Development of DNA Vaccine Targeting E6 and E7 Proteins of Human Papillomavirus 16 (HPV16) and HPV18 for Immunotherapy in Combination with Recombinant Vaccinia Boost and PD-1 Antibody

Abstract

Immunotherapy for cervical cancer should target high-risk human papillomavirus types 16 and 18, which cause 50% and 20% of cervical cancers, respectively. Here, we describe the construction and characterization of the pBI-11 DNA vaccine via the addition of codon-optimized human papillomavirus 18 (HPV18) E7 and HPV16 and 18 E6 genes to the HPV16 E7-targeted DNA vaccine pNGVL4a-SigE7(detox)HSP70 (DNA vaccine pBI-1). Codon optimization of the HPV16/18 E6/E7 genes in pBI-11 improved fusion protein expression compared to that in DNA vaccine pBI-10.1 that utilized the native viral sequences fused 3′ to a signal sequence and 5′ to the HSP70 gene of Mycobacterium tuberculosis. Intramuscular vaccination of mice with pBI-11 DNA better induced HPV antigen-specific CD8+ T cell immune responses than pBI-10.1 DNA. Furthermore, intramuscular vaccination with pBI-11 DNA generated stronger therapeutic responses for C57BL/6 mice bearing HPV16 E6/E7-expressing TC-1 tumors. The HPV16/18 antigen-specific T cell-mediated immune responses generated by pBI-11 DNA vaccination were further enhanced by boosting with tissue-antigen HPV vaccine (TA-HPV). Combination of the pBI-11 DNA and TA-HPV boost vaccination with PD-1 antibody blockade significantly improved the control of TC-1 tumors and extended the survival of the mice. Finally, repeat vaccination with clinical-grade pBI-11 with or without clinical-grade TA-HPV was well tolerated in vaccinated mice. These preclinical studies suggest that the pBI-11 DNA vaccine may be used with TA-HPV in a heterologous prime-boost strategy to enhance HPV 16/18 E6/E7-specific CD8+ T cell responses, either alone or in combination with immune checkpoint blockade, to control HPV16/18-associated tumors. Our data serve as an important foundation for future clinical translation. IMPORTANCE Persistent expression of high-risk human papillomavirus (HPV) E6 and E7 is an obligate driver for several human malignancies, including cervical cancer, wherein HPV16 and HPV18 are the most common types. PD-1 antibody immunotherapy helps a subset of cervical cancer patients, and its efficacy might be improved by combination with active vaccination against E6 and/or E7. For patients with HPV16+ cervical intraepithelial neoplasia grade 2/3 (CIN2/3), the precursor of cervical cancer, intramuscular vaccination with a DNA vaccine targeting HPV16 E7 and then a recombinant vaccinia virus expressing HPV16/18 E6-E7 fusion proteins (TA-HPV) was safe, and half of the patients cleared their lesions in a small study (NCT00788164). Here, we sought to improve upon this therapeutic approach by developing a new DNA vaccine that targets E6 and E7 of HPV16 and HPV18 for administration prior to a TA-HPV booster vaccination and for application against cervical cancer in combination with a PD-1-blocking antibody.

Design of a multi-epitope vaccine against cervical cancer using immunoinformatics approaches

“…Second, E6 and E7 proteins are essential for the initiation and maintenance of HPV-associated malignancies, cancer progression, and escape from the immune system 47 . Third, E6 and E7 proteins are viral antigens that are not subject to central tolerance by human immune systems 48 . Therefore, in the present study, the E6 and E7 oncoproteins from HPV16 were chosen as the target antigens for epitope prediction.…”

Development of a Spontaneous HPV16 E6/E7-Expressing Head and Neck Squamous Cell Carcinoma in HLA-A2 Transgenic Mice

“…3A ). Vaccinations with TA-HPV alone have previously failed to generate detectable HPV antigen-specific CD8 + T cell-mediated immune responses ( 16 ). As shown in Fig.…”

“…To establish the HPV16 E6 peptide (aa 72 to 80)-specific CD8 + T cell line, splenocytes from female C57BL/6 mice vaccinated with pcDNA3-CRT/HPV16 E6(R55K) DNA, followed by boost vaccination with TA-HPV by skin scarification at a dose of 5 × 10 4 PFU/mouse ( 16 ) 1 week later, were harvested 1 week after the last vaccination. The splenocytes were stimulated with irradiated, HPV16 E6 peptide (aa 72 to 80)-loaded TC-1 cells in the presence of murine IL-2 (20 U/mL) and were restimulated once a week.…”

Therapeutic cancer vaccines: From biological mechanisms and engineering to ongoing clinical trials

“…Therefore, other HPV vaccination strategies have begun evaluation. DNA vaccination targeting HPV proteins E6 and E7, both associated with HPV 16 and 18, results in poor immunogenicity, possibly because the virus evades host recognition [26] . To overcome this obstacle, DNA vaccination is combined with other immunotherapies.…”