Biological activity and preclinical efficacy of azetidinyl pyridazines as potent systemically-distributed stearoyl-CoA desaturase inhibitors

Article abstract of DOI:10.1016/j.bmcl.2010.10.107

Potent and orally bioavailable SCD inhibitors built on an azetidinyl pyridazine scaffold were identified. In a one-month gDIO mouse model of obesity, we demonstrated that there was no therapeutic index even at low doses; efficacy in preventing weight gain

Full text of DOI:10.1016/j.bmcl.2010.10.107

Bioorganic & Medicinal Chemistry Letters 21 (2011) 479–483
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Biological activity and preclinical efficacy of azetidinyl pyridazines
as potent systemically-distributed stearoyl-CoA desaturase inhibitors
⇑
Elise Isabel , David A. Powell, W. Cameron Black, Chi-Chung Chan, Sheldon Crane, Robert Gordon,
Jocelyne Guay, Sebastien Guiral, Zheng Huang, Joël Robichaud, Kathryn Skorey, Paul Tawa, Lijing Xu,
Lei Zhang, Renata Oballa
Merck Frosst Centre for Therapeutic Research, PO Box 1005, Pointe-Claire-Dorval, Québec, Canada H9R 4P8
a r t i c l e i n f o
a b s t r a c t
Article history:
Potent and orally bioavailable SCD inhibitors built on an azetidinyl pyridazine scaffold were identified. In
a one-month gDIO mouse model of obesity, we demonstrated that there was no therapeutic index even at
low doses; efficacy in preventing weight gain tracked closely with skin and eye adverse events. This was
attributed to the local SCD inhibition in these tissues as a consequence of the broad tissue distribution
observed in mice for this class of compounds. The search for new structural scaffolds which may display
a different tissue distribution was initiated. In preparation for an HTS campaign, a radiolabeled azetidinyl
pyridazine displaying low non-specific binding in the scintillation proximity assay was prepared.
Ó 2010 Elsevier Ltd. All rights reserved.
Received 14 September 2010
Revised 20 October 2010
Accepted 21 October 2010
Available online 26 October 2010
Keywords:
Stearoyl-CoA desaturase inhibitors
SCD
Pyridazine
Azetidine
Systemically-distributed
The incidence of obesity and diabetes occurrence worldwide
continues to increase at an alarming pace. The World Health Orga-
nization (WHO) has estimated that 1.6 billion adults are currently
overweight with at least 400 million of these being classified as ob-
ese.1 There is a strong risk factor for developing type 2 diabetes in
obese individuals.2 It has been hypothesized that de-regulation of
stearoyl-CoA desaturase-1 enzyme (SCD, a delta-9 desaturase)
may play an important role in the development of these metabolic
disorders.3 SCD catalyzes the formation of a cis-double bond at the
C-9 position of C-16 palmitoyl-CoA and C-18 stearoyl-CoA into
palmitoleoyl-CoA and oleoyl-CoA, respectively.4 Four SCD isoforms
have been characterized in mouse. Although they display a similar
desaturation activity towards stearoyl-CoA and palmitoyl-CoA,
they have a different tissue distribution.5,6 So far, two isoforms,
SCD1 and SCD5, have been characterized in humans. Human
SCD1 is the ortholog of the mouse SCD1, whereas SCD5 is not an
ortholog of any of the four mouse enzyme and is highly expressed
in brain and pancreas.7 Besides delta-9 desaturases, there are two
other desaturases in humans: delta-5 and delta-6. Since these two
desaturases are involved in the synthesis of highly unsaturated
fatty acids which are esterified into phospholipids and contribute
to maintain membrane fluidity, selectivity against delta-5 and del-
ta-6 is essential to avoid undesirable toxicities.8 Recently, we and
others disclosed several new classes of potent SCD inhibitors which
are capable of inhibiting SCD activity in vivo (Fig. 1).9–12 We dem-
onstrated that efficacy in preventing weight gain in either a grow-
ing or established diet induced obesity model (gDIO or eDIO)
tracked closely with partial eye closure and alopecia. We now re-
port a complementary series, the azetidinyl pyridazines. The pur-
pose of this work was to identify a series that would allow for a
therapeutic window by a titration of the dose of a systemically-dis-
tributed SCD inhibitor to the minimal amount required for efficacy
in a gDIO model. Additionally, as a result of the unique physical
properties of these inhibitors, specifically their low affinity for
polyvinyltoluene (PVT) and yttrium silicate, a member of this ser-
ies served as a key component in a recently-developed SPA binding
assay.
Azetidinyl pyridazines were synthesized following the general
sequence described in Scheme 1. The commercially available
3-methyl-6-chloropyridazine was oxidized to the corresponding
carboxylic acid 2. The carboxylic acid was then converted into an
acid chloride which was immediately reacted with acetic hydra-
zine in the presence of an organic base to obtain 3 in satisfactory
yield over 2 steps. Compound 3 was reacted with Burgess’s reagent
(methyl N-(triethylammoniumsulfonyl)carbamate) at high tem-
perature in the microwave to perform a dehydration/cyclization
and provide the oxadiazole 4. The commercially available Boc-pro-
tected 3-azetidinol 5 was reacted with phenols 6 under modified
Mitsunobu conditions to yield ethers 7.13 The Boc protecting group
was removed in the presence of anhydrous HCl, and in most cases,
the azetidine hydrochloride salt was isolated directly by filtration.
⇑
Corresponding author. Tel.: +1 514 428 3655; fax: +1 514 428 2624.
E-mail addresses: elise_isabel@merck.com, isabel.e@videotron.ca (E. Isabel).
0960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.10.107

480
E. Isabel et al. / Bioorg. Med. Chem. Lett. 21 (2011) 479–483
O
O
S
N
S
H2N
HO
N
N
N
CF3
N
O
CF3
N
N
H
N
N
N
N
N
S
N
CF3
O
N
MF-152 (ref. 9a)
MF-438 (ref. 9b)
Xenon/ Novartis (ref.10)
O
N
N
N
O
F
Cl
S
N
N
O
N N
O
N
O
CF3
N
Daiichi (ref. 11)
Figure 1. Recently published SCD inhibitors.
N
N
N
N
N
Cl
A (ref. 9c)
Abbott (ref. 12)
appropriate azetidines 8 was successfully achieved to provide the
pyridazines 11a–c. Acylhydrazides 12a–c were obtained by react-
ing the methyl esters with a large excess of hydrazine in EtOH as
a co-solvent. The terminal nitrogen of the hydrazines 12a–c was
then acylated with acetoxyacetyl chloride to give the doubly acyl-
ated hydrazides 13a–c. Compounds were then subjected to a dehy-
dration/cyclization sequence with the Burgess’s reagent to yield
the oxadiazoles 14a–c. Finally, the deprotection of the acetate
group on the primary alcohol with hydrazine provided compounds
15a–c.
O
O
O
N
a
c
b
Cl
Cl
Cl
Cl
N
N
N
1
HO
N
N
2
HN
NH
N N
3
N
N
4
O
O
Boc
N
HN
O
R
R
R
OH
Boc
N
OH
+
e
d
H
Cl
f
5
6a-i
7a-i
8a-i
O
N
O
R
Azetidine inhibitors 9a–i and 15a–c were tested in a rat liver
microsomal enzyme assay.14 The cellular potencies against SCD
and selectivities against the delta-5 and delta-6 desaturases (data
N
N
N
N
9a-i
not shown, all analogs tested were inactive, >2 lM) were measured
Scheme 1. Reagents and conditions: (a) potassium dichromate (1.2 equiv), H2SO4,
50 °C, 4 h, 40% (b) (i) oxalyl chloride (1.1 equiv), DMF (cat.), toluene (ii) acetic
hydrazine (1.2 equiv), Hunig’s base (2.0 equiv), dichloromethane, 0 °C, 1.5 h, 54% (c)
Burgess’s reagent (1.5 equiv), THF, 150 °C (microwave), 30 min, 97% (d) phenol (1.2
equiv), 1,10-(azodicarbonyl)dipiperidine (1.2 equiv), tri-n-butylphosphine (1.2
equiv), THF, 16 h, 75 °C, 47% (e) 4 M HCl in dioxane (5.0 equiv), dichloromethane,
83% (f) 4 (1.0 equiv), K2CO3 (3.0 equiv), dioxane, 110 °C, 2 days, 40%. Yields given for
R = ortho-CF3.
in a HepG2 whole cell assay.15 The results of these experiments are
shown in Table 1. When compared to MF-438, 9a was approxi-
mately 12-fold less potent in the enzyme assay but had a compara-
ble potency in the cellular assay. The SAR in this series followed a
similar trend to those previously reported, with ortho-substitution
on the aryl ether ring providing the most potent compounds.9 Com-
pound 9g with a meta-substituted trifluoromethyl group was far
less active in the enzyme assay and addition of an ortho-substituent
to the meta-substituted CF3 (9h) was not sufficient to restore po-
tency. Evaluation of the pharmacokinetics properties of 9d and 9e
in mice revealed that these azetidinyl analogs have a similar long
half-life as the piperidine derivatives (7 h versus 6.4 h for MF-
438).16 LC-MS analysis of the whole blood samples identified a
M+16 metabolite circulating with the parent compound. Based on
previous observations with MF-438 and A, it was suspected that this
metabolite was being generated by the enzymatic oxidation of the
methyl moiety of the oxadiazole to the corresponding alcohol.9 Con-
sidering that this alcohol functionality did not negatively impact the
potency of previous analogs, these primary alcohols (15a–c) were
prepared so that they could be evaluated. Compound 15a exhibited
a comparable potency to MF-438 against the rat enzyme with a sim-
ilar bioavailability and half-life. When comparing 15a to 9a, it seems
clear that adding the primary alcohol moiety provided a marked
improvement in intrinsic potency. Compounds 15b and 15c had
similar potencies in the enzyme assay, but there was almost a
10-fold difference in the whole cell assay, 15b being the most potent
(5.6 nM).
The synthesis was completed by a thermal displacement step in
refluxing dioxane to afford the desired compounds (9a–i) in
acceptable yields.
SCD inhibitors 15a–c were synthesized according to the se-
quence of reactions shown in Scheme 2. The carboxylic acid 2
was converted into an acid chloride with oxalyl chloride in the
presence of a catalytic amount of DMF. After 1 h, the volatile com-
ponents were removed under reduced pressure. The acid chloride
was dissolved in toluene, cooled to 0 °C and MeOH was added to
provide the methyl ester 10. Thermal displacement with the
O
O
O
O
O
a
b
N
O
Cl
Cl
R
N
N
N
N
HO
N
N
2
N
10
N
11a-c
c
O
O
N
O
N
O
d
R
R
HN
N
N
HN
Ac
O
NH
NH2
13a-c
12a-c
Having identified the azetidinyl pyridazines as potent and
bioavailable SCD inhibitors, we next sought to choose a suitable
inhibitor for evaluation in an in vivo anti-obesity mouse model
study and assess if a therapeutic window could be established.
Lead compounds 15a and 15c were further profiled in a mouse li-
ver pharmacodynamic model (mLPD) in order to measure in vivo
inhibition of liver SCD activity.14 We had previously found that
compounds which exhibit a high inhibition in our mLPD demon-
strate efficacy in anti-obesity models. Inhibitors 15a and 15c were
administered orally in 1% methocel. One hour later, 14C-stearic acid
(SA) was intravenously administered in 60% PEG-200. Two hours
following the administration of the tracer, livers were harvested.
O
e
Ac
O
N
O
O
N
HO
f
N
O
R
N
O
R
N
N
N
N
N
N
15a-c
14a-c
Scheme 2. Reagents and conditions: (a) (i) oxalyl chloride (1.1 equiv), DMF (cat.),
toluene, 1 h (ii) MeOH (28 equiv), toluene, 2 h, 0 °C, 70%; (b) 8 (1.0 equiv), K2CO3
(3.0 equiv), dioxane, 110 °C, 2 days, 50%; (c) hydrazine (20 equiv), EtOH, 16 h; (d)
acetoxyacetyl chloride (1.2 equiv), dichloromethane/water 2:3, 2 h, used unpuri-
fied; (e) Burgess’s reagent (1.2 equiv), THF, 150 °C (microwave), 30 min, 51%; (f)
hydrazine (10 equiv), MeOH, 16 h, 75%. Yields given for R = ortho-CF3.

E. Isabel et al. / Bioorg. Med. Chem. Lett. 21 (2011) 479–483
481
Table 1
Potency and mouse pharmacokinetics data of azetidinyl piperazine SCD inhibitors
O
N
X
N
O
R
N
N
N
X= H, OH
Compd
X
R
Rat SCD enz. IC50 (nM)a
HepG2 cell IC50 (nM)a
Mouse PK (%F; t1/2 in h)b
MF-438
9a
9b
H
H
H
2-CF3
2-CF3
2-Br
2.3
27
21
21
9.0
23
73; 6.4
75; 2.8
n.d.
9c
H
2-I
7.2
22
n.d.
9d
9e
9f
9g
9h
9i
15a
15b
15c
H
H
H
H
H
H
OH
OH
OH
2-Br, 5-F
2-Br, 4-F
2-Cl, 5-CF3
3-CF3
2-Cl, 3-CF3
2-Cl, 4-F, 6-Cl
2-CF3
28
67
19
40
97; 6.9
91; 7.7
n.d
n.d
n.d.
163
1992
3385
8167
2.3
n.d.
n.d.
n.d.
n.d.
13
n.d.
57; 5.0
61; 4.4
100; 2.2
2-Br, 5-F
2-Br
3.4
4.9
5.6
50
a
IC50’s are an average of at least two independent titrations.
10 mg/kg PO (1% methocel, n = 2) and 2 mg/kg IV (60% PEG-200, n = 2).
b
After a basic hydrolysis of liver lipids, samples were analyzed with
a radiometric detector. Since SCD effects the conversion of stearic
to oleic acid (OA), low levels of OA characterize a potent SCD inhib-
itor. As reported in Table 2, 15c was demonstrated to be a more po-
tent inhibitor of liver SCD activity in vivo as compared to 15a.
Compound 15c achieved a >90% suppression of oleic acid produc-
tion at a dose of 1 mg/kg, 3-fold lower than that required to
observed comparable effect on oleic acid production with 15a. A
tissue distribution study (Fig. 2) with 15c was conducted and con-
firmed that 15c retains a systemically-distributed tissue profile
with a 4.5-fold higher drug concentration being observed in the
liver compared to plasma, similar to MF-438 and A. Compound
15c, as a result of its potent in vivo activity, was evaluated in a
one-month growing diet induced obesity model (gDIO) in mice.
Our goal was to down titrate 15c to see if at low exposure it was
possible to avoid adverse effects previously observed with A
(0.2 mg/kg) and MF-438 (5 mg/kg). The doses chosen for this study
with 15c were 0.1 and 0.3 mg/kg qd. The highest dose was selected
in order to provide full inhibition of liver activity (as demonstrated
in the mLPD assay) while the low dose was chosen at 1/3 of the
high dose where only partial inhibition of liver SCD activity is ob-
served. Results are shown in Figure 3. When 15c was administered
at 0.1 mg/kg/day, there was a moderate, but statistically significant
improvement in body weight over the control group (P <0.01) after
28 days. No eye adverse effects (AE’s) were observed with 0.1 mg/
kg/day of 15c for the duration of the study.17 However after
3 weeks of treatment, skin AE’s were observed. With a dose of
0.3 mg/kg/day of 15c, AE’s were comparable to what was observed
Table 2
mLPD assay: in vivo SCD inhibition of 15a and 15c
Compd
Dose (mg/kg)
% Inhibitiona
Plasma and liver levels (lM)
15a
15a
15a
15c
15c
15c
3.0
1.0
0.3
1.0
0.3
0.1
97% (P = 0.044)
NS
NS
92% (P <0.001)
74% (P <0.001)
NS
1.9; 15.8
0.01; 7.9
0; 1.2
1.4; 11.9
0.1; 7.2
0; 2.7
NS = Non-significant.
a
SCD activity index (ratio of 14C-oleic acid/14C-stearic acid) in hydrolyzed liver
lipids; n = 5 per group.
Tissue distribution of 15c in Mice
P.O. 10 mg/kg (10 mL/kg) in 1% Methocel, 2 hours after dosage
45
40
35
30
25
20
15
10
5
O
N
HO
N
O
Br
N
N
N
0
Plasma
Left Median
Liver Lobe
Left
Epididymal
Fat
Heart
Left Brain
Cerebellum Skin
(Shaved)
Figure 2. Tissue distribution 2 h post-dose at 10 mg/kg PO in C57BL6 mice fed on normal chow diet.

482
E. Isabel et al. / Bioorg. Med. Chem. Lett. 21 (2011) 479–483
Figure 3. Body weight gain, eye AE’s and skin AE’s of C57BL6 mice fed on high fat diet and dosed qd with 15c compared to two control arms: fed on high fat and normal chow
over 4 weeks; n = 8 per group; HFD-veh = high fat diet, vehicle treated control group; ND-veh = normal diet, vehicle treated control group.
previously with MF-438 and A. Based on this data, it was con-
cluded that obtaining a beneficial effect for reduced weight gain
without experiencing AE’s with a systemically-distributed com-
pound was likely not possible (Fig. 3).
non-specific binding with various types of scintillation proximity
beads. The azetidinyl piperazines display increased water solubil-
ity and possess a measured log D7.4 of less than 1.0, in contrast
to all other SCD inhibitors available in-house. A preliminary test
in the presence of TBS buffer18 in a plastic eppendorf tube demon-
strated that the azetidinyl piperazines 15a and 15c were recovered
in high yields. For inhibitors tested from other series available in-
house (results not shown) there was less than 30% recovery
(Table 3).19 Once it was discovered that 15a and 15c were adhering
less to plastic, 15c was used to test two types of scintillation beads:
PVT and yttrium. From this experiment, PVT-WGA and yttrium sil-
icate-antimouse beads were chosen for further investigation. Even
though 15c had a slightly greater recovery in the first round of
experiment (15a: 69% versus 15c: 79% recovery), it became clear
that it would be a greater synthetic challenge to obtain a radiola-
beled version of 15c while maintaining the ortho-bromine. We
decided to revisit 15a under the best conditions identified with
Since the azetidinyl pyridazine series and other systemically-
distributed series were not providing an adequate therapeutic win-
dow, there was a need to identify new scaffolds. We felt that new
scaffolds which offer a different tissue distribution profile may pro-
vide a suitable therapeutic window to enable clinical development.
A high throughput screening campaign using a scintillation prox-
imity binding assay (SPA) was put in place to evaluate our in-house
small molecule collection for activity against SCD. A suitable com-
pound to support the development of a SPA binding assay had to be
identified. Such a compound should have good potency against
SCD (i.e., <50 nM) and give low non-specific binding with the
scintillation proximity beads. Unfortunately, previously identified
series demonstrated significant binding to plate and gave high
Table 3
Identification of a suitable compound for the development of a SPA binding assay
% Recovery in TBS
% Recovery for beads and media with 15ca
% Recovery with optimized conditions
15c
9c
15a
Cond. 1
70
Cond. 2
66
Cond. 3
50
Cond. 4
96
Cond. 5
98
Cond. 6
89
Cond. 7
112
15a PVT
15a Yttrium
Concn (nM)
79
42
69
29
84
a
Cond. 1: 3 mg of PVT-PEI-WGA type A beads into 200
supernatant (ꢀ150 L) and analyze by LC-MS using a standard curve. Cond. 2: 3 mg of PVT-PEI-WGA type B beads and proceed as in cond. 1. Cond. 3: 3 mg of PVT-WGA beads
and proceed as in cond. 1. Cond. 4: 9.5 mg of yttrium silicate-WGA beads and proceed as in cond. 1. Cond. 5: 9.5 mg of yttrium silicate-poly-l-lysine beads and proceed as in
cond. 1. Cond. 6: 100 L of yttrium silicate-antimouse beads + 5 L of TBS buffer. Add 1 L of 15c (10 M DMSO). Shake 5 min, centrifuge at 12,000 rpm for 1 min, remove
supernatant (ꢀ90 L). Cond. 7: 100 L of yttrium silicate-antimouse beads + 5
lL of TBS buffer. Add 2 lL of 15c (10 lM DMSO). Shake for 5 min, centrifuge at 12,000 rpm for 1 min, remove
l
l
l
l
l
l
l
lL of 0.15 mg/mL mouse-anti-Flag antibody and proceed as in cond. 6.
Br
Br
Br
Br
N
Br
a
b
Br
+
OH
Boc
N
HO
HN
O
CF3
CF3
Boc
N
O
17
CF3
5
HCl
16
O
18
O
N
O
CF3
N
O
O
CF3
O
MeO
N
HN
NH2
N
N
N
c
d
Cl
Br
Br
MeO
N
10
N
19
20
Br
Br
e, f, g
O
N
O
N
HO
HO
N
O
CF3
N
CF3
h
N
N
N
N
N
T
Br
15% T incorporation
21
specific activity= 4.6 Ci/mmol
hot 15a
T
Br
Scheme 3. Reagents and conditions: (a) 5 (2.3 equiv) phenol (1.0 equiv), 1,10-(azodicarbonyl)dipiperidine (3.0 equiv), tri-n-butylphosphine (3.0 equiv), THF, 16 h, 75 °C, 85%;
(b) 4 M HCl in dioxane (5.0 equiv), dichloromethane, 4 h, 71%; (c) 18 (1.0 equiv), 10 (1.5 equiv), K2CO3 (3.0 equiv), dioxane, 110 °C, 24 h, 74%; (d) hydrazine (20 equiv), EtOH,
24 h, 92%; (e) acetoxyacetyl chloride (1.2 equiv), dichloromethane/water 2:3, 0.5 h, 0 °C, used crude; (f) Burgess’s reagent (1.3 equiv), dioxane, 120 °C, 2 h, 26% over 2 steps;
(g) hydrazine (10 equiv), MeOH, 16 h, 75%; (h) Pd(OH)2 on carbon (2.6 equiv), Et3N (13 equiv), T2 (60 mm Hg), EtOH/THF 5:1, 16 h, 35%.

E. Isabel et al. / Bioorg. Med. Chem. Lett. 21 (2011) 479–483
3. Liu, G. Expert Opin. Ther. Patents 2009, 19, 1169.
483
15c. Although low recovery was obtained with 15a in the presence
of PVT beads, an acceptable 84% recovery was observed with yt-
trium beads. Following this, a radiolabeled synthesis of 15a was
designed (Scheme 3). A Mitsunobu-type reaction between 5 and
1620 was performed in refluxing THF for 2 days to give the ether
17 in an 85% yield. The Boc group was cleaved in a few hours at
room temperature in presence of HCl to provide the hydrochloride
salt 18 as a white solid. The azetidine 18 and the chloropyridazine
10 were reacted with potassium carbonate in refluxing dioxane for
24 h. The intermediate 19 was obtained in a 74% yield and was fur-
ther reacted with excess hydrazine to provide the hydrazide 20 in a
high yield of 92%. The terminal nitrogen of 20 was reacted with
acetoxyacetyl chloride to give the doubly acylated intermediate
which was used crude for the cyclization with Burgess’s reagent.
This provided the oxadiazole 21 in a 26% yield over two steps.
The primary alcohol was deprotected in presence of a large excess
of hydrazine. The last step of the synthesis was the incorporation of
two tritium atoms on 21. This was achieved with an excess of
Pd(OH)2 on carbon and T2 in a 5:1 EtOH/THF solvent mixture. Com-
pound 15a was obtained in 35% yield after HPLC purification with
15% tritium incorporation. This compound was an effective ligand
in an in-house SPA-based HTS campaign.21
4. Ntambi, J. M. Prog. Lipid Res. 1995, 34, 139.
5. Ntambi, J. M.; Miyrazaki, M. Curr. Opin. Lipidol. 2003, 14, 255.
6. Miyazaki, M.; Jacobson, M. J.; Man, W. C.; Cohen, P.; Asilmaz, E.; Friedman, J.;
Ntambi, J. M. J. Biol. Chem. 2003, 278, 33904. and references cited therein.
7. Wang, J.; Yu, L.; Schmidt, R. E.; Su, C.; Huang, X.; Gould, K.; Cao, G. Biochem.
Biophys. Res. Commun. 2005, 332, 735.
8. Nakamura, M. T.; Nara, T. Y. Annu. Rev. Nutr. 2004, 24, 345.
9. (a) Li, C. S.; Belair, L.; Guay, J.; Murgasova, R.; Sturkenboom, W.; Ramtohul, Y.
K.; Zhang, L.; Huang, Z. Bioorg. Med. Chem. Lett. 2009, 19, 5214; (b) Léger, S.;
Black, W. C.; Deschênes, D.; Dolman, S.; Falgueyret, J.-P.; Gagnon, M.; Guiral, S.;
Huang, Z.; Guay, J.; Leblanc, Y.; Li, C. S.; Masse, F.; Oballa, R. M.; Zhang, L. Bioorg.
Med. Chem. Lett. 2010, 20, 499; (c) Ramtohul, Y. K.; Black, W. C.; Chan, C.-C.;
Crane, S.; Guay, J.; Guiral, S.; Huang, Z.; Oballa, R. M.; Xu, L.; Zhang, L.; Li, C. S.
Bioorg. Med. Chem. Lett. 2010, 20, 1593.
10. Dales, N.; Zhang, Z.; Fonarev, J.; Fu, J.; Kamboj, R.; Kodumuru, V.; Pokrovskaia,
N.; Sun, S. WO 2008/074835.
11. Uto, Y.; Ueno, Y.; Kiyotsuka, Y.; Miyazawa, Y.; Kuruta, H.; Ogata, T.; Yamada,
M.; Deguchi, T.; Konishi, M.; Takagi, T.; Wakimoto, S.; Ohsumi, J. Eur. J. Med.
Chem. 2010, 45, 4788.
12. Liu, G.; Lynch, J. K.; Freeman, J.; Liu, B.; Xin, Z.; Zhao, H.; Serby, M. D.; Kym, P.
R.; Suhar, T. S.; Smith, H. T.; Cao, N.; Yang, R.; Janis, R. S.; Krauser, J. A.; Cepa, S.
P.; Beno, D. W. A.; Sham, H. L.; Collins, C. A.; Surowy, T. K.; Camp, H. S. J. Med.
Chem. 2007, 50, 3086.
13. Tsunoda, T.; Yamamiya, Y.; Itô, S. Tetrahedron Lett. 1993, 34, 1639.
14. Li, C. S.; Ramtohul, Y., Huang, Z.; Zhang, L.; Lachance, N. WO 2006/130986.
15. Zhang, L.; Ramtohul, Y.; Gagné, S.; Styhler, A.; Wang, H.; Guay, J.; Huang, Z. J.
Biomol. Screening 2010, 15, 169.
16. Bateman, K. P.; Castonguay, G.; Xu, L.; Rowland, S.; Nicoll-Griffith, D. A.; Kelly,
N.; Chan, C.-C. J. Chromatogr. B 2001, 754, 245.
17. A scoring system was implemented to evaluate the severity of the AE’s: a score
of 1 represents light AE’s whereas severe AE’s were given a score of 3.
18. 50 mM of tris-(hydroxymethyl)aminomethane and 150 mM of NaCl. pH
adjusted to 7.6 with HCl.
In conclusion, we demonstrated that with a systemically-dis-
tributed azetidinyl pyridazine SCD inhibitor it was not possible
to maintain weight gain benefits in mice without also negatively
affecting skin and eye functions. As a result, there is a need to iden-
tify new scaffolds displaying a differing tissue distribution profile
and the preparation of a radiolabeled SCD inhibitor which serves
as a probe for a SPA binding assay in an HTS campaign is disclosed.
Full details about this work will be reported shortly.
19. Two sets of samples were analyzed. Sample set 1 was a 100 nM solution of the
SCD inhibitor in 100
100 nM solution of the inhibitors in 100
prepared in plastic eppendorfs. For each sample, 45
out using MAXYMUM RECOVERY (marketed by AXYGEN) 200
glass vial with built-in insert. Fourty-five micro liters of acetonitrile
l
L of a 1:1 EtOH/TBS buffer mixture. Sample set 2 was a
L of TBS buffer. Solutions were
L of solution was pipetted
L pipette tips to
l
l
l
a
a
containing and internal standard (IS) were added. Vials were capped, vortexed
and used as is. Each sample was done in duplicate. For data analysis, the
References and notes
concentrations from the sample set
1 were set at 100 nM (no external
quantitative standards) and runs from sample set
comparison of peak area (standardized with an IS).
2 were then a direct
1. World Health Organization, Fact sheet No. 311, September 2006 (data available
at www.who.int).
20. Adams, A. D.; Green, A.I.; Szewczyk, J. W. WO 2007/064553.
21. Skorey, K. in preparation.
2. World Health Organization, Fact sheet No. 312, November 2009 (data available
at www.who.int).