Reduced Renal Clearance of Cefotaxime in Asians with a Low-Frequency Polymorphism of OAT3 (SLC22A8)

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ABSTRACT

Organic anion transporter 3 (OAT3, SLC22A8), a transporter expressed on the basolateral membrane of the proximal tubule, plays a critical role in the renal excretion of organic anions including many therapeutic drugs. The goal of this study was to evaluate the in vivo effects of the OAT3-Ile305Phe variant (rs11568482), present at 3.5% allele frequency in Asians, on drug disposition with a focus on cefotaxime, a cephalosporin antibiotic. In HEK293- Flp-In cells, the OAT3-Ile305Phe variant had a lower maximum cefotaxime transport activity, Vmax, [159 ± 3 nmol*(mg protein)- 1/min (mean ± SD)] compared with the reference OAT3 [305 ± 28 nmol*(mg protein) 1/min, (mean ± SD), p < 0.01], whereas the Michaelis-Menten constant values (Km) did not differ. In healthy volunteers, we found volunteers that were heterozygous for the Ile305Phe variant and had a significantly lower cefotaxime renal clearance (CLR; mean ± SD: 84.8 ± 32.1 mL/min, n = 5) compared with volunteers that were homozygous for the reference allele (158 ± 44.1 mL/min, n = 10; p = 0.006). Furthermore, the net secretory component of cefotaxime renal clearance (CLsec) was reduced in volunteers heterozygous for the variant allele [33.3 ± 31.8 mL/min (mean ± SD)] compared with volunteers homozygous for the OAT3 reference allele [97.0 ± 42.2 mL/min (mean ± SD), p = 0.01]. In summary, our study suggests that a low-frequency reduced-function polymorphism of OAT3 associates with reduced cefotaxime CLR and CLsec .©2013 Wiley Periodicals, Inc. and the American Pharmacists Association J Pharm Sci 102:3451–3457, 2013

Section snippets

INTRODUCTION

OAT1 (SLC22A6) and OAT3 (SLC22A8) are well- studied organic anion transporters known to reabsorb and secrete endogenous organic anions (e.g., pyruvate, α-ketoglutarate, oxalate, urate, and estrone sulfate) and exogenous anions (e.g., methotrexate, ciprofloxacin, acyclovir, captopril, and thiazides) across the basolateral membrane of the proximal tubule.1., 2., 3., 4. In vivo studies in Oat3 knockout mice demonstrate the importance of OAT3 in the renal elimination of drugs. The area under the

RESULTS AND DISCUSSION

In vitro cefotaxime transport was measured using HEK293-Flp-in cells stably transfected with empty vector (EV), reference SLC22A8 cDNA or with OAT3- Ile305Phe variant cDNA, reference SLC22A6 (OAT1), SLC22A7 (OAT2), and SLC22A11 (OAT4) cDNA subcloned into the mammalian expression vector pcDNA5/FRT. These cell lines, which were previously generated in our laboratory,10., 24., 25., 26. took up radiolabeled model substrates of organic anion transporters (Fig. S1). Cefotaxime uptake studies were

CONCLUSIONS

Cefotaxime is a third-generation cephalosporin with broad-spectrum antibacterial activity. In this study, we show that cefotaxime is a substrate of OAT3 and that the low-frequency OAT3 variant, Ile305Phe, found in Asians (~ 3.5% allele frequency), is associated with significant reduced cefotaxime uptake and reduced Vmax in in vitro kinetic studies. Our cell surface biotinylation study suggests that the mechanism of the reduced cefotaxime Vmax in OAT3-Ile305Phe cells is because of lower surface

ACKNOWLEDGMENTS

This work was supported by a grant from the National Institutes of Health (NIH; GM61390). C.B. was supported by National Research Service Award T32 GM07546 from the NI H. This work was also made possible in part by core services provided by the Clinical and Translational Science Institute (CTSI), Clinical Research Center at San Francisco General Hospital, funded by the National Center for Research Resources (NIH MO1-RR00083-44). We thank all staff at UCSF Drug Studies Unit for developing the

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