A Stem-Like Tumor Cell Population Defines a Therapeutically Vulnerable Niche in SMARCB1-Deficient Sinonasal Carcinoma

SMARCB1-deficient sinonasal carcinoma (SDSC) is a rare, highly aggressive malignancy with dismal prognosis, limited therapeutic options and no established preclinical models. To address this gap, we combined single-nucleus RNA sequencing (snRNAseq), spatial transcriptomics, and ex vivo patient-derived tissue slice culture (TSC) to dissect intratumoral heterogeneity, microenvironmental niches, and treatment vulnerabilities in an index case of SDSC. snRNAseq revealed three transcriptionally distinct malignant subpopulations. Among these, we identified two specialized subsets: one marked by ALDH1A1, a gene associated with stemness, and another characterized by NTN4 expression. Spatial analysis mapped these populations to discrete tumor niches. Notably, the ALDH1A1+ compartment localized to a basal layer with intermingled TP63 expressing basal cells adjacent to stroma, showed reduced proliferative activity and displayed transcription factor activity consistent with a stem-like state. Ex vivo TSC drug testing revealed striking selective vulnerability: the mTOR inhibitor Sapanisertib induced extensive tumor necrosis and completely ablated both ALDH1A1+ and NTN4+ subpopulations. Gene set enrichment analysis provided mechanistic insight, confirming significant enrichment of PI3K-AKT-mTOR signaling specifically in ALDH1A1+ tumor cells. Additionally, Sapanisertib markedly reduced endothelial cells, consistent with anti-angiogenic effects. To validate the broader relevance of these findings, we analyzed an independent cohort of 12 SDSC, including recurrent tumors. ALDH1A1 expression was present in all cases across three spatial patterns, with particularly high levels in recurrences compared to primaries, supporting a recurrence-founder role. Furthermore, strong mesothelin expression was observed in both the index patient as well as a subset of the retrospective cohort, suggesting additional therapeutic opportunities with mesothelin-directed antibody-drug conjugates (ADC). Together, this study provides the most comprehensive molecular and functional characterization of SDSC to date. By uncovering a stem-like, ALDH1A1+ niche vulnerable to mTOR inhibition as well as mesothelin as a potential target of ADCs, our work highlights actionable biology in this otherwise lethal and understudied tumor entity. This repository holds unprocessed Seurat objects for the snRNAseq and Xenium spatial transcriptomics datasets generated in this study.