Marijke Niens

HLA Class II Expression by Hodgkin Reed-Sternberg Cells Is an Independent Prognostic Factor in Classical Hodgkin's Lymphoma

PURPOSE The neoplastic Hodgkin Reed-Sternberg (HRS) cells in classical Hodgkins lymphoma (cHL) are derived from B cells. The frequency of HLA class II downregulation and its effect on prognosis are unknown. PATIENTS AND METHODS Immunohistochemistry results for HLA class II were evaluated in 292 primary cHL patients in a population-based approach. Patients were diagnosed between 1989 and 2000 in the northern part of the Netherlands. Median age at diagnosis was 38 years (range, 8 to 88 years); 63% had Ann Arbor stage I or II, 24% stage III, and 13% stage IV disease. Median follow-up was 7.1 years. For 168 patients, HLA genotype data were available. RESULTS Lack of HLA class II cell-surface expression on HRS cells was observed in 41.4% and was more common in patients with extranodal disease, patients with Epstein-Barr virus-negative disease, and patients with HLA class I-negative HRS cells. Alleles of three microsatellite markers in the HLA class II region were associated with presence or absence of protein expression. In univariate analyses, lack of HLA class II expression coincided with adverse outcome (5-years failure free survival [FFS], 67% v 85%; P = .001; 5-years age and sex matched relative survival (RS), 80% v 90%; P = .027). This effect remained in multivariate analyses for FFS with a hazard ratio of 2.40 (95% CI, 1.45 to 3.98) and RS with a relative excess risk of death of 2.55 (95% CI, 1.22 to 5.31). CONCLUSION Lack of membranous HLA class II expression by HRS cells in diagnostic lymph node specimens is an independent adverse prognostic factor in cHL.

Serum chemokine levels in Hodgkin lymphoma patients: highly increased levels of CCL17 and CCL22

Hodgkin lymphoma (HL) is characterized by a minority of neoplastic Hodgkin‐Reed Sternberg (HRS) cells surrounded by a non‐neoplastic reactive infiltrate. As immunological mechanisms appear to be crucial in classical HL pathogenesis, altered serum chemokine levels might be related to disease activity. Serum levels of nine chemokines were examined in 163 untreated HL patients and 334 controls. We investigated single nucleotide polymorphisms (SNPs) for association with serum CCL17 (thymus and activation‐regulated chemokine, TARC) levels and HL susceptibility. Serum CCL17 and CCL22 (macrophage‐derived chemokine, MDC) levels were significantly increased in 82% and 57% of the HL patients. Nodular sclerosis cases showed increased serum CCL17 and CCL22 levels (P < 0·001) and serum levels were correlated with Ann Arbor stage. Of nine patients with pre‐ and post‐treatment serum samples, the majority showed decreased CCL17 and CCL22 levels after treatment. HRS cells expressed CCL17 and CCL22 in 77% and 75% of 74 cases. Three SNPs showed a trend of increased serum CCL17 levels with minor alleles in controls, but were not associated with HL susceptibility. CCL17 and CCL22 were the only chemokines with increased serum levels in the vast majority of HL patients, which provides further insight into the molecular mechanism(s) leading to infiltrations of reactive lymphocytes in HL.

HLA-G protein expression as a potential immune escape mechanism in classical Hodgkin's lymphoma.

Classical Hodgkins lymphoma (cHL) is characterized by the presence of an abundant reactive infiltrate, lacking effective cytotoxic responses. Especially in Epstein-Barr virus (EBV)-negative cHL, the neoplastic Hodgkin-Reed-Sternberg (HRS) cells have lost protein expression of major histocompatibility complex (MHC) class I, enabling escape from cytotoxic T lymphocyte (CTL) responses. However, downregulation of MHC class I generally induces natural killer (NK) cell activation. The paucity of NK cells in the reactive infiltrate of cHL and the systemic NK cell deficiency observed in cHL patients led us to investigate the expression of human leukocyte antigen (HLA)-G, which is known to inhibit NK-cell- and CTL-mediated cytotoxicity. By immunohistochemistry, HLA-G protein was expressed by HRS cells in 54% (95/175) of cHL cases. This expression was associated with absence of MHC class I on the cell surface of HRS cells (P < 0.001) and EBV-negative status (P < 0.001). Previously, genetic markers located in the proximity of the HLA-A and HLA-G genes had been shown to be associated with susceptibility to EBV-positive cHL. In the present study, these markers associated with MHC class I protein expression but not with presence of HLA-G. Our results suggest that induction of HLA-G protein expression in HRS cells contributes to the modulation of immune responses observed in cHL.

Genetic susceptibility to Hodgkin's lymphoma associated with the human leukocyte antigen region

Abstract:  Based on the presence of an abundant inflammatory infiltrate, expression of a broad spectrum of cytokines and the professional antigen presenting phenotype of Hodgkin Reed‐Sternberg cells it can be anticipated that immunological mechanisms play a major role in the pathogenesis of Hodgkins lymphoma (HL). Genetic susceptibility to HL probably relates to functionality of the immune system and the large number of associations with the human leukocyte antigen (HLA) region in family and population‐based studies supports this relation. In Epstein–Barr virus (EBV) positive HL cases, which usually demonstrate HLA class I expression, HRS cells should be able to present EBV derived antigenic peptides and trigger the immune system. This process depends on the affinity of the HLA binding groove for binding immunogenic peptides and thus on the HLA alleles. It can be anticipated that certain combinations of alleles predispose to or protect from the development of EBV positive HL. In EBV negative HL cases other antigenic peptides, related to malignant transformation, in combination with other HLA alleles may be involved. In addition, differential attraction and activation of inflammatory cells may influence HL subtype. In this article, possible roles of HLA in HL pathogenesis are explored and genetic associations of HLA with HL are reviewed and commented on.

The human leukocyte antigen class I region is associated with EBV-positive Hodgkin's lymphoma : HLA-A and HLA Complex Group 9 are putative candidate genes

Various studies have indicated that the human leukocyte antigen (HLA) region is associated with Hodgkins lymphoma. We recently showed a specific association of the HLA class I region with EBV-positive Hodgkins lymphoma cases. One haplotype of two consecutive microsatellite markers (D6S265 and D6S510) was overrepresented in the patient group, whereas another haplotype was underrepresented. Here, we did fine mapping of this region of ∼400 kb as a next step to find the causative single-nucleotide polymorphism(s) (SNP). To select candidate SNPs for screening the total study population, several known SNPs were determined by sequencing two individuals homozygous for either of the above-mentioned associated haplotypes. Seven SNPs displayed different alleles in these two individuals and were therefore analyzed in the total study population, including 238 Hodgkins lymphoma patients and 365 family-based controls. All seven SNPs showed significant association with the EBV-positive patient group. Two of these SNPs were analyzed in a Scottish Hodgkins lymphoma population and revealed significant associations as well. The associated SNPs are located nearby two putative candidate genes: HLA-A and HLA complex group 9. HLA-A represents the most interesting target because of its consistent expression in EBV-positive Hodgkins lymphoma cases and its ability to present EBV-derived peptides to cytotoxic T cells. (Cancer Epidemiol Biomarkers Prev 2006;15(11):2280–4)

A factorial experiment for optimizing the PCR conditions in routine genotyping

Although most PCRs would produce proper PCR products when first tried, a general optimization is required to yield the best results. This optimization is often achieved by changing one factor at a time. However, this may lead to suboptimal results, since interactions between conditions are difficult to detect with this approach. In the present study, we describe the factorial optimization of PCR conditions for microsatellite genotyping, by introducing small systematic variations in conditions during the genotyping process. The hypothesis was that small changes will not affect genotyping results, but will provide information about the optimality of current conditions. The conditions to vary were the concentrations of buffer, MgCl2, dNTPs, primers, Taq polymerase and DNA, the annealing temperature (Ta) and the number of cycles. We show that, by a 28 factorial experiment, it is possible to identify not only the factors responsible for obtaining good results, but also those responsible for bad results. However, the condition leading to the highest signals is not necessarily the best operational condition. The best operational condition is minimally sensitive to random pipetting fluctuations and yields reliable genotypes as well.

The human leukocyte antigen region and colorectal cancer risk

PURPOSERecently, we found a certain haplotype in the human leukocyte antigen Class III subregion to be associated with breast cancer. Epidemiologic studies have shown that breast cancer and colorectal cancer have several risk factors in common. In view of these studies and because polymorphisms located in the human leukocyte antigen III region have been found to be associated with colorectal cancer, we wondered whether the same region also is involved in colorectal cancer susceptibility.METHODSThe human leukocyte antigen region was genotyped with 14 microsatellite markers in germline DNA from 643 colorectal cancer patients and 841 family-based controls. Association analyses and the Haplotype Sharing Statistic were used to search for differences between patients and controls. Subgroup analyses were performed for gender, age at diagnosis, and localization of the tumor.RESULTSThe Haplotype Sharing Statistic analysis revealed neither a difference in mean haplotype sharing between all patients and controls, nor in any of the subgroups. The single allele, genotype, and two-locus association analyses for all patients and for the different subgroups did not show an association with colorectal cancer for the 14 microsatellite markers. Also, no association was observed between the tumor necrosis factor-beta polymorphism and colorectal cancer.CONCLUSIONSNo association was observed between commonly occurring haplotypes and alleles in the human leukocyte antigen region and colorectal cancer risk.