Design, Characterization, and Evaluation of scFvCD133/rGelonin: A CD133-Targeting Recombinant Immunotoxin for Use in Combination with Photochemical Internalization

Journal of Clinical Medicine

卷:9

MichaelG. Rosenblum¹  LawrenceH. Cheung¹  PålKristian Selbo²  Anette Weyergang²  CathrineElisabeth Olsen²  Kristian Berg²  DanielA. Vallera³ 

[1] Department of Experimental Therapeutics, The University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA;

[2] Department of Radiation Biology, Institute for Cancer Research, Oslo University Hospital, The Norwegian Radium Hospital, N-0310 Oslo, Norway;

[3] Department of Therapeutic Radiology-Radiation Oncology, University of Minnesota, Masonic Cancer Center, Minneapolis, MN 55455, USA;

关键词: cancer stem cells;    cd133;    prominin-1;    photochemical internalization;    photodynamic therapy;    immunotoxins;    ribosome-inactivating proteins;    drug delivery;   

DOI  :  10.3390/jcm9010068

来源: DOAJ

【 摘 要 】

The objective of this study was to develop and explore a novel CD133-targeting immunotoxin (IT) for use in combination with the endosomal escape method photochemical internalization (PCI). scFvCD133/rGelonin was recombinantly constructed by fusing a gene (scFvCD133) encoding the scFv that targets both non-glycosylated and glycosylated forms of both human and murine CD133/prominin-1 to a gene encoding the ribosome-inactivating protein (RIP) gelonin (rGelonin). RIP-activity was assessed in a cell-free translation assay. Selective binding and intracellular accumulation of scFvCD133/rGelonin was evaluated by flow cytometry and fluorescence microscopy. PCI of scFvCD133/rGelonin was explored in CD133^high and CD133^low cell lines and a CD133^neg cell line, where cytotoxicity was evaluated by the MTT assay. scFvCD133/rGelonin exhibited superior binding to and a higher accumulation in CD133^high cells compared to CD133^low cells. No cytotoxic responses were detected in either CD133^high or CD133^low cells after 72 h incubation with <100 nM scFvCD133/rGelonin. Despite a severe loss in RIP-activity of scFvCD133/rGelonin compared to free rGelonin, PCI of scFvCD133/rGelonin induced log-fold reduction of viability compared to PCI of rGelonin. Strikingly, PCI of scFvCD133/rGelonin exceeded the cytotoxicity of PCI of rGelonin also in CD133^low cells. In conclusion, PCI promotes strong cytotoxic activity of the per se non-toxic scFvCD133/rGelonin in both CD133^high and CD133^low cancer cells.

【 授权许可】

Unknown