Human concentrative nucleoside transporter 3 is a determinant of fludarabine transportability and cytotoxicity in human renal proximal tubule cell cultures

Interpatient variability in renal elimination of fludarabine (9-beta-D-arabinosyl-2-fluoroadenine) by renal human nucleoside transporters (hNTs) may contribute to unpredictable toxicities including rare nephrotoxicities. This study assessed relationships between hNT levels and fludarabine uptake and cytotoxicity in cultures of human renal proximal tubule cells (hRPTCs) that produce multiple transporter types.hRPTC cultures were established from ten different individuals and their hNT characteristics were assessed by measuring RNA expression by TaqMantrade mark reverse transcriptase polymerase chain reaction, protein abundance by quantitative immunoblotting of cell surface protein preparations, and uptake by radiolabeled nucleoside uptake assays. Fludarabine cytotoxicity against hRPTC cultures was quantified using methoxyphenyl tetrazolium inner salt (MTS) assays.RNA, protein and activities for human equilibrative NT 1 (hENT1) and 2 (hENT2) and human concentrative NT 3 (hCNT3) were identified in cultures of hRPTCs from ten different individuals. Significant differences in hCNT3 activities were exhibited among hRPTC cultures and correlated positively with cell surface levels of hCNT3 protein, but did not correlate with hCNT3 mRNA levels.Observed differences in hCNT3-mediated uptake activities, hNT-mediated fludarabine uptake activities, and fludarabine cytotoxicities correlated positively with each other, suggesting that hCNT3 is a primary determinant of fludarabine uptake and cytotoxicity in hRPTC cultures. Variations in hCNT3 abundance in renal proximal tubules, and hence nucleoside reabsorption, may explain interpatient variability in fludarabine's pharmacokinetics and toxicities.