Cytotoxic activity of a recombinant GnRH-PAP fusion toxin on human tumor cell lines.
Pokeweed antiviral protein (PAP), a ribosome-inactivating protein isolated from the leaves of Phytolacca americana, reveals potent antiviral activity against viruses or cytotoxic action against cells once inside the cytoplasm. Therefore PAP is a good candidate to be used as an immunotoxin. We constructed a bacterial expression plasmid encoding PAP as a fusion protein with gonadotropin-releasing hormone (GnRH), a neuropeptide with receptor sites on several gynaecologic tumors. The resulting recombinant toxin was produced in Escherichia coli and accumulated in inclusion bodies. After purification under denaturing conditions, renaturated GnRH-PAP shows an IC(50) of 3 nM on in vitro translation assays and selectively inhibits the growth of the GnRH receptor positive Ishikawa cell line (ID(50) of 15 nM); on the other hand, neither GnRH nor PAP alone had any effect.
Schlick
J
J
Laboratoire de Biochimie et Biologie Moléculaire, Université de Franche-Comté, 16 route de Gray, 25030 Besançon, France. jlsk3@netcourrier.com
Dulieu
P
P
Desvoyes
B
B
Adami
P
P
Radom
J
J
Jouvenot
M
M
D000970:Antineoplastic Agents / Q000494:pharmacology*; D004354:Drug Screening Assays, Antitumor; D015870:Gene Expression; D007987:Gonadotropin-Releasing Hormone / Q000235:genetics / Q000494:pharmacology*; D006801:Humans; D000922:Immunotoxins / Q000235:genetics / Q000302:isolation & purification / Q000494:pharmacology*; D009699:N-Glycosyl Hydrolases; D010940:Plant Proteins / Q000235:genetics / Q000494:pharmacology*; D011966:Receptors, LHRH / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000302:isolation & purification / Q000494:pharmacology; D054789:Ribosome Inactivating Proteins, Type 1; D014407:Tumor Cells, Cultured
D000970:Antineoplastic Agents; D000922:Immunotoxins; D010940:Plant Proteins; D011966:Receptors, LHRH; D011993:Recombinant Fusion Proteins; D054789:Ribosome Inactivating Proteins, Type 1; D007987:Gonadotropin-Releasing Hormone; D009699:N-Glycosyl Hydrolases; C024275:pokeweed antiviral protein