33fusion derived oncogenic programs shape the immune landscape in translocation renal cell carcinoma

Abstract Background Renal cell carcinoma (RCC) comprises a heterogeneous group of cancers with diverse molecular drivers and clinical behaviors. Despite this diversity, the majority of RCC cases are treated using therapeutic strategies developed for the most common subtype, clear cell RCC (ccRCC), due to limited understanding of subtype-specific biology. Translocation RCC (tRCC) is a rare and clinically aggressive RCC subtype defined by gene fusions involving MiT/TFE family transcription factors, most commonly TFE3. Currently, there are no molecularly tailored treatments for tRCC, and standard-of-care therapies utilized for other RCC subtypes are typically less effective in tRCC. Emerging data suggest that tRCC is molecularly distinct from other RCC subtypes. However, an incomplete understanding of both tumor-intrinsic drivers and the tumor microenvironment (TME) features of tRCC presents barriers to developing effective therapeutics for this cancer. Methods The study employed an integrative multi-omics approach to dissect the cellular and molecular landscape of tRCC. The tRCC cohort consisted of 16 tumor samples from 15 patients, analyzed using single-nucleus RNA sequencing (snRNA-seq), single-nucleus ATAC sequencing (snATAC-seq), spatial transcriptomics, and T cell receptor (TCR) sequencing. Comparative analyses were performed against single-cell datasets from ccRCC samples, and bulk RNA-sequencing datasets were integrated for validation. Differential gene expression and chromatin accessibility analyses were conducted to identify tRCC-specific transcriptional programs and regulatory elements. The tumor cell-of-origin was inferred by comparing tumor transcriptomes to a single-cell atlas of normal human kidney tissue. Tumor-intrinsic transcriptional programs and intra-tumoral heterogeneity were delineated using non-negative matrix factorization methods. Finally, we characterized the immune landscape and cellular crosstalk within the TME by integrating snRNA-seq data, immune deconvolution of bulk RNA-seq data, TCR sequencing, and spatial transcriptomics. Results Our study revealed that tRCC and ccRCC likely share a common cell of origin in the VCAM1-positive proximal tubule cells but diverge significantly at the molecular level due to their distinct oncogenic drivers. Tumors from tRCC demonstrated profound differences in transcriptional pathways compared to ccRCC, including upregulation of oxidative phosphorylation, respiration, and lysosomal pathways driven by TFE3 fusions (Figure A). Notably, tRCC tumors showed higher activity of a pigmentation-related MITF-like transcriptional program (Figure B). Chromatin accessibility profiling revealed selective enrichment of regulatory elements associated with TFE3 and epithelial-mesenchymal transition (EMT), underscoring the unique epigenomic landscape of tRCC. Despite the genetically quiescent landscape, we identified six conserved oncogenic meta-programs shared across tRCC tumors, describing the intra-tumoral heterogeneity. Among them, EMT and proximal tubule programs were largely mutually exclusive and modulated by the level of TFE3 activity, with high TFE3 expression driving EMT and suppressing epithelial identity (Figure C). The tRCC tumor microenvironment was characterized by limited infiltration of cytotoxic CD8+ T cells compared to ccRCC, many of which displayed dysfunctional phenotypes. Additionally, the TME was enriched for immunosuppressive tumor-associated macrophages and matrix-associated fibroblasts. Spatial and ligand-receptor interaction analyses revealed several suppressive cellular interactions, including those involving COL1A1-ITGAV and SPP1-ITGAV axes, which may promote EMT and reinforce local immune suppression (Figure D). Conclusions This study defines the fusion-driven oncogenic programs, intratumoral heterogeneity, and a profoundly immunosuppressive TME of tRCC, which likely underlie its poor response to immunotherapy. Our findings provide a framework for rational therapeutic development, including strategies targeting fusion-mediated transcriptional programs and reprogramming the tumor microenvironment to overcome immune resistance.