Synthesis and SAR of novel, 4-(phenylsulfamoyl)phenylacetamide mGlu 4 positive allosteric modulators (PAMs) identified by functional high-throughput screening (HTS)

Article abstract of DOI:10.1016/j.bmcl.2010.07.007

Herein we disclose the synthesis and SAR of a series of 4-(phenylsulfamoyl)phenylacetamide compounds as mGlu₄ positive allosteric modulators (PAMs) that were identified via a functional HTS. An iterative parallel approach to these compounds culminated in the discovery of VU0364439 (11) which represents the most potent (19.8 nM) mGlu₄ PAM reported to date.

Full text of DOI:10.1016/j.bmcl.2010.07.007

Bioorganic & Medicinal Chemistry Letters 20 (2010) 5175–5178
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Synthesis and SAR of novel, 4-(phenylsulfamoyl)phenylacetamide mGlu4
positive allosteric modulators (PAMs) identified by functional
high-throughput screening (HTS)
Darren W. Engers a,b, Patrick R. Gentry a,b, Richard Williams a,b, Julie D. Bolinger a, C. David Weaver a,b
,
a,b,
Usha N. Menon a, P. Jeffrey Conn a,b, Craig W. Lindsley a,b,c, Colleen M. Niswender a,b, , Corey R. Hopkins
*
*
a Department of Pharmacology, Vanderbilt University Medical Center, Nashville, TN 37232, USA
b Vanderbilt Program in Drug Discovery, Nashville, TN 37232, USA
c Department of Chemistry, Vanderbilt University, Nashville, TN 37232, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
Herein we disclose the synthesis and SAR of a series of 4-(phenylsulfamoyl)phenylacetamide compounds
as mGlu4 positive allosteric modulators (PAMs) that were identified via a functional HTS. An iterative par-
allel approach to these compounds culminated in the discovery of VU0364439 (11) which represents the
most potent (19.8 nM) mGlu4 PAM reported to date.
Received 20 May 2010
Revised 29 June 2010
Accepted 2 July 2010
Available online 8 July 2010
Ó 2010 Elsevier Ltd. All rights reserved.
Keywords:
mGlu
Metabotropic glutamate receptor 4
Functional high-throughput screen (HTS)
L
-Glutamic acid (glutamate) is the primary excitatory neuro-
for mGlu4 positive allosteric modulation (Fig. 1).11–19 With the
exception of VU0361737,15 2, these disclosed mGlu4 PAMs have
been deficient in their CNS penetration, limiting their usefulness
for in vivo studies.
A functional high-throughput screening (HTS) campaign initi-
ated at Vanderbilt to identify novel mGlu4 PAMs has led to the
identification of many of these previously reported ligands
(Fig. 1). In addition to the compounds already reported, there were
a number of 4-(phenylsulfamoyl)phenylacetamides, represented
by 4 (Figs. 2 and 3). Based on our previous results, we first changed
the 2-furyl amide of 4 to the 2-pyridyl amide, 5, and a 30-fold in-
crease in potency was observed. At this point we embarked on a
lead identification program to determine if this structural class of
compounds would be a tractable lead for our mGlu4 PAM program.
transmitter in synaptic pathways in the CNS.1 There are two major
classes of glutamate receptors, divided based upon the mechanism
of activation. Ionotropic glutamate receptors (iGlu’s) are ligand-
gated ion channels, whereas metabotropic glutamate receptors
(mGlu’s) do not form an ion channel pore, but rather are a type
of G protein-coupled receptor (GPCR).2,3 The mGlu’s are members
of the group C family of GPCRs and are themselves categorized into
three families based on their receptor structure and physiological
activity: Group I (mGlu1 and mGlu5), Group II (mGlu2 and mGlu3),
and Group III (mGlu4, mGlu6, mGlu7, and mGlu8). This last group of
mGlu’s (Group III) has historically been the least studied, mainly
due to a dearth of selective ligands, especially allosteric modula-
tors (modulators that act via a site other than the orthosteric
site).4,5 We have been interested in one particular Group III recep-
tor, mGlu4, due to the potential therapeutic implications of this
receptor in disease states such as epilepsy6–8 and Parkinson’s
disease.4,5
O
OH
N
Until recently, the most well characterized mGlu4 positive allo-
steric modulator (PAM) has been the compound PHCCC,9,10 1, a
partially selective mGlu4 potentiator. However, more recently,
our laboratories and others have expanded the list of novel probes
OH
O
Cl
O
O
O
N
HN
Cl
O
N
H
HN
VU0361737, 2
Cl
VU0155041, 3
* Corresponding authors. Tel.: +1 615 343 4303; fax: +1 615 343 3088 (C.M.N.);
tel.: +1 615 936 6892; fax: +1 615 936 4381 (C.R.H.).
E-mail addresses: colleen.niswender@vanderbilt.edu (C.M. Niswender), corey.r.-
hopkins@vanderbilt.edu (C.R. Hopkins).
(−)-PHCCC,1
Figure 1. Recently reported mGlu4 PAMs.20
0960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.07.007

5176
D. W. Engers et al. / Bioorg. Med. Chem. Lett. 20 (2010) 5175–5178
Table 1
O
O
O
O
SAR analysis of sulfonamide substitution
S
S
O
N
O
N
O
H
O
H
S
R1
N
O
N
H
N
H
O
N
N
N
H
R2
VU0130734, 4
hEC = 5960 nM
GluMax = 52.0% (PHCCC)
VU0361591, 5
hEC = 159 nM
GluMax = 60.5% (PHCCC)
50
50
Figure 2. HTS Hit (VU0130734, 4) to lead (VU0361591, 5).
Compound R1
R2 EC50 (nM)
%
a,11
hmGlu4
PHCCCa,11
5
2,4,6-Trimethylphenyl
Phenyl
2-Chlorophenyl
2-Fluorophenyl
2,5-Difluorophenyl
4-Chloro-3-
trifluoromethylphenyl
3-Chloro-4-fluorophenyl
Cyclohexyl
Cyclopropyl
Cyclobutyl
Cyclopropylmethyl
2-Methoxyethyl
H
H
H
H
H
H
159
177
27
206
325
Inactive
60.5
69.2
88.2
91.7
87.6
19.5
250
+EC20 glutamate
8a
8b
8c
8d
8e
-EC20 glutamate
200
150
100
50
8f
H
H
H
H
H
H
Inactive
1030
>10,000
>5000
>5000
>10,000
19.8
63.1
23.7
64.4
70.2
22.1
8g
8h
8i
8j
8k
0
-9.5
-8.5
-7.5
-6.5
-5.5
-4.5
Log VU0361591 (5), [M]
∗
∗
8l
372
35.8
15.4
Figure 3. Concentration-dependent potentiation of glutamate in mGlu4/Gqi5 CHO
cells by 5. In the absence of an EC20 concentration of glutamate, 5, does not elicit
receptor activation.
∗
∗
8m
Inactive
∗
∗
8n
Inactive
12.0
The initial SAR centered around the right-side sulfonamide por-
tion of the molecule. To that end, compounds were synthesized in a
three step protocol starting with the commercially available 4-
nitrophenylsulfonyl chloride. Thus, the sulfonyl chloride was re-
acted with an appropriate amine (pyridine, DCM; 67–97%) yielding
7. Next, the nitro compound was reduced to the aniline (H2, Pd/C or
Ra–Ni; >95%) which was then acylated with picolinyl chloride giv-
ing the desired compounds, 8a–n, in high overall yield (Scheme 1).
The SAR analysis of the compounds is outlined in Table 1. The
pharmacological assay utilized to assess mGlu4 PAM activity (cal-
cium mobilization11) shows day-to-day variability in the maximal
PAM response; for this reason, all data have been normalized to the
% response of the control PAM, PHCCC, to compare relative efficacy.
Changing from the 2,4,6-trimethylphenyl sulfonamide, 5, to the
less sterically hindered phenyl sulfonamide, 8a, provided a com-
pound that was equipotent (159 nM vs 177 nM) and exhibited sim-
ilar maximal response. Evaluating other aryl substituents led to the
2-chlorophenyl sulfonamide, 8b, which provided a dramatic in-
crease in potency (27 nM, 88.2%); one of the most potent mGlu4
PAMs reported to date. However, going from the 2-chloro to 2-flu-
oro, 8c, resulted in an erosion of potency (27 nM vs 206 nM), but
minimal effect on maximal potentiation. Further substitution
around the phenyl ring resulted in less active (8d, 325 nM,
87.6%), or inactive compounds (8e and 8f) with slight modification.
Going away from arylsulfonamides to alkyl or cycloalkyl sulfona-
a
Compounds were tested using a 1:3 serial dilutions starting at 30 lM. Data
represent the average of triplicate determinations performed on one day. For %
PHCCC, the maximal response elicited by compound was divided by the response of
the control PAM, PHCCC, on that same day.
mides resulted in reduced potency and maximal effect (8g–8n).
Dialkylation of the sulfonamide led to reduced activity (to inactive)
and much reduced max values (8l–8n).
Table 2
Heteroarylamide evaluation, 9a–g
O
O
S
O
N
R1 Cl
R
N
H
9a-g
a,11
Compound
R
R1
H
EC50 (nM) hmGlu4
% PHCCCa,11
90.0
F
N
∗
9a
122
O
N
∗
9b
9c
9d
9e
9f
H
Inactive
478
8.6
102.9
79.4
54.2
31.1
25.4
N
∗
H
N
∗
∗
N
H
77.4
269
R1
O
X
O
S
HN
R2
O
S
O
S
S
R1
Cl
N
H
R2
O2N
pyridine, DCM;
S
O2N
X
6
7
67-97%
N
∗
Me
Me
990
O
O
N
N
R1
1. H2, Pd/C or Ra-Ni;
>95%
∗
O
N
N
R2
9g
686
S
N
H
X
2. Picolinoyl chloride,
DIEA, DCM
a
Compounds were tested using a 1:3 serial dilutions starting at 30 lM. Data
8a-n
represent the average of triplicate determinations performed on one day. For %
PHCCC, the maximal response elicited by compound was divided by the response of
the control PAM, PHCCC, on that same day.
Scheme 1. Synthesis of 4-sulfamoylphenylpicolinamides, 8a–n. All library com-
pounds were purified by mass-directed prep LC where required.21,22

D. W. Engers et al. / Bioorg. Med. Chem. Lett. 20 (2010) 5175–5178
5177
Table 4
Next, we focused on our best compound (2-chlorophenylsulf-
In vitro pharmacokinetic evaluation of select compounds
onamide, 8b) and evaluated alternative amide moieties (Table 2).
Based on our previous work, we chose a small heteroaryl subset
for evaluation. Small substitution on the pyridyl ring was tolerated
(9a, 6-fluoro, 122 nM, 90.0%); however, a larger group led to an
inactive compound (9b, 6-methoxy, inactive). The other six- and
five-membered heterocycles were also well-tolerated (9c–9e).
Next, having evaluated cycloalkylamines as the sulfonamide por-
tion, we wanted to evaluate simple methylated compounds. Utiliz-
ing the active 2-pyrimidine and 2-thiazole amides, the N–Me
arylsulfonamides were synthesized and evaluated and there was
a ꢀ5- to 10-fold loss of potency, as well as a dramatic decrease
in maximal response, as compared to PHCCC (9f, 990 nM, 31.1%;
9g, 686 nM, 25.4%).
Compound HLM (%
remaining)
RLM (%
remaining)
PPB (r) (%
free)
PPB (h) (%
free)
4
5
nd
nd
0.02
0.84
2.5
0.1
1.3
0.63
11.9
93.3
25.8
0.02
1.2
8.2
0.3
0.3
0.1
2.7
37
12.7
0.1
nd
2.2
0.1
0.1
0.1
1.7
0.5
0.3
0.1
nd
8b
9d
9f
9g
10
11
13
3.3
0.08
0.05
0.02
0.05
Lastly, modifications were made to the internal phenyl ring in
an attempt to improve potency and maximal response (Table 3).
3-Methoxy substitution, 10, on both the internal phenyl ring and
sulfonamide ring was well-tolerated (132 nM, 112.1%) and gave
a compound equipotent to the lead compound, 5. Substituting
a 3-chloro in the internal phenyl ring gave a compound equipo-
tent to our best compound, 8b, (11, 19.8 nM, 102.3%). Moving
the chloro to the 2-position (relative to the amide, 12) led to
an inactive compound suggesting that steric bulk ortho to the
acetamide is not tolerated. These substituents could potentially
hinder the NH group from making a key interaction in the bind-
ing site. Rearranging the orientation of the sulfonamide and
chloro group led to a 36-fold loss of potency (13, 731 nM,
54.9%). Removal of the NH group altogether and replacement
with a methylene group also led to a much less potent com-
pound (14, 237 nM, 46.3%). Finally, reversal of the methylene
and sulfone moieties led to a compound with much decreased
potency and maximal response (15, 2170 nM, 46.5%) suggesting
the need for the NH and orientation of the sulfone directly at-
tached to the internal phenyl ring.
Having a number of very potent and efficacious compounds in
hand, we next wanted to evaluate in vitro pharmacokinetic (PK)
properties. The compounds were evaluated for their metabolic sta-
bility in both human and rat microsomes, as well as their protein
binding in both species.15 As can be seen in Table 4, these com-
pounds all exhibited poor microsomal stability in both human
and rat. Although the initial screening hit, 4, showed good stability
in RLM (93.3% remaining); all other compounds tested were very
unstable with less than 30% remaining after the incubation period
of 60 min. Metabolite ID analysis showed oxidation of both the
internal phenyl ring, as well as oxidation of the pyridine ring (data
not shown). In addition, with the exception of compounds 4, 9g,
and 10, these compounds were highly protein bound in the rat,
with even less free fraction observed in the human. Due to these
disappointing in vitro PK results, these compounds were not pro-
gressed further into in vivo testing.
Table 3
Further SAR of 2-pyridylamide-4-phenylsulfamoyl compounds
Compound
Structure
EC50 (nM)
hmGlu4
%
PHCCCa,11
a,11
In summary, we report a series of small molecule mGlu4 posi-
tive allosteric modulators identified through a functional HTS lead.
These compounds represent a series of 2-pyridylamide-4-phenyl-
sulfonamide compounds that possess excellent in vitro potency
(VU0364439, 11, 19.8 nM) and maximal response relative to the
control PAM, PHCCC. Unfortunately, these compounds possess less
than ideal PK properties preventing their use as in vivo tools; how-
ever, we anticipate that these compounds will inform the mGlu4
community with more in vitro tool compounds.
O
O
O
O
S
O
N
N
10
132
112.1
102.3
H
O
N
H
O
O
S
O
N
N
H
11
19.8
Cl
N
H
Cl
O
Acknowledgments
S
O
N
N
H
12
13
14
Inactive
731
8.1
54.9
46.3
Cl
Cl
N
H
The authors thank Qingwei Luo and Rocio Zamorano for techni-
cal assistance with pharmacology assays and Emily L. Days, Tasha
Nalywajko, Cheryl A. Austin, and Michael Baxter Williams for their
critical contributions to the HTS portion of the project and Matt
Mulder, Chris Denicola, and Sichen Chang for the purification of
compounds utilizing the mass-directed HPLC system. This work
was supported by the National Institute of Mental Health, National
Institute of Neurological Disorders, and Stroke, the Michael J. Fox
Foundation, the Vanderbilt Department of Pharmacology, and the
Vanderbilt Institute of Chemical Biology.
Cl
Cl
S
O
N
H
N
N
H
O
O
O
O
S
O
N
237
Cl
Cl
N
H
References and notes
O
N
S
15
2170
46.5
O
O
N
H
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a
Compounds were tested using a 1:3 serial dilutions starting at 30 lM. Data
represent the average of triplicate determinations performed on one day. For %
PHCCC, the maximal response elicited by compound was divided by the response of
the control PAM, PHCCC, on that same day.

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