PT - JOURNAL ARTICLE AU - Michele Visentin AU - Min-Hwang Chang AU - Michael F Romero AU - Rongbao Zhao AU - I David Goldman TI - Substrate- and pH- specific antifolate transport mediated by OATP2B1-SLCO2B1 AID - 10.1124/mol.111.074823 DP - 2011 Jan 01 TA - Molecular Pharmacology PG - mol.111.074823 4099 - http://molpharm.aspetjournals.org/content/early/2011/10/20/mol.111.074823.short 4100 - http://molpharm.aspetjournals.org/content/early/2011/10/20/mol.111.074823.full AB - Human OATP2B1 (OATP-B; SLCO2B1) is expressed in the apical membrane of the small intestine and the hepatocyte basolateral membrane and transports structurally diverse organic anions with a wide spectrum of pH sensitivities. This paper describes highly pH-dependent OATP2B1-mediated antifolate transport and compares this property with that of BSP, a preferred substrate. At pH 5.5 and low substrate concentrations (~2.5 μM), only [3H]pemetrexed influx (in contrast to methotrexate (MTX), folic acid and reduced folates) could be detected in OATP2B1-transfected HeLa R1-11 cells that lack endogenous folate-specific transporters. Influx was optimal at pH 4.5-5.5, falling precipitously with an increase in pH > 6.0; BSP influx was independent of pH. Influx of both substrates at low pH was markedly inhibited by the proton ionophore, FCCP; BSP influx was also suppressed at pH 7.4. At 300 μM MTX, influx was one-third of that of pemetrexed; influx of folic acid, 5-methyltetrahydrofolate, or 5-formyltetrahydrofolate was not detected; there were similar findings in OATP2B1-expressing Xenopus oocytes. The pemetrexed influx Km was ~ 300 μM; the raltitrexed influx Ki was ~ 70 μM at pH 5.5. Stable expression of OAPT2B1 in HeLa R1-11 cells resulted in substantial raltitrexed, but modest pemetrexed, growth inhibition consistent with their affinities for this carrier. Hence, OATP2B1 represents a low-affinity transport route for antifolates (relative affinities: raltitrexed>pemetrexed>MTX), at low pH. In contrast, the high affinity of this transporter for BSP relative to antifolates appears to be intrinsic to the carrier binding site independent of the proton concentration.