Antitumor Efficacy of the Novel RAF Inhibitor GDC-0879 Is Predicted by BRAF Mutational Status and Sustained Extracellular Signal-Regulated Kinase/Mitogen-Activated Protein Kinase Pathway Suppression

Oncogenic activation of the BRAF serine/threonine kinase has been associated with initiation and maintenance of melanoma tumors. As such, development of pharmacologic agents to target RAF proteins or their effector kinases is an area of intense investigation. Here we report the biological properties of GDC-0879, a highly selective, potent, and orally bioavailable RAF small-molecule inhibitor. We used extracellular signal-regulated kinase (ERK)-1/2 and mitogen-activated protein kinase/ERK kinase (MEK)-1/2 phosphorylation as biomarkers to explore the relationship between tumor outcome and pharmacodynamic inhibition of the RAF-MEKERK pathway. In GDC-0879–treated mice, both cell line– and patient-derived BRAF tumors exhibited stronger and more sustained pharmacodynamic inhibition (>90% for 8 hours) and improved survival compared with mutant KRAS–expressing tumors. Despite the involvement of activated RAF signaling in RAS-induced tumorigenesis, decreased time to progression was observed for some KRAS-mutant tumors following GDC-0879 administration. Moreover, striking differences were noted for RAF and MEK inhibition across a panel of 130 tumor cell lines. Whereas GDC-0879– mediated efficacy was associated strictly with BRAF status, MEK inhibition also attenuated proliferation and tumor growth of cell lines expressing wild-type BRAF (81% KRAS mutant, 38% KRAS wild type). The responsiveness of BRAF melanoma cells to GDC-0879 could be dramatically altered by pharmacologic and genetic modulation of phosphatidylinositol 3-kinase pathway activity. These data suggest that GDC-0879–induced signaling changes are dependent on the point of oncogenic activation within the RAS network. Taken together, these studies increase our understanding of the molecular determinants for antitumor efficacy resulting from RAF pathway inhibition and have implications for therapeutic intervention in the clinic. [Cancer Res 2009;69(7):OF1–10] Introduction RAF protein kinases have been implicated in cellular responses relevant to tumorigenesis, including cell proliferation, invasion, survival, and angiogenesis (1, 2). The RAF family is composed of three members, ARAF, BRAF, and RAF1, which play a pivotal role in transducing signals from RAS to downstream kinases, mitogenactivated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK) kinase (MEK)-1/2 and ERK-1/2. Activating somatic mutations in BRAF are frequently observed in several tumor types, including malignant melanoma (3) and colorectal carcinoma (4). Almost 90% of these BRAF mutations are a T1799A transversion in exon 15 that results in a ValGlu (V600E) amino acid substitution in the activation segment of the kinase. This mutation obviates the requirement for the Thr599 and Ser602 phosphorylation that occurs during normal BRAF activation and leads to constitutive kinase activity that is 500-fold greater than that of wild-type protein (5, 6). In melanoma, BRAF mutation has been reported in premalignant atypical or dysplastic nevi and may thereby implicate BRAF activation as an initiating event in tumorigenesis (7). Tumor maintenance of melanoma cell lines in vivo is also highly dependent on BRAF (8, 9). Experiments with BRAF-specific inducible short hairpin RNA (shRNA) show that BRAF silencing in an established tumor inhibits further tumor progression, and in some tumor xenograft models the loss of BRAF resulted in complete tumor regression (9). This effect was caused by loss of proliferation, increased apoptosis, and macrophage infiltration. Taken together, BRAF represents an excellent target for anticancer therapy based on epidemiology, preclinical target validation, and drugability of protein kinases. There are currently numerous efforts to develop therapeutic agents to target BRAF or its downstream kinases (10). The first RAF inhibitor to enter clinical development was the multikinase inhibitor Nexavar (sorafenib tosylate), which is now approved for the treatment of patients with advanced renal cell carcinoma and unresectable hepatocellular carcinoma. However, the broad selectivity profile of this inhibitor makes it unsuitable for proofof-concept evaluation of RAF kinase inhibition in tumors (11). Recently, more selective RAF inhibitors have been disclosed, and chemical inhibition of BRAF enzymatic activity using these compounds can result in diminished proliferation and survival of tumor cells in vitro (12, 13). However, only very limited in vivo data have been published for these inhibitors. For instance, daily administration of SB-590885 resulted in a slight delay of tumor growth for one BRAF-mutant melanoma xenograft model (12). This effect was consistent with the modest reduction (50–80%) in Note: Supplementary data for this article are available at Cancer Research Online (http://cancerres.aacrjournals.org/). Requests for reprints: Marcia Belvin, Genentech, Inc., 1 DNA Way, South San Francisco, CA 94080. Phone: 650-467-7346; Fax: 650-225-1411; E-mail: [email protected] or Lori Friedman, Phone: 650-467-1926; E-mail: [email protected]. I2009 American Association for Cancer Research. doi:10.1158/0008-5472.CAN-08-3563 www.aacrjournals.org OF1 Cancer Res 2009; 69: (7). April 1, 2009 Research Article Published Online First on March 10, 2009 as 10.1158/0008-5472.CAN-08-3563 Research. on April 19, 2017. © 2009 American Association for Cancer cancerres.aacrjournals.org Downloaded from Published OnlineFirst March 10, 2009; DOI: 10.1158/0008-5472.CAN-08-3563