Engineering Transcription Factors that Sense Histone Modifications in Cancer Cells

Mounting evidence from genome-wide comparisons of chromosome packaging and gene expression in healthy versus cancer cells suggests that epigenetic hyper-repression, rather than genetic mutation in many cases, supports cancer aggressiveness. Aberrant behavior of the chromatin system (genomic DNA, and nuclear RNA and proteins) has been implicated as a driver of metastasis and drug resistance. Since the early 1990’s small compounds have been used to disrupt hyper-repressed chromatin to simultaneously activate sets of therapeutic genes in cancer cells. However, it is difficult to customize the biological activity of these small compound inhibitors, and they do not directly mediate RNA PolII activity at silenced tumor suppressor genes. To address these limitations, our lab has designed synthetic reader-actuator (SRA) fusion proteins that bind epigenetic marks within chromatin. The first SRA we have designed and tested in cancer cells, “Polycomb-based transcription factor” (PcTF), reads histone modifications through a protein-protein interaction between its N-terminal Polycomb chromodomain (PCD) motif and trimethylated lysine 27 of histone H3 (H3K27me3). The C-terminal VP64 domain of PcTF recruits endogenous activators to silenced targets. We identified a set of 104 genes that become consistently activated from 24 - 72 hours after PcTF-overexpression in a triple negative breast cancer cell line (BT-549). PcTF-positive BT-549 cells show reduced proliferation and viability in vitro. We propose that SRA-responsive genes, which includes anti-proliferation, immune response, and epithelial cell fate genes, are part of a ‘druggable’ anti-cancer gene module. These genes are rarely mutated in basal tumors and therefore may produce a therapeutic effect when they become activated. This talk will review our published results for BT-549, new data from additional TNBC lines, progress in engineering enhanced SRA activity, and analysis of 99 genes that are repressed in both cell lines and basal tumors to support clinical translation of our research.