Mechanisms of APOBEC3A-induced cancer evolution and cancer vulnerability

PROJECT SUMMARY/ABSTRACT Cancer evolution driven by DNA-mutating APOBEC enzymes (apolipoprotein B mRNA editing enzyme, catalytic polypeptide-like), and the emergence of resistance to therapy, present a pressing clinical challenge. In particular, the APOBEC3A (A3A) enzyme is implicated in a range of cancers and is hypothesized to drive resistance to

targeted anticancer therapies. This project proposes to explore the mechanisms of A3A activity during targeted therapy, its substrate preferences, and potential methods of inhibiting its activity to slow cancer evolution and impede the emergence of drug resistance. Aim 1 focuses on investigating the role of A3A in the emergence of

drug resistance, hypothesizing that A3A expression is upregulated during targeted therapy, leading to APOBEC- signature mutations. We will test this using lung cancer cell-line models and targeted therapies, and will use CRISPR technology to genetically eliminate A3A and monitor the impact on the emergence of drug-resistant

cells. Aim 2 seeks to determine the genomic, transcriptomic, and proteomic targets of A3A and other APOBECs. Our previous work suggests that APOBEC mutagenesis and RNA editing are directed by local and mesoscale structural constraints. We will conduct computational analyses to identify hotspots of A3A activity in DNA and

RNA and their subsequent protein effects, refining our understanding of APOBEC mutagenesis and identifying super-optimal A3A substrates. Aim 3 proposes to evaluate the efficacy of hairpins and hairpin-like molecules as A3A inhibitors. We anticipate that A3A's affinity for hairpin substrates may render it susceptible to inhibition by

hairpins or hairpin-like molecules. Testing will be performed in vitro and in cells to assess whether hairpin molecules can inhibit endogenous cellular A3A activity and slow drug resistance evolution. Through a combination of computational and experimental approaches, this research will advance our understanding of

APOBEC mutagenesis, potentially offering novel diagnostics, research tools, and treatments for APOBEC-driven cancers.