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

Abstract

OATP1B1 and OATP1B3 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 OATP1B1 and OATP1B3 degradation and determined the effects of proteasome inhibitors on OATP1B1- and OATP1B3-mediated transport. Co-immunoprecipitation of FLAG-OATP1B1/1B3 and HA-ubiquitin was observed in human embryonic kidney (HEK) 293 cells co-transfected with FLAG-tagged OATP1B1/OATP1B3 and hemagglutinin (HA)-tagged ubiquitin, suggesting that OATP1B1 and OATP1B3 can be ubiquitin-modified. Although blocking proteasome activity by bortezomib treatment (50 nM, 7 h) increased the endogenous ubiquitin-conjugated FLAG-OATP1B1 and FLAG-OATP1B3 in HEK293-FLAG-OATP1B1 and–OATP1B3 cells, such treatment did not affect the total protein levels of OATP1B1 and OATP1B3, suggesting that the UPS plays a minor role in degradation of OATP1B1 and OATP1B3 under current constitutive conditions. Pretreatment with bortezomib (50–250 nM, 2–7 h) significantly decreased transport of [3H]CCK-8, a specific OATP1B3 substrate, in HEK293-OATP1B3 and human sandwich-cultured hepatocytes (SCH). However, bortezomib pretreatment had negligible effects on the transport of [3H]E217βG and [3H]pitavastatin, dual substrates of OATP1B1 and OATP1B3, in HEK293-OATP1B1/1B3 cells and/or human SCH. Compared with vehicle control treatment, bortezomib pretreatment significantly decreased the maximal transport velocity (Vmax) of OATP1B3-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 MG132, epoxomicin, and carfilzomib also significantly decreased OATP1B3-mediated [3H]CCK-8 transport. In summary, the current studies for the first time report ubiquitination of OATP1B1 and OATP1B3 and the apparent substrate-dependent inhibitory effect of bortezomib on OATP1B3-mediated transport. The data suggest that bortezomib has a low risk of causing OATP-mediated DDIs.

Regulation of Organic Anion Transporting Polypeptides (OATP) 1B1- and OATP1B3-Mediated Transport: An Updated Review in the Context of OATP-Mediated Drug-Drug Interactions

“…Ubiquitination is an important post-translational modification that regulates many cellular processes including signal transduction, cell cycle control and transcriptional regulation through mediating proteasome degradation of proteins and the maintenance of protein homeostasis [ 137 ]. A recent study reported that both OATP1B1 and OATP1B3 can be ubiquitin-conjugated in HEK293 cells over-expressing OATP1B1 or OATP1B3 [ 138 ]. However, the effects of ubiquitination on OATP1B1 and OATP1B3 transport function have not been demonstrated and further studies are warranted.…”

“…In OATP1B1- and OATP1B3-expressing stable cell lines and human SCH, treatment with lysosome inhibitor chloroquine markedly increased protein levels of OATP1B1 and 1B3, suggesting that the lysosome plays an important role in degradation of OATP1B1 and OATP1B3 [ 138 , 161 ]. The estimated maximum unbound concentration of chloroquine at the inlet to the liver is ~56.8 μM after a 600 mg single dose in human [ 161 ].…”

“…Bortezomib is the first-in-class proteasome inhibitor approved as the first-line treatment option for multiple myeloma and second-line treatment option for mantle cell lymphoma [ 171 ]. Although OATP1B1 and OATP1B3 are ubiquitinated proteins in transporter overexpressing HEK293 cells [ 138 ], treatment with bortezomib did not affect the total protein levels of OATP1B1 and OATP1B3, suggesting that the proteasome is likely to play a minor role in degradation of OATP1B1 and OATP1B3 under constitutive conditions.…”

“…Bortezomib is not a competitive inhibitor of OATP1B1 or OATP1B3 at clinically relevant concentrations up to 98 nM [ 138 ]. Interestingly, pretreatment with bortezomib decreased OATP1B3-mediated transport in a substrate-dependent manner.…”

Proteasome Inhibitors Bortezomib and Carfilzomib Stimulate the Transport Activity of Human Organic Anion Transporter 1

“…Carfilzomib inhibited the chymotrypsin-like activity of the 20S proteasome in the kidney by 50~60% in rats (FDA, 2012). Recent studies showed that bortezomib can affect the activities of copper transporter 1, ATP-binding cassette transporter A1 (ABCA1) and ABCG1, metal transporter ZIP14 and organic anion transporting polypeptide (OATP) 1B3 (Alam et al, 2017;Jandial et al, 2009;Ogura et al, 2011;Zhao et al, 2014). However, the potential of these antitumor drugs to modulate OAT1 activity has not been explored.…”