Tracing the evolutionary trajectory of CDK4/6 inhibitor resistance in oestrogen receptor positive breast cancer

Understanding tumour evolution is key to overcoming resistance to CDK4/6 inhibitors (CDK4/6i) in oestrogen receptor positive breast cancer. While genetic changes contribute, many patients lack identifiable mutations, suggestive of non-genetic epigenetic reprogramming and transcriptional plasticity. Here, we integrate expressed barcode lineage tracing, single-cell and bulk profiling, together with mathematical modelling, to study the evolution of resistance to the clinically approved CDK4/6i; Abemaciclib and Palbociclib. Our findings show that the evolutionary path to resistance to CDK4/6i is dependent on both the specific drug and the cancer cell model and can be driven by heritable epigenetic changes, independent of other acquired genetic driver alterations. We identify epigenetically driven transcriptional cell states that pre-exist as minority sub-populations in parental cells that become enriched in CDK4/6i resistant cells and are characterised by increased enhancer accessibility despite resistant cells showing overall reduced global accessibility. These transcriptional states show open chromatin in parental cells, suggestive of the presence of pre-existing epigenetically ‘primed’ cells. Finally, we generate and validate a core set of genes predictive of neoadjuvant endocrine + CDK4/6i response. Functional genomic assessment of candidate genes from this signature highlights potential collateral vulnerabilities in resistant cells. Overall, our study highlights the functional role of epigenetic adaptation in driving CDK4/6i resistance.