(P 234) GENOMIC LANDSCAPE OF CLEAR CELL SARCOMA

Objective: Clear Cell Sarcoma (CCS) represent a rare soft tissue sarcoma with an unusually aggressive phenotype (frequent in-transit, lymphatic and early distant metastases). Translocations resulting in chimeric genes fusing EWSR1 with either ATF1 or rarely CREB1 or CREM are the defining molecular events. As of today, no effective treatment is available and 5-year survival is 59%. Bioinformatics analyses and novel disease models are urgently needed to identify and validate therapeutic vulnerabilities.

Methods: Next generation sequencing (NGS), including whole genome, exome, and transcriptome data of 23 CCS patients and three CCS cell lines were collected. Somatic copy number alterations (SCNA) were found using CNVkit [https://cnvkit.readthedocs.io/en/stable/] and GISTIC2.0 [https://broadinstitute.github.io/gistic2/], and gene fusion detection was performed using Arriba [https://github.com/suhrig/arriba]. A new variant calling approach [https://github.com/snakemake-workflows/dna-seq-varlociraptor] was optimized on a training data set using a cohort of GIST (Gastrointestinal Stromal Tumor) samples with various common and uncommon, somatic and germline variants. Click-iT EdU Flow Cytometry Cell Proliferation Assay (Thermo Fisher) was used to evaluate palbociclib response in the cell lines.

Results: Variant calling revealed few variants in cancer associated genes. SCNA analyses revealed 9p21.3 loss in 12/23 patients, and smallest region of overlap (SRO) analyses implicated CDKN2A as the principal 9p21.3 deletion target, which was also found in two out of three CCS cell lines. Arriba revealed no recurrent or apparent oncogenic gene fusions beyond EWSR1 fusions. The CDK4/6 inhibitor palbociclib had antiproliferative activity in cell lines with CDKN2A loss but not in the CDKN2A-positive subline.

Conclusion: These studies confirm EWSR1 fusion as the central genomic alteration in CCS. Recurrent loss of CDKN2A in about half of patients represents an additional genomic event. Future studies should focus on better understanding the oncogenic consequences of EWSR1 fusions, to identify therapeutic vulnerabilities.