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RG7204 (PLX4032), a Selective BRAFV600E Inhibitor, Displays Potent Antitumor Activity in Preclinical Melanoma Models

The BRAF(V600E) mutation is common in several human cancers, especially melanoma. RG7204 (PLX4032) is a small-molecule inhibitor of BRAF(V600E) kinase activity that is in phase II and phase III clinical testing. Here, we report a preclinical characterization of the antitumor activity of RG7204 using established in vitro and in vivo models of malignant melanoma. RG7204 potently inhibited proliferation and mitogen-activated protein/extracellular signal-regulated kinase (ERK) kinase and ERK phosphorylation in a panel of tumor cell lines, including melanoma cell lines expressing BRAF(V600E) or other mutant BRAF proteins altered at codon 600. In contrast, RG7204 lacked activity in cell lines that express wild-type BRAF or non-V600 mutations. In several tumor xenograft models of BRAF(V600E)-expressing melanoma, we found that RG7204 treatment caused partial or complete tumor regressions and improved animal survival, in a dose-dependent manner. There was no toxicity observed in any dose group in any of the in vivo models tested. Our findings offer evidence of the potent antitumor activity of RG7204 against melanomas harboring the mutant BRAF(V600E) gene.

Abstract 2156: Preclinical combinations of vemurafenib, a selective BRAF inhibitor, with other targeted therapies in BRAFV600E colorectal cancer models

Mutations in BRAF at codon 600 promote catalytic activity and are associated with 8% of all human (solid) tumors, including about 10% of colorectal cancers (CRCs). Vemurafenib (RG7204, PLX4032) is a first-in-class, BRAF-specific small molecule inhibitor that dose-dependently inhibits tumor growth in BRAFV600E CRC xenografts. Despite these encouraging preclinical findings, unlike the remarkable responses observed in melanoma, single agent vemurafenib in a Phase I extension trial of 21 patients with previously treated metastatic CRC resulted in a modest 5% response rate. We therefore explored a range of combinations of vemurafenib with different novel targeted therapies including a MEK inhibitor, an AKT inhibitor, a PI3K inhibitor, an mTOR/PI3K dual inhibitor and an EGFR inhibitor in BRAFV600E CRC cell lines. The consequences of these combinations on proliferation were analyzed by MTT assay and the combined effect was determined by combination index (CI) calculated using CalcuSyn software. Western analysis and Annexin V staining were utilized to evaluate combination effects on downstream signaling events and apoptosis induction. Optimized doses of both vemurafenib and a MEK inhibitor were tested as single agents and in combination in CRC xenograft models in nude mice. Synergistic anti-proliferative effects were observed with combinations of vemurafenib and other targeted therapies in the BRAFV600E positive CRC cell lines tested. More effective signaling inhibition and apoptosis induction were observed with the combinations than either agent alone. Combining vemurafenib with a MEKi delivered greater anti-tumor activity and increased life span of animals in the LS411N CRC xenograft model. In a BRAF-mutant CRC xenograft model with inherent resistance to vemurafenib (RKO), tumor growth inhibition by vemurafenib was enhanced by combining with an AKT inhibitor. These in vitro and in vivo data suggest that the administration of vemurafenib in combination with novel targeted therapies may be effective in delivering enhanced and sustained clinical antitumor efficacy in colorectal cancers harboring the BRAFV600E mutation. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2156. doi:1538-7445.AM2012-2156

Abstract 2562: Efficacy of RG7204 (PLX4032), a selective BRAFV600E inhibitor, in combination with pegylated interferon alpha-2a (Pegasys), paclitaxel, carboplatin, and anti-VEGF MAb B20-4.1 in the LOX-IMVI human melanoma xenograft model

Few drugs are currently approved for the treatment of melanoma, including dacarbazine (chemotherapy) and interferon alpha (immunotherapy). However, only a small fraction of patients with metastatic melanoma respond to dacarbazine and a survival benefit has not been shown with this or any other chemotherapy regimen. Taxanes have demonstrated some activity in melanoma, including modest response rates in combination with carboplatin or bevacizumab. These results contrast with the extraordinary Phase I clinical data reported with the BRAFV600E kinase inhibitor RG7204 (PLX4032), which has demonstrated a remarkable response rate (N Engl J Med 363(9):809-819, 2010). Here we report our preclinical investigation of RG7204 in combination with the biologics interferon alpha-2a (Pegasys) or B20-4.1 (B20) (mouse/ human cross-reactive anti-VEGF MAb), and the chemotherapies paclitaxel and carboplatin in the BRAFV600E mutant LOX-IMVI melanoma model in nude mice.The combination of RG7204 and paclitaxel provided superior survival as compared to either monotherapy. Conversely, the combination of RG7204 plus carboplatin or RG7204 plus B20 did not extend survival as compared to RG7204 monotherapy. When RG7204, paclitaxel, and B20 were given in triplet combination, no improvement in survival was observed over any doublet. Similarly, the triplet combination with RG7204, carboplatin and B20 failed to demonstrate improved survival over any doublets or RG7204 monotherapy. Although complete suppression of tumor growth was readily achieved with either RG7204 or Pegasys monotherapy, the combination provided a remarkable survival benefit with median survival extended by nearly a year as compared to either monotherapy. Taken together, these data support clinical testing of RG7204 with interferon alpha in melanoma patients harboring BRAFV600E mutation in order to extend survival, and may allow for use of lower dose interferon. Clinical investigation of RG7204 and paclitaxel combination may also be warranted. While our data do not support combined therapy with RG7204 plus anti-VEGF therapy, it should be noted that the LOX model was resistant to B20 monotherapy, and therefore these findings might not apply to tumors with intrinsic sensitivity to anti-VEGF therapy. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2562. doi:10.1158/1538-7445.AM2011-2562