N-Heteroaryl glycinamides and glycinamines as potent NPY5 antagonists

Article abstract of DOI:10.1016/j.bmcl.2011.06.078

Subtype specific ligands are needed to evaluate the therapeutic potential of modulating the brain's neuropeptide Y system. The benzothiazepine glycinamide 1a was identified as an NPY5 antagonist lead. While having acceptable solubility, the compound was f

Full text of DOI:10.1016/j.bmcl.2011.06.078

Bioorganic & Medicinal Chemistry Letters 21 (2011) 5573–5576
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
N-Heteroaryl glycinamides and glycinamines as potent NPY5 antagonists
Lingyun Wu a, , Kai Lu a, Mahesh Desai a, Mathivanan Packiarajan a, Amita Joshi a,
⇑
Mohammad R. Marzabadi a, Vrej Jubian a, Kim Andersen a, Gamini Chandrasena a, Noel J. Boyle b,
Mary W. Walker b
a Department of Chemical & Pharmacokinetic Sciences, 215 College Road, Paramus, NJ 07652, USA
b Neuroinflammation Disease Biology Unit, Lundbeck Research USA, 215 College Road, Paramus, NJ 07652, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
Subtype specific ligands are needed to evaluate the therapeutic potential of modulating the brain’s neu-
ropeptide Y system. The benzothiazepine glycinamide 1a was identified as an NPY5 antagonist lead.
While having acceptable solubility, the compound was found to suffer from high clearance and poor
exposure. Optimization efforts are described targeting improvements in potency, microsomal stability,
and PK properties. The low microsomal stability and poor PK properties were addressed through the opti-
mization of the sulfonyl urea and replacement of the benzothiazepinone with other N-heteroaryl glyci-
namides. For example, the analogous benzoxazine glycinamide 2e has improvements in both affinity
(human Y5 Ki 4 nM vs 1a 27 nM) and microsomal stability (human CLint 2.5 L/min vs 1a 35 L/min). How-
ever the brain penetration (B/P 43/430 nM at 10 mg/kg PO) remained an unresolved issue. Further opti-
mization by decreasing the hydrogen bond donating properties and PSA provided potent and brain
penetrant NPY5 antagonists such as 5f (human Y5 Ki 9 nM, B/P 520/840 nM 10 mg/kg PO).
Ó 2011 Elsevier Ltd. All rights reserved.
Received 3 June 2011
Accepted 17 June 2011
Available online 25 June 2011
Keywords:
NPY5
N-Aryl glycinamide
Microsomal stability
Brain penetration
Neuropeptide Y (NPY) is a 36 amino acid neuropeptide widely
expressed in the central and peripheral nervous systems.1 It has
been shown that NPY influences a diverse array of biological func-
tions including blood pressure, food intake, circadian rhythms, and
stress sensitivity.2–4 NPY reportedly exerts its effects through the
interaction with specific receptors of the G-protein coupled recep-
tor (GPCR) superfamily. Five NPY receptor subtypes have been
identified to date (Y1, Y2, Y4, Y5, and Y6).5 The NPY5 receptor is
well known for its role in appetite, and there are multiple studies
supporting its potential utility in treating obesity.6–8 Based on its
expression in the limbic system, we hypothesized that the NPY5
receptor might also modulate stress sensitivity.9 In order to better
understand the function of the NPY5 receptor, selective agonists
and antagonists are required. Many NPY5 chemotypes have been
reported10 and two compounds, MK0557 and Velneperit, from
Merck and Shionogi, respectively, have advanced to human clinical
trials. Both compounds induced statistically significant weight loss
in obese subjects.11,12 In our attempts to identify novel antagonist
chemotypes for the NPY5 receptor we identified through screening
the moderately potent benzothiazepinone glycinamide 1a (Fig. 1).
Further profiling showed that 1a is not stable in microsomal incu-
bations and has poor pharmacokinetic properties. However, com-
pound 1a represents an attractive lead with good solubility
report our work to overcome the clearance and PK liabilities culmi-
nating in the discovery of a new class of structurally novel hetero-
cyclic glycinamides and their reduced analogs as NPY5 antagonists.
The compounds presented in this paper were synthesized fol-
lowing the methodology as described in Schemes 1 and 2. The ben-
zothiazepine and benzothiazepinone analogs were synthesized as
outlined in Scheme 1. The condensation of 2-aminothiophenol
with a methacrylate afforded benzothiazepinones (10), which
could be reduced to the benzothiazepines (11).13 Alkylation with
a-bromomethyl acetate followed by hydrolysis gave the carboxylic
acids (13), which may be oxidized to the sulfoxides (14) or sulfones
(15). The amines (21) were prepared from the commercially
Cl
H
H
N
N
N
N
H
N
H
N
S
O
O
O
N
S
O
S
O O
O O
2e
1a
, Y5 Ki = 27 nM
F
rCLint = 230 mL/min
hCLint = 35 L/min
F
F
H
Brain (1hr, 1 mg/kg, PO) 0 ng/g
Plasma (1 hr, 1 mg/kg, PO) 0.75 ng/mL
N
N
H
(Sol7.4 = 440 lg/mL) and an optimal Log D = 2.4. In this letter, we
O
N
S
O O
5f
⇑
Corresponding author. Tel.: +1 201 350 0362; fax: +1 201 261 0623.
E-mail address: lwu@lundbeck.com (L. Wu).
Figure 1. Lead optimization of NPY5 antagonist 1a.
0960-894X/$ - see front matter Ó 2011 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2011.06.078

5574
L. Wu et al. / Bioorg. Med. Chem. Lett. 21 (2011) 5573–5576
Table 1
O
SAR of benzothiazepine and benzothiazepinone glycinamides
OEt
Y
H
N
Y
N
R2
O
OMe
a
NH2
SH
c
+
R1
H
N
R1
O
R1
S
S
R2
R2
8
9
N
12
H
X
R2
O
N
R3
10
11
Y = O
Y = H2
b
Y
S
R1
O O
OH
Y
H
N
b,c
b,d
Compd
R1, R2
R3
X
Y
hY5, Kia
(nM)
hCLint
rCLint
N
X
g
d
N
H
(L/min)
(mL/min)
X
O
Y
N
R3
R2
R1
S
1a
1b
1c
1d
1e
1f
Me, Me
H, H
NMe2
NMe2
Me
Et
cPr
iPr
S
S
S
S
S
S
S
O
O
O
O
O
O
H2
H2
27
420
1900
200
210
19
35
8
—
—
—
23
10
38
230
160
—
—
—
420
56
430
R1
R2
O O
1
13
14
X = S
X = SO
15 X = SO2
e
f
Me, Me
Me, Me
Me, Me
Me, Me
Me, Me
Me, Me
O
O
MeO
HO
1g
1h
iPr
iPr
6.9
23
H
N
X
Y
d
N
X
Y
N
X
Y
g
SO2
c
R
R
R
Binding affinity was determined using a [125I]PYY radioligand binding assay to
a
16
17
18
membranes from cells expressing human NPY5 receptor.9 Data shown as average of
two or more assays.
b
H
N
A 1 lM test compound in either rat or human (pooled) liver microsomes
R
2
3
X = O
X = S
N
(0.5 mg/mL) incubated at 37 °C for 0, 5, 15, 30, 60 min with NADPH regenerating
system. Following incubation, the acetonitrile quenched reaction samples were
centrifuged to afford the supernatant for LC/MS analysis to determine the remained
drug concentrations for T1/2 estimation. The clearances were estimated according to
Ref. 15.
H
N
X
O
R3
Y
S
4 X = CH2
O O
H2N
c
HOOC
HOOC
Maximum human liver blood flow = 1.5 L/min.
Maximum rat liver blood flow = 20 mL/min.
i, j
h
H
N
H
d
R3
N
R3
NH2
S
S
O O
O O
19
20
21
Scheme 1. Reagents and conditions: (a) microwave 220 °C13 15–20%; (b) LiAlH4,
of representative compounds (1a–f) is outlined in Table 1. Not
surprisingly, removal of the gem-dimethyl group improved human
microsomal stability albeit at the expense of potency (1b). Replace-
ment of the sulfonyl urea with a methanesulfonamide (1c) resulted
in a dramatic loss of potency. However, increasing the steric bulk of
the sulfonamide restored potency ( 1d, 1e, 1f). Reduction of the
benzothiazepinone (1f) to a benzothiazepine (1g) improved both
potency and microsomal stability while oxidation to a sulfone neg-
atively impacted these properties (1h). Although the potency and
stability of 1a could be improved, the microsomal stability was still
unsatisfactory. We envisioned that replacement of the lipophilic
benzothiazepinone ring system with six-membered N-substituted
heterocycles would improve stability. The isopropyl sulfonamide
(R3) was kept constant in this analog series as SAR studies in a re-
lated chemotype demonstrated the value of this moiety to the
affinity for the receptor (unpublished results).
THF, reflux, 90%; (c) -bromo methyl acetate, TBAB, KOH, THF; (d) LiOH, MeOH, 30–
a
50% for two steps; (e,f) mCPBA, 10–30%; (g) compound 21, EDCI, DMAP, DCM, 10–
50%; (h) R3SO2Cl, NaOH, H2O, 5 °C or rt 30–50%, (i) DPPA, Et3N, BnOH/toluene,
reflux, 80–90%; (j) Pd/C, H2, EtOH, 95%.
O
O
H
N
Cl
R
H
N
N
X
N
b
H
N
a
R
R
X
S
X
22
O O
23
24
H
R
5
X = O
X = S
N
c, d
N
6
H
X
N
7 X = CH2
S
O O
Scheme 2. Reagents and conditions: (a)
a-chloro acetyl chloride, Et3N, DCM; (b)
The binding and in vitro clearance data of the N-heteroaryl gly-
cinamides are summarized in Table 2. The key replacement of the
benzothiazepinone with a benzoxazinone isostere having a lower
c Log D7.4 showed retention of affinity (2a). Fluorine substitution
at the 4-position (2b) reduced affinity while enhancing microsomal
stability relative to 1f. Fluorine substitution at the 5-position (2c)
improved both affinity and stability. Affinity was further enhanced
with a 5-chloro substituent (2d), at the expense of lower human
microsomal stability, consistent with higher lipophilicity. Reduc-
tion of the benzoxazinone (2d) to the benzoxazine (2e) was well
tolerated. Replacement of the benzoxazinone with benzothiazinone
(3a) and benzothiazine (3b) generated very potent compounds but
with poor microsomal stability. Oxidation of the metabolically la-
bile sulfide (3b) to sulfone (3c) increased the microsomal stability
while maintaining the binding affinity. The dihydroquinolinone
4a and dihydroquinoline 4b were also found to be potent Y5 antag-
onists with improved microsomal stability versus 1f.
compound 21, K2CO3, NaI, EtOH, reflux, 30–50% for two steps; (c) BH3ÁTHF, THF,
reflux; (d) HCl, reflux, 10–70% for two steps.
available amino acid (19). Treatment with a sulfonyl chloride or
sulfamyl chloride generated the carboxylic acids (20), which were
converted via Curtius rearrangement to the corresponding amines
(21). Coupling of amines (21) with the carboxylic acids (13–15)
generated the target compounds (1a–h). The synthesis of N-het-
eroaryl glycinamides (2–4) involved the same synthetic methodol-
ogy as for 1. Starting from the commercial available or known
starting materials (16), the glycinamides (2–4) were obtained with
moderate overall yield.
The glycinamines (5–7) were prepared from benzazines (22)14
as shown in Scheme 2. Treatment of 22 with a-chloroacetyl chlo-
ride afforded the intermediates 23, which were subsequently
heated with amines (21) in the presence of potassium carbonate
to give amides (24). Reduction with borane resulted in the target
molecules (5–7).
Our initial SAR efforts focused on modifying the benzothiazepi-
none and sulfonyl urea moieties of the lead molecule 1a. The SAR
Selected compounds were evaluated for their in vivo pharmaco-
kinetic properties. The plasma and brain levels measured 1 h after
dosing at 10 mg/kg PO are listed in Table 3. While the six-
membered N-heterocyclic analogs (2d, 3c, 4a) showed improved

L. Wu et al. / Bioorg. Med. Chem. Lett. 21 (2011) 5573–5576
5575
Table 2
Table 4
SAR of N-heteroaryl glycinamides 2, 3, and 4
SAR of N-heteroaryl glycinamines 5, 6, and 7
H
H
N
R
R
N
N
N
H
N
H
N
X
X
O
Y
S
S
O O
O O
b,c
b,d
hY5Kia (nM)
c Log D7.4
hCLint
rCLint
b,c
b,d
Compd
R
X
Y
hY5 Kia (nM) c Log D7.4 hCLint
rCLint
Compd
X
R
(L/min)
(mL/min)
(L/min)
(mL/min)
5a
5b
5c
5d
5e
5f
6
O
O
O
O
O
O
S
CH2
4-F
170
110
88
60
14
9.2
370
4.4
3.5
3.6
3.2
4.1
4.1
4.4
4.6
4.7
—
—
—
—
1.5
2.6
—
—
—
—
—
32
7.6
—
1f
—
H
—
O
O
O
O
O
S
—
O
O
O
O
H2
O
H2
19
21
97
3.3
1.6
1.9
1.9
2.5
3.4
2.8
4.0
2.2
2.7
4.1
23
—
420
—
4,5-F
5-OMe
4-Cl
5-Cl
5-CF3
5-Cl
2a
2b
2c
2d
2e
3a
3b
3c
4a
4b
4-F
5-F
5-Cl
5-Cl
5-Cl
5-Cl
5-Cl SO2 H2
5-Cl CH2
5-Cl CH2 H2
1.7
0.6
2.4
2.5
4.0
6.1
1.9
0.8
5.1
32
11
17
28
140
75
23
40
85
8.4
1.0
3.5
0.8
0.6
1.8
3.4
0.9
7
5-Cl
2.1
31
S
Binding affinity was determined using a [125I]PYY radioligand binding assay to
a
O
membranes from cells expressing human NPY5 receptor.9 Data shown as average of
two or more assays.
b
Binding affinity was determined using a [125I]PYY radioligand binding assay to
a
See Table 1 for protocols and reference.
Maximum human liver blood flow = 1.5 L/min.
Maximum rat liver blood flow = 20 mL/min.
c
membranes from cells expressing human NPY5 receptor.9 Data shown as average of
two or more assays.
d
See Table 1 for protocols and reference.15
b
c
Maximum human liver blood flow = 1.5 L/min.
Maximum rat liver blood flow = 20 mL/min.
d
C-5 position (5f) afforded a compound with improved potency
and similar microsomal stability. Based on its potency compound
5f was advanced into pharmacokinetic studies. We were pleased
to discover that 5f has comparable plasma exposure to the previous
glycinamide compounds but with improved overall brain exposure
(Table 3).
In summary, the novel benzothiazepinone NPY5 antagonist lead
1a was identified through a screening program. The low micro-
somal stability and poor PK properties were addressed through a
systematic SAR study to investigate the value of this chemotype
as potential antagonists. Optimization of the sulfonyl urea and
replacement of the benzothiazepinone with other N-heteroaryl
glycinamides resulted in improvements in affinity and microsomal
stability. However brain penetration remained an unresolved issue.
Further optimization of the aryl moiety provided potent and brain
penetrant NPY5 antagonists such as 5f, which have allowed for
continued understanding of the pharmacology of this receptor.
The details of the potential of these analogs for the treatment of
various mood and anxiety disorders will be the subject of future
communications.
Table 3
Plasma and brain exposure, 10 mg/kg, PO, 1 ha
Compound
1a
2d
33
481
0.07
2e
43
430
0.10
3c
7.5
28
0.27
4a
16
879
0.02
5f
Brain (nM)
Plasma (nM)
B/P
0
520
840
1.5
0
0.62
a
Femoral artery cannulated Sprague–Dawley male rats were purchased from
Taconic Laboratories. The average body weight was 300–350 g. Each compound that
was dissolved in 20% 2-hydroxypropyl-b-cyclodextrin solution was orally admin-
istered to three rats (N = 3) at a dose of 10 mg/kg. Blood samples were collected at
four time points: predose (0 h), 1 h, 2 h, and 4 h. The rats were then sacrificed, and
the brain tissues were collected and immediately stored at À80 °C. Plasma samples
were obtained by centrifuging the blood samples. To increase bioanalysis
throughput, an equal amount of N = 3 plasma or brain samples at the same time
point, which were collected from three different rats, were pooled together. Each
pooled rat brain sample was homogenized in an aqueous solution. Protein precip-
itation of the pooled plasma or homogenized brain sample afforded a supernatant
that was analyzed by LC/MS/MS, an Agilent 1100 HPLC (Agilent Technologies, Palo
Alto, CA) and a TSQ Quantum MS (ThermoFinnigan, San Jose, CA). Compound
concentrations in the plasma and brain matrices were quantified using Thermo-
Finnigan Xcalibur.
Acknowledgements
We thank Manuel Cajina, Asanthi Pieris, and Martha Vallejo for
providing bioanalysis, PK and metabolic stability data, respectively.
In addition Chi Zhang for providing Phys-Chem data, and John Pet-
erson and Andrew White for helpful suggestions.
exposure relative to 1a, the brain to plasma (B/P) ratios are low.
Various factors may contribute to the low brain penetration.16,17
We postulated that reduction of the glycinamide carbonyl would
enhance blood brain barrier permeability by lowering the PSA
(e.g., TPSA dropped from 88 to 71 by reducing 2e to 5e)18 and
decreasing the hydrogen bond donating properties of the amide
N–H,19 a strategy which has been shown to improve central expo-
sure in other systems.20
The N-heteroaryl glycinamines 5–7 (Table 4) were less potent
than the analogous glycinamides, but with comparable microsomal
stability (5e vs 2e, 7 vs 4b). Surprisingly, the benzothiazine 6 was
much less potent than the corresponding benzoxazines and dihy-
droquinolines, presumably the larger sulfur atom changes the SAR
of the phenyl substitution in the more conformationally flexible
glycinamine series. Substitution at the C-5 position (5e) is preferred
over C-4 substitution (5d) suggesting that bulky and electron with-
drawing groups at the C-5 position are optimal for the glycinamine
series possibly due to their hydrophobic interaction with the bind-
ing pocket and impact on the basicity. Introduction of a CF3 at the
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