Synovial Sarcoma Chromatin Dynamics Reveal a Continuum in SS18:SSX Reprograming.

Synovial sarcoma (SyS) is an aggressive soft-tissue malignancy characterized by a pathognomonic chromosomal translocation leading to the formation of the SS18::SSX fusion oncoprotein. SS18::SSX associates with mammalian BAF complexes suggesting deregulation of chromatin architecture as the oncogenic driver in this tumour type. To examine the epigenomic state of SyS we performed comprehensive multi-omics analysis on 52 primary pre-treatment human SyS tumours. Our analysis revealed a continuum of epigenomic states across the cohort at fusion target genes independent of rare somatic genetic lesions. We identify cell-of-origin signatures defined by enhancer states and reveal unexpected relationships between H2AK119Ub1 and active marks. The number of bivalent promoters, dually marked by the repressive H3K27me3 and activating H3K4me3 marks, has strong prognostic value and outperforms tumor grade in predicting patient outcome. Finally, we identify SyS defining epigenomic features including H3K4me3 expansion associated with striking promoter DNA hypomethylation in which SyS displays the lowest mean methylation level of any sarcoma subtype. We explore these distinctive features as potential vulnerabilities in SyS and identify H3K4me3 inhibition as a promising therapeutic strategy.

Targeting of SUMOylation leads to cBAF complex stabilization and disruption of the SS18::SSX transcriptome in Synovial Sarcoma.

Synovial Sarcoma (SS) is driven by the SS18::SSX fusion oncoprotein and is ultimately refractory to therapeutic approaches. SS18::SSX alters ATP-dependent chromatin remodeling BAF (mammalian SWI/SNF) complexes, leading to the degradation of canonical (cBAF) complex and amplified presence of an SS18::SSX-containing non-canonical BAF (ncBAF or GBAF) that drives an SS-specific transcription program and tumorigenesis. We demonstrate that SS18::SSX activates the SUMOylation program and SSs are sensitive to the small molecule SAE1/2 inhibitor, TAK-981. Mechanistically, TAK-981 de-SUMOylates the cBAF subunit SMARCE1, stabilizing and restoring cBAF on chromatin, shifting away from SS18::SSX-ncBAF-driven transcription, associated with DNA damage and cell death and resulting in tumor inhibition across both human and mouse SS tumor models. TAK-981 synergized with cytotoxic chemotherapy through increased DNA damage, leading to tumor regression. Targeting the SUMOylation pathway in SS restores cBAF complexes and blocks the SS18::SSX-ncBAF transcriptome, identifying a therapeutic vulnerability in SS, positioning the in-clinic TAK-981 to treat SS.

Targeting of SUMOylation leads to cBAF complex stabilization and disruption of the SS18::SSX transcriptome in Synovial Sarcoma.

Synovial Sarcoma (SS) is driven by the SS18::SSX fusion oncoprotein. and is ultimately refractory to therapeutic approaches. SS18::SSX alters ATP-dependent chromatin remodeling BAF (mammalian SWI/SNF) complexes, leading to the degradation of canonical (cBAF) complex and amplified presence of an SS18::SSX-containing non-canonical BAF (ncBAF or GBAF) that drives an SS-specific transcription program and tumorigenesis. We demonstrate that SS18::SSX activates the SUMOylation program and SSs are sensitive to the small molecule SAE1/2 inhibitor, TAK-981. Mechanistically, TAK-981 de-SUMOylates the cBAF subunit SMARCE1, stabilizing and restoring cBAF on chromatin, shifting away from SS18::SSX-ncBAF-driven transcription, associated with DNA damage and cell death and resulting in tumor inhibition across both human and mouse SS tumor models. TAK-981 synergized with cytotoxic chemotherapy through increased DNA damage, leading to tumor regression. Targeting the SUMOylation pathway in SS restores cBAF complexes and blocks the SS18::SSX-ncBAF transcriptome, identifying a therapeutic vulnerability in SS, positioning the in-clinic TAK-981 to treat SS.

Aberrant gene activation in synovial sarcoma relies on SSX specificity and increased PRC1.1 stability.

The SS18-SSX fusion drives oncogenic transformation in synovial sarcoma by bridging SS18, a member of the mSWI/SNF (BAF) complex, to Polycomb repressive complex 1 (PRC1) target genes. Here we show that the ability of SS18-SSX to occupy H2AK119ub1-rich regions is an intrinsic property of its SSX C terminus, which can be exploited by fusion to transcriptional regulators beyond SS18. Accordingly, SS18-SSX recruitment occurs in a manner that is independent of the core components and catalytic activity of BAF. Alternative SSX fusions are also recruited to H2AK119ub1-rich chromatin and reproduce the expression signatures of SS18-SSX by engaging with transcriptional activators. Variant Polycomb repressive complex 1.1 (PRC1.1) acts as the main depositor of H2AK119ub1 and is therefore required for SS18-SSX occupancy. Importantly, the SSX C terminus not only depends on H2AK119ub1 for localization, but also further increases it by promoting PRC1.1 complex stability. Consequently, high H2AK119ub1 levels are a feature of murine and human synovial sarcomas. These results uncover a critical role for SSX-C in mediating gene deregulation in synovial sarcoma by providing specificity to chromatin and further enabling oncofusion binding by enhancing PRC1.1 stability and H2AK119ub1 deposition.

Optimization of the Enrichment of Circulating Tumor Cells for Downstream Phenotypic Analysis in Patients with Non-Small Cell Lung Cancer Treated with Anti-PD-1 Immunotherapy.

The current study aimed at the optimization of circulating tumor cell (CTC) enrichment for downstream protein expression analyses in non-small cell lung cancer (NSCLC) to serve as a tool for the investigation of immune checkpoints in real time. Different enrichment approaches-ficoll density, erythrolysis, their combination with magnetic separation, ISET, and Parsortix-were compared in spiking experiments using the A549, H1975, and SKMES-1 NSCLC cell lines. The most efficient methods were tested in patients (n = 15) receiving immunotherapy targeting programmed cell death-1 (PD-1). Samples were immunofluorescently stained for a) cytokeratins (CK)/epithelial cell adhesion molecule (EpCAM)/leukocyte common antigen (CD45), and b) CK/programmed cell death ligand-1 (PD-L1)/ indoleamine-2,3-dioxygenase (IDO). Ficoll, ISET, and Parsortix presented the highest yields and compatibility with phenotypic analysis; however, at the patient level, they provided discordant CTC positivity (13%, 33%, and 60% of patients, respectively) and enriched for distinct CTC populations. IDO and PD-L1 were expressed in 44% and 33% and co-expressed in 19% of CTCs. CTC detection was associated with progressive disease (PD) (p = 0.006), reduced progression-free survival PFS (p = 0.007), and increased risk of relapse (hazard ratio; HR: 10.733; p = 0.026). IDO-positive CTCs were associated with shorter PFS (p = 0.039) and overall survival OS (p = 0.021) and increased risk of death (HR: 5.462; p = 0.039). The current study indicates that CTC analysis according to distinct immune checkpoints is feasible and may provide valuable biomarkers to monitor NSCLC patients treated with anti-PD-1 agents.

Evaluation of the detection of Toll-like receptors (TLRs) in cancer development and progression in patients with colorectal cancer.

BACKGROUND: Toll-like receptors (TLRs) play essential role in innate and acquired immunity, are expressed in various cell types, and are associated with altered susceptibility to many diseases, and cancers. The aim of this study was to investigate TLR2 (-196 to-174del), TLR4 (Asp299Gly and Thr399Ile) and TLR9 (T1237C and T1486C) gene polymorphisms at risk of colorectal cancer (CRC) development and progression. METHODS: Peripheral blood was obtained from 397 patients with adjuvant (stage II/III, n = 202) and metastatic (n = 195) CRC. Moreover, blood samples from 50 healthy volunteers and 40 patients with adenomatous polyps were also included as control groups. DNA from patients and controls was analyzed using PCR and PCR-RFLP for genotyping functional polymorphism within TLR2, TLR4 and TLR9 genotypes. RESULTS: TLR2-196 to-174del/del genotype was detected in 76.6% of the patients and was significantly higher that the controls groups (p<0.001). TLR4 Asp299Gly, TLR4 Thr399Ile, TLR9 T1237C and T1486C homozygous genotypes were detected in 70.5%, 70.5%, 61.5% and 61.5% of the patients respectively, and were also significantly higher than that in the control groups (p<0.001). All polymorphisms detected were also significantly associated with the metastatic disease (p<0.001) leading to shorter overall survival (p<0.001); whereas, TLR4 Asp299Gly and Thr399Ile polymorphisms were significantly associated with KRAS mutations. CONCLUSIONS: The detection of higher frequencies of the TLR2, TLR4 and/or TLR9 polymorphisms in CRC patients compared with the control groups highlight the role of these polymorphism in CRC development and cancer progression.