaminohexanoic acid was
attached to the N-termini of the obtained di-, tri-, and
tetrathiazolecarboxamide derivatives 12 followed by
condensation of the C-terminal carboxyl group with
N,N-dimethyl-2-aminopropane, methylamine, or N-tri-
phenylmethyl-1,6-diaminohexane. At the last stage, de-
tritylation of the synthesized compounds by TFA was
carried out. EDTA-derivatives of di- and trithiazole-
carboxamides were obtained. As a result, compounds
with di€erent numbers of thiazolecarboxamide units in
the peptide and the substitutes in the side chain were
obtained. The number of thiazole units in the synthe-
sized oligopeptides was estimated by NMR spectro-
scopy by comparing the integrals of proton signals of
thiazole and those of the additional groups attached to
the N- or C-terminus of the compounds. The elution
time of the compounds on reverse-phase chromato-
graphy gradually increased with the lengthening of the
oligopeptides. The structures of the thiazolecarbox-
amide analogues are given in Figure 2. We did not try to
prepare longer oligocarboxamides (penta- and hexa-
thiazolecarboxamides) because of their low solubility in
Heteroanalogues of distamycin and netropsin in which
one of the methylpyrrole units is substituted by thiazole
have been synthesized. Studies related to the site-speci-
®city of a thiazole-containing oligopeptide analogue of
netropsin (compound 3 in Fig. 1) demonstrated that this
compound bound to 4±5 base pairs of the minor
groove.13,14 Compound 3 has higher binding anity for
13
.
sites with high content of G C pairs; in addition, it
prefers an alternate pyrimidine sequence instead of a
homopolymeric sequence. There are also reports show-
ing the in¯uence of the methyl substituents of a thia-
zole-containing lexitropsin on the mode of binding to
DNA.29
HIV-1 reverse transcriptase (RT) is an enzyme playing a
crucial role in the replication of the retrovirus.32 RT
catalyzes three reactions, all essential, to transform the
single-stranded retroviral RNA genome into a double-
stranded proviral DNA. The enzyme is multifunctional
and exhibits three enzymatic activities: RNA-dependent
DNA polymerase, DNA-dependent DNA polymerase
and ribonuclease H (RNase H). HIV-1 RT starts minus-
strand DNA synthesis from tRNALys3 as primer and the
plus-strand RNA genome as template. After digestion
.
of the RNA template from the duplex RNA DNA, the
synthesis of the plus-strand DNA proceeds using the
minus-strand DNA as template. Thus, during replication,

V. A. Ryabinin et al. / Bioorg. Med. Chem. 8 (2000) 985±993
987
Figure 2. Basic scheme of the synthesis of oligo-1,3-thiazolecarboxamide derivatives.
7,26,31,33
.
DNA DNA duplexes,
organic solvents and water. According to preliminary
DNA footprinting data, oligothiazolecarboxamides
there are no reports
concerning the interaction of such compounds with
.
interacted with DNA sites containing both A T and
.
.
RNA RNA or RNA DNA duplexes. As HIV-1 RT
interacts with various template±primers, such as
.
G C pairs (to be published elsewhere).
.
.
.
RNA RNA, RNA DNA and DNA DNA during viral
replication, it seemed interesting to study the interaction
of the thiazolecarboxamides with all these nucleic acids.
This was done by determining the eciency of the
interaction between the thiazolecarboxamides and HIV-1
Interaction of derivatives with HIV-1 RT
While distamycin and its analogues are known to inter-
act predominantly with the minor groove of the B-form

988
V. A. Ryabinin et al. / Bioorg. Med. Chem. 8 (2000) 985±993
shown by the results with activated DNA or poly-
.
(dA) d(pT)16.
.
In the presence of RNA DNA [poly(A) d(pT)16] or
.
.
.
DNA RNA [poly(dA) (pU)16] template±primers, the
inhibition of reverse transcription was less important
.
than in the case of DNA DNA, although compounds
18c and 18d showed good inhibitory properties (Table 1).
The anity of the thiazole-containing compounds for
nucleic acids depended essentially on the structure of
the additional groups at the N- and C-termini of oligo-
carboxamides, and on the number of thiazole cycles.
For tri- and tetrameric oligomers (compounds 18c
and 18d), the inhibition was enhanced by one order of
magnitude.
.
In the case of a template±primer RNA RNA such as
.
poly(A) (pU)16, the inhibitory e€ect of the thiazole-
containing derivatives in the RT-catalyzed reaction was
extremely low (IC50=1.5±7.7 mM). Apparently, this
inhibition was independent of either the number of
amino acid units in the oligopeptide or the structure of
the additional groups within these compounds.
Our results showed that the e€ects of the thiazole deri-
vatives were similar to those of distamycin only in the
case of DNA DNA and RNA RNA template±primer
duplexes. A di€erent behavior was found, however,
between the thiazole derivatives and distamycin in the
Figure 3. Reverse transcription in the presence of thiazolecarbox-
amides or distamycin. Relative rates of RT-catalyzed polymerization
(V) versus concentration of distamycin (*) or tetrathiazole carbox-
amide NH2(Tz)4N(CH3)2 (compound 18d) (&), in the presence of
poly(A)-d(pT)16 template±primer complex. Arrows indicate the IC50
values.
.
.
.
.
case of RNA DNA or DNA RNA: some thiazole deri-
vatives inhibited the polymerization reaction on poly-
.
.
RT in the presence of di€erent duplexes used as model
(dA) d(pT)16 and poly(dA) (pU)16 at concentrations
that were 1±2 orders of magnitude lower than that of
distamycin.
.
of nucleic acids: poly(A) oligo(U), poly(A) oligo(dT),
.
.
.
poly(dA) oligo(U), poly(dA) oligo(dT) and activated
DNA.
We compared logically our values to results described
for a compound of the same nature. Anyway, the IC50
values for thiazole compounds are higher than what is
found for a standard inhibitor of the HIV-1 RT like
AZTTP.
The eciency of the interaction was determined by
estimating the concentration of the thiazolecarboxamide
giving 50% inhibition (IC50) of the RT-dependent
polymerization. Experiments were done in parallel in
the presence of distamycin used as reference of a small
groove binding molecule. A typical experiment on the
determination of the IC50 value for the thiazole-con-
taining oligopeptides is shown in Figure 3. The IC50
values for all the compounds analyzed are given in
Table 1. The thiazole-containing oligopeptides were
better inhibitors of reverse transcription than distamycin
The features governing the interaction of DNA with
di€erent compounds in solution cannot always be
extrapolated to the interaction of DNA with an enzyme,
as the latter causes substantial structural changes in the
nucleic acid regions situated within the protein globule
(for a review see ref 34). For example oligonucleotides
containing the phenazinium group at the 50-end
.
in the presence of a template±primer DNA DNA, as
Table 1. E€ect of thiazolecarboxamide derivatives on HIV-1 RTa
Compound
Nꢀ thiazole
units
IC50 (mM)
.
Poly(dA) d(pT)16
.
Poly(A) d(pT)16
.
Poly(dA) (pU)16
.
Poly(A) (pU)16
Activated DNA
18b
18c
18d
19b
19c
20
21
Distamycin
2
3
4
2
3
2
4
Ð
10
10
10
50
10
50
50
50
970
700
10
20
10
350
650
860
250
40
20
350
180
250
9000
5000
460
20
40
500
870
360
120
7500
7700
5500
6800
6000
2700
2800
5000
4800
aReverse transcription was performed as described in the Experimental using di€erent template±primers as indicated. IC50 corresponds to the con-
centration of thiazolcarboxamide derivative inhibiting 50% of the HIV-1 RT activity. IC50 values for the distamycin are indicated.

V. A. Ryabinin et al. / Bioorg. Med. Chem. 8 (2000) 985±993
989
demonstrated a similar anity for complementary
Conclusion
DNA and RNA in solution, while the anity of the
ethidium derivative of d(pT)8 was about 2 orders of
magnitude higher for poly(A) than that for poly(dA)
templates in their complexes with HIV RT.35 This seems
to result as a consequence of a change in the structure of
the nucleic acid after binding to the enzyme.
We synthesized di€erent oligothiazolecarboxamide
derivatives containing up to four thiazole cycles. These
compounds were inhibitors of HIV-1 RT when
assayed in the presence of di€erent template±primers:
DNA DNA, DNA RNA and RNA DNA. Under the
same conditions, distamycin inhibited the enzyme only
.
with DNA DNA duplexes. The results presented here
implicate that interactions of HIV-1 RT with the sec-
ondary structures of the primer±template may play a
signi®cant role in its inhibition by thiazole derivatives.
.
.
.
The formation of a complex between ds DNA and
enzymes involved in replication substantially diminishes
intrastrand complementary interactions, apparently as a
result of an increase of the distance between the com-
plementary strands and alteration of their structural
characteristics as compared with those in solution. Such
a change of the complex is promoted by binding of the
template and primer on the enzyme through e€ective
interactions with the protein. It is interesting to recall
that there is a correlation between the interaction of
distamycin with ds DNA in solution, and in the com-
plex with HIV-1 RT.
Experimental
Chemistry
TLC analysis was carried out on precoated plates of
silica gel 60F254 (Merck). Puri®cation was perfomed on
1
silica gel (40±60 mm). H NMR spectra were recorded
Why can thiazole-containing oligopeptides interact bet-
with a Bruker AM-200 apparatus. Reverse phase HPLC
was performed with Altex dual-pump system and
2.5Â25 cm with Diasorb 130 C16T (BioChemMack).
All the chemicals used were obtained from Fluka.
EDTA-bisanhydride was prepared according to ref 43.
.
ter with RNA DNA complexes than distamycin? And
why do all compounds analyzed have a low anity for
.
the RNA RNA duplex?
Di€erent reports have focused on the signi®cance of
HIV-1 RT primer±template interactions. It is known
Ethyl 2-amino-1,3-thiazole-4-carboxylate (4). This was
prepared as described in ref 44. After recrystallization
from EtOH the yield of pure compound was 62%. H
NMR (CDCl3) d 7.42 (s, 1H), 7.02 (b s, 2H), 4.26 (q,
2H, J=7.0 Hz), 1.29 (t, 3H, J=7.0 Hz).
.
.
that DNA DNA and DNA RNA duplexes in solution
can be either in B or in A form, whereas A-form (11
1
36,37
.
residues per turn) is characteristic of RNA RNA.
Based on kinetic and thermodynamic data it can be
assumed that a duplex containing a template in A-like
form and a primer in B-like form is optimal for RT or
DNA polymerases such as mammalian DNA poly-
merase b.38,39 X-ray analysis data of HIV-1 RT and
polymerase b complexes with nucleic acids strongly
support this idea.40 42 A template±primer in complex
with HIV-1 RT binds to the enzyme in a catalytically
relevant fashion. The template±primer conforms more
closely in structure to a classical A-form DNA near the
polymerase active site and to B-form DNA near the
RNase H active site. RT must be able to induce a sig-
2-Triphenylmethylamino-4-carboxy-1,3-thiazol (5). A
mixture of compound 4 (7.4 g, 43 mmol) and tri-
phenylmethyl chloride (12.0 g, 43 mmol) was dissolved
in 60 mL of dry pyridine and stirred for 16 h at room
temperature. The precipitate was ®ltered o€, washed
with pyridine (2Â10 mL), and the ®ltrate was evapo-
rated under reduced pressure up to 40±60 mL. Ten
grams of NaOH in 100 mL of 60% EtOH was added to
the residue. The mixture was stirred until precipitate
formation and left at room temperature for 20 h. The
mixture was cautiously acidi®ed with acetic acid to pH
5±6. The precipitate was collected by ®ltration, washed
with water (10Â50 mL) and acetone (2Â25 mL). The
yield of product 5 was 12.6 g (72%).1H NMR (CDCl3) d
9.46 (bs, 1H), 7.27±7.40 (m, 15H), 7.27 (s, 1H).
.
ni®cant bend in the DNA DNA duplex such that the
helical axes of the A-form and B-form portions make an
angle of about 40±45ꢀ. For DNA polymerase b a similar
situation was found: the DNA duplex presented an
A-form in the vicinity of the active site, while double
stranded DNA outside the active site presented a
B-form. Thus, the adaptation of DNA or RNA to the
optimal conformation for the enzyme may be a very ®ne
process.
1,2,3-Benzotriazol-1-yl-2-triphenylmethylamino-1,3-thia-
zol-4-carboxylate (6). Compound 5 (7.20 g, 18.6 mmol)
was rapidly stirred with 1-hydroxy-1,2,3-benzotriazole
(2.52 g, 18.6 mmol) in dry dichloromethane (200 mL),
and DCC (3.90 g, 19.0 mmol) was added to the mixture
in small portions at room temperature. After stirring the
mixture for 4 h, the precipitated urea was ®ltered o€
and solution was poured into hexane (200 mL). The
precipitate was collected and dried in vacuo over SiO2.
The resulting crude product 6 (9.0 g, 96%) was su-
Assuming that thiazole oligopeptides have an increased
.
anity for the minor groove of the RNA DNA
duplexes when complexed to RT, the low anity of
.
these compounds for RNA RNA can be explained by
the inability of the latter to e€ectively adapt to the
RT-dependent speci®c conformation. Thus, it seems
possible to use the thiazole-containing oligopeptides for
detection of speci®c nucleic acid conformation (A- and
B-forms) after binding with enzymes.
1
ciently pure and could be used directly for synthesis. H
NMR (CDCl3) d 7.65 (s, 1H), 7.28±7.65 (m, 4H), 7.32
(s, 15H), 7.08 (s, 1H).

990
V. A. Ryabinin et al. / Bioorg. Med. Chem. 8 (2000) 985±993
1
Ethyl-2-[2-(triphenylmethylamino)-1,3-thiazol-4-carbox-
amido]-1,3-thiazole-4-carboxylate (7a). A mixture of
compound 4 (1.83 g, 10.6 mmol), compound 6 (5.35 g,
10.6 mmol) and NEt3 (2.6 mL, 18 mmol) in dry DMF
(25 mL) was stirred for 1 h at 50 ꢀC. The greenish mixture
was allowed to react at 50 ꢀC without stirring for 16 h.
The mixture was evaporated under reduced pressure.
The residue was puri®ed by column chromatography;
the resulting product 7a was eluted by chloroform:
18.6 g (95%). H NMR ((CD3)2CO) d 7.05±7.43 (m,
15H), 4.02 (q, 2H, J=7.0 Hz), 2.19 (t, 2H, J=7.0 Hz),
2.08 (t, 2H, J=7.0 Hz), 1.28±1.60 (m, 6H), 1.20 (t, 3H,
J=7.0 Hz).
6-Triphenylmethylaminohexanoic acid (9). Triphenyl-
methyl chloride (26.8 g, 97 mmol) was added to a solution
of compound (8) (18.5 g, 95 mmol) and NEt3 (30 mL) in
dry CH2Cl2 at 0 ꢀC. The mixture was stirred for 3 h at
room temperature. The precipitate was ®ltered o€, the
®ltrate was washed with water (2Â50 mL) and then
evaporated. The residue was dissolved in ethanol (150
mL), and a mixture of NaOH (27 g) and H2O (30 mL)
was added dropwise at room temperature. The mixture
was stirred for 4 h at room temperature, evaporated to
100 mL, water (200 mL) was added to the residue and
the mixture was cautiously acidi®ed with acetic acid to
pH 5±6. The product was extracted with CHCl3 (2Â100
mL), the organic layer was washed with water (3Â200
mL) and dried over Na2SO4. After evaporation, practi-
cally pure 9 was obtained (24.8 g, 65%). 1H NMR
((CD3)2CO) d 7.12±7.52 (m, 15H), 2.26 (t, 2H, J=7.0
Hz), 2.10 (t, 2H, J=7.0 Hz), 1.28±1.67 (m, 6H).
1
hexane (1:1) (5.0 g, 88%). H NMR (CDCl3), d 10.40
(bs, 2H), 7.81 (s, 1H), 7.31 (s, 15H), 7.26 (s, 1H), 6.60 (s,
1H), 4.38 (q, 2H, J=7.0 Hz), 1.37 (t, 3H, J=7.0 Hz).
Ethyl-2-[2-[2-(triphenylmethylamino)-1,3-thiazol-4-car-
boxamido]-1,3-thiazol-4-carboxamido]-1,3-thiazole-4-car-
boxylate (7b). Compound 7a (2.6 g, 4.8 mmol) dissolved
in CHCl3 (30 mL) was treated with TFA (15 mL) for 1 h
at room temperature. The mixture was evaporated
under reduced pressure. Twenty milliliters of water were
added to the residue. The precipitate was ®ltered,
washed with 5% NaHCO3 (20 mL), rinsed with water
(4Â20 mL), dried on a ®lter, washed with ether and
dried in vacuo over SiO2. The resulting amine (1.03 g),
compound 6 (1.82 g, 3.5 mmol) and NEt3 (0.7 mL, 5.0
mmol) in dry DMF (5 mL) were stirred for 1 h at 50 ꢀC.
The greenish mixture was allowed to react at 50 ꢀC
without stirring for 16 h. The mixture was evaporated
under reduced pressure. The residue was puri®ed by
column chromatography using chloroform as eluent
giving the product 7b (1.76 g, 55%). 1H NMR
((CD3)2CO) d 10.90 (bs, 1H), 9.80 (bs, 1H), 8.14 (s, 1H),
8.21 (s, 1H), 8.04 (s, 1H), 7.25±7.56 (m, 16H), 4.32 (q,
2H, J=7.0 Hz), 1.34 (t, 3H, J=7.0 Hz).
1,2,3-Benzotriazol-1-yl-6 triphenylmethylaminohexyl-
carboxylate (10). DCC (3.70 g, 17.9 mmol) in small
portions was added to compound 9 (6.36 g, 17.0 mmol)
and 1-hydroxy-1,2,3-benzotriazole (2.42 g, 17.9 mmol)
in dry dichloromethane (100 mL) at room temperature.
After the mixture was stirred for 3 h, the precipitated
urea was ®ltered o€. Filtrate was evaporated and dried
in vacuo over SiO2. The resulting crude product 10 (8.27
g, 99%) was suciently pure and could be used directly
for synthesis. 1H NMR ((CD3)2CO) d 7.12±7.91 (m,
19H), 2.25 (t, 2H, J=7.0 Hz), 2.16 (t, 2H, J=7.0 Hz),
1.28±1.67 (m, 6H).
Ethyl-2-[2-[2-[2-(triphenylmethylamino)-1,3-thiazol-4-car-
boxamido]-1,3-thiazol-4-carboxamido]-1,3-thiazol-4-car-
boxamido]-1,3-thiazole-4-carboxylate (7c). Compound
7b (0.88 g, 1.3 mmol) in CHCl3 (15 mL) was treated
with TFA (7 mL) for 1 h at room temperature. The
mixture was evaporated under reduced pressure. Ten
milliters of water were added to the residue. The pre-
cipitate was ®ltered, washed with 5% NaHCO3 (5 mL),
rinsed with plenty of water, dried on a ®lter, washed
with ether (4Â5 mL) and dried in vacuo over SiO2. The
resulting amine (0.51 g), compound 6 (1.55 g, 3.1 mmol)
and NEt3 (0.6 mL, 4.2 mmol) in dry DMF (6 mL) were
stirred for 1 h at 50 ꢀC. The greenish mixture was
allowed to react without stirring at 50 ꢀC for 16 h. The
mixture was evaporated under reduced pressure. The
residue was puri®ed by column chromatography using
chloroform:DMF (9:1) as eluent giving the product 7c
(0.49 g, 48%). 1H NMR (CDCl3:(CD3)2SO=1:1) d
7.84 (s, 1H), 7.81 (s, 1H), 7.68 (s, 1H), 7.62 (s, 1H),
7.05±7.25 (m, 16H), 4.17 (q, 2H, J=7.0 Hz)), 1.20 (t, 3H,
J=7.0 Hz).
6-Triphenylmethylaminohexylcarboxy derivatives (11a±d).
Compound 7a±c (1.0 mmol) was dissolved in the mix-
ture of CHCl3 (15 mL) and TFA (7 mL). After 1 h at
room temperature the mixture was evaporated under
reduced pressure, and water (10 mL) was added to the
residue. The precipitate was ®ltered, washed with 5%
NaHCO3 (5 mL), rinsed with plenty of water, dried on
®lter, washed with ether (4Â10 mL) and dried in vacuo
over SiO2. The resulting amine or 1.0 mmol compound
4, compound 10 (1.02 g, 2.1 mmol) and NEt3 (0.3 mL)
in dry DMF (6 mL) were stirred for 10 h at 60 ꢀC. The
brownish mixture was evaporated under reduced pres-
sure. The residue was puri®ed by column chromato-
graphy using chloroform as eluent. The yield of pure
compounds 11a, 11b, 11c, 11d was 85, 76, 58 and 63%,
respectively.
1H NMR of compound 11a ((CD3)2CO) d 11.07 (bs,
1H), 8.02 (s, 1H), 7.1±7.5 (m, 15H), 4.30 (q, 2H, J=7.0
Hz), 2.91 (bs, 2H), 2.57 (t, 2H, J=7.0 Hz), 1.46±1.80
(m, 6H), 1.32 (t, 3H, J=7.0 Hz).
Ethyl ester-6-aminohexanoic acid hydrochloride (8).
Thionyl chloride was added dropwise to the solution of
6-aminohexanoic acid (13.1 g, 100 mmol) in dry ethanol
(120 mL) at temperature 0±5 ꢀC. The mixture was
stirred for 1 h at room temperature, ®ltered o€ and the
®ltrate was evaporated. The crude product was recrys-
tallized from EtOH. The yield of pure compound 8 was
1H NMR of compound 11b ((CD3)2CO) d 11.29 (bs, 1H),
10.77 (bs, 1H), 8.09 (s, 1H), 8.01 (s, 1H), 7.1±7.5 (m,
16H), 4.29 (q, 2H, J=7.0 Hz), 2.93 (bs, 2H), 2.58 (t, 2H,
J=7.0 Hz), 1.49±1.79 (m, 6H), 1.33 (t, 3H, J=7.0 Hz).

V. A. Ryabinin et al. / Bioorg. Med. Chem. 8 (2000) 985±993
991
1H NMR of compound 11c ((CD3)2CO) d 11.44 (bs,
1H), 10.85 (bs, 2H), 8.19 (s, 1H), 8.10 (s, 1H), 8.03 (s,
1H), 7.1±7.5 (m, 16H), 4.30 (q, 2H, J=7.0 Hz), 2.90 (bs,
2H), 2.60 (t, 2H, J=7.0 Hz), 1.30±1.84 (m, 6H), 1.33 (t,
3H, J=7.0 Hz).
1H), 7.15±7.50 (m, 15H), 3.25 (s, 3H), 2.75 (t, 2H), 2.33
(t, 2H), 1.90±2.10 (m, 2H), 1.70±1.80 (m, 4H).
1H NMR of compound 14c (DMF-d7) d 8.38 (s, 1H),
8.03 (bs, 1H), 7.83 (s, 1H), 7.15±7.55 (m, 15H), 2.61 (t,
2H), 2.11 (s, 3H), 2.09 (t, 2H), 1.3±1.8 (m, 6H).
1H NMR of compound 11d (DMF-d7) d 11.30 (bs, 1H),
10.60 (bs, 1H), 8.44 (s, 1H), 8.39 (s, 1H), 8.30 (s, 1H),
8.20 (s, 1H), 7.1±7.5 (m, 16H),4.35 (q, 2H, J=7.0 Hz),
2.90 (bs, 2H), 2.60 (t, 2H, J=7.0 Hz), 1.20±1.90 (m,
6H), 1.36 (t, 3H, J=7.0 Hz)
1-Triphenylmethyl-1,6-diaminohexane (15). Triphenyl-
methyl chloride (6.6 g, 24 mmol) in 50 mL dry CH2Cl2
was added to a solution of 1,6-diaminohexane (16.6 mg,
140 mmol) in dry CH2Cl2 (100 mL). The mixture was
stirred for 4 h at 20 ꢀC. Product was puri®ed by column
chromatography using chloroform:EtOH:NEt3 (94:5:1)
as eluent. The yield of compound 15 was 2.73 g (7.7
mmol, 32%).
Preparation of acids 12a±d. 4 M NaOH (10 mL) was
added to the solution of ethyl ester (11a±d) (3 mmol) in
ethanol (20 mL). The mixture was stirred for 12 h at
20 ꢀC, diluted with water (30 mL) and cautiously acid-
i®ed with HCl to pH 5±6. The precipitated acid was
collected, washed with water (6Â10 mL) and dried in
vacuo over SiO2. The resulting acids (90±95%) were
suciently pure and could be used directly for synthesis.
1H NMR ((CD3)2CO) d 7.10±7.60 (m, 15H), 3.09 (t,
7.84, 2H, J=7.2 Hz), 2.10 (t, 7.84, 2H, J=7.2 Hz), 1.52
(bm, 4H), 1.32 (bm, 4H).
Preparation of compound 16. To the mixture of acid 12c
(145 mg, 0.23 mmol) and 1-hydroxy-1,2,3-benzotriazole
(40 mg, 0.26 mmol) in dry CH2Cl2 (5 mL) was added
DCC (54 g, 0.26 mmol) at room temperature. After
stirring for 4 h, the amine 15 (93 mg, 0.26 mmol) and
70 ml NEt3 were added and the reaction mixture was
boiled for 4 h. The yield of pure compound 16 after
column chromatography (0±5% gradient of ethanol in
chloroform) was 187 mg (0.19 mmol, 84%).
N,N-Dimethyl-1,3-propylendiamine derivatives (13a±d)
and methylamine derivatives. DCC (0.4 g, 2.0 mmol) in
small portions was added to the mixture of acid 12a±c
(1.0 mmol) and 1-hydroxy-1,2,3-benzotriazole (0.3 g,
2.0 mmol) in dry DMF (3 mL) at room temperature.
After stirring for 2 h, N,N-dimethyl-1,3-diaminopro-
pane (0.2 mL) or methylamine hydrochloride (0.4 g, 6
mmol) and NEt3 (5 mL) was added and reaction mix-
ture was left for 12 h. The precipitate was ®ltered o€,
washed with CH2Cl2 (5 mL). Combined ®ltrates diluted
with CHCl3 (50 mL) were washed with 5% NaHCO3
(2Â15 mL), water (3Â25 mL), dried over Na2SO4 and
evaporated. The yield of products 13a, 13b, 13c, 13d and
14a, 14b, 14c after column chromatography (0±20%
gradient of EtOH in CHCl3:NEt3 (95:5) was 60, 49, 55,
35, 65, 52 and 40%, respectively.
1H NMR ((CD3)2CO) d 11.46 (bs, 1H), 10.50 (bs, 1H),
7.84 (s, 1H), 7.10±7.50 (m, 30H), 7.98 (s, 1H), 3.40 (q,
2H, J=6.5 Hz), 2.84 (t, 2H, J=7.0 Hz), 2.64 (t, 2H,
J=7.0 Hz), 2.14 (t, 2H, J=7.0 Hz), 0.94±1.90 (m, 14H).
Preparation of compound 17. DCC (93 g, 0.45 mmol)
was added to the mixture of acid 12d (260 mg, 0.30
mmol) and 1-hydroxy-1,2,3-benzotriazole (69 mg, 0.45
mmol) in dry DMF (5 mL) at room temperature. After
stirring for 4 h, NH2CH2CH(OH)CH2OH (120 mg, 1.30
mmol) was added and the reaction mixture was left for
4 h at 60oC. The yield of pure compound 17 after col-
umn chromatography (0±15% gradient of EtOH in
CHCl3:NEt3 (95:5)) was 110 mg (0.12 mmol, 40%).
1H NMR of compound 13a ((CD3)2CO) d 8.10 (bs, 1H),
7.87 (s, 1H), 7.15±7.55 (m, 15H), 3.54 (bt, 2H), 2.64 (bs,
2H), 2.49 (bt, 2H), 2.17 (s, 6H), 1.10±2.10 (m, 10H).
1H NMR of compound 13b ((CD3)2CO) d 8.17 (bs, 1H),
7.96 (s, 1H), 7.86 (s, 1H), 7.15±7.55 (m, 15H), 3.54 (bt,
2H), 2.64 (bs, 2H), 2.49 (bt, 2H), 2.17 (s, 6H), 1.10±2.10
(m, 10H).
1H NMR ((CD3)2CO) d 11.48 (bs, 1H), 10.75 (bs,
2H), 10.30 (bs, 1H), 8.18 (s, 1H), 8.09 (bs, 1H), 8.02
(s, 1H), 8.01 (s, 1H), 7.82 (s, 1H), 7.10±7.50 (m, 15H),
4.51 (bs, 1H), 3.88 (bs, 1H), 3.60 (m, 4H), 2.60 (t, 2H,
J=7.0 Hz), 2.15 (t, 2H, J=7.0 Hz), 1.70 (m, 4H), 1.45
(m, 2H).
1H NMR of compound 13c ((CD3)2CO) d 8.00 (s, 1H),
7.90 (s, 1H), 7.85 (s, 1H), 7.15±7.55 (m, 15H), 3.58
(b.quin, 2H), 2.60 (bm, 2H), 2.33 (bm, 2H), 2.07 (s, 6H),
1.10±2.10 (m, 10H).
1H NMR of compound 13d (DMF-d7) d 8.40 (s, 1H),
8.31 (s, 1H), 8.28 (s, 1H), 8.10 (s, 1H), 7.1±7.5 (m, 15H),
3.83 (bs, 2H), 2.55 (bs, 2H), 2.30 (bs, 2H), 2.13 (s,6H),
1.1±2.0 (m, 10H).
Preparation of compounds 18±21. Compounds 13, 14, 16
or 17 (100 mg) were dissolved in the mixture of CHCl3
(2 mL) and TFA (1 mL) and, after 1 h at room tem-
perature, the mixture was evaporated under reduced
pressure and washed with ether (4Â5 mL), dissolved in
0.1% TFA and puri®ed by HPLC (0±15% gradient
CH3CN, 0.1% TFA 50±80%). The yield of products
18a, 18b, 18c, 18d, 19a, 19b, 19c, 20, 21 after column
chromatography (0±20% gradient of EtOH in CHCl3:
NEt3 (95:5) was 80, 75, 71, 50, 72, 60, 55, 51 and 45%,
respectively.
1H NMR of compound 14a (CCl4) d 10.78 (bs, 1H), 7.82
(bs, 1H), 7.74 (s, 1H), 7.20±7.50 (m, 15H), 3.27 (s, 3H),
2.80 (bt, 2H), 2.35 (bt, 2H), 2.01 (bm, 2H), 1.77 (bm, 4H).
1H NMR of compound 14b ((CD3)2CO) d 11.49 (bs,
1H), 10.62 (bs, 1H), 8.35 (bs, 1H), 7.78 (bs, 1H), 7.75 (s,

992
V. A. Ryabinin et al. / Bioorg. Med. Chem. 8 (2000) 985±993
1H NMR of compound 18a ((CD3)2SO) d 11.48 (bs,
1H), 7.84 (s, 1H), 3.56 (t, 2H, J=7.0 Hz), 2.64 (t, 2H,
J=7.0 Hz), 2.53 (t, 2H, J=7.0 Hz), 2.20 (s, 6H), 2.10 (t,
2H, J=7.0 Hz), 1.40±1.90 (m, 8H).
KCl (in the case of DNA DNA and DNA RNA tem-
plate±primer duplexes, KCl was omitted), 0.5 mM
EDTA, 5 mM Mg(OAc)2, 1 mM dithiothreitol, 40 mM
[3H]dTTP (1.50 mCi) or in case of activated DNA all
four dNTPs: dATP, dCTP, dGTP and [3H]dTTP. The
polymerization reaction was initiated by addition of RT
(2±5 nM). Samples were taken from the reaction mixture
at 2±10 min intervals and further treatment of the reac-
tion was carried out as described.48
.
.
1H NMR of compound 18b ((CD3)2SO) d 12.30 (bs,
1H), 11.48 (bs, 1H), 7.98 (s, 1H), 7.84 (s, 1H), 3.54 (t,
2H, J=7.0 Hz), 2.65 (t, 2H, J=7.0 Hz), 2.53 (t, 2H,
J=7.0 Hz), 2.17 (s, 6H), 2.11 (t, 2H, J=7.0 Hz), 1.40±
1.90 (m, 8H).
Reaction mixtures contained each one of the template±
primer duplexes in the following optimal concentra-
1H NMR of compound 18c ((CD3)2SO) d 12.40 (bs,
2H), 11.48 (bs, 1H), 7.99 (s, 1H), 7.98 (s, 1H), 7.84 (s,
1H), 3.55 (t, 2H, J=7.0 Hz), 2.65 (t, 2H, J=7.0 Hz),
2.52 (t, 2H, J=7.0 Hz), 2.18 (s, 6H), 2.10 (t, 2H, J=7.0
Hz), 1.40±1.90 (m, 8H).
tions38 expressed as A260 units/mL: 0.5 poly(A) d(pT)16,
.
.
1.5 poly(A) (pU)16, 1.5 poly(dA) (pU)16, 2.0 poly(dA)
.
.
d(pT)16 or 2.0 activated DNA. The ratio of the primer
to template concentration (mononucleotide concentra-
tion) was 1:5 in all cases except for activated DNA.
1H NMR of compound 18d ((CD3)2SO) d 12.40 (bs,
2H), 12.20 (bs, 1H), 11.48 (bs, 1H), 8.01 (s, 1H), 7.99 (s,
1H), 7.98 (s, 1H), 7.84 (s, 1H), 3.56 (t, 2H, J=7.0 Hz),
2.63 (t, 2H, J=7.0 Hz), 2.54 (t, 2H, J=7.0 Hz), 2.18 (s,
6H), 2.11 (t, 2H, J=7.0 Hz), 1.40±1.90 (m, 8H).
Initial rates of the polymerization reaction were deter-
mined from the tangents of the data curves at zero time.
The oligopeptide concentration inhibiting the poly-
merization rate to 50% (IC50) was estimated from the
dependencies of the initial rates. The statistical error
was estimated using the data from 3±4 experiments.
1H NMR of compound 19a (D2O) d 7.69 (s, 1H), 3.00
(t, 2H, J=7.0 Hz), 2.86 (s, 3H), 2.51 (t, 2H, J=7.0 Hz),
1.69 (bruin, 4H, J=7.0 Hz), 1.43 (bruin, 4H, J=7.0 Hz).
Acknowledgements
1H NMR of compound 19b ((CD3)2SO) d 12.43 (bs,
1H), 11.95 (bs, 1H), 8.33 (s, 1H), 8.00 (bs, 1H), 7.85 (s,
1H), 4.30 (bs, 2H), 2.81 (s, 3H), 2.16 (t, 2H, J=7.0 Hz),
1.68 (m, 4H, J=7.0 Hz), 1.39 (m, 4H, J=7.0 Hz).
This work was supported in part by the program
INTAS-RFBR (95-1214), the Russian Fund of Basic
Research (RFBR) (96-04-49953, 97-04-49915), the Cen-
tre National de la Recherche Scienti®que (CNRS
France), PICS-RFBR (Nꢀ 571) and the French Agency
for Research against AIDS (ANRS). The authors are
grateful to G. V. Gursky and S. L. Grokhovsky for the
site-speci®city assay.
1H NMR of compound 19c ((CD3)2SO) d 12.57 (bs,
1H), 12.43 (bs, 1H), 11.95 (bs, 1H), 8.34 (s, 1H), 8.33 (s,
1H), 8.00 (bs, 1H), 7.85 (s, 1H), 4.40 (bs, 2H), 2.79 (s,
3H), 2.12 (t, 2H, J=7.0 Hz), 1.61 (m, 4H, J=7.0 Hz),
1.35 (m, 4H, J=7.0 Hz).
1H NMR of compound 20 ((CD3)2SO) d 12.40 (bs, 1H),
12.00 (bs, 1H), 8.19 (s, 1H), 7.88 (s, 1H), 3.48 (t, 2H,
J=7.0 Hz), 3.02 (t, 2H, J=7.0 Hz), 2.52 (t, 2H, J=7.0
Hz), 2.14 (t, 2H, J=7.0 Hz), 1.20±1.80 (m, 14H).
References
1. Larder, B. A.; Darby, G.; Richman, D. D. Science 1989,
243, 1731.
2. Larder, B. A.; Kemp, S. D. Science 1989, 246, 1155.
3. Bailly, C.; Chaires, J. B. Bioconjug. Chem. 1998, 9, 513.
4. Di Marco, A.; Gaetani, M.; Orezzi, P.; Arcamone, F. Can-
cer Chemother. Rep. 1962, 18, 15.
5. Julia, M.; Preau-Joseph, N. P. Bull. Soc. Chim. Fr. 1967, 97,
4348.
1H NMR of compound 21 ((CD3)2CO:D2O=1:1) d 7.94
(s, 1H), 7.92 (s, 1H), 7.90 (s, 1H), 7.73 (s, 1H), 3.81 (bs,
1H), 3.51 (m, 4H), 2.94 (t, 2H, J=7.0 Hz), 2.38 (t, 2H,
J=7.0 Hz), 1.63 (m, 4H) 1.37 (m, 2H).
6. Penco, S.; Redaelli, S.; Arcamone, F. Gazz. Chim. Ital.
1967, 97, 1110.
7. Arcamone, F.; Penco, S.; Delle Monache, F. Gazz. Chim.
Ital. 1969, 99, 620.
8. Lown, J. W. Antiviral Res. 1992, 17, 179.
9. Krowicki, K.; Lown, J. W. J. Org. Chem. 1987, 52, 3493.
10. Rao, K. E.; Dusgupta, D.; Sasisekharan, V. Biochemistry
1988, 27, 3018.
11. Rao, K. E.; Krowicki, K.; Burckhart, G.; Zimmer, C.;
Lown, J. W. Chem. Res. Toxicol. 1991, 4, 241.
12. Ding, L.; Grehn, L.; De Clercq, E.; Andrei, G.; Snoeck,
R.; Balzarini, J.; Fransson, B.; Ragnarsson, U. Acta Chem.
Scand. 1994, 48, 498.
Reverse transcriptase assay
Nucleotides, polynucleotides, salts and distamycin were
obtained from Sigma. [3H]dTTP was obtained from
Radioisotop (Russia) or ICN Biomedicals (France). All
oligodeoxyribonucleotides were synthesized as in ref 45
and were homogeneous according to reverse-phase and
ion-exchange chromatography. Oligoribonucleotides
were prepared as reported before.46 Calf thymus DNA
was hydrolyzed by DNase I to obtain activated DNA.48
Electrophoretically homogeneous HIV-1 RT p66/p51
was prepared as previously described.47
13. Plouvier, B.; Bailly, C.; Houssier, R.; Rao, K. E.; Lown, J.
W.; Heinichart, J. P. Nucleic Acids Res. 1991, 19, 5821.
14. Plouvier, B.; Bailly, C.; Houssier, R.; Heinichart, J. P.
Heterocycles 1991, 32, 693.
RT assay was performed at 30 ꢀC. The reaction mixture
(50 mL) contained 50 mM Tris±HCl, pH 8.0, 80 mM

V. A. Ryabinin et al. / Bioorg. Med. Chem. 8 (2000) 985±993
993
15. Wade, W. S.; Mrksich, M.; Dervan, P. B. Biochemistry
1993, 32, 11385.
34. Nevinsky, G. A. Mol. Biol.(Mosk.) 1995, 29, 6.
35. Khodyreva, S. N.; Podust, V. N.; Sergeev, D. S.; Ivanova,
E. M.; Frolova, E. I.; Koshkin, A. A.; Godovikova, T. S.;
Zarytova, V. F.; Richetti, M.; Lavrik, O. I. Mol. Biol. Rep.
1993, 18, 43.
36. Reid, R. S.; Stuart, A. L.; Gupta, S. V.; Latimer, L. J.;
Haug, B. L.; Lee, J. S. Nucleic Acids Res. 1987, 15, 4325.
37. Matthew, J. B.; Richards, F. M. Biopolymers 1984, 23, 2743.
38. Nevinsky, G. A.; Andreola, M-L.; Yamkovoy, V. I.;
Levina, A. S.; Barr, P. J.; Tarrago-Litvak, L.; Litvak, S. Eur.
J. Biochem. 1992, 207, 351.
16. Rao, K. E.; Sasisekharan, V. Indian J. Chem. 1990, 29B, 503.
17. Rao, K. E.; Sasisekharan, V. Indian J. Chem. 1990, 29B, 508.
18. Rao, K. E.; Bathini, Y. J. W. J. Org. Chem. 1990, 55, 728.
19. Mote, Jr, J.; Ghanouni, P.; Reines, D. A. J. Mol. Biol.
1994, 236, 725.
20. Filipowsky, M. E.; Kopka, M. L.; Brazilzison, M.; Lown,
J. W.; Dickerson, R. E. Biochemistry 1996, 35, 15397.
21. Chen, A. Y.; Yu, C.; Gatto, B.; Liu, L. F. Proc. Natl.
Acad. Sci. USA 1993, 90, 8131.
22. Kittler, L.; Bell, A.; Baguley, B. C.; Lober, G. Bioch. Mol.
Biol. Int. 1996, 40, 263.
39. Kolocheva, T. I.; Nevinsky, G. A. Mol. Biol. (Mosk.)
1993, 27, 1368.
23. Trauger, J. W.; Baird, E. E.; Dervan, P. B. Nature 1996,
382, 559.
40. Jacobo-Molina, A.; Ding, J.; Nanni, R.; Clark, A.; Lu, X.;
Tantillo, C.; Williams, R.; Kamer, G.; Ferris, A.; Clark, P.;
Hizi, A.; Hughes, S.; Arnold, E. Proc. Natl. Acad. Sci. USA
1993, 49, 6320.
41. Patel, P.; Jacobo-Molina, A.; Ding, J.; Tantillo, C.; Clark,
A.; Raag, R.; Nanni, R.; Hughes, S.; Arnold, E. Biochemistry
1995, 34, 5351.
24. Volpe, D. A.; Du, D. L.; Zurlo, M. G.; Mongelli, N.;
Murphy, M. J. Invest. New Drugs 1992, 10, 255.
25. Broggini, M.; Coley, H. M.; Mongelli, N.; Pesenti, E.;
Wyatt, M. D.; Hartley, J. A.; Dincalci, M. Nucleic Acids Res.
1995, 23, 81.
26. Lown, J. W. Drug Dev. Res. 1995, 34, 145.
27. Clanton, D. J.; Buckheit, R. W.; Terpening, S. J.; Kiser,
R.; Mongelli, N.; Borgia, A. L.; Schultz, R.; Narayanan, V.;
Bader, J. P.; Rice, W. G. Antiviral Res. 1995, 27, 335.
28. Baraldi, P. G.; Beria, I.; Cacciari, B.; Capolongo, L.;
Cozzi, P.; Mongelli, N.; Romagnoli, R.; Spalluto, G. Bioorg.
Med. Chem. Lett. 1996, 11, 1247.
29. Plouvier, B.; Houssin, R.; Helbecque, N.; Colson, P.;
Houssier, C.; Henichart, J. P.; Bailly, C. Anti-Cancer Drug
Design 1995, 10, 155.
30. Plouvier, B.; Houssin, R.; Hecquet, B.; Colson, P.; Hous-
sier, C.; Waring, M. J.; Henichart, J. P.; Bailly, C. Bioconjug.
Chem. 1994, 5, 475.
31. Lown, J. W. J. Mol. Recognit. 1994, 7, 79.
32. Retroviruses; Con, J. M.; Hughes, S. H.; Varmus, H. E.,
Eds.; Cold Spring Harbor Laboratory Press, 1997.
33. Wyatt, M. D.; Garbiras, B. J.; Lee, M.; Forrow, S. M.;
Hartley, J. A. Bioorg. Med. Chem. Lett. 1994, 4, 801.
42. Pelletier, H.; Sawaya, M. R.; Kumar, A.; Wilson, S. H.;
Kraut, J. Science 1994, 264, 1891.
43. Eckelman, W. C.; Karesh, S. M.; Reba, R. C. J. Pharm.
Sci. 1975, 64, 704.
44. Sprague, J. M.; Lincoln, R. M.; Ziegler, C. J. Am. Chem.
Soc. 1946, 68, 266.
45. Zarytova, V. P.; Ivanova, E. M.; Romanenko, V. P.
Bioorg. Chem. (Mosk.) 1985, 11, 815.
46. Mudrakovskaya, A. V.; Yamkovoy, V. I. In The Enzymes
of Microorganisms and Biopolymer Degradation; Debabov, V.
G., Gordon, I. O., Eds.; NPO Medbioeconomics: Moscow,
1990; pp 199±206.
47. Sallafranque-Andreola, M. L.; Robert, D.; Barr, P. J.;
Fournier, M.; Litvak, S.; Sarih-Cottin, L.; Tarrago-Litvak, L.
Eur. J. Biochem. 1989, 184, 367.
48. Nevinsky, G. A.; Veniaminova, A. G.; Levina, A. S.;
Podust, V. N.; Lavrik, O. I.; Holler, E. Biochemistry 1990, 29,
1200.