Discovery of a potent HIV integrase inhibitor that leads to a prodrug with significant anti-HIV activity

Article abstract of DOI:10.1021/ml2001246

Worldwide research efforts in drug discovery involving HIV integrase have produced only one compound, raltegravir, that has been approved for clinical use in HIV/AIDS. As resistance, toxicity, and drug-drug interactions are recurring issues with all class

Full text of DOI:10.1021/ml2001246

Letter
pubs.acs.org/acsmedchemlett
Discovery of a Potent HIV Integrase Inhibitor That Leads to a
Prodrug with Significant anti-HIV Activity
Byung I. Seo, Vinod R. Uchil, Maurice Okello, Sanjay Mishra, Xiao-Hui Ma, Malik Nishonov,
Qingning Shu, Guochen Chi, and Vasu Nair*
Department of Pharmaceutical and Biomedical Sciences and the Center for Drug Discovery, University of Georgia, Athens,
Georgia 30602, United States
S
* Supporting Information
ABSTRACT: Worldwide research efforts in drug discovery involving HIV integrase have
produced only one compound, raltegravir, that has been approved for clinical use in HIV/
AIDS. As resistance, toxicity, and drug−drug interactions are recurring issues with all classes of
anti-HIV drugs, the discovery of novel integrase inhibitors remains a significant scientific
challenge. We have designed a lead HIV-1 strand transfer (ST) inhibitor (IC50 70 nM),
strategically assembled on a pyridinone scaffold. A focused structure−activity investigation of
this parent compound led to a significantly more potent ST inhibitor, 2 (IC50 6 ± 3 nM).
Compound 2 exhibits good stability in pooled human liver microsomes. It also displays a
notably favorable profile with respect to key human cytochrome P450 (CYP) isozymes and
human UDP glucuronosyl transferases (UGTs). The prodrug of inhibitor 2, i.e., compound 10, was found to possess remarkable
anti-HIV-1 activity in cell culture (EC50 9 ± 4 nM, CC50 135 ± 7 μM, therapeutic index = 15 000).
KEYWORDS: Integrase inhibitor, pyridinone, CYP/UGT profile, anti-HIV prodrug
favorable in vitro drug−drug interaction profile with respect
he retroviral enzyme, HIV-1 integrase, which is encoded at
Tthe 3′-end of the pol gene of the human immunodeficiency to key cytochrome P450 (CYP) and UDP glucuronosyltrans-
virus (HIV), is essential for HIV replication and is a significant
target for the discovery and development of anti-HIV
therapeutic agents.1−11 However, research efforts in the area
of anti-HIV integrase inhibitors for the treatment of acquired
immunodeficiency syndrome (AIDS) have resulted in only one
compound, raltegravir (Isentress), that has been approved by
the FDA for the clinical treatment of HIV-AIDS.7,8 However, as
resistance, toxicity, and drug−drug interactions are recurring
issues with all classes of anti-HIV drugs, the discovery of new,
anti-HIV active integrase inhibitors remains a significant
scientific challenge. HIV-1 integrase is a 32 kDa protein,1,12,13
which catalyzes the incorporation of HIV DNA into host
chromosomal DNA through a specifically defined sequence of
reactions, which involves 3′-processing and a key strand transfer
(ST) step.1,3,12−15 Initiation of integration occurs in the
cytoplasm, where a complex is formed between viral cDNA,
previously produced by reverse transcription, and HIV
integrase. Following this is site-specific endonucleatic cleavage
of two nucleotides from each 3′-end of double-stranded viral
DNA, which produces truncated viral DNA with terminal
CAOH-3′ (3′-processing). The next step, ST, occurs in the
nucleus and involves staggered nicking of chromosomal DNA
and joining of each 3′-end of the recessed viral DNA to the 5′-
ends of the host DNA, followed by repair/ligation. The ST step
is carried out after transport of the processed, preintegration
complex from the cytoplasm into the nucleus. Both 3′-
processing and ST steps require divalent metal ion cofactors.
To explore whether a significantly anti-HIV active integrase
inhibitor could be discovered that would also possess a
ferase (UGT) isozymes, we carried out the design of such an
inhibitor from a lead compound discovered in our laboratory.
This lead compound was 4-(1,5-dibenzyl-1,2-dihydro-2-oxopyr-
idin-3-yl)-2-hydroxy-4-oxobut-2-enoic acid (1, Figure 1), which
Figure 1. Structure of lead compound 1.
was an inhibitor of the ST step of HIV-1 integrase (IC50
70 nM). Using compound 1 as a starting point, we undertook
lead optimization studies on 1.16
In the discovery of lead compound 1, it was established that
the specific nature of the modified nucleobase scaffold (i.e., the
pyridinone ring) and the nature of the substituents on the
scaffold (the functional components as well as the hydrophobic
benzyl groups) were critical for integrase inhibitory activity. For
this reason, we focused our optimization studies on substituents
on the hydrophobic phenyl groups of the pyridinone scaffold.
Received: May 26, 2011
Accepted: October 5, 2011
Published: October 5, 2011
© 2011 American Chemical Society
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ACS Medicinal Chemistry Letters
Letter
Table 1. In Vitro Anti-HIV Data for Analogues of Compound 1
Table 1. continued
Table 1. continued
aryl ring A
aryl ring B
aryl ring A
aryl ring B
ab
,
ab
,
entry
o
m
p
o
m
p
EC50 (μM)
entry
o
m
p
o
m
p
EC50 (μM)
1
H
F
H
H
H
H
H
H
H
H
Cl
H
H
F
H
H
F
H
H
H
H
F
H
H
H
H
H
H
H
H
H
H
H
H
Cl
Cl
H
Cl
Cl
Cl
H
Cl
H
Cl
H
H
Cl
Cl
Cl
Me
H
H
H
H
H
H
H
F
2.1
0.9
1.0
0.6
0.7
1.8
0.8
1.6
0.8
>5.0
0.3
2.0
1.1
1.2
0.7
2.4
>5.0
2.5
0.7
>5.0
0.6
1.4
1.4
0.8
2.4
2.5
1.4
1.4
0.6
30
31
32
33
34
35
36
H
F
Me
Cl
H
H
H
H
F
H
H
H
H
H
H
F
Cl
H
F
2.2
0.7
0.9
1.4
1.4
1.2
0.7
5.1
1.5
0.8
2.2
1.9
1.9
0.9
0.9
0.7
1.1
0.8
1.0
0.8
1.5
0.6
0.7
>5.0
1.3
1.0
0.5
1.3
2
H
H
H
F
3
H
F
F
5-Cl
Cl
H
4
F
F
Cl
Cl
H
5
F
H
F
H
6
H
H
H
H
H
F
OMe
F
H
H
H
H
H
F
H
H
Cl
H
H
H
H
H
H
H
H
H
H
H
F
7
Cl
H
H
F
8
Me
F
F
37 2,6-di-F
38 2,6-di-F
H
H
H
H
F
H
H
F
Cl
H
9
H
H
F
10
11
12
13
14
15
16
17
18
19
20
21
22
OMe
F
OMe
F
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
H
F
H
H
F
Cl
H
F
H
H
H
F
H
H
H
H
H
H
H
H
F
H
H
H
F
H
F
H
H
H
H
F
Cl
Cl
H
Cl
H
H
H
H
Cl
H
H
H
F
H
H
F
Cl
H
F
F
H
H
F
Me
F
H
H
H
H
H
H
F
F
F
H
F
H
Me
Me
Me
F
F
H
F
H
F
CN
H
H
F
H
H
H
F
H
H
F
F
H
H
F
F
H
Cl
F
H
H
H
H
H
F
H
Cl
H
Me
H
H
H
H
H
H
H
H
F
F
H
H
F
H
H
H
H
H
H
F
H
F
Cl
H
H
F
F
23 2,6-di-F
H
H
Cl
H
Cl
Cl
H
H
H
H
H
F
F
H
F
24
25
26
27
28
H
F
F
H
H
H
H
H
H
H
Me
Me
F
COOH
H
H
F
H
F
H
H
F
Cl
H
H
H
H
F
H
F
H
H
H
F
H
F
Me
H
F
T2a9ble F1. continued
F
a
b
EC50 values are the average of three determinations. Standard deviations for the EC50 are within 31% of the average. EC50 = concentration for 50%
inhibition of the replication of HIV-1.
In the subsequent study, we examined the effects of various
substituents, e.g., methoxy, chloro, alkyls, and mixed halo/alkyl
and others, on the phenyl rings and their effect on the
enzymology involving ST step inhibition. There was considerable
variation in the ST inhibitory activity for these compounds
(IC50 <10 nM to >1500 nM). Fluoro substitution IC50 data,
however, were more compelling. Among this entire group of
fluorinated compounds, the difluoro, trifluoro, and tetrafluoro
substituted compounds all had ST inhibitory IC50 values falling
in the range of <10 nM, showing significant improvement over
lead compound 1. Within this group of fluorinated compounds,
the trifluoroaryl (o- and o,p) and tetrafluoroaryl (o,p and o,p)
substituted analogues (involving both phenyl rings) were the
most active in terms of the integrase IC50 and IC90 data (≤6 and
<100 nM, respectively). While the detailed reason for the
increase in inhibitory potency with appropriate fluorine
substitution is not fully understood; hydrophobic and/or
electrostatic interactions may contribute.17−19 In the next level
of lead optimization, we investigated the antiviral cell culture data
for these compounds. The results are summarized in Table 1
and show that the anti-HIV-1 EC50 values were largely in the
1−3 μM range. However, two compounds emerged from these
studies that exhibited anti-HIV EC50 values of 500 nM or less.
They were 4-(5-(2,4-difluorobenzyl)-1-(2-fluorobenzyl)-2-oxo-
1,2-dihydropyridin-3-yl)-2-hydroxy-4-oxobut-2-enoic acid
(2, entry 56, Table 1) and 4-(1,5-bis(2,4-difluorobenzyl)-2-
oxo-1,2-dihydropyridin-3-yl)-4-hydroxy-2-oxobut-3-enoic acid
(entry 11, Table 1). Their ST inhibition IC50 data were 6 ±
3 nM and 5.5 ± 1.5 nM, respectively. The eventual selection
of compound 2 over entry 11 as the key compound to move
forward is discussed in the prodrug section below.
A highly efficient synthesis of compound 2 (Scheme 1) was
developed in our laboratory. Only seven steps (aromatic
nucleophilic addition, demethylation/deoxygenation, radical
bromination, benzylation, palladium-catalyzed cross-coupling,
Claisen condensation, and acid-catalyzed hydrolysis)20−26 were
required for the total synthesis of 2 from commercially available
5-bromo-2-methoxypyridine (3). The overall yield from 3 was 37%.
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ACS Medicinal Chemistry Letters
Letter
Scheme 1. Methodology Developed for the Synthesis of
a
Integrase Inhibitor 2
Figure 2. Stability of 2 in pooled human liver microsomes monitored
by HPLC/UV.
metabolized by different CYP isoforms. In addition, compound 2
did not exhibit any activation of these CYP isozymes. Thus, our
studies suggest that this integrase-based, anti-HIV compound is
anticipated to have a favorable drug interaction profile with
respect to key CYP isozymes.
Because isozymes of UGT also play an important role in
determining drug−drug interactions, we investigated the substrate
activity of 2 toward key human UGTs.27,31 Compound 2 was not
a substrate for the following key UGT isozymes: 1A1, 1A4, 1A6,
1A9, and 2B7. In comparison, the major mechanism for rapid
clearance of raltegravir in humans is through UGT 1A1-mediated
glucuronidation.27 The integrase inhibitor, S/GSK 1349572, is
also a substrate for UGT 1A1, and its primary route of
metabolism and subsequent clearance is glucuronidation.10
Antiviral data in cell culture of compound 2 revealed a
significant disconnect of almost two or more orders of
magnitude between the anti-HIV-1 activity (EC50 500 nM,
MAGI cells) and the ST inhibition data for 2 (IC50 6 nM). For
HIV-1 integrase inhibitors, there is normally a reasonably
strong correlation between ST IC50 data and cell culture EC50
data.3,5 Because the disconnect between the IC50 and EC50 data
for compound 2 and also other compounds, including entry 11
(Table 1), appeared to be a problem associated with the cellular
permeability of the inhibitors, we examined prodrugs of these
compounds. The isopropyl ester prodrug, 10 (Figure 3), was
a
Abbreviations: (i) tert-butyl methyl ether (TBME); (ii) trimethylsilyl
chloride (TMSCl), triethylsilane (TES), trifluoroacetic acid (TFA);
(iii) N-bromosuccinimide (NBS); (iv−v) dimethylformamide (DMF);
(vi) tetrahydrofuran (THF).
Stability studies of compound 2 in pooled human liver
microsomes were carried out by preincubation, initiation with
NADPH, incubation at 37 °C, quenching of samples at various
time intervals with cold acetonitrile, centrifugation to remove
precipitated proteins, and finally HPLC analysis that utilized UV
detection.27−29 These studies revealed that integrase inhibitor 2
was relatively stable in human liver microsomes, exhibiting an in
vitro half-life of >3 h, as evidenced from HPLC data, which
showed that 80% of compound 2 remained after the 3 h
incubation in microsomes (Figure 2). The key metabolite, which
was slowly produced, was identified by HPLC and HRMS data
to be the product of the retro-Claisen cleavage of the diketo
group of 2 to produce the acetyl pyridinone 8.
The in vitro drug interaction profile involving key cytochrome
P450 (CYP) isozymes28−30 and appropriate substrates in pooled
human liver microsomes with varying concentrations of 2,
followed by kinetic analysis (Table 2), showed that compound 2
was not an inhibitor of CYP3A4 and CYP2D6 isozymes and was
a very weak inhibitor of the CYP2C8 isozyme (Table 1). These
key isozymes account for a total of over 80% of drugs that are
Figure 3. Structures of prodrug 10 and raltegravir.
Table 2. IC50 Data for Inhibition of Key Cytochrome P450 Isozymes
CYP450 isozyme
substrate/stock solution
conc (μM)
protein (mg/mL)
incubation (min)
IC50 data (μM)
CYP3A4
CYP3A4
CYP2D6
CYP2C8
testosterone (50 mM)
triazolam (50 mM)
100
200
200
200
0.3
0.4
2.0
0.4
30
30
60
30
>200
>200
>200
>65
dextromethorphan (50 mM)
amodiaquine (5 mM)
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ACS Medicinal Chemistry Letters
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easily synthesized from compound 2 through acid-catalyzed
esterification with 2-propanol. The cLog P values for
compounds 2 and the isopropyl ester 10 are 2.38 and 4.24,
respectively, suggesting that compound 10 is significantly less
polar than compound 2 and thus would be expected to be more
cellularly permeable. The antiviral data confirmed this, as
isozymes, as it was not an inhibitor or activator of these
isozymes. Also of significance was the observation that inhibitor
2 was not a substrate of important human UGTs. A prodrug of
2, i.e., compound 10, exhibited remarkable anti-HIV-1 activity
in cell culture (EC50 = 9 nM, CC50 = 135 μM). The therapeutic
or selectivity index of prodrug 10, which was 15 000, was also a
notable finding. Further biological studies are in progress.
prodrug 10 exhibited remarkable anti-HIV-1 activity (EC50
=
9 ± 4 nM, MAGI cells). This was the best activity achieved of
all of the prodrugs studied in this work (EC50 values ranged
from 9 to 46 nM for the isopropyl ester prodrugs). The
isopropyl ester prodrug of entry 11 (Table 1) was significantly
less active than compound 10, exhibiting an EC50 of 46 ±
18 nM (MAGI cells). The overall performance of the assay was
validated by the MOI-sensitive positive control compound,
raltegravir, which exhibited the expected level of antiviral
activity (EC50 < 6 nM).7 Cell viability data for 10 showed only
low toxicity at higher test concentrations (CC50 = 135 ± 7 μM,
CC90 > 200 μM), although a CC90 was not reached at the
highest test concentration (200 μM). It is of relevance to
mention that the EC50 and EC90 data for 10 correlate
exceptionally well with the ST inhibition IC50 and IC90 data
for 2 (6 and 97 nM, respectively). The therapeutic (selectivity)
index TI (CC50/EC50) for 10 was 15 000. Finally, isopropyl
ester 10 was a poor inhibitor of HIV integrase in enzymatic
studies (IC50 > 475 nM), suggesting that the anti-HIV
activity of 10 was most likely the result of its hydrolysis in
cell culture to produce the cellularly active anti-HIV compound
2. Consistent with this conclusion was our observation that
CYP and UGT studies on compound 10 were precluded by
its rapid hydrolysis in human liver microsomes to produce
compound 2 (100% conversion in 15 min).
ASSOCIATED CONTENT
* Supporting Information
Antiviral assays, microsome stability assays, cytochrome P450
inhibition assays, UDP-glucuronosyltransferase substrate assays,
synthesis of integrase inhibitor and its prodrug, analytical data
for all compounds, and HPLC purity trace of integrase
inhibitor. This material is available free of charge via the
Internet at http://pubs.acs.org.
■
S
AUTHOR INFORMATION
Corresponding Author
*E-mail: vnair@rx.uga.edu.
Funding
This project was supported by research grant RO1 AI 43181
(NIAID) and shared equipment grant IS10RR016621 (NCRR)
from the National Institutes of Health. Its contents are solely
the responsibility of the authors and do not necessarily
represent the official views of the NIH. We also thank the
Terry Chair Endowment, the Georgia Research Alliance, and
the University of Georgia for additional research support.
■
ACKNOWLEDGMENTS
■
Some of the data cited here were determined at Inhibitex, Inc.,
Alpharetta, GA, and at the Southern Research Institute,
Frederick, MD, and we express our thanks to them.
While a number of structurally diverse compounds have been
reported to be inhibitors of HIV integrase, the data of two of
these compounds (Figure 4), that have received considerable
REFERENCES
■
(1) Frankel, A. D.; Young, J. A. T. HIV-1: Fifteen Proteins and an
RNA. Annu. Rev. Biochem. 1998, 67, 1−25.
(2) Trono, D.; Van Lint, C.; Rouzioux, C.; Verdin, E.; Barre-Sinoussi,
F.; Chun, T.-W.; Chomont, N. HIV Persistence and the Prospect of
Long-Term Drug-Free Remissions for HIV Infected Individuals.
Science 2010, 329, 174−180.
(3) Nair, V.; Chi, G. HIV Integrase Inhibitors as Therapeutic Agents
in AIDS. Rev. Med. Virol. 2007, 17, 277−295.
(4) Nair, V.; Chi, G.; Ptak, R.; Neamati, N. HIV Integrase Inhibitors
with Nucleobase Scaffolds: Discovery of a Highly Potent Anti-HIV
Agent. J. Med. Chem. 2006, 49, 445−447.
(5) Pommier, Y.; Johnson, A. A.; Marchand, C. Integrase Inhibitors
to Treat HIV/AIDS. Nat. Rev. Drug Discovery 2005, 4, 236−248.
(6) Nair, V. Novel Inhibitors of HIV Integrase: the Discovery of
Potential Anti-HIV Therapeutic Agents. Frontiers Med. Chem. 2005, 2,
3−20.
(7) Summa, V.; Petrocchi, A.; Bonelli, F.; Crescenzi, B.; Donghi, M.;
Ferrara, M.; Fiore, F.; Gardelli, C.; Paz, O. G.; Hazuda, D. J.; Jones, P.;
Kinzel, O.; Laufer, R.; Monteagudo, E.; Muraglia, E.; Nizi, E.; Orvieto,
F.; Pace, P.; Pescatore, G.; Scarpelli, R.; Stillmock, K.; Witmer, M. V.;
Rowley, M. Discovery of Raltegravir, a Potent, Selective Orally
Bioavailable HIV-Integrase Inhibitor for the Treatment of HIV-AIDS
Infection. J. Med. Chem. 2008, 51, 5843−5855.
(8) Laufer, R.; Paz, O. G.; DiMarco, A.; Bonelli, F.; Monteagudo, E.;
Summa, V.; Rowley, M. Quantitative Prediction of Human Clearance
Guiding the Development of Raltegravir (MK-0518, Isentress) and
Related HIV Integrase Inhibitors. Drug Metab. Dispos. 2009, 37, 873−
883.
Figure 4. Two well-known β-diketo compounds.
attention and that have a relationship, albeit peripheral, to the
compounds described herein, are worthy of mention. Both
compounds are ST inhibitors [IC50 20 nM (S-1360) and
170 nM (L-731,988)]. The in vitro anti-HIV data for the
β-diketo triazole, S-1360,32 showed an EC50 of 140 nM and a
CC50 of 110 μM (PBMC), resulting in a TI of 790. Compound
L-731,988 is somewhat less active (EC50 1 μM in MT-4 cells).33
The CC50 value was not given.
In summary, our search for new integrase-based, anti-HIV
compounds led to the discovery of a highly potent ST inhibitor
of HIV-1 integrase, 2 (IC50 6 nM). This compound was
relatively stable in pooled human liver microsomes (80% of
compound remained after incubation for 3 h). It displayed a
favorable interaction profile with respect to key human CYP
(9) Garvey, E. P.; Johns, B. A.; Gartland, M. J.; Foster, S. A.; Miller,
W. H.; Ferris, R. G.; Hazen, R. J.; Underwood, M. R.; Boros, E. E.;
880
dx.doi.org/10.1021/ml2001246|ACS Med. Chem. Lett. 2011, 2, 877−881

ACS Medicinal Chemistry Letters
Letter
Thompson, J. B.; Weatherhead, J. G.; Koble, C. S.; Allen, S. H.;
Schaller, L. T.; Sherrill, R. G.; Yoshinaga, T.; Kobayashi, M.; Wakasa-
Morimoto, C.; Miki, S.; Nakahara, K.; Noshi, T.; Sato, A.; Fujiwara, T.
The Naphthyridinone GSK364735 is a Novel, Potent Human
Immunodeficiency Virus Type I Integrase Inhibitor and Antiretroviral.
Antimicrob. Agents Chemother. 2008, 52, 901−908.
(10) Min, S.; Song, I.; Borland, J.; Chen, S.; Lou, Y.; Fujiwara, T.;
Piscitelli, S. C. Pharmacokinetics and Safety of S/GSK1349572, a
Next-Generation HIV Integrase Inhibitor, in Healthy Volunteers.
Antimicrob. Agents Chemother. 2010, 54, 254−258.
(29) Baranczewski, P.; Stanczak, A.; Sundberg, K.; Svensson, R.;
Wallin, A.; Jansson, J.; Garberg, P.; Postlind, H. Introduction to In
Vitro Estimation of Metabolic Stability and Drug Interactions of New
Chemical Entities in Drug Discovery and Development. Pharmacol.
Rep. 2006, 58, 453−472.
(30) Chauret, N.; Gauthier, A.; Martin, J.; NicollGriffith, D. A. Vitro
Comparison of Cytochrome P450-Mediated Metabolic Activities in
Human, Dog, Cat, and Horse. Drug Metab. Dispos. 1997, 25, 1130−
1136.
(31) Williams, J. A.; Hyland, R.; Jones, B. C.; Smith, D. A.; Hurst, S.;
Goosen, T. C.; Peterkin, V.; Koup, J. R.; Ball, S. E. Drug-Drug
Interactions for UDP-Glucuronosyltransferase Substrates: A Pharma-
cokinetics Explanation for Typically Observed Low Exposure (AUCi/
AUC) Ratios. Drug Metab. Dispos. 2004, 32, 1201−1208.
(32) Yoshinaga, T.; Sato, A.; Fujishita, T.; Fujiwara, T. S-1360: In
Vitro Activity of a New HIV-1 Integrase Inhibitor in Clinical
Development. Presented at the 9th Conference on Retroviruses and
Opportunistic Infections, Seattle, WA; February 24−28, 2002;
Abstract 8, p 55. Data given for compound S-1360 are those cited
by Billich, A. S-1360 Shionogi-GlaxoSmithKline. Curr. Opin. Invest.
Drugs 2003, 4, 206−209.
(11) Klibanov, O. M. Elvitegravir, an Oral HIV Integrase Inhibitor,
for the Potential Treatment of HIV Infection. Curr. Opin. Invest. Drugs
2009, 10, 190−200.
(12) Asante-Appiah, E.; Skalka, A. M. HIV-1 Integrase: Structural
Organization, Conformational Changes, and Catalysis. Adv. Virus Res.
1999, 52, 351−369.
(13) Esposito, D.; Craigie, R. HIV Integrase Structure and Function.
Adv. Virus Res. 1999, 52, 319−333.
(14) Liao, C.; Marchand, C.; Burke, T. R.; Pommier, Y.; Nicklaus, M.
C. Authentic HIV-1 Integrase Inhibitors. Future Med. Chem. 2010, 2,
1107−1122.
(33) Hazuda, D. J.; Felock, P.; Witmer, M.; Wolfe, A.; Stillmock, K.;
Grobler, J. A.; Espeseth, A.; Gabryelski, L.; Schleif, W.; Blau, C.; Miller,
M. D. Inhibitors of Strand Transfer that Prevent Integration and
Inhibit HIV-1 Replication in Cells. Science 2000, 287, 646−650.
(15) Hare, S.; Gupta, S. S.; Valkov, E.; Engelman, A.; Cherepanov, P.
Retroviral Intasome Assembly and Inhibition of DNA Strand Transfer.
Nature 2010, 464, 232−236.
(16) Cox, A. G.; Nair, V. Novel HIV Integrase Inhibitors with Anti-
HIV Activity: Insights into Integrase Inhibition from Docking Studies.
Antiviral Chem. Chemother. 2006, 17, 343−353.
(17) Urban, J. J.; von Tersch, R. L.; Famini, G. R. Effect of Fluorine
Substitution on Phenol Acidities in the Gas Phase and in Aqueous
Solution. A Computational Study Using Continuum Solvation Models.
J. Org. Chem. 1994, 59, 5239−5245.
(18) Resnati, G. Synthesis of Chiral and Bioactive Fluoroorganic
Compounds. Tetrahedron 1993, 49, 9385−9445.
(19) DiMagno, S.; Sun, H. The Strength of Weak Interactions:
Aromatic Fluorine in Drug Design. Curr. Top. Med. Chem. 2006, 6,
1473−1482.
(20) Boros, E. E.; Burova, S. A.; Erickson, G. A.; Johns, B. A.; Koble,
C. S.; Kurose, N.; Sharp, M. J.; Tabet, E. A.; Thompson, J. B.; Toczko,
M. A. A Scaleable Synthesis of Methyl 3-Amino-5-(4-fluorobenzyl)-2-
pyridinecarboxylate. Org. Process Res. Dev. 2007, 11, 899−902.
(21) Singh, B.; Bacon, E. R.; Lesher, G. Y.; Robinson, S.; Pennock, P.
O.; Bode, D. C.; Pagani, E. D.; Bentley, R. G.; Connell, M. J.; Hamel,
L. T.; Silver, P. J. Novel and Potent Adenosine 3′,5′-Cyclic Phosphate
Phosphodiesterase-III Inhibitors: Thiazolo[4,5-b][1,6]naphthyridin-2-
ones. J. Med. Chem. 1995, 38, 2546−2550.
(22) Kowalski, P. Reactions of 2-Aminopyridine with Benzyl-
ChlorideBenzylation of Pyridine Ring. Pol. J. Chem. 1984, 58,
959−960.
(23) 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.
(24) Nair, V.; Turner, G. A.; Buenger, G. S.; Chamberlain, S. D. New
Methodologies for the Synthesis of C-2 Functionalized Hypoxanthine
Nucleosides. J. Org. Chem. 1988, 53, 3051−3057.
(25) Jiang, X. H.; Song, L. D.; Long, Y. Q. Highly Efficient
Preparation of Aryl Beta-Diketo Acids with Tert-butyl Methyl Oxalate.
J. Org. Chem. 2003, 68, 7555−7558.
(26) Uchil, V.; Seo, B.; Nair, V. A Novel Strategy to Assemble the
Beta-Diketo Acid Pharmacophore of HIV Integrase Inhibitors on
Purine Nucleobase Scaffolds. J. Org. Chem. 2007, 72, 8577−8579.
(27) Kassahun, K.; McIntosh, I.; Cui, D.; Hreniuk, D.; Merschman,
S.; Lasseter, K.; Azrolan, N.; Iwamoto, M.; Wagner, J. A.; Wenning, L.
A. Metabolism and Disposition in Humans of Raltegravir (MK-0518),
an Anti-AIDS Drug Targeting the Human Immunodeficiency Virus 1
Integrase Enzyme. Drug Metab. Dispos. 2007, 35, 1657−1663.
(28) Ortiz de Montellano, P. R., Ed. Cytochrome P450: Structure,
Mechanism, and Biochemistry, 3rd ed.; Kluwer Academic/Plenum:
New York, 2005.
881
dx.doi.org/10.1021/ml2001246|ACS Med. Chem. Lett. 2011, 2, 877−881