Development of a Rat Sandwich-Cultured Hepatocytes Model Expressing Functional Human Organic Anion Transporting Polypeptide (OATP) 1B3: A Potential Screening Tool for Liver-Targeting Compounds

Purpose: Organic anion transporting polypeptide (OATP) 1B3 transports many clinically important drugs, including statins, from blood into the liver. It exclusively expresses in human liver under normal physiological conditions. There is no rodent ortholog of human OATP1B3. Tissue targeting of therapeutic molecules mediated by transporters, including liver-targeting via liver-specific OATPs, is an emerging area in drug development. Sandwich-cultured primary hepatocytes (SCH) are a well characterized in vitro model for assessment of hepatic drug uptake and biliary excretion. The current study was designed to develop a novel rat SCH model expressing human OATP1B3 to study the hepatic disposition of OATP1B3 substrates. Methods: Primary rat hepatocytes transduced with adenoviral vectors expressing FLAG-tagged OATP1B3 (Ad-OATP1B3), a control vector Ad-LacZ, or that were non-transduced were cultured in a sandwich configuration. FLAG immunoblot and immunofluorescence-staining determined expression and localization of OATP1B3. Uptake of [3H]-cholecystokinin octapeptide (CCK-8), a specific OATP1B3 substrate, was determined. Taurocholate (TC) is a substrate routinely used in SCH to assess biliary excretion via bile canaliculi (BC) and is also a substrate of OATP1B3. [3H]-TC accumulation in cells+BC, cells, biliary excretion index (BEI) and in vitro Clbiliary were determined using B-CLEAR® technology. Results: OATP1B3 protein was extensively expressed and primarily localized on the plasma membrane in day 4 Ad-OATP1B3-transduced rat SCH. [3H]-CCK-8 accumulation in cells+BC was significantly greater (~5-13 folds, p<0.001) in day 4 SCH with vs. without Ad-OATP1B3-transduction. Expressing OATP1B3 in rat SCH significantly increased [3H]-TC accumulation in cells+BC and cells, without affecting BEI and in vitro Clbiliary. Conclusions: Rat SCH expressing human OATP1B3-is a novel in vitro model allowing simultaneous assessment of hepatic uptake, hepatocellular accumulation and biliary excretion process of a human OATP1B3 substrate. This model could be a potential tool for screening for liver-targeting compounds mediated by OATP1B3.

Wilson’s Disease: Facing the Challenge of Diagnosing a Rare Disease

Wilson disease (WD) is a rare disorder caused by mutations in ATP7B, which leads to the defective biliary excretion of copper. The subsequent gradual accumulation of copper in different organs produces an extremely variable clinical picture, which comprises hepatic, neurological psychiatric, ophthalmological, and other disturbances. WD has a specific treatment, so that early diagnosis is crucial to avoid disease progression and its devastating consequences. The clinical diagnosis is based on the Leipzig score, which considers clinical, histological, biochemical, and genetic data. However, even patients with an initial WD diagnosis based on a high Leipzig score may harbor other conditions that mimic the WD’s phenotype (Wilson-like). Many patients are diagnosed using current available methods, but others remain in an uncertain area because of bordering ceruloplasmin levels, inconclusive genetic findings and unclear phenotypes. Currently, the available biomarkers for WD are ceruloplasmin and copper in the liver or in 24 h urine, but they are not solid enough. Therefore, the characterization of biomarkers that allow us to anticipate the evolution of the disease and the monitoring of new drugs is essential to improve its diagnosis and prognosis.

Assessing the Functional Redundancy between P-gp and BCRP in Controlling the Brain Distribution and Biliary Excretion of Dual Substrates with PET Imaging in Mice

P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP) are co-localized at the blood–brain barrier, where they display functional redundancy to restrict the brain distribution of dual P-gp/BCRP substrate drugs. We used positron emission tomography (PET) with the metabolically stable P-gp/BCRP substrates [11C]tariquidar, [11C]erlotinib, and [11C]elacridar to assess whether a similar functional redundancy as at the BBB exists in the liver, where both transporters mediate the biliary excretion of drugs. Wild-type, Abcb1a/b(−/−), Abcg2(−/−), and Abcb1a/b(−/−)Abcg2(−/−) mice underwent dynamic whole-body PET scans after i.v. injection of either [11C]tariquidar, [11C]erlotinib, or [11C]elacridar. Brain uptake of all three radiotracers was markedly higher in Abcb1a/b(−/−)Abcg2(−/−) mice than in wild-type mice, while only moderately changed in Abcb1a/b(−/−) and Abcg2(−/−) mice. The transfer of radioactivity from liver to excreted bile was significantly lower in Abcb1a/b(−/−)Abcg2(−/−) mice and almost unchanged in Abcb1a/b(−/−) and Abcg2(−/−) mice (with the exception of [11C]erlotinib, for which biliary excretion was also significantly reduced in Abcg2(−/−) mice). Our data provide evidence for redundancy between P-gp and BCRP in controlling both the brain distribution and biliary excretion of dual P-gp/BCRP substrates and highlight the utility of PET as an upcoming tool to assess the effect of transporters on drug disposition at a whole-body level.

Implementation of mini-invasive methods of bile drainage in the treatment of blastomatous mechanical jaundice

Tumor diseases of the BPDZ (biliopancreatoduodenal zone) occupy one of the leading places among the causes of disability and mortality of the population, both according to global and domestic statistics, while in 85–90 % of cases, patients with distal tumor biliary obstruction are detected. The most common symptom of malignant tumors of periampullary localization is OJ(obstructive jaundice), which is found in 86–95 % of patients and becomes the first manifestation of the disease in 65–70 % of cases. Purpose. Optimization of surgical treatment in surgically incurable patients with obstructive blastomatous jaundice. Materials and methods. The study was clinical in nature, it was carried out on the basis of CCH №17 (Kharkov, Ukraine) and the State Institution “Institute of General and Emergency Surgery named after V. I. Zaitsev National Academy of Medical Sciences of Ukraine ") and consisted of two stages. The main task of the first stage was a retrospective assessment of the results of antegrade and retrograde minimally invasive interventions performed in the period from 2006 to 2011 in 122 patients. The main objective of the second stage of the study was to assess the results of surgical treatment of 75 patients with blastomatous breast, who were treated from 2012 to 2019. using the developed treatment algorithm. Results. At the first stage of the study, the effectiveness of endoscopic transpapillary and transhepatic biliary excretion methods in patients with distal malignant obstruction of the gastrointestinal tract was compared, for which patients who underwent minimally invasive interventions were divided into two subgroups: group A (63 observations) biliary excretion, and in group b (59 observations) as biliary decompression antegrade percutaneous-transhepatic methods of drainage of bilious ways were applied. Endoscopic transpapillary surgery showed itself to be, not inferior to open surgery in effectiveness of biliary decompression, instead it has a number of advantages, such as low trauma, relative safety, low complication and mortality. Conclusions. The developed algorithm of bile excretion in incurable patients with blastomatous MF allowed to minimize the number of postoperative complications to 4.0 %, to avoid the development of transient hyperamylasemia and postmanipulation pancreatitis. Endoscopic retrograde methods of RV prosthetics as the final stage of surgical treatment in this category of patients have certain advantages over antegrade methods of external drainage, primarily due to the preservation of the natural passage of bile in the duodenum and better adaptation and tolerability by patients

Sitosterolemia: Inherited Disorder of Plant Sterols Absorption and Biliary Excretion

Sitosterolemia is a rare, inherited, autosomal recessive disorder of lipid metabolism, we report on the diagnosis and treatment of a child with sitosterolemia misdiagnosed as a skin disease.