A Vascular Endothelial Growth Factor Receptor-2 Kinase Inhibitor Potentiates the Activity of the Conventional Chemotherapeutic Agents Paclitaxel and Doxorubicin in Tumor Xenograft Models Stuart Emanuel, Robert H. Gruninger, Angel Fuentes-Pesquera, Peter J. Connolly, Jennifer A. Seamon, Susan Hazel, Rose Tominovich, Beth Hollister, Cheryl Napier, Michael R. D’Andrea, Michael Reuman, Gilles Bignan, Robert Tuman, Dana Johnson, David Moffatt, Mark Batchelor, Anne Foley, James O’Connell, Rodger Allen, Martin Perry, Linda Jolliffe, and Steven A. Middleton Cancer Therapeutics Research (S.E., R.H.G., A. F.-P., P.J.C., J.A.S., M.R., G.B., L.J., S.A.M.) and Oncology Research (R.To., M.R.D., R.Tu., D.J.], Johnson & Johnson Pharmaceutical Research and Development, Raritan, New Jersey; Institute of Medical and Veterinary Science, Adelaide, Australia (S.H.); Piedmont Research Center, Morrisville, North Carolina (B.H., C.N.); and Celltech Research, Limited, Slough, United Kingdom (D.M., M.B., A.F., J.O., R.A., M.P.) Received March 19, 2004; accepted June 9, 2004 ABSTRACT Inhibition of angiogenesis may have wide use in the treatment of cancer; however, this approach alone will not cause tumor regres- sion but may only slow the growth of solid tumors. The clinical potential of antiangiogenic agents may be increased by combining them with conventional chemotherapeutics. 4-[4-(1-Amino-1- methylethyl)phenyl]-2-[4-(2-morpholin-4-yl-ethyl)phenylamino]py- rimidine-5-carbonitrile (JNJ-17029259) represents a novel struc- tural class of 5-cyanopyrimidines that are orally available, selective, nanomolar inhibitors of the vascular endothelial growth factor receptor-2 (VEGF-R2) and other tyrosine kinases involved in angiogenesis, such as platelet-derived growth factor receptor, fibroblast growth factor receptor, VEGF-R1, and VEGF-R3, but have little activity on other kinase families. At nanomolar levels, JNJ-17029259 blocks VEGF-stimulated mitogen-activated pro- tein kinase signaling, proliferation/migration, and VEGF-R2 phos- phorylation in human endothelial cells; inhibits the formation of vascular sprouting in the rat aortic ring model of angiogenesis; and interferes with the development of new veins and arteries in the chorioallantoic membrane assay. At higher concentrations of 1 to 3 M, this compound shows antiproliferative activity on cells that may contribute to its antitumor effects. JNJ-17029259 delays the growth of a wide range of human tumor xenografts in nude mice when administered orally as single-agent therapy. Histological examination revealed that the tumors have evidence of reduced vascularity after treatment. In addition, JNJ-17029259 enhances the effects of the conventional chemotherapeutic drugs doxoru- bicin and paclitaxel in xenograft models when administered orally in combination therapy. An orally available angiogenesis inhibitor that can be used in conjunction with standard chemotherapeutic agents to augment their activity may have therapeutic benefit in stopping the progression of cancer and preventing metastasis. The vascular endothelial growth factor receptor-2 (VEGF- R2) mediates the process of angiogenesis by inducing the survival, proliferation, and migration of endothelial cells (Millauer et al., 1993). Angiogenesis is prominent during embryonic development, but in a normal adult, it occurs only as a result of injury or during endometrial turnover. How- ever, several pathological conditions initiate the growth of new blood vessels, including diabetic retinopathy, age-re- lated macular degeneration, endometriosis, rheumatoid ar- Article, publication date, and citation information can be found at http://molpharm.aspetjournals.org. doi:10.1124/mol.104.000638. ABBREVIATIONS: VEGF-R2, vascular endothelial growth factor receptor-2; PDGF-R, platelet-derived growth factor receptor-; EGF-R, epidermal growth factor receptor; FGF-R, fibroblast growth factor receptor; HASMC, human aortic smooth muscle cell; HUVEC, human umbilical vein vascular endothelial cells; PLC-, phospholipase C-; CDK, cyclin-dependent kinase; HER2, human epidermal growth factor receptor-2; FGF-R2, fibroblast growth factor receptor-2; CAM, chorioallantoic membrane; JNJ-17029259, 4-[4-(1-amino-1-methylethyl)phenyl]-2-[4-(2- morpholin-4-yl-ethyl)phenylamino]pyrimidine-5-carbonitrile; MAP, mitogen-activated protein; DMSO, dimethyl sulfoxide; PBS, phosphate-buff- ered saline; ERK, extracellular signal-regulated kinase; ANOVA, analysis of variance; SU5416, semaxanib; E64, trans-epoxysuccinyl-L-leu- cylamino(4-guanidino)butane; ZD6474, [N-(4-bromo-2-fluorophenyl)-6-methoxy-7-[(1-methylpiperidin-4-yl)methoxy]quinazolin-4-amine]; PTK787, (1-[4-chloroanilino]-4-[4-pyridylmethyl] phthalazine succinate). 0026-895X/04/6603-635–647$20.00 MOLECULAR PHARMACOLOGY Vol. 66, No. 3 Copyright © 2004 The American Society for Pharmacology and Experimental Therapeutics 638/1171386 Mol Pharmacol 66:635–647, 2004 Printed in U.S.A. 635 at ASPET Journals on June 20, 2016 molpharm.aspetjournals.org Downloaded from