Combination of cell penetrating peptides and heterologous DNA prime/protein boost strategy enhances immune responses against HIV-1 Nef antigen in BALB/c mouse model

Abstract

To develop a strong HIV specific T-cell response, the HIV-1 Tat and Nef regulatory proteins have been known as attractive antigenic candidates in vaccine design. A peptide transduction domain of Tat (48–60 aa) could act to deliver other therapeutic molecules into different cells. In this line, several cell-penetrating peptides (CPPs) have been designed to transfer DNA, siRNA, polypeptides and proteins into cells through non-covalent approach such as CADY and PEP families. Some studies showed that the endogenous adjuvants including heat shock protein Gp96 could stimulate antigen-specific T cell immune responses. In this study, different Nef DNA and protein constructs were generated, and their abilities were evaluated to induce T cell immune responses and humoral immunity in mouse model. A novel prime-boost immunization approach was also used in which the priming injections consisted of Nef/Tat (PTD)-Nef DNA plus Gp96 DNA followed by Nef/Tat (PTD)-Nef protein formulated in Freund’s adjuvant or the Pep-1 and Cady-2 CPPs. Generally, our results indicated that Tat (PTD)-Nef fusion DNA or protein could significantly elicit higher humoral and cellular immune responses than Nef DNA or protein, respectively. Analysis of the immune responses demonstrated that the Tat (PTD)-Nef+Gp96 DNA prime/Tat (PTD)-Nef protein+Cady-2 boost regimen significantly enhanced the Nef/Tat (PTD)-Nef-specific T cell responses. This modality induced high levels of IgG2a and IFN-γ directed toward Th1 responses, and also Cytotoxic T Lymphocytes (CTLs) activity as compared to other immunization strategies. The immunostimulatory properties of Pep-1 and Freund’s adjuvant were almost similar in different immunization strategies. These findings showed that the use of Tat (PTD)-Nef antigen in prime-boost strategy along with Gp96 adjuvant and Cady-2 CPP may have practical implications for developing HIV-1 vaccine in large animal model.