A divalent immunotoxin formed by the disulfide bond between hinge regions of fab domain

SeongHyeok Choi, JiEun Kim, YongChan Lee, Young J. Jang, Ira Pastan, MuHyeon Choe

Research output: Contribution to journalArticle

7 Citations (Scopus)

Abstract

Recombinant immunotoxins are hybrid cytotoxic proteins designed to selectively kill cancer cells. A divalent immunotoxins, [B3(FabH1)-PE38]2, was constructed by recombining Fab domain of B3 antibody as a celltargeting domain and Pseudomonas exotoxin A (PE) as a cytotoxic domain. Monoclonal antibody, B3, is the murine antibody (IgG1 κ) directed against LewisY -related carbohydrate antigens, which are abundant on the surface of many carcinomas. Fab fragment of this antibody was used in this study with the modified hinge sequence where last two cysteines out of three were mutated to serine. PE is a 66 kDa bacterial toxin that kills eukaryotic cells by inhibiting protein synthesis with ADP ribosylation of ribosomal elongation factor 2 (EF2). Fc region of B3 antibody was substituted with the truncated form of PE (38 kDa, PE38) on DNA level. [B3(FabH1)-PE38)2 was formed by disulfide bond between cysteines in the modified hinge region of B3(FabH1)-PE38. Each polypeptide for recombinant immunotoxins was overexpressed in Escherichia coli and collected as inclusion bodies. Each inclusion body was solubilized and refolded, and cytotoxic effects were measured. Divalent immunotoxins, [B3(FabH1)-PE38]2, had ID50 values of about 10 ng/mL on A431 cell lines and about 4 ng/mL on CRL1739 cell lines. Control immunotoxins, B3(scFv)-PE40, had ID50 values of about 28 ng/mL on A431 cell lines and about 41 ng/mL on CRL1739 cell lines. Divalent immunotoxins, [B3(FabH1)-PE38]2, had higher cytotoxic effects than B3(scFv)-PE40 control immunotoxins.

Original languageEnglish
Pages (from-to)1361-1365
Number of pages5
JournalBulletin of the Korean Chemical Society
Volume22
Issue number12
Publication statusPublished - 2001 Dec 20

Fingerprint

Immunotoxins
Hinges
Disulfides
Cells
Antibodies
Cysteine
Peptide Elongation Factor 2
Bacterial Toxins
Immunoglobulin Fab Fragments
Adenosine Diphosphate
Serine
Escherichia coli
Proteins
Immunoglobulin G
Monoclonal Antibodies
Carbohydrates
Antigens
Peptides
DNA

Keywords

  • [B3(FabH1)-PE38]
  • B3 antibody
  • Divalent immunotoxin
  • Pseudomonas exotoxin A, Antibody refolding

ASJC Scopus subject areas

  • Chemistry(all)

Cite this

A divalent immunotoxin formed by the disulfide bond between hinge regions of fab domain. / Choi, SeongHyeok; Kim, JiEun; Lee, YongChan; Jang, Young J.; Pastan, Ira; Choe, MuHyeon.

In: Bulletin of the Korean Chemical Society, Vol. 22, No. 12, 20.12.2001, p. 1361-1365.

Research output: Contribution to journalArticle

Choi, SeongHyeok ; Kim, JiEun ; Lee, YongChan ; Jang, Young J. ; Pastan, Ira ; Choe, MuHyeon. / A divalent immunotoxin formed by the disulfide bond between hinge regions of fab domain. In: Bulletin of the Korean Chemical Society. 2001 ; Vol. 22, No. 12. pp. 1361-1365.
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