Inhibitors of HIV-1 attachment: The discovery and structure-activity relationships of tetrahydroisoquinolines as replacements for the piperazine benzamide in the 3-glyoxylyl 6-azaindole pharmacophore

Article abstract of DOI:10.1016/j.bmcl.2015.11.009

6,6-Fused ring systems including tetrahydroisoquinolines and tetrahydropyrido[3,4-d]pyrimidines have been explored as possible replacements for the piperazine benzamide portion of the HIV-1 attachment inhibitor BMS-663068. In initial studies, the tetrahydroisoquinoline compounds demonstrate sub-nanomolar activity in a HIV-1 pseudotype viral infection assay used as the initial screen for inhibitory activity. Analysis of SARs and approaches to optimization for an improved drug-like profile are examined herein.

Full text of DOI:10.1016/j.bmcl.2015.11.009

Bioorganic & Medicinal Chemistry Letters xxx (2015) xxx–xxx
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Inhibitors of HIV-1 attachment: The discovery and structure–activity
relationships of tetrahydroisoquinolines as replacements for the
piperazine benzamide in the 3-glyoxylyl 6-azaindole pharmacophore
a
a
a
a
b
Jacob J. Swidorski a, , Zheng Liu , Zhiwei Yin , Tao Wang , David J. Carini , Sandhya Rahematpura ,
Ming Zheng b, Kim Johnson b, Sharon Zhang c, Pin-Fang Lin c, Dawn D. Parker b, Wenying Li b,
Nicholas A. Meanwell a, Lawrence G. Hamann a,y, Alicia Regueiro-Ren a
⇑
a Department of Discovery Chemistry, Bristol-Myers Squibb Research and Development, 5 Research Parkway, Wallingford, CT 06492, USA
b Department of Pharmaceutical Candidate Optimization, Bristol-Myers Squibb Research and Development, 5 Research Parkway, Wallingford, CT 06492, USA
c Department of Virology, Bristol-Myers Squibb Research and Development, 5 Research Parkway, Wallingford, CT 06492, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
6,6-Fused ring systems including tetrahydroisoquinolines and tetrahydropyrido[3,4-d]pyrimidines have
been explored as possible replacements for the piperazine benzamide portion of the HIV-1 attachment inhi-
bitor BMS-663068. In initial studies, the tetrahydroisoquinoline compounds demonstrate sub-nanomolar
activity in a HIV-1 pseudotype viral infection assay used as the initial screen for inhibitory activity. Analysis
of SARs and approaches to optimization for an improved drug-like profile are examined herein.
Received 24 August 2015
Revised 2 November 2015
Accepted 4 November 2015
Available online xxxx
Keywords:
HIV-1
Ó 2015 Elsevier Ltd. All rights reserved.
Entry
Attachment
Tetrahydroisoquinoline
Tetrahydropyrido[3,4-d]pyrimidines
BMS-663068
O
O
OMe
O
O
N
R
OMe
N
N
X
N
O
N
H
N
N
N
N
Y
H
N
N
N
Z
N
1
BMS-626529
X, Y, Z = C, N; R = H, OH, alkyl
Despite the considerable advances in the treatment of HIV-1, in
recent years HIV-1 drug resistance has become of increasing
concern with antiretroviral (ARV) naïve patients infected with drug
resistant virus at a rate of 10–17% in developed countries.1 In addi-
tion to treatment-naïve patients with pre-existing virus mutations,
acquired resistances are still emerging after treatment with the
current standard of care regimens. Although rates of resistance
development have slowed as combination antiretroviral therapy
(cART) agents have been developed,2 there is still a need for new
ARV agents to treat patients who have developed resistance to
current therapies, particularly the highly treatment experienced
group. New ARV agents with mechanistically distinct modes of
action that can offer complementary resistance profiles to
marketed drugs will be essential to treat patients with resistant
HIV-1infections.3 HIV-1 attachment inhibitors (AIs) provide a
differentiated target from the core cART agents currently being
⇑
Corresponding author.
y
Current address: Novartis Institutes for BioMedical Research, 250 Massachusetts
Ave., Cambridge, MA 02139, USA.
http://dx.doi.org/10.1016/j.bmcl.2015.11.009
0960-894X/Ó 2015 Elsevier Ltd. All rights reserved.
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2
J. J. Swidorski et al. / Bioorg. Med. Chem. Lett. xxx (2015) xxx–xxx
O
O
O
O
OMe
OMe
OH
N
b
or
c
5
6
HN
a
N
N
N
N
H
H
N
N
N
N
Br
Br
2
3
4
N
N
O
N
O
O
O
OMe
OMe
N
N
N
N
N
H
H
N
N
N
N
N
N
N
5
6
Scheme 1. Synthesis of 3-glyoxylyl azaindole phenyl and pyridyl tetrahydroisoquinolines. Reagents and conditions: (a) TBTU, DIEA, DMF, rt, 71 h, 88%; (b) 2-tri-n-
butylstannylpyridine, Pd(PPh3)4, 1,4-dioxane, 100 °C, 4 days, 26%; (c) phenylboronic acid, PdCl2(dppf)-CH2Cl2 adduct, K2CO3, 1,4-dioxane/water (4:1), 85 °C, 2 h, 48%.
Table 1
Antiviral activity, cytotoxicity, in vitro metabolic stability, solubility properties and c Log P data for tetrahydroisoquinoline-based HIV-1 attachment inhibitors
HLM% rem at 10 minc
RLM% rem at 10 minc
Solubility pH 6.5d
(lg/mL)
c Log Pe
a
b
Compound
EC50 (nM)
CC50
(
lM)
1
5
6
0.10
0.09
0.02
>300
>33.3
>11.1
97
11
10
96
8
5
22 (c, pH 5.7)
19 (c)
<1 (a)
0.86
2.26
3.54
a
HIV-1 pseudotype assay performed at least in duplicate (averaged) utilizing HeLa CD4 CCR5 cells and HIV-1 JRFL virus envelope (Ref. 5). Experimental details are reported
in the Supporting information.
b
Concentration of drug associated with a 50% reduction in HeLa cell viability in the absence of pseudotype virus performed at least in duplicate (averaged) and in parallel
with the pseudotype assay (Ref. 5).
c
Metabolic stability experimental determination is detailed in the Supporting information. % Remaining at 10 min was extrapolated into low clearance (>85% remaining at
10 min), medium clearance (61–84% remaining at 10 min) and high clearance (<61% remaining at 10 min). Reference agents are referred to in the Supporting information.
d
Solubility of crystalline (c) or amorphous (a) material at pH = 6.5 unless stated otherwise stated.
c Log P values were calculated using Cambridgesoft ChemBioDraw version 12.0.
e
prescribed.4 Attachment inhibitors have been shown to interfere
with the first step of the HIV-1 entry process by binding to the viral
TFA
Boc
SnBu3
N
H2N
Boc
glycoprotein (gp120) and stabilizing
a
conformation not
N
a
b
recognized by CD4.5 Attachment of gp120 to the cellular CD4
receptor is disrupted by the envelope changes and, consequently,
the downstream processes which allow for viral genetic material
to be released into the host cell cytosol are interfered with, hinder-
ing viral replication.6–8
N
+
N
N
Br
7
8
9
10
In previous communications, we have detailed chemical
approaches to HIV-1 AIs that developed structure–activity
relationships (SARs) and which delivered several compounds into
clinical trials.9–21 Recently we have detailed the discovery of 1 as
a potent HIV-1 attachment inhibitor and the clinical form of the
molecule, a phosphate prodrug, BMS-663068.22,23 The phosphate
prodrug of 1 has progressed in the clinic through Phase IIb studies
with encouraging results that support its continued development
in Phase III trials.24 In humans, benzamide hydrolysis was observed
as a metabolic pathway, which spurred interest in preparing non-
amides so we analyzed a full range of structural modifications in
pursuit of a differentiated clinical candidate. Previous efforts com-
municated by our group12 and, more recently, Tuyishime et al.25,26
have shown that specific modifications to the piperazinamide moi-
ety could afford compounds with excellent antiviral potency; how-
ever, all of these reports described amide derivatives that could
potentially be hydrolyzed through similar metabolic processes.
Alternatively, we found the tetrahydroisoquinoline (THIQ) to have
a topology that could position an aromatic group in a similar posi-
tion to the phenyl group of the benzamide of HIV AIs similar to 1.
We envisioned functionalization of either the THIQ or the core to
improve solubility and perhaps preclude the use of a prodrug. THIQ
Scheme 2. Synthesis of pyridyl tetrahydroisoquinoline 10, TFA salt. Reagents and
conditions: (a) Pd(PPh3)4, 1,4-dioxane, 110 °C, 65 h, 91%; (b) TFA, CH2Cl2, rt, 6 h,
100%.
6 (EC50 = 0.02 nM) showed an improvement in potency in the
in vitro HIV-1 pseudotype assay used as the initial screen for inhi-
bitory activity in the program compared to the clinical candidate 1,
EC50 = 0.10 nM. In addition, the THIQ was an attractive motif
because it allowed for modifications at several positions which
could potentially modulate lipophilicity of the molecule and/or
influence solubility, permeability and metabolic stability of the
parent compound.27
Taking into account prior SARs, our initial molecules incorpo-
rated the 3-methyl-1,2,4-triazole at the 7-position of the 3-glyoxy-
lyl azaindole core. The synthetic approach to construct the 3-
glyoxylyl azaindole phenyl and pyridyl THIQs is illustrated in
Scheme 1. With 2 in hand from previous efforts,23 the amide was
constructed utilizing an O-(benzotriazol-1-yl)-N,N,N0,N0-tetram-
ethyluronium tetrafluoroborate (TBTU)-mediated coupling with 3
to give the 5-bromo THIQ 4. Installation of the pyridyl moiety
was accomplished using Stille conditions to attain 5, while the
phenyl group was installed via a Suzuki coupling with the phenyl
boronic acid to give 6.
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J. J. Swidorski et al. / Bioorg. Med. Chem. Lett. xxx (2015) xxx–xxx
3
O
O
O
O
O
O
OMe
OMe
OMe
Br
OH
HN
N
N
a
b, c, d, or e
+
N
N
N
N
H
N
N
H
H
Br
R
N
11
12
N
N
10
b
c
d
e
N
N
X
X
R =
N
NH
O
NH
Z
N
N
N,C
N,C
EtOOC
Y
Y
f
N
H
N
N
H
O
g
h
N
N
N
H
N
H
COOH
CONR1R2
Scheme 3. Synthesis of C-7 modified 3-glyoxylyl azaindole pyridyl tetrahydroisoquinolines. Reagents and conditions: (a) TBTU, DIEA, DMF, rt, 70 h, 75%; (b) 3-
(tributylstannyl)-1H-pyrazole-5-carboxylate, Pd(PPh3)4, 1,4-dioxane, 110 °C, 17 h, 30%; (c) 14, 17, and 18; heterocycle, copper (0), pyridine, 145 °C; (d) compounds 15, 16, 19,
and 20; heterocycle, Cu(I)iodide, K2CO3, (1R,2S)-N1,N2-dimethylcyclohexane-1,2-diamine, 1,4-dioxane, 100 °C; (e) compounds 26 and 27; amine, CO (80 psi), Pd(PPh3)4, water,
triethylamine, 80 °C, 16 h; (f) compound 23; methylamine (2 M in THF), 40–60 °C, 6.5 days; (g) compound 22; methanol, water, K2CO3, 60 °C, 2 h; (h) compounds 24 and 25;
amine, TBTU, DIEA, DMF, rt.
Compounds 5 (EC50 = 0.09 nM) and 6 (EC50 = 0.02 nM) were
examined in the pseudotype assay and both showed improved
potency when compared to 1 (EC50 = 0.10 nM). Additionally, 5
ester with methylamine (23), hydrolysis (22), or hydrolysis
followed by TBTU-mediated coupling of the free carboxylic acid
(24 and 25).
demonstrated improved solubility (19
over 6 (<1 g/mL as an amorphous solid) and similar solubility to 1
(22 g/mL at pH 5.7 as a crystalline solid). Unfortunately, both 5
l
g/mL as a crystalline solid)
Substitution of the C-7 position of the azaindole (Table 2) with a
1,2,3 triazole, as in 13, did not improve the in vitro metabolic sta-
bility or aqueous solubility compared to the prototype 5. Examples
14–18 explored variation of the lipophilicity by changing the sub-
stituents attached to the triazole. The more lipophilic compounds
15 and 16 showed very poor solubility along with a poor in vitro
metabolic profile. The less lipophilic compounds 14 and 18 exhib-
ited better solubility and slightly improved metabolic stability.
Consistent with previous SAR, the unsubstituted 1,2,4-triazole 14
lost potency. The methoxymethyl-substituted triazole 17 did not
benefit from the decreased lipophilicity as HLM and RLM stability
for this compound was still very poor; however, solubility was
slightly improved. Substitution at the 5-position of the triazole in
18 did improve metabolic stability slightly, but potency decreased
dramatically due to disruption in the planarity of the heterocyclic
ring and the azaindole.20 The N-linked pyrazoles 19 and 20 also
probed variation of lipophilicity but failed to show an improve-
ment in metabolic stability. Variations of C-linked pyrazoles were
examined in the context of compounds 21–25. Although improved
compared to 5, the in vitro metabolic stability of 21 (18/17% parent
remaining after a 10 min incubation with microsomes) was still
too low to progress into more advanced studies. Carboxylic acid
22 showed a great improvement in metabolic stability (95/91%
parent remaining after 10 min incubation with microsomes), but
the solubility of the compound was still less than ideal. Methyl
amide 23 showed a slight loss of potency and solubility while
amides 24 and 25 had solubility so low that the metabolic stability
wasn’t determined. The series of C-7 substitutions concluded with
the two C-7 amides 26 and 27. While HLM stability improved
slightly, and methyl amide 26 did show an improvement in
l
l
and 6 were quickly metabolized upon incubation with human or
rat liver microsomes, with only 5–11% of parent drug remaining
after the 10 min incubation period (Table 1). We have shown pre-
viously that solubility and metabolic stability, as well as potency
can be affected by the substituent in the 7-position of the azaindole
moiety.20 Therefore, we designed a series of compounds that
would vary the lipophilicity of the molecule through modifications
of the substituent at the C-7 position of the 6-azaindole ring.
Earlier SAR developed in the program led us to focus on introduc-
ing triazoles and pyrazoles preferentially bearing polar sub-
stituents, as well as direct attachment of amides to the azaindole
core. Because the initial assembly of 5 and 6 installed the THIQ late
in the synthesis, an alternative approach was developed to enable
C-7 substitution late in the synthetic sequence. The 5-pyridyl THIQ
9 was assembled via a Stille coupling of 7 with 8 (Scheme 2). The
Boc group was removed by treatment with TFA in dichloromethane
to give 10. Amide formation was accomplished through a TBTU-
mediated coupling of 10 with 1128 to yield the C-7 azaindole bro-
mide 12 that proved to be a useful intermediate for subsequent
modifications. The brominated core could be elaborated using Ull-
man conditions to introduce N-linked heterocycles (13–20), Stille
conditions for the C-linked pyrazoles 21–25, or carbonylation con-
ditions to prepare the C-7 amides 26 and 27 (Scheme 3). The C-7
pyrazole 21 was formed by elaborating the pyrazole prior to the
Stille coupling by reducing the ethyl ester of 3-(tributylstannyl)-
1H-pyrazole-5-carboxylate with LAH. The other C-linked pyrazoles
were further modified post-coupling through amination of the
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4
J. J. Swidorski et al. / Bioorg. Med. Chem. Lett. xxx (2015) xxx–xxx
Table 2
Antiviral activity, cytotoxicity, in vitro metabolic stability and calculated Log P data for C-7 substituted azaindole HIV-1 attachment inhibitors
O
O
OMe
N
N
N
H
R
N
Compound
R
EC50 (nM)
0.06
CC50
37.5
(lM)
HLM/RLM% remaining at 10 min
7/0
Solubility*
18
(l
g/mL)
c Log P
N
13
3.14
N
N
N
N
14
15
0.48
0.14
162
9/20
7/8
30
<1
1.99
3.01
N
N
N
N
>3.7
CF3
N
N
16
0.14
>11.1
7/5
3
2.71
N
CF2CH3
N
N
17
18
19
0.54
53.4
0.41
>100
119
5/9
27
37
<1
1.79
1.79
4.28
N
CH2OMe
CH2OMe
N
29/33
1/1
N
N
N
N
>1.23
CF3
N
N
20
21
22
23
24
0.13
0.08
0.15
0.14
0.37
>33.3
190
3/1
ND
20
7
2.31
2.51
3.77
3.42
2.57
CH2OH
N
18/17
95/91
6/6
HN
CH2OH
N
HN
>300
28.4
COOH
N
8
HN
CONHMe
CONMe2
N
>33.3
ND
2
HN
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J. J. Swidorski et al. / Bioorg. Med. Chem. Lett. xxx (2015) xxx–xxx
5
Table 2 (continued)
Compound
R
EC50 (nM)
0.10
CC50
(lM)
HLM/RLM% remaining at 10 min
ND
Solubility*
(
lg/mL)
c Log P
N
25
>33.3
1
2.86
HN
CONHC2H4OH
26
0.62
0.44
>100
20/3
33
4
2.74
2.18
CONHMe
CONHCH2CH2OH
27
>33.3
24/28
*
Solubility of amorphous solids measured at pH = 6.5.
O
O
O
O
Bn
Bn
HN
HN
N
N
a
b
c
d
28
29
30
31
OH
OTBDMS
O
OTBDMS
OH
O
SnBu3 Bn
N
HN
HN
Bn
N
e
f, g
+
+
OTf
32
33
34
35
36
Scheme 4. Synthesis of 1- and 1,10-modified tetrahydroisoquinolines. Reagents and conditions: (a) TBDMS-Cl, 1H-imidazole, DMAP, DMF, rt, 22.5 h, 97% (b) NaHMDS, benzyl
bromide, THF, rt, 41 h; (c) TBAF (1 M in THF), THF, rt, 45 min; (d) Tf2O, pyridine, CH2Cl2, 0 °C-rt, 1 h, 11% over three steps; (e) tri-2-furylphosphine, LiCl, Pd2(dba)3, NMP, 85 °C,
15.5 h, 77%; (f) methyltitanium triisopropoxide (1 M in THF), EtMgBr (1 M in THF), THF, ꢀ78 °C to 50 °C, 23 h, 23%; (g) 10% Pd/C, MeOH, H2 (1 atm), 4.5 h, 1:0.57 ratio based on
1H NMR integration, 100%.
solubility, both compounds showed diminished antiviral potency.
Although this series of compounds did not improve metabolic sta-
bility as much as initially hoped, there seemed to be a general
trend that compounds with lower c Log P values had improved
metabolic stability. The only compound with a major improvement
in metabolic stability, acid 22, lacked the potency of the best com-
pounds and also had inadequate passive membrane permeability
with PAMPA assay values of 0 nm/s at pH = 7.4 and 32 nm/s at
pH = 5.5. Based on these results, we decided to pursue in vitro bio-
transformation studies with 5 in an effort to determine where the
metabolic soft spots in the molecule were located (see Fig. 1).
In vitro biotransformation studies with 5 and several related
analogs identified the benzylic position adjacent to the N atom of
the THIQ as the labile metabolic site for this chemotype. The hemi-
aminal 37 was the only metabolite produced after incubating 5
with human liver microsomes. In order to obtain a sufficient
amount of 37 to test for biological activity and metabolic profiling,
the biotransformation of 5 into 37 was accomplished preparatively
using a liver S9 fraction from Aroclor 1254-induced rats incubated
with compound at 37 °C for 4.5 h.29 Hemiaminal 37 not only had a
lower calculated c Log P than the parent (1.6 vs 2.26, Table 3), but
both metabolic stability and solubility were greatly improved.
Unfortunately, however, there was more than a 50-fold decrease
in antiviral potency compared with 5.
In an attempt to improve the metabolic stability by blocking the
benzylic position, cyclopropyl analog 38 was prepared. Preparation
of the substituted THIQ was accomplished by protecting the pheno-
lic alcohol of the commercially available 5-hydroxy-3,4-dihydroiso-
quinolin-1(2H)-one (28) as the TBDMS ether followed by benzyl
protection of the amide to give 30 (Scheme 4). The TBDMS ether
was removed with TBAF and the phenol was converted into triflate
32. Modified Stille conditions30 allowed for efficient cross-coupling
to install the phenyl ring. The benzyl-protected amide was then
converted to the aminocylopropane 35 utilizing conditions devel-
oped by Kulinkovich and de Meijere followed by hydrogenolysis
of the benzyl protecting group.31 During the formation of the
aminocylopropane 35, the amino ethyl analog 36 was also formed.
Since separation of these two compounds was difficult, the mixture
of 35 and 36 was directly coupled to core 2 using a TBTU-mediated
coupling and the fully elaborated products 38 and 39 were separa-
ble by preparative HPLC. While the stability of 38 in HLM was only
marginally better than 6, potency declined by 460-fold. Ethyl
derivative 39, which could in principle deter metabolism by
increasing steric hindrance around the labile site, did not offer
492.18
N
464.18
C27H22O3N7
C26H22O2N7
N
O
O
OH
209.11
-H2O C14H13N2
O
N
N
H
N
N
N
256.08
C12H10O2N5
+H2O
274.09
C12H12O3N5
Figure 1. In vitro biotransformation analysis of 5.
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6
J. J. Swidorski et al. / Bioorg. Med. Chem. Lett. xxx (2015) xxx–xxx
Table 3
Antiviral activity, cytotoxicity, in vitro metabolic stability and calculated Log P data for modifications to the 1-position of the THIQ ring
O
O
R1
OMe
R2
N
N
N
H
N
N
Z
N
Compound
R1
H
R2
Z
EC50 (nM)
CC50
(lM)
HLM/RLM% remaining at 10 min
Solubility (
lg/mL)
c Log P
37
38
39
OH
N
CH
CH
4.62
9.20
0.71
>100
5.45
24.7
79/72
30/6
21/5
70 (c)
<1 (a)
<1 (a)
1.60
3.86
4.59
Cyclopropyl
Ethyl
H
HCl
SnBu3
R
R
Bn
N
O
Bn
N
Bn
O
HN
R
H2N
N
N
N
a
b
+
N
NH
N
CO2Et
Cl
R = H
50 R = Me
40
41
45
49
8
R = H
42 R = Me
43
R = H
46 R = Me
47
51
R = c-Pr
44 R = NMe2
R = c-Pr
48 R = NMe2
R = c-Pr
52 R = NMe2
R
R
Bn
N
N
N
HN
c
d
N
N
N
N
53
57
R = H
54 R = Me
55
R = H
58 R = Me
59
R = c-Pr
56 R = NMe2
R = c-Pr
60 R = NMe2
Scheme 5. Synthesis of 2-substituted-4-(pyridin-2-yl)-5,6,7,8-tetrahydropyrido[3,4-d]pyrimidines. Reagents and conditions: (a) NaOMe, MeOH, rt; (b) POCl3, DIEA, toluene,
reflux; (c) Palladium(II)acetate, tri-2-furylphosphine, 1,4-dioxane, reflux; (d) DIEA, DCM, 1-chloroethyl chloroformate, 0 °C, 1 h then rt, isolated HCl salts.
H
N
HCl
SnBu3
Bn
O
Bn
N
Cl
Bn
Bn
O
O
N
N
N
N
c
a
b
+
N
NH2
61
NH
N
H2N
CO2Et
O
Cl
40
62
63
8
N
N
Cl
H
H
N
Bn
N
N
N
N
N
HN
N
d
e
N
N
HCl
N
64
65
66
N
N
Scheme 6. Synthesis of 2-amino-substituted 4-(pyridin-2-yl)-5,6,7,8-tetrahydropyrido[3,4-d]pyrimidines. Reagents and conditions: (a) sodium methoxide, MeOH, reflux
18 h, 58%; (b) POCl3, reflux, 18 h, 11.5%, (c) palladium(II)acetate, tri-2-furylphosphine, 1,4-dioxane, reflux, 5 h, 71%; (d) N,N-dimethylethylenediamine, acetonitrile, reflux,
26 h, 68%; (e) 20% palladium hydroxide on carbon, 1 N HCl, methanol, H2 (1 atm), rt, 5 days, 100%.
any advantage in terms of improving metabolic stability. Addition-
ally, both 38 and 39 showed elevation in cytotoxicity (5.45 M for
38 and 24.7 M for 39) in the assay. These compounds showed that
even small structural changes at the 1-position of the THIQ ring can
cause a dramatic loss in potency and that metabolism in these
molecules can shift to other positions if a single soft spot is blocked.
Only two compounds, 22 and 37, showed a significantly
improved metabolic profile. Both of these compounds contained
polar groups, with 22 containing a free carboxylic acid on the
pyrazole at C-7 while 37 is hydroxylated at the 1-position of the
THIQ ring. With these data in hand, we decided to pursue a series
of compounds that would increase the polarity of the THIQ ring. Cal-
culated c Log P values show a considerable decrease in lipophilicity
when the THIQ ring is replaced by a tetrahydropyrido[3,4-d]pyrim-
idine heterocycle. Further adjustments in lipophilicity could then be
realized via substitution at the 2-position of the bicyclic ring.
The synthesis of the bicyclic ring system is illustrated in
Scheme 5.32 Analogs 57–60 were constructed through cyclization
l
l
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J. J. Swidorski et al. / Bioorg. Med. Chem. Lett. xxx (2015) xxx–xxx
7
Table 4
Antiviral activity, cytotoxicity, in vitro metabolic stability and calculated Log P data for tetrahydropyrido[3,4-d]pyrimidine modifications
O
O
OMe
N
N
R
N
N
H
N
N
N
N
N
Compound
R
EC50 (nM)
CC50
(
lM)
HLM/RLM% remaining at 10 min
Solubility (lg/mL)
c Log P
67
68
69
70
71
H
Me
0.17
0.42
0.02
0.03
69.2
>33.3
>11.1
>3.7
>3.7
52.8
79/81
ND
ND
ND
ND
3 (a)
3 (a)
<1 (a)
<1 (a)
267 (c)
0.12
0.62
1.07
1.04
1.17
Cyclopropyl
NMe2
NHCH2CH2NMe2
with commercially-available 40 and the substituted amidine or
guanidine 41–44 to provide the pyrimidinones 45–48. The pyrim-
idinone intermediate was then treated with POCl3 to give the chlo-
rides 49–52. Stille coupling with 3-(tributylstannyl)pyridine
followed by deprotection of the amine gave the elaborated tetrahy-
dropyrido[3,4-d]pyrimidines 57–60 as their respective HCl salts.
Alternatively, the bicyclic ring was constructed by reacting 40 with
urea to give the tetrahydropyridopyrimidinedione 62 (Scheme 6).
The dione 62 was converted to dichloride 63 through treatment
with POCl3. Stille coupling was accomplished selectively at the
4-position which then allowed substitution of the remaining
chloride with N,N-dimethylethylenediamine to give the fully
elaborated tetrahydropyrido[3,4-d]pyrimidine 65. Deprotection
via hydrogenolysis gave 66 as the HCl salt. The final deprotected
amines 57–60 and 66 were individually coupled with 2 using TBTU
to give 67–71.
action will be a critical component of new HIV-1 treatments.
HIV-1 attachment inhibitors show a differentiated mode of action
from the current standard of care, and could be a component, espe-
cially in the population of highly treatment-experienced patients
where drug resistance is more prevalent. In this communication
we have examined replacements for the piperazine benzamide
moiety, a critical structural component in HIV-1 attachment inhi-
bitors. With favorable in vitro antiviral potency, the THIQ ring sys-
tem was a promising starting point for modifications that could
potentially allow for a low dose formulation without the need for
a prodrug. THIQ analogs with modified heterocycles at the C-7
position of the azaindole varied the lipophilicity of the molecule
but were unable to achieve the desired improvements to aqueous
solubility and had only modest improvements in metabolic stabil-
ity. Modification of the THIQ ring aimed at sterically blocking
metabolism only slightly improved the in vitro metabolic profile
of the series as a consequence of redirected metabolism but did
not improve aqueous solubility. Introducing heteroatoms into the
THIQ heterocycle led us to explore the tetrahydropyrido[3,4-d]
pyrimidine series which lowered the c Log P of the molecule sub-
stantially. This series retained targeted antiviral potency and 67
showed a remarkable improvement in metabolic stability. How-
ever, further profiling of 67 in vivo revealed properties inferior to
1. With these studies, we have demonstrated that a series of 6,6
The unsubstituted analog in the tetrahydropyrido[3,4-d]pyrim-
idine series, 67, showed promise as the antiviral potency was
retained (EC50 = 0.17 nM) and the metabolic profile was improved
considerably (79/81% parent remaining after a 10 min incubation
with microsomes), albeit with a decrease in solubility (3 lg/mL)
when compared to 5 (Table 4). The lower c Log P of 67 (0.12) seems
to correlate with an improvement in free fraction when compared
with 5 (8% vs 2%). Methyl substitution at the 2-position of the
tetrahydropyrido[3,4-d]pyrimidine ring in 68 showed lower
antiviral potency (EC50 = 0.42 nM) while substitution with cyclo-
propyl (69) or dimethylamino (70) groups increased potency
(EC50 = 0.02 and 0.03 nM, respectively) but showed even less desir-
bicyclic systems is
a suitable isosteric replacement for the
piperazine benzamide moiety in HIV-1 attachment inhibitors.
Ultimately, the superior overall profile of the clinical candidate 1
precluded any further development of these bicyclic analogs.
able aqueous solubility (<1 lg/mL). Large polar substituents such
as in 71 exhibited reduced antiviral activity (EC50 = 69 nM)
Acknowledgements
associated with a significant improvement in aqueous solubility
(267
l
g/mL).
The authors gratefully acknowledge Dr. John F. Kadow for his
analysis and review of the manuscript and Dr. Stella Huang for
her support of spectroscopic data.
Further profiling of 67 showed adequate passive membrane
permeability with PAMPA assay values of 294 nm/s at pH = 7.4
and 244 nm/s at pH = 5.5;33 consequently, this compound was
advanced into rat PK studies. IV plasma clearance and the volume
of distribution of 67 were 5.2 mL/min/kg (indicating low clearance)
and 0.33 L/kg, respectively, after dosing at 1 mg/kg. The resulting
mean terminal half life was 1.1 h and 67 had good oral exposure,
Supplementary data
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.bmcl.2015.11.
009.
with AUClast = 41 lM * h and Cmax = 10.0 lM after dosing at
5 mg/kg using a poly(ethylene glycol) 400 (PEG 400) and ethanol
(90:10 v/v) vehicle. Overall, the IV and PO profile is very promising
although inferior to that of 1 which exhibited a t1/2 = 5.9 h, an
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Please cite this article in press as: Swidorski, J. J.; et al. Bioorg. Med. Chem. Lett. (2015), http://dx.doi.org/10.1016/j.bmcl.2015.11.009

8
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Please cite this article in press as: Swidorski, J. J.; et al. Bioorg. Med. Chem. Lett. (2015), http://dx.doi.org/10.1016/j.bmcl.2015.11.009