Transport and activation of S-(1,2-dichlorovinyl)- l-cysteine and N-acetyl- S-(1,2-dichlorovinyl)- l-cysteine in rat kidney proximal tubules

Abstract

An important step in understanding the mechanism underlying the tubular specificity of the nephrotoxicity of toxic cysteine conjugates is to identify the rate-limiting steps in their activation. The rate-limiting steps in the activation of toxic cysteine conjugates were characterized using isolated proximal tubules from the rat and 35S-labeled S-(1,2-dichlorovinyl)- l-cysteine (DCVC) and N-acetyl- S-(1,2-dichlorovinyl)- l-cysteine (NAC-DCVC) as model compounds. The accumulation by tubules of 35S radiolabel from both DCVC and NAC-DCVC was time and temperature dependent and was mediated by both Na +-dependent and independent processes. Kinetic studies with DCVC in the presence of sodium revealed the presence of two components with apparent K m and V max values of (1) 46 μ m and 0.21 nmol/mg·min and (2) 2080 μ m and 7.3 nmol/mg·min. NAC-DCVC uptake was via a single system with apparent K m and V max values of 157 μ m and 0.65 nmol/mg·min, respectively. Probenecid, an inhibitor of the renal organic anion transport system, inhibited accumulation of radiolabel from NAC-DCVC, but not from DCVC. The covalent binding of 35S label to cellular macromolecules was much greater from [ 35S]DCVC than from NAC-[ 35S]DCVC. Analysis of metabolites showed that a substantial amount of the cellular NAC-[ 35S]DCVC was unmetabolized while [ 35S]DCVC was rapidly metabolized to bound 35S-labeled material and unidentified products. The data suggest that DCVC is rapidly metabolized following transport, but that activation of NAC-DCVC depends on a slower rate of deacetylation. The results are discussed with regard to the segment specificity of cysteine conjugate toxicity and the role of disposition in vivo in the nephrotoxicity of glutathione conjugates.