HCV epitope, homologous to multiple human protein sequences, induces a regulatory T cell response in infected patients.

BACKGROUND & AIMS: Spontaneous resolution of hepatitis C virus (HCV) infection depends upon a broad T cell response to multiple viral epitopes. However, most patients fail to clear infections spontaneously and develop chronic disease. The elevated number and function of CD3(+)CD4(+)CD25(+)FoxP3(+) regulatory T cells (T(reg)) in HCV-infected patients suggest a role of Treg cells in impaired viral clearance. The factors contributing to increased Treg cell activity in chronic hepatitis C cases remain to be delineated. METHODS: Immunoinformatics tools were used to predict promiscuous, highly-conserved HLA-DRB1-restricted immunogenic consensus sequences (ICS), each composed of multiple T cell epitopes. These sequences were synthesized and added to cultures of peripheral blood mononuclear cells (PBMCs), derived from patients who resolved HCV infection spontaneously, patients with persistent infection, and non-infected individuals. The cells were collected and following 5days incubation, quantified and characterized by flow cytometry. RESULTS: One immunogenic consensus sequence (ICS), HCV_G1_p7_794, induced a marked increase in Treg cells in PBMC cultures derived from infected patients, but not in patients who spontaneously cleared HCV or in non-infected individuals. An analogous human peptide (p7_794), on the other hand, induced a significant increase in Treg cells among PBMCs derived from both HCV-infected and non-infected individuals. JanusMatrix analyses determined that HCV_G1_p7_794 is comprised of Treg cell epitopes that exhibit extensive cross-reactivity with the human proteome. CONCLUSIONS: A virus-encoded peptide (HCV_G1_p7_794) with extensive human homology activates cross-reactive CD3(+)CD4(+)CD25(+)FoxP3(+) natural Treg cells, which potentially contributes to immunosuppression and to the development of chronic hepatitis C.

Dendritic cell-mediated, DNA-based vaccination against hepatitis C induces the multi-epitope-specific response of humanized, HLA transgenic mice.

Hepatitis C virus (HCV) is the etiologic agent of chronic liver disease, hepatitis C. Spontaneous resolution of viral infection is associated with vigorous HLA class I- and class II-restricted T cell responses to multiple viral epitopes. Unfortunately, only 20% of patients clear infection spontaneously, most develop chronic disease and require therapy. The response to chemotherapy varies, however; therapeutic vaccination offers an additional treatment strategy. To date, therapeutic vaccines have demonstrated only limited success. Vector-mediated vaccination with multi-epitope-expressing DNA constructs alone or in combination with chemotherapy offers an additional treatment approach. Gene sequences encoding validated HLA-A2- and HLA-DRB1-restricted epitopes were synthesized and cloned into an expression vector. Dendritic cells (DCs) derived from humanized, HLA-A2/DRB1 transgenic (donor) mice were transfected with these multi-epitope-expressing DNA constructs. Recipient HLA-A2/DRB1 mice were vaccinated s.c. with transfected DCs; control mice received non-transfected DCs. Peptide-specific IFN-γ production by splenic T cells obtained at 5 weeks post-immunization was quantified by ELISpot assay; additionally, the production of IL-4, IL-10 and TNF-α were quantified by cytokine bead array. Splenocytes derived from vaccinated HLA-A2/DRB1 transgenic mice exhibited peptide-specific cytokine production to the vast majority of the vaccine-encoded HLA class I- and class II-restricted T cell epitopes. A multi-epitope-based HCV vaccine that targets DCs offers an effective approach to inducing a broad immune response and viral clearance in chronic, HCV-infected patients.