Regulation of OATP1B1 and OATP1B3 by lysine acetylation and lysine deacetylase inhibitors - Organic anion transporting polypeptides (OATP)1B1 and OATP1B3 (abbreviated as OATP1B1/3) are liver- specific drug transport proteins that mediate uptake, from blood into the liver, of a diverse array of endogenous compounds, environmental toxins, and many clinically important drugs (e.g., lipid-lowering statins and anticancer agents). OATP1B1/3 are important determinants of transport-mediated drug-drug interactions (DDIs) resulting in severe side effects, such as statin-induced rhabdomyolysis, a sometimes-fatal muscle toxicity. Dysfunction of OATP1B1/3 significantly contributes to altered drug disposition and adverse drug events. Our long-term goal is to delineate the molecular mechanisms underlying drug/toxin disposition through OATP1B1/3, and to predict and mitigate OATP-mediated drug-drug and drug-disease interactions. Although it is evident that factors (drugs, aging, disease) modulating OATP1B1/3 function could cause drug-drug or drug-disease interactions with OATP1B1/3 substrates, unfortunately, our ability to predict such interactions is hampered due the dearth of information on OATP1B1/3 regulation. In particular, modulating lysine acetylation, a major post-translational modification known to alter function of numerous target proteins, including histone for epigenetic regulation, has not been investigated for OATP1B1/3. The overall objective of this application is to determine the molecular mechanisms governing the regulation of OATP1B1/3 by lysine acetylation, and to evaluate the impact of lysine deacetylase (KDAC) inhibitors (KDACIs) on OATP1B1/3 expression and function. Our central hypothesis is that OATP1B1/3 are lysine-acetylated proteins, deacetylation of which involves KDAC6; OATP1B1/3 can be regulated at the epigenetic level by KDACIs targeting histone deacetylation and at the post-translational level by KDAC6 inhibition. Our novel preliminary data show that 1) OATP1B1/3 are lysine-acetylated; 2) mutagenesis mimicking hyper-acetylation of OATP1B1 and specific KDAC 6 inhibition significantly reduces OATP1B1/3 transport function; and 3) FDA-approved pan KDAC inhibitors (KDACIs) inhibiting histone deacetylation induce mRNA and protein levels of OATP1B1/3. Guided by preliminary results, two specific Aims are outlined. In Aim 1, we will elucidate the role of KDAC6 in regulation of OATP1B1/3 lysine acetylation and transporter function. In Aim 2, we will dissect the regulation of OATP1B1/3 by pan KDACI drugs and the interplay between epigenetic and posttranslational regulation of OATP1B1/3 via K-Ac. A combination of proteomics, biochemical, and genetic engineering approaches will be utilized in cell lines and in the physiologically relevant sandwich-cultured primary human hepatocytes. The outcomes of these experiments will elucidate novel posttranslational and epigenetic mechanism(s) involved in regulating OATP1B1/3. The knowledge gained from these studies will be invaluable toward the rational design of novel drugs and inhibitors to optimize drug therapy while avoiding unwanted drug interactions. This work will enhance our ability to predict altered OATP1B1/3 function by lysine-deacetylase modulators (e.g., drugs/candidates that are HDAC inhibitors/activators and liver disease states).
