Cefazolin administration and 2-methyl-1,3,4-thiadiazole-5-thiol (MTD) in human tissue

Cephalosporin antibiotic administration can lead to the formation of heterocyclic thiol metabolites that have been associated with hypoprothrombinemia and hemorrhage. The cefazolin structure includes a heterocyclic thiol, MTD, that can inhibit the γ-carboxylation of glutamate required to produce active clotting factors. We set out to determine whether MTD might be present in tissue from patients treated with cefazolin prior to clinically-indicated surgery. To test that hypothesis, we took advantage of the fact that heterocyclic thiols can be S-methylated by thiopurine methyltransferase (TPMT). As a first step, recombinant human TPMT was used to S-methylate MTD. MTD was a TPMT substrate with an apparent K_m of 63 μM. TPMT was then used to radioactively label a methyl acceptor substrate present in liver and kidney cytosol from patients treated preoperatively with cefazolin. Pooled renal cytosol was then used to isolate the methylated product by reverse-phase HPLC. The methylated product in kidney cytosol coeluted with methylated MTD during HPLC. When this methylated product was subjected to tandem mass spectroscopy, it was identified as S-methyl-MTD. MTD, an inhibitor of clotting factor activation, is present in the tissues of patients treated with cefazolin.