Osteosarcoma tumors maintain intra-tumoral transcriptional heterogeneity during bone and lung colonization

Abstract

Background Tumors are complex tissues containing collections of phenotypically diverse malignant and nonmalignant cells. We know little of the mechanisms that govern heterogeneity of tumor cells nor of the role heterogeneity plays in overcoming stresses, such as adaptation to different microenvironments. Osteosarcoma is an ideal model for studying these mechanisms—it exhibits widespread inter- and intra-tumoral heterogeneity, predictable patterns of metastasis, and a lack of clear targetable driver mutations. Understanding the processes that facilitate adaptation to primary and metastatic microenvironments could inform the development of therapeutic targeting strategies. Results We investigated single-cell RNA-sequencing profiles of 47,977 cells obtained from cell line and patient-derived xenograft models as cells adapted to growth within primary bone and metastatic lung environments. Tumor cells maintained phenotypic heterogeneity as they responded to the selective pressures imposed during bone and lung colonization. Heterogenous subsets of cells defined by distinct transcriptional profiles were maintained within bone- and lung-colonizing tumors, despite high-level selection. One prominent heterogenous feature involving glucose metabolism was clearly validated using immunofluorescence staining. Finally, using concurrent lineage tracing and single-cell transcriptomics, we found that lung colonization enriches for multiple clones with distinct transcriptional profiles that are preserved across cellular generations. Conclusions Response to environmental stressors occurs through complex and dynamic phenotypic adaptations. Heterogeneity is maintained, even in conditions that enforce clonal selection. These findings likely reflect the influences of developmental processes promoting diversification of tumor cell subpopulations, which are retained, even in the face of selective pressures.

Enhanced Biomimetics of Three-Dimensional Osteosarcoma Models: A Scoping Review

“…To grasp the complexity of the OS TME, exploring its heterogeneity is essential, and single-cell RNA sequencing can serve as a promising tool to unveil valuable insights into cellular diversity within OS TME [ 93 ]. This contributes to the exploration of cellular mechanisms, cancer evolution, and the detection of drug-resistant cancer cell population, facilitating the development of therapeutic targeting strategies [ 93 , 94 ]. Considering the intrinsic heterogeneity and diverse subtypes of osteosarcoma [ 95 ], patient-derived 3D OS models offer a potential biomimetic model, capturing the diversity of this tumour.…”

Immunohistochemical Study of Histone Protein 3 Modification in Pediatric Osteosarcoma Identifies Reduced H3K27me3 as a Marker of Poor Treatment Response

“…Another limitation is that our study used a TMA-based assay, which is not able to assess for possible intra-tumoral heterogeneity. Epigenetic regulation is known have a spatial heterogeneity in many tumors [34], and this could be magnified as a source of sampling error in an OS TMA with known epigenetic and histologic heterogeneity [35][36]. Given the findings in this study, future studies may benefit from both bulk and spatially resolved analytics, including transcriptomic and proteomic studies to determine if epigenetic regulation can be used in a prognostic or predictive fashion.…”

Children's Oncology Group's 2023 blueprint for research: Bone tumors

“…Data regarding whether the DNA damage leading to the complex osteosarcoma genome is an ongoing process after diagnosis 50 are emerging as are data regarding epigenetic and metabolic acclimations of metastasis and progression. 51,52 Immune therapies have been and are being actively investigated with single-agent immune-checkpoint inhibitors lacking a clinical signal alone. 53 A potentially promising approach is antibody-drug conjugates targeting membrane proteins selectively expressed on osteosarcoma cells.…”

“…Emerging approaches to target chromosomal instability through targeting kinases involved in DNA‐damage repair and exploring strategies related to BRCA1 functional deficiency such as PARP inhibitors 49 may have relevance in osteosarcoma given the presence of mutational signature 3 in 30% of cases. Data regarding whether the DNA damage leading to the complex osteosarcoma genome is an ongoing process after diagnosis 50 are emerging as are data regarding epigenetic and metabolic acclimations of metastasis and progression 51,52 . Immune therapies have been and are being actively investigated with single‐agent immune‐checkpoint inhibitors lacking a clinical signal alone 53 .…”