(P 262) ESTABLISHMENT AND CHARACTERIZATION OF THE FIRST RADIATION-INDUCED, PATIENT-DERIVED ANGIOSARCOMA CELL LINE

Objective: Radiation-induced breast angiosarcoma (RIBAS) represents an unmet medical need, characterized by high recurrence rate, limited therapeutic options and dismal prognosis in advanced/metastatic stages. To the best of our knowledge, angiosarcoma preclinical models available worldwide are limited to five UV-induced cutaneous angiosarcoma cell lines, and one patient-derived xenograft model for radiation-induced angiosarcoma. Our objective was to develop a bona fide RIBAS cell line (RIAS001), describing its immunophenotypic features, tumorigenicity in mice, and RNA expression profile.

Methods: RIBAS cells were harvested from pleural effusion of a patient affected by metastatic RIBAS, cultured (named as RIAS001 cell line), and implanted subcutaneously into the right flank of 7-week old NOD-SCID IL2R γnull mice (cell-line derived xenograft; CDX). Total RNA was extracted from cell lines and snap-frozen tumor tissues; total RNA Sequencing was performed by Illumina TruSeq Stranded Total RNA gold Kit and NovaSeq6000 System. RIAS001 cells were characterized by immunohistochemistry, immunofluorescence, transcriptomic profile.

Results: We successfully isolated RIBAS cells from the pleural effusion of a 63-year old woman affected by secondary, radiation-induced angiosarcoma of the breast. We cultured cells in vitro, establishing doubling time and optimizing cell culture conditions with different culture media and growth factors.
In detail, we tested M199 (HUVEC cell culture medium), DMEM KnockOut (optimal for primary patient-derived cell lines), and DMEM High Glucose, which proved as the most suited for RIAS001 growth. From the cultured cells (RIAS001), a xenograft mouse model (2 generations) was established, and its morphological and immunohistochemical features were compared to the patient tumor sample as shown in Figure 1. Both the cell line and the CDX retained immunophenotypic angiosarcoma features, as shown by immunofluorescence at confocal microscopy (Figure 2) and immunohistochemistry (Figure 1). The RNA expression profile of RIAS001 and RIAS001-CDX was analyzed in comparison to HUVEC cells, and to other cell lines and CDX derived from non-angiosarcoma STS. The analysis confirmed high expression of CD31, ERG, and MYC in RIAS001/RIAS001-CDX.

Conclusion: We established the first bona fide RIBAS cell line described so far (RIAS001). We have shown tumorigenicity in mice, and retained morphological and immunophenotypic features of RIAS001/RIAS001-CDX compared to the patient’s tumor samples. Therefore, RIAS001 represents a valuable and versatile preclinical radiation-induced angiosarcoma model, which could foster future angiosarcoma research.