title: Construction, expression, and activity of a novel immunotoxin comprising a humanized antiepidermal growth factor receptor scFv and modified Pseudomonas aeruginosa exotoxin A.
abstract: Overexpression of epidermal growth factor receptor (EGFR) plays a significant role in the development and metastasis of many solid tumors. Strategies based on anti-EGFR immunotoxins have shown promising results in several studies, but immunogenicity of antibody and toxin moieties is a limitation of this type of therapeutics. In the present study, a novel humanized anti-EGFR immunotoxin (huscFv-PE25) was developed by genetic fusing of a humanized anti-EGFR single-chain variable fragment (huscFv) with a modified Pseudomonas aeruginosa exotoxin A (PE25KDEL). The reactivity and toxicity of this immunotoxin with tumor cells were assessed by dot-blot, enzyme-linked immunosorbent assay, and MTT procedures. Results of enzyme-linked immunosorbent assay and dot-blot assay indicated that the immunotoxin recognizes and efficiently binds to EGFR-overexpressing tumor cells. MTT assay showed a specific growth-inhibitory effect of huscFv-PE25 on EGFR-overexpressing A431 cells, without any inhibitory effect on EGFR-negative cells. In conclusion, the results of this study indicated that huscFv-PE25 can recognize and exert an inhibitory effect on EGFR-overexpressing cancer cells, despite its smaller size and lower immunogenicity. This may provide a basis for the development of novel clinical therapeutic agents against EGFR-overexpressing tumor cells.
authors.affiliation: aDepartment of Medical Biotechnology, Faculty of Advanced Medical Sciences bDrug Applied Research Center cStudent Research Committee dBiotechnology Research Center eInfection and Tropical Disease Research Center, Tabriz University of Medical Sciences, Tabriz fCardiogenetic Research Laboratory, Rajaie Cardiovascular Medical and Research Center, Iran University of Medical Sciences, Tehran, Iran.
authors.forename: Bahman
authors.initials: B
authors.lastname: Akbari
authors.forename: Safar
authors.initials: S
authors.lastname: Farajnia
authors.forename: Nosratollah
authors.initials: N
authors.lastname: Zarghami
authors.forename: Nejat
authors.initials: N
authors.lastname: Mahdieh
authors.forename: Mohammad
authors.initials: M
authors.lastname: Rahmati
authors.forename: Shiva A
authors.initials: SA
authors.lastname: Khosroshahi
authors.forename: Abolfazl
authors.initials: A
authors.lastname: Barzegar
authors.forename: Leila
authors.initials: L
authors.lastname: Rahbarnia
mesh_terms: D000072283:A549 Cells; D036002:ADP Ribose Transferases / Q000737:chemistry / Q000494:pharmacology*; D000818:Animals; D001427:Bacterial Toxins / Q000737:chemistry / Q000494:pharmacology*; D016466:CHO Cells; D045744:Cell Line, Tumor; D000068818:Cetuximab / Q000494:pharmacology; D003412:Cricetulus; D004797:Enzyme-Linked Immunosorbent Assay; D066246:ErbB Receptors / Q000096:biosynthesis / Q000276:immunology*; D005098:Exotoxins / Q000737:chemistry / Q000494:pharmacology*; D007135:Immunoglobulin Variable Region / Q000737:chemistry / Q000276:immunology; D000922:Immunotoxins / Q000737:chemistry* / Q000276:immunology / Q000302:isolation & purification / Q000494:pharmacology*; D008175:Lung Neoplasms / Q000188:drug therapy* / Q000201:enzymology / Q000276:immunology; D057127:Single-Chain Antibodies / Q000737:chemistry / Q000276:immunology; D012878:Skin Neoplasms / Q000188:drug therapy* / Q000201:enzymology / Q000276:immunology; D037521:Virulence Factors / Q000737:chemistry / Q000494:pharmacology*; D000097668:Pseudomonas aeruginosa Exotoxin A
chemical_list: D036002:ADP Ribose Transferases; D001427:Bacterial Toxins; D000068818:Cetuximab; D066246:ErbB Receptors; D005098:Exotoxins; D007135:Immunoglobulin Variable Region; D000922:Immunotoxins; D057127:Single-Chain Antibodies; D037521:Virulence Factors; D000097668:Pseudomonas aeruginosa Exotoxin A; C512478:EGFR protein, human