EWSR1::ATF1 hyperactivation via TRIM11 depletion triggers lethal metabolic rewiring in clear cell sarcoma

Clear cell sarcoma (CCS) is an ultra-rare and aggressive soft tissue sarcoma driven by the EWSR1::ATF1 fusion oncoprotein. Currently, no effective and well-tolerated treatment options are available to CCS patients. Direct inhibition of fusion transcription factors is precluded by the absence of defined ligand-binding pockets, whereas their abundance is under the control of the ubiquitin-proteasome system (UPS), offering an alternative avenue for biology-guided treatment. Here we show the abundance of EWSR1::ATF1 is tightly regulated by the UPS, and that release of this constraint is lethal to CCS cells. A reporter-based CRISPR/Cas9 knockout screen of UPS components identified TRIM11 as an E3 ligase that regulates turnover of EWSR1::ATF1, which it recognises as a neosubstrate without altering wild-type EWSR1 or ATF1. Depletion of TRIM11 increased nuclear fusion protein abundance, amplified the fusion-driven transcriptional programs and reduced viability. Proteomic analysis identified the cataplerotic enzyme phosphoenolpyruvate carboxykinase 1 (PCK1), a direct EWSR1::ATF1 target, as the key mediator of lethal effects of fusion hyperactivation. Extracellular flux analysis, isotope tracing and quantitative metabolomics revealed excessive cataplerotic carbon withdrawal and impaired oxidative phosphorylation as the underlying mechanism. In patient tumors, high PCK1 expression associated with metastatic transcriptome programs and disease presentation, as well as shorter survival. Taken together, we report that hyperactivation of EWSR1::ATF1 opens a metabolic vulnerability, establishing TRIM11 as an actionable dependency and PCK1 as a candidate prognostic biomarker in CCS.

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Publication Details

Journal
bioRxiv (Cold Spring Harbor Laboratory)
Published
2026-11-10
DOI
https://doi.org/10.64898/2026.11.06.764260
Primary Topic
Sarcoma Diagnosis and Treatment
Type
preprint
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preprint

EWSR1::ATF1 hyperactivation via TRIM11 depletion triggers lethal metabolic rewiring in clear cell sarcoma

Nicola Biondi, Francisco Yanqui-Rivera, Nischalan Pillay, Martin Schneider et al.
bioRxiv (Cold Spring Harbor Laboratory)
Sarcoma Diagnosis and Treatment
preprint

EWSR1::ATF1 hyperactivation via TRIM11 depletion triggers lethal metabolic rewiring in clear cell sarcoma

Nicola Biondi, Francisco Yanqui-Rivera, Nischalan Pillay, Martin Schneider, Sandra Strauß, Julien Picotto, Gunhild Mechtersheimer, Sebastian Bauer, Shiva Ahmadi, Michelle Neßling, Mikolaj Michal Slabicki, Akanksha Farswan, Claudia Scholl, Kevin B. Jones, Anushka Deshpande, Adarsh Raveendran, Dario L. Frey, Mohamad Saoud, Ashraf Yusuf Rangrez, Laura Mayerhofer, Priya Chudasama, Michael Buettner, Dawid Krzeciesa, Karsten Richter, Liang Chen, Stefan Froehling, Susanne Grünewald, Christopher Davies, Daniel Hughes, Christiane Optiz, Kwang Seok Lee, Kamini Kaushal, Wenxin Chao, Kilian Bales, Sumathi Vaiyapuri
preprint en

Abstract

Clear cell sarcoma (CCS) is an ultra-rare and aggressive soft tissue sarcoma driven by the EWSR1::ATF1 fusion oncoprotein. Currently, no effective and well-tolerated treatment options are available to CCS patients. Direct inhibition of fusion transcription factors is precluded by the absence of defined ligand-binding pockets, whereas their abundance is under the control of the ubiquitin-proteasome system (UPS), offering an alternative avenue for biology-guided treatment. Here we show the abundance of EWSR1::ATF1 is tightly regulated by the UPS, and that release of this constraint is lethal to CCS cells. A reporter-based CRISPR/Cas9 knockout screen of UPS components identified TRIM11 as an E3 ligase that regulates turnover of EWSR1::ATF1, which it recognises as a neosubstrate without altering wild-type EWSR1 or ATF1. Depletion of TRIM11 increased nuclear fusion protein abundance, amplified the fusion-driven transcriptional programs and reduced viability. Proteomic analysis identified the cataplerotic enzyme phosphoenolpyruvate carboxykinase 1 (PCK1), a direct EWSR1::ATF1 target, as the key mediator of lethal effects of fusion hyperactivation. Extracellular flux analysis, isotope tracing and quantitative metabolomics revealed excessive cataplerotic carbon withdrawal and impaired oxidative phosphorylation as the underlying mechanism. In patient tumors, high PCK1 expression associated with metastatic transcriptome programs and disease presentation, as well as shorter survival. Taken together, we report that hyperactivation of EWSR1::ATF1 opens a metabolic vulnerability, establishing TRIM11 as an actionable dependency and PCK1 as a candidate prognostic biomarker in CCS.

bioRxiv (Cold Spring Harbor Laboratory)
University College London Hospitals NHS Foundation Trust (GB), German Cancer Research Center (DE), Heidelberg University (DE), Huntsman Cancer Institute (US), Royal Orthopaedic Hospital (GB), University Hospital Heidelberg (DE), Massachusetts General Hospital (US), National Center for Tumor Diseases (DE), Royal National Orthopaedic Hospital NHS Trust (GB), Essen University Hospital (DE), German Centre for Cardiovascular Research (DE), University College London (GB), University of Duisburg-Essen (DE)
Sarcoma Diagnosis and Treatment
DOISourceOpenAlex
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