Long-Term Protection Against Human Papillomavirus E7-Positive Tumor by a Single Vaccination of Adeno-Associated Virus Vectors Encoding a Fusion Protein of Inactivated E7 of Human Papillomavirus 16/18 and Heat Shock Protein 70

We investigated a gene vaccine strategy against human papillomavirus (HPV)-induced cancer and premalignant diseases, using adeno-associated virus (AAV) vector encoding the viral E7 oncoproteins as the tumor antigens from HPV serotypes 16 (HPV16) and 18 (HPV18). Genetically inactivated E7 proteins were fused with a heat shock protein 70 (hsp70) to minimize the risk of cell transformation and enhance immune responses. The fusion protein gene was packaged in AAV serotype 1 or 2 (AAV1 or 2) for efficient in vivo gene expression. Our results showed that after a single intramuscular injection, the AAV1 vector elicited stronger HPV-specific cytotoxic T lymphocyte (CTL) responses and interferon-g secretion when compared with the AAV2 vector. Prophylactic immunization with AAV1 protected 100% of the mice from tumor growth for more than 1 year, whereas all the control mice immunized with either a LacZ vector or saline grew large tumors and died within 6 weeks after inoculation of E7-positive tumor cell line TC-1. In addition, this single-dose AAV1 vaccination completely protected the mice against second and third challenges with higher numbers of TC-1 cells. Despite lower CTL responses against the E7 antigens, AAV2 vector prophylactic immunization was also sufficient to protect 100% of the mice against the initial and second tumor challenges and 70% of the mice against the third challenge. In addition, therapeutic immunization with AAV1 after palpable tumor formation inhibited tumor growth and caused tumor regression in some mice. Thus, our studies support the potential of AAV vectors as a genetic vaccine for the prevention and treatment of HPV-induced malignancies.