Abstract T101

Single-Nuclei and Spatial Transcriptomic Analysis of Nodular Lymphocyte-Predominant Hodgkin Lymphoma (NLPHL)

Nodular lymphocyte-predominant Hodgkin lymphoma (NLPHL) typically exhibits favorable prognosis compared to classical Hodgkin lymphoma. However, a subset of patients experience disease relapse, which can be difficult to manage, particularly in early relapses. The molecular mechanisms underlying relapse remain poorly understood. Characterization of lymphoma (LP) cells and their spatial interactions within the tumor microenvironment is crucial for understanding relapse biology and developing precision medicine approaches.

We performed single-nuclei RNA sequencing and spatial transcriptomic profiling on FFPE samples from NLPHL patients stratified by relapse status, revealing cellular heterogeneity at single cell resolution. Nuclei from 17 patients (n=9 R, n=8 NR) were analyzed, yielding 80,855 high-quality nuclei after quality control. Following Harmony integration (by patient) to correct batch effects, nuclei were clustered using Seurat's Louvain algorithm to identify nine distinct major cell types using marker gene combinations. Single-nuclei analysis revealed distinct transcriptional profiles distinguishing LP cells from normal B cells, with candidate biomarker genes showing expression differences between relapse and non-relapse patients. CellChat analysis based on snRNA-seq data identified complex intercellular communication networks involving multiple ligand-receptor pairs across different cell types. Spatial transcriptomics was performed using the 10x Genomics Xenium platform, followed by cell type annotation through label transfer from snRNA-seq data, identifying 30 cellular subtypes at single-cell resolution and revealing the characteristic popcorn-like structures where LP cells are encircled by T cells. Our integrated approach combining single-cell transcriptomics with spatial validation provides comprehensive insights into NLPHL relapse mechanisms at cellular and molecular levels. The spatially validated intercellular communication networks suggest candidate therapeutic targets, potentially informing future approaches in NLPHL treatment.