1. The effect of direct translocation across endosomes on the cytotoxicity of the recombinant protein e23sFv-Fdt-casp6 to HER2 positive gastric cancer cells
- Author
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Ren, Jun-Lin, Meng, Yan-Ling, Hu, Bin, Jia, Lin-Tao, Zhang, Rui, Xu, Yan-Ming, Xie, Qiao-Sheng, Zhang, Ying-Qi, Jin, Bo-Quan, Chen, Si-Yi, Wang, Tao, and Yang, An-Gang
- Subjects
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ENDOSOMES , *RECOMBINANT proteins , *CANCER cells , *HER2 protein , *METASTASIS , *PROGNOSIS , *PSEUDOMONAS , *APOPTOSIS - Abstract
Abstract: 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. [Copyright &y& Elsevier]
- Published
- 2011
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