Development of Allosteric Small Molecule APOBEC3B Inhibitors from In Silico Screening.

APOBEC3B cytosine deaminase contributes to the mutational burdens of tumors, resulting in tumor progression and therapy resistance. Small molecule APOBEC3B inhibitors have potential to slow or mitigate these detrimental outcomes. Through molecular dynamics (MD) simulations and computational solvent mapping analysis, we identified a novel putative allosteric pocket on the C-terminal domain of APOBEC3B (A3Bctd), and virtually screened the ChemBridge Diversity Set (N~110,000) against both the active and potential allosteric sites. Selected high-scoring compounds were subsequently purchased, characterized for purity and composition, and tested in biochemical assays, which yielded 13 hit compounds. Orthogonal NMR assays verified binding to the target protein. Initial selectivity studies suggest these compounds preferentially target A3Bctd over related deaminase APOBEC3A (A3A), and MD simulations indicate this selectivity may be due to the steric repulsion from H56 that is unique to A3A. Taken together, our studies represent the first virtual screening effort against A3Bctd that has yielded candidate inhibitors suitable for further development.

Mutational signature-based identification of DNA repair deficient gastroesophageal adenocarcinomas for therapeutic targeting.

Homologous recombination (HR) and nucleotide excision repair (NER) are the two most frequently disabled DNA repair pathways in cancer. HR-deficient breast, ovarian, pancreatic and prostate cancers respond well to platinum chemotherapy and PARP inhibitors. However, the frequency of HR deficiency in gastric and esophageal adenocarcinoma (GEA) still lacks diagnostic and functional validation. Using whole exome and genome sequencing data, we found that a significant subset of GEA, but very few colorectal adenocarcinomas, show evidence of HR deficiency by mutational signature analysis (HRD score). High HRD gastric cancer cell lines demonstrated functional HR deficiency by RAD51 foci assay and increased sensitivity to platinum chemotherapy and PARP inhibitors. Of clinical relevance, analysis of three different GEA patient cohorts demonstrated that platinum treated HR deficient cancers had better outcomes. A gastric cancer cell line with strong sensitivity to cisplatin showed HR proficiency but exhibited NER deficiency by two photoproduct repair assays. Single-cell RNA-sequencing revealed that, in addition to inducing apoptosis, cisplatin treatment triggered ferroptosis in a NER-deficient gastric cancer, validated by intracellular GSH assay. Overall, our study provides preclinical evidence that a subset of GEAs harbor genomic features of HR and NER deficiency and may therefore benefit from platinum chemotherapy and PARP inhibitors.