title: Enhanced generation of cytotoxic T lymphocytes by increased cytosolic delivery of MHC class I epitope fused to mouse heat shock protein 70 via polyhistidine conjugation.
abstract: Heat shock protein 70 (Hsp70)-associated antigens in a soluble form have been shown to elicit strong antigen-specific cytotoxic T lymphocyte (CTL) responses following immunization without any adjuvants. In order to improve the potential of Hsp70, we genetically designed a novel Hsp70-based antigen delivery system, in which the model MHC class I epitope of ovalbumin (OVA) (SIINFEKL; OVA257-264) was fused to mouse Hsp70. To facilitate the cytosolic delivery of the peptide following Hsp receptor-mediated endocytosis, polyhistidine of 25 or 50 residues was further fused to the fusion protein. Each fusion protein was then expressed in E. coli and purified. When added to DC2.4 cells, a mouse dendritic cell line, the fusion protein containing polyhistidine of 25 residues was efficiently taken up by the cells and efficiently distributed to the cytosol. The fusion protein also exhibited a significantly improved efficacy of MHC class I-restricted presentation of antigen. Vaccination of mice with the polyhistidine fusion protein generated strong antigen-specific CTL responses and antitumor activity. These findings suggest that polyhistidine fusion is a useful strategy to increase the potential of Hsp-based vaccination.
authors.affiliation: Department of Biopharmaceutics and Drug Metabolism, Graduate School of Pharmaceutical Science, Kyoto University, Sakyo-ku, Kyoto, Sakyo-ku, Japan.
authors.forename: Seiji
authors.initials: S
authors.lastname: Takemoto
authors.forename: Makiya
authors.initials: M
authors.lastname: Nishikawa
authors.forename: Takayuki
authors.initials: T
authors.lastname: Otsuki
authors.forename: Ayumi
authors.initials: A
authors.lastname: Yamaoka
authors.forename: Kazuki
authors.initials: K
authors.lastname: Maeda
authors.forename: Atsushi
authors.initials: A
authors.lastname: Ota
authors.forename: Yoshinobu
authors.initials: Y
authors.lastname: Takakura
mesh_terms: D000818:Animals; D017951:Antigen Presentation / Q000276:immunology; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D003600:Cytosol / Q000187:drug effects* / Q000276:immunology; D003713:Dendritic Cells / Q000187:drug effects / Q000276:immunology; D016503:Drug Delivery Systems; D000939:Epitopes / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D005260:Female; D050883:HSC70 Heat-Shock Proteins / Q000008:administration & dosage / Q000737:chemistry* / Q000276:immunology / Q000494:pharmacology; D006639:Histidine / Q000008:administration & dosage / Q000737:chemistry* / Q000276:immunology / Q000494:pharmacology; D015395:Histocompatibility Antigens Class I / Q000008:administration & dosage / Q000737:chemistry* / Q000276:immunology / Q000494:pharmacology; D007376:Interleukin-2 / Q000276:immunology; D051379:Mice; D008810:Mice, Inbred C57BL; D009368:Neoplasm Transplantation; D009369:Neoplasms / Q000188:drug therapy / Q000276:immunology; D010047:Ovalbumin / Q000276:immunology; D011993:Recombinant Fusion Proteins / Q000008:administration & dosage / Q000737:chemistry* / Q000276:immunology / Q000494:pharmacology; D013602:T-Lymphocytes, Cytotoxic / Q000187:drug effects* / Q000276:immunology
chemical_list: D000939:Epitopes; D050883:HSC70 Heat-Shock Proteins; D015395:Histocompatibility Antigens Class I; C494372:Hspa8 protein, mouse; D007376:Interleukin-2; D011993:Recombinant Fusion Proteins; C033223:polyhistidine; D006639:Histidine; D010047:Ovalbumin