Evidence for distinct mechanisms of immune suppression in EBV-positive and EBV-negative Hodgkin lymphoma

Abstract

Epstein Barr Virus (EBV) has been recognized for its ability to transform B lymphocytes and for its association with different types of cancers including Hodgkin lymphoma. In addition, EBV may also modulate the microenvironment of HL. In this study, we aimed to investigate the prevalence of EBV among HL cases in Ethiopia and to assess the tissue cellular composition of EBV-related and EBV-unrelated cases. We constructed a tissue microarray (TMA) of 126 consecutive cases of classical HL (CHL) and nodular lymphocyte predominant HL (NLPHL) from a tertiary cancer centre, Tikur Anbessa Hospital, Addis Ababa, Ethiopia, and evaluated a panel of immunohistochemical markers. The quantification of immune cells was performed using HALO 2.3, a platform for image analysis from Indica Lab Inc. A total of 77/126 (61.1%) of HL cases expressed LMP1/EBER. Infiltration of CD8+, T-bet+ and FoxP3+ cells was higher in the microenvironment of EBV-related CHL, with P values of <0.001, <0.001 and <0.016, respectively. In contrast, the expression of PD1 was higher in the microenvironment of EBV-unrelated CHL cases (P < 0.001). Unlike in Western countries, the majority of HL cases in Ethiopia were associated with EBV. As FoxP3+ and PD1-expressing cells are thought to participate in down regulation of the immune response by different mechanisms, this finding highlights the previously unrecognized possibility that distinct immunosuppressive mechanisms may be ongoing within EBV positive and negative HL types. This may have important prognostic and therapeutic implications.

Keywords: Epstein Barr Virus; HL microenvironment; immune suppression.

Fig. 1

Association of age groups with LMP1 expression on HL (P = 0.02)

Fig. 2

Differences in the expression of LMP1 between HL subtypes (P < 0.001)

Fig. 3

Mean differences of immune markers expression on the cells of the tumor microenvironment of HL. Panel a through d represent data of all HL cases (CHL & NLPHL); (a) mean differences between CD3+ and CD20+ cells, (b) proportion of CD3, CD4, and CD8 and the mean difference between CD4+ and CD8+ cells, (c) proportion of T-cell subsets markers; C-maf, T-bet, FoxP3 and PD1, (d) mean difference between C-maf+ and T-bet+ cells. Panel e through i show differences of immune markers expression between CHL & NLPHL; (e) proportion of CD3+ and CD20+ cells in CHL and NLPHL subtypes, (f) proportion of CD3+, CD4+ and CD8+ cells in cHL and NLPHL subtypes, (g) proportion of C-maf+, T-bet+, FoxP3+ and PD1+ cells in CHL and NLPHL, (h) Difference of FoxP3+ cells between CHL and NLPHL, (i) Difference of PD1+ cells between CHL and NLPHL. P values are depicted where statistically significant.

Fig. 4

Mean differences of immune markers expression on the cells of the microenvironment (TME) of EBV-related and EBV-unrelated CHL cases. (a) Proportion of CD3+ and CD20+ cells, (b) Proportion of CD3+, CD4+ and CD8+ cell, (c) Difference in CD8+ cells proportion in the TME of the two groups, (d) Difference between EBV-related and EBV-unrelated CHL in CD4:CD8 ratio, (e) Proportion of C-maf+, T-bet+, FoxP3+ and PD1+ cells, (f) C-maf+ cells proportion in the TME of the two groups, (g) Difference in T-bet+ cells proportion between the two groups, (h) Difference in FoxP3+ cells proportion between the two groups, (i) Difference in PD1+ cells proportion in the TME between the two groups. P values are depicted where statistically significant. Data from panels f through i represent the same data from panel e but restratified by marker. (EBER: Epstein Barr Virus encoding-RNA, LMP1: Latent membrane protein 1)