Piperidine-based heterocyclic oxalyl amides as potent p38α MAP kinase inhibitors

Article abstract of DOI:10.1016/j.bmcl.2009.12.031

The design and synthesis of a new class of p38α MAP kinase inhibitors based on 4-fluorobenzylpiperidine heterocyclic oxalyl amides are described. Many of these compounds showed low-nanomolar activities in p38α enzymatic and cell-based cytokine TNFα produc

Full text of DOI:10.1016/j.bmcl.2009.12.031

Bioorganic & Medicinal Chemistry Letters 20 (2010) 1059–1062
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Piperidine-based heterocyclic oxalyl amides as potent p38
kinase inhibitors
a MAP
*
Babu J. Mavunkel, John J. Perumattam, Xuefei Tan , Gregory R. Luedtke, Qing Lu, Don Lim, Darin Kizer,
*
Sundeep Dugar, Sarvajit Chakravarty, Yong-jin Xu, Joon Jung, Albert Liclican, Daniel E. Levy , Jocelyn Tabora
Departments of Medicinal Chemistry and Biology, Scios, Inc., 6500 Paseo Padre Parkway, Fremont, CA 94555, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
The design and synthesis of a new class of p38a MAP kinase inhibitors based on 4-fluorobenzylpiperidine
heterocyclic oxalyl amides are described. Many of these compounds showed low-nanomolar activities in
Received 22 October 2009
Revised 4 December 2009
Accepted 7 December 2009
Available online 11 December 2009
p38 enzymatic and cell-based cytokine TNF production inhibition assays. The optimal linkers between
a
a
the piperidine and the oxalyl amide were found to be [6,5] fused ring heterocycles. Substituted indoles
and azaindoles were favored structural motifs in the cellular assay.
Ó 2009 Elsevier Ltd. All rights reserved.
Keywords:
p38
Piperidine
Heterocyclic
MAP
Kinase
Inhibitor
Oxalyl
Amide
Inflammatory
The pro-inflammatory cytokines, tumor necrosis factor alpha
(TNF ), and interleukin 1 beta (IL-1b) are known to be involved
vated, translates multiple stimuli to multiple responses and results
in the release of a pro-inflammatory cassette of cytokines including
a
in the pathogenesis of inflammatory disorders such as rheumatoid
arthritis (RA), Crohn’s disease, and psoriasis.1–4 The FDA approval
of biologics (EnbrelÒ, RemicadeÒ, HumiraÒ, and KineretÒ) specifi-
cally targeting these cytokines by mimicking their receptors5–8
have demonstrated the effectiveness of this treatment paradigm.
However, like many biological agents, these drugs have disadvan-
tages relating to high costs and inconvenient dosing regimens.
Thus, there remain unmet needs for developing safe, effective,
and orally active small molecule inhibitors that serve the same
therapeutic functions.
IL-1b, IL-6, and TNFa 10
. As p38a activation is not involved in nor-
mal physiology, it is believed that inhibition of this kinase target
could result in a reduction of the levels of these cytokines that
are thought to play a pathophysiological role in many inflamma-
tory diseases. Therefore, p38a MAP kinase is considered to be a po-
tential therapeutic target for the treatment of inflammatory
diseases such as RA11,12 and has been the focus of many clinical
candidates by various pharmaceutical companies in the past
decades.13,14
We previously reported the discovery of a novel series of 4-flu-
p38 MAP kinase is a serine–threonine protein kinase and is
identified initially as the molecular target of a pyridylimidazole
class of compounds. These compounds are known to inhibit the
orobenzylpiperidine indole-based p38
resented by Figure 1.15 The proposed binding mode of 1 in the ATP
site of p38 MAP kinase (Fig. 2) shows this class of molecules
a MAP kinase inhibitors rep-
a
biosynthesis of TNF
lated human monocytes.9 Among the four known isoforms of p38
, b, , and d), the and b isoforms are the most widely expressed.
While the function of the b isoform is not well understood, the
a and IL-1b in lipopolysaccharide (LPS)-stimu-
(a
c
a
a
isoform has been shown to be a control point which, when acti-
* Corresponding authors. Tel.: +1 650 704 3051 (D.E.L.); +1 011 86 22 66282937
(X.T.).
Figure 1. Structures and potencies of 4-fluorobenzylpiperidine indole analogs, 1
and 2.
E-mail addresses: Xuefei827@gmail.com (X. Tan), del345@gmail.com (D.E. Levy).
0960-894X/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2009.12.031

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B. J. Mavunkel et al. / Bioorg. Med. Chem. Lett. 20 (2010) 1059–1062
Table 1
p38
a
enzyme and cellular activity for compounds 1, 2, and 4-9
25
26
Compd Linker
p38a
IC50
(l
M,
dWBA IC50
(lM,
n = 3)
n = 3)
1
2
0.018
0.10
0.03
0.010
4
5
0.945
>10
17%@1
ND
lM
6
0.10
26%@0.3
ND
lM
7
8
9
0.090
0.60
0.06
Figure 2. Proposed binding mode of piperidine indole analog 1 in the ATP site of
p38a MAP kinase.
10%@1
0.048
lM
forming a critical hydrogen bond with the kinase backbone at
hinge amino acid Met-109.16 Additionally, the 4-fluorobenzyl
group occupies an adjacent hydrophobic pocket that is both critical
for activity and, as shown in the literature,17–19 is a crucial region
relating to kinase specificity. Substitutions such as chlorine, meth-
oxy and methyl ortho to the amide moiety at indole position 6
were found to improve p38
a enzymatic potency presumably due
to the restricted conformation along the carbonyl aryl C–C bond.
An oxalyl amide modification at C-3 of the indole was found to fur-
ther improve enzyme activity. While findings in several of these
areas will be the focus of future publications, this Letter describes
structure–activity data relating to the heterocyclic core only.
Due to a lack of X-ray crystallographic data at the time of our
studies, the exact role of the oxalyl amide was unknown and atten-
tion was focused on whether an indole was the optimal linker join-
ing the piperidine amide to the oxalyl amide. Thus, a number of
heterocyclic analogs with either different lengths or different elec-
tronic properties relative to indole were proposed. Representative
analogs, compounds 4–9, are illustrated in Table 1.
4-Fluorobenzylpiperidine 3 was a common intermediate used
in the synthesis of each analog, and was prepared by reacting
diethyl 4-fluorobenzylphosphonate with BOC-protected piperi-
done via a Horner–Emmons reaction, followed by catalytic hydro-
genation of the double bond and Boc-deprotection (Scheme 1).
Where direct acylation of heteroaromatics with oxalyl chloride
could be utilized, the preparation of target compounds was
straightforward as illustrated in Scheme 2 for compounds 1, 2,
and 9.
Scheme 1. Preparation of 4-fluorobenzylpiperidine. Reagents and conditions: (a)
NaH, DMF then 4-oxopiperidine-1-tert-butyl carbamate, 48%; (b) H2, Pd/C, then
HCl/ether, 98%.
For the second set of conditions, a mild basic transformation
was developed for the preparation of compound 6 (Scheme 4). As
shown, the indazole 3-aldehyde intermediate was prepared in a
one step process from its corresponding indole derivative via dou-
ble nitrosation followed by ring rearrangement.24 The aldehyde
group was then protected as a dithiane prior to introduction of
the methyl group at N-1 of the indazole. Treatment with butyl lith-
ium followed by quenching with dimethylcarbamyl chloride gave
the protected oxalyl amide. Upon deprotection, compound 6 was
obtained.
Flipped indole analog 7 was prepared according to Scheme 5 by
first reacting 4-nitrophenylglyoxylic acid with dimethyl amine fol-
lowed by SnCl2 reduction to give corresponding aniline. Reductive
amination with 1,1-dimethoxyacetone followed by treatment with
AlCl3 resulted in cyclization forming the desired indole core. Subse-
quent reaction with phosgene followed by quenching with 4-fluor-
obenzylpiperidine gave the desired oxalyl amide.
For heterocycles inert to treatment with oxalyl chloride, an
aldehyde was chosen as a precursor for the oxalyl moiety and
one of two coupling protocols were utilized. In the first set of con-
ditions, reaction of an aldehyde with TMSCN followed by treat-
ment with acid and subsequent base hydrolysis provided an
hydroxy aryl acetic acid. After first forming a methyl ester, PCC oxi-
dation of the hydroxyl group provided the -keto ester. Hydrolysis
with NaOH and coupling with dimethyl amine then gave the de-
sired -keto amide. Both compounds 4 and 5 were prepared using
this route (Scheme 3).
a-
a
Finally, regarding heterocyclic systems such as benzofuran that
could not be incorporated utilizing an aldehyde route, synthesis
was achieved through oxidation of a methylene group to an oxalyl
moiety (Scheme 6). Beginning with methyl 2,4-dihydroxybenzoate,
a

B. J. Mavunkel et al. / Bioorg. Med. Chem. Lett. 20 (2010) 1059–1062
1061
Scheme 4. Reagents and conditions: (a) EDCI, DMAP, 6-chloro-1H-indole-5-
carboxylic acid,20 Et3N, DMF, 81%; (b) 6 N HCl, NaNO2, 56%; (c) HS(CH2)3SH, BF3,
90%; (d) NaH, DMF then MeI, 94%; (e) n-BuLi, THF then ClCONMe2, 73%; (f)
(CF3CO)2IPh, CH3CN–H2O, 90%.
Scheme 2. Reagents and conditions: (a) 6-chloro-1H-indole-5-carboxylic acid,20
EDCI, DMAP, DIPEA, DCM, 81%; (b) oxalyl chloride then dimethyl amine, 72–87%; (c)
6-methoxy-1H-indole-5-carboxylic acid,20 EDCI, DMAP, DIPEA, DCM, 90%; (d) TBTU,
6-methoxy-1H-pyrrolo[2,3-b]pyridine-5-carboxylic acid,21 Et3N, DMF, 83%.
Scheme 5. Reagents and conditions: (a) dimethyl amine, EDCI, THF, 79%; (b)
SnCl2ÁH2O, concd HCl, 60 °C, 85%; (c) 1,1-dimethoxyacetone, anhydrous Na2SO4,
AcOH, NaBH(OAc)3, 87%; (d) AlCl3, DCM, 60 °C, 24%; (e) phosgene, pyridine, 0 °C
then 4-fluorobenzypiperidine, 58%.
reaction with 4-bromo-but-2-enoic acid methyl ester followed by
palladium-mediated cyclization yielded the desired benzofuran
core. SeO2 oxidation of the b-carbon then gave the methyl oxalate.
Finally, hydrolysis with LiOH and coupling with dimethyl amine
gave compound 8.
The p38a activities for analogs 1, 2, and 4–9 are shown in Table
1. Compared to the initial 6-substituted indole leads, analogs with
different length of linkers such as phenyl (4) and 2H-chromene (5)
showed a significant decrease in p38a enzymatic activity, indicat-
ing the [6,5] fused ring systems maintained the preferred spacing
between the amide and oxalyl amide. Furthermore, the flipped
Scheme 3. Reagents and conditions: (a) EDCI, DMAP, 5-formyl-2-methoxy-benzoic
acid,22 Et3N, DMF, 86%; (b) TMSCN, n-BuLi; (c) concd HCl, heat; (d) KOH, MeOH, 75–
84% (three steps); (e) HCl, MeOH, reflux; (f) PCC, DCM; (g) NaOH, MeOH–H2O; (h)
EDCI, dimethyl amine, 30–50% (four steps); (i) EDCI, DMAP, 5-formyl-2-trifluoro-
methyl-2H-chromene-3-carboxylic acid,23 Et3N, DMF, 81%.
2-methyl-1H-indole analog (7) still had reasonably good p38
a
enzyme activity, when compared to the initial indole analogs
(1 and 2).
In our subsequent [6,5] fused ring systems, it is clearly shown
that both polarity and electronics play important roles in modulat-
ing enzymatic and, in particular, cellular activity. Indoles and
azaindoles are clearly preferred systems than indazole and benzofu-
ran in both enzyme and cellular assays. Additionally, 6-methoxy
substitutions result in better cellular activities than 6-chloro substi-
tutions. In the case of 7-azaindole analog 9, the cellular activity is al-
most the same as its enzymatic activity. An anomaly in the SAR is the
the desired methyl 4-hydroxy-2-methoxybenzoate was prepared
by sequential protection of two hydroxyl groups through benzyla-
tion and methylation. Following deprotection of the benzyl group
and ring iodination, the ester was hydrolyzed and the resulting
acid was coupled with 4-fluorobenzyl piperidine. Subsequent

1062
B. J. Mavunkel et al. / Bioorg. Med. Chem. Lett. 20 (2010) 1059–1062
7. Lichtenstein, G. R.; Panaccione, R.; Mallarkey, G. Ther. Adv. Gastroenterol. 2008,
1, 43.
8. Furst, D. E. Clin. Ther. 2004, 26, 1960.
9. Lee, J. C.; Laydon, J. T.; McDonnell, P. C.; Gallagher, T. F.; Kumar, S.; Green, D.;
McNutly, D.; Blumenthal, M. J.; Heyes, J. R.; Landvatter, S. W.; Strickler, J. E.;
McLaughlin, M. M.; Siemens, I. R.; Fisher, S. M.; Livi, G. P.; White, J. R.; Adams, J.
L.; Young, P. R. Nature 1994, 372, 739.
10. Foster, M. L.; Halley, F.; Souness, J. E. Drug News Perspect. 2000, 13, 488.
11. Kumar, S.; Boehm, J.; Lee, J. C. Nat. Rev. Drug Disc. 2003, 2, 717.
12. Saklatvala, J. Curr. Opin. Pharmacol. 2004, 4, 372.
13. Dominguez, C.; Powers, D. A.; Tamayo, N. Curr. Opin. Drug Discov. Devel. 2005, 8,
421.
14. Lee, M.; Dominguez, C. Curr. Med. Chem. 2005, 12, 2979.
15. Mavunkel, B. J.; Chakravarty, S.; Perumattam, J. J.; Luedtke, G. R.; Lu, Q.; Dugar,
S.; Laney, M.; Henson, M.; Protter, A.; Xu, Y.-J.; Schreiner, G. F.; Almirez, R.;
Scott, B.; Lewicki, J.; Moore, A.; Lee, S.; Brahn, E.; Liu, D. Y. International
Symposium on Advances in Synthetic, Combinatorial and Medicinal Chemistry,
Moscow, Russia, 2004.
16. Docking studies were conducted using a high-resolution p38
a X-ray structure
(PDB ID: 1ove). The extra-precision mode of Glide, a ligand–receptor docking
tool (Schrodinger, LLC), was used to generate a poses for piperidine indole
analog 1. Poses were further filtered by visual inspection.
17. Davies, S. P.; Reddy, H.; Caivano, M.; Cohen, P. Biochem. J. 2000, 351, 95.
18. Wang, Z.; Canagarajah, B. J.; Boehm, J. C.; Kassisa, S.; Cobb, M. H.; Young, P. R.;
Abdel-Meguid, S.; Adams, J. L.; Goldsmith, E. J. Structure 1998, 6, 1117.
19. Lisnock, J.; Tebben, A.; Frantz, B.; O’Neill, E. A.; Croft, G.; O’Keefe, S. J.; Li, B.;
Hacker, C.; De Laszlo, S.; Smith, A.; Libby, B.; Liverton, N.; Hermes, J.; LoGrasso,
P. Biochemistry 1998, 37, 16573.
20. Mavunkel, B. J.; Chakravarty, S.; Perumattam, J. J.; Dugar, S.; Lu, Q.; Liang, X.
WO 2000/071535, 2000.
21. Mavunkel, B. J.; Perumattam, J. J.; Lu, Q.; Dugar, S.; Goyal, B.; Wang, D. X.;
Chakravarty, S.; Luedtke, G. R.; Nashashibi, I.; Tester, R.; Tan, X. US Patent
Application 2005/288299, 2005.
22. Nomura, M.; Takahashi, Y.; Tanase, T.; Miyachi, H.; Tsunoda, M.; Ide, T.;
Murakami, K. WO 2000/ 075103, 2000.
23. Carter, J. S.; Obukowicz, M. G.; Devadas, B.; Talley, J. J.; Brown, D. L.; Graneto, M.
J.; Bertenshaw, S. R.; Rogier, D. J., Jr.; Nagarajan, S. R.; Hanau, C. E.; Hartmann, S.
J.; Ludwig, C. L.; Metz, S. WO 98/47890, 1998.
24. Büchi, G.; Lee, G. C. M.; Yang, D.; Tannenbaum, S. R. J. Am. Chem. Soc. 1986, 108,
4115.
Scheme 6. Reagents and conditions: (a) K2CO3, Bu4NCl, KI, benzyl bromide,
acetone, 81%; (b) NaH, MeI, 95%; (c) Pd/C, H2 (1 atm.), MeOH, 84%; (d) BTMAICl2,
Na2CO3, 65%; (e) KOH, MeOH–H2O, 89%; (f) 3, TBTU, Et3N, DMF, 93%; (g) 4-bromo-
but-2-enoic acid methyl ester, K2CO3, acetone, 69%; (h) Pd(OAc)2, Na2CO3, HCOONa,
BnEt3NCl, DMF, 81%; (i) SeO2, dioxane, 77%; (j) LiOH, THF–H2O, 100%; (k) EDCI, Et3N,
dimethyl amine, 87%.
25. Procedure for human p38
a kinase assay: Compounds were assayed using an
6-methoxy-benzofuran analog 8, which exhibits a 60-fold reduction
in vitro method measuring the incorporation of radiolabeled ATP into a peptide
substrate. Compounds were dissolved in DMSO and diluted into water to the
desired concentrations. Compounds were mixed with the enzyme reagent, and
in potency in the p38
In summary, we designed and synthesized a number of 4-fluor-
obenzylpiperidine based heterocyclic oxalyl amides as potent p38
a enzyme assay compared to indole 2.
the reactions were initiated by the addition of
containing 200 M biotin–peptide substrate and 0.6 mM ATP (+100
32P-ATP). Final assay conditions were 25 mM MOPS, pH 7.0, 26.25 mM b-
glycerol phosphate, 80 mM KCl, 22 mM MgCl2, 3 mM MgSO4, 1 mg/ml gelatin,
0.625 mM EGTA, 1 mM DTT, 50 M peptide substrate, 150 M ATP, and 5 nM
p38 . After 60 min incubation at 30 °C, the reactions were stopped by the
addition of 10 l per reaction of 1.5% phosphoric acid. A portion of each of the
a
4Â substrate cocktail
a
l
l
Ci/ml
MAP kinase inhibitors. The optimal linker length between the
piperidine amide and oxalyl amide as well as the significance of
electronic properties of different heterocycles were studied in both
p38a enzymatic and cellular based assays and confirm the impor-
tance of the indole core.
c
-
l
l
a
l
reactions was transferred into the well of a streptavidin-coated Flash Plate
(Perkin Elmer). The plates were washed 3Â in PBS containing 0.01% Tween, and
sealed. Counts incorporated were determined on
a scintillation counter.
References and notes
Relative enzyme activity at each compound concentration was calculated by
subtracting background counts (counts measured in the absence of enzyme)
from each result, and comparing the resulting counts to those obtained in the
absence of inhibitor. The IC50 was determined to be the concentration of
compound which reduced the incorporation of ATP into the substrate by 50%,
when compared with control reactions containing no inhibitor.
1. Smolen, J. S.; Steiner, G. Nat. Rev. Drug Disc. 2003, 2, 473.
2. Smith, R. J. Drug Discovery Today 2005, 10, 1598.
3. Cominelli, F. N. Eng. J. Med. 2004, 351, 2045.
4. Sticherling, M. Drug Discovery Today 2005, 2, 275.
5. Jarvis, B.; Faulds, D. Drugs 1999, 57, 945.
26. The LPS/TNF
a Human Whole Blood Assay was run as described in: Mavunkel,
B.; Dugar, S.; Luedtke, G.; Tan, X.; McEnroe, G. US Patent 6,696,443, 2004.
6. Choy, E. H.; Panayi, G. S. N. Eng. J. Med. 2001, 344, 907.