Characterising the immunopeptidome in hepatitis B
Characterising the immunopeptidome in hepatitis B Hepatitis B (HBV) is the leading cause of hepatocellular carcinoma (HCC) worldwide. There is presently no cure for HBV, and treatment options aim to reduce the risk of liver related complications and HCC. Immunotherapies have not been successful in achieving the long-term immune control required for functional cure. The display of HBV-derived peptides on major histocompatibility class I (MHC-class I, human leukocyte antigen (HLA) class I in humans) molecules on the cell surface is crucial to mount antiviral immune cell mediated responses. Immunogenic peptides are generated and selected via the HLA class I antigen presentation pathway (APP). Within this pathway, two homologous molecules tapasin and tapasin-related molecule TAPBPR optimize the peptide repertoire presented on HLA class I on the cell surface. TAPBPR is involved in both loading optimal peptides and preventing sub-optimal peptides from loading (filtering) and has been shown to restrict the peptide repertoire on HLA -A, -B and -C molecules. By using an immunopeptidomics approach, I aimed to identify novel HBV-derived epitopes presented on HLA class I molecules which could be utilised in new peptide-based immunotherapies, to induce broad T cell responses and achieve functional cure. I hypothesised that depletion of TAPBPR would alter the HBV peptide repertoire presented to immune cells. I then subsequently explored whether recombinant TAPBPR can be utilised to render HBV infected cells more immunogenic. I first conducted a comprehensive literature review on HLA presented HBV peptides which revealed that the vast majority of HBV-derived peptides characterised to date are restricted to HLA-A allotypes and are not representative of HBV endemic regions. Furthermore, the majority of studies over the last two decades have used computational methods to identify novel HLA class I presented HBV peptides. Based on the information obtained, cell lines overexpressing single HLA class I of choice and individual HBV proteins were generated and expanded for analysis by mass spectrometry (immunopeptidomics). This led to the discovery of novel HBV-derived peptides presented on HLA class I molecules. Despite successfully establishing the HLA class I immunopeptidome in HBV infected human liver tissue, no HBV-derived peptides were identified using an immunopeptidomics approach. In order to investigate the impact of the loss of TAPBPR on the HBV immunopeptidome, TAPBPRKO cells expressing individual HBV proteins were generated and expanded for immunopeptidomics analysis. Non-quantitative and semi-quantitative immunopeptidomics analysis revealed that TAPBPR filters more HBV-derived peptides than it loads and is able to shape the HBV peptide repertoire on HLA-A, -B and -C molecules. TAPBPR was also observed to reduce the abundance of virtually all HBV-derived peptides and change the abundance hierarchy of HBV peptides. Finally, by using recombinant soluble TAPBPR fused to a nanobody targeting emerald green fluorescent protein (sTAPBPR-GFPNB ), highly efficient loading of HBV-derived peptides was observed on a wide range of HLA class I allotypes on target positive cells. This has exciting translational potential to be an important therapeutic tool to trigger broad immune responses against HBV to achieve functional cure.
hepatitis B, MHC class I antigen presentation, TAPBPR
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