Little is known about the gene expression profile and significance of the rosetting CD4 þ CD26À T cells in classical Hodgkin's lymphoma (cHL). To characterize these T cells, CD4 þ CD26À and CD4 þ CD26 þ T-cell populations were sorted from lymph node (LN) cell suspensions from nodular sclerosis HL (NSHL) and reactive LNs. mRNA profiles of stimulated and resting cell subsets were evaluated with quantitative RT-PCR for 46 genes. We observed a higher percentage of CD4 þ CD26À T cells in NSHL than in reactive LNs. The resting CD4 þ CD26À T cells in NSHL showed higher mRNA levels of CD25, CTLA4, OX40 and CCR4 compared with in LNs, supporting a regulatory T-cell (Treg) type, and this was validated by immunohistochemistry. Moreover, these cells showed low or no expression of the Th1-or Th2-related cytokines IL-2, IFN-g, IL-13, IL-12B, IL-4, and IL-5, and the chemoattractant receptor CRTH2. Besides Tregs, Th17 cells may exist in NSHL based on the significantly higher IL-17 mRNA level for both T-cell populations in NSHL. Upon stimulation in vitro, lack of upregulation of mRNA levels of most cytokine genes indicated an anergic character for the CD4 þ CD26À T-cell subset. Anergy fits with the Treg profile of these cells, probably explaining the immunosuppressive mechanism involved in NSHL.

Since Hodgkin and Reed-Sternberg (HRS) cells of Hodgkin lymphoma (HL) generally have immuno-1-4 Immunohistochemical analysis of the neoplastic cells of CHL has revealed an immunophenotype that is very specific, but highly unusual for a B-cell-derived tumor. The so-called Hodgkin and Reed-Sternberg (HRS) cells typically express CD30 and CD15, have inconsistent expression of CD20 and CD79a, and typically lack J-chain and immunoglobulins. 5,6 The neoplastic cells of NLPHL, the so-called lymphocytic and histiocytic (L&H)-type HRS cells, do express classical B-cell markers like CD20, CD79a, and J-chain 7 but lack CD30 and CD15.

Little is known about the cytokine profile of nodular lymphocyte predominance Hodgkin's lymphoma (NLPHL) and the significance of the characteristic rosetting CD4(+)/CD57(+) T cells. We analysed the T lymphocyte populations isolated from lymph node suspensions from five patients with NLPHL, two with follicular hyperplasia and progressive transformation of germinal centres (PTGC), three with classical Hodgkin's lymphoma (CHL) and five with hyperplasia of the tonsil. We sorted the T cells based on expression of CD3, CD4 and CD57 by flow cytometry and evaluated the cytokine mRNA profiles of the T cells with quantitative RT-PCR. NLPHL cases were as rich in T cells as the CHL cases, but all NLPHL cases had a much higher frequency of CD4(+)/CD57(+) T cells. In contrast to the CD4(+)/CD57(+) T cells from tonsils, IL2 and IL4 mRNAs were consistently absent from the CD4(+)/CD57(+) T cells of NLPHL. Even after stimulation, no IL4 transcripts could be detected in the CD4(+)/CD57(+) T cells of NLPHL. On the other hand, IFNgamma transcripts were elevated in NLPHL and PTGC T cell subsets as compared to tonsillar T cell subsets. IL13 mRNA was exclusively produced by the T cells of CHL cases, indicating that IL13 may be a key cytokine in CHL. In conclusion, elevated levels of CD4(+)/CD57(+) T cells are characteristic of NLPHL and these T cells display a distinct cytokine mRNA profile.

Mantle cell lymphoma (MCL) is characterized by genetic instability and a poor prognosis. Many blastoid variants are (hypo)tetraploid and have an even worse prognosis. We investigated the role of signalling by mitogen-activated protein kinases (MAPKs) in MCL. As compared to normal tonsil B cells, MCL cells showed higher activation of the JNK MAPK in both an MAPK array and a sandwich ELISA assay. Immunohistochemistry showed overexpression of phospho (p)-JNK (Thr183/Tyr185) in 30 of 37 MCL cases. Inhibition of p-JNK with SP600125 resulted in growth arrest in all four MCL cell lines (Jeko-1, HBL-2, UPN-1, Granta-519), which could be partly reversed by the addition of CD40L and IL-4. Furthermore, SP600125 led to G2/M phase arrest on day 1 and a striking increase in endoreduplication on day 2 and day 3, which was confirmed by karyotype analysis. G2/M arrest was associated with down-regulation of EGR1 and p21 protein expression. SP600125-induced polyploidy could be blocked by the BCL-2 inhibitor YC137. These data suggest that constitutive JNK activity is necessary to promote proliferation and maintain diploidy in MCL. JNK inhibition leads to cell cycle deregulation and endoreduplication, mimicking the tetraploid state seen in a subset of MCL cases. Thus, our data also provide an experimental model to study polyploid MCL cells.

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Current genomic models in diffuse large B-cell lymphoma (DLBCL) are based on single tumor biopsies, which might underestimate heterogeneity. Data on mutational evolution largely remains unknown. An exploratory study using whole exome sequencing on paired (primary and relapse) formalin fixed paraffin embedded DLBCL biopsies (n = 14) of 6 patients was performed to globally assess the mutational evolution and to identify gene mutations specific for relapse samples from patients treated with rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisolone. A minority of the mutations detected in the primary sample (median 7.6%, range 4.8–66.2%) could not be detected in the matching relapse sample. Relapsed DLBCL samples showed a mild increase of mutations (median 12.5%, range 9.4–87.6%) as compared to primary tumor biopsies. We identified 264 genes possibly related to therapy resistance, including tyrosine kinases (n = 18), (transmembrane) glycoproteins (n = 73), and genes involved in the JAK-STAT pathway (n = 7). Among the potentially resistance related genes were PIM1, SOCS1, and MYC, which have been reported to convey a risk for treatment failure. In conclusion, we show modest temporal heterogeneity between paired tumor samples with the acquisition of new mutations and identification of genes possibly related to therapy resistance. The mutational evolution could have implications for treatment decisions and development of novel targeted drugs.

To identify genes involved in the pathogenesis of classical Hodgkin lymphoma (cHL), we performed serial analysis of gene expression (SAGE) and array-based comparative genomic hybridization (aCGH). Comparison of SAGE libraries of cHL cell lines L428 and L1236 with that of germinal centre B cells revealed consistent overexpression of only 14 genes. In contrast, 141 genes were downregulated in both cHL cell lines, including many B cell and HLA genes. aCGH revealed gain of 2p, 7p, 9p, 11q and Xq and loss of 4q and 11q. Eighteen percent of the differentially expressed genes mapped to regions with loss or gain and a good correlation was observed between underexpression and loss or overexpression and gain of DNA. Remarkably, gain of 2p and 9p did not correlate with increased expression of the proposed target genes c-REL and JAK2. Downregulation of many genes within the HLA region also did not correlate with loss of DNA. FSCN1 and IRAK1 mapping at genomic loci (7p and Xq) that frequently showed gain were overexpressed in cHL cell lines and might be involved in the pathogenesis of cHL.

There are several indications that classical Hodgkin lymphoma (cHL) and at least a proportion of cases of Primary Mediastinal B cell Lymphoma (PMBL) are derived from B cells at similar stages of differentiation and share common pathogenic mechanisms. The first indication was the existence of mediastinal grey zone lymphomas as identified in the 4th International Symposium on HL, with clinical, histological and immunohistochemical features intermediate between cHL and PMBL. Second, both tumor types resemble a cell that is developmentally situated in‐between the germinal center reaction and a plasma cell. Third, cHL and PMBL were found to have similar gene expression profiles, including the lack of immunoglobulin expression and low levels of B cell receptor signalling molecules, and the secretion of molecules like the chemokine TARC and the prominent expression of IL‐13 receptors. Fourth, both entities were found to have common genomic aberrancies, notably in 2p15 and 9p24, the sites of the REL oncogene and the tyrosine kinase gene JAK2, respectively. Further comparison of both lymphoma types may provide further insight in the pathogenic mechanisms and allow the design of diagnostic algorithms to sort out the small number of so‐called mediastinal grey zone lymphomas, that appear to be intermediate between PMBL and cHL.