Efeitos da eletroporação <i>in vivo</i> na resposta imunológica induzida por uma vacina de DNA contra tumores induzidos por HPV-16.

Abstract

SALES, N. S. Effects of the in vivo electroporation in the induced immune response by DNA vaccines against induced tumors by HPV-16. 2015. 63 p. Masters thesis (Microbiology) -Instituto de Ciências Biomédicas, Universidade de São Paulo, São Paulo, 2015.Cervical cancer is the third leading cause of cancer death among women in the world and the fourth cause of cancer death in women in Brazil. It's main etiological agent is human papilloma virus (HPV). More than 100 types of this virus have been identified and about 40 of them infect anogenital mucosal tissues. Among them, they are classified as high or low risk viruses according to their oncogenic potential. Among the high-risk virus types, HPV-16 and HPV-18 are considered the main cause of cervical cancer, being responsible for around 85% of the cases. Efforts are being made in the search for immunotherapeutic strategies against HPV-induced tumors, since the prophylactic vaccines now commercialized, are not able to benefit already infected individuals or patients with established cancer. In this context, DNA vaccines have demonstrated good efficacy against these tumors in murine studies, and are considered safe, stable and capable of inducing specific immune response. However, these formulations have shown immunogenicity in humans, requiring the use of approaches that increase the efficacy of these vaccines, such as the use of adjuvants, codon optimization and the use of different plasmid delivery systems. The in vivo electroporation (EP) is a method for delivering DNA vaccines, consisting on the application of electric pulses that open transient pores in the cell membrane and facilitate DNA entry into the cell. This technique has been extensively studied for its ability to potentiate the effect of DNA vaccines. Our group has developed a DNA vaccine expressing the E7 protein of HPV-16 fused to glycoprotein D (gD) of herpes simplex virus type 1 (HSV-1) (pgDE7h). The use of EP for the delivery of 50 µg of the vaccine by the intramuscular route (im) increased the therapeutic effect in mice challenged with the TC-1 tumor cell line that expresses the HPV-16 E6 and E7 proteins, generating full anti-tumor protection. The vaccine combined to EP induced increased cell transfection by plasmid DNA and the migration of pro-inflammatory cells to the inoculation site. It was also observed an enhanced activation of CD8 + E7-specific cells, with greater cytotoxic ability and capacity of secreting IFN-γ, IL-2 and TNF-α simultaneously. It was also observed an increased frequency of CD8 + T cells with greater avidity and effector memory phenotype in animals subjected to the electroporation immunization system. The use of EP in immunizations through the intradermal (id) route was not able to improve the vaccine effectiveness. In conclusion, the present results consolidated the EP as a powerful technique to potentiate the therapeutic effect induced by the pgDE7h vaccine administered by intramuscular injection reinforcing its potential for use in future clinical trials. APC -Antigen pr...