Although multiple studies have investigated TME and PD-L1 expression in B-cell lymphomas, only limited, small studies have been conducted in BL.14−16
In the present study, we extensively evaluated the TME composition, activation status and expression of inhibitory immune checkpoints both on the inflammatory infiltrate and neoplastic cells of BL tumors including eBL, sBL and idBL cases. Thus, we investigated PD-L1 expression and the contribution of EBV in fostering the activation of the PD1-PD-L1 axis. The influence of the microenvironment on cell proliferation and destruction varies greatly according to the inherent histotype of the lymphoma cell type.29−31 In particular, Hodgkin lymphoma (HL) tissue often consists of relatively few monoclonal cancer cells but at least 90% non-malignant cells (e.g., regulatory T cells), contributing to a rather unique surrounding immune ecosystem. On the other hand, BL seems to be largely devoid of such a supportive cellular environment, although the high content of Tumor-Associated Macrophages (TAMs) might play a distinct, specific, important role in neoplastic progression through secretion of chemokines, cytokines and immune checkpoint-associated proteins as PD-L1.12,13
In the recent decade a model has been developed to describe the complex mechanism of macrophage activation as a polarization towards two opposite states, namely M1 and M2, with pro-inflammatory and pro-tumoral properties respectively.32−36 TAM density in particular M2-TAMs have been associated with tumor progression and poor prognosis in DLBCL. 37, 38
In the present work, we identified a polarization towards a M2 phenotype of TAMs in all cases by applying three different approaches (CIBERSORT, mIHC and mIF) regardless of the subtype of BL, and thus, of EBV status.
These cells, intimately associated with the neoplastic cells, constituted also the major source of PD- L1, which may inhibit the overall inflammatory response and allow the neoplastic cells to evade antitumor immunity. However, the lower rate of PD-L1 expression on TAMs in sBL, as compared with eBL and idBL, which are frequently associated with EBV, may suggest a role of the virus in inducing PDL1 expression.
PD-L1 is a major regulator of T cell function and, after engaging PD-1, leads to an altered functional state of T-cells, namely T cell exhaustion.39 In this regard, we found that in eBL and idBL the vast majority of the CD8 + infiltrating T cells expressed PD-1 highlighting an adaptive immune response resistance mechanism in such cases.
However, the role of PD-1 expression in Tumor Infiltrating Lymphocytes (TILs) on both lymphoid and epithelial malignancies is controversial.38 PD-1 expression in CD8 + cells has been associated with the selective suppression of cytotoxic lymphocytes in EBV positive nasopharyngeal carcinoma.40 On the other hand, the PD-1 + TILs have also been described to lack Tim-3 expression in papilloma virus positive cancers, and thus possibly representing activated T-cells.41
Emerging evidence in EBV-related malignancies indicates that the virus possesses the ability to actively shape the tumor microenvironment, and favours its escape from anti-tumor immune responses through a variety of complex mechanisms.18 EBV may induce a strong up-regulation of PD-L1 expression both directly on the surface of human primary monocytes, or indirectly on neoplastic cells, through its viral proteins LMP-1 and LMP-2 which interfere with downstream cellular signalling (i.e. AP1; JAK/STAT) 42,43 to induce an immune tolerant niche for EBV- related tumors.44−46
Although PD-L1 expression has been largely investigated in B cell lymphomas, the distinction of its expression in cellular microenviroment and/or in tumour cells has not been made in most studies.1−4
Here we showed that EBV in BL might induce PD-L1 expression on tumor cells in a minority of cases characterized by a non-canonical latency with LMP2A positivity. On the other hand, it might influence PD-L1 upregulation on TAMs also in cases with canonical EBV latency I. However, the prevalence of M2 macrophages as primary constituent of the TME in BL is a constant finding in all BL subtypes and thus, macrophage polarization towards a pro-tumoral state seems an event related to the intrinsic characteristics of the tumor.