MECHANISM FOR THE ANTITHYROID ACTION OF MINOCYCLINE

Administration of minocycline (MN), a tetracycline antibiotic, produce s a black pigment in the thyroids of humans and several species of exp erimental animals and antithyroid effects in rodents. We have previous ly shown that these effects appear to be related to interactions of MN with thyroid peroxidase (TPO), the key enzyme in thyroid hormone synt hesis. In the present study, the mechanisms for inhibition of TPO-cata lyzed iodination and coupling reactions by MN were investigated. MN wa s stable in the presence of TPO and H2O2, but adding iodide or a pheno lic cosubstrate caused rapid conversion to several products. TPO-depen dent product formation, characterized by on-line LC-APCI/MS and H-1-NM R, involved oxidative elimination to form the corresponding benzoquino ne with subsequent dehydrogenation at the aliphatic 4-(dimethylamino) group. Addition of thiol-containing polymers (bovine serum albumin or thiol-agarose chromatographic beads) had a minimal effect on MN oxidat ion by TPO, but substantially reduced product formation and produced c oncomitant losses in free thiols. Covalent bonding through a thioether linkage of a reactive intermediate, the benzoquinone iminium ion, was inferred from these findings. Iodide- and phenolic cosubstrate-depend ent oxidation of tetracycline to demethylated and dehydrogenated produ cts was also observed, although at a slower rate than MN. The products and kinetics observed with MN were consistent with oxidation of MN by either the enzymatic iodinating species formed by reaction of TPO com pound I with iodide or phenoxyl radicals/cations generated by TPO-medi ated oxidation of a phenolic cosubstrate. The proposed reaction mechan ism is consistent with alternate substrate inhibition of TPO-catalyzed iodination of tyrosyl residues in thyroglobulin (Tg) by MN, as previo usly reported. Furthermore, the observed phenoxyl radical-mediated oxi dation of MN is consistent with its previously reported potent inhibit ion of the coupling of hormonogenic iodotyrosine residues in Tg in the reaction that forms thyroid hormones. The proposed reaction mechanism also implicates a reactive benzoquinone iminium ion intermediate that could be important in toxicity of MN.