Topological importance of CD8+ T-cell enrichment in the tumor microenvironment of classic Hodgkin lymphoma

Classic Hodgkin lymphoma (CHL) histologically consists of Hodgkin Reed-Sternberg (HRS) cells and the tumor microenvironment (TME), but the relationship between TME characteristics and clinical features of CHL remains unclear. We aimed to investigate the effects of the TME structure on the outcomes of patients with CHL. We performed a high-throughput analysis of HRS cells and their topological relationship with the reactive immune cells in the TME. After multiplexed immunofluorescence labeling against CD4, CD8, CD30, CD68, CD163, PD-1, and PD-L1, visual images were analyzed. Phenotypes were assigned to all reactive cells, such as CD4⁺ and CD8⁺ T-cells and macrophages. Since the densities of PD1⁺/CD4⁺ T-cells, CD8⁺ T-cells, and PD-L1⁺ macrophages were significantly higher in the area < 60 μm than in the area < 120 μm from each HRS cell in 45 tissue samples from 34 patients with CHL, we further analyzed the TME-component cells by focusing on the 60 μm radius in the initial samples. TMEs containing > 15 CD8⁺ T-cells were associated with a significantly better 3-year progression-free survival than those with ≤ 15 CD8⁺ T-cells (100% vs. 53%, p = 0.006). In comparison with TMEs containing ≤ 15 CD8⁺ T-cells, TMEs containing > 15 CD8⁺ T-cells had significantly more PD-L1^- macrophages (mean 3 vs. 1 cell, p = 0.015) and fewer PD-1⁺/CD4⁺ T-cells (mean 16 vs. 28 cells, p = 0.036). Epstein-Barr virus positivity in HRS cells was significantly associated with a higher number of macrophages in the 60 μm radius area. In conclusion, the TME structure in patients with CHL can differ, enabling precision therapies.