Construction, Characterization and Immunogenicity of Human Immunodeficiency Virus-Like Particles
A vaccine expressing virus-like particles is an attractive candidate for the development of an effective vaccine for human immunodeficiency virus type 1 (HIV-1). A single vaccine plasmid was constructed to express HIV-1 Gag, Pol, Env, Tat, Rev and Vpu. Safety mutations and deletions were introduced into the VLP DNA to generate a vaccine insert that was non-infectious. The 5¡¦and 3¡¦ long terminal repeats, integrase, vif, vpr and nef were removed to further enhance the safety of the vaccine insert. Moreover, mutations were introduced into nucleocapsid and reverse transcriptase to severely restrict viral RNA packaging and to abolish RT and RNase H activity. Virus-like particles were efficiently released from primate cells, but particles were not produced in rodent cells. Therefore, purified particles were used as the inoculum to test the immunogenicity of the VLP vaccines in a rodent system. Systemic and mucosal immune responses to HIV-1 were enhanced by intranasal immunization of purified VLPs expressed from the virally-regulated multi-gene DNA vaccine. VLPs were co-immunized with cytosine-phosphate-guanosine oligodeoxynucleotides (CpG ODNs) to enhance the immune response to HIV-1 gene products. VLPs elicited specific immunity to HIV-1 antigens in both the systemic and mucosal immune compartments. Anti-Env antibodies were detected in the sera, as well as in the washes from harvested lungs, intestines and vagina from immunized mice. In addition, Env- and Gag-specific IFN-×-secreting splenocytes were elicited in the mice vaccinated with VLPs. Co-inoculation of CpG ODNs with VLPs significantly enhanced both arms of the immune response. In addition, these particulate immunogens were compared to soluble proteins (Gag and Env). Mice immunized with soluble protein alone or co-vaccinated with CpG ODNs elicited much lower immune responses compared to VLP-vaccinated mice. Specifically, CTLs were induced by vaccination with VLPs, whereas the immune response elicited by soluble proteins (+/- CpG ODNs) was almost exclusively antibody-mediated. Overall, a vaccine expressing virus-like particles is one of the most promising alternatives to replication-competent virus in eliciting high levels of cross-reactive neutralizing antibodies in combination with a robust cell-mediated response against multiple viral antigens to protect an HIV-infected host from life-long infection or disease.
Advisor:Velpandi Ayyavoo; Sharon Hillier; Kelly Stefano Cole; Ronald Montelaro; Ted Ross
School:University of Pittsburgh
School Location:USA - Pennsylvania
Source Type:Master's Thesis
Keywords:molecular virology and microbiology
Date of Publication:08/11/2005