ATR and PKMYT1inhibition re-sensitize a subset of TNBC patient-derived models to carboplatin and induce mitotic catastrophe

Triple negative breast cancer (TNBC) is associated with poor prognosis and is mainly treated with chemotherapy-based regimens, often including carboplatin. Resistance to carboplatin is a common clinical issue that is either initially present or develops with treatment. Overcoming this resistance is a significant clinical challenge, which highlights the need for novel therapeutic strategies. We used a pooled shRNA screening approach with a chemoresistant TNBC patient-derived xenograft (PDX) cell (PDXC) line to identify targets whose knockdown would enhance the efficacy of carboplatin. This screening led to the identification of the ATR (ataxia telangiectasia and Rad3-related) gene as a key therapeutic vulnerability. Inhibiting ATR with BAY1895344 or AZD6738 re-sensitized carboplatin-resistant PDXCs and PDXs to carboplatin, resulting in an increase in DNA damage, and apoptosis. ATR inhibition prevents carboplatin-resistant cells from effectively engaging the S and G2/M checkpoints required for DNA repair, leading to mitotic catastrophe. We further identified that the addition of ATR inhibitors to carboplatin enabled FOXM1-targeted gene program leading to premature passage into mitosis. Moreover, targeting PKMYT1, a regulator of cyclin-dependent kinase 1 (CDK1) controlling the G2/M checkpoint, through knockdown or with the novel PKMYT1 inhibitor RP-6306, also enhanced carboplatin efficacy in our TNBC PDXC. Molecular factors associated with response to the ATR inhibitor/carboplatin combination included low RNA levels of PKMYT1. These results underscore the pivotal roles of ATR and PKMYT1 in mediating resistance to carboplatin in TNBC and support targeting these pathways to overcome carboplatin resistance in this disease.