Induction of broad multifunctional CD8+ and CD4+ T cells by hepatitis B virus antigen-based synthetic long peptides ex vivo

Introduction: Therapeutic vaccination based on synthetic long peptides (SLP®) containing both CD4+ and CD8+ T cell epitopes is a promising treatment strategy for chronic hepatitis B infection (cHBV). Methods: We designed SLPs for three HBV proteins, HBcAg and the non-secreted proteins polymerase and X, and investigated their ability to induce T cell responses ex vivo. A set of 17 SLPs was constructed based on viral protein conservation, functionality, predicted and validated binders for prevalent human leukocyte antigen (HLA) supertypes, validated HLA I epitopes, and chemical producibility. Results: All 17 SLPs were capable of inducing interferon gamma (IFNɣ) production in samples from four or more donors that had resolved an HBV infection in the past (resolver). Further analysis of the best performing SLPs demonstrated activation of both CD8+ and CD4+ multi-functional T cells in one or more resolver and patient sample(s). When investigating which SLP could activate HBV-specific T cells, the responses could be traced back to different peptides for each patient or resolver. Discussion: This indicates that a large population of subjects with different HLA types can be covered by selecting a suitable mix of SLPs for therapeutic vaccine design. In conclusion, we designed a set of SLPs capable of inducing multifunctional CD8+ and CD4+ T cells ex vivo that create important components for a novel therapeutic vaccine to cure cHBV.</p

Macrophages inhibit human osteosarcoma cell growth after activation with the bacterial cell wall derivative liposomal muramyl tripeptide in combination with interferon-γ

Drug Delivery Technolog

Molecular Mimicry of Human Cytochrome P450 by Hepatitis C Virus at the Level of Cytotoxic T Cell Recognition

Hepatitis C virus (HCV) is thought to be involved in the pathogenesis of autoimmune hepatitis (AIH) type 2, which is defined by the presence of type I antiliver kidney microsome autoantibodies directed mainly against cytochrome P450 (CYP)2D6 and by autoreactive liver infiltrating T cells. Virus-specific CD8+ cytotoxic T lymphocytes (CTLs) that recognize infected cells and contribute to viral clearance and tissue injury during HCV infection could be involved in the induction of AIH. To explore whether the antiviral cellular immunity may turn against self-antigens, we characterized the primary CTL response against an HLA-A*0201–restricted HCV-derived epitope, i.e., HCV core 178–187, which shows sequence homology with human CYP2A6 and CYP2A7 8–17. To determine the relevance of these homologies for the pathogenesis of HCV-associated AIH, we used synthetic peptides to induce primary CTL responses in peripheral blood mononuclear cells of healthy blood donors and patients with chronic HCV infection. We found that the naive CTL repertoire of both groups contains cross-reactive CTLs inducible by the HCV peptide recognizing both CYP2A6 and CYP2A7 peptides as well as endogenously processed CYP2A6 protein. Importantly, we failed to induce CTLs with the CYP-derived peptides that showed a lower capacity to form stable complexes with the HLA-A2 molecule. These findings demonstrate the potential of HCV to induce autoreactive CD8+ CTLs by molecular mimicry, possibly contributing to virus-associated autoimmunity

Frequent detection of human papillomavirus 16 E2-specific T-helper immunity in healthy subjects

The incidence of genital human papillomavirus (HPV) infections is high in young, sexually active individuals. Most infections are cleared within 1 year after infection. The targets for the cellular immune response in this process of viral clearance remain to be identified, but the expression pattern of the E2 protein in early infection and low-grade cervical intraepithelial neoplasia renders this early protein a candidate antigen. Therefore, we studied the HPV16 E2-specific T-cell responses in more detail. Very strong proliferative responses against one or more peptide-epitopes derived from this antigen can be found in peripheral blood mononuclear cell cultures of approximately half of the healthy donors. Additional analysis revealed that at least a majority of these responses represent reactivity by memory CD4(+) T-helper (Th) 1-type cells capable of secreting IFN-gamma on antigenic stimulation. Interestingly, all of the E2 peptides against which strong responses were detected are clustered in the key functional domains of the E2 protein, which are conserved to considerable extent between HPV types. This suggests that HPV16 E2-specific Th memory may be installed through encounter with HPV types other than HPV16. Indeed, one HPV16 E2-specific Th clone was found to cross-react against homologuous peptides from other HPV types, but three other Th clones failed to show similar cross-reactivity. Therefore, part of the HPV16 E2-specific Th memory may relate to previous encounter of other HPV types, whereas the majority of the immune repertoire concerned is most likely established through infection with HPV16 itself. Our data are the first to reveal that the T-cell repertoire of healthy donors can contain particularly high frequencies of E2-specific memory Th cells and suggest that boosting of this immunity can be used for preventive and therapeutic vaccination against HPV-induced lesion

Frequent display of human papillomavirus type 16 E6-specific memory t-Helper cells in the healthy population as witness of previous viral encounter

Genital human papillomavirus (HPV) infection is common and the majority of infected individuals successfully deal with this virus. Clearance of HPV is presumably mediated by T cells but HPV-16-specific T-cell memory was usually detected in patients with progressive disease and not in healthy subjects, suggesting that HPV-immunity comes too late. We now show the presence of HPV-16 E6-specific memory T-helper (Th) responses in a major fraction (12 of 20) of healthy individuals by application of the IFN-gamma-ELISPOT assay. Although nearly all E6-peptides were recognized, the majority of the responders targeted peptide sequences of the COOH-terminal half (E6(81-158)) of HPV-16 E6. In a direct comparison, the presence of HPV-16 E6-specific T cells coincided with HPV-16 E2-specific T-cell reactivity in healthy individuals, whereas hardly any HPV-16 E7-specific Th immunity was found. This indicates that the induction of T-cell reactivity against HPV-16 E7 is suboptimal during infection when compared with that against HPV-16 E2 and HPV-16 E6. In conclusion, the presence of HPV-16 E6-specific Th memory in the healthy population demonstrates that HPV infection leads to T-cell immunity against immediate early proteins expressed during infection. Because this HPV-16 E6-specific T-cell immunity was frequently detected in healthy subjects, our data suggest that the observed IFN-gamma-producing proliferating T cells circulating in the peripheral blood play a role in protection against persistent HPV infection and associated development of malignancie