Uptake of Chemically Reactive, DNA-Damaging Sulfuric Acid Esters into Renal Cells by Human Organic Anion Transporters

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Abstract
The procarcinogen 1-methylpyrene is activated by hepatic enzymes via 1-hydroxymethylpyrene to 1-sulfooxymethylpyrene (1-SMP), a highly reactive and mutagenic metabolite. Previously, high levels of 1-SMP DNA adducts were observed in rat kidneys after intraperitoneal administration of 1-hydroxymethylpyrene or 1-SMP. This study examined whether organic anion transporters (OAT) that are expressed at the basolateral membrane of proximal tubule cells are involved in uptake of SMP. Human epithelial kidney (HEK293) cells that stably express human OAT1 (hOAT1) and hOAT3 were used. Stable isomers of 1-SMP, (2-SMP and 4-SMP) competitively inhibited the uptake of characteristic substrates p-aminohippurate for hOAT1 and estrone sulfate for hOAT3. Both inhibitors exhibited high affinity for hOAT1 (Ki = 4.4 μM for 2-SMP; Ki = 5.1 μM for 4-SMP) as well as hOAT3 (Ki = 1.9 μM for 2-SMP; Ki = 2.1 μM for 4-SMP). The uptake rate of 4-SMP (at a concentration of 10 μM) by hOAT1- and hOAT3-expressing cells was 3.0 and 1.6 times higher, respectively, than in control cells. Uptake of the reactive isomer 1-SMP was investigated using as the end point the level of DNA adducts that were formed in the cells. After exposure to 1-SMP (10 μM), the DNA adduct level was 4.6 and 3.0 times higher in hOAT1- and hOAT3-expressing cells, respectively, than in control cells. The enhanced DNA adduct formation in hOAT-expressing cells was abolished in the presence of the OAT inhibitor probenecid. This study indicates that OAT can mediate the basolateral uptake of reactive sulfuric acid esters into proximal tubule cells and thereby participate in kidney cell damage by these compounds.