IMPDH as a biological probe for RNA antiviral drug discovery: Synthesis, enzymology, molecular docking, and antiviral activity of new ribonucleosides with surrogate bases

Article abstract of DOI:10.1080/15257770701490506

Our interest in the discovery of molecules with antiviral activity against RNA viruses led us to the design of ribonucleosides with surrogate bases with the intent of using inhibition of inosine monophosphate dehydrogenase (IMPDH) as a probe for antiviral drug discovery. A general methodology for the preparation of these compounds is discussed. Kinetic parameters of the inhibition studies with IMPDH, which were carried out spectrophotometrically by monitoring the formation of NADH, are given. Antiviral information and correlation of activity with IMPDH inhibition are discussed. Copyright Taylor & Francis Group, LLC.

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Impdh As A Biological Probe For Rna
Antiviral Drug Discovery: Synthesis,
Enzymology, Molecular Docking,
And Antiviral Activity Of New
Ribonucleosides With Surrogate Bases
Vasu Nair a , Xiaohui Ma a , Qingning Shu a , Fan Zhang a , Vinod Uchil
a & Govardhan R. Cherukupalli a
a Center for Drug Discovery and the Department of Pharmaceutical
and Biomedical Sciences, University of Georgia, Athens, Georgia,
USA
Published online: 10 Dec 2007.
To cite this article: Vasu Nair , Xiaohui Ma , Qingning Shu , Fan Zhang , Vinod Uchil & Govardhan
R. Cherukupalli (2007) Impdh As A Biological Probe For Rna Antiviral Drug Discovery: Synthesis,
Enzymology, Molecular Docking, And Antiviral Activity Of New Ribonucleosides With Surrogate Bases,
Nucleosides, Nucleotides and Nucleic Acids, 26:6-7, 651-654, DOI: 10.1080/15257770701490506
To link to this article: http://dx.doi.org/10.1080/15257770701490506
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Nucleosides, Nucleotides, and Nucleic Acids, 26:651–654, 2007
Copyright ꢀ Taylor & Francis Group, LLC
C
ISSN: 1525-7770 print / 1532-2335 online
DOI: 10.1080/15257770701490506
IMPDH AS A BIOLOGICAL PROBE FOR RNA ANTIVIRAL DRUG
DISCOVERY: SYNTHESIS, ENZYMOLOGY, MOLECULAR DOCKING,
AND ANTIVIRAL ACTIVITY OF NEW RIBONUCLEOSIDES WITH
SURROGATE BASES
Vasu Nair, Xiaohui Ma, Qingning Shu, Fan Zhang, Vinod Uchil,
and Govardhan R. Cherukupalli
Center for Drug Discovery and the Department of
2
Pharmaceutical and Biomedical Sciences, University of Georgia, Athens, Georgia, USA
Our interest in the discovery of molecules with antiviral activity against RNA viruses led us
2
to the design of ribonucleosides with surrogate bases with the intent of using inhibition of inosine
monophosphate dehydrogenase (IMPDH) as a probe for antiviral drug discovery. A general method-
ology for the preparation of these compounds is discussed. Kinetic parameters of the inhibition studies
with IMPDH, which were carried out spectrophotometrically by monitoring the formation of NADH,
are given. Antiviral information and correlation of activity with IMPDH inhibition are discussed.
Keywords IMPDH inhibitors; synthesis; antiviral
INTRODUCTION
The enzyme, inosine monophosphate dehydrogenase (IMPDH; EC
1.1.1.205), catalyses the oxidative conversion of inosine 5ꢁ-monophosphate
(IMP) to xanthosine 5ꢁ-monophosphate (XMP) with the involvement of the
coenzyme, nicotinamide adenine dinucleotide (NAD+).[1,2] IMPDH is an
important target for the discovery of antiviral, anticancer, and immuno-
suppressive agents.[3] Consistent with this is the observation that some in-
hibitors of IMPDH have been found to have anticancer, antiviral and im-
munosuppressive activity.[4−7] IMPDH is a sulfhydryl enzyme in which the
Cys-331 residue in the active site may act as a nucleophilic participant in
interactions with inhibitors that carry Michael acceptors at appropriate po-
sitions on the nucleobase.[7−9] Participation of the Cys-331 is also consis-
tent with the mechanism of substrate action of IMPDH, which involves
This project was supported by Grant No. AI056540 from the NIH. Its contents are solely the respon-
sibility of the authors and do not necessarily represent the official views of the NIH. We thank Drs. E.
Kern, R. Sidwell, and J. Huggins for antiviral screening results.
Address correspondence to Vasu Nair, Department of Pharmaceutical and Biomedical Sci-
ences, Room 320, R.C. Wilson PH, The University of Georgia, Athens, GA 30602-2352, USA. E-mail:
vnair@rx.uga.edu
651

652
V. Nair et al.
FIGURE 1 Target molecules of this investigation.
covalent interaction of the enzyme and coenzyme (NAD+) complex with
the 2-position of IMP.[1] In an ongoing drug discovery program on RNA
antiviral compounds in our laboratory,[7,8,10,11] we have utilized IMPDH
as a probe for the initial identification of potential antiviral molecules
(Figure 1).
RESULTS AND DISCUSSION
A general and efficient synthetic approach to compounds of Figure 1
is illustrated in Scheme 1. Guanosine (11) was converted to the 2-iodo
SCHEME 1 A concise synthetic approach to C-2 functionalized inosine analogues.

IMPDH as Biological Probe
653
TABLE 1 Data on inhibition of IMPDH by C-2 functionalized IMP analogues
b
c
Inhibitors
Ki (µM)a
k
inact(s−1
)
k
on(M−1s−1
)
1
2
3
5
6
8
9
10
3.98
—
4.25
0.029
—
0.013
0.027
—
—
0.023
0.030
0.73 × 104
2.12 × 104
0.33 × 104
2.67 × 104
—
1.11
no inhibition
81.8 (reversible)
74.4
—
3.05 × 102
—
4.70
aKi = Ki,app /(1 + [IMP])/Km).
bc
k
= kinact/Ki .
on
compound 13 by acetylation and deamination/halogenation reactions. The
key step in the synthesis was the palladium-catalyzed cross-coupling of inter-
mediate 13 with functionalized stannanes[12] to give 14. Deprotection under
standard conditions gave the target molecules 15. For compounds, 8 and 9,
the methodology used has been described previously by us.[10]
Data from inhibition studies with inosine monophosphate dehydroge-
nase (IMPDH) from E. coli are summarized in Table 1. Details of the proce-
dure for the enzyme kinetics are discussed.[8,11] It is clear from the data that
most of the target compounds are strong irreversible inhibitors of IMPDH.
FIGURE 2 Docking results of inhibitor 1 in the active site of IMPDH. The phosphate group is locked
into position by polar interactions with Ile330, Gly366, Ser388 and Tyr411. The ribose hydroxyls form
hydrogen bonds with Ser68 and Asp364. The backbone N-H of Gly415 and Met414, and C=O of
Gln441stabilizes the base moiety by forming hydrogen bonds. The S atom of active residue Cys331
+
˚
is 4.0 A away from the 2-vinyl group terminal carbon. Cofactor NAD stacks with the base moiety of
inhibitor.

654
V. Nair et al.
Antiviral data on the target nucleosides correlate well with inhibition of
IMPDH by their monophosphates (see[7] for general mechanistic explana-
tion). For example, the parent nucleosides of compounds 1, 2, 3, 5, and 10
show moderate to good antiviral activity against a number of RNA viruses
and also against a few DNA viruses. Antiviral studies are continuing and the
completed results will be published elsewhere.
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545–560.
2. Goldstein, B.M.; Colby, T.D. IMP dehydrogenase: Structural aspects of inhibitor binding. Curr. Med.
Chem. 1999, 6, 519–536.
3. De Clerq, E. Strategics in the design of antiviral drugs. Nature Rev. Drug Discovery 2002, 11, 13–25.
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tumor and antiviral agents. Curr. Med. Chem. 1999, 6, 599–614.
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fluorovinyl] inosine 5ꢁ-monophosphate. Bioorg. Med. Chem. Lett. 2003, 13, 645–647.
12. Nair, V.; Turner, G.A.; Chamberlain, S.D. Novel approaches to functionalized nucleosides via
palladium-catalyzed cross-coupling with organostannanes. J. Am Chem. Soc. 1987, 109, 7223–7224.