Low-Dose Adenovirus Vaccine Encoding Chimeric Hepatitis B Virus Surface Antigen-Human Papillomavirus Type 16 E7 Proteins Induces Enhanced E7-Specific Antibody and Cytotoxic T-Cell Responses

Abstract

Induction of effective immune responses may help prevent cancer progression. Tumor-specific antigens, such as those of human papillomaviruses involved in cervical cancer, are targets with limited intrinsic immunogenicity. Here we show that immunization with low doses (10 6 infectious units/dose) of a recombinant human adenovirus type 5 encoding a fusion of the E7 oncoprotein of human papillomavirus type 16 to the carboxyl terminus of the surface antigen of hepatitis B virus (HBsAg) induces remarkable E7-specific humoral and cellular immune responses. The HBsAg/E7 fusion protein assembled efficiently into virus-like particles, which stimulated antibody responses against both carrier and foreign antigens, and evoked antigen-specific kill of an indicator cell population in vivo. Antibody and T-cell responses were significantly higher than those induced by a control adenovirus vector expressing wild-type E7. Such responses were not affected by preexisting immunity against either HBsAg or adenovirus. These data demonstrate that the presence of E7 on HBsAg particles does not interfere with particle secretion, as it occurs with bigger proteins fused to the C terminus of HBsAg, and results in enhancement of CD8 ؉ -mediated T-cell responses to E7. Thus, fusion to HBsAg is a convenient strategy for developing cervical cancer therapeutic vaccines, since it enhances the immunogenicity of E7 while turning it into an innocuous secreted fusion protein.Tumor cells of certain types of cancer express proteins, designated as tumor-specific antigens (TSAs), which are not present in nontumor cells. In neoplasias caused by oncoviruses, such as cervical cancers associated with human papillomavirus type 16 (HPV-16) and liver cancers caused by the hepatitis B and C viruses, the viral proteins represent TSAs. A natural mechanism for elimination of chronically infected or transformed cells is activation of cytotoxic T lymphocytes (CTLs) specific for the viral proteins. However, such proteins, are in general weak immunogens and do not induce adequate activation of antigen-specific T cells.The E6 and E7 products of HPV-16 induce transformation by blocking p53 and retinoblastoma (Rb)-mediated cell cycle control pathways, respectively, and by activating cyclins E and A (44). These proteins are constitutively expressed, albeit at low levels, in preneoplastic as well as cancer tissues and, therefore, represent persistent TSAs. Several lines of evidence suggest that E7 may be an effective immunological target for vaccines against oncogenic HPVs. Cell-mediated immunity to E7 has been demonstrated in HPV-mediated intraepithelial lesions of the uterine cervix (2, 31). Cytolytic T cells to HPV-16 E7 have been found in the blood of women with HPV-16-positive cervical neoplasia (20), and lymphoproliferative responses to E7 were found to inversely correlate with viral load (21). In addition, most cervical intraepithelial lesions caused by HPV regress spontaneously, and the phenomenon is accompanied by macrophage and CD4 ϩ

Hepatitis B Surface Antigen-based Vaccines for Human Neoplastic Disease

“…One particular study that fused HBsAg to either E7 or an N-terminally truncated E7 (Cid-Arregui et al, 2003), illustrated the formation of VLPs; however the VLP concentration was so low (~20ng/mL) that large scale production, with significant concentrating protocols would be required to generate enough antigen to deliver as a vaccine. The low yield of VLPs is likely to have contributed to the decision to use a viral vaccine vector to deliver the rHBsAg vaccine (Baez-Astua et al, 2005;Limbach & Richie, 2009). In accordance with results reported in this chapter, the concentration of VLPs in a similar study (Cid-Arregui et al, 2003) was below the detection range of the Vidas assay (Figure 3.20).…”

“…Chimeric HBsAg-based vaccines elicit CTL responses against numerous foreign antigens (Valenzuela et al, 1985;Rutgers et al, 1988;Vreden et al, 1991;Gordon et al, 1995;Gordon et al, 1996;Bisht et al, 2001;Bisht et al, 2002;Berkower et al, 2004;Puaux et al, 2004;Marsac et al, 2005;Michel et al, 2007) including HPV-16 E7 (Baez-Astua et al, 2005). DNA vaccines are cheap to produce and are effective at inducing CTL responses in animal models (Rath et al, 2005;Payette et al, 2006;Muthumani et al, 2008b;Laddy et al, 2009;Leitner et al, 2009).…”

“…The E7 protein of HPV-16 was attached to the C-terminus of HBsAg which retained VLP structure and antigenicity (Baez-Astua et al, 2005). When HBsAg was fused to the merozoite surface protein 1 of P. vivax , the ability to form VLPs was retained.…”

“…Whole E7, or E7 fused to HBsAg has been delivered to mice by an adenovirus vector system that demonstrated significant E7-specific immune responses which were enhanced by a fusion to HBsAg (Baez-Astua et al, 2005). When a replicon was used to deliver E6-E7 fusion antigens which were either mutated to eliminate oncogenicity or left native, protection was afforded against HPV-associated tumour challenge (Cassetti et al, 2004).…”

“…These vaccines rely on a structural protein which can self assemble into viral capsid structures (VLPs). Examples include the L1 protein of papillomavirus (Warrino et al, 2005) and HBsAg (Baez-Astua et al, 2005), as a fusion protein with E6 or E7 (reviewed in Stanley, 2002). A recent vaccine clinical trial used HPV16 L1 and E7 fusion VLPs (Kaufmann et al, 2007).…”