The effect of direct translocation across endosomes on the cytotoxicity of the recombinant protein e23sFv-Fdt-casp6 to HER2 positive gastric cancer cells

Biomaterials. 2011 Oct;32(30):7641-50. doi: 10.1016/j.biomaterials.2011.06.071. Epub 2011 Jul 20.


HER2-positive cancers represent a class of malignancies with high metastasis and poor prognosis. We previously generated the e23sFv-PEA II-casp6 recombinant, which contains an anti-HER2 single-chain antibody (e23sFv), a Pseudomonas exotoxin A translocation domain (PEA II), and a constitutively active caspase-6 (casp6), and demonstrated its potent selective anti-tumor activities. In this study, we generated a smaller-sized recombinant e23sFv-Fdt-casp6, in which the PEA II domain was replaced with the furin cleavage sequence from diphtheria toxin (Fdt), and explored its translocation pathway and specific killing mechanism. We found that e23sFv-Fdt-casp6 proteins, following their receptor-mediated endocytosis in HER2-positive gastric cancer cells, underwent furin-mediated cleavage in endosome and engaged in direct translocation of the released C-terminal fragment (active caspase-6) instead of via the trans-Golgi and the endoplasmic reticulum (ER) pathway. The active caspase-6 cleaved its well-documented substrate, Lamin A, and subsequently triggered the apoptosis of cancer cells. The e23sFv-Fdt-casp6 proteins produced from genetically modified cells showed a selective cytotoxicity to cultured HER2-positive gastric cancer cells. Similar to the results of our previous research on e23sFv-PEA II-casp6, the delivery of liposome-encapsulated e23sFv-Fdt-casp6 constructs in tumor-adjacent muscles also inhibited tumor growth and prolonged animal survival in a nude mouse xenograft tumor model. Moreover, e23sFv-Fdt-casp6 proteins were also cytotoxic to trastuzumab-resistant gastric cancer cells characterized by downregulated HER2 expression. Accordingly, e23sFv-Fdt-casp6 recombinant provides a promising therapeutic alternative for HER2-positive and trastuzumab-resistant gastric cancers.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antibodies / genetics
  • Antibodies / immunology
  • Antibodies / therapeutic use*
  • Apoptosis / drug effects
  • Caspase 6 / genetics
  • Caspase 6 / therapeutic use*
  • Cell Line, Tumor
  • Cytosol / metabolism
  • Diphtheria Toxin / genetics
  • Diphtheria Toxin / therapeutic use*
  • Endosomes / metabolism*
  • Furin / metabolism
  • Humans
  • Lamin Type A / metabolism
  • Liposomes
  • Mice
  • Mice, Nude
  • Protein Transport
  • Receptor, ErbB-2 / immunology
  • Receptor, ErbB-2 / metabolism*
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / therapeutic use*
  • Stomach Neoplasms / drug therapy*
  • Stomach Neoplasms / metabolism


  • Antibodies
  • Diphtheria Toxin
  • Lamin Type A
  • Liposomes
  • Recombinant Fusion Proteins
  • Receptor, ErbB-2
  • Furin
  • Caspase 6