Identification of nonabsorbable inhibitors of the scavenger receptor-BI (SR-BI) for tissue-specific administration

Article abstract of DOI:10.1016/j.bmcl.2016.03.025

The identification of a low-permeability scavenger receptor BI (SR-BI) inhibitor starting from the ITX-5061 template is described. Structure-activity and structure-permeability relationships were assessed for analogs leading to the identification of compound 8 as a potent and nonabsorbable SR-BI inhibitor.

Full text of DOI:10.1016/j.bmcl.2016.03.025

Bioorganic & Medicinal Chemistry Letters 26 (2016) 1901–1904
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Identification of nonabsorbable inhibitors of the scavenger
receptor-BI (SR-BI) for tissue-specific administration
⇑
Steven M. Sparks , Huiqiang Zhou, Claudia Generaux, Lindsey Harston, David Moncol,
Channa Jayawickreme, Janet Parham, Patrick Condreay, Thomas Rimele
GlaxoSmithKline, Enteroendocrine Discovery Performance Unit and Platform Technology and Science, 5 Moore Drive, Research Triangle Park, NC 27709, United States
a r t i c l e i n f o
a b s t r a c t
Article history:
The identification of a low-permeability scavenger receptor BI (SR-BI) inhibitor starting from the ITX-
5061 template is described. Structure–activity and structure–permeability relationships were assessed
for analogs leading to the identification of compound 8 as a potent and nonabsorbable SR-BI inhibitor.
Ó 2016 Elsevier Ltd. All rights reserved.
Received 14 December 2015
Revised 7 March 2016
Accepted 8 March 2016
Available online 9 March 2016
Keywords:
Scavenger receptor class B type I
SR-BI inhibitor
Nonabsorbed SR-BI inhibitor
Intestinal SR-BI
Introduction
sporozoites, the causative agents of malaria, into hepatocytes has
also been postulated to occur through interaction with SR-BI.11–13
The scavenger receptor class B type I (SR-BI) is a multifunctional
transmembrane receptor present in a variety of tissues and cell
types where its main functions govern the selective uptake of ster-
ols and lipids.1–4 SR-BI has the highest expression in tissues regu-
lating sterol metabolism including the liver, intestine, and the
steroidogenic tissues, the adrenal glands, ovaries and testes. The
receptor has been widely studied for its role in cholesterol and
cholesterol ester exchange between cells and lipoprotein particles
including high-density lipoprotein (HDL). With these functions SR-
BI plays a key role in the process of reverse cholesterol transport in
the liver, where it provides a mechanism for the reuptake of
cholesterol from HDL.5 SR-BI has also been implicated in inflamma-
tory processes governed by oxidized lipids via the receptor’s recog-
nition of oxidized lipoprotein particles including oxidized low
density lipoprotein (OxLDL).6
Our interest in SR-BI results from the numerous papers detail-
ing the intestinal physiology mediated by the receptor where it
also acts as a mediator of sterol and hydrophobic small molecule
(vitamins and carotenoids) uptake.14–17 Postprandial micelles
(PPM) composed of dietary lipids resulting from the breakdown
of triglycerides to fatty acids and monacylgycerol mix with bile
salts in the lumen to form micelles which are recognized by SR-BI
at the enterocyte brush border membrane. Upon PPM binding,
SR-BI acts as a lipid sensor which initiates apolipoprotein B
trafficking from the apical membrane within the enterocytes and
is thus involved in intestinal lipoprotein production.18 Indeed,
upregulation of intestinal SR-BI under conditions of insulin
resistance in rodents is associated with the overproduction of
apolipoprotein B48-containing lipoproteins from the intestine.19
In addition, overexpression of SR-BI has been shown to accelerate
lipid absorption in rodents.20
While intestinal modulation of SR-BI may provide beneficial
effects for ailments caused by dysregulated intestinal lipid meta-
bolism including dyslipidemia, diabetes and obesity, to date only
tissue selective transgenic modulation has allowed for the study
of the receptor at the intestinal level. To further explore the biology
of SR-BI in the intestine, we sought to develop low-permeability
inhibitors of the receptor. These compounds are designed to be
orally administered and nonabsorbable, eliciting their potential
therapeutic effects via inhibition of SR-BI locally in the GI-
tract.21,22
In addition to lipid particle binding and the subsequent role
played in lipid metabolism, SR-BI has been implicated in the entry
of viral particles and parasites into host cells.7,8 The receptor’s role
as a key co-factor required for Hepatitis C Virus (HCV) entry has
been extensively studied, with inhibitors of SR-BI now advancing
in clinical trials as HCV entry inhibitors.9,10 Entry of Plasmodium
⇑
Corresponding author at present address: Viamet Pharmaceuticals, 4505
Emperor Blvd., Suite 300, Durham, NC 27703, United States. Tel.: +1 919 467
8539x317.
E-mail address: ssparks@viamet.com (S.M. Sparks).
http://dx.doi.org/10.1016/j.bmcl.2016.03.025
0960-894X/Ó 2016 Elsevier Ltd. All rights reserved.

1902
S. M. Sparks et al. / Bioorg. Med. Chem. Lett. 26 (2016) 1901–1904
After a survey of known SR-BI inhibitors,23–28 we choose ITX-
5061 (1) as our starting point (Fig. 1). ITX-5061 (1) was developed
as a p38 mitogen-activated protein kinase (MAPK) inhibitor which
showed increased HDL-cholesterol levels in humans that was sub-
O
a or a, b
O
Br
R
O
O
O
S
O
O O
S
N
N
H
N
N
H
H
H
OMe
O
O
OMe
O
sequently attributed to the compound’s activity as
a SR-BI
9
13 16
-
D
,
inhibitor.29 Owing to the compound’s high polar surface area
(tPSA = 123 Å2), we surmised that addition of polar or charged
functionalities off of either the sulfonamide or morpholino termini
of ITX-5061 would limit cellular permeation and allow for the
identification of a nonabsorbed SR-BI inhibitor.
O
Br
c, b, d, e
O
O
N3
O
S
O
HO
N
H
N
H
O
OMe
O
F
E
Chemistry for the preparation of modified sulfonamide analogs
of ITX-5061 is shown in Scheme 1. Alkylation of 1-napthol fol-
lowed by Friedel–Crafts acylation and saponification provided the
desired ketoacids A. Coupling with 5-t-butyl-2-methoxybenzene-
1,2-diamine followed by treatment with the appropriate sulfonyl
chloride afforded the substituted sulfonamides B. For compounds
5–8, SEM protection of the sulfonamide nitrogen was conducted
to prevent intramolecular cyclization of the 3-chloropropylsulfon-
amide intermediate. Subsequent treatment with the desired ami-
nes followed by deprotection provided the final compounds C. In
select cases ester saponification was required to afford the final
carboxylic acid-containing analogs.
Chemistry for the preparation of preparation of morpholino
replacements of ITX-5061 is shown in Scheme 2. Bromide deriva-
tive D was treated directly with substituted amines to provide
either tertiary amine 15, quaternary amines 9 and 13, or interme-
diate esters which required saponification to yield the desired car-
boxylic acids 14 and 16. Amide derivatives were formed starting
from bromide E. Azide displacement followed by saponification,
amide bond formation, and sulfonamide formation provided inter-
mediate F. Reduction followed by coupling and deprotection
afforded amides 11 and 12.
O
O
f, g, h
or f, i, b
O
N
H
R'''
O
S
O
N
N
H
H
OMe
O
11-12
Scheme 2. Reagents and conditions: (a) amine, CH3CN, 90 °C; (b) LiOH, H2O, THF;
(c) NaN3, DMF, 90 °C; (d) 5-t-butyl-2-methoxybenzene-1,3-diamine, HATU, DIEA,
DMF; (e) CH3SO2Cl, pyr; (f) PPh3, H2O, THF; (g) RCO2H, HATU, DIEA; (h) TFA;
(i) RCOCl, DIEA.
ethylindocarbocyanine perchlorate (DiI) from HDL particles into
the human osteosarcoma U2OS cell line overexpressing human
SR-BI.24 The potency of the compounds was determined using a
ten-point dose response curve and reported as a pIC50. Compounds
with acceptable inhibition of DiI uptake were progressed into a cel-
lular permeability assay utilizing Madin-Darby canine kidney
(MDCK) cells. Passive permeation was evaluated in this assay by
using a P-glycoprotein (PGP) inhibitor added to the apical media
to provide an apparent permeability (Papp) value.30 Compounds
with low to medium permeability (Papp < 20 nM/s) were assessed
for lower GI stability by incubation in rat cecal contents under an
anaerobic atmosphere. This luminal stability assay was devised
to measure the potential for gut bacterial metabolism of test
The analogs were evaluated for their ability to block the cellular
uptake of the fluorescent lipid 1,10-dioctadecyl-3,3,30,30-tetram-
O
Table 1
O
N
O
S
O
SR-BI inhibition: sulfonamide SAR, SPR, and luminal stability
O
N
N
H
H
OMe
O
1
O
SR-BI pIC50 = 7.0
O
N
O
O
tPSA = 123 Å2
S
O
R
N
N
H
H
OMe
O
Figure 1. SR-BI inhibitor ITX-5061 (1).
a
Compound
R
pIC50
MDCK
Papp
(nM/s)
Luminal
stabilityb
T½ (h)
b
O
OH
O
R
a, b, c
HO
1
2
Methyl
Propyl
HO2C
7.0 0.2
7.3 0.4
126
—
O
A
3
6.7 0.1
2
1
>12
>12
Cl-
N+
HO
HO
HO2C
HO2C
OH
N
4
6.8 0.3
d, e
O
O
R
O
S
O
R'
N
N
5
6
7.4 0.5
5.3 0.1
8
11
—
H
H
N
N
O
O
B
—
f, g, h
or f, g, h, c
O
H
N
O
R
O
S
O
HO
R''2N
7
8
7.3 0.3
6.8 0.2
<1
<1
11
N
N
H
H
OH
TFA-
O
O
C
N+
>12
Scheme 1. Reagents and conditions: (a) RCH2CH2X, K2CO3, DMF; (b) ClCOCO2Et,
AlCl3, DCM; (c) LiOH, H2O, THF; (d) 5-t-butyl-2-methoxybenzene-1,3-diamine,
HATU, DIEA, DMF; (e) R0SO2Cl, pyr; (f) For R0 = (CH2)3Cl, SEM-Cl, Et3N, 0 °C; (g) KI,
R002NH, CH3CN, 80 °C; (h) HCl, MeOH or HF-pyr.
a
pIC50 values are expressed as the mean of P2 replicates.
—Indicates not tested.
b

S. M. Sparks et al. / Bioorg. Med. Chem. Lett. 26 (2016) 1901–1904
1903
Table 2
Encouraged by the results obtained with the sulfonamide ana-
logs, we next explored modifications to the morpholine moiety
(Table 2). While direct quaternization of the morpholino-group
(9) provided a 1-log reduction in potency, extending the morpho-
line group out by two additional methylene groups (10) main-
tained potency equivalent to 1. Amide containing compounds 11
and 12 highlighted that a terminal amine was well tolerated while
a terminal carboxylic acid was not, however glycine amide 11 suf-
fered from rapid degradation in the luminal stability assay. Quater-
nary amine 13 showed moderate potency with diacid 14
reinforcing that acidic functionality did not provide potent inhibi-
tion of SR-BI. Diol 15 provided the desired potency (pIC50 = 7.1) but
suffered from moderate permeability (Papp = 45 nM/s). Piperidine
derivative 16 provided further evidence that acidic functionality
was not well tolerated.
With a stable of molecules meeting our desired in vitro profile
for further evaluation as nonabsorbed SR-BI inhibitors, we pro-
gressed select compounds into DMPK studies (Table 3). Utilizing
male CD rats, portal and jugular drug blood concentrations were
measured following oral dosing (30 mg/kg) out to a 24 h timepoint.
Fecal drug recovery was evaluated for compounds with portal vein
drug concentrations of less than 100 ng/mL with the goal of achiev-
ing greater than 80% recovery for progression. In general, portal
Cmax values were below the 100 ng/mL cutoff with the exception
of compound 16 (Portal Cmax = 2003 ng/mL), which we hypothesize
may achieve higher drug levels due to active transport. The fecal
drug recovery of the remaining compounds identified quaternary
amines 8 and 9 as promising candidates for evaluation as nonab-
sorbed SR-BI inhibitors, with compound 8 affording the best over-
all profile with higher potency (pIC50 = 6.8) and lower variability in
the fecal recovery assay (fecal drug recovery = 83 3%) compared
with amine 9. Finally, the aqueous solubility of solid compound 8
in simulated fasted intestinal fluid was measured and showed
SR-BI inhibition: sulfonamide SAR, SPR, and luminal stability
O
O
R
O
O
S
N
N
H
H
OMe
O
a
Compound
R
pIC50
MDCK
Papp
(nM/s)
Luminal
stabilityb
T½ (h)
b
TFA-
N+
9
6.0 0.1
<1
>12
O
N
10
11
7.3 0.3
7.3 0.2
—
3
—
3
O
NH2
O
O
N
H
O
12
13
6.0 0.4
6.3 0.2
3
11
N
H
TFA-
N+
OH
<1
>12
N
CO2H
CO2H
OH
OH
14
15
16
<4.5
—
45
5
—
N
N
7.1 0.1
6.0 0.4
>12
>12
CO2H
a
pIC50 values are expressed as the mean of P2 replicates.
—Indicates not tested.
b
excellent solubility (FaSSIF solubility = 808 lg/mL).
In summary, optimization of the structure–activity and struc-
ture–permeability relationships starting from the ITX-5061 tem-
plate afforded compound 8 as a nonabsorbed inhibitor of SR-BI.
Following assessment of SR-BI inhibition, in vitro permeability
and GI-stability were assessed to identify low permeability-high
stability compounds which were progressed into in vivo DMPK
studies keying on identification of compounds with low portal vein
drug concentrations. Finally, fecal drug recovery allowed for the
identification of compound 8 as an optimal agent to study the bio-
logical effects of small molecule SR-BI inhibition in the GI-tract.
Further studies evaluating the gastrointestinal effects of SR-BI inhi-
bition utilizing nonabsorbed inhibitor 8 will be reported in due
course.
Table 3
DMPK properties of select SR-BI inhibitorsa,b,c
Compound
Portal
Cmax (ng/mL)
Systemic
Cmax (ng/mL)
Fecal drug
recoveryb (%)
8
9
12
13
16
77
28
25
51
2003
13
10
3
13
590
83
3
84 34
46 17
61 14
—
a
Dosed via oral gavage at 30 mg/kg to male CD rats in 0.5% HPMC/0.1% Tween
80/water.
b
Fecal recovery values are expressed as the mean standard error of P2
replicates.
c
—Means not determined.
Supplementary data
compounds. Compounds with a half-life (T½) of 11 h or greater
were considered for progression.
Supplementary data (experimental procedures for compound 8
and assays protocols) associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.bmcl.2016.03.
025.
Initial studies explored modifications to the sulfonamide por-
tion of ITX-5061 (Table 1). ITX-5061 (1) was evaluated to establish
a baseline for potency and permeability and profiled with a pIC50 of
7 and high permeability, with a Papp value of 126 nM/s. Propyl ana-
log 2 was profiled to assess the possibility of extending the sulfon-
amide group and was equipotent, suggesting further modifications
off the sulfonamide region would be tolerated. Adjusting the
oxidation state of the terminal carbon atom to a carboxylic acid
(3) provided acceptable potency with low permeability and
good GI-stability. Quaternary amines (4 & 8) were well tolerated
and provided low permeability and excellent in vitro GI-stability.
Amines extended by a propyl linker terminated with diol- (5) or
triol-functionality (7) provided excellent potency, however,
adjustment of the oxidation state to the dicarboxylic acid (6)
greatly eroded activity.
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