Novel Tricyclic Pyroglutamide Derivatives as Potent RORγt Inverse Agonists Identified using a Virtual Screening Approach

Article abstract of DOI:10.1021/acsmedchemlett.0c00496

Employing a virtual screening approach, we identified the pyroglutamide moiety as a nonacid replacement for the cyclohexanecarboxylic acid group which, when coupled to our previously reported conformationally locked tricyclic core, provided potent and selective RORγt inverse agonists. Structure-activity relationship optimization of the pyroglutamide moiety led to the identification of compound 18 as a potent and selective RORγt inverse agonist, albeit with poor aqueous solubility. We took advantage of the tertiary carbinol group in 18 to synthesize a phosphate prodrug, which provided good solubility, excellent exposures in mouse PK studies, and significant efficacy in a mouse model of psoriasis.

Full text of DOI:10.1021/acsmedchemlett.0c00496

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Letter
Novel Tricyclic Pyroglutamide Derivatives as Potent RORγt Inverse
Agonists Identified using a Virtual Screening Approach
Qingjie Liu,* Douglas G. Batt, Carolyn A. Weigelt, Shiuhang Yip, Dauh-Rurng Wu, Max Ruzanov,
John S. Sack, Jinhong Wang, Melissa Yarde, Sha Li, David J. Shuster, Jenny H. Xie, Tara Sherry,
Mary T. Obermeier, Aberra Fura, Kevin Stefanski, Georgia Cornelius, Purnima Khandelwal,
Joseph A. Tino, John E. Macor, Luisa Salter-Cid, Rex Denton, Qihong Zhao, and T. G. Murali Dhar
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ABSTRACT: Employing a virtual screening approach, we identified the pyroglutamide moiety as a nonacid replacement for the
cyclohexanecarboxylic acid group which, when coupled to our previously reported conformationally locked tricyclic core, provided
potent and selective RORγt inverse agonists. Structure−activity relationship optimization of the pyroglutamide moiety led to the
identification of compound 18 as a potent and selective RORγt inverse agonist, albeit with poor aqueous solubility. We took
advantage of the tertiary carbinol group in 18 to synthesize a phosphate prodrug, which provided good solubility, excellent exposures
in mouse PK studies, and significant efficacy in a mouse model of psoriasis.
KEYWORDS: RORγt, inverse agonist, pyroglutamide, virtual screening, phosphate prodrug
inflammatory bowel disease, and multiple sclerosis. The utility
of down-regulation of IL-17 as a treatment for autoimmunity
has been validated in animal models as well as clinically by
anti-IL-17 antibodies (such as secukinumab and ixekizumab)
which have been approved for clinical use.5 Thus, considerable
effort has been expended to identify small molecules, which
would down-regulate IL-17 through modulation of RORγt.6−8
Previous reports from our laboratories have detailed the
discovery of several series of small-molecule inverse agonists
with potential utility in the treatment of autoimmune
diseases.9−11 In particular, compounds bearing carboxylic
acid groups have proven to be very potent and selective
RORγt inverse agonists, culminating in identification of the
clinical candidate BMS-986251 (1).11 As part of our continued
pursuit of RORγt inverse agonists, we wanted to identify
compounds lacking the carboxylate moiety due to the potential
he retinoic acid-related orphan receptor (ROR) family
Tcomprises three nuclear hormone receptors, RORα,
Figure 1. Previously reported RORγt inverse agonists
RORβ, and RORγ. These receptors are widely expressed in
many tissues and are involved in numerous regulatory
functions.1 In contrast, the expression of RORγt, a 498
amino acid splice variant of RORγ, is restricted to lymphoid
cells. Acting as an important transcription factor in thymic
tissue, RORγt mediates the differentiation of naive T cells into
Th17 cells and regulates their maintenance and production of
pro-inflammatory cytokines, including IL-17, IL-22, and GM-
CSF.2−4 IL-17 has been implicated in the pathology of multiple
autoimmune diseases, including rheumatoid arthritis, psoriasis,
Received: September 11, 2020
Accepted: November 3, 2020
https://dx.doi.org/10.1021/acsmedchemlett.0c00496
© XXXX American Chemical Society
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A

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Figure 2. From virtual lead to tricyclic pyroglutamide series 4.
suitable for evaluation in animal models of autoimmune
diseases (Figure 1).
Virtual screening, wherein a library of hypothetical
molecules is evaluated in silico for interaction with a biological
target of interest, can be an efficient way to narrow the focus of
a discovery effort and guide the actual preparation of
compounds more likely to show interesting biological activity.
Because of the huge number of possible nonacidic substituents
which could be envisioned to replace the cyclohexylcarboxylate
moiety of 1, we undertook a virtual screening effort by creating
a virtual library of compounds with nonacidic moieties linked
to our previously reported and optimized (S)-3-((4-
fluorophenyl)sulfonyl)-3-(4-(perfluoropropan-2-yl)phenyl)-
pyrrolidine moiety.9 Potential lead structures were docked into
the ligand binding domain (LBD) of RORγt from an in-house
X-ray crystal structure with a related compound in the ligand
binding pocket using Glide SP.14 Constraints and post
minimization were not required, and binding poses were
ranked by the Glide docking score. Of the ∼60 000 virtual
analogues screened, the top-scoring 400 were evaluated
visually. Many were discarded for lack of additional binding
interactions with the LBD, or for lack of synthetic versatility to
enable SAR studies. One of the most attractive candidates was
the (S)-pyroglutamide analogue 3 (Figure 2). In the binding
model, the left-hand portion of 3 was predicted to bind in the
hydrophobic pocket, making good contacts with the side
chains of Trp317 on helix 3, His479 on helix 11, Ile400 and
Phe401 on helix 7, and Met365 and Arg364 on helix 5, as
previously reported (Figure 3).9 In addition, (a) the
benzamide moiety was predicted to project into a hydrophobic
pocket, (b) the benzamide carbonyl was predicted to form a
Figure 3. Model of compound 3 in the binding pocket of RORγt.
liability of acyl glucuronide formation with carboxylic
acids.12,13 A previously reported nonacidic compound 2
showed good potency and selectivity, along with useful
pharmacokinetics and pharmacodynamics.10 Here, we report
our continued efforts to find additional nonacid-containing
analogues as inverse agonists of RORγt with in vitro profiles
Table 1. Pyrrolidinone N-Substitution
a
ab
,
bc
,
d
compd
R
RORγt GAL4 EC50 (nM)
IL-17 hWB EC50 (nM)
LM t1/2 (min) (h, m, r)
clogP
5
6
7
8
9
10
CH2-4-F-Ph
CH2-4-CN-Ph
CH2CH2CN
CH2COOH
iPr
7.0 1.4
30 29
18 11
1300 1500
13 1.1
8.5 6.5
57 4.1
nt
30 15
nt
61 28
81 5.4
24, 29, 35
58, 74, 61
>120, 79, 45
nt
50, 52, 45
>120, 75, 55
6.35
6.06
4.25
3.93
5.25
4.47
Me
a
b
c
Mean EC50 values standard deviation for two or more experiments, except as noted. nt = not tested. Half-life in human, mouse, and rat liver
microsomes log P calculated using the ChemAxon calculator15
d
B
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Table 2. Pyrrolidinone Ring Substitution
a
b
c
Mean EC50 values standard deviation for two or more experiments, except as noted. Half-life in human, mouse, and rat liver microsomes log P
calculated using the ChemAxon calculator15
hydrogen bond to the backbone NH of Glu379, and (c) the
side chain of Arg367 shifted to form a hydrogen bond to the
pyroglutamate carbonyl. In the event, preparation of this
compound confirmed inverse agonist potency for the (S)-
pyroglutamide in the RORγt GAL4-Luciferase reporter assay
(GAL4, Figure 2). In contrast, the corresponding (R)-
pyroglutamide analogue was over 30-fold less potent (EC50
1.5 μM, compared to 46 nM for 3). The ease of synthesis of
pyroglutamic acid derivatives and the potential for additional
pyroglutamide ring substitution were also appealing features of
this lead, which was selected for further study.
We opted to explore the pyroglutamide structure−activity
relationship (SAR) in our tricyclic series (generic structure 4,
Figure 2), since conformational restraint significantly improved
C
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a
Table 4. PK Profile of Compound 18 in BALB/c Mice
b
c
route
IV
2
PO
dose (mg/kg)
Cmax (μM)
C24h (μM)
4
2.2 0.6
0.62 0.21
4.3 2.3
43.9
Tmax (h)
AUC72h (μM-h)
AUCtot (μM-h)
T1/2 (h)
23.0
23.7
15
44.4
CL (mL/min/kg)
Vss (L/kg)
F (%)
2.0
2.7
94
a
b
12 time points, 3 mice/time point/route. Solution vehicle: 2.5%
c
NMP; 67.5% PEG 400; 4.5% Pluronic F-68; 25.5% water. Solution
vehicle: 5% NMP; 76% PEG 400; 19% TPGS.
Figure 4. Co-crystal structure of compound 18 with the LBD of
RORγt (PDB ID 7JYM).
Figure 5. Phosphate prodrug of 18.
Table 3. In Vitro Profile of Compound 18
RORγt GAL4 EC50 (nM)
RORα GAL4 EC50 (nM)
16
9
>10 000
RORβ GAL4 EC50 (nM)
IL-17 hWB EC50 (nM)
mouse Th17 EC50 (nM)
PXR EC50 (nM)
LXRα EC50 (nM)
LXRβ EC50 (nM)
rCYP IC50 (nM)
1A2
8400 2100
43 24
25 20
>50 000
>7500
>7500
>20 000
>20 000
4900
2C8
2C9
2C19
2D6
3A4
>20 000
>20 000
>20 000
>100 000
>120, >120, >120
2.5, 1.3
Figure 6. Oral efficacy of prodrug 19 in mouse acanthosis. Doses are
of parent equivalent.
HepG2 EC50 (nM)
LM t1/2 (min) h, m, r
protein binding, % free (h, m)
CACO-2 A-B, B-A (nm/s)
CACO-2 efflux ratio
solubility (mg/mL):
@pH 1 (0.1 N HCl solvent)
@pH 4 (50 nM acetate buffer)
@pH 6.5 (50 nM acetate buffer)
fluorine substituent to temper the lipophilicity (clogP 6.06, vs
6.35 for 5) improved the metabolic half-life. Since the clogP for
this compound was still quite high, and the benzylic methylene
linker was felt to be a metabolic liability, smaller and less
lipophilic substituents were explored. The propiononitrile
derivative 7 (clogP 4.25) was still potent in the GAL4 assay
with slightly improved hWB potency. Importantly, the human
metabolic half-life was significantly improved, although there
was very little improvement for this parameter in rodents. The
acidic compound 8, however, lost GAL4 potency dramatically.
This was not surprising since this part of the LBD of RORγt is
hydrophobic, based on the aforementioned model (Figure 3).
Smaller substituents (isopropyl 9 and methyl 10) on the
nitrogen atom of pyroglutamide revealed that larger groups
were not required for good GAL4 potency, while moderate
hWB potency and some degree of metabolic stability
(particularly in human microsomes) could be maintained.
Overall, the SAR of the N-substituent suggested that a wide
variety of groups were tolerated, leading to compounds with
moderate to good potency in the GAL4 and hWB assays,
93, 190
2.0
<0.001
<0.001
<0.001
the RORγt potency in both GAL4 and a human whole blood
assay (hWB) by about 50-fold over that of the unconstrained
series.10 We initially explored variation of the pyrrolidinone N-
substituent, as shown in Table 1. Since the binding model for 3
suggested that lipophilic substituents would be beneficial (vide
supra), the 4-fluorobenzyl compound 5 was prepared and
found to be very potent in GAL4 (EC50 7 nM) and moderately
potent in hWB with an EC50 of 57 nM. However, it had poor
metabolic stability, as indicated by short half-lives following
incubation with human, mouse, and rat liver microsomes
(LM). A second benzyl analogue 6 with a nitrile replacing the
D
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Table 5. Exposure of Compound 18 following Dosing Prodrug 19 in Mouse Acanthosis
a
dose (mg/kg QD)
5
15
45
274.3
162.9
8.34 2.26
0.91 0.33
5.82 1.10
13.9×
day 1 AUCtot (μM-h)
day 9 AUC24 (μM-h)
day 9 Cmax (μM)
day 1 C24h (μM)
day 9 C24h (μM)
13.9
9.0
30.9
45.2
0.65 0.19
0.21 0.08
0.15 0.08
0.4×
2.58 0.36
0.48 0.14
1.36 0.56
3.2×
b
day 9 C24h fold mouse Th17 IC90
a
Prodrug was dosed to provide the indicated amount of parent after phosphate hydrolysis; solution vehicle: 20% of 0.1 M acetate buffer; 80% PEG
b
400. Protein binding adjusted mouse Th17 IC90 = 0.42 μM
†
Scheme 1. General Synthesis of N-Substituted Pyroglutamic Acid Intermediates
†
Reagents and conditions: (a) NaH, THF or DMF, rt; (b) LiOH, THF:MeOH:H2O (3:1:1), rt, 32-69% for 2 steps; (c) NaOH, H2O, 50 °C; (d)
concentrated HCl, acetone, reflux, 60% for 2 steps; (e) TBDMS-Cl, Et3N, imidazole, THF-DMF, rt, quantitative yield; (f) RuO2·H2O, NaIO4,
EtOAc-H2O, rt, 89-98%; (g) TFA, DCM, 0 °C - rt, quantitative yield; (h) Cs2CO3, CH3I, 50 °C, 37%; (i) NaH, CH3I, DMF, 50 °C, 26%; (j)
DAST, DCM, 0 °C to rt, 34%; (k) 4M HCl in dioxane, DCM, rt, 63%.
although metabolic stability remained a significant issue,
particularly in mouse which was the species for preclinical
efficacy studies.
pronounced in human in the case of difluoro analogue 16. The
latter may be due to the increase in lipophilicity (clogP 5.32, vs
4.41 for the corresponding monofluoro analogues.)
Additional substitution on the pyrrolidinone ring was
explored in order to examine effects on potency and metabolic
stability (Table 2). The (4S)-hydroxy substituted compound
11 lost potency by 3-fold in both GAL4 and hWB without
improving metabolic stability. The corresponding methoxy
derivative 12 regained whole blood potency but at the expense
of reduction in metabolic stability, probably because of
oxidative demethylation. Inversion of the hydroxyl group of
11 with a fluoro moiety provided the (4R) derivative 13, with
potency in hWB improved over the unsubstituted analogue 10.
Gratifyingly, the metabolic stability was restored to the level
seen for 10. On the basis of the well-precedented improvement
in stability toward oxidative dealkylation seen with deuterium
substitution,16 the trideuteromethyl analogue 14 was prepared
and showed a dramatic improvement in stability in both rodent
species. Curiously, the epimeric fluoride 15, while retaining
potency, showed a drop-off in stability, which was even more
While 14 was a promising compound with desirable potency
and excellent microsomal stability, very poor aqueous solubility
(<1 μg/mL) imposed a significant hurdle for in vivo evaluation.
Because of the beneficial effect of the (4R)-fluoro substituent
on both hWB potency and rodent metabolic stability, we
briefly re-examined variation of the pyrrolidinone N-
substituent in the context of this improvement. A pendant
hydroxyl group was considered as a possible solution to the
solubility issue, since this would provide a handle for potential
solubilizing prodrugs17 should that become desirable. The
easily prepared 2-hydroxyethyl derivative 17 was intriguing in
that no significant change in potency relative to 14 was seen,
although metabolic stability in rodents was adversely effected.
However, replacement of the primary carbinol of 17 with the
dimethyl tertiary carbinol provided 18, with both potency and
metabolic stability virtually identical to those achieved with 14.
An X-ray cocrystal structure of 18 bound in the LBD of RORγt
E
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‡
Scheme 2. Synthesis of Final Compounds 18 and 19
‡
Reagents and conditions: (m) HATU, DIEA, DMF, rt, 35%; (n) K2CO3, iPrOH, 100 °C, 51%; (o) 5-methyl-1H-tetrazole, dibenzyl
diisopropylphosphoramidite, DCM, 0 °C − rt; (p) 35% H2O2 in water, DCM, 0 °C, 89% for two steps o & p; (q) H2, Pd−C, MeOH/EtOAc, rt,
78%.
was obtained (Figure 4, PDB ID 7JYM), and it was gratifying
to see the binding pose of this compound to be as predicted for
compound 3 (Figure 3). In addition, the pyroglutamide
fluorine substituent makes favorable contacts with Cα and the
side chain methylenes of Arg364. The gem-dimethyl moiety
makes beneficial lipophilic contacts with the side chains of
Gln286, Leu287, and Phe377, directing the hydroxyl group
toward solvent. Although no appropriate water molecules were
observed in the crystal structure, the distances from the
pyrrolidinone oxygen to the side chain of Arg367 (4.3 Å) and
to the backbone carbonyl oxygen of Arg364 (3.7 Å) suggest
the possibility of water-mediated hydrogen bonding between
the lactam and the protein.
Because of its promising profile, 18 was further profiled in
vitro, and these results are summarized in Table 3. Compound
18 was very selective against other nuclear hormone receptors
(RORα, RORβ, PXR, LXRα, and LXRβ) and was devoid of
recombinant cytochrome P450 enzyme inhibition (rCYP),
except for weak inhibition of the 2C9 isozyme. In addition, 18
showed no effect on HepG2 liver cells (EC50 > 100 μM),
suggesting a reduced liability for liver toxicity relative to
another nonacidic compound (2, HepG2 EC50 42 μM).
Compound 18 had reasonable permeability in CACO-2 cells,
with no significant efflux. Pharmacokinetic studies of 18 in
BALB/c mice (Table 4) showed good oral exposure, with
the parent compound. At 25 mg/kg (parent equivalent), a dose
which could not be achieved with the poorly soluble parent,
the prodrug provided a near-dose proportional increase in
exposure (C24h 4.26 0.63 μM).
These promising results prompted us to evaluate compound
19 in the mouse acanthosis model, a preclinical model of
psoriasis.18,19 The prodrug was dosed at 6, 17, and 50 mg/kg,
equivalent to 5, 15, and 45 mg/kg of parent, calculated on the
basis of the PK results for 19. Acanthosis was induced with
recombinant humanized IL-23 injected every other day into
the right ear of C57bl/6 female mice, until the last injection on
day 9. A starting baseline measurement of ear thickness (before
the first injection) was made on day 0, and ear thickness was
then measured every other day, prior to the next ear injection.
Compound 19 was dosed orally approximately 2 h before the
first IL-23 injection and continued once daily until day 9. The
placebo/control group was dosed with blank vehicle, and a
recombinant dual chain human anti-IL-23 adnectin was
administered SC as a positive control at doses of 3 mg/kg
on days 0, 3, and 7 (which gave maximal efficacy based on
other studies). No significant weight loss was seen at any of the
doses, suggesting that the compound was well tolerated. At the
two higher doses, trough blood levels of the parent 18 (Table
5) exceeded the protein binding adjusted IC90 for this
compound in a mouse Th17 reporter assay by 3- and 6-fold,
respectively. (No intact phosphate prodrug was observed in
blood samples.) Gratifyingly, as shown in Figure 6, the 15 and
45 mg (parent equivalent) doses of prodrug 19 resulted in
significantly reduced ear swelling (38% and 67%, respectively,
on day 9), with the highest dose comparable in efficacy to the
IL-23 antibody.
excellent bioavailability (94%), low clearance (CL), long T1/2
,
and a good peak-to-trough ratio (3.5). However, 18 had very
poor aqueous solubility (Table 3), which would severely limit
dosing for in vivo efficacy and tolerability studies.
As a preliminary approach to improving solubility, the
phosphate ester 19 (Figure 5) was prepared as a potential
prodrug. Prodrug 19, as expected, provided a dramatic increase
in solubility at pH 6.5 (≥4.7 mg/mL). This enabled in vivo
studies in BALB/c mice with 19 using higher doses than were
possible with 18 itself, while also avoiding the use of N-
methylpyrrolidine (NMP) and emulsifier in the dosing
solution (vehicle: 20% of 0.1 M acetate buffer, 80% PEG
400). At 5 mg/kg (parent equivalent), 19 showed roughly the
same trough concentration of 18 (0.76 0.13 μM) as when 18
was dosed directly (1.08 0.15 μM), with 67% exposure of
Three general routes (Scheme 1) were used to make the N-
substituted pyroglutamic acid intermediates for coupling to the
tricyclic core (the preparation of which has been previously
described).10 Alkylation of commercially available methyl or
ethyl (S)-5-oxopyrrolidine-2-carboxylate 20 by an alkyl halide
followed by saponification of ester 21 provided substituted
(S)-pyroglutamic acid 22. The N-substituted pyroglutamic acid
derivatives could also be obtained by the cyclization of N-alkyl
glutamate esters or N-alkylglutamic acid,20−22 which were
F
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Authors
obtained by reductive alkylation of diethyl L-glutamate
hydrochloride or L-glutamic acid with an aldehyde or ketone,
or by another N-alkylation method. For example, conjugate
addition of glutamic acid 23 to acrylonitrile provided 24, which
was cyclized to provide 25. Pyroglutamic acids bearing an
additional ring substituent were prepared from commercially
available Boc-protected hydroxyproline ester 26.23−25 Follow-
ing protection of the hydroxy group as the TBDMS ether,
ruthenium(IV) oxide oxidation to lactam 27 and deprotection
provided 28. Monomethylation using cesium carbonate as
base, or dimethylation with sodium hydride provided, after
saponification, 29 or 30, respectively. The ring fluorine was
introduced by treatment of 26 with DAST, followed by
oxidation, deprotection, and alkylation as above to provide 34.
Coupling of the acids to the tricyclic amine core to give final
products was generally accomplished using HATU or BOP.
The synthesis of compound 18 and its prodrug 19 is
outlined in Scheme 2. Following coupling of the intermediate
unsubstituted pyroglutamic acid intermediate 33 (Scheme 1)
with amine 35,6 the resulting intermediate 36 was alkylated
with isobutylene oxide to give 18 as the major product.
Compound 18 was converted to dibenzyl phosphite 37, which
was then oxidized to dibenzyl phosphate 38 in a one-pot two-
step reaction. Reductive removal of the dibenzyl groups
afforded the phosphate prodrug 19.
In summary, from a virtual screening exercise, pyroglutamide
analogues emerged as interesting leads for identifying novel
nonacid containing RORγt inverse agonists. Optimization of
the pyroglutamide moiety in the context of our conformation-
ally locked tricyclic series led to the identification of compound
18 as a potent and selective compound with an attractive in
vitro profile. In the mouse acanthosis model of psoriasis, 18
(dosed as the more soluble phosphate prodrug 19) provided
good dose-dependent efficacy. Although a significant improve-
ment was achieved in the in vitro HepG2 liver cell toxicity
assay over a previous nonacidic lead, suggesting reduced
potential for liver injury, further evaluation of this compound
was precluded by the identification of a potential hERG
inhibition issue (hERG patch clamp EC50 1.9 μM). However,
the current work demonstrated that useful pharmacokinetics
and efficacy could be achieved with nonacidic analogues of our
clinical compound 1 and that a good solubility could be
obtained in this lipophilic series without resort to incorpo-
ration of a carboxylic acid moiety. Further exploration based
on these results will be reported elsewhere.
Douglas G. Batt − Research and Early Development, Bristol
Myers Squibb Company, Princeton, New Jersey 08540-4000,
United States
Carolyn A. Weigelt − Research and Early Development,
Bristol Myers Squibb Company, Princeton, New Jersey
08540-4000, United States
Shiuhang Yip − Research and Early Development, Bristol
Myers Squibb Company, Princeton, New Jersey 08540-4000,
United States
Dauh-Rurng Wu − Research and Early Development, Bristol
Myers Squibb Company, Princeton, New Jersey 08540-4000,
United States
Max Ruzanov − Research and Early Development, Bristol
Myers Squibb Company, Princeton, New Jersey 08540-4000,
United States
John S. Sack − Research and Early Development, Bristol Myers
Squibb Company, Princeton, New Jersey 08540-4000,
United States
Jinhong Wang − Research and Early Development, Bristol
Myers Squibb Company, Princeton, New Jersey 08540-4000,
United States
Melissa Yarde − Research and Early Development, Bristol
Myers Squibb Company, Princeton, New Jersey 08540-4000,
United States
Sha Li − Research and Early Development, Bristol Myers
Squibb Company, Princeton, New Jersey 08540-4000,
United States
David J. Shuster − Research and Early Development, Bristol
Myers Squibb Company, Princeton, New Jersey 08540-4000,
United States
Jenny H. Xie − Research and Early Development, Bristol
Myers Squibb Company, Princeton, New Jersey 08540-4000,
United States
Tara Sherry − Research and Early Development, Bristol Myers
Squibb Company, Princeton, New Jersey 08540-4000,
United States
Mary T. Obermeier − Research and Early Development,
Bristol Myers Squibb Company, Princeton, New Jersey
08540-4000, United States
Aberra Fura − Research and Early Development, Bristol Myers
Squibb Company, Princeton, New Jersey 08540-4000,
United States
Kevin Stefanski − Research and Early Development, Bristol
Myers Squibb Company, Princeton, New Jersey 08540-4000,
United States
Georgia Cornelius − Research and Early Development, Bristol
Myers Squibb Company, Princeton, New Jersey 08540-4000,
United States
ASSOCIATED CONTENT
■
sı
* Supporting Information
The Supporting Information is available free of charge at
https://pubs.acs.org/doi/10.1021/acsmedchemlett.0c00496.
Purnima Khandelwal − Research and Early Development,
Bristol Myers Squibb Company, Princeton, New Jersey
08540-4000, United States
Joseph A. Tino − Research and Early Development, Bristol
Myers Squibb Company, Princeton, New Jersey 08540-4000,
United States
John E. Macor − Research and Early Development, Bristol
Myers Squibb Company, Princeton, New Jersey 08540-4000,
United States
Luisa Salter-Cid − Research and Early Development, Bristol
Myers Squibb Company, Princeton, New Jersey 08540-4000,
United States
Description of the virtual screening protocols and
complete experimental details and characterization for
all new compounds (PDF)
AUTHOR INFORMATION
■
Corresponding Author
Qingjie Liu − Research and Early Development, Bristol Myers
Squibb Company, Princeton, New Jersey 08540-4000, United
States; orcid.org/0000-0002-8512-3979; Phone: 609-
252-4794; Email: qingjie.liu@bms.com
G
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Rex Denton − Research and Early Development, Bristol Myers
Squibb Company, Princeton, New Jersey 08540-4000,
United States
TPGS
Trp
Vss
tocopheryl polyethylene glycol succinate
tryptophan
volume of distribution.
Qihong Zhao − Research and Early Development, Bristol
Myers Squibb Company, Princeton, New Jersey 08540-4000,
United States
T. G. Murali Dhar − Research and Early Development, Bristol
Myers Squibb Company, Princeton, New Jersey 08540-4000,
United States; orcid.org/0000-0003-0738-1021
REFERENCES
■
(1) Cook, D. N.; Kang, H. S.; Jetten, A. M. Retinoic acid-related
orphan receptors (RORs): regulatory functions in immunity,
development, circadian rhythm, and metabolism. Nucl. Receptor Res.
2015, 2, 101185.
(2) Ivanov, I. I.; McKenzie, B. S.; Zhou, L.; Tadokoro, C. E.;
Lepelley, A.; Lafaille, J. J.; Cua, D. J.; Littman, D. R. The orphan
nuclear receptor RORγt directs the differentiation program of
proinflammatory IL-17+ T helper cells. Cell 2006, 126, 1121.
(3) Korn, T.; Bettelli, E.; Oukka, M.; Kuchroo, V. K. IL-17 and Th17
cells. Annu. Rev. Immunol. 2009, 27, 485.
Complete contact information is available at:
https://pubs.acs.org/10.1021/acsmedchemlett.0c00496
Notes
The authors declare no competing financial interest.
ABBREVIATIONS
(4) Lee, J. S.; Cua, D. J. The emerging landscape of RORγt biology.
Immunity 2014, 40, 451.
(5) Balato, A.; Scala, E.; Balato, N.; Caiazzo, G.; Di Caprio, R. D.;
Monfrecola, G.; Raimondo, A.; Lembo, S.; Ayala, F. Biologics that
inhibit the TH17 pathway and related cytokines to treat inflammatory
disorders. Expert Opin. Biol. Ther. 2017, 17, 1363.
(6) Dhar, T. G. M.; Zhao, Q.; Markby, D. W. Targeting the nuclear
hormone receptor RORγt for the treatment of autoimmune and
inflammatory disorders. Annu. Rep. Med. Chem. 2013, 48, 169.
(7) Fauber, B. P.; Magnuson, S. Modulators of the nuclear receptor
retinoic acid receptor-related orphan receptor-c (RORγ or RORc). J.
Med. Chem. 2014, 57, 5871.
(8) Pandya, V. B.; Kumar, S.; Sachchidanand; Sharma, R.; Desai, R.
C. Combating autoimmune diseases with retinoic acid receptor-
related orphan receptor-γ (RORγ or RORc) inhibitors: hits and
misses. J. Med. Chem. 2018, 61, 10976.
(9) Duan, J. J.-W.; Lu, Z.; Jiang, B.; Stachura, S.; Weigelt, C. A.;
Sack, J. S.; Khan, J.; Ruzanov, M.; Galella, M. A.; Wu, D.; Yarde, M.;
Shen, D.; Shuster, D. J.; Borowski, V.; Xie, J. H.; Zhang, L.; Vanteru,
S.; Gupta, A. K.; Mathur, A.; Zhao, Q.; Foster, W.; Salter-Cid, L. M.;
Carter, P. H.; Dhar, T. G. M. Structure-based discovery of phenyl (3-
phenylpyrrolidin-3-yl)sulfones as selective, orally active RORγt
inverse agonists. ACS Med. Chem. Lett. 2019, 10, 367.
■
Arg
arginine
AUC
Boc
area under the curve
tert-butyloxycarbonyl
benzotriazol-1-yloxytris(dimethylamino)-
phosphonium hexafluorophosphate
concentration at 24 h
maximum concentration
clearance
calculated log partition coefficient
dimethylaminosulfur trifluoride
dichloromethane
diisopropylethylamine
N,N-dimethylformamide
50% efficacious concentration
ethyl acetate
BOP
C24
Cmax
CL
clogP
DAST
DCM
DIEA
DMF
EC50
EtOAc
F
bioavailability
hWB
GAL4
Gln
human whole blood assay
GAL4-luciferase reporter assay
glutamine
Glu
glutamic acid
(10) Marcoux, D.; Duan, J. J.-W.; Shi, Q.; Cherney, R. J.; Srivastava,
A. S.; Cornelius, L.; Batt, D. G.; Liu, Q.; Beaudoin-Bertrand, M.;
Weigelt, C. A.; Khandelwal, P.; Vishwakrishnan, S.; Selvakumar, K.;
Karmakar, A.; Gupta, A. K.; Basha, M.; Ramlingam, S.; Manjunath,
N.; Vanteru, S.; Karmakar, S.; Maddala, N.; Vetrichelvan, M.; Gupta,
A.; Rampulla, R. A.; Mathur, A.; Yip, S.; Li, P.; Wu, D.-R.; Khan, J.;
Ruzanov, M.; Sack, J. S.; Wang, J.; Yarde, M.; Cvijic, M. E.; Li, S.;
Shuster, D. J.; Borowski, V.; Xie, J. H.; McIntyre, K. W.; Obermeier,
M. T.; Fura, A.; Stefanski, K.; Cornelius, G.; Hynes, J.; Tino, J. A.;
Macor, J. E.; Salter-Cid, L.; Denton, R.; Zhao, Q.; Carter, P. H.; Dhar,
T. G. M. Rationally designed, conformationally constrained inverse
agonists of RORγt  identification of a potent, selective series with
biologic-like in vivo efficacy. J. Med. Chem. 2019, 62, 9931.
(11) Cherney, R. J.; Cornelius, L. A. M.; Srivastava, A.; Weigelt, C.
A.; Marcoux, D.; Duan, J. J.-W.; Shi, Q.; Batt, D. G.; Liu, Q.; Yip, S.;
Wu, D.; Ruzanov, M.; Sack, J.; Khan, J.; Wang, J.; Yarde, M.; Cvijic,
M. E.; Mathur, A.; Li, S.; Shuster, D.; Khandelwal, P.; Borowski, V.;
Xie, J.; Obermeier, M.; Fura, A.; Stefanski, K.; Cornelius, G.; Tino, J.
A.; Macor, J. E.; Salter-Cid, L.; Denton, R.; Zhao, Q.; Carter, P. H.;
Dhar, T. G. M. Discovery of BMS-986251: a clinically viable, potent,
and selective RORγt inverse agonist. ACS Med. Chem. Lett. 2020, 11,
1221.
GM-CSF granulocyte-macrophage colony-stimulating factor
HATU hexafluorophosphate azabenzotriazole tetramethyl
uronium
His
IL
histidine
interleukin
Ile
IV
isoleucine
intravenous
LBD
Leu
LM
LXR
Met
NMP
PEG
Phe
PO
PXR
QD
rCYP
ROR
Rt
ligand binding domain
leucine
liver microsomes
liver X receptor
methionine
N-methylpyrrolidinone
polyethylene glycol
phenylalanine
per os (oral)
pregnane X receptor
per day
recombinant cytochrome P450
receptor-related orphan receptor
room temperature
structure−activity relationship
(12) Walles, M.; Brown, A. P.; Zimmerlin, A.; End, P. New
perspectives on drug-induced liver injury risk assessment of acyl
glucoronides. Chem. Res. Toxicol. 2020, 33, 1551.
(13) Shipkova, M.; Armstrong, V. W.; Oellerich, M.; Wieland, E.
Acyl glucuronide drug metabolites: toxicological and analytical
implications. Ther. Drug Monit. 2003, 25, 1.
SAR
TBDMS tert-butyldimethylsilyl
TFA
TH17
THF
trifluoroacetic acid
T helper 17 cell
tetrahydrofuran
̈
(14) Glide and Maestro; Schrodinger, LLC: New York, NY, 2020.
H
https://dx.doi.org/10.1021/acsmedchemlett.0c00496
ACS Med. Chem. Lett. XXXX, XXX, XXX−XXX

ACS Medicinal Chemistry Letters
pubs.acs.org/acsmedchemlett
Letter
(15) ChemAxon. https://docs.chemaxon.com/display/docs/LogP_
and_logD_calculations.html.
(16) Pirali, T.; Serafini, M.; Cargnin, S.; Genazzani, A. A.
Applications of deuterium in medicinal chemistry. J. Med. Chem.
2019, 62, 5276.
(17) Jornada, D. H.; dos Santos Fernandes, G.; Chiba, D. E.;
Ferreira de Melo, T. R.; dos Santos, J. L.; Chung, M. C. The prodrug
approach: a successful tool for improving drug solubility. Molecules
2016, 21, 42.
(18) Zheng, Y.; Danilenko, D. M.; Valdez, P.; Kasman, I.; Eastham-
Anderson, J.; Wu, J.; Ouyang, W. Interleukin-22, a TH17 cytokine,
mediates IL-23-induced dermal inflammation and acanthosis. Nature
2007, 445, 648.
(19) Chan, J. R.; Blumenschein, W.; Murphy, E.; Diveu, C.;
Wiekowski, M.; Abbondanzo, S.; Lucian, L.; Geissler, R.; Brodie, S.;
Kimball, A. B.; Gorman, D. M.; Smith, K.; de Waal Malefyt, R.;
Kastelein, R. A.; McClanahan, T. K.; Bowman, E. P. IL-23 stimulates
epidermal hyperplasia via TNF and IL-20R2-dependent mechanisms
with implications for psoriasis pathogenesis. J. Exp. Med. 2006, 203,
2577.
(20) McKinney, L. L.; Uhing, E. H.; Setzkorn, E. A.; Cowan, J. C.
Cyanoethyllation of Alpha Amino Acids. I Monocyanoethyl
Derivatives. J. Am. Chem. Soc. 1950, 72, 2599.
(21) Rigo, B.; Couturier, D.; Kolocouris, N. Studies on
pyrrolidinones. Michael reaction of glutamic acid and diethylgluta-
mate. J. Heterocycl. Chem. 1986, 23, 1769.
(22) Marchalin, S.; Kadlecikova, K.; Bar, N.; Decroix, B. The
improved synthesis of enantiopure (S)-N-arylmethyl-5-oxoprolines.
Synth. Commun. 1998, 28, 3619.
(23) Hamed, R. B.; Henry, L.; Gomez-Castellanos, J. R.; Mecinovic,
J.; Ducho, C.; Sorensen, J. L.; Claridge, T. D. W.; Schofield, C. J.
Crotonase Catalysis Enables Flexible Production of Functionalized
Prolines and Carbapenams. J. Am. Chem. Soc. 2012, 134, 471.
(24) Velaparthi, U.; Liu, P.; Wittman, M. D.; Langley, D. R. Triazine
kinase inhibitors. Patent No. WO 09/015254, 2009.
(25) Rey, Y. P.; Gilmour, R. Modulating NHC catalysis with
fluorine. Beilstein J. Org. Chem. 2013, 9, 2812.
I
https://dx.doi.org/10.1021/acsmedchemlett.0c00496
ACS Med. Chem. Lett. XXXX, XXX, XXX−XXX