Neo-morphic missense mutant p53 proteins and the co-drivers promoting cell invasion

abstract: Phenotypic and molecular profiling demonstrates a high degree of heterogeneity in the breast tumors. TP53 tumor suppressor is mutated in 30% of all breast tumors and the mutation frequency in basal-like subtype is as high as 80% and co-exists with several other somatic mutations in different genes. It was hypothesized that tumor heterogeneity is a result of a combination of neo-morphic functions of specific TP53 driver mutations and distinct co-mutations or the co-drivers for each type of TP53 mutation. The 10 most common p53 missense mutant proteins found in breast cancer patients were ectopically expressed in normal-like mammary epithelial cells and phenotypes associated with various hallmarks of cancer examined. Supporting the hypothesis, a wide spectrum of phenotypic changes in cell survival, resistance to apoptosis and anoikis, cell migration, invasion and polarity was observed in the mutants compared to wildtype p53 expressing cells. The missense mutants R248W, R273C and Y220C were most aggressive. Integrated analysis of ChIP and RNA seq showed distinct promoter binding profiles of the p53 mutant proteins different than wildtype p53, implying altered transcriptional activity of mutant p53 proteins and the phenotypic heterogeneity of tumors. Enrichment and model-based pathway analyses revealed dysregulated adherens junction and focal adhesion pathways associated with the aggressive p53 mutants. As several somatic mutations co-appear with mutant TP53, we performed a functional assay to fish out the relevant collaborating driver mutations, the co-drivers. When PTEN was deleted by CRISPR-Cas9 in non-invasive p53-Y234C mutant cell, an increase in cell invasion was observed justifying the concept of co-drivers. A genome wide CRISPR library-based screen on p53-Y234C and R273C cells identified separate candidate co-driver mutations that promoted cell invasion. The top candidates included several mutated genes in breast cancer patients harboring TP53 mutations and were associated with cytoskeletal and apoptosis resistance pathways. Overall, the combined approach of molecular profiling and functional genomics screen highlighted distinct sets of co-driver mutations that can lead to heterogeneous phenotypes and promote aggressiveness in cells with different TP53 mutation background, which can guide development of novel targeted therapies.Dissertation/ThesisDoctoral Dissertation Biochemistry 201