Construction and characterization of attenuated recombinant Listeria monocytogenes delivering HPV16 E7
【Abstract】 Listeria monocytogenes (LM) is an excellent tumor vaccine vector. In this study, recombinant LM expressing human papillomavirus type 16 (HPV16) E7 protein, a vaccine candidate was constructed and the biological characteristics were determined. Through homologous recombination, E7 gene was cloned in frame with the LM4 hly promoter-signal sequence, and introduced into the LM4 genome. The recombinants, constructed as LM4△hly::E7, were plasmid- and antibiotic-resistant gene-free. LM4△hly::E7 could express and secrete a 66 kDa E7-LLO fusion protein, which had immunological activities. Proliferation of LM4△hly::E7 in RAW264.7 macrophages was observed under confocal laser scanning microscope. Specific anti-E7 antibodies could be detected by ELISA in mice immunized with LM4△hly::E7. The LD50 of LM4△hly::E7 was 3.863 × 109 CFU in C57BL/6 mice with intraperitoneal immunization, which was 104 times higher than that of wild type LM4. No obvious pathological changes were detected in the immunized mice. These results suggest that LM4△hly::E7 expressing E7-LLO fusion protein may be a safe and promising vector-vaccine candidate for cervical cancer, and may also be important biological materials for anti-tumor research.
【Keywords】 attenuated Listeria monocytogenes; HPV16 E7; macrophage cell line; safety;
(Translated by Chengji)
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