ALBERT

Description: Introduction The human leucocyte antigen-G (HLA-G) is a nonclassical class Ib molecule that suppresses various immune cell functions and may contribute to immune escape and cancer development. HLA-G polymorphisms, especially HLA-G-725(C/G/T) 5’URR and HLA-G 14bp del/ins 3’UTR, might influence the expression of HLA-G transcript and protein, and in consequence, affect the biological features of HLA-G. Therefore, we investigated whether these two polymorphisms, which seem to be functionally relevant, may play a role in susceptibility to chronic lymphocytic leukemia (CLL) and a clinical course of the disease. So far, no studies have reported any potentially impact of HLA-G polymorphisms on lymphoid neoplasms. Methods HLA-G725(C/G/T) 5’URR and HLA-G 14bp del/ins 3’UTR polymorphisms were genotyped in 167 previously untreated patients with CLL. The control group consisted of 98 randomly selected blood donors. Results Strong linkage disequilibrium between HLA-G-725(C/G/T) and HLA-G 14 bp del/ins was observed (D’=1.0 and r2=0.2). Six distinct haplotypes, including G/del, C/del, T/del, G/ins, C/ins, T/ins were found in the CLL patients. Among the controls only five haplotypes were found due to the T/ins haplotype not being observed. The probability of the occurrence of G/ins and T/ins haplotypes was higher in the CLL than in the controls (p= 0.01). The analysis of the prognostic significance of diplotypes, as well as the previously reported correlations between HLA-G genotypes and HLA-G expression in vitro and in vivo, allowed us to identify the HLA-G diplotype-based risk groups. The low-risk (LR) group comprised CC/del-del, CC/del-ins and CC/ins-ins diplotypes, and the high risk (HR) group included GG/del-del, GG/del-ins, GG/ins-ins, GC/del-del, GC/del-ins, GC/ins-ins, TT/del-del, TT/del-ins, TT/ins-ins and CT/del-ins diplotypes. The patients carrying LR diplotypes presented a higher 3-year treatment-free survival (TFS) (56.7%, 95% CI 47-66) than those with HR diplotypes (38.6%, 95% CI 27-52; p= 0.005). Additionally in the group of mutated IGHV patients, subjects carrying LR diplotypes presented a higher probability of 3-year TFS than those with HR diplotypes (68.5% vs 43.2%; p= 0.04). In regard to overall survival (OS), the estimated 5-year OS rates were 95.6% (95% CI 89-98) and 74.2% (95% CI 57-86) in the LR and HR group respectively (p= 0.005). Moreover, among the unmutated IGHV patients, those carrying LR diplotypes had a better 5-year OS compared to the patients with HR diplotypes (87.1% vs 71%; p= 0.02). Multivariate analysis demonstrated the IGHV mutation status (p= 0.005) and HLA-G diplotype-based risk groups (p= 0.01) to be independent factors predicting OS. Conclusions The results suggest the potential role of HLA-G and its polymorphisms in CLL. The inherited ability of the host to increase expression of the HLA-G antigen might contribute to the escape of CLL cells of the immuno-surveillance of the host and in turn to disease progression and the worse outcome for patients with CLL. Disclosures: No relevant conflicts of interest to declare.

Description: Abstract 2421 Background: Tumor necrosis factor (TNF)–α is an important pro-inflammatory cytokine involved in the modulation of lymphoma development and the balance between cell-mediated and humoral immunity. Deregulated concentrations of TNF–α have been detected in patients with lymphoma and were associated with an adverse prognosis. Evidence that the single-nucleotide polymorphism (SNP) in TNF promoter, TNF −308G〉A, could be the susceptibility locus for non-Hodgkin lymphoma (NHL) has been provided by case-control studies. Therefore we tested the hypothesis that TNF −308G〉A influences clinical course of B-cell chronic lymphocytic leukemia (CLL). Patients and Methods: We genotyped TNF −308G〉A (rs1800629) in 278 newly diagnosed patients with CLL and 192 ethnically-matched healthy individuals using the 7900 HT Real-Time (Applied Biosystems, USA). Some randomly selected DNA samples were analysed by direct sequencing using 3130xl Genetic Analyzer (Applied Biosystems, USA). Sequence data were based on the NCI SNP500 website. The IgVH mutation status in CLL patients was performed according to the protocol described by van Dongen et al. Serum samples from the 153 newly diagnosed CLL patients were collected at the time of diagnosis and tested by an enzyme-linked immunosorbent assay (ELISA) kit for human TNF (Quantikine, R&D Systems, USA). Results: The TNF −308G〉A allelic frequencies and distributions were consistent with Hardy-Weinberg equilibrium, and did not differ significantly between CLL patients and the control group. There were no significant differences between TNF allelic or genotype distributions and clinical characteristics of CLL patients at diagnosis, including age, clinical stage according to Rai classification, serum LDH and β2-microglobulin levels, surface CD38 expression, ZAP-70 expression, Döhner's cytogenetic groups, and IgVH mutation status. Neither of assessed TNF polymorphic variants was associated with response to first-line treatment, progression free survival (PFS) nor treatment free survival (TFS) that was measured from time point of diagnosis to first therapy. With a median follow-up of surviving patients of 52 months (range 1–209 months), the group of patients with TNF (−308A) allele (TNF−308AA or TNF−308AG genotypes) had significantly shorter overall survival (OS) compared to those carrying TNF (−308GG) genotype (p=0.01, log–rank test). To further characterize the prognostic impact of genetic variation in TNF on CLL patients survival, we divided the patients according to the IgVH mutation status into a IgVH unmutated (homology ≥98%) and a IgVH mutated (homology A polymorphism did not influence survival of patients with the IgVH mutated gene. To further investigate this difference, we analyzed the impact of TNF SNP on serum TNF-α levels in CLL patients at the time of diagnosis. We found that high TNF-α levels, greater than the median (16.84 pg/mL) value, correlated with stages III and IV according to Rai classification (p=0.001, chi2 test), elevated serum levels of LDH (p=0.01) or β2-microglobulin (p=0.001), CD38 expression ≥30% (p=0.001), ZAP-70 expression 〉20% (p=0.01) as well as with TNF (−308AA) genotypes (p=0.04). The patients with high TNF-α levels had significantly shorter TFS (p