Tricyclic HIV integrase inhibitors: VI. SAR studies of 'benzyl flipped' C3-substituted pyrroloquinolines

Article abstract of DOI:10.1016/j.bmcl.2008.12.079

A series of C3 halobenzyl-substituted tricyclic HIV integrase inhibitors was prepared. Improvement in cell-based inhibitor potency was observed in comparison to previously disclosed tricyclic pyrroloquinolines carrying the 'halobenzyl tail' at the lactam

Full text of DOI:10.1016/j.bmcl.2008.12.079

Bioorganic & Medicinal Chemistry Letters 19 (2009) 1187–1190
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Tricyclic HIV integrase inhibitors: VI. SAR studies of ‘benzyl flipped’
C3-substituted pyrroloquinolines
*
Sammy Metobo, Michael Mish , Haolun Jin, Salman Jabri, Rachael Lansdown, Xiaowu Chen,
Manuel Tsiang, Matthew Wright, Choung U. Kim
Gilead Sciences, Medicinal Chemistry, Inc. 333 Lakeside Drive, Foster City, CA 94404, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
A series of C3 halobenzyl-substituted tricyclic HIV integrase inhibitors was prepared. Improvement in
cell-based inhibitor potency was observed in comparison to previously disclosed tricyclic pyrroloquino-
lines carrying the ‘halobenzyl tail’ at the lactam nitrogen. Animal PK for several of the C3-substituted
inhibitors was examined, with a dihaloaryl analog achieving good balance in protein-shifted EC50 and
t1/2 in animal PK studies.
Received 20 November 2008
Accepted 18 December 2008
Available online 25 December 2008
Keywords:
AIDS
Ó 2008 Elsevier Ltd. All rights reserved.
HIV
Integase inhibitors
Tricyclic pyrroloquinoline
C3 Benzyl SAR
Introduction: The causative pathogen of AIDS, human immuno-
deficiency virus (HIV), requires three essential enzymes encoded
in the HIV pol gene for replication. While there are many licensed
therapeutics that target protease (PR) and reverse transcriptase
(RT), by comparison it has been only recently that promising
integrase (IN) inhibitors have been advanced through clinical
development.1
Previous reports from our group have examined the SAR of IN
inhibitors based on a pyrroloquinoline-containing tricyclic scaffold
such as 1 (Fig. 1).2 Here, we report the discovery of a next-genera-
tion class of analogs in the pyrroloquinoline series, shown gener-
ally as 2, where the benzyl group is appended to the pyridine C3
position. These new inhibitors display improved potency and
in vivo PK when compared to many of the previously disclosed
analogs from our research program.
The rationale to pursue this SAR was provided by a few key con-
siderations. Based on examination of our integrase/DNA active site
model, initially developed for earlier leads in this program,3 we
hypothesized that these ‘benzyl flipped’ analogs would possess
excellent inhibitor potency. Figure 2 presents an overlay of the pre-
viously reported analog 1, with a prototypical ‘flipped tail’ inhibitor
9. It can be seen that the integrase active site can accommodate the
new inhibitor 9 especially well if the scaffold binds in such a way
as to orient the benzyl tail into the ‘lipophilic pocket’ of the enzyme
that our model has postulated previously.
F
SO2CH3
N
X
N
"benzyl
flip"
Ar
Y
N
N
N
O
O
OH
OH
2
1
X = OCH3 or CH3NSO2CH3
Y = H or CH3
Figure 1. Tricyclic pyrroloquinoline inhibitor 1, and general ‘benzyl flipped’
inhibitor structure 2.
Additionally, protein-shifted inhibitor potency and metabolic
stability were recognized as essential considerations in lead iden-
tification. Evaluation of these new ‘benzyl flipped’ analogs was
undertaken as part of our ongoing effort to develop a new series
of potent IN inhibitors with excellent PK.
Synthesis, results and discussion: Our synthesis plan for the series
hinged on the use of a C3-substituted pyridine dielectrophile4 as
the reacting partner in a Dieckmann condensation to quickly build
up the tricyclic core of the scaffold. The synthesis of analog 8 is
shown in Scheme 1, where condensation between N-methyl suc-
cinimide and the pyridine dicarboxylate 3 is carried out to give
bis-phenol 4. Treatment with excess triisopropylsilyl (TIPS) chlo-
ride in the presence of TEA gives the intermediate bis-ether, which
then furnishes the mono-TIPS ether 6 upon reaction with an equiv-
alent of water added to the reaction mixture. Reduction of the
imide to the lactam with LiBH4, followed by alkylation of the free
* Corresponding author. Tel.: +1 650 522 5316; fax: +1 650 522 5169.
E-mail address: mmish@gilead.com (M. Mish).
0960-894X/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2008.12.079

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S. Metobo et al. / Bioorg. Med. Chem. Lett. 19 (2009) 1187–1190
yield intermediate 10. Further elaboration via lactam N-alkylation
with p-F benzyl bromide gave, after PMB removal, the bis-p-F ben-
zyl analog 11.
Synthesis of these first analogs in the ‘flipped-tail’ series was
followed by efforts that focused on two additional elements of
SAR. Primarily, we examined leads featuring a C5-N-methyl, N-me-
syl group (exemplified by previously reported compounds in the
program such as 1). The impact of a dihalogenated aryl ‘tail’ was
also of interest based on results from our original inhibitor series.
In the context of our earlier work, both lines of SAR were expected
to result in meaningful improvements in potency and PK behavior.
The synthesis of these latter analogs is detailed in Scheme 2.
Here, Dieckmann condensation of N-DMB-imide with a nitrile-es-
ter 12 led to aniline 14. Mesylation followed by reduction provided
lactam 18. Subsequent DMB removal, PMB recapping of the C8-
phenol, lactam N-methylation, and final deprotection gave analog
20. A similar synthetic sequence involving dihaloaryl ‘tail’-bear-
ing-pyridine 13 ultimately furnished analog 21.
Figure 2. Overlay of ‘flipped-tail’ compound 9 (white scaffold) onto 1 (orange
Comparison of strand transfer IC50 and antiviral EC50 values
(Table 1) for C5-methoxy analogs 8, 9 and 11 showed a trend
where a decrease in potency is seen upon increasing the size of
the lactam N-substituent.
Within this set, the most potent analog in the series is the free-
lactam compound 9. Despite the presence of two aryl ‘tails’ in
compound 11, a moderate level of both enzymatic and cell-based
antiviral activity was maintained. This observation suggests that
the presence of two extended lipophilic moieties on the pyrrolo-
quinoline core results in a comparatively poor inhibitor fit relative
scaffold) docked into integrase/DNA complex model.
phenol with MeI gave the final compound 8 after TIPS hydrolysis
employing TFA/TES in dichloromethane (DCM).
For the related analogs 9 and 11, the N-3,5-dimethoxybenzyl
(DMB) protected imide was utilized in the Dieckmann condensa-
tion. The synthetic sequence is analogous to the one previously de-
scribed, until unmasking of the DMB-lactam via heating in TFA/THF
(to furnish analog 9) is followed by recapping the C8-phenol to
OH
O
O
O
O
a
b
R
N
N
F
N
F
O
OH
O
3
4 R=CH3
5 R=DMB
OTIPS
O
OH
O
c
R
N
R
N
N
F
N
F
O
OTIPS
O
OTIPS
6 R=CH3
7 R=DMB
d
O
R
N
N
F
O
O
OH
8 R=CH3
9 R=H
H
N
e
N
F
O
OPMB
10
f
O
N
N
F
O
OH
11
F
Scheme 1. Reagents and conditions: (a) N-methyl succinimide, NaOMe, THF, reflux, 24 h, 66%; (b) TIPSCl (2 equiv), DMF, DIPEA; (c) i—H2O, 16 h, 84% across 2 steps; ii—LiBH4,
THF/MeOH, rt–60 °C, 76–85%; (d) i—MeI, NaH, 80–85%; ii—TFA/THF/H2O, 82–88%; (e) PMBBr, CS2CO3,DMF, 44%; (f) i—NaH, p-F benzylbromide, DMF, 65%; ii—TFA/TES/DCM,
82%.

S. Metobo et al. / Bioorg. Med. Chem. Lett. 19 (2009) 1187–1190
1189
NH2
O
NC
R
R
F
a
b
DMB
N
O
N
F
N
O
O
OH
14 R=H
15 R=Cl
12 R=H
13 R=Cl
NHMs
NHMs
O
R
F
R
F
c
DMB
N
DMB
N
N
N
O
O
OH
OTIPS
18 R=H
19 R=Cl
16 R=H
17 R=Cl
d
NMs
OH
R
F
N
N
O
20 R=H
21 R=Cl
Scheme 2. Reagents and conditions: (a) LiHMDS (2.5 equiv), THF, 0 oC–rt, 24 h, 50–86%; (b) i—TIPSCl (1.2 equiv), DMF, TEA, 65–75%; ii—MsCl, TEA (excess, then KOtBu), 60–
65%; (c) LiBH4, THF/MeOH, rt–80 oC; (d) i—MeI, Cs2CO3, 74–82% across 2 steps; ii–TFA/TES/DCM, rt to 70 oC, 90–94%; iii—PMBBr, Cs2CO3, DMF, 40–52%, (iv) MeI, NaH, DMF,
70–78% (v) TFA/TES/DCM, 80%.
Table 1
Table 2
In vitro data for C3-substituted tricyclic inhibitorsa
Comparison of PK results for new analogs 20 and 21 in rat and dog, presented
alongside PK data for clinically approved IN inhibitor raltegravir (RAL data is adapted
from Ref. 1)
Compound
IC50 (nM)
EC50, (10% FBS)c
EC50 (HSP)d
b
8
9
11
20
21
100
14
600
80
20
16
105
2
nd
27
nd
7
Compound
Rat
(h)
Dog
F (%)
T
CL (L/h/kg)
F (%)
T
(h)
½
CL (L/h/kg)
½
Raltegravir
20
21
45
12
11
2
0.4
1.7
2.4
0.49
0.22
69
47
60
11 (b T1/2
)
0.36
0.13
0.12
265
1.1
35
4.4
8.8
a
Values are means of at least two experiments, given in nM, nd, not determined.
b
c
Ref. 6a.
Ref. 6b.
F (%): fraction available to the general circulation upon oral dosing of test com-
pounds as compared to i.v. dosing, calculated based on AUC from i.v. and p.o.
groups, expressed as %; CL: total body clearance obtained from i.v. dosing groups.
All compounds were dosed as free parent in a solution form (EtOH, PG, PEG400; and
d
HSP, human serum proteins adjusted EC50, obtained by assaying compounds in
the presence of physiological concentrations of human serum albumin and AAG;
see Ref. 6c for details.
citric acid; pH 3.3 for iv and pH 2.2 for po); and T was generated from the i.v.
½
dosing group. The data represent the mean value obtained from three animals in
each study. Data adapted from Ref. 1.
to inhibitors carrying only one benzyl ‘tail’.5 On the other hand,
evaluation of the C5-aza substituted, ‘flipped-tail’ analogs showed,
most notably in the case of analog 20, significant improvement in
the EC50 and protein shifted EC50 values.
Conclusions: The synthetic work presented above enabled the
examination of a new series of pyrroloquinoline HIV integrase
inhibitors that moved the ‘benzyl tail’ portion of the inhibitor to
the C3-position of the inhibitor scaffold. This change to the tricyclic
scaffold was found to be well-tolerated with respect to the result-
ing potency of the compounds as measured by the integrase strand
transfer and tissue culture anti-HIV assays. PK evaluation of a sub-
set of these leads showed that fine-tuning of both the C5-substitu-
ent and the aryl portion of the ‘benzyl tail’ proved beneficial in
terms of identifying diahaloaryl ‘flipped tail’ analog 21. This lead
showed good oral bioavailability (F), low in vivo clearance, and
high antiviral activity. These results support possible further clini-
cal evaluation of this new series.
Of the compounds thus obtained, we chose to advance several
that showed compelling potency in the protein shifted assay to
evaluation in rat and dog PK studies (summarized in Table 2).
The resulting rat and dog PK for analogs 20 and 21 are presented
alongside the animal PK data reported for the clinically approved
IN inhibitor raltegravir (MK-0518).1 While the bioavailability of
this new tricyclic series is comparable to that of the reference com-
pound, the clearance of 20 was seen to be significantly lower than
that of raltegravir in both species. Further gains in the DMPK prop-
erties of the ‘benzyl flipped’ pyrroloquinoline leads were seen with
the dihaloaryl-tail compound 21. Specifically, the in vivo clearance
was further driven down in rat as compared to the 4-fluorobenzyl
analog 20. Overall, 21 showed improvement in terms of t1/2 in both
rat and dog, while at the same time showing protein-shifted po-
tency that was only moderately attenuated when compared to
the related compound 20.
Acknowledgments
The authors are grateful to Xubin Zheng and Bing Lu for analyz-
ing samples in PK studies; Vahid Zia and co-workers in the formu-
lation group for supporting the PK studies; Gregg Jones and Fang
Yu for carrying out biological assays.

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S. Metobo et al. / Bioorg. Med. Chem. Lett. 19 (2009) 1187–1190
substituent of 11 is replaced with a proton, illustrates a large decrease in the
References and notes
potency of 11 compared to either of these nearly identical analogs that carry
only a single benzyl tail group.
6. (a) Strand transfer assay modified from
1. To date, only a single HIV integrase inhibitor, MK-0518 (Raltegravir), has gained
FDA approval for marketing in the US, while many others continue to advance
through late-stage human clinical trials. See: Summa, V.; Petrocchi, A.; Bonelli,
F.; Benedetta Crescenzi, B.; Donghi, M.; Ferrara, M.; Fiore, F.; Gardelli, C.;
Gonzalez Paz, O.; 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.; Michael Rowley, M. J. Med. Chem. 2008, 513, 5843.
2. (a) Jin, H.; Cai, R. Z.; Schacherer, L.; Jabri, S.; Tsiang, M.; Fardis, M.; Chen, X.;
Chen, J. M.; Kim, C. U. Bioorg. Med. Chem. Lett. 2006, 16, 3989. and the references
cited within; (b) Fardis, M.; Jin, H.; Jabri, S.; Cai, R. Z.; Mish, M.; Tsiang, M.; Kim,
C. U. Bioorg. Med. Chem. Lett. 2006, 16, 4031; (c) Metobo, S.; Jin, H.; Tsiang, M.;
Kim, C. U. Bioorg. Med. Chem. Lett. 2006, 16, 3985; (d) Jin, H.; Wright, M.; Pastor,
R.; Mish, M.; Metobo, S.; Jabri, S.; Lansdown, R.; Cai, R. Z.; Pyun, P.; Tsiang, M.;
Chen, X.; Kim, C. U. Bioorg. Med. Chem. Lett. 2008, 16, 3989.
3. Chen, X.; Tsiang, M.; Yu, F.; Hung, M.; Jones, G. S.; Zeynalzadegan, A.; Qi, X.; Jin,
H.; Kim, C. U.; Swaminathan, S.; Chen, J. M. J. Mol. Biol. 2008, 380, 504. and
references cited therein.
4. The preparation of pyridine dielectrophile 3 has been reported in Johns, B.; Boros,
E.; Kawasuji, T.;Koble, C. S.; Kurose, N.;Murai, H.;Sherrill, R.;Weatherhead, J. W.O.
Patent 2005/077050(A2), 2005, while the preparation of pyridines 12 and 13 is
based on chemistry reported in Cai, Z.; Jabri, S.; Jin, H.; Lansdown, R.; Metobo, S.;
Mish, M.; Pastor, R. W.O. Patent 2007/136714(A2), 2007.
a previous report (Hazuda et al.,
Nucleic Acid Res. 1994, 22, 1121). Biotinylated donor DNA was bound to
Reacti-Bind High Binding Capacity Streptavidin coated white plates. DIG-
tagged target DNA with anti-DIG antibody-conjugated horse radish
peroxidase detection was used. (b) For antiviral assay, 50 ll of 2Â test
concentration of 5-fold serially diluted drug in culture medium were added to
each well of a 96-well plate (9 concentrations) in triplicate. MT-2 cells were
infected with HIV-1 IIIB at an m.o.i. of 0.01 for 3 h. Fifty microliters of
infected cell suspension in culture medium (ꢀ1.5 Â 104 cells) were then
added to each well containing the drug dilutions. The plates are incubated at
37 °C for 5 days. One hundred microliters of CellTiter-GloTM Reagent (catalog
# G7571, Promega Biosciences, Inc., Madison, WI) were then added to each
well. Cell lysis was allowed to complete by incubating at room temperature
for 10 min. Chemiluminescence was then read. For the cytotoxicity assay, the
protocol is identical to that of the antiviral assay, except that uninfected cells
and
a
3-fold
serial
dilution
of
drugs
were
used.
(c) The effect of compounds binding to serum protein components was
evaluated by determining the antiviral EC50 in MT-2 cells in 10% FBS in the
presence or absence of serum concentrations of HSA (35 mg/ml) or
(1.5 mg/ml). From the EC50 data in the presence of each individual protein,
the EC50 resulting from the combined effect of both proteins (as in serum)
can be calculated. The derivation of the appropriate equation for this
calculation can be made through competitive binding assumptions.
a1-AGP
5. Consideration of the ST IC50 of compound 11 alongside either 9, or a related
inhibitor originally reported in Ref. 2a wherein the bulky C3-p-fluorobenzyl