O1116: Differences in Metastatic Ability of Osteosarcoma Is Related to Extracellular Vesicles

Background: Osteosarcoma (OS) is a highly metastatic sarcoma affecting adolescents. Metastasis at diagnosis carries high mortality, and the most common site of metastasis is in the lungs. Even for patients without metastasis, recurrence is common and most often recurs at in the lungs rather than at the primary site. Extracellular Vesicles (EVs) have been linked to tumor growth and metastases in many different cancers. Understanding mechanisms of metastases can help better target metastatic disease and catch micrometastases at an earlier stage, thus, decreasing mortality for these patients. The aim of this study is to elucidate the impact of extracellular vesicles on osteosarcoma metastasis.

Methods: Two murine OS cell lines, K7M2, with high metastatic potential, and K12, with low metastatic potential were compared. EVs from these cell lines were collected via sequential ultracentrifugation and quantified using nano-tracking analysis. EVs were dyed with fluorescent dye and injected into mice. Mice were subsequently euthanized, and their organs were harvested for fluorescent measurement. EVs were additionally co-cultured with RAW 264.7 macrophages and cell markers analyzed via flow cytometry. Supernatants from treated cells were collected and analyzed for IL-10 levels by ELISA.

Results: EVs from highly metastatic, K7M2, OS cell lines are released in higher quantity than from lower metastatic, K12, cell lines (Figure 1A). EVs from the higher metastatic cell line traffic to the lungs at higher quantities compared to the lower metastatic cell line or controls (Figure 1B and C). EVs from these cell lines also differ functionally in their effect on macrophages in vitro, with higher change in polarization from M1 to M2, seen on flow cytometry (Figure 2A). IL-10 release, a marker of M2-like macrophages, was 7.7 times higher in M1 macrophages stimulated with K7M2 compared to M1 macrophages stimulated with K12, supporting M2 polarization from the EVs.

Conclusion: The highly metastatic OS cell line releases more EVs, traffics to the lungs in vivo and causes macrophages to polarize more toward M2. Greater expression of M2 phenotype corresponds with tumor associated macrophages. Elucidating the mechanism of EVs from OS may explain differences in pulmonary metastatic potential of OS cells and may guide future diagnostic or therapeutic tools in managing osteosarcoma.