Construction and characterization of the recombinant immunotoxin RTA-4D5-KDEL targeting HER2/neu-positive cancer cells and locating the endoplasmic reticulum

Applied Microbiology and Biotechnology
Pengyan JiaoYuhong Ren

Abstract

The specific targeting of immunotoxins enables their wide application in cancer therapy. The A-chain of the ricin protein (RTA) is an N-glycosidase that catalyzes the removal of adenine from the 28S rRNA, preventing protein translation and leading to cell death. Ricin is highly toxic but can only exert its toxic effects from within the cytoplasm. In this study, we linked the anti-HER2 single-chain variable fragment 4D5 scFv and the endoplasmic reticulum-targeting peptide KDEL to the C-terminal of the RTA to construct immunotoxin RTA-4D5-KDEL. In vitro experiments showed that the anticancer effect of RTA-4D5-KDEL towards ovarian cancer cells SKOV-3 increased 440-fold and 28-fold relative to RTA and RTA-4D5, respectively. RTA-4D5-KDEL had a strong inhibitory effect on HER2-overexpressing SKOV-3 cells and caused little damage to normal HEK-293 cells and H460 lung cancer cells. Immunofluorescence experiments showed that the immunotoxin RTA-4D5 could specifically bind to SKOV-3 cells, but not to normal cells HEK-293. The immunotoxin RTA-4D5-KDEL could rapidly localize the recombinant protein to the endoplasmic reticulum. These results suggest that the recombinant immunotoxin RTA-4D5-KDEL has a strong inhibitory effect on ovarian can...Continue Reading

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Citations

Jun 21, 2019·Toxins·Natalia Sowa-RogozińskaMonika Słomińska-Wojewódzka
Dec 4, 2019·Cancer Letters·Maria J PiñaF Javier Arias
Sep 8, 2021·Cancer Cell International·Reza MahmoudiMohammad Rahmati

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Methods Mentioned

BETA
PCR
flow cytometry
confocal microscopy
flow
confocal

Software Mentioned

Flow Jo

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