Cytotoxicity of oxidative metabolites of procainamide

Indirect evidence suggests that metabolism of procainamide (PA) may be necessary for generation of its autoimmunity-inducing capacity. Reactive metabolites of PA produced by hepatic mixed function oxidases have been identified by their capacity to bind proteins covalently. The present study extends these findings by comparing the toxicity of various chemically synthesized metabolites of PA to a variety of cell lines and primary cultures. PA and its N-acetyl- and N-acetyl-N-oxide-derivatives were nontoxic to S49.1 cells at concentrations up to 5 mM, whereas nitro-Pa (4-nitro-N-(diethylaminoethyl)benzamide hydrochloride) and azoxy-PA (bis-N,N'-(diethylaminoethyl)-4,4'-azoxydibenzamide) displayed partial cytotoxicity at 1 and 0.1 mM, respectively. (The therapeutic range of PA is 0.02-0.05 mM.). In contrast, hydroxylamine-PA (4-hydroxylamino-N-(diethylaminoethyl)benzamide hydrochloride) was highly cytotoxic, producing a TD50 between 0.002 and 0.045 mM among the eight cell lines tested. Hydroxylamine-PA sensitivity correlated with cell reducing capacity, suggesting that redox cycling contributes to cytotoxicity. The hydroxylamine metabolite of PA undergoes spontaneous air oxidation to the nitroso-derivative (nitroso-PA). When this oxidation was minimized by rapid manipulations or inhibited by ascorbic acid, cytotoxicity was reduced or eliminated, suggesting that the nitroso-derivative may be the toxic metabolite. Quiescent peripheral blood lymphocytes, especially T-cells, were relatively insensitive to hydroxylamine-PA when evaluated by dye exclusion 1 day after drug exposure. However, massive DNA strand breaks were detectable in these cells immediately after drug exposure, and resting lymphocytes treated with 0.01 mM hydroxylamine-PA died slowly during the subsequent week.(ABSTRACT TRUNCATED AT 250 WORDS)