A Synthetic Chromatin Reader-Effector Activates Key Silenced Tumor Suppressor Genes in Triple Negative Breast Cancer Cells

Epigenetic control has arisen as a driver and druggable vulnerability in many hard-to-treat cancers. The development of epigenetic therapy is advancing as a way to reactivate tumor suppressor genes (TSGs) as an alternative to traditional chemotherapies. Triple negative breast cancer (TNBC) often shows relapse after chemotherapy and is untreatable with hormone therapy, but has shown modest responses to leading epigenetic drugs in vitro, in vivo, and in clinical trials. Currently, the design of epigenetic treatments for TNBC is limited to small molecule inhibitors that bind chromatin-modifying enzymes. Critical limitations such as passive gene regulation and off-target interactions outside of chromatin prevent epigenetic therapy from meeting its full potential. Our overall goal is to engineer synthetic reader-effectors to override aberrant gene expression profiles in cancer. In TNBC, CBX reader-repressor proteins are overexpressed and silence the expression of TSGs. In this study, we compare the anti-cancer effect of a synthetic reader-activator, Polycomb-based Transcription factor (PcTF), versus small molecule inhibitors. PcTF is designed to bind a biochemical signature in silenced chromatin (histone H3K27me3) and induce immediate activation of nearby genes. To support clinical translation, we identified a set of 99 TSGs that are silenced in TNBC patient samples and in model TNBC cell lines. In PcTF-expressing cells, we observed immediate activation of several of these TSGs accompanied by reduced viability in 12 - 24 hours, which suggests a faster anti-cancer effect than the epigenetic inhibitor compounds that typically require 3 - 7 days of treatment. In conclusion, we have established a strategy to compare the impact of chromatin engineering versus epigenetic drugs on gene expression and cancer cell viability in vitro. These results lead the way towards investigating cancer apoptosis in vitro, and inhibition of metastasis in a mouse model (currently underway).