Small Molecule Therapeutics BGB-283, a Novel RAF Kinase and EGFR Inhibitor, Displays Potent Antitumor Activity in BRAF-Mutated Colorectal Cancers Zhiyu Tang 1 , Xi Yuan 2 , Rong Du 2 , Shing-Hu Cheung 2 , Guoliang Zhang 3 , Jing Wei 2 , Yuan Zhao 2 , Yingcai Feng 4 , Hao Peng 4 , Yi Zhang 4 , Yunguang Du 4 , Xiaoxia Hu 1 , Wenfeng Gong 1 ,Yong Liu 1 , Yajuan Gao 1 , Ye Liu 4 , Rui Hao 4 , Shengjian Li 4 , Shaohui Wang 3 , Jiafu Ji 5 , Lianhai Zhang 5 , Shuangxi Li 5 , David Sutton 1 , Min Wei 4 , Changyou Zhou 3 , Lai Wang 1 , and Lusong Luo 2 Abstract Oncogenic BRAF, which drives cell transformation and prolif- eration, has been detected in approximately 50% of human malignant melanomas and 5% to 15% of colorectal cancers. Despite the remarkable clinical activities achieved by vemurafenib and dabrafenib in treating BRAF V600E metastatic melanoma, their clinical efficacy in BRAF V600E colorectal cancer is far less impressive. Prior studies suggested that feedback activation of EGFR and MAPK signaling upon BRAF inhibition might contribute to the relative unresponsiveness of colorectal cancer to the first-generation BRAF inhibitors. Here, we report characterization of a dual RAF kinase/ EGFR inhibitor, BGB-283, which is currently under clinical inves- tigation. In vitro, BGB-283 potently inhibits BRAF V600E -activated ERK phosphorylation and cell proliferation. It demonstrates selec- tive cytotoxicity and preferentially inhibits proliferation of cancer cells harboring BRAF V600E and EGFR mutation/amplification. In BRAF V600E colorectal cancer cell lines, BGB-283 effectively inhibits the reactivation of EGFR and EGFR-mediated cell proliferation. In vivo, BGB-283 treatment leads to dose-dependent tumor growth inhibition accompanied by partial and complete tumor regressions in both cell line-derived and primary human colorectal tumor xenografts bearing BRAF V600E mutation. These findings support BGB-283 as a potent antitumor drug candidate with clinical potential for treating colorectal cancer harboring BRAF V600E muta- tion. Mol Cancer Ther; 14(10); 2187–97. Ó2015 AACR. Introduction The MAPK pathway plays an essential role in regulating cell proliferation and survival. Activation of the RAS–RAF–MEK–ERK kinase cascade by external stimuli transduces signals from the plasma membrane into the cell nucleus to control gene expression and determine cell fate (1). Aberrant activation of the MAPK signal transduction pathway is frequently found in different types of cancers, contributing to increased cell division, suppressed apoptosis, and enhanced cell motility and invasion (2, 3). BRAF, one of the three members of the RAF kinase family, has been identified as a target for cancer therapy (4, 5). A sequencing screen of 923 cancer samples detected mutations in the BRAF gene in approximately 50% of human malignant melanomas and 15% of colorectal cancers (6–11), with the V600E mutation accounting for at least 90% of oncogenic BRAF mutations (6). This V600E mutation introduces a negative charge in the BRAF kinase domain that mimics and bypasses the phosphorylation required for BRAF activation, which is normally achieved through growth factor– activated receptor tyrosine kinases. As a result, this "gain of function" of BRAF gives rise to a constitutive MAPK signaling that promotes tumor progression, in which BRAF V600E activates MEK1/2 in a RAS-independent manner (12–15). Small molecules that selectively target mutant BRAF exhibit good efficacy and yield impressive clinical responses in melano- ma patients with BRAF V600E mutation. The RAF inhibitors that selectively inhibit BRAF V600E tumors, vemurafenib (PLX4032) and dabrafenib (GSK 2118436), have generated objective response rates from 50% to 70%, respectively, in early clinical trials treating metastatic melanoma (5, 16, 17). The clinical experience with these BRAF inhibitors in benefiting melanoma patients confirms BRAF V600E as a bona fide oncogenic target and validates the utility of cancer therapies that target BRAF and MAPK signaling (4, 5). However, these first-generation BRAF inhibitors still have limitations, including development of cutaneous squa- mous cell carcinomas and treatment-related keratoacanthomas (KA) due to paradoxical activation of MAPK signaling. In addi- tion, they have inadequate clinical activity outside of melanoma 1 Department of In Vivo Pharmacology, BeiGene (Beijing) Co., Ltd., Beijing, P.R. China. 2 Department of Discovery Biology, BeiGene (Beijing) Co., Ltd., Beijing, P.R. China. 3 Department of Chemistry, BeiGene (Beijing) Co., Ltd., Beijing, P.R. China. 4 Department of Molec- ular Sciences, BeiGene (Beijing) Co., Ltd., Beijing, P.R. China. 5 Key Laboratory of Carcinogenesis and Translational Research (Ministry of Education), Department of Surgery, Peking University Cancer Hospital and Institute, Beijing, P.R. China. Note: Supplementary data for this article are available at Molecular Cancer Therapeutics Online (http://mct.aacrjournals.org/). Z. Tang and X. Yuan contributed equally to this article. Corresponding Author: Lusong Luo, BeiGene (Beijing) Co., Ltd., No. 30 Science Park Road, Zhong-Guan-Cun Life Science Park, Changping District, Beijing 102206, P.R. China. Phone: 86-10-5895-8201; Fax: 86-10-5895-8088; E-mail: lusong.luo@beigene.com doi: 10.1158/1535-7163.MCT-15-0262 Ó2015 American Association for Cancer Research. 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