Quantitation of EBV-DNA In Peripheral Blood In Hodgkin Lymphoma: Associations with Other Biomarkers and Patient Characteristics

Abstract 2678

The Epstein-Barr virus (EBV) is present in the malignant Hodgkin/Reed-Sternberg (HRS) cells of 20–40% cases of Hodgkin lymphoma (HL) in western countries. Detection and quantitation of cell-free plasma EBV-DNA has been proposed as a biomarker in EBV-associated malignancies, including HL. We analyzed the presence of EBV in HRS cells, peripheral blood compartments, and correlated these data to EBV serology, other circulating biomarkers and clinical characteristics. In a single institution cohort of 93 HL patients, EBV-DNA was quantitated using real-time PCR for the EBNA region in whole blood (WB, n=69), plasma (PL, n=75) and peripheral blood mononuclear cells (MNC, n=74). Cell-free DNA levels were determined using a real-time PCR for the globin gene (n=84), cytokine concentrations were measured in 85 patients. EBV status of HRS cells was studied using in-situ hybridization with EBER. EBV-DNA was positive in 32% WB, 21% PL and 20% MNC at HL diagnosis, with concordant WB and PL results in 86% of cases. EBER was present in HRS cells in 32% of cases. EBV-DNA copy number over 1500/ml was predictive for the presence of EBER in HRS cells in 86% and 88% WB and PL, respectively, while EBV copy numbers <1500 had only 55% and 57% PPV for WB and PL. Older patients (>50 years) were frequently EBER-positive (66%), with higher EBV copy numbers (median 12800/ml), while younger patients (<50 years) were less frequent EBER-positive (23%), and copy numbers were lower (median 1200/ml) (p=0.01). EBER-positive HL cases frequently had EBNA-1 antibody titers < 100 U/ml when compared to EBER-negative cases (44% vs. 13%, p=0.01), and there was a trend for a negative correlation between EBV copy number and EBNA1 titers (p=0.07). In patients with EBER+ HL the plasma EBV-DNA copy number correlated to the level of circulating cell-free DNA (rho=0.65), IL-10 levels (rho=0.87), IL-6 levels (rho=0.74) and inflammation parameters, including CRP (rho=0.79) and ESR (rho=0.57) (all p<0.02), and were higher in stage IV disease (p=0.02). In conclusion, a high EBV-copy number in whole blood and/or plasma at diagnosis is a good indicator for EBV-association of HL, and EBV copy numbers strongly correlate to other biological parameters of disease activity. Moreover, the inverse correlation between EBV copy number and EBNA1 antibody titers may indicate a reduction in immune surveillance favoring the expansion of EBV.