Primary central nervous system (PCNS)-lymphoma is an Epstein–Barr virus (EBV)-associated opportunistic malignancy of HIV-infected persons [1,2]. HIV-infected individuals progressing to PCNS-lymphoma were recently found to lack EBV-specific CD4+ T cells irrespective of their absolute CD4+ T cell counts . Here, we tested the hypothesis that lack of CD4+ T cell-help for EBV-specific B cells impacts on EBV-specific antibody production, translating into changes in the humoral immuneresponse.
Six individuals with confirmed diagnosis of PCNS-lymphoma and pre-disease blood samples available were identified within the Swiss HIV Cohort Study databank . In these six cases, levels of viral capsid antigen (VCA)- and EBV-nuclear antigen (EBNA)-specific immunoglobilin (Ig)G were assessed. In four of the cases, also cytomegalovirus (CMV) IgG levels were assessed (two cases were CMV negative). A total of 38 samples were analyzed; the median number of samples per patient was six (range 4 to 11), taken at approximately 6-monthly intervals and covering a mean period of 3.7 years (range 1.7–6.8 years). The study was Institutional Review Board-approved, conducted according to the principles expressed in the Helsinki declaration, and written informed consent was obtained from all participants. Antibody levels were determined in serum samples using commercially available enzyme-linked immunosorbent assay kits (ImmunoWell IgG test, GenGio, San Diego, California, USA). In these same samples, we also performed quantitative EBV polymerase chain reaction (PCR) analyses. Viral DNA quantification was performed using routine TaqMan real-time PCR technique (Applied Biosystems, Foster City, California, USA). Median IgG levels and trends over time, as well as EBV DNA levels, were compared with matched HIV-infected control-individuals (1: 1 matching, 41 samples from six control patients). Clinical characteristics of case and control individuals are summarized in Table 1. Repeated IgG measurements per patient were summarized by their medians and a slope over time to indicate potential time trends. The slope was estimated by linear regression. Median values and slopes of cases and matched controls were compared with a paired Wilcoxon signed rank test or a paired t-test, as indicated. All reported P-values are based on two-sided tests and performed at the 5% significance level. Statistical analyses were performed with R version 2.3.1 software (R Foundation for Statistical Computing; http://www.R-project.org).
Overall median VCA levels were 1347.25 U/ml (range 1196–1728 U/ml) in cases and 1292.75 U/ml (566.5–1452 U/ml) in controls. The median (as well as mean) value in cases was always higher than in the respective controls (P = 0.03, Wilcoxon test). The median difference between cases and respective controls was 150.25 (range 1–694 U/ml) (Fig. 1a). There was no noticeable time-trend in VCA values over time: Median slopes per year were −6.9 U/ml (range −232.4 to +32.6 U/ml) in cases, and −7.2 U/ml (−100.4 to +175.0 U/ml) in controls, and cases and controls did not significantly differ with respect to their slopes (P = 1, Wilcoxon test). Overall median EBNA levels were 2274.5 U/ml (range 1615–2866.5 U/ml) in cases and 2136.5 U/ml (1007.5–2450 U/ml) in controls. Except for one case–control pair, the median value in cases was always higher than in the respective controls (P = 0.06, Wilcoxon test). The median difference between cases and their respective controls was 420.25 U/ml (range −59.5 to +925 U/ml) (Fig. 1b). Again, there were no noticeable time-trends (data not shown). Overall median optical densities (ODs) of CMV IgG measurements were 3.06 (range 1.99–3.12) for cases, and 3.11 (1.02–3.23) for controls. Differences in ODs between cases and respective controls were −0.15, −0.10, 0.03 and 0.97 (P = 0.53, t-test) (Fig. 1c), with no noticeable time-trends (data not shown). In none of 38 samples from individuals later developing PCNS-lymphoma EBV DNA was detected. In the control group, one of 41 samples tested positive for EBV DNA, which is in line with previous data .
Presuming that the humoral immuneresponse is important in controlling latent EBV infection , these data seem to indicate that exposure to viral antigens in these CD4+ T cell-‘helpless’ individuals is triggering an inefficient (i.e. qualitatively impaired) antibody-response. Intriguingly, a recent study in patients developing nasopharyngeal carcinoma (another EBV-associated malignancy) also found increased levels of EBV VCA-directed antibodies . The nature of the postulated qualitative defect in antibody-production in PCNS-lymphoma patients remains to be determined.
To our knowledge, this is the first controlled study in this setting, and unique in that patients were assessed longitudinally over several years prior to the development of disease. The limitation of this study is the small sample size, and larger, ideally prospective studies will need to further address the role of quantitative versus qualitative differences in the pathogen-specific humoral immuneresponse.
We thank J. A. Schifferli and P. Erb for discussion and critical review of the manuscript. The members of the Swiss HIV Cohort Study are M. Battegay, E. Bernasconi, J. Böni, H. Bucher, Ph. Bürgisser, S. Cattacin, M. Cavassini, R. Dubs, M. Egger, L. Elzi, P. Erb, M. Fischer, M. Flepp, A. Fontana, P. Francioli, H. Furrer, M. Gorgievski, H. Günthard, B. Hirschel, I. Hösli, Ch. Kahlert, L. Kaiser, U. Karrer, O. Keiser, C. Kind, Th. Klimkait, B. Ledergerber, B. Martinez, N. Müller, D. Nadal, M. Opravil, F. Paccaud, G. Pantaleo, L. Perrin, J.-C. Piffaretti, M. Rickenbach, C. Rudin, P. Schmid, D. Schultze, J. Schüpbach, R. Speck, P. Taffé, P. Tarr, A. Telenti, A. Trkola, P. Vernazza, R. Weber, S. Yerly.
Sponsorship: This work was supported by a Swiss HIV Cohort Study-grant and a Swiss National Science Foundation Professorship-grant to C.H. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
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