In vitro and in vivo characterization of the cytomegalovirus and polyomavirus BK specific immune response

Abstract

During my PhD thesis several aspects of the interaction of Cytomegalovirus and Polyomavirus BK with the host’s immune system were examined (see list of publications). The overall aim was to compare immune response in healthy individuals and kidney transplant recipients with or without viral replication. In healthy individuals, Polyomaviruses BK and JC infect 80% and 60%, respectively. For CMV seroprevalences may reach up to 80%. Intermittent virus shedding in urine is observed for BKV in 7%, JCV in 19% and CMV in 0%. However, no virus replication in plasma was detected. Posttransplant, mainly due to prolonged immune suppression the amount and function of CMV- and BKV-specific T-cells is impaired. Calcineurin inhibitors lead to a direct reduction of INFγ production of virus-specific T-cells, whereas antiproliferative immunosuppressives reduce the expansion capacity in a dosedependent manner. This may be a major reason for uncontrolled virus replication. The humoral response reflects the amount of recent antigen exposure and does not directly indicate protection from virus replication. Virus-specific cellular immune responses would probably allow to assessing the risk of future replication. Overall the importance of CMV and BKV specific T-cells posttransplant in controlling virus replication was examined. For both viruses we could calculate a protective threshold of virus-specific T-cells. CMV-pp65 specific CD4 T-cells above 0.03% were significantly associated with no CMV replication during the next eight weeks. Additionally, below this cut-off, CMV seropositive recipients developed more often GCV-resistant CMV replication. During BKV replication, patients with more than 69 BKV-LT specific T-cells per 1 Mio PBMCs were significantly more often showing decreasing BKV loads in plasma. As virus-specific T-cells seem to be crutial in reducing virus replication, and reduction of immune suppression harbours the risk of acute rejection, a booster vaccine could be a new therapeutic option. A booster vaccine could probably elevate the amount of virus-specific T-cells above a critical threshold of protection from disease, despite effects of immune suppression. We tried to identify immunodominant regions with the CMV pp65 and BKV LT proteins. We used a combined approach of computer prediction algorithms and experimental verification. Epitope mapping of BKV LT with computer prediction revealed several clusters, which could be immunodominant and also potentially be processed and recognized in various HLA backgrounds. The identified cluster regions were synthesised as 15 and 25mers. Expansion and re-stimulation with predicted epitopes could so far confirm the HLA A and B-specific prediction of single 9mers covered by the larger 15 and 25mer sequences. However, other HLA types need to be tested for direct stimulation and expansion potential of the predicted epitopes. Additionally, tetramer staining should be performed for verification. Based on this research, we will be able to improve current immune monitoring and probably a high-specific peptide-based vaccine against BKV LT could be developed and be used to increase the amount of BKV-LT specific T-cells. Another potentially therapeutic agent could be the blockade of PD1 ligand. PD1 expression in chronic virus infection lead to impaired CD8+ T-cell function. CMV-specific CD4 T-cells treated with an inhibitory antibody against PD1 ligand, and thereby activating CD4+ T-cells, lead to a increase of the expansion capacity. We have shown, that the anti-PD1 ligand antibody increases various cytokines. This could be also tested for BK virus. Measurement of virus-specific T-cells may replace serological assays in the future, due to a better correlation to effective antiviral control, which can be used as monitoring tool during infection and post-vaccination.