Characterization of a novel sarcoma cell line with an EWSR1::POU2AF3 fusion.

Sarcomas with an EWSR1::POU2AF3(COLCA2) fusion are a very recently described entity of preferentially sinonasal origin and with undifferentiated round/spindle cell morphology. We established a novel cell line (PF1095) carrying a EWSR1::POU2AF3 fusion from the malignant pleural effusion of a 25-year-old sarcoma patient. The patient was first diagnosed with poorly differentiated neuroendocrine carcinoma based on tumor cell morphology and positivity to markers such as EMA, synaptophysin, and CD56. Later, the EWSR1 translocation was identified in the tumor cells with unknown partners and the patient received chemotherapy according to the Ewing 2008 protocol in combination with surgery and proton beam radiotherapy. At the time of cell line establishment, the disease progressed to pleural sarcomatosis with pleural effusion. In the cell line, we identified POU2AF3 as a fusion partner of EWSR1 and a TP53 frameshift deletion. Next, we determined the sensitivity of PF1095 cells to the currently approved chemotherapies in comparison to two conventional Ewing sarcoma lines (EW-7 and MHH-ES1) with the two most frequent EWSR::FLI1 fusions. Finally, we tested potential new combination therapies. We performed cell viability, proliferation, and cell cycle assays. We found that the proliferation rate of PF1095 cells was much slower than the EWSR1::FLI1 fusion lines and they also had a lower sensitivity to both irinotecan and doxorubicin treatment. Expression level of SLFN11, a predictor of sensitivity to DNA damaging agents, was also lower in PF1095 cells. Combination treatment with the PARP inhibitors olaparib and irinotecan or doxorubicin synergistically reduced cell viability and induced cell death and cell cycle arrest. This unique cell model provides an opportunity to test therapeutic approaches preclinically for this novel and aggressive sarcoma entity.