Abstract 5025: Single cell RNA sequencing reveals degenerative mechanisms in osteoblastic osteosarcoma and targetability of senescent subpopulations
Authors: Saul Dominik , Kosinsky Robyn Laura 
Published: June 2022

Osteosarcoma is the most common primary solid tumor of the bone, typically affecting children and adolescents. Surgery and adjuvant chemotherapy result in an unsatisfying 5-year survival rate of &lt;25%. Although immune adjuvant strategies are promising, recent developments targeting the specialized tumor microenvironment by aiming at osteoclasts and checkpoint inhibitors focus on the individual immune biology. We aimed to mirror the tumor microenvironment and pseudotemporal development of osteoblasts during their transformation to osteoblastic osteosarcoma cells. For this purpose, single cell RNA-sequencing (scRNA-seq) profiles of 166,012 osteoblastic osteosarcoma and 49,555 healthy bone cells isolated from 28 patients were analyzed. In pseudotemporal analyses, we identified developmental divergence points with distinct transcriptional patterns purporting paths of devolution or giving rise to proliferative arrest. We found the biologically predicted clinging conjunction between osteoclasts, osteoblasts and B cells as regulatory T cells. In addition, we discovered intratumoral cellular communication patterns with ligand-receptor interactions and secreted signaling analysis between certain pillars of heterogeneous subpopulations, illustrating the importance of the local immune-genomic landscape. Upon several experimentally verified pathways, the TGF-β interaction was paramount. A mathematical assembly of degenerated cell populations revealed canonical markers of an osteoblastic cancerous state, out of which the vast majority has been associated with tumorigenesis in earlier studies. Intriguingly, nonnegative matrix factorization gave rise to a spatially localized senescent cell state, potentially accessible to new senolytic compounds which are currently in clinical trials for aging-related diseases. This senescent state with proliferative arrest and the secretion of a senescence-associated secretory phenotype (SASP), has not been described in osteosarcoma so far. In addition, we identified novel pathways of intertumoral communication in these distinct clusters, which can be exploited therapeutically, and validated these with a single-cell guided pipeline to repurpose drugs and their combinations. Thus, we were able to define a temporal pattern for the development of osteoblastic osteosarcoma. A local microenvironment with gradual states of devolution, and distinct cell-cell communicational patterns was established, out of which TGF-β warrants further experimental as therapeutical exploitation. Finally, we curtailed degenerated and, surprisingly, senescent subpopulations, each of which were predicted to vary in therapeutic response, mirroring the clinical observed intratumoral heterogeneity and potentially guiding the way to an individualized treatment regimen.

Citation Format: Dominik Saul, Robyn Laura Kosinsky. Single cell RNA sequencing reveals degenerative mechanisms in osteoblastic osteosarcoma and targetability of senescent subpopulations [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5025.