Abstract LB-020: Characterization of a novel BRAF in-frame deletion reveals a distinct mutational activating mechanism for oncogenic kinases
Authors: Foster Scott , Ozen Aysegul , Whalen Dan , Song Kyung , Hatzivassiliou Georgia , Hymowitz Sarah , Skelton Nick , Malek Shiva 
Published: August 2015

Abstract
Kinase domain mutations are frequent drivers of many different types of cancer. While extensive studies of kinases such as BRAF and EGFR have provided insight into the mechanistic basis for many oncogenic point mutations such as the commonly occurring BRAFV600E and EGFRL858R mutations, the activation mechanism of short, in-frame deletions (such as the common EGFR exon19 deletion in non-small cell lung cancer) remains to be fully understood. In this work, we have discovered the activation mechanism for novel in-frame deletions in BRAF that have been reported in a small subset of thyroid cancers (TCGA Thyroid data set). These BRAF deletions structurally align with the EGFR exon19 deletion (most common being ΔELREA) and a similar deletion recently identified in HER2-positive breast tumor samples (ΔLRENT). These deletions target the β3-αC loop within the N-lobe of the kinase domain, which provides essential flexibility to the αC-Helix allowing the kinase to toggle between inactive and active conformations. Using molecular modeling studies, we demonstrate that all of these deletions genetically compromise this flexibility and constitutively “snap” the αC-Helix in the active conformation. Further characterization of the BRAF deletions shows that similar to BRAFV600E mutations, the β3-αC deletions are kinase-activating, CRAF-independent, and confer dimer-independent activity. Interestingly, cells with this deletion are sensitive to BRAF kinase inhibitors that bind to the active conformation (such as GDC-0879), but are innately resistant to the BRAF kinase inhibitor vemurafenib (which binds BRAF in the αC-Helix inactive, outward-shifted conformation). Similarly, modeling predicts all known β3-αC deletions would confer innate resistance to αC-Helix outward-shifting drugs such as vemurafenib or lapatinib. Further, we speculate that a β3-αC deletion could confer acquired resistance in vemurafenib-treated or lapatanib-treated patients. While many of the more common oncogenic point mutations have been suggested to function in part by destabilizing the inactive conformation and hence promoting the active conformation, it is becoming increasingly clear that oncogenic events that alter the αC-Helix region to directly promote the active conformation provide an alternative mechanism of kinase activation in cancer. Taken together our work underscores the importance of conformation specific kinase inhibitors to target mutationally activated kinases in cancer.
Citation Format: Scott Foster, Aysegul Ozen, Dan Whalen, Kyung Song, Georgia Hatzivassiliou, Sarah Hymowitz, Nick Skelton, Shiva Malek. Characterization of a novel BRAF in-frame deletion reveals a distinct mutational activating mechanism for oncogenic kinases. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr LB-020. doi:10.1158/1538-7445.AM2015-LB-020