Characterization of a Novel Human Tumor Necrosis Factor‐α Mutant with Increased Cytotoxic Activity

• Published in: Cancer science 1995-01, Vol.86 (1), p.72-80
• Main Authors: Masegi, Tsukio, Kato, Arata, Kitai, Kazuo, Fukuoka, Masami, Ogawa, Hiroko, Ichikawa, Yataro, Nakamura, Satoshi, Watanabe, Naoki, Niitsu, Yoshiro
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Animal behavior; Antineoplastic agents; Base Sequence; Biological and medical sciences; Cell Death; Chemical Phenomena; Chemistry, Physical; Cross-Linking Reagents; Cross‐linking; Cytotoxic activity; Cytotoxicity; Fibroblasts; Humans; Immunotherapy; Lipoprotein Lipase - metabolism; Medical sciences; Molecular Sequence Data; Mutagenesis; Mutants; Pancreatic carcinoma; Pharmacology. Drug treatments; Plasmids; Protein engineering; Receptor binding; Receptors, Tumor Necrosis Factor - metabolism; Recombinant Proteins - metabolism; Recombinant Proteins - pharmacology; Structure-Activity Relationship; TNF mutant; Trimers; Tumor Cells, Cultured; Tumor necrosis factor; Tumor necrosis factor receptors; Tumor Necrosis Factor-alpha - chemistry; Tumor Necrosis Factor-alpha - genetics; Tumor Necrosis Factor-alpha - pharmacology; Tumor necrosis factor-TNF
• ISSN: 0910-5050; 1347-9032; 1349-7006; 1876-4673
• DOI: 10.1111/j.1349-7006.1995.tb02990.x

Various novel recombinant human tumor necrosis factor‐α (TNF) mutants were prepared using protein engineering techniques, and their cytotoxic activity was compared with that of the intact form of TNF (intact TNF). Mutant 471 (a TNF mutant molecule with the deletion of 7 amino acids at the amino‐terminal and the substitution of Pro8Ser9Asp10 by ArgLysArg) had a 6‐fold higher cytotoxic activity against murine L929 cells. The mutant TNF had an increased ability to bind to TNF receptor on murine L929 fibroblasts cells. A cross‐linking study revealed that mutant 471 had an increased ability to form an active trimer. Mutant 471 also showed higher cytotoxic activity against human KYM myosarcoma cells and human MIA PaCa‐2 pancreatic carcinoma cells. The possible cachectin activity of the mutant was almost the same as that of intact TNF. These results suggest that mutant 471 might be a more promising candidate as an anticancer agent than intact TNF.