The early evolutionary landscape of osteosarcoma provides clues for targeted treatment strategies.

Osteosarcomas are aggressive primary tumors of bone that are typically detected in locally advanced stages, however which genetic mutations drive the cancer before its clinical detection remain unknown. To identify these events, we perform longitudinal genome-sequencing analysis of 12 patients with metastatic or refractory osteosarcoma. Phylogenetic and molecular clock analyses are carried out next to identify actionable mutations, and these are validated by integrating data from additional 153 osteosarcomas and pre-existing functional evidence from mouse PDX models. We found that the earliest and thus clinically most promising mutations affect the cell cycle G1 transition, which is guarded by cyclins D3, E1 and cyclin-dependent kinases 2, 4 and 6. Cell cycle G1 alterations originate no more than a year before the primary tumor is clinically detected and occur in >90 and 50% proportions of patients of the discovery and validation cohorts, respectively. In comparison, other cancer driver mutations could be acquired at any evolutionary stage and often do not become pervasive. Consequently, our data support that the repertoire of actionable mutations present in every osteosarcoma cell is largely limited to cell cycle G1 mutations. Since they occur in mutually exclusive combinations favoring either CDK2 or CDK4/6 pathway activation we propose a new genomically-based algorithm to direct patients to correct clinical trial options. This article is protected by copyright. All rights reserved.