**1. Introduction**

Organic anion transporter 3 (OAT3), which is encoded by the SLC22A8 gene, is primarily expressed at the basolateral membrane of kidney proximal tubules, and actively translocates corresponding substrates from the blood into renal tubule epithelial cells. Those substrates are then effluxed out of the apical membrane into urine by other transporters [1–3]. OAT3 is involved in the renal elimination of various kinds of important clinical drugs from the kidney, such as anticancer agents (e.g., methotrexate), antivirals (e.g., tenofovir, valacyclovir), antibiotics (e.g., benzylpenicillin, cefotaxime), antihypertensives (e.g., furosemide, sitagliptin), H2 receptor antagonists (e.g., cimetidine, famotidine), and nonsteroidal anti-inflammatory drugs (e.g., ketoprofen, ibuprofen) [4–6]. Therefore, the renal OAT3 function is a critical determinant in drug clearance out of the body, and in the pharmacokinetic and pharmacodynamic properties of drugs, which ultimately affect the drugs' efficacy and systemic or renal toxicity.

Combination therapies by coadministration of different drugs are often used for treatment of a single or multiple diseases. If one drug is an inhibitor, substrate, or inducer of OAT3, it will inhibit uncompetitively or competitively, or stimulate the renal transport and

**Citation:** Fan, Y.; Liang, Z.; Zhang, J.; You, G. Oral Proteasomal Inhibitors Ixazomib, Oprozomib, and Delanzomib Upregulate the Function of Organic Anion Transporter 3 (OAT3): Implications in OAT3-Mediated Drug-Drug Interactions. *Pharmaceutics* **2021**, *13*, 314. https://doi.org/10.3390/ pharmaceutics13030314

Academic Editor: Dong Hyun Kim

Received: 31 July 2020 Accepted: 24 February 2021 Published: 28 February 2021

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excretion of other clinical substrates, cause potential drug-drug interactions (DDIs), and sequent intra- and interindividual variation in clinical response to drugs [6,7]. Transportermediated clinical DDIs have attracted the attention of academic, industrial, and regulatory agencies. OAT3-mediated DDIs abundantly exist between imipenem-cilastatin, piperacillintazobactam, bezafibrate-mizoribine, puerarin-methotrexate, benzylpenicillin–acyclovir, etc., and markedly alter the pharmacokinetic parameters of affected drugs [8–12]. Through inhibition of OAT1/3, probenecid, wedelolactone, and wogonin prevented the kidney accumulation of aristolochic acid and related nephropathy, apigenin- or cilastatin-ameliorated imipenem, or diclofenac-induced nephrotoxicity [13–17].

The transporter expression and function may be modulated by certain drugs, phytomedicines, or xenobiotics, resulting in altered disposition of clinical substances, which is an indirect manner of obtaining transporter-mediated DDIs, in contrast to direct interaction with the transporter-like inhibitors or substrates [4]. For example, administration of 1 <sup>α</sup>,25-dihydroxyvitamin D3, mercuric chloride, or methotrexate decreased OAT3 expression in crude or basolateral membranes of rat kidneys; while the renal expression was increased in normal rats by ochratoxin A treatment, in diabetic rats by insulin or atorvastatin plus insulin treatment, or in obese rats by prebiotic *Lactobacillus paracasei* HII01 or xylooligosaccharide treatment [18–25].

OAT3 expression and activity can be regulated through posttranslational modifications, including phosphorylation, ubiquitination, and SUMOylation [26–28]. As ubiquitination of OAT3 is an initiating process that triggers the internalization of OAT3 from the plasma membrane to intracellular endosomes, it is a critical molecular mechanism for OAT3 regulation [29,30]. Our lab demonstrated that activation of protein kinase C (PKC) could enhance OAT3 ubiquitination, and accelerate OAT3 internalization and subsequent degradation [27]. The transport activity and quantity of OAT3 on the plasma membrane were then reduced. Since proteasome inhibition can affect ubiquitination of targeted proteins and degradation, modulation of proteasome activity could potentially interfere with the physiological function of transporters. Proteasome inhibitors have shown to influence the copper transporter 1, Na+/H+ exchanger-3, ATP-binding cassette transporters A1 (ABCA1) and ABCG1, organic-anion-transporting polypeptide (OATP) 1B3, metal transporter ZIP14, and OAT1 [31–36]. However, it is not clear whether OAT3 can be regulated by controlling proteasome activity. Ixazomib, oprozomib, and delanzomib are oral proteasome inhibitors that target the ubiquitin–proteasome system for multiple myeloma treatment. In the present study, we investigated the influence of ixazomib, oprozomib, and delanzomib on OAT3 expression and transport activity, and elucidated the underlying mechanisms.

#### **2. Materials and Methods**
