Topological analysis reveals a PD-L1 associated microenvironmental niche for Reed-Sternberg cells in Hodgkin lymphoma

Signaling between programmed cell death protein 1 (PD-1) and the programmed cell death -1 ligands (PD-1 ligands, PD-L1, PD-L2) is essential for malignant Hodgkin Reed-Sternberg (HRS) cells to evade anti-tumor immunity in classical Hodgkin lymphoma (cHL). Copy number alterations of 9p24.1 /CD274(PD-L1)/PDCD1LG2(PD-L2) contribute to robust PD-L1 and PD-L2 expression by HRS cells. PD-L1 is also expressed by non-malignant tumor-associated macrophages (TAMs) but the relationships between PD-L1+ HRS cells, PD-L1+ TAMs, and PD-1+ T-cells remain undefined. We used multiplex immunofluorescence and digital image analysis to examine the topography of PD-L1+ and PD-1+ cells in the tumor microenvironment (TME) of cHL. We find that the majority of PD-L1 in the TME is expressed by the abundant PD-L1+ TAMs which physically co-localize with PD-L1+ HRS cells in a microenvironmental niche. PD-L1+ TAMs are enriched for contacts with T-cells and PD-L1+ HRS cells are enriched for contacts with CD4+ T-cells, a subset of which are PD-1+. Our data define a unique topology of cHL in which PD-L1+ TAMs surround HRS cells and implicate CD4+ T-cells as a target of PD-1 blockade.