Construction and characterization of an attenuated recombinant Listeria monocytogenes vector vaccine delivering HPV16 E7 / 生物工程学报

ABSTRACT

Listeria monocytogenes (L. monocytogenes, LM) is an excellent tumor vaccine vector. In this study, recombinant LM vaccine candidate expressing human papillomavirus type 16 (HPV16) E7 protein was constructed and its charactericts were determined. Through homologous recombination, E7 gene was cloned in frame with the LM4 Phly promoter-signal sequence, and introduced into the chromosome of LM4. The recombinant strain named LM4△hly::E7 with the plasmid-free and antibiotic-resistant gene-free was constructed. LM4△hly::E7 could express and secrete E7-LLO fusion protein; its size is 66 kDa and has immunological activity. Furthermore, LM4△hly::E7 could multiply in RAW264.7 macrophages by confocal laser scanning microscope. Additionally, LM4△hly::E7 could induce specific antibodies against E7 in immunized mice in ELISA. Also, the 50% lethal dose (LD₅₀) of LM4△hly::E7 strain was 3.863×10⁹ CFU (Colony-Forming Units) in C57BL/6 mice with intraperitoneal immunization, which was more attenuated than wild type LM4. Mice immunized with LM4△hly::E7 did not show obvious pathological change. These data show that LM4△hly::E7 expressing E7-LLO fusion protein has good safety, which may provide the materials for research of antitumor effect and would be a promising vaccine candidate for cervical cancer.