title: Transfection of complementary DNAs for the heavy and light subunits of human gamma-glutamylcysteine synthetase results in an elevation of intracellular glutathione and resistance to melphalan.
abstract: Although glutathione (GSH) has long been implicated in resistance to certain common chemotherapeutic agents, including alkylating agents, platinum analogues, and doxorubicin, evidence establishing a direct role in the resistant phenotype has been lacking. We cotransfected COS cells with the cDNAs for the two subunits of gamma-glutamylcysteine synthetase (GCS), which catalyzes the rate-limiting step in the de novo synthesis of GSH and is itself up-regulated in some drug-resistant tumor cells. Transfection resulted in increased GCS activity and elevated GSH levels (up to 2.6-fold). Cotransfection with the two subunits greatly enhanced the synthetic efficiency of the heavy subunit. A direct correlation (P < 0.01) between intracellular GSH levels and the LD99 dose of melphalan was observed, signifying that elevation of the thiol secondary to GCS expression is sufficient to confer the resistance phenotype.
authors.affiliation: Department of Human Oncology, University of Wisconsin Medical School, Madison 53792, USA.
authors.forename: R T
authors.initials: RT
authors.lastname: Mulcahy
authors.forename: H H
authors.initials: HH
authors.lastname: Bailey
authors.forename: J J
authors.initials: JJ
authors.lastname: Gipp
mesh_terms: D000818:Animals; D018906:Antineoplastic Agents, Alkylating / Q000494:pharmacology*; D002384:Catalysis; D002460:Cell Line; D018076:DNA, Complementary / Q000235:genetics*; D019008:Drug Resistance, Neoplasm / Q000502:physiology; D005721:Glutamate-Cysteine Ligase / Q000235:genetics*; D005978:Glutathione / Q000096:biosynthesis*; D006801:Humans; D007424:Intracellular Fluid / Q000378:metabolism; D007668:Kidney / Q000166:cytology; D046911:Macromolecular Substances; D008558:Melphalan / Q000494:pharmacology*; D014162:Transfection
chemical_list: D018906:Antineoplastic Agents, Alkylating; D018076:DNA, Complementary; D046911:Macromolecular Substances; D005721:Glutamate-Cysteine Ligase; D005978:Glutathione; D008558:Melphalan