Hemodynamic effects of potent and selective JNK inhibitors in anesthetized rats: Implication for targeting protein kinases in metabolic diseases

Article abstract of DOI:10.1016/j.bmcl.2006.10.013

The hemodynamic effects of a series of potent and selective 4-aminopyridine carboxamide-based pan-JNK inhibitors were assessed in an anesthetized rat model. The effects of these agents on mean arterial pressure, heart rate, cardiac contractility, and peri

Full text of DOI:10.1016/j.bmcl.2006.10.013

Bioorganic & Medicinal Chemistry Letters 17 (2007) 495–500
Hemodynamic effects of potent and selective JNK inhibitors
in anesthetized rats: Implication for targeting protein
kinases in metabolic diseases
Gang Liu,a,* Hongyu Zhao,a Bo Liu,a Zhili Xin,a Mei Liu,a Michael D. Serby,a
Nathan L. Lubbers,b Deborah L. Widomski,b James S. Polakowski,b David W. A. Beno,c
James M. Trevillyana and Hing L. Shama
aMetabolic Disease Research, Global Pharmaceutical Research and Development, Abbott Laboratories, Abbott Park,
IL 60064-6101, USA
bIntegrative Pharmacology, Global Pharmaceutical Research and Development, Abbott Laboratories, Abbott Park,
IL 60064-6101, USA
cExploratory Pharmacokinetics, Global Pharmaceutical Research and Development, Abbott Laboratories, Abbott Park,
IL 60064-6101, USA
Received 25 September 2006; revised 5 October 2006; accepted 5 October 2006
Available online 10 October 2006
Abstract—The hemodynamic effects of a series of potent and selective 4-aminopyridine carboxamide-based pan-JNK inhibitors were
assessed in an anesthetized rat model. The effects of these agents on mean arterial pressure, heart rate, cardiac contractility, and
peripheral vascular resistance are described, and the implication for targeting protein kinases in metabolic diseases is discussed.
ꢀ 2006 Elsevier Ltd. All rights reserved.
The c-Jun N-terminal kinases (JNKs) are members of
the mitogen-activated protein kinase (MAPK) family
and regulate signal transduction in response to environ-
mental stress, cytokines, and free fatty acids. Three dis-
tinct genes encoding JNKs have been identified (jnk1,
jnk2, and jnk3), and at least 10 different splicing isoforms
exist in mammalian cells.1 Up-regulation of JNK expres-
sion and activity has been implicated in a number of dis-
ease states, including diabetes, obesity, inflammation,
auto-immune diseases, and neuro-degenerative diseas-
es.2 Therefore, small molecule inhibitors of JNKs have
been pursued in a number of laboratories as potential
therapeutic agents for the aforementioned ailments.3
We are particularly intrigued by the potential of inhibit-
ing JNK1 activity for boosting the metabolic insulin-sig-
naling cascade and ameliorating insulin resistance.4
Obesity and type 2 diabetes are prevalent and serious
metabolic diseases. The expenses associated with treat-
ing these metabolic diseases, including diabetes and
obesity, exceed 20% of the national health care budget.
However, obesity and type 2 diabetes are still regarded
as chronic, non-life-threatening diseases. Because the
treatment regimen tends to be chronic, the safety
requirement on these drugs is extremely high.
Recently, cardiovascular safety has attracted both scien-
tific and media attention wꢁith the high-profile withdrꢁaw-
al of propulsid (Cisapride )5 and rofecoxib (Vioxx ).6
In addition, reports of deleterious cardiovascular effects
of several marketed protein tyrosine kinase inhibitors,
including a monoclonal antibody,7 have appeared in
the literature. The chronic use of imatinib (Gleevecꢁ),
a Bcr-Abl tyrosine kinase inhibitor, has been linked to
a striking 50% decrease in the pumping capacity of the
heart, and the development of severe congestive heart
failure in some patients.8 The unanticipated cardiotoxic-
ity of imatinib highlights the need for more vigorous
preclinical and clinical evaluation of the cardiovascular
liabilities of new protein kinase inhibitors.
Keywords: c-Jun N-terminal kinase inhibitor; Anesthetized rat model;
Cardiotoxicity; Metabolic diseases.
*
Corresponding author. Tel.: +1 847 935 1224; fax: +1 847 938
1674; e-mail: gang.liu@abbott.com
0960-894X/$ - see front matter ꢀ 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2006.10.013

496
G. Liu et al. / Bioorg. Med. Chem. Lett. 17 (2007) 495–500
NH2
NH2
N
O
O
N
S
Cl
O
S
O
O
H
H
N
N
N
O
O
2
O
1
IC50 = 0.019 μM vs. JNK1
IC50 = 0.12 μM vs. JNK2
IC50 = 0.024 μM vs. JNK1
IC50 = 0.074 μM vs. JNK2
Figure 1. Selected 4-aminopyridine carboxamide JNK inhibitors reported from our laboratories.
We recently reported a series of 4-aminopyridine carb-
oxamides, represented by 1 and 2 (Fig. 1), as potent
JNK1 and JNK2 inhibitors with minimal cross-activity
against other kinases.9 These compounds also were
shown to exhibit excellent selectivity (>10,000-fold) over
hERG channel in a dofetilide binding assay.
per assay than the anesthetized dog model. In addition,
easier access to animals and reduced instrumentation
contribute to a greater capacity for compound through-
put in the anesthetized rat assay compared to the dog as-
say. Such assays have been successfully implemented in
the lead optimization programs for melanin concentrat-
ing hormone receptor 1 (MCH1) antagonists13 and tran-
sient receptor potential channel vanilloid 1 (TRPV1)
receptor antagonists.14
Because of the stringent safety requirements for the
treatment of chronic diseases such as type 2 diabetes,
and the potential cardiovascular liabilities associated
with protein kinase inhibitors, we initiated an assess-
ment of the anesthetized in vivo hemodynamic effects
of these JNK inhibitors early in our lead optimization
process.
Therefore, from a technical standpoint, we hypothesized
that the inactin-anesthetized rat model could be used as
an effective screening model to establish structure–activ-
ity relationships (SAR) regarding the cardiovascular ef-
fects of our pan-JNK inhibitors. For the sake of direct
comparison, 30 lg/mL (plasma concentration) was tar-
geted as a 30· therapeutic concentration. Aminopyri-
The pentobarbital anesthetized dog has been widely
accepted as the gold standard in assessing the cardiovas-
cular effects of new chemical entities (NCEs).10 Howev-
er, since we identified the need to screen JNK inhibitors
for cardiovascular liabilities at a much earlier stage in
our lead optimization program, we sought to implement
a higher throughput and less compound-intensive,
whole-animal model that could be applied as an early
screening tool.
dine carboxamide
1 was selected as an initial
compound for evaluation in the anesthetized rat model.
Compound 1 represents a potent JNK inhibitor with a
cellular EC50 value of 318 nM against c-Jun phosphory-
lation in HepG2 cells. In the inactin-anesthetized rat
cardiovascular screening model,15 aminopyridine 1 de-
creased MAP beginning with the 10 mg/kg infusion,
the maximum decrease was 41% at 90 min (Fig. 1).
The MAP of two animals fell below the minimum pres-
sure limit during the last 5 min of the final infusion. HR
began to decrease slightly with the 30 mg/kg infusion
while there was a slight increase in cardiac contractility
(dP/dt). Maximum change for both parameters was less
than 15%. PVR began to decrease during the 10 mg/kg
infusion, the maximum decrease was 25% at 90 min.
These preliminary data suggest that as the concentration
increases, compound 1 may be acting, at least partially,
as a peripheral vasodilator (decrease in vascular resis-
tance). This is supported by the minimal effects on HR
and cardiac contractility. The decrease in vascular resis-
tance contributes to the decreases in MAP. The end of
infusion (EOI) plasma concentration at 90 minutes
was slightly lower than projected, but still within the rel-
evant range. The hemodynamic profile of the other ac-
tive JNK inhibitor 2 is similar to that of 1, in terms of
the changes in MAP and PVR (Table 1).
The acutely instrumented anesthetized rat cardiovascu-
lar model11,12 is a predictive in vivo model of cardiovas-
cular function that effectively interrogates lead
molecules for undesirable cardiovascular effects. Studies
are conducted under anesthesia (inactin) using a series
of intravenous infusions and periodic collection of small
(150 lL) blood samples to assess active drug levels, pro-
viding concentration-response functions for clinically
relevant endpoints of mean arterial pressure (MAP),
heart rate (HR), cardiac contractility, and peripheral
vascular resistance (PVR). The predictive value of the
model arises from its ability to identify compounds that
produce effects on cardiovascular function at relevant
doses or plasma concentrations, results that would pre-
clude them from further characterization.
In this screening paradigm, a compound producing an
effect greater than 15% compared to vehicle within an
estimated 30-fold window is unacceptable for further
characterization. Also, if MAP decreases below the min-
imum pressure limit (70 mmHg), the infusion is stopped
and a blood sample collected. This minimum pressure
level was implemented to ensure a functional cardiovas-
cular system for determination of drug plasma levels.
To understand whether the hemodynamic changes in-
duced by 1 and 2 are related to the general structure
of aminopyridine or related to the inhibition of JNK sig-
naling pathway, we also profiled two inactive analogs (3
and 4) from the same series. As shown in Table 1, 4-
hydroxypyridine 3 only reduced MAP slightly in rats
at higher EOI plasma levels, while having no effect on
One significant advantage of the anesthetized rat model
is that it requires significantly less compound (<200 mg)

G. Liu et al. / Bioorg. Med. Chem. Lett. 17 (2007) 495–500
497
Table 1. Hemodynamic effects of active JNK1 inhibitors versus inactive analogs in anesthetized rat model
Class
Compound
JNK1
IC50 (lM)
Maximum change
in MAPa
Maximum
change in HRa
Maximum change
in dP/dta
Maximum change
in PVRa
EOI plasma
90 min (lg/mL)b
Active
1
2
3
4
0.019
0.024
>10
#41%
#20%
# <15%
NE
# <15%
# <15%
NE
" <15%
" <15%
NE
#25%
#51%
NE
19.3
14.7
39.9
23.6
Inactive
6.4
NE
"41%
#46%
NE, no effect.
a Mean values based on the results from three animals.
b Mean values based on concentrations from three animals.
the other hemodynamic parameters measured. On the
other hand, N-methylated carboxamide 4 showed a dif-
ferent CV profile from that exhibited by the active
aminopyridine 1, producing a large decrease in PVR
(46%). The large compensatory increase in cardiac con-
tractility (41% increase) likely contributed to maintain-
ing MAP. These results suggest that the observed
hemodynamic effects of 1 and 2 may not be linked to
the general structure of aminopyridine carboxamide.
However, we could not rule out the possible linkage be-
tween the hemodynamic effects and JNK activity inhibi-
tion, or potential secondary pharmacodynamic
properties (off-target effects) of these structures.
Considerable effort was made to assign the observed car-
diovascular toxicity to cross-activity at specific recep-
tors, transporters, and ion channels via a CEREP-
panel18 affinity screening. Unfortunately, we were un-
able to identify a molecular target for which activity of
the studied compound 1 would correlate with the ob-
served hemodynamic effects, as cross-activity of 1 was
minimal against 75 different receptors, transporters,
and ion channels (<30% inhibition of control-specific
binding at 10 lM). Furthermore, we could not attribute
the cardiovascular toxicity to the general state of non-
JNK kinase inhibition either. As shown in the Supple-
mentary material, 1 is an extremely selective ATP com-
petitive JNK inhibitor, exhibiting no significant
inhibitory activity toward any other kinases in a panel
of 77 kinases19 at the highest concentration tested except
CSF1R and ERK2, but still with a selectivity window
greater than 100-fold (Fig. 2).
The synthesis of active analogues has been reported pre-
viously.9,16 The preparation of inactive analog 3 is de-
scribed in Scheme 1. Starting from the readily
available a-oxoketene dithoacetal 5,17 cyclization with
cyanoacetamide yielded a hydroxypyridine sodium salt,
which was alkylated with i-propyl iodide to give highly
functionalized pyridine 6. Hydrolysis of the acetal and
Ag2O oxidation of the resulting aldehyde provided the
acid 7. Condensation of 7 with p-methylsulfonylbenzyl-
amine generated amide 8. Oxidation of sulfide with m-
CPBA to sulfone, followed by hydrolysis, afforded the
4-hydroxypyridine 3. Straightforward methylation of 1
provided N-methyl carboxamide 4.
We then launched a broader screening campaign to
identify active JNK inhibitors with an improved cardio-
vascular profile over that of 1. Nine potent JNK inhib-
itors from the aminopyridine series were evaluated in the
anesthetized rat screening to establish structural param-
eters responsible for cardiovascular liabilities. Since the
decrease in MAP was produced by the decrease in
PVR for both 1 and 2, the model was streamlined to
S
S
N
S
S
N
c,d
e
a,b
O
O
O
HO
N
O
N
O
O
O
O
5
6
7
OH
N
S
N
N
O S
O
O S
O
f,g
H
H
N
N
O
N
O
O
O
8
3
NH2
N
h
O S
O
1
N
N
O
O
4
Scheme 1. Reagents and conditions: (a) cyanoacetamide, Na, t-BuOH, reflux, 5 h; (b) i-PrI, DMF, 120 ꢂC, 30 min, 80%; (c) 1 N aq HCl, THF, 65 ꢂC,
2 h, 90%; (d) AgNO3, aq KOH, EtOH, rt, over night, 75%; (e) p-methylsulfonylbenzyl amine, TBTU, Et3N, DMF, rt, 30 min, 81%; (f) m-CPBA,
CH2Cl2, rt, over night; (g) 3 N aq NaOH, 1,4-dioxane, 80 ꢂC, 3 h, 65% over two steps; (h) NaH, DMF, MeI, rt, 72%.

498
G. Liu et al. / Bioorg. Med. Chem. Lett. 17 (2007) 495–500
Change in Mean Arterial Pressure Change in Heart Rate
Baseline
3
10
30
mg/kg
3
10
30
mg/kg
Baseline
50
25
0
100
75
50
25
0
-25
-50
-75
-100
-25
-50
-30
0
30
60
90
-30
0
30
60
90
Time (Minutes)
Time (Minutes)
Change in dP/dt @ 50 mmHg
Change in Peripheral Vascular Resistance
Baseline
3
10
30
mg/kg
Baseline
10
3
30
mg/kg
4
2
2000
0
0
-2
-4
-2000
-30
0
30
60
90
-30
0
30
60
90
Time (Minutes)
Time (minutes)
Figure 2. Hemodynamic effects induced by 1 in anesthetized rats. The red line represents those of the vehicle (10% DMSO in PEG400), and the blue
line represents those of compound 1.
maximize throughput by focusing on MAP, HR, and
contractility. The results are summarized in Table 2.
Once again, one common hemodynamic change was ob-
served in these studies: a consistent, across-the-board
decrease of MAP induced by these analogues (9a–9i)
at comparable plasma concentrations, regardless of the
structural variations at the 2-, 5-, and 6-position of the
aminopyridine core. The effects on HR and cardiac con-
tractility were generally mild to non-existent, with the
exception of 9a, which induced a large increase of heart
contractility. It is worth noting that for potent JNK
inhibitors 9g and 9i, the decrease in MAP was very close
to the acceptable range (<15%), while having no effects
on HR and contractility. However, the EOI plasma con-
centrations were found to be slightly lower than those
achieved by 1 and 2.
Table 2. Hemodynamic effect profiling of additional JNK inhibitors in anesthetized rat
NH2
R2
R1
H
R3
N
N
O
9a-i
Compound R1
R2
R3
JNK1
IC50 (lM) change in change
Maximum Maximum Maximum EOI plasma
change
90 min (lg/mL)b
MAPa
in HRa
in dP/dta
9a
9b
9c
9d
–OEt
–CN 2-Sulfonamidophenyl–
–Cl 2-Sulfonamidophenyl–
0.012
0.023
0.030
0.061
#31%
#20%
#23%
#20%
"22%
"13%
NE
"56%
NE
25.8
12.0
–OEt
–OEt
–OEt
–CN 4-Chloro-3-pyridyl–
–CN N-(5-Nitropyridin-2-
yl)aminomethyl–
# >15%
"12%
NE
9e
9f
–OEt
–OEt
–CN 4-(Phenylsulfinyl)phenyl–
–CN 4-(Phenylsulfonyl)phenyl– 0.13
0.050
#24%
#24%
#24%
#18%
#23%
#30%
#23%
NE
#11%
NE
"24%
NE
NE
9g
9h
9i
Cyclopropylmethoxy –CN 4-(Methylsulfonyl)phenyl– 0.023
Cyclopentane-amino –CN 4-(Methylsulfonyl)phenyl– 0.096
8.2
10.5
#6%
NE
"13%
NE
NE
3-Thieno
NA
NA
–CN 4-(Methylsulfonyl)phenyl– 0.081
0.002
0.034
10
11
#22%
#11%
9.0
#20%
NA, no applicable; NE, no effect.
a Mean values based on the results from three animals.
b Mean values based on concentrations from three animals.

G. Liu et al. / Bioorg. Med. Chem. Lett. 17 (2007) 495–500
499
NH
N
O
OH
N
O
H
N
N
N
N
N
H
Cl
O
10 (Merck)
IC50 ~ 0.002 μM
vs. JNK1 & 3
11 (Hoffman LaRoche)
IC50 = 0.034 μM
vs. JNK1
N
Cl
H
Figure 3. Selected literature JNK inhibitors evaluated in anesthetized rat model.
We also evaluated a number of JNK inhibitors reported
in the literature in the anesthetized rat model. The
hemodynamic effects of two of them are summarized
in Table 2. The imidazole pyrimidine 1020 (Fig. 3) was
shown to have severe cardiovascular effects. It induced
a large decrease in MAP (30%) during the last infusion
period. The MAP of two animals fell below the mini-
mum pressure limit during the last 5 min of the final
infusion. HR was also lowered by 22% in the final infu-
sion period. Since 10 was a non-selective JNK and p38
inhibitor, it was difficult to interpret the origin of the
hemodynamic effects of 10. Oxindole 1121 (Fig. 3) de-
creased MAP and cardiac contractility by 23% and
20% at 90 min (end of infusion), respectively.
In summary, we have described the application of estab-
lished in vivo anesthetized rat cardiovascular safety
screening in the JNK1 inhibitor lead selection process.
This report highlights the hemodynamic effects of the
aminopyridine JNK inhibitors including hypotension
and a reduction in vascular resistance. Such findings
underscore the value and importance of positioning in vi-
vo cardiovascular safety screening early in the lead selec-
tion process for kinase programs. Our finding, coupled
with recent observation of cardiac toxicity on the other
marketed protein kinase inhibitors, calls for more vigor-
ous screening of future kinases inhibitors, particularly
for the chronic indications such as metabolic diseases.
Thus, we discovered a hypotensive effect of a series of
highly potent and selective JNK inhibitors in vivo.
The decrease in MAP is likely driven in large part by
the decrease in peripheral vascular resistance. Even
though the hypotensive effect could be attenuated, we
could not completely eliminate it through structural
modification. A similar hypotensive effect was also ob-
served with two structurally distinct classes of literature
JNK inhibitors. Interestingly, our results with imatinib
in anesthetized dog also indicated that it decreases arte-
rial pressure with concomitant reduction in systemic
vascular resistance (Dr. Ryan M. Fryer, unpublished
internal results). The mechanism of action for the car-
diovascular effect is currently not understood. The re-
cent study with imatinib suggested that protein kinases
might be involved in certain hitherto unknown cytopro-
tective functions in the heart.8
Acknowledgments
The authors thank Ms. Sanyi Wang, Ms. Jill Clampit,
and Dr. Eric Johnson for the execution of the binding,
cellular, and selectivity assays.
Supplementary data
Supplementary data associated with this article can be
found, in the online version, at doi:10.1016/
j.bmcl.2006.10.013.
References and notes
1. Gupta, S.; Barrett, T.; Whitmarsh, A. J.; Cavanagh, J.;
Sluss, H. K.; Derijard, B.; Davis, R. J. EMBO J. 1996, 15,
2760.
Protein kinase inhibitors have become the single largest
class of drug discovery targets in the last 10 years. Most
of the kinase inhibitors are developed for life-threaten-
ing oncology indications. However, there is also great
interest in developing kinase inhibitors for more chron-
ic, non-life-threatening indications, such as auto-im-
mune diseases, diabetes, and obesity. Preclinical
evaluations of the cardiovascular effects of protein ki-
nase inhibitors have not been widely reported. This re-
port highlighted the potential cardiovascular liability
associated with serine/threonine kinase JNK inhibitors,
potentially other classes of protein kinase inhibitors as
well. Such potential issue should establish the impor-
tance of early screening for cardiovascular safety in ki-
nase inhibitor programs, particularly in the area of
chronic disease indications, such as diabetes and obesity.
2. Manning, A. M.; Davis, R. J. Nat. Rev. Drug Discovery
2003, 2, 554.
3. Liu, G.; Rondinone, C. M. Curr. Opin. Investig. Drugs
2005, 6, 979.
4. Wellen, K. E.; Hotamisligil, G. S. J. Clin. Invest. 2005,
115, 1111.
5. Paakkari, I. Toxicol. Lett. 2002, 127, 279.
6. (a) Solomon, S. D.; McMurray, J. J. V.; Pfeffer, M. A.;
Witters, J.; Fowler, R.; Finn, P.; Anderson, W. F.;
Zauber, A.; Hawk, E.; Bertagnolli, M. N. Engl. J. Med.
2005, 352, 1071; (b) Levesque, L. E.; Brophy, J. M.;
Zhang, B. Ann. Intern. Med. 2005, 142, 481.
7. Smith, K. L.; Dang, C.; Seidman, A. D. Expert Opin. Drug
Saf. 2006, 5, 619.
8. Kerkela, R.; Grazette, L.; Yacobi, R.; Iliescu, C.; Patten,
R.; Beahm, C.; Walters, B.; Shevtsov, S.; Pesant, S.;
Clubb, F. J.; Rosenzweig, A.; Salomon, R. N.; Van Etten,

500
G. Liu et al. / Bioorg. Med. Chem. Lett. 17 (2007) 495–500
R. A.; Alroy, J.; Durand, J. B.; Force, T. Nat. Med. 2006,
12, 908.
dt50), is derived from LVP. Via laparotomy, peripheral
blood flow (PBF) was measured by placing a cuff type flow
probe around the upper abdominal aorta just below the
diaphragm and above the renal arteries. Posthoc, periph-
eral vascular resistance, PVR, was calculated as
PVR = MAP/PBF. PVR is being used as a rat approxi-
mation of dog systemic vascular resistance. Rats were
randomly chosen for each treatment group. Baseline levels
for all parameters were within normal range for each rat.
Following a 30-min control period, compounds or vehicle
(10% DMSO/PEG400; 1 mL/kg) were administered over
three ascending doses, each delivered over a 30-min
infusion at doses of 3, 10, and 30 mg/kg, respectively. A
small volume (150 lL) blood sample was collected at the
end of each infusion for drug level determination. For
each parameter the baseline for each animal was defined as
the time 0 reading. For all post treatment time points, the
change from baseline for each animal was calculated for
each parameter.
9. Zhao, H.; Serby, M. D.; Xin, Z.; Szczepankiewicz, B. G.;
Liu, M.; Abad-Zapatero, C.; Fry, E. H.; Johnson, E. F.;
Wang, S.; Pederson, T.; Gum, R. J.; Haasch, D. E.;
Clampit, J. E.; Rondinone, C.; Trevillyan, J. M.; Sham, H.
L.; Liu, G. J. Med. Chem. 2006, 49, 4455.
10. Greaves, P.; Williams, A.; Eve, M. Nat. Rev. Drug
Discovery 2004, 3, 226.
11. Polakowski, J. S.; Segreti, J. A.; Cox, B. F.; Hsieh, G. C.;
Kolasa, T.; Moreland, R. B.; Brioni, J. D. Clin. Exp.
Pharm. Phys. 2004, 31, 837.
12. Reinhart, G. A.; Fryer, R. M.; Osinski, M. A.; Polakow-
ski, J. S.; Cox, B. F.; Gintant, G. A. Curr. Opin. Chem.
Biol. 2005, 9, 1.
13. Kym, P. R.; Souers, A. J.; Campbell, T. J.; Lynch, J. K.;
Judd, A. S.; Iyengar, R.; Vasudevan, A.; Gao, J.;
Freeman, J. C.; Wodka, D.; Mulhern, M.; Zhao, G.;
Wagaw, S. H.; Napier, J. J.; Brodjian, S.; Dayton, B. D.;
Reilly, R. M.; Segreti, J. A.; Fryer, R. M.; Preusser, L. C.;
Reinhart, G. A.; Hernandez, L.; Marsh, K. C.; Sham, H.
L.; Collins, C. A.; Polakowski, J. S. J. Med. Chem. 2006,
49, 2339.
16. Liu, G.; Zhao, H.; Liu, B.; Xin, Z.; Mei Liu; Kosogof, C.;
Szczepankiewicz, B. G.; Wang, S.; Clampit, J. E.; Gum, R.
J.; Haasch, D. L.; Trevillyan, J. M.; Sham, H. L. Bioorg.
Med. Chem. Lett. 2006, 16, 5723.
14. Drizin, I.; Gomtsyan, A.; Bayburt, E. K.; Schmidt, R. G.;
Zheng, G. Z.; Perner, R. J.; DiDomenico, S.; Koenig, J.
R.; Turner, S. C.; Jinkerson, T. K.; Brown, B. S.; Keddy,
R. G.; McDonald, H. A.; Honore, P.; Wismer, C. T.;
Marsh, K. C.; Wetter, J. M.; Polakowski, J. S.; Segreti, J.
A.; Jarvis, M. F.; Faltynek, C. R.; Lee, C. H. Bioorg. Med.
Chem. 2006, 14, 4740.
15. The methodology of the anesthetized rat model is
summarized here: male Sprague–Dawley rats were anes-
thetized with the long-acting barbiturate, inactin (100 mg/
kg). Catheters were placed in both femoral arteries; one
for measurement of MAP and HR, the other for collection
of blood samples. Additional catheters were placed in the
femoral vein for compound administration and saline
infusion (to maintain hydration). A specialized transducer
tip catheter was advanced into the left ventricle of the
heart for measurement of left ventricular pressure (LVP).
The index of cardiac contractility, dP/dt at 50 mmHg (dP/
17. Mahata, P. K.; Syam Kumar, U. K.; Sriram, V.; Ila, H.;
Junjappa, H. Tetrahedron 2003, 59, 2631.
18. Compound 1 was tested for % inhibition at 10 lM in a
standard panel of 75 receptors, transporters, and ion
channels offered from CEREP (<www.cerep.com/>).
19. Experimental details for kinase selectivity screening can be
found in reference: Szczepankiewicz, B. G.; Kosogof, C.;
Nelson, L. T. J.; Liu, G.; Liu, B.; Zhao, H.; Serby, M. D.;
Xin, Z.; Liu, M.; Gum, R. J.; Haasch, D. L.; Wang, S.;
Clampit, J. E.; Johnson, E. F.; Lubben, T. H.; Stashko, M.
A.; Olejniczak, E. T.; Sun, C.; Dorwin, S. A.; Haskins, K.;
Abad-Zapatero, C.; Fry, E. H.; Hutchins, C. W.; Sham,
H. L.; Rondinone, C. M.; Trevillyan, J. M. J. Med. Chem.
2006, 49, 3563.
20. Scapin, G.; Patel, S. B.; Lisnock, J.; Becker, J. W.;
LoGrasso, P. V. Chem. Biol. 2003, 10, 705.
21. Luk, K.-C.; Mahaney, P.E.; Mischke. S. G. WO-0035906,
2000.