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 opt...