Chromatin signalling pathways and FANCE amplification affect ATR inhibitor sensitivity in metastatic breast cancer

Metastatic breast cancer remains incurable, as many patients develop therapy resistance. Loss of ATM/p53 increases reliance on the ATR pathway, positioning ATR inhibitors (ATRi) as promising therapeutics. However, targeting a single pathway often leads to resistance due to tumour heterogeneity or alternative signalling mechanisms. Here, we combine chromatin-enrichment proteomics and phospho-proteomics with genome sequencing data in the ATRi-sensitive triple negative breast cancer (TNBC) cell line MDA-MB-453 to map adaptive responses to the ATR inhibitor AZD6738. We identify chromatin-associated activation of survival pathways, including AKT1, RPTOR (mTORC1), CDK4/5, and OGFR, alongside hyperphosphorylation of MKI67, SAFB2, and CHD4, indicating ATRi sensitivity. Complementary siRNA screening of DNA damage response (DDR) genes reveals that amplification of Fanconi anaemia (FA) pathway gene FANCE increases sensitivity to AZD6738. Analysis of breast cancer datasets highlights frequent FANCE amplification in metastatic patients, particularly in circulating tumour cells. Strikingly, pharmacological inhibition of the FA pathway (UBE2T/FANCL-IN-1) synergises with AZD6738. Together, our findings define adaptive resistance mechanisms to ATR inhibition and nominate FA pathway blockade as a rational combination strategy. Overall, our work provides fundamental insight into the complexity of DDR in metastatic breast cancer and offers a platform for mechanistic investigation, which can be exploited in cancer therapy.