Dual PI3K/mTOR inhibition is required to combat resistance to CDK4/6 inhibitor and endocrine therapy in <i>PIK3CA</i> -mutant breast cancer

Combined CDK4/6 inhibitor (CDK4/6i) and endocrine therapy (ET) improves outcomes in advanced estrogen receptor-positive (ER⁺) breast cancer, but emergence of resistance to this combination underscores the pressing need for alternative therapeutic strategies. A promising approach involves adding an inhibitor of the PI3K/AKT/mTOR pathway to the standard combined CDK4/6i and ET, but selecting the most effective inhibitors and their optimal combinations has proven to be challenging. Here, we compared the efficacy of various triple combinations using single- or dual-point PI3K/AKT/mTOR pathway inhibitors in breast cancer cell lines, cell line xenografts, patient-derived xenografts, and organoids resistant to CDK4/6i and ET and exhibiting <i>PIK3CA</i>, <i>PTEN</i>, or <i>AKT1</i> mutations. <i>PIK3CA</i>-mutant, <i>PTEN</i>-wild type, CDK4/6i-resistant, and ET-resistant models required the addition of the dual PI3K/mTOR inhibitor gedatolisib to effectively impede tumor growth by blocking the HIF-1α pathway through both mTORC1 inhibition and PI3K/AKT-mediated modulation of GSK3α/β activity. Conversely, <i>PIK3CA</i>-wild type, <i>PTEN</i>-null cells benefited from triple combinations incorporating either the AKT inhibitor capivasertib or the dual mTORC1/2 inhibitor sapanisertib to block tumor growth. In addition, gedatolisib reduced viability of <i>PIK3CA</i>- or <i>AKT1</i>-mutant and <i>PTEN</i>-wild type CDK4/6i-resistant patient-derived organoids compared with the α-specific PI3K inhibitor alpelisib. Our data support the higher efficacy of gedatolisib over alpelisib in ER⁺ breast tumors harboring alterations of the PI3K/AKT/mTOR pathway including <i>PIK3CA</i> or <i>AKT1</i> mutations.