Primary Hodgkin lymphomas (HL) contain small numbers of neoplastic Reed-Sternberg (RS) cells within an extensive host inflammatory infiltrate, which includes few cytotoxic T-cells. The lack of an effective host anti-tumor response prompts speculation regarding the mechanisms that HL RS cells use to evade immune surveillance. To identify novel HL-specific T-cell inhibitory molecules, we compared the gene expression profiles of a series of HL and diffuse large B-cell lymphoma (DLBCL) cell lines. The HL cell lines overexpressed galectin-1 (GAL1), a carbohydrate-binding lectin that selectively induces the apoptosis of cytotoxic T cells. GAL1 mRNA abundance was 5- to 33- fold higher in HL cell lines than in DLBCL lines. GAL1 protein expression was also uniformly high in HL cell lines and low or undectable in DLBCL lines by western blotting.

Immunohistochemical staining of primary tumors revealed abundant GAL1 expression in HL RS cells whereas DLBCLs were uniformly negative. To elucidate the mechanisms responsible for GAL1 over expression in HL, we analyzed the GAL1 locus and identified a candidate AP1 enhancer element 1.5 kb downstream of the GAL1 transcription start site. The predicted enhancer element increased the expression of a reporter gene (luciferase) 5-fold in a HL cell line but had no effect in either a DLBCL or fibroblast cell line. In electrophoretic mobility studies, AP1 bound to the predicted GAL1 enhancer element in a cell-type specific manner. In addition, mutation of the AP1 binding site in the GAL1 enhancer abolished its activity. Since the AP1 components, cJUN and JUN-B, are known to be overexpressed in HL, we analyzed cJUN and JUN-B copy numbers and transcript abundance in the HL cell lines using 100K high-density SNP arrays and oligonucleotide RNA microarrays. We found increased copy numbers of cJUN and/or JUN-B in all HL cell lines tested and confirmed these findings using fluorescence in situ hybridization (FISH). Increased cJUN and JUN-B copy numbers were associated with significantly higher transcript abundance in the HL cell lines. In additional functional assays, we confirmed previous reports that recombinant GAL1 induced the apoptosis of CD3+ PHA-stimulated lymphoblasts in a dose- and time-dependent manner. Taken together, these studies indicate that HL RS cells selectively over express GAL1, at least in part, via AP1-mediated induction of the GAL1 enhancer. Given the profound effects of GAL1 on cytotoxic T cells, RS-specific expression of GAL1 likely inhibits the host anti-tumor response in HL.

Disclosure: No relevant conflicts of interest to declare.

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