title: Potent killing of HBV-related hepatocellular carcinoma by a chimeric protein of anti-HBsAg single-chain antibody and truncated Bid.
abstract: Targeted therapy is needed for hepatitis B virus (HBV)-mediated hepatocellular carcinoma (HCC) which shows overexpression of HBV surface antigen (HBsAg). We previously developed scFv15, a human single-chain antibody against HBsAg. Here we tested the strategic feasibility of scFv15-mediated delivery of apoptotic effectors for HBsAg-targeted HCC therapy and application of HA2 motif of influenza hemagglutinin to enhance endosome escape and antitumor effect. A class of HBsAg-targeted immunoproapoptotic molecule was generated by sequentially fusing scFv15, the furin-cleavable motif from diphtheria toxin (Fdt), HA2 and a truncated apoptotic protein Bid (tBid). The resulting scFv15-Fdt-HA2-tBid was prokaryotically expressed and functionally characterized for HBsAg-binding capacity, endosome escape activity and antitumor effect as compared with scFv15-Fdt-tBid. Both scFv15-Fdt-HA2-tBid and scFv15-Fdt-tBid retained affinity and specificity for HBsAg, and bound and selectively killed HBsAg-positive HCC cells via apoptosis. Notably, the IC50 of scFv15-Fdt-HA2-tBid in HBsAg-positive PLC/PRF/5 cells was 10 times lower than that of scFv15-Fdt-tBid. In vivo imaging of antitumor activity demonstrated 95% growth inhibition of orthotopic HCC by scFv15-Fdt-HA2-tBid compared with 75% suppression by scFv15-Fdt-tBid. This study represents an extended application of the immunoproapoptotic strategy in the treatment of HBsAg-positive HCC and shows significant potential of HA2 as a functional enhancer for endosome-encapsulated antibody-conjugates.
authors.affiliation: The State Key Laboratory of Cancer Biology, Department of Biochemistry and Molecular Biology, Fourth Military Medical University, Xi'an 710032, China.
authors.forename: Bo
authors.initials: B
authors.lastname: Yan
authors.forename: Qing
authors.initials: Q
authors.lastname: Ouyang
authors.forename: Zhining
authors.initials: Z
authors.lastname: Zhao
authors.forename: Feng
authors.initials: F
authors.lastname: Cao
authors.forename: Tao
authors.initials: T
authors.lastname: Wang
authors.forename: Xiaofei
authors.initials: X
authors.lastname: Jia
authors.forename: Yanling
authors.initials: Y
authors.lastname: Meng
authors.forename: Shuai
authors.initials: S
authors.lastname: Jiang
authors.forename: Jiayun
authors.initials: J
authors.lastname: Liu
authors.forename: Rui
authors.initials: R
authors.lastname: Chen
authors.forename: Lintao
authors.initials: L
authors.lastname: Jia
authors.forename: Rui
authors.initials: R
authors.lastname: Zhang
authors.forename: Weihong
authors.initials: W
authors.lastname: Wen
authors.forename: Boquan
authors.initials: B
authors.lastname: Jin
authors.forename: Siyi
authors.initials: S
authors.lastname: Chen
authors.forename: Jing
authors.initials: J
authors.lastname: Zhao
authors.forename: Angang
authors.initials: A
authors.lastname: Yang
mesh_terms: D000368:Aged; D000595:Amino Acid Sequence; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D051023:BH3 Interacting Domain Death Agonist Protein / Q000378:metabolism*; D006528:Carcinoma, Hepatocellular / Q000188:drug therapy* / Q000473:pathology / Q000821:virology; D045744:Cell Line, Tumor; D011992:Endosomes / Q000187:drug effects / Q000378:metabolism; D005260:Female; D019793:Fluorescein / Q000378:metabolism; D006514:Hepatitis B Surface Antigens / Q000276:immunology*; D006515:Hepatitis B virus / Q000187:drug effects / Q000502:physiology*; D006801:Humans; D008113:Liver Neoplasms / Q000188:drug therapy* / Q000473:pathology / Q000821:virology; D008297:Male; D051379:Mice; D008875:Middle Aged; D008969:Molecular Sequence Data; D011485:Protein Binding / Q000187:drug effects; D021381:Protein Transport / Q000187:drug effects; D011993:Recombinant Fusion Proteins / Q000494:pharmacology / Q000627:therapeutic use*; D057127:Single-Chain Antibodies / Q000494:pharmacology / Q000627:therapeutic use*
chemical_list: D051023:BH3 Interacting Domain Death Agonist Protein; D006514:Hepatitis B Surface Antigens; D011993:Recombinant Fusion Proteins; D057127:Single-Chain Antibodies; D019793:Fluorescein