Left-Right Asymmetry in the Breast TME is Associated with Distinct Bioelectric and Epigenetic Tumor Features

The tumor microenvironment (TME) plays a pivotal role in cancer progression. Here, we report a left-right (L-R) asymmetry in the breast TMEs. Bioinformatic analyses of RNASeq data and histological data from TCGA, revealed that R-sided tumors exhibit higher stromal content and are enriched in cancer-associated fibroblasts (CAFs). These differences were validated in paired L-R PDX and xenograft models, where R-tumors showed increased α-SMA and PDGFRα expression. Associated with laterality, we observed an asymmetry in tumor bioelectric properties: in both PDX and xenograft models, as well as in cells cultured with L-R conditioned media, L-sided tumors consistently displayed a depolarized membrane potential relative to their R counterparts. In-vitro, these features were shown to be epigenetically regulated - abolished by the DNA methyltransferase inhibitor decitabine - and reversible upon inversion of the conditioned media laterality. Methylome analyses in animal models revealed that L-tumors are hypermethylated at ion channel genes, particularly gap junction components, correlating with reduced expression. A computational model further demonstrated that bistable states of membrane potential and methylation can spontaneously emerge depending on initial conditions and gap junction coupling strength. Together, these results uncover L-R asymmetry as an overlooked layer of breast tumor heterogeneity which could serve as proof of principle for other tumors in bilateral organs and could have therapeutic implications based on their differential bioelectric and epigenetic profiles.