Pyrimido[4,5-d]azepines as potent and selective 5-HT2C receptor agonists: Design, synthesis, and evaluation of PF-3246799 as a treatment for urinary incontinence

Article abstract of DOI:10.1016/j.bmcl.2010.11.120

New pyrimido[4,5-d]azepines 7 are disclosed as potent 5-HT_2C receptor agonists. A preferred example, 7b had minimal activation at either the 5-HT_2A or 5-HT_2B receptors combined with robust efficacy in a preclinical canine model of stress urinary incontinence (SUI) and attractive pharmacokinetic and safety properties. Based on this profile, 7b (PF-3246799) was identified as a candidate for clinical development for the treatment of SUI. In addition, it proved to be critical to build an understanding of the translation between recombinant cell-based systems, native tissue preparations and in vivo preclinical models. This was a significant undertaking and proved to be crucial in compound selection.

Full text of DOI:10.1016/j.bmcl.2010.11.120

Bioorganic & Medicinal Chemistry Letters 21 (2011) 2715–2720
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Pyrimido[4,5-d]azepines as potent and selective 5-HT2C receptor agonists:
Design, synthesis, and evaluation of PF-3246799 as a treatment
for urinary incontinence
Mark D. Andrews a, , Paul V. Fish a, , Julian Blagg a, Tiffini K. Brabham b, Paul E. Brennan a, Alison Bridgeland c,
Alan D. Brown a, Peter J. Bungay d, Kelly M. Conlon c, Nicholas J. Edmunds c, Kerry af Forselles c,
Colleen P. Gibbons e, Martin P. Green a, Giles Hanton b, Mark Holbrook b, Alan S. Jessiman a, Karin McIntosh f,
Gordon McMurray c, Carly L. Nichols a, James A. Root c, R. Ian Storer a, Michael R. Sutton b, Robin V. Ward c,
Dominique Westbrook c, Gavin A. Whitlock a
⇑
⇑
a Worldwide Medicinal Chemistry, Pfizer Global Research and Development, Sandwich Laboratories, Sandwich, Kent CT13 9NJ, UK
b Drug Safety Research & Development, Pfizer Global Research and Development, Sandwich Laboratories, Sandwich, Kent CT13 9NJ, UK
c Genitourinary Biology, Pfizer Global Research and Development, Sandwich Laboratories, Sandwich, Kent CT13 9NJ, UK
d Pharmacokinetics, Dynamics & Metabolism, Pfizer Global Research and Development, Sandwich Laboratories, Sandwich, Kent CT13 9NJ, UK
e CVMED Biology, Pfizer Global Research and Development, Groton laboratories, 558 Eastern Point Road, Groton, CT 06340, USA
f Pharmaceutical Sciences, Pfizer Global Research and Development, Sandwich Laboratories, Sandwich, Kent CT13 9NJ, UK
a r t i c l e i n f o
a b s t r a c t
Article history:
New pyrimido[4,5-d]azepines 7 are disclosed as potent 5-HT2C receptor agonists. A preferred example, 7b
had minimal activation at either the 5-HT2A or 5-HT2B receptors combined with robust efficacy in a pre-
clinical canine model of stress urinary incontinence (SUI) and attractive pharmacokinetic and safety
properties. Based on this profile, 7b (PF-3246799) was identified as a candidate for clinical development
for the treatment of SUI. In addition, it proved to be critical to build an understanding of the translation
between recombinant cell-based systems, native tissue preparations and in vivo preclinical models. This
was a significant undertaking and proved to be crucial in compound selection.
Received 21 October 2010
Revised 23 November 2010
Accepted 29 November 2010
Available online 4 December 2010
Keywords:
Pyrimido[4,5-d]azepines
5-HT2C receptor agonists
PF-3246799
Ó 2010 Elsevier Ltd. All rights reserved.
Urinary incontinence
The neurotransmitter serotonin (5-HT) mediates its effects
through at least 14 different receptor subtypes that have been clas-
sified into seven major families, 5-HT1–7. The 5-HT2 family has
three members 2A, 2B, and 2C and, unlike 5-HT2A and 5-HT2B
receptors, the expression of 5-HT2C receptors appears to be re-
stricted to the central nervous system (CNS). 5-HT2C receptor ago-
nists have become attractive drug targets that have potential use in
the treatment of a number of conditions including obesity, psychi-
atric disorders, sexual dysfunction, and urinary incontinence.1
Selectivity over agonism at the 5-HT2A and 5-HT2B receptors would
be a key objective because 5-HT2A agonists can potentially be
hallucinogenic and have cardiovascular (CV) effects,2 whereas
5-HT2B agonism has been associated with heart valvulopathy and
pulmonary hypertension.3
The search for potent and selective 5-HT2C agonists has identi-
fied lorcaserin (1) (APD-356; Arena) which has completed phase 3
clinical trials for the treatment of obesity4 and vabicaserin (2)
(SCA-136; Pfizer) as a potential therapy for schizophrenia.5 Further-
more, several small molecule 5-HT2C agonists have been reported to
be in early clinical development or undergoing preclinical optimi-
zation and evaluation.6
Me
H
N
Cl
H
NH
N
H
1: APD-356
2: SCA-136
lorcaserin
vabicaserin
⇑
Corresponding authors. Tel.: +44(0)1304 648831 (M.D.A.), +44(0)1304 644589
(P.V.F.).
At Pfizer, we have disclosed several new templates as 5-HT2C
receptor agonists7–14 and some of these compounds have now pro-
gressed to clinical trials. As part of our research efforts to identify
E-mail addresses: mark.d.andrews@pfizer.com (M.D. Andrews), paul.fish@pfi-
zer.com (P.V. Fish).
0960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.11.120

2716
M. D. Andrews et al. / Bioorg. Med. Chem. Lett. 21 (2011) 2715–2720
Table 1
potential new 5-HT2C agonist drug candidates, we adopted a strat-
egy of exploring multiple chemical templates in order to increase
our chances of having compounds survive to become advanced
clinical candidates. In this Letter, we disclose pyrimido[4,5-d]-
azepines (7) as potent and selective 5-HT2C agonists.
5-HT2C activity for compounds 5–7a,b,c
Compound
x
5-HT2C
EC50 (nM)
Emax (%)
5a
5b
5c
0
1
2
3
—
—
—
Inactive
64d
NH
NH
N
N
Aromatic tethered heterocyclic piperazines 3 have been identi-
fied as potent 5-HT2C agonists.7a However, some compounds in this
series were shown to be mutagenic in the Ames assay and mecha-
nistic studies proposed bioactivation of the piperazine ring as the
likely cause.7b In order to try and eliminate the activity in the Ames
assay, whilst retaining excellent 5-HT2C agonist potency, we sought
to remove the piperazine substituent and replace it with a fused
six- or seven-membered ring containing a basic amine group
(Fig. 1). Although benzazepines 4 were known to be active as
5-HT2C ligands, we wished to retain a heterocyclic azine core in
order to minimize the lipophilicity as it was anticipated that this
would lead to improved drug-like properties. The tetrahydro-pyri-
dopyrimidines 5, 6 and tetrahydro-pyrimidoazepines 7 were then
selected based on anticipated ease of synthesis.
50d
100
( )x
N
N
5d
1900
6a
6b
6c
0
1
2
3
380
—
92
27d
—
—
Inactive
Inactive
( )x
6d
7a
7b
7c
0
1
2
—
4.5
Inactive
95
N
NH
110
99
( )x
N
a
See Ref. 15 for complete details of assay conditions.
Values (EC50, Emax) are geometric means of 2–4 experiments. Differences of
b
<2-fold should not be considered significant.
c
5-HT2C high receptor expression cell-line (see text).
% Activation at 10 lM.
d
Given the structural changes relative to the starting point 3 we
were unsure what length of linker would be required between the
pyrimidine and the pendant aromatic ring and so compounds were
synthesised with a directly attached phenyl ring (5–7: x = 0) and
with 1 to 3 methylene spacers (5–7: x = 1–3).
5-HT2C potency (EC50 1900 and 380 nM respectively) in the high
expression cell-line (Table 1). However, this activity almost disap-
peared in the low expression cell-line with these compounds only
The 5-HT2C agonist activity of target compounds 5–7 (Tables 1
and 2) was evaluated by measuring the ability to induce a fluores-
cence based calcium mobilization signal in a FLIPR assay employ-
ing recombinant CHO K1 cells expressing the human 5-HT2C
receptor.15 Midway through our research program it proved neces-
sary to re-express this cell-line with lower 5-HT2C receptor density
to give better translation between in vitro and in vivo outcomes;
these have been designated as ‘high’ and ‘low’ receptor expression
cell-lines respectively. As would be anticipated, the high expres-
sion cell-line was more highly amplified and showed compounds
to be more potent (EC50) with higher intrinsic activity (Emax) ago-
nists than the low expression cell-line. Agonist activity at the
5-HT2B receptor was measured in a similar manner with recombi-
nant cell-based systems expressing the human 5-HT2B receptor.
Selected compounds were also tested for their ability to inhibit
binding of [3H]-meselurgine at the human 5-HT2C receptor utiliz-
ing SPA technology and cellular membrane preparations generated
from recombinant Swiss 3T3 cells.15
showing weak activation of the 5-HT2C receptor at 10 lM. Results
with the pyrimidoazepines 7 were much more encouraging, with
both benzyl (7b: x = 1, EC50 = 4.5 nM) and phenethyl substituents
(7c: x = 2, EC50 = 110 nM) giving significant agonist activity in the
high expression cell-line. Once again there was a drop-off in activ-
ity when 7b and 7c were screened in the low expression cell-line
with 7c becoming a weaker partial agonist (EC50 4800 nM, Emax
40%). However, 7b was found to retain both good potency and
intrinsic activity (EC50 190 nM, Emax 75%) which identified azepine
7b as a preferred template for optimization from this set. Further-
more, 7b was determined to be negative in a panel of in vitro
genetic toxicology studies (Ames, micronucleus) either with or
without metabolic activation.
At the outset of our work, there were very few reports of
pyrimido[4,5-d]azepines 7 and these were restricted to the patent
literature for targets other than 5-HT2C receptor agonists.16 Hence,
we initiated a lead-to-candidate research program to explore the
structure-activity relationships (SAR) of 7 with the objective of
seeking potent 5-HT2C agonists with minimal activity at either
the 5-HT2A or 5-HT2B receptors. Furthermore, target compounds
were designed to have drug-like properties consistent with CNS
target space.17
Initial results with the tetrahydro-pyridopyrimidines 5 and 6
were disappointing, with only 5d and 6a showing some modest
N
Pyrimido[4,5-d]azepine target compounds 7 were prepared
using a short 4-step synthesis as described in Scheme 1. Condensa-
tion of phenylacetamidines 8 with azepine ketoester 9,18 followed
by cyclization, created the 4-pyrimidones 10 which then under-
went chlorination to the corresponding 4-chloropyrimidines 11
by treatment with POCl3. Dechlorination of 11 with zinc19 gave
pyrimidines 12 and finally deprotection of the azepine N-benzyl
protecting group gave 7. For some examples, it was possible to con-
vert 11 directly to 7 by hydrogenolysis.
R
NH
N
N
O
R
NH
3: Piperazines
4: Benzazapines
e.g. CP-809101: R = 3-Cl
The SARs were initially focused on exploring substituents on the
phenyl ring of 7 with respect to activation of the 5-HT2C and 5-HT2B
receptors (Table 2). A range of groups were tolerated at either the
2-, 3-, or 4-positions with only the 2-Cl (7j) and 2-CF3 (7m) losing
5-HT2C activity compared to 7b (R = H). Although 5-HT2C agonist
activity could be retained with a number of groups, these substit-
uents invariably introduced an unacceptable level of 5-HT2B activa-
tion with some compounds becoming more active at the 5-HT2B
receptor (7k,l). Hence, the initial lead 7b emerged as having a
superior profile from this set.
( )n
NH
( )m
N
R
( )x
N
5: 5,6,7,8-tetrahydropyrido[4,3-d]pyrimidines, m = 1; n = 0
6: 5,6,7,8-tetrahydropyrido[3,4-d]pyrimidines, m = 0; n = 1
7: 6,7,8,9-tetrahydro-5H-pyrimido[4,5-d]azepines, m = n = 1
Figure 1. Design of pyrimidine targets 5–7.

M. D. Andrews et al. / Bioorg. Med. Chem. Lett. 21 (2011) 2715–2720
2717
Table 2
5-HT2C and 5-HT2B activity for compounds 1 and 7a,b,c
N
NH
R
N
7
Compound
R
c log P
5-HT2C
5-HT2C
5-HT2B
EC50 (nM)
Emax (%)
Ki (nM)
EC50 (nM)
Emax (%)
APD-356 (1)d
—
H
2-F
3-F
3.2
1.6
1.8
1.8
1.8
2.1
2.1
2.1
2.3
2.3
2.3
2.5
2.5
210
190
194
125
310
330
210
100
—
84
75
65
67
60
70
84
63
38e
56
65
15e
44
167
160
NTf
NT
470
305
244
NT
1850
84
NT
NT
308
—
—
60–70e
33–50e
43
66
90
56
62
80
36
7b
7d
7e
7f
7g
7h
7i
7j
7k
7l
220
183
610
1200
410
164
178
30
4-F
2-Me
3-Me
4-Me
2-Cl
3-Cl
4-Cl
2-CF3
3-CF3
180
192
—
50
60
—
160
62
7m
7n
15e
63
300
a
b
c
See Ref. 15 for complete details of assay conditions.
Values (EC50, Emax, Ki) are geometric means of 2–4 experiments. Differences of <2-fold should not be considered significant.
5-HT2C low receptor expression cell-line (see text).
Data are presented for comparison in a common assay format. Literature values for APD-356 (1): h5-HT2C EC50 9 nM, Emax 100%, Ki 15 nM; h5-HT2B EC50 943 nM, Emax
d
100%. See Ref. 4b.
e
% Activation at 10 lM.
NT denotes not tested.
f
EtO2C
NH
N
R
+
NH2.HCl
Ph
O
8
9
a
Cl
O
b
N
HN
N
N
R
R
Ph
Ph
N
N
11
10
c
f
d or e
N
N
N
NH
R
R
Ph
N
N
12
7
Scheme 1. General synthesis of pyrimidines 7. Reagents and conditions: (a) NaOEt, EtOH, reflux, 18 h; (b) POCl3, Et4N+Clꢀ, MeCH2CN, 100 °C, 18 h; (c) Zn metal, THF, concd
NH3 (aq), reflux, 6 h; (d) For R – Cl: NH4þHCO2ꢀ, Pd/C, MeOH, reflux, 6 h; (e) for R = Cl: (i) 1-chloroethyl chloroformate, 1,8-bis(dimethylamino)naphthalene, CH2Cl2, rt, 18 h;
(ii) MeOH, reflux, 2 h; (f) H2 (50 psi), Pd/C, EtOH, 50 °C.
Compound 7b was then evaluated in additional in vitro phar-
macokinetic and pharmacology screens (Table 3). Azepine 7b has
good metabolic stability in both HLM and human hepatocytes con-
sistent with low predicted clearance, weak CYP450 enzyme inhibi-
tion and good permeability. There was no evidence for recognition
and efflux by the P-glycoprotein (P-gp) transporter as measured by
transit performance in the MDCK-MDR1 cell-line. Compound 7b
had low ion channel activity as measured by binding to represen-
tative potassium, sodium and calcium channels.
Pharmacological evaluation for activity at the 5-HT2A receptor
was measured in a FLIPR assay employing Swiss 3T3 cells express-
ing the recombinant human 5-HT2A receptor, in vitro tissue prepa-
rations with canine femoral artery, and in vivo models.20 Screening
of 7b in the 5-HT2A recombinant assay gave a significant response
with an EC50 68 nM and Emax 82% (Table 3). However, our experi-
ence has taught us that this was a highly expressed/coupled
cell-line which over estimates 5-HT2A activity and that the tissue
preparation assay was a better predictor of in vivo outcomes.

2718
M. D. Andrews et al. / Bioorg. Med. Chem. Lett. 21 (2011) 2715–2720
Table 3
Physicochemical properties, in vitro activation of 5-HT2 receptors, ADME profiles and
ion channel binding affinities of 7ba
7b
Physicochemical properties
mol wt
c log P
log D7.4
TPSA (Å2)
pKa
239
1.6
0.2
38
8.5
5-HT2 receptor activationb
2C: EC50 (Emax
2C: EC50 (Emax
2C: Ki
)
)
4.5 nM (95%)c
190 nM (75%)d
160 nM
2A: EC50 (Emax
)
68 nM (82%)c
2B: EC50 (Emax
)
33–50% @ 10 l
Mc
Figure 2. Effect of 5-HT (h, n = 32), norfenfluramine (
D
, n = 6), APD-356 (1)( ,
r
ADME profile
n = 7), PF-3246799 (7b)(s, n = 12) and DMSO (ꢁ, n = 8) in human colon as a
e
HLM, Clint
(l
l/min/mg)
<7
<5
measure of 5-HT2B activity.
l/min/106 cells)
e
H.hepatocytes, Clint
(
l
CYP1A2 inhibition, IC50 (nM)
CYP2C9 inhibition, IC50 (nM)
CYP2C19 inhibition, IC50 (nM)
CYP2D6 inhibition, IC50 (nM)
CYP3A4 inhibition, IC50 (nM)
PAMPA, Papp ꢁ 10ꢀ6 cm sꢀ1
>30,000
>30,000
>30,000
>30,000
>30,000
28
Table 4
Rat and dog pharmacokinetic data for 7b
Rat
Dog
MDCK-MDR1, AB/BA Papp ꢁ 10ꢀ6 cm sꢀ1
29/35
Liver microsomes, Clint
Hepatocytes, Clint
Plasma protein binding (%)
(l
l/min/mg)
28
15
0
<7
<5
0
(l
l/min/106 cells)
Ion channel binding affinities
K+ hERG, IC50 (nM)
>22,300
>10,000
>10,000
>10,000
>10,000
Na+ site 2, IC50 (nM)f
Intravenous dose (mg/kg)
1.0a
2.8
51
0.030c
3.4
11.6
3.5
Ca2+ L-site (diltiazem), IC50 (nM)f
Ca2+ L-site (verapamil), IC50 (nM)f
Ca2+ L-site (DHP), IC50 (nM)f
Elimination half-life, T1/2 (h)
Plasma clearance, Cl (ml/min/kg)
Volume of distribution, Vd (l/kg)
8.9
a
Oral dose (mg/kg)
Cmax (ng/ml)
Tmax (h)
AUC0?1 (ng/h/ml)
Bioavailability, Fo (%)
2b
49
0.25
290
44
—
—
—
—
—
See Ref. 28 for definition of terms.
5-HT2 receptor activation EC50 and Emax values are geometric means of at least
b
four experiments. Differences of <2-fold should not be considered significant.
c
High receptor expression cell-line (see text).
Low receptor expression cell-line (see text).
Intrinsic clearance, Clint
Data from Cerep, <20% I @ 10,000 nM.
d
e
a
b
c
Single intravenous bolus administration to male CD rat (n = 1).
Single oral administration to male CD rat (n = 2).
Single intravenous infusion over 1 h to female beagle dog (n = 2).
f
Evaluation of 7b in the canine femoral artery gave a much weaker
response of EC50 3400 nM and Emax 51%. Screening of 7b in the rat
head-twitch model21 at 10 mg/kg (po, n = 8) gave no response and
7b had no significant effect on blood pressure or heart rate during a
CV assessment in an anaesthetized dog model up to 0.5 mg/kg (iv
infusion over 60 min, n = 4); hence, we concluded that 7b had no
significant 5-HT2A activity.
Compounds with weak activity in the 5-HT2B recombinant cell-
based assay were then screened for 5-HT2B activity in in vitro tis-
sue preparations with human colon.22–24 Compound 7b was
screened alongside APD-356 (1) with 5-HT and norfenfluramine
acting as positive controls (Fig. 2). Compound 7b showed minimal
activation of the 5-HT2B receptor compared to 5-HT and norfenflur-
amine and was significantly less active than APD-356 (1) in this
intravenous administration to dog, plasma clearance of 7b was
low relative to liver blood flow resulting in an elimination half-life
of 3.4 h. These results established that 7b had pharmacokinetic
properties compatible with evaluation in rodent and canine mod-
els of disease.
The metabolic fate of 7b was investigated in HLM. The major
metabolic routes were identified as N-hydroxylation of the azepine
group to give hydroxylamine 13 and p-hydroxylation of the phenyl
ring to give 14 (Fig. 3). Several minor metabolites were due to
mono-oxidation of the phenyl ring and benzylic methylene group.
The metabolism of 7b was investigated in individual CYP450 re-
combinant enzymes which identified that CYPs 2D6, 2B6, 2C8,
and 2C19 are likely to be the major CYP450 enzymes contributing
to the CYP-mediated clearance of 7b.
model. On average, 7b demonstrated <20% activation at 10
hence, we concluded that 7b had minimal 5-HT2B activity.
lM;
Compound 7b was screened for off-target pharmacology
against a panel of 110 receptors, enzymes and ion channels (CEREP,
Pharmacokinetic data for 7b was generated in vivo in rat and
dog (Table 4). Following single intravenous administration of 7b
to rat, plasma clearance was moderate relative to liver blood flow
and volume of distribution was moderate resulting in an elimina-
tion half-life of 2.8 h. There was also a renal component to clear-
ance with approx. 10% of parent being excreted unchanged in
urine. Following single oral administration to rat, 7b was rapidly
absorbed and showed moderate oral bioavailability (40%) suggest-
ing complete absorption from the gut, based on blood clearance of
51 ml/min/kg and an assumed liver blood flow of 70 ml/min/kg.
Good CNS exposure was confirmed in rat (10 mg/kg po, t =
30 min, n = 8) with free drug levels in brain similar to free drug lev-
els in plasma ([brain]free/[plasma]free = 0.6). Following single
Bioprint™) and was found to have binding affinities of IC50 >2.5
except for the muscarinic (M1/3–5 0.3–1.2 M), 5-HT3 (2.3 M) and
5-HT6 (1.8 M) receptors. Further evaluation in functional assays
showed no agonist activation of the hM1/4/5 receptor at concentra-
lM
l
l
l
tions up to 10
with IC50 = 6.8
l
l
M and only weak antagonism at the hM4 receptor
M. CV risk through direct interaction with cardiac
ion channels was viewed to be minimal from in vitro experiments
where 7b had no significant effect on either the hERG channel as
measured by functional blockade in patch-clamp experiments
(IC50 >100
through functional blockade in the isolated rabbit cardiac myocyte
(up to 100 M).
lM) or the cardiac sodium channel (Nav 1.5) as assessed
l

M. D. Andrews et al. / Bioorg. Med. Chem. Lett. 21 (2011) 2715–2720
2719
development for the treatment of SUI. In addition, it proved to be
critical to build an understanding of the translation between re-
combinant cell-based systems, native tissue preparations and
in vivo preclinical models. This was a significant undertaking and
proved to be crucial in compound selection. Second generation
pyrimido[4,5-d]azepines have been identified with improved ago-
nist activity at the 5-HT2C receptor combined with no measurable
activation of the 5-HT2B receptor.15 These studies will be the sub-
ject of future publications.
N
NH
N
7b
Major pathways
CYP2D6
CYP2B6
CYP2C8
CYP2C19
Acknowledgments
R2
N
We thank David Batchelor, David Gordon, Russell Horne, Debbie
Lovering, Ed Pegden, Michael Ralph, Adam Stennett and Simon
Wheeler for compound synthesis. We are grateful to Jummai
Guillabert, Raj Logan, Anthony Payne, Lucy Rogers, Sunny Z. Sun,
Christine Tyman and Kuresh Youdim for additional screening data.
We acknowledge the excellent support from John Deering and
Trevor Newbury for large scale hydrogenation services and Laure
Hitzel and Juliet Love of the Separation and Analytical Services
group.
N
R1
N
13: R1 = OH; R2 = H
14: R1 = H; R2 = OH
Figure 3. In vitro metabolism of 7b in HLM.
The evaluation of 7b in a preclinical in vivo canine model of
stress urinary incontinence (SUI) showed a robust dose-dependant
response in improving urethral tone that was superior to APD-356
(1) at similar free plasma concentrations of drug (Fig. 4).25,26
Azepine 7b increased peak urethral pressure (PUP) by 20–30% at
a free plasma concentration of 20–45 nM which compared favor-
ably with other 5-HT2C agonists10–12 and drugs that have been
evaluated in this model, for example, duloxetine (SNRI);27 PF-
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sub-chronic rat (4-day, up to 30 mg/kg) and dog (7-day, up to
10 mg/kg) exploratory toxicology studies and salt screening with
7b identified the hemi-fumarate salt as a crystalline, non-hygro-
scopic solid form (mp 210 °C) with excellent aqueous solubility
(>10 mg/mL at pH 1.5–11.7) and stability (pH 1–12).
In summary, we have identified new pyrimido[4,5-d]azepines 7
as potent 5-HT2C receptor agonists. A preferred example 7b had
minimal activation at either the 5-HT2A or 5-HT2B receptors, com-
bined with robust efficacy in a preclinical canine model of SUI and
attractive pharmacokinetic and safety properties. Based on this
profile, 7b (PF-3246799)15 was identified as a candidate for clinical
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Figure 4. The effect of APD-356 (1) and PF-3246799 (7b) on peak urethral pressure
(PUP) as percent change relative to control in the female anaesthetised dog (n P 4;
iv infusion). Each bar represents the mean response SEM versus mean free plasma
concentration. Dog 5-HT2C functional potency, APD-356 (1): EC50 = 16.2 nM (82%)
(n = 3); PF-3246799 (7b): EC50 = 10.8 nM (81%) (n = 3).

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