Ecological interactions in breast cancer: Cell facilitation promotes growth and survival under drug pressure

The interplay of positive and negative interactions between drug sensitive and resistant cells determines the efficacy of treatment in heterogeneous cancer cell populations. We studied isogenic estrogen receptor positive (ER+) breast cancer cell lineages sensitive and resistant to ribociclib-induced CDK4/6 inhibition in monoculture and coculture, finding that sensitive cells grow and compete more effectively in the absence of treatment. During treatment with ribociclib, sensitive cells survive and proliferate better when grown together with resistant cells than when grown in monoculture, termed facilitation in ecology. Several lines of evidence indicate estradiol production by resistant cells as the mechanism of facilitation, with both liquid chromatography-tandem mass spectrometry (LC-MS/MS) assays and single cell RNA-seq showing higher production by resistant cells, and sensitive cells shifting phenotype to become more like resistant cells when grown in coculture. Adding estradiol in monoculture provides sensitive cells with partial resistance, and cancels facilitation in coculture. Mathematical modeling quantifies the strength of competition and facilitation, and predicts that blocking facilitation has the potential to control both resistant and sensitive cell populations and inhibit the emergence of a refractory population. Overall design: Spheroids of different composition (100% sensitive, 50% sensitive – 50% resistant, 100% resistant) were initiated from Venus-labeled CAMA-1 and mCherry-labeled CAMA-1_ribociclib_resistant cells and were subjected to 1 uM ribociclib treatment. After 11 days, speroids were harvested, washed and cell suspensions were viably frozen for single cell RNA-seq analysis.