Methylglyoxal bis(guanylhydrazone) (MGBG) and alpha-difluoromethylornithine (DFMO) have anti-neoplastic effects on many tissues, plus antomicrobial chemotherapeutic actions. Both drugs block polyamine biosynthesis in the putrescine --> spermidine --> spermine pathway. DFMO irreversibly inhibits ornithine decarboxylase, which converts ornithine --> putrescine. MGBG functions primarily as a spermidine analog and inhibits the S-adenosylmethionine decarboxylase required for the conversion of spermidine --> spermine. The polyamines (spermine, spermidine, putrescine) function in the cell to stabilize nucleic acids and are essential for cell viability. Recent studies implicate polyamines in control of exocytosis in some cells. The main hypotheses of the grant are that 1] polyamines are required for exocytosis of norepinephrine (NE) from adrenal medulla; 2] DFMO and MGBG interfere with exocytosis; and 3] DFMO and MGBG can be used to understand the role of polyamines in regulated exocytosis, synthesis, and uptake of catecholamines in PC12 cells (cultured adrenal medullary pheochromocytoma). [3H]NE uptake will be used as a marker for catecholamine uptake and release. Exocytosis will be stimulated by KCl depolarization, plus nicotinic, muscarinic and mixed cholinergic agonists. Digitonin-permeabilized cells will be used to examine the role of Ca++ in release. High performance liquid chromatography (HPLC) will be used to quantify endogenous polyamines, NE, dopamine (DA) and adenine nucleotides from extracted PC12 cells, after treatments with cytostatic and cytotoxic doses of one or both drugs. The data will increase the understanding of the use of these drugs as antineoplastic and antimicrobial (anti-trypanosomes) agents.
