Discovery and optimization of pyrazoline compounds as B-Raf inhibitors

Bioorganic and Medicinal Chemistry Letters

Volumn 20, Issue 16, 2010, Pages 4800-4804

  Duffey, Matthew O ^a   Adams, Ruth ^a   Blackburn, Christopher ^a   Chau, Ryan W ^a   Chen, Susan ^a   Galvin, Katherine M ^a   Garcia, Khristofer ^a   Gould, Alexandra E ^a   Greenspan, Paul D ^a   Harrison, Sean ^a   Huang, Shih Chung ^a   Kim, Mi Sook ^a   Kulkarni, Bheemashankar ^a   Langston, Steven ^a   Liu, Jane X ^a   Ma, Li Ting ^a   Menon, Saurabh ^a   Nagayoshi, Masayuki ^a   Rowland, R Scott ^a   Vos, Tricia J ^a   Xu, Tianlin ^a   Yang, Johnny J ^a   Yu, Shaoxia ^a   Zhang, Qin ^a   more..

^a MILLENNIUM PHARMACEUTICALS INC   (United States)

Author keywords

B Raf; B Raf inhibitor; Pyrazoline

Indexed keywords

B RAF KINASE; B RAF KINASE INHIBITOR; PYRAZOLINE DERIVATIVE; PROTEIN KINASE INHIBITOR; PYRAZOLE DERIVATIVE; RECOMBINANT PROTEIN;

ANIMAL EXPERIMENT; ARTICLE; CHEMICAL MODIFICATION; DRUG POTENCY; DRUG SELECTIVITY; DRUG SOLUBILITY; DRUG SYNTHESIS; ENZYME INHIBITION; HIGH THROUGHPUT SCREENING; NONHUMAN; RAT; STRUCTURE ACTIVITY RELATION; AMINO ACID SUBSTITUTION; ANTAGONISTS AND INHIBITORS; BINDING SITE; CHEMISTRY; COMPUTER SIMULATION; DRUG SCREENING; GENETICS; METABOLISM; SYNTHESIS;

AMINO ACID SUBSTITUTION; BINDING SITES; COMPUTER SIMULATION; DRUG EVALUATION, PRECLINICAL; PROTEIN KINASE INHIBITORS; PROTO-ONCOGENE PROTEINS B-RAF; PYRAZOLES; RECOMBINANT PROTEINS; STRUCTURE-ACTIVITY RELATIONSHIP;

EID: 77955423078     PISSN: 0960894X     EISSN: None     Source Type: Journal    
DOI: 10.1016/j.bmcl.2010.06.113     Document Type: Article

References (18)

1. M.J. Robinson, and M.H. Cobb Curr. Opin. Cell Biol. 9 1997 180
2. S.J. Heidorn, C. Milagre, S. Whittaker, A. Nourry, I. Niculesu-Davas, N. Dhomen, J. Hussain, J.S. Reis-Filho, C.J. Springer, C. Pritchard, and R. Marais Cell 140 2010 209
3. P.I. Poulikakos, C. Zhang, G. Bollag, K.M. Shokat, and N. Rosen Nature 464 2010 427
4. G. Hatzivassiliou, K. Song, I. Yen, B.J. Brandhuber, D.J. Anderson, R. Alvarado, M.J.C. Ludlam, D. Stokoe, S.L. Gloor, G. Vigers, T. Morales, I. Aliagas, B. Liu, S. Sideris, K.P. Hoeflich, B.S. Jaiswal, S. Seshagiri, H. Koeppen, M. Belvin, L.S. Friedman, and S. Malek Nature 464 2010 431
5. H. Davies, G.R. Bignell, C. Cox, P. Stephens, S. Edkins, S. Clegg, J. Teague, H. Woffendin, M.J. Garnett, W. Bottomley, N. Davis, E. Dicks, R. Ewing, Y. Floyd, K. Gray, S. Hall, R. Hawes, J. Hughes, V. Kosmidou, A. Menzies, C. Mould, A. Parker, C. Stevens, S. Watt, S. Hooper, R. Wilson, H. Jayatilake, B.A. Busterson, C. Cooper, J. Shipley, D. Hargrave, K. Pritchard-Jones, N. Maitland, G. Chenevix-Trench, G.J. Riggins, D.D. Bigner, G. Palmieri, A. Cossu, A. Flanagan, A. Nicholson, J.W.C. Ho, S.Y. Leung, S.T. Yuen, B.L. Weber, H.F. Seigler, T.L. Darrow, H. Paterson, R. Marais, C.J. Marshall, R. Wooster, M.R. Stratton, and P.A. Futreal Nature 417 2002 949
6. S.R. Hingorani, M.A. Jacobetz, G.P. Robertson, M. Herlyn, and D.A. Tuveson Cancer Res. 63 2003 5198
7. K. Flaherty, I. Puzanov, J. Sosman, K. Kim, A. Ribas, G. McArthur, R.J. Lee, J.F. Grippo, K. Nolop, and P. Chapman J. Clin. Oncol. 27 2009 15s
8. Raf activity was determined using a 384 well streptavidin coated flashplate. Reactions were performed with biotinylated peptide substrate at 30 °C for 3 h in a final volume of 30 μL. After addition of stop the reaction was incubated on a flashplate for 2 h. The flashplates were read on a Topcount counter.
9. The whole cell ELISA assay utilized a human melanoma cell line (A375) possessing the mutation BRaf V600E. A375 cells seeded overnight were incubated with Raf inhibitors for 3 h. At the end of the incubation, the cells were fixed, permeabilized, blocking buffer added and the plates were incubated overnight. After the blocking buffer was discarded, the plates were incubated with anti-phospho-ERK antibody for 1 h followed by treatment with anti-horse radish peroxidase. Optical density was read at 650 nm for tetramethylbenzidine as substrate.
10. C. Blackburn, M.O. Duffey, A.E. Gould, B. Kulkarni, J.X. Liu, S. Menon, M. Nagayoshi, T.J. Vos, and J. Williams Bioorg. Med. Chem. Lett. 20 2010 4795
11. P.T.C. Wan, M.J. Garnett, S.M. Roe, S. Lee, D. Niculescu-Duvaz, V.M. Good Cancer Genome Project C.M. Jones, C.J. Marshall, C.J. Springer, D. Barford, and R. Marais Cell 116 2004 855
12. We initially used a homology model because no DFG-in structures were available. When these structures became available, they were found to be nearly identical to the homology model. Initial attempts to dock 2 in the DFG-out conformation proved to be unsatisfactory.
13. A.C. Annigeri, and S. Siddappa Indian J. Chem. 1 1963 484
14. J. Lange, H. Coolen, H. van Stuivenberg, J. Dijksman, A. Herremans, E. Ronken, H. Keizer, K. Tipker, A. McCreary, W. Veerman, H. Wals, B. Stork, P. Verveer, A. den Hartog, N. de Jong, T. Adolfs, J. Hoogendoorn, and C. Kruse J. Med. Chem. 47 2004 627
15. 50: 14 μM). Due to ease of synthesis and time consuming methods required for HPLC separation we evaluated all compounds as racemic mixtures.
16. Solubility was determined by a turbidimetric solubility assay which provides the kinetic solubility of compounds in 50 mM potassium phosphate buffer at pH 6.8.
17. 50: 27 nM).
18. Pharmacokinetic experiments were performed by dosing either intravenously (0.5 mg/kg or 1 mg/kg) or orally (10 mg/kg) in male SD rats.