Intrinsic epigenetic state of primary osteosarcoma drives metastasis
Introduction
Osteosarcoma (OS) is a primary malignant bone tumor commonly affecting children, adolescents, and young adults. In the last 50 years, the survival of patients with non-metastatic OS has significantly improved from approximately 20% to greater than 70%. However, approximately 20-25% of patients with OS present with metastatic disease and the outcomes for patients with metastatic or relapsed OS has remained unchanged over the past 30-40 years, with overall survival rates of less than 30%. Consequently, a better understanding of the molecular mechanisms that support metastasis is essential to improve the clinical outcome of patients with OS.
Methods
To investigate the differences in the active chromatin of OS, we utilized patient-derived xenografts (PDX) derived from tumor biopsies of patients with OS. The PDX’s used in this study were grouped into the following categories based on disease presentation: localized - tumor obtained from the primary site in a patient without evidence of metastatic disease; primary met - tumor obtained from the primary site in patient with evidence of metastasis; local control - tumor obtained from primary site through surgical resection after neoadjuvant chemotherapy; and distal met - metastatic tumor obtained from a distal site (e.g., lung). We integrated active chromatin landscapes defined by histone H3 lysine acetylated chromatin (H3K27ac) and chromatin accessibility profiled by assay for transposase-accessible chromatin (ATAC) with gene expression across a cohort of localized (n = 3), primary met (n = 4), local control (n = 4) and distal met (n = 2) PDX’s.
Results
Primary OS tumors from patients with metastasis (primary met) have a distinct active chromatin landscape compared to primary tumors from patients without metastatic disease (localized). The difference in chromatin activity shapes the transcriptional profile of OS. We identified novel candidate genes involved in OS pathogenesis and metastasis, including PPP1R1B, PREX1 and IGF2BP1, which exhibit increased chromatin activity in primary met along with higher transcript levels. We further showed that loss of PREX1 in primary met OS cells significantly diminishes the proliferation, invasion, migration, and colony formation capacity. Overall, differential chromatin activity in primary met occurs in proximity of genes regulating actin cytoskeleton organization, cellular adhesion, and extracellular matrix suggestive of their role in facilitating OS metastasis. Furthermore, chromatin profiling of tumors from metastatic lung lesions noted increases in chromatin activity in genes involved in cell migration and key intracellular signaling cascades, including the Wnt pathway.
Conclusion
The successful treatment of metastatic disease remains an extremely difficult challenge for solid tumor patients, including osteosarcoma. The identification of key underlying molecular mechanisms responsible for predisposing and enabling the tumor cell to enhance its metastatic potential remains elusive. Through integration of RNA-Seq, ATAC-seq, and ChIP-Seq for H3K27Ac, we identified significant differential epigenomic landscapes for primary tumors obtained from patients with localized and metastatic OS, with noted differential H3K27Ac activity across the genome for those patients with upfront metastatic disease. Our identification of novel candidate metastatic-promoting genes that are present in primary metastatic bone tumors, such as PPP1R1B, PREX1 and IGF2BP1, provides the framework for subsequent functional genomic studies into the role of these genes in metastatic osteosarcoma. Thus, this data demonstrates that metastatic potential is intrinsically present in primary metastatic tumors and the cellular chromatin profiles further adapt to allow for successful dissemination, migration, and colonization at the distal metastatic site.