Shamsutdinova, D. | Search bioRxiv

bioRxiv2026

Spatially-Resolved Multiomic Atlas of Leiomyosarcoma Identifies Two Clinically Relevant Epigenetically-Driven Cell States

Leiomyosarcoma is a smooth muscle-derived malignancy marked by significant clinical heterogeneity. The extent and nature of cellular heterogeneity and molecular underpinnings remain poorly understood. To address this at transcriptomic and epigenomic levels, we performed single-nucleus multiome sequencing on untreated primary leiomyosarcoma tissues. Malignant cells segregated almost exclusively into two previously unrecognized and epigenetically distinct states: a dedifferentiated, mesenchymal-like subtype (MES) and a differentiated smooth muscle-enriched subtype (SMC). Chromatin accessibility profiling revealed strong enrichment of nuclear factor I (NFI) transcription factor motifs in MES cells, whereas AP-1 family motifs--most prominently FOSL2--were selectively accessible in SMC cells. Established leiomyosarcoma cell lines faithfully recapitulated these subtypes, and targeted depletion of NFI or AP-1 factors suppressed proliferation, invasion, and in vivo tumor growth, demonstrating functional dependency on these transcriptional programs. Spatial transcriptomics across 328 tissue cores from 128 leiomyosarcomas showed that immunosuppressive macrophages preferentially cluster around MES regions, revealing a subtype-specific tumor-immune niche. Clinically, MES-dominant tumors were associated with significantly worse patient outcomes. Through an epigenetic inhibitor screen, we identify and validate SMARCA4/2 inhibition as a promising therapeutic vulnerability for MES leiomyosarcomas. Together, this work defines two epigenetically driven, transcription factor-regulated, and clinically relevant states of leiomyosarcoma, revealing mechanistic underpinnings of tumor heterogeneity and uncovering actionable therapeutic strategies. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=93 HEIGHT=200 SRC="FIGDIR/small/726988v1_ufig1.gif" ALT="Figure 1"> View larger version (36K): org.highwire.dtl.DTLVardef@132928eorg.highwire.dtl.DTLVardef@133e3b7org.highwire.dtl.DTLVardef@1ab4398org.highwire.dtl.DTLVardef@e2de25_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2025

EWS::WT1 Isoform-Dependent Regulation of Neogenes in Desmoplastic Small Round Cell Tumors

Desmoplastic small round cell tumor (DSRCT) is a rare, aggressive sarcoma characterized by the pathognomonic EWS::WT1 fusion protein (FP), an oncogenic chimeric transcription factor (OCTF) resulting from the t(11;22)(p13;q12) translocation. Recent studies have identified "neogenes" (NGs), genes normally silent in normal tissues but transcriptionally activated by OCTFs, as potential tumor-specific markers in fusion-driven cancers. In this study, we investigated the expression and regulation of DSRCT-specific NGs (DSRCT_NGs) using multimodal data across different cohorts of patients, PDX, and cell line data. We evaluated bulk and single-nucleus RNA sequencing of patient specimens from MD Anderson Cancer Center, revealing the robust ability for DSRCT_NGs to distinguish FP-positive DSRCT from samples failing detection of the EWS::WT1 FP. To elucidate the regulatory role of the EWS::WT1 FP in driving NG expression, we performed knockdown experiments in four DSRCT cell lines. This consistently resulted in a reduction of DSRCT_NG expression. Isoform-specific expression of EWS::WT1 in LP9 and MeT-5A mesothelial cells revealed that the E-KTS isoform of EWS::WT1 predominantly drives DSRCT_NG expression. Mechanistically, ATAC-seq and ChIP-seq analyses demonstrated that EWS::WT1 directly binds to accessible chromatin regions near NG transcription start sites, enriched for WT1 motifs and active histone marks. Integration of Hi-ChIP data further revealed that EWS::WT1 facilitates long-range enhancer-promoter looping at DSRCT_NG loci, promoting the expression of nearby genes. Collectively, these findings establish DSRCT_NGs as direct transcriptional outputs of the EWS::WT1 FP and implicate their loci as regulatory regions of the DSRCT transcriptome. Their fusion-dependent expression, chromatin accessibility, and promoter-enhancer connectivity underscore their potential utility as highly specific biomarkers and therapeutic targets in DSRCT.

genomics↗