Evaluation of T-cell and B-cell epitopes and design of multivalent vaccines against HTLV-1 diseases
Human T-cell lymphotropic virus type I (HTLV-1) is a C type retrovirus that is the causative agent of an aggressive T-cell malignancy, adult T-cell leukemia/lymphoma (ATLL). The virus is also implicated in a number of inflammatory disorders, the most prominent among them being HTLV-1 associated myelopathy or tropical spastic paraparesis (HAM/TSP). HTLV-1, like many viruses that cause chronic infection, has adapted to persist in the face of an active immune response in infected individuals. The viral transactivator Tax is the primary target of the cellular immune response and humoral responses are mainly directed against the envelope protein. Vaccination against HTLV-1 is a feasible option as there is very little genetic and antigenic variability. Vaccination regimes against chronic viruses must be aimed at augmenting the immune response to a level that is sufficient to clear the virus. This requires that the vaccine delivers a potent stimulus to the immune system that closely resembles natural infection to activate both the humoral arm and the cellular arm. It is also clear that multicomponent vaccines may be more beneficial in terms of increasing the breadth of the immune response as well as being applicable in an outbred population. As a step in this direction, this dissertation work first describes the identification and evaluation of human T-cell and B-cell epitope based vaccines that are capable of inducing robust immune responses. A novel strategy was designed for delivery of multiple Tax protein derived CTL epitopes into the same antigen presenting cell for the simultaneous priming of anigen specific T-cells. This design allowed antigen processing by cellular proteasomes to efficiently liberate the individual minimal epitopes endogenously for presentation via MHC Class I. This type of construct was effective in vivo in HLA-A*0201 transgenic mice in inducing cellular immune responses against all individual epitopes. A statistically significant reduction in viral replication was observed in mice that were immunized with a multiepitope Tax CTL construct and challenged with recombinant Tax vaccinia virus, by the induction of antiviral IFN-g and cytolysis of infected target cells. This reduction was determined to be the result of immune responses specifically targeting the Tax protein and was dependent on the presence of CD8+ T-cells. Studies were also undertaken to engineer peptides that have a high propensity to fold into native protein like structure for the induction of antibodies that have a high affinity for the complex transient secondary structure adopted by the native protein. A single matrix multicomponent template strategy was applied to design a peptide from the central region of the TM (residues 347-374) that forms a parallel trimeric hairpin structure. Mutational studies have implicated this region to be critical for the fusion process after receptor binding. The template design served to constrain the individual peptide strands at one end and bring them in close proximity to promote hydrophobic interactions to initiate a coiled coil formation. Key mutations in the native peptide sequence were also made to increase hydrophobic interactions. We observed that the mutations combined with the template design resulted in a peptide that had a high helical content in aqueous medium and antibodies raised against the peptide recognized native viral protein with higher affinity compared with the wild type template peptide antibodies. These results suggested the presence of native sequence like conformation. Furthermore, a chimeric peptide incorporating a “promiscuous” T-helper epitope derived from the region of chain reversal in the TM subunit (residues 392-415 of gp21) implicated in the fusion process was tested for its ability to induce neutralizing antibodies. This peptide induced high titered antibodies that were capable of inhibiting HTLV-1 infected cell induced syncytia formation. These data, taken together may have implications in the development of multivalent vaccines against HTLV-1.
School:The Ohio State University
School Location:USA - Ohio
Source Type:Master's Thesis
Keywords:peptide vaccine design multivalent hla a2 1 transgenic mice antibodies coiled coil b cell epitopes t
Date of Publication:01/01/2003