Actionable fusions and rearrangements can be efficiently identified by Hi-C whole genome sequencing in lung tumors

Introduction: Molecular testing in lung and other solid tumors has led to the identification of driver mutations that can be effectively targeted by new therapeutics. In addition to single point mutations that activate signaling proteins such as EGFR and KRAS, the presence of various gene fusions that lead to activation of key oncogenic drivers have become prominent drug targets. Detection of these fusions by RNA sequencing is the current gold standard but evidence suggests that fusions are still missed for various reasons. This study aims to improve detection rate of targetable fusions and rearrangements in solid tumors using a new method called Hi-C sequencing. Methods: Hi-C sequencing is a novel whole genome DNA-sequencing assay for detection of structural variation based on unique Hi-C chemistry which leverages sequencing of linked pairs of reads which occur nearby one another in 3-dimensional and linear space, from FFPE samples. Linking reads amplifies the rearrangement signal by providing many more read pairs spanning the breakpoint and also overcoming masked fusions resulting from breakpoints in non-unique sequences. Results: In a set of 139 NSCLC samples, Hi-C sequencing demonstrated 100% concordance with FISH and/or RNA sequencing results in cases with known fusions (19/19, including 11 ALK , 2 MET , 2 ROS , 2 MET , 1 NTRK , 1 NRG1 and 1 RET fusions). The next cohort of 97 samples were previously determined by standard DNA and RNA sequencing to be negative for drivers such as EGFR and KRAS mutations and fusions of ALK, MET, NTRK, RET, and ROS . In this cohort, we have detected biomarkers related to drug sensitivity in 16 cases. These include targetable fusions such as NTRK2 (1), NRG1 (1), PRKCA (1), and ERBB2 ( 1 ). Furthermore, we detected loss of function variants indicating sensitivity to checkpoint inhibitors (2 cases), or PARP inhibitors (6). In addition, noncanonical fusions were detected in NRG1 (1), and ALK (1), both retaining their functional domains and potentially indicating sensitivity to inhibitors. Conclusions: Encouraging results from this study suggest that Hi-C sequencing may be a valuable tool in the molecular classification of NSCLC and other solid tumors and could lead to improvement in patient care. Hi-C can detect gene fusions and rearrangements that are known to drive cancer and may be used for therapy selection, including in cases which were negative by standard genetic testing.