Replicating Ad-recombinants encoding non-myristoylated rather than wild-type HIV Nef elicit enhanced cellular immunity

Objective: To determine if immunization with non-myrisloylated nef would elicit enhanced cellular immune responses resulting from improved presentation of Nef pepticles by MHC-I on the cell surface, and enhanced T-cell help. Design: The myristoylation site of HIV and SIV Nef is required for several Nef functions that modulate the immune response in an infected host, including downregulation of MHC-I, MHC-II, and CD4, and increased expression of the invariant chain on the cell surface. We constructed replication-competent Ad5- and Ad7-HIV recombinants encoding wild-type nef (nef(WT)) or a nef mutant (nef(NM)) lacking 19 amino-terminal amino acids, including the myristoylation site, and sequentially immunized chimpanzees mucosally, Methods: Peripheral blood lymphocytes were evaluated over the immunization course for Nef-specific cellular immune responses by interferon (IFN)-gamma ELISPOT and T-cell proliferation assays. Nef-specific CD4 and CD8 memory T cells that produced intracellular IFN-gamma, interleukin-2, and tumor necrosis factor (TNF)-alpha were assessed by flow cytometry. Results: In comparison to immunization with Ad-HIVnef(WT), Ad-HIVnef(NM) elicited statistically significant increases in numbers of IFN-gamma-secreting cells after the Ad7-HIVnefNm immunization and increased T-cell proliferative responses following both Ad5- and Ad7-HIVnefNm immunizations. Nef-specific CD4 and CD8 memory T-cell populations secreting TNF-alpha were also significantly increased in the Ad-HIVnefNM immunization group. Conclusions: The results support the hypothesis that immunization with Ad-recombinants encoding HIVnef(NM) rather than HIVnef(WT) elicits enhanced cellular immunity resulting from improved antigen presentation and greater T-cell help.

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