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 PD-1 ligands (PD-L1, PD-L2) is essential for malignant Hodgkin Reed-Sternberg (HRS) cells to evade antitumor 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 nonmalignant tumor-associated macrophages (TAMs), but the relationships among PD-L11 HRS cells, PD-L11 TAMs, and PD-11 T cells remain undefined. We used multiplex immunofluorescence and digital image analysis to examine the topography of PD-L11 and PD-11 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-L11 TAMs, which physically colocalize with PD-L11 HRS cells in a microenvironmental niche. PD-L11 TAMs are enriched for contacts with T cells, and PD-L11 HRS cells are enriched for contacts with CD41 T cells, a subset of which are PD-11. Our data define a unique topology of cHL in which PD-L11 TAMs surround HRS cells and implicate CD41 T cells as a target of PD-1 blockade

Spatial signatures identify immune escape via PD-1 as a defining feature of T-cell/histiocyte-rich large B-cell lymphoma

T-cell/histiocyte-rich large B-cell lymphoma (TCRLBCL) is an aggressive variant of diffuse large B-cell lymphoma (DLBCL) characterized by rare malignant B cells within a robust but ineffective immune cell infiltrate. The mechanistic basis of immune escape in TCRLBCL is poorly defined and not targeted therapeutically. We performed a genetic and quantitative spatial analysis of the PD-1/PD-L1 pathway in a multi-institutional cohort of TCRLBCLs and found that malignant B cells harbored PD-L1/PD-L2 copy gain or amplification in 64% of cases, which was associated with increased PD-L1 expression (P = .0111). By directed and unsupervised spatial analyses of multiparametric cell phenotypic data within the tumor microenvironment, we found that TCRLBCL is characterized by tumor-immune “neighborhoods” in which malignant B cells are surrounded by exceptionally high numbers of PD-L1–expressing TAMs and PD-1+ T cells. Furthermore, unbiased clustering of spatially resolved immune signatures distinguished TCRLBCL from related subtypes of B-cell lymphoma, including classic Hodgkin lymphoma (cHL) and DLBCL-NOS. Finally, we observed clinical responses to PD-1 blockade in 3 of 5 patients with relapsed/refractory TCRLBCL who were enrolled in clinical trials for refractory hematologic malignancies (NCT03316573; NCT01953692), including 2 complete responses and 1 partial response. Taken together, these data implicate PD-1 signaling as an immune escape pathway in TCRLBCL and also support the potential utility of spatially resolved immune signatures to aid the diagnostic classification and immunotherapeutic prioritization of diverse tumor types. Key Points: • Spatially resolved signatures of PD-1/PD-L1 signaling in the tumor microenvironment define T-cell/histiocyte-rich large B-cell lymphoma. • Three of 5 patients with relapsed/refractory TCRLBCL showed objective clinical responses to single-agent PD-1 blockade (pembrolizumab)