Design and syntheses of melanocortin subtype-4 receptor agonists. Part 2: Discovery of the dihydropyridazinone motif

Article abstract of DOI:10.1016/j.bmcl.2005.06.033

Optimization of the biological activity of a new class of non-peptidyl, pyridazinone derived human melanocortin subtype-4 receptor agonists is disclosed.

Full text of DOI:10.1016/j.bmcl.2005.06.033

Bioorganic & Medicinal Chemistry Letters 15 (2005) 4023–4028
Design and syntheses of melanocortin subtype-4 receptor
agonists. Part 2: Discovery of the dihydropyridazinone motif
Feroze Ujjainwalla,a,* Daniel Warner,a Christine Snedden,a Ricky D. Grisson,a
Thomas F. Walsh,a Matthew J. Wyvratt,a Rubana N. Kalyani,b Tanya MacNeil,b
Rui Tang,b David H. Weinberg,b Lex Van der Ploegc and Mark T. Gouletc
aDepartment of Medicinal Chemistry, Merck Research Laboratories, Rahway, NJ 07065-0900, USA
bDepartment of Obesity and Metabolic Research, Merck Research Laboratories, Rahway, NJ 07065-0900, USA
cMerck Research Laboratories-Boston, 33 Avenue Louis Pasteur, Boston, MA 02115, USA
Received 9 April 2005; accepted 7 June 2005
Available online 7 July 2005
Abstract—Optimization of the biological activity of a new class of non-peptidyl, pyridazinone derived human melanocortin subtype-
4 receptor agonists is disclosed.
Ó 2005 Elsevier Ltd. All rights reserved.
The association between the melanocortin subtype-4
receptor (MC4R) pathway and the modulation of feed-
ing has been established through the course of a number
of elegant genetic and pharmacological studies, involv-
ing both animals and humans.1,2 These studies, in part,
have provided the impetus in drug discovery to identify
novel, small molecule agonists of the hMC4R for the
potential treatment of human obesity.3 In a recent re-
port from this laboratory, we described the discovery
of a structurally unique pyridazinone derived class of
functionally selective (vs hMC3R and hMC5R) hMC4R
agonists, exemplified by compounds 1–3 (Fig. 1).4
In this report, we disclose our endeavors to optimize the
biological activity of these compounds, with particular
emphasis on improving functional potency and selectiv-
ity. This article presents the syntheses and structure–
activity relationships (SARs) of both pyridazin-3(2H)-
ones and 4,5-dihydropyridazin-3(2H)-ones as potent
and subtype selective hMC4R agonists.
Figure 1. Human MC4R agonist 1, 2, and 3.
potency and selectivity. Assembly of these compounds
was executed through a late stage amide bond coupling
of the pyridazinone and the aryl piperidine/pyrrolidine
halves. The requisite aryl piperidine/pyrrolidine frag-
ments were synthesized using the procedures described
in Scheme 1.
The general synthetic approaches to the pyridazinones
presented in this study are illustrated in Schemes 1–4.
We envisioned that modification of the aryl ring at-
tached to either the piperidine of 1 and 2 or the pyrrol-
idine of 3 would be a fertile area in which to investigate
Preparation of the 4-aryl piperidine fragment 9 began
with commercially available piperidone 4 (Scheme 1).
From the outset of the synthesis, we elected to replace
the N-benzyl protecting group with a t-butyl carbamate
derivative, since this would accommodate potentially
reducible functionality under catalytic hydrogenation
*
Corresponding author. Tel.: +1 732 594 1719; fax: +1 732 594
2210; e-mail: f_ujjainwalla@merck.com
0960-894X/$ - see front matter Ó 2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2005.06.033

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F. Ujjainwalla et al. / Bioorg. Med. Chem. Lett. 15 (2005) 4023–4028
8 as a single cis diastereoisomer. Ultimately, a trans
disposition of the carboxamide and aryl groups was re-
quired, and although a direct conversion of 7 to trans-8
was not achievable in our hands, we discovered that 7
could be converted to an enriched mixture of trans-8/
cis-8 (P3:1, respectively) in the presence of magnesium
metal in methanol.6 Exposure of trans-8/cis-8 to sodium
in methanol afforded exclusively trans-8, which upon
base catalyzed hydrolysis of the ester functionality fur-
nished the racemic trans carboxylic acid 9.
3-Aryl piperidine 11 was prepared in an analogous fash-
ion to 9, starting with commercially available piperidone
10 (Scheme 1).
4-Aryl pyrrolidine 14 was prepared according to a
concerted cycloaddition strategy (Scheme 1). Thus,
azomethine-ylide [3+2] cycloaddition of benzyl-(meth-
oxymethyl)[(trimethylsilyl)methyl]amine and
a trans
arylacrylate in the presence of a catalytic amount of tri-
fluoroacetic acid afforded trans pyrrolidine 12.7 Replace-
ment of the N-benzyl protecting group with a Boc
derivative for the aforementioned reasons, followed by
base catalyzed hydrolysis of the ester functionality pro-
vided the racemic trans carboxylic acid 14.
Scheme 1. Reagents and conditions: (i) (a) H2 (50 psi), Pd/C, EtOH,
H2O, rt; (b) (Boc)2O, NaHCO3, NaCl, CHCl3, H2O, 60 °C; (ii) Tf2O,
DIPEA,CH2Cl2, ꢀ78 °C; (iii) arylboronic acid, Pd(dppf)Cl2, EtOH,
toluene, 2 M NaHCO3, 80 °C; (iv) Mg, MeOH, rt; (v) (a) Na, MeOH,
65 °C; (b) 5 N NaOH, MeOH, 65 °C; (vi) cat. TFA, CH2Cl2, 0 °C; (vii)
(a) HCO2NH4, Pd(OH)2/C, MeOH, 65 °C; (b) (Boc)2O, NaHCO3,
NaCl, CHCl3, H2O, 60 °C.
Investigation of an aryl piperazine variant of 1 was also
of interest, and the requisite coupling fragment was
prepared according to the general method delineated be-
low (Scheme 2). Arylation of known piperazine 15
according to the Evans–Chan8 protocol followed by
saponification of the ester functionality afforded the
racemic carboxylic acid 17.
Scheme 2. Reagents and conditions: (i) arylboronic acid, Cu(OAc)2,
˚
Acylation of 184 with racemic acid 9 afforded an equi-
molar mixture of the target pyridazinone 19 and the
related trans diastereoisomer 20 (Scheme 3). These com-
pounds were conveniently separated by normal phase
flash chromatography on silica gel stationary phase.
Acid catalyzed removal of the Boc group afforded pyrid-
pyridine, 4 A sieves, CH2Cl2, rt; (ii) 5 N NaOH, MeOH, 65 °C.
Scheme 3. Reagents and conditions: (i) 9, EDC, HOBT, CH2Cl2,
N-methylmorpholine, rt; (ii) HCl, EtOAc, rt.
conditions, and facilitate late stage ease of removal.
Accordingly, palladium-catalyzed hydrogenolysis of 4
followed by BOC protection afforded piperidone 5. For-
mation of the enol triflate 6 and subsequent Suzuki-
Miyaura5 cross-coupling with an arylboronic acid
afforded 4-aryl tetrahydropiperidine 7. Reduction of
the tetrasubstituted alkene functionality under atmo-
spheric pressure of hydrogen furnished aryl piperidine
Scheme 4. Reagents and conditions: (i) Zn, AcOH, 120 °C, 56–71%;
(ii) preparative normal phase chiral HPLC, ChiralcelÒ OD stationary
phase, 50% EtOH/hexanes mobile phase, 36% (25, faster eluting
diastereoisomer), 34% (26, slower eluting diastereoisomer).

F. Ujjainwalla et al. / Bioorg. Med. Chem. Lett. 15 (2005) 4023–4028
4025
Table 2. Binding affinity and functional activity of compounds at the
human MC4R10,11
azinones 21 and 22, respectively. Pyridazinones 27–55
were prepared uneventfully according to this general
method.
The synthetic strategy to dihydropyridazinones 25 and
26 is presented in Scheme 4. Reduction of 23 in the
presence of zinc metal and acetic acid provided 24
as a statistically equivalent mixture of C-4 epimers,9
which was resolved by preparative normal phase chiral
high performance liquid chromatography (HPLC).
Dihydropyridazinones 60–69 were prepared in an
analogous fashion.
Compound
R
Binding IC50
(nM)
cAMP EC50
(nM) (%max)
2
38
39
40
41
42
43
44
45
Ph
130
58
240 (85%)
150 (101%)
2400 (30%)
110 (101%)
550 (75%)
710 (75%)
120 (81%)
36% at 10 lM
630 (62%)
Ph-4-Cl
Ph-2,4-diCl
Ph-4-F
36
66
Pyridazinones 1 (Table 1), 2 (Table 2), and 3 (Table 3)
were disclosed in an earlier report,4 and are displayed
here for comparative purposes. Investigation of the aryl
substituent attached to piperidine ring revealed a
marked preference for 4-substitution compared to 3-
substitution, as exemplified by compounds 23 and 27
versus 30 and 28, respectively (Table 1). In the case of
fluorine substitution, the 2-fluoro analog 31 was also
examined and showed binding affinity comparable to
that of 23, but intermediary functional activity between
23 and 30. Notably, while compounds such as 29, 32,
and 34 all demonstrated comparable binding affinity to
hMC4R relative to 23, they were significantly less effica-
cious hMC4R agonists. Since the binding and functional
assays are inherently different, it was not unexpected to
observe compounds which showed differences in binding
affinity to, and activation of the hMC4R. However, the
particularly large disconnect in IC50 versus EC50 for
compounds such as 32, 34, and especially 29 was inter-
esting considering the subtle structural differences
between the compounds. The binding affinity
(IC50 = 3.2 nM) and lack of functional activity (2% at
10 lM) observed with 29 suggested that this compound
might be a functional antagonist of hMC4R. Pyridazi-
Ph-2-F
100
200
45
Ph-4-CF3
Ph-4-CN
CH2Ph
750
390
CH2Ph-4-F
Table 3. Binding affinity and functional activity of compounds at the
human MC4R10,11
Compound
R
Binding IC50
(nM)
cAMP EC50
(nM) (%max)
3
46
47
48
49
50
51
Ph
Ph-2-F
Ph-4-F
98
160
70
400 (103%)
710 (82%)
110 (84%)
200 (81%)
230 (70%)
750 (67%)
430 (67%)
Ph-2,4-diF
Ph-4-Cl
Ph-4-Me
Ph-4-OMe
60
45
28
19
Table 1. Binding affinity and functional activity of compounds at the
human MC4R9,10
none 29 was indeed found to be a functional antagonist
demonstrating KB = 3.1 0.7 nM.12 In the 3-aryl piper-
idine and 4-aryl pyrrolidine series, a much closer corre-
lation between hMC4R binding affinity and functional
activity was observed (Tables 2 and 3). The 4-fluoro
and 4-chloro modifications were found to display the
optimum blend of hMC4R binding affinity, functional
activity, and receptor agonism, and overall, the 4-fluor-
ophenyl substitution pattern was preferred in each of the
three series. A more detailed hMCR activity profile of
pyridazinones 23, 40, and 47 is shown in Table 4. Pyrid-
azinone 23 showed the best combination of hMC4R
binding affinity, functional activity, and functional selec-
tivity (vs hMC3R and hMC5R) relative to 40 and 47,
although these differences while statistically significant
were subtle.
Compound
R
Binding IC50
(nM)
cAMP EC50
(nM) (%max)
1
27
28
29
23
30
31
32
33
34
35
36
37
Ph
Ph-4-Cl
Ph-3-Cl
33
2.4
43
180 (77%)
210 (39%)
680 (37%)
2% at 10 lM
61 (71%)
Ph-2,4-diCl
Ph-4-F
Ph-3-F
3.2
11
490
13
29% at 10 lM
550 (58%)
Ph-2-F
Ph-4-CF3
Ph-3-CF3
Ph-4-Me
2-Naphthyl
1-Naphthyl
3-Thienyl
3.8
100
4.7
17
>5000 (24%)
40% at 10 lM
1100 (47%)
12% at 10 lM
22% at 10 lM
490 (82%)
Replacement of the piperidine ring in 23 with a pipera-
zine variant resulted in a significant attenuation of
hMC4R functional activity, irrespective of the chirality
of the carboxamide group (52 and 53, Table 5). While
89
130

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F. Ujjainwalla et al. / Bioorg. Med. Chem. Lett. 15 (2005) 4023–4028
Table 4. Human MCR activity profile of 23, 40, and 4710,11
Table 6. Binding affinity and functional activity of 56–59 at the human
MC4R10,11
Compound Receptor Binding IC50
(nM)
cAMP EC50
(nM) (%max)
23
40
47
MC3
MC4
MC5
MC3
MC4
MC5
MC3
MC4
MC5
940 92
11 1.2
6
61 19 (71 3%)
2% at 10 lM
840 92
40% at 4 lM
66 20
7
1% at 10 lM
45 5% at 10 lM
110 17(101 5%)
1200 210
>3000
70 13
1 1% at 10 lM
2400 790 (24 2%)
110 35 (84 3%)
5
1400 150
1% at 10 lM
Table 5. Binding affinity and functional activity of 52–55 at the human
MC4R10,11
Compound
Binding IC50 (nM)
cAMP EC50 (nM) (%max)
56
57
58a
59
28
290
93
470 (60%)
9% at 10 lM
410 (94%)
320 (59%)
4.7
a 1:1 mixture of trans diastereoisomers.
Compound
C-30
stereo
C-300
stereo
Binding
IC50 (nM)
cAMP EC50
(nM) (%max)
52
53
54
55
R
R
S
S
R or S
S or R
R or S
S or R
950
28
6% at 10 lM
1100 (75%)
with a prerequisite that overall molecular weight be con-
served. With this condition in mind, installation of a ste-
reogenic center at the C-4 position of the pyridazinone
core emerged as a seductive option, since the vectoral
disposition of the SAR-sensitive C-4 grouping would
be different from a center of tetrahedral (sp3) geometry
versus trigonal (sp2) geometry. This led to the C-4,5-
dihydropyridazinone modification, which for complete-
ness, was investigated in two stereochemical series. The
first, involved 23, 40, and 47, which possessed the pre-
ferred stereochemical disposition between the carbox-
amide and aryl groups on the piperidine/pyrrolidine
nucleus, and the second, involved the related trans dia-
stereoisomers, which featured an antipodal arrangement
of these groups (Table 7). Since the reductions of 23, 40,
and 47 were stereo-random, a set of two diastereoiso-
mers was generated for each compound, and these were
epimeric at the C-4 position. Dihydropyridazinones 26,
63, and 67, which originated from 23, 40, and 47, respec-
tively, showed improved hMC4R binding affinity com-
pared to their respective pyridazinone counterparts. In
the case of 26, ꢁ7-fold improvement in hMC4R binding
affinity was observed relative to 23, whereas for 63 ver-
sus 40 and 67 versus 47, the changes in hMC4R binding
affinity were less pronounced. HMC4R functional activ-
ity and receptor agonism were essentially unchanged by
partial saturation of the pyridazinone core. The full
hMCR activity profile of 26, 63, and 67 is illustrated
in Table 8, which demonstrates the inherent potential
of the dihydropyridazinone modification to modulate
hMC4R binding affinity, functional activity, receptor
agonism, and subtype selectivity (Table 8).
1100
1000
9% at 10 lM
38% at 10 lM
53 demonstrated comparable hMC4R binding affinity
relative to 23, ꢁ20-fold attenuation in functional activi-
ty was observed, again highlighting the difference be-
tween binding to, and activation of the hMC4R.
Reversal of the stereochemical orientation of the methyl
group in the tether amplified this drop in potency (54
and 55). Elimination of the carbonyl functionality
(amide) resulted in attenuated hMC4R functional activ-
ity (56), which was further dramatized in the related
trans diastereoisomer 57 (Table 6). Pyridazinones 56
and 57 were prepared according to the Fukuyama meth-
od.13 Thus, the trans racemic primary alcohol derived
from borane reduction of carboxylic acid 9 (Ar = 4-
fluorophenyl) was coupled with the 2,4-dinitrobenzene-
sulfonamide derivative of amine 18. Subsequent diaste-
reoisomer separation and desulfonylation afforded
diamines 56 and 57. Capping the carboxamide function-
ality or the piperidine moiety with a methyl group did
not lead to compounds with substantially improved
hMC4R binding affinity or functional activity (58 and
59, respectively). N-Methyl amide 58 was prepared by
acylation of the amino methyl derivative of 18 with car-
boxylic acid 9 (Ar = 4-fluorophenyl), while 59 was pre-
pared using a reductive amination reaction between 23
and paraformaldehyde.
To optimize the functional selectivity of the pyridazi-
none series of compounds, we conceptualized the idea
of introducing additional chirality to this lead design,
In conclusion, we have described the design and asym-
metric synthesis of a new class of non-peptidyl, MC4R

F. Ujjainwalla et al. / Bioorg. Med. Chem. Lett. 15 (2005) 4023–4028
4027
Table 7. Binding affinity and functional activity of compounds at the
human MC4R10,11
Table 8. Human MCR activity profile of 26, 63, and 6710,11
Compound Receptor Binding IC50 cAMP EC50
(nM)
(nM) (%max)
12 7% at 10 lM
2% at 10 lM
40 7.0 (67 2%)
2400 390 (27 2%)
220 41 (33 8%)
1100 280 (21 3%)
30 5.2 (98 3%)
26
63
67
MC1B14
MC3
MC4
230 0.0
420 47
1.5 0.2
5
MC5
1100 140
1100 64
4700 92
22 2.8
MC1B14
MC3
MC4
Compound C-4 stereo Ra,b
Binding cAMP EC50
IC50 (nM) (nM) (%max)
MC5
3100 290
290 93
7
2% at 10 lM
MC1B14
MC3
MC4
240 31 (18 2%)
10 2% at 10 lM
66 19 (90 3%)
1600 1000
18 3.6
690 132
25
26
60
R or S
S or R
R or S
26
390 (72%)
40 (67%)
MC5
5
1% at 10 lM
References and notes
1.5
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560
20% at 10 lM
61
62
63
64
65
66
67
68
69
S or R
R or S
S or R
R or S
S or R
R or S
S or R
R or S
S or R
690
540
22
4000 (32%)
480 (100%)
30 (98%)
1300
280
47
2000 (16%)
1400 (33%)
1200 (90%)
66 (98%)
18
780
430
4000 (26%)
740 (48%)
a Absolute stereochemistry.
b Ar signifies 4-fluorophenyl.
4. Ujjainwalla, F.; Warner, D.; Walsh, T. F.; Wyvratt, M. J.;
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percentage of cAMP accumulation at 10 lM of compound
relative to a-MSH. For a detailed description of the assay
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11. All compounds are single enantiomers, >95% pure
(HPLC) and were tested as either TFA or HCl salts.
12. Functional antagonism was determined by measuring the
ability of different concentrations (from 5 to 1500 nM) of
compound 29 to shift the [125I]NDP-aMSH agonist dose–
response curve. KB of 3.1 0.64 nM was calculated from
Schild analysis.
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10. (a) All data represent mean values from at least two
separate experiments. SD (n = 2) or SEM (n = 3) were
generally 30% of the mean, unless specifically indicated
otherwise. hMC4R binding IC50 is defined as the concen-
tration of compound which can inhibit binding of
[
125I]NDP-a-MSH by 50% from membranes prepared
from CHO cells expressing human MC4R. hMC4R cAMP
EC50 is defined as the inflection point of the cAMP dose–
response curve for any given compound. %Max is the