Protective immunity with an E1 multivalent epitope DNA vaccine against cottontail rabbit papillomavirus (CRPV) infection in an HLA-A2.1 transgenic rabbit model

Abstract

Cottontail rabbit papillomavirus (CRPV)/rabbit model is widely used to study pathogenesis of papillomavirus infections and malignant tumor progression. Recently, we established HLA-A2.1 transgenic rabbit lines and demonstrated efficacy for the testing of immunogenicity of a well-known A2-resticted epitope (HPV16E7/82-90) [Hu J, Peng X, Schell TD, Budgeon LR, Cladel NM, Christensen ND. An HLA-A2.1-transgenic rabbit model to study immunity to papillomavirus infection. J Immunol 2006;177(11):8037-45]. In the present study, we screened five HLA-A2.1 restricted epitopes from CRPVE1 (selected using online MHCI epitope prediction software) and constructed a multivalent epitope DNA vaccine (CRPVE1ep1-5). CRPVE1ep1-5 and a control DNA vaccine (Ub3) were then delivered intracutaneously onto normal and HLA-A2.1 transgenic rabbits, respectively, by a helium-driven gene-gun delivery system. One, two or three immunizations were given to different groups of animals from both New Zealand White outbred and EIII/JC inbred genetic background. Two and three immunizations with CRPVE1ep1-5 DNA vaccine provided complete protection against viral DNA infection of HLA-A2.1 transgenic rabbits from both genetic backgrounds but not in the control-vaccinated groups. One immunization, however, failed to protect HLA-A2.1 transgenic rabbits against viral DNA infection. This study further demonstrated that the HLA-A2.1 transgenic rabbits can be used to test the immunogenicity of HLA-A2.1 restricted epitopes identified by MHCI epitope predication software.

Figure 1

A) T2 binding assay for the peptides. NDCP1-P5 represented CRPVE1/161–169, CRPVE1/245–253, CRPVE1/42–50, CRPVE1/303–311 and CRPVE1/149–157 respectively. NDCP6 (HPV16E7/82–90) and medium were positive and negative controls. T2 cells were pulsed with each peptide (10μM) and MFI of A2 expression were detected with flow cytometry. B). HLA-A2.1/peptide complex stabilization assay. NC34 (HTLV-1 Tax/67–75) and NC36 (HPV16E7/82–90) were used as controls. T2 cells were cultured with the peptide overnight and the peptide was removed. At 2, 4 and 6 hours after the peptide removal, T2 cells were labeled for A2 expression.

Figure 2

Papilloma outgrowth in outbred HLA-A2.1 transgenic and control rabbits after viral DNA challenge. HLA-A2.1 transgenic and control outbred rabbits immunized with CRPVE1ep1-5 three (A, B) and two times (C, D) were challenged with wild type (A, C) and codon optimized (B, D) CRPV DNA. HLA-A2.1 transgenic rabbits were completely protected from both wild type and codon optimized CRPV DNA challenge in both experiments. Significantly smaller papillomas were found in all immunized A2 rabbits (P<0.01, unpaired student t test).

Figure 3

Papilloma outgrowth in EIII/JC inbred HLA-A2.1 transgenic and control rabbits after viral DNA challenge. HLA-A2.1 transgenic and control inbred rabbits immunized with CRPVE1ep1-5 three (A, B) and two times (C, D) were challenged with wild type (A, C) and codon optimized (B, D) CRPV DNA. HLA-A2.1 transgenic rabbits were completely protected from both wild type and codon optimized CRPV DNA challenge in both experiments. Significantly smaller papillomas were found in all immunized A2 rabbits (P<0.01, unpaired student t test).