Abstract 2975: Synthetic lethality in synovial sarcoma: SS18-SSX fusions and DNA damage response (DDR) inhibitors
Authors: Fleuren Emmy DG , Jones Sam , Frankum Jessica , Williamson Chris , Menon Malini , Shipley Janet , Ashworth Alan , Graaf Winette van der , Lord Christopher J. 
Published: August 2018

Aim: Synovial sarcoma (SS) is an aggressive soft-tissue malignancy predominantly affecting the young and characterised by pathognomonic SS18-SSX fusions. Treatment options are limited and therapeutic targeting of SS18-SSX oncoproteins has not yet been achieved. We carried out large-scale, multiplatform functional genomic screenings followed by extensive in vitro and in vivo validation studies to identify therapeutically actionable dependencies in SS with the highest translational potential.

Methods: High-throughput drug sensitivity and siRNA screens including 79 clinically relevant compounds (0.5-1000nM) and siRNAs targeting 1600 different genes, respectively, were carried out in SS cell lines (SYO-1, HS-SY-II, Aska-SS, Yamato-SS and CME-1) and &gt;115 cancer cell lines of other histologies in triplicate in 5-day survival assays. SS drug and siRNA sensitivity data was compared to sensitivity profiles of non-SS tumour cell lines (Mann-Whitney test). Validation inhibitory and mechanistic assays were performed with selected drugs and siRNA in SS cells, non-SS cells ectopically expressing SS18-SSX fusion proteins, and relevant other controls. Presence of replication fork stress biomarkers was assessed by (fluorescent) immunohistochemistry and DNA fibre analysis.

Results: Both siRNA and drug assays identified a series of genes involved the DNA damage response (DDR) to be candidate genetic dependencies in SS. siRNA screen hits included ATR, the ATR-activating proteins RAD9A and RAD18, and the two tumour suppressor genes BRCA1 and BRCA2. The drug screen identified a dependency on two clinical PARP inhibitors (PARPi); talazoparib and rucaparib. ATR and PARP sensitivity in SS cells was comparable to that of tumour cells harbouring known molecular defects associated with sensitivity. A series of validation assays using clinical ATR (ATRi) and PARP inhibitors (PARPi) confirmed a synthetic lethal effect in SS tumour cell lines in vitro, which was confirmed for ATRi in patient-derived xenografts in vivo. Ectopic expression of SS18-SSX fusion proteins in non-SS cell lines elicited ATRi and PARPi sensitivity, which was mediated by a reduction in SS18 and SMARCB1 protein levels. ATRi sensitivity in SS was characterised by an increase in biomarkers of replication fork stress (increased yH2AX, decreased replication fork speed and increased R-loops), an apoptotic response and was found to be dependent upon Cyclin E expression. Finally, we found that combinations with cisplatin or PARP inhibitors enhanced the anti-tumour cell effect of ATRi, suggesting that either single agent ATRi or combination therapy involving ATRi might be further assessed as a candidate approach for SS treatment.

Conclusions: Our integrated analysis identified an unexpected dependency of SS tumour cells on DDR mediators. Importantly, sensitivity was synthetic lethal with SS18-SSX oncoproteins and is clinically actionable.

Citation Format: Emmy DG Fleuren, Sam Jones, Jessica Frankum, Chris Williamson, Malini Menon, Janet Shipley, Alan Ashworth, Winette van der Graaf, Christopher J. Lord. Synthetic lethality in synovial sarcoma: SS18-SSX fusions and DNA damage response (DDR) inhibitors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2975.