Abstract

PurposeNovel covalent inhibitors of KRASG12C have shown limited response rates in KRASG12C mutant (MT) colorectal cancer (CRC) patients. Thus, novel KRASG12C inhibitor combination strategies that can achieve deep and durable responses are needed. Experimental designSmall molecule KRASG12C inhibitors AZ1569 and AZ8037 were employed. To identify novel candidate combination strategies for AZ1569, we performed RNA sequencing, siRNA and high-throughput drug screening. Top hits were validated in a panel of KRASG12CMT CRC cells and in vivo. AZ1569-resistant CRC cells were generated and characterised. ResultsResponse to AZ1569 was heterogeneous across the KRASG12CMT models. AZ1569 was ineffective at inducing apoptosis when used as single-agent or combined with chemotherapy or agents targeting the EGFR/KRAS/AKT axis. Using a systems biology approach, we identified the anti-apoptotic BH3-family member BCL2L1/Bcl-xL as a top hit mediating resistance to AZ1569. Further analyses identified acute increases in the pro-apoptotic protein BIM following AZ1569 treatment. ABT-263 (Navitoclax), a pharmacological Bcl-2 family-inhibitor that blocks the ability of Bcl-xL to bind and inhibit BIM, led to dramatic and universal apoptosis when combined with AZ1569. Furthermore, this combination also resulted in dramatically attenuated tumour growth in KRASG12CMT xenografts. Finally, AZ1569-resistant cells showed amplification of KRASG12C, EphA2/c-MET activation, increased pro-inflammatory chemokine profile and cross-resistance to several targeted agents. Importantly, KRAS amplification and AZ1569-resistance were reversible upon drug withdrawal, arguing strongly for the use of drug holidays in the case of KRAS amplification. ConclusionsCombinatorial targeting of Bcl-xL and KRASG12C is highly effective, suggesting a novel therapeutic strategy for KRAS G12CMT CRC patients.