LBA2 - Investigation of non-V600 BRAF mutations commonly found in NSCLC for their sensitivity to Dabrafenib or Trametinib

Abstract

ABSTRACT Background: The most common BRAF mutations in non-small cell lung cancer (NSCLC) are non-V600 in contrast to melanoma. BRAF pathway inhibitors have not been systematically investigated in non-V600 mutations in vitro and in the clinic. We tested the effect of two clinically available BRAF pathway inhibitors (Trametinib and Dabrafenib) on a subset of clinically identified BRAF mutations in a cohort of lung cancers enriched for adenocarcinoma in patients with no or limited smoking history. Methods: NSCLC tumor samples (FFPE) were tested for the presence of EGFR, KRAS, NRAS, HRAS and BRAF mutations by DGGE or NGS-based methods. We generated 15 BRAF expression plasmids, harboring the mutations found in the given cohort and others described in the literature. BRAF mutants were subjected to an in vitro kinase assay. BRAF constructs were also expressed in HEK293T cells (with and without wt-CRAF) to study their impact on ERK signaling and determine the effect of inhibitors. Results: Among 229 NSCLC patients, 12 patients (5.2%) were found to harbor a BRAF mutation in their tumor: V600 (25%), G469A (16.7%), G469V (8.3%), D594N (25%), D594E (8.3%), G596C (8.3%) and G466V (8.3%). Mutations were characterized as activating or kinase-impaired (in-vitro kinase assay). Kinase-impaired BRAF mutants could still activate the ERK pathway in a CRAF-dependent manner, more than wt-BRAF/wt-CRAF co-transfectant. A MEK inhibitor (Trametinib) and a selective BRAF-inhibitor (Dabrafenib) were tested at clinically relevant doses on HEK293T transfectants (either expressing BRAF mutant alone or together with CRAF). ERK signaling induced by activating mutations was reduced in response to both inhibitors separately. Trametinib inhibited the CRAF-dependent ERK signaling induced by impaired-kinase BRAF mutations. Dabrafenib activated the ERK pathway in cells expressing only CRAF as well as cells co-expressing a kinase-impaired BRAF mutation and CRAF. Conclusions: This study predicts sensitivity of activating non-V600 BRAF mutations in lung cancer to Trametinib or Dabrafenib. Targeting kinase impaired BRAF mutations which signal through CRAF using Dabrafenib will require the addition of Trametinib.