Flavonoid conjugates interact with organic anion transporters (OATs) and attenuate cytotoxicity of adefovir mediated by organic anion transporter 1 (OAT1/SLC22A6)

Abstract

Flavonoids are conjugated by phase II enzymes in humans to form glucuronidated and sulfated metabolites that are excreted in urine via the kidney. In this study, we examined the interaction between metabolites of quercetin and isoflavonoids found in vivo with human organic anion transporters 1 (OAT1) and 3 (OAT3) and their potential in attenuating OAT-induced cytotoxicity of adefovir. Accumulation of flavonoid conjugates was studied in human embryonic kidney 293H cells overexpressing OAT1 or OAT3. OAT1-overexpressing cells exhibited an increased uptake of the sulfated conjugates, genistein-4′- O-sulfate and quercetin-3′- O-sulfate. OAT3-overexpressing cells demonstrated enhanced uptake of glucuronide conjugates, such as daidzein-7- O-glucuronide, genistein-7- O-glucuronide, glycitein-7- O-glucuronide and quercetin-3′- O-glucuronide. Position of conjugation was important since quercetin-3- O-glucuronide and quercetin-7- O-glucuronide were poorly transported. Kinetic analysis revealed high affinity uptake of quercetin-3′- O-sulfate by OAT1 ( K m = 1.73 μM; V max = 105 pmol/min/mg). OAT3 transported isoflavone glucuronides with lower affinity ( K m = 7.9–19.1 μM) but with higher V max (171–420 pmol/min/mg). Quercetin-3′- O-sulfate strongly inhibited OAT1-mediated p-aminohippuric acid uptake with an IC 50 of 1.22 μM. Transport of 5-carboxyfluorescein by OAT3 was potently inhibited by quercetin-3- O-glucuronide, quercetin-3′- O-glucuronide and quercetin-3′- O-sulfate (IC 50 = 0.43–1.31 μM). In addition, quercetin-3′- O-sulfate was shown to effectively reduce OAT1-mediated cytotoxicity of adefovir, an antiviral drug, in a dose-dependent manner. These data suggest that OAT1 and OAT3 are responsible for basolateral uptake of flavonoid conjugates in kidney, and flavonoid conjugates inhibit OAT1 and OAT3 activity at physiologically relevant concentrations. Interaction with OATs limits systemic availability of flavonoids and may be a mechanism of food–drug interaction via inhibition of renal uptake.