Integrated Genomic and Proteomic Analysis Reveals T‐B Lymphocyte Signatures in the MYCN Driven “Immune Desert” of Specific Neuroblastoma Subtypes

Abstract

ABSTRACT Aims This study aims to systematically dissect how MYCN amplification shapes the immunosuppressive tumor microenvironment (TME) in high‐risk neuroblastoma, elucidating key mechanisms underlying immune evasion. Methods We performed an integrated multi‐omics analysis of bulk RNA‐seq (n = 721), single‐cell RNA‐seq (n = 9), proteomic data (n = 49) and spatial transcriptomics (Visium, with external validation in melanoma). Analyses included unsupervised clustering, cell–cell communication inference, transcriptional regulatory network reconstruction, and spatial proximity assessment to map the immune landscape. Results A distinct molecular subtype (Class C), defined by MYCN amplification and poor prognosis, exhibited a comprehensive “immune desert” phenotype characterized by low immune scores and minimal leukocyte infiltration. Single‐cell analysis confirmed significant depletion of T and B lymphocytes within the Class C TME. Dysregulated transcriptional networks were identified, including upregulation of REL and EOMES in T cells—with EOMES potentially driving exhaustion via regulation of Transient Receptor Potential (TRP) genes, and REL inhibition enhancing cytotoxic function in vitro. A unique immunosuppressive B‐cell subset (B7) engaged in enhanced crosstalk with exhausted T cells and harbored a MYC‐centered network linked to cell cycle dysregulation and poor survival. Spatial transcriptomics revealed significant proximity between B7‐active regions and Treg/exhaustion‐enriched areas, externally validated in melanoma. Proteomic data validated elevated REL expression in MYCN‐amplified tumors. Conclusion This work delineates the immunosuppressive architecture of MYCN‐driven neuroblastoma, revealing novel regulatory nodes within specific lymphocyte compartments. Integrating single‐cell, spatial, and proteomic evidence, we propose REL inhibition as a therapeutic candidate, the EOMES/TRP axis as a bioinformatically supported hypothesis, and the B7/MYC hub as a hypothesis supported by transcriptomic and spatial evidence.