Epigenetic analysis of osteosarcoma to define subclasses relevant to chemoresistance and metastasis

SUMMARY OS is a bone cancer seen mostly in children and young adults. It is among the most genomically complex tumors with aneuploidy, structural rearrangement and copy number being the hallmarks of the OS genome. Currently, treatment consists of highly toxic chemotherapy and there are no available targeted therapies. In addition, all patients with conventional OS are treated the same and there are no known upfront predictors of response or clinically relevant subtypes other than % necrosis in response to chemotherapy. Using epigenetics, we have uncovered a potential novel way to subclassify OS into subtypes driven by distinct transcription factor networks. These subtypes were initially identified using ATACseq and verified using other epigenetic marks (H3K27ac). In this application we propose to characterize the biological basis for these subtypes and identify subtype-specific vulnerabilities. We will also further evaluate initial data indicating that these subtypes have differential drug response. In Aim 1, we will define the core regulatory circuitry (CRC) that drives epigenetic heterogeneity in OS and define transcription factor vulnerabilities for OS subtypes using pooled focused CRISPRi screens in vitro and in vivo. In Aim 2, we will use multiomics and spatial transcriptomics to characterize intratumoral heterogeneity in OS. Finally, in Aim 3 we will determine the value of the epigenetic approach to intratumoral heterogeneity in developing new strategies for combination therapy in OS. Overall, our studies represent an innovative and potentially transformative approach to a disease that has seen no progress in therapy in 40 years.