Absence of P-glycoprotein transport in the pharmacokinetics and toxicity of the herbicide paraquat.

Genetic variation in the multidrug resistance gene ABCB1, which encodes the efflux transporter P-glycoprotein (P-gp), has been associated with Parkinson disease. Our goal was to investigate P-gp transport of paraquat, a Parkinson-associated neurotoxicant. We used in vitro transport models of ATPase activity, xenobiotic-induced cytotoxicity, transepithelial permeability, and rhodamine-123 inhibition. We also measured paraquat pharmacokinetics and brain distribution in Friend leukemia virus B-type (FVB) wild-type and P-gp-deficient (mdr1a(-/-)/mdr1b(-/-)) mice following 10, 25, 50, and 100 mg/kg oral doses. In vitro data showed that: 1) paraquat failed to stimulate ATPase activity; 2) resistance to paraquat-induced cytotoxicity was unchanged in P-gp-expressing cells in the absence or presence of P-gp inhibitors GF120918 [N-(4-[2-(1,2,3,4-tetrahydro-6,7-dimethoxy-2-isoquinolinyl)ethyl]-phenyl)-9,10-dihydro-5-methoxy-9-oxo-4-acridine carboxamide] and verapamil-37.0 [95% confidence interval (CI): 33.2-41.4], 46.2 (42.5-50.2), and 34.1 µM (31.2-37.2)-respectively; 3) transepithelial permeability ratios of paraquat were the same in P-gp-expressing and nonexpressing cells (1.55 ± 0.39 and 1.39 ± 0.43, respectively); and 4) paraquat did not inhibit rhodamine-123 transport. Population pharmacokinetic modeling revealed minor differences between FVB wild-type and mdr1a(-/-)/mdr1b(-/-) mice: clearances of 0.47 [95% confidence interval (CI): 0.42-0.52] and 0.78 l/h (0.58-0.98), respectively, and volume of distributions of 1.77 (95% CI: 1.50-2.04) and 3.36 liters (2.39-4.33), respectively; however, the change in clearance was in the opposite direction of what would be expected. It is noteworthy that paraquat brain-to-plasma partitioning ratios and total brain accumulation were the same across doses between FVB wild-type and mdr1a(-/-)/mdr1b(-/-) mice. These studies indicate that paraquat is not a P-gp substrate. Therefore, the association between ABCB1 pharmacogenomics and Parkinson disease is not attributed to alterations in paraquat transport.

Influence of substrate complexity on the diastereoselective formation of spiroiminodihydantoin and guanidinohydantoin from chromate oxidation.

Chromate is a human carcinogen with a poorly defined mechanism of DNA damage. In vitro and prokaryotic studies have shown that DNA damage may occur via the formation of the hydantoin lesions guanidinohydantoin (Gh) and spiroiminodihydantoin (Sp) from further oxidation of 8-oxo-7,8-dihydroguanine (8oxoG). The unusual structure of these lesions coupled with their enhanced mutagenicity make them attractive for study with regard to their role in chromate-induced cancer. We have studied the formation of Gh versus Sp and their associated diastereomers following oxidation by model Cr(V) complexes and from in situ chromate reduction by ascorbate and glutathione. Identification of the two optically assigned diastereomers of Sp (R-Sp and S-Sp) as well as the two diastereomers of Gh (Gh1 and Gh2, not yet optically assigned) was carried out using increasingly sterically hindered substrates (nucleoside --> ssDNA --> dsDNA). Lesion formation and diastereomeric preference were found to be highly oxidant- and substrate-dependent. The Ir(IV)-positive control showed a shift from near equal levels of Gh and Sp and near equal levels of all four diastereomers in the nucleoside to all Gh formation in dsDNA, with a 5-fold enhancement in Gh2 over Gh1. The two model Cr(V) complexes used in this study, Cr(V)-salen and Cr(V)-ehba, showed opposite trends going from nucleoside to dsDNA with Cr(V)-salen giving enhanced Sp formation (with mainly R-Sp formed) and the Cr(V)-ehba having an oxidation profile nearly identical to that of Ir(IV). The two chromate reduction systems, Cr(6+)/ascorbate and Cr(6+)/glutathione, designed to model the intracellular reduction of chromate, showed lower levels of oxidation in all substrates. Notable in this group was the shift in the formation of the lesions to essentially all Sp for the Cr(6+)/ascorbate system with the most sterically hindered substrate, dsDNA. These results, when coupled with the known diastereomeric preference for excision of hydantoin lesions by the hNEIL1 enzyme, show the importance of defining both levels of lesion formation and diastereomeric preference of formation with regard to their potential impact on chromate carcinogenesis.