Targeted Transcriptomic Profiling Identifies Biomarkers and Molecular Features Associated With Immunotherapy Response...

Classic Hodgkin lymphoma (CHL) is a highly curable disease. However, a subset of patients develops relapsed/refractory disease, underscoring the need for predictors of treatment response. Although immune-checkpoint inhibitors have improved outcomes, many patients still experience suboptimal responses. A deeper understanding of the CHL transcriptomic landscape may improve patient stratification and guide treatment approaches. Targeted mRNA-NGS was performed on formalin-fixed paraffine-embedded CHL-tissue using the HTG EdgeSeq Precision Immune-Oncology Panel (1,392 genes) in treatment-naive and post-chemotherapy relapsed treated with nivolumab-based protocols. Differential gene expression and pathway enrichment analyses were conducted to characterize tumor-intrinsic, and microenvironmental features associated with disease biology and treatment response. Across all CHL samples (n = 25) we observed overexpression of immune-checkpoint and immunoregulatory genes (CD274, CTLA-4, IL6, IL13), and markers of macrophage-rich/matrix-remodeling microenvironment (CD163, MMP2, TIMP1), alongside loss of B-cell identity and tumor suppression programs. Pathways analysis revealed activation of IL6/JAK/STAT3, TNF-α/NF-κB, KRAS-UP, and Inflammatory response, with reduced proliferative and metabolic activity. Immunotherapy non-responders, both in the overall (n = 5; 20%), and the treatment-naïve CHL (n = 2; 12%) displayed a proliferative, cytokine-activated, and immunologically cold phenotype. This includes shared overexpression of FADD, NFKB1, and TGFB1 and downregulation of SMAD7, together with activation of E2F, G2M, MYC, and PI3K/AKT/mTOR pathways and negative enrichment of TNF-α/NF-κB, IL6/JAK/STAT3, and Inflammatory response. Post-chemotherapy relapsed (n = 8) exhibited adaptative immunoregulatory and stress response signatures (IL15, TNFSF18, MAPK8, FUT4), along with reduced immune activation and loss of tumor-suppressor signaling. This study highlights the biological complexity and transcriptional diversity underlying CHL and its tumor microenvironment. Immunotherapy non-responders exhibited an "oncogene-driven" transcriptional profile independent of previous treatment status, whereas responders showed an "inflammatory immune-scape" phenotype, offering insight into mechanisms of resistance. Despite the limited sample size, these findings provide a foundation for larger precision-focused studies aimed at improving outcomes in CHL.