SSX-mediated chromatin engagement and targeting of BAF complexes activates oncogenic transcription in synovial sarcoma.

Synovial sarcoma (SS) is a soft-tissue malignancy driven by a recurrent chromosomal translocation (t(X;18)) that uniformly produces the SS18-SSX oncogenic fusion protein. SS18 is a core subunit of the mammalian SWI/SNF (BAF) complexes, which remodel nucleosomes in an ATP-dependent manner and antagonistically oppose gene-silencing activity of polycomb-repressive complexes to maintain transcriptional control throughout development and differentiation. We previously discovered that in SS, incorporation of the oncogenic SS18-SSX fusion into BAF complexes leads to eviction of the tumor-suppressor BAF47 (INI1/SMARCB1) subunit, and aberrant activation of polycomb target genes by displacement of H3K27me3-mediated repression. However, uncoupling the oncogenic consequences of two co-occurrent BAF complex perturbations, gain of 78- amino acids of SSX to SS18 and loss of BAF47, has remained a challenge for the field. To identify effective targeted therapeutics for this patient population, it is critical that we understand the contribution of the gain- versus loss-of-function properties of these molecular events in this malignancy. Here we demonstrate that the SSX 78aa tail engages mononucleosomes and targeted, quantitative mass spectrometry proteomics reveals preferential engagement to nucleosomes decorated with histone modifications associated with transcriptional repression. Using biochemical affinity assays, we find that SSX dramatically increases the affinity of SS18-SSX-containing BAF complexes for chromatin, thereby decreasing the dynamic mobility of BAF complexes. Furthermore, we show that SS18-SSX-containing BAF complexes possess a broader genomic footprint and exhibit distinct chromatin localization in that expression of SS18-SSX drives a near complete retargeting of BAF complexes genome-wide. SS18-SSX directs BAF complexes to polycomb-repressed sites to activate embryonic development and neuronal gene pathways hallmark to SS primary tumors. This targeting by SSX results in a transcriptional signature markedly distinct from sarcomas such as malignant rhabdoid tumors, which are driven solely by biallelic loss of BAF47. Moreover, using CRISPR/Cas9-mediated KO of BAF47 in SS cell lines, we show that the proliferative arrest of SS cell lines upon suppression of SS18-SSX is independent of BAF47 reassembly into BAF complexes, thereby demonstrating that SSX targeting of BAF complexes drives oncogenesis in a manner distinct from BAF47 loss. Taken together, these studies uncover a novel functionality of the SSX tail that is required for SS oncogenesis, and inform the selection of appropriate targeted therapeutic agents for this gain-of-function BAF complex-driven cancer. This abstract is also being presented as Poster B25. Citation Format: Matthew J. McBride, John L. Pulice, Robert T. Nakayama, Nazar Mashtalir, Davis R. Ingram, Jacob D. Jaffe, Jack F. Shern, Javed Khan, Jason L. Hornick, Alexander J. Lazar, Cigall Kadoch. SSX-mediated chromatin engagement and targeting of BAF complexes activates oncogenic transcription in synovial sarcoma [abstract]. In: Proceedings of the AACR Conference on Advances in Sarcomas: From Basic Science to Clinical Translation; May 16-19, 2017; Philadelphia, PA. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(2_Suppl):Abstract nr PR11.