Treatment with proteasome inhibitor bortezomib decreases organic anion transporting polypeptide (OATP) 1 B3-mediated transport in a substrate-dependent manner

OATP1B1 and OATP1 B3 mediate hepatic uptake of many drugs (e. g., statins) and can mediate transporter-mediated drug-drug-interactions (DDIs). Bortezomib is the first-in-class proteasome inhibitor drug approved by the U. S. Food and Drug Administration for the treatment of multiple myeloma. The potential of bortezomib to cause OATP-mediated DDIs has not been assessed. The current study investigated the involvement of the ubiquitin-proteasome system (UPS) in OATP1 B1 and OATP1 B3 degradation and determined the effects of proteasome inhibitors on OATP1 B1 -and OATP1 B3-mediated transport. Co-immunoprecipitation of FLAG-OATP1 Bi/1 B3 and HA-ubiquitin was observed in human embryonic kidney (HEK) 293 cells co-transfected with FLAG-tagged OATP1 B1/OATP1 B3 and hemagglutinin (HA)-tagged ubiquitin, suggesting that OATP1 B1 and OATP1 B3 can be ubiquitin-modified. Although blocking proteasome activity by bortezomib treatment (50 nM, 7 h) increased the endogenous ubiquitin-conjugated FLAG-OATP1 B1 and FLAG-OATP1 B3 in HEK293FLAG- OATP1 1 and-OATP1 B3 cells, such treatment did not affect the total protein levels of OATP1 B1 and OATP1 B3, suggesting that the UPS plays a minor role in degradation of OATP1 B1 and OATP1 B3 under current constitutive conditions. Pretreatment with bortezomib (50-250 nM, 2-7 h) significantly decreased transport of [3H] CCK-8, a specific OATP1 B3 substrate, in HEK293-OATP1 B3 and human sandwich-cultured hepatocytes (SCH). However, bortezomib pretreatment had negligible effects on the transport of [3H] E2 17O3G and [3H] pitavastatin, dual substrates of OATP1 B1 and OATP1 B3, in HEK293OATP1 B1/1 B3 cells and/or human SCH. Compared with vehicle control treatment, bortezomib pretreatment significantly decreased the maximal transport velocity (Vmax) of OATP1 B3-mediated transport of CCK-8 (92.25 +/- 14.2 vs. 133.95 +/- 15.5 pmol/mg protein/min) without affecting the affinity constant (Km) values. Treatment with other proteasome inhibitors MG1 32, epoxomicin, and carfilzomib also significantly decreased OATP1 B3mediated [3H] CCK-8 transport. In summary, the current studies for the first time report ubiquitination of OATP1 B1 and OATP1 B3 and the apparent substrate-dependent inhibitory effect of bortezomib on OATP1 B3-mediated transport. The data suggest that bortezomib has a low risk of causing OATP-mediated DDIs.