Ribavirin uptake by human erythrocytes and the involvement of nitrobenzylthioinosine-sensitive (es)-nucleoside transporters.
1. The major toxicity associated with oral therapy with ribavirin is anaemia, which has been postulated to occur as a result of accumulation of ribavirin triphosphate interfering with erythrocyte respiration. The objective of this study was to determine the mechanism by which ribavirin enters into erythrocytes. 2. Entry into human erythrocytes was examined by measuring influx rates of [3H]-ribavirin alone and with the inhibitor nitrobenzylthioinosine (NBMPR), and by investigating the inhibitory effects of nucleoside and nucleobase permeants on ribavirin transport, by use of inhibitor oil-stop methods. Transport mechanisms were further characterized by assessment of substrates to cause countertransport of ribavirin in preloaded erythrocytes, and by measuring the effects of ribavirin on [3H]-NBMPR binding to erythrocyte membranes. 3. Human erythrocytes had a saturable influx mechanism for ribavirin (Km at 22 degrees C of 440+/-100 microM) which was inhibited by nanomolar concentrations of NBMPR (IC50 0.99+/-0.15 nM). Nucleosides also inhibited the influx of ribavirin (adenosine more effective than uridine) but the nucleobases hypoxanthine and adenine had no effect. In addition, uridine caused the countertransport of ribavirin in human erythrocytes. Entry of ribavirin into horse erythrocytes, a cell type that lacks the NBMPR-sensitive (es) nucleoside transporter, proceeded slowly and via a pathway that was resistant to NBMPR inhibition. Ribavirin was a competitive inhibitor of adenosine influx (mean Ki 0.48+/-0.14 mM) and also inhibited NBMPR binding to erythrocyte membranes (mean Ki 2.2+/-0.39 mM). 4. These data indicate that ribavirin is a transported permeant for the es nucleoside transporter of human erythrocytes. There was no evidence for ribavirin entering cells via a nucleobase transporter.
Jarvis
S M
SM
Department of Biosciences, University of Kent, Canterbury, UK.
Thorn
J A
JA
Glue
P
P
D000241:Adenosine / Q000378:metabolism; D000818:Animals; D000998:Antiviral Agents / Q000097:blood*; D002352:Carrier Proteins / Q000378:metabolism*; D033722:Equilibrative-Nucleoside Transporter 2; D004910:Erythrocyte Membrane / Q000378:metabolism; D004912:Erythrocytes / Q000378:metabolism*; D006736:Horses; D006801:Humans; D066298:In Vitro Techniques; D008565:Membrane Proteins / Q000378:metabolism*; D012254:Ribavirin / Q000097:blood*; D013868:Thioinosine / Q000031:analogs & derivatives / Q000378:metabolism; D014529:Uridine / Q000097:blood
D000998:Antiviral Agents; D002352:Carrier Proteins; D033722:Equilibrative-Nucleoside Transporter 2; D008565:Membrane Proteins; C438654:SLC29A2 protein, human; D013868:Thioinosine; D012254:Ribavirin; C001789:4-nitrobenzylthioinosine; D000241:Adenosine; D014529:Uridine