The breadth of CD8+ T cell responses in chronic and resolved HBV infection

Introduction: Approx. 350 million people are chronically infected with hepatitis B virus (HBV) worldwide. HBV-specific CD8+ T cells are crucial for viral clearance. T cell based immunotherapies are thus a promising strategy for clearance of chronic HBV infection. However, little is known about HBV-specific CD8+ T cell epitopes. Therefore, we aim to identify novel HBV-specific CD8+ T cell epitopes and to differentiate the CD8+ T cell repertoire in chronically HBV infected patients compared to patients who spontaneously eliminated the virus.

Methods: PBMC from 70 patients chronically infected with HBV genotype D and 16 patients with resolved HBV infection were screened in an ELISpot-Assay using overlapping peptides (OLPs, 18mers) covering the full HBV genotype D proteome. Positive ELISpot responses were confirmed by intracellular interferon-γ (IFN-γ) staining. Afterwards, epitopes were finemapped and HLA-restriction was determined using EBV-immortalized B cells. Additionally, viral sequences were determined in chronically infected patients and HLA associated sequence polymorphisms (“footprints”) were identified as indicators for escape mutations.

Results: Both the immunological and the virological analysis revealed new HBV-specific CD8+ T cell epitopes. Interestingly, chronically infected patients targeted epitopes in polymerase, core- and X-protein but not in HBsAg, whereas patients with resolved HBV infection additionally targeted epitopes in HBsAg. 50% of the patients showed at least one CD8+ T cell response to an OLP. In total, 70 epitopes could be detected.

Furthermore, most of the footprints were found in polymerase and core, but only few and weak footprints were found in HBsAg. This goes along with the lack of IFN-γ responses in the HBsAg, indicating that in patients with chronic HBV infection, CD8+ T cell responses do not target HBsAg and thus do not drive viral escape in this viral region.

Conclusion: By performing both a comprehensive analysis of HBV-specific CD8+ T cell responses as well as analysis of HLA class I footprints, we were able to identify novel HBV-specific CD8+ T cell epitopes and to demonstrate viral escape in some of these epitopes. In addition, the lack of responses to HBsAg in patients with chronic infection implies a dominant role of the HBsAg in immune regulation and persistence of the virus.

Corresponding author: Ehrenmann, Philipp S.

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