DOCSLIB.ORG

Effect of MRP4 G487E Variant on Atpase Activity

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Effect of MRP4 G487E Variant on Atpase Activity Copyright 2015 by Jessica L. Friedman ii Abstract ATP-binding cassette (ABC) transporters are a diverse family of transmembrane proteins that facilitate the ATP powered translocation of substrates across cellular membranes. Multidrug resistance-associated protein 4 (MRP4) is a highly polymorphic, human ABC transporter with a wide array of substrates, including, antiviral drugs and prostaglandins. Previously, altered antiviral drug transport capacities of nonsynonymous single nucleotide polymorphic MRP4 variants, relative to reference MRP4 transport activity, were reported. This project focused on characterizing the ATP hydrolysis capabilities of the G487E MRP4 variant (rs11568668); G487 is located in an ATP binding cassette, also known as the nucleotide binding domain, of MRP4. Previously published works report a 0.008 minor allele frequency in Asian American individuals (n = 120) and show decreased transport activity with G487E. Novel data on G487E MRP4 activity from this study corroborate the decreased drug transport findings. This study characterized prostaglandin E2 – stimulated ATP hydrolysis by human MRP4 reference and G487E variant expressed in insect cells. MRP4 specific activity was determined by its selective inhibition with beryllium fluoride. Reference MRP4 reached a maximal ATP hydrolysis velocity of 27.4 ± 1.07 nmoles Pi/mg protein/minute; for comparison, ATP hydrolysis in Sf21 membranes absent of exogenous transporters reached a maximum velocity of 18.4 ± 0.55 nmoles Pi/mg protein/minute. The G487E MRP4 variant had an intermediate Vmax value of 24.4 ± 0.93 nmoles Pi/mg protein/minute. As measured by an ANOVA, Vmax values for the Sf21, MRP4 reference and G487E MRP4 membranes were significantly different (p < 0.0001). In contrast, Km values were similar for the three sample types and ranged from 0.34 ± 0.16 µM for reference MRP4 to 0.44 ± 0.18 µM for G487E MRP4. Structural rationales for these data iii were explored using novel homology models of the first nucleotide binding domain of MRP4. The G487E mutation was determined to only affect the MRP4 binding affinity of ATP; prostaglandin E2 binding was not impacted. These results suggest that the reduced transport phenotype of G487E MRP4 relative to reference MRP4 is due, at least in part, to impaired ATP hydrolysis. Acknowledgements This document marks the end of an incredible journey in academic research and the beginning of an exciting new career in pharmaceutical consulting. I want to thank my friends, family, and mentors who enhanced this experience and helped make this transition possible. To my thesis committee, thank you for your support in my decision to leave UCSF a few years earlier than originally planned. Deanna Kroetz, thank you for being an incredibly supportive PI and friend. Despite my career interests being far outside of your expertise, you facilitated the series of informational interviews that led me to health care consulting and gave me the time to explore and enter this career path. Thank you for your encouragement through this entire project; I am thrilled that I could leave the lab with detailed instructions for the successful execution of MRP4 ATPase assays and preliminary MRP4 variant ATPase activity data. Jaime Fraser, thank you for your continual support through my UCSF journey, from my recruitment in 2013 to my Masters thesis submission in 2015. Kathy Giacomini, thank you for your insight into career transitions in non-research science careers and your feedback on my Masters thesis. To my PSPG cohort, thank you for being involved in every step of this journey – becoming a California resident, balancing fellowship applications, rotations, project proposals, with journal clubs and 220 presentations, surviving TAing pharmacy students, and solidifying our next career steps. iv To my friends and family, thank you for always being a phone call, text, or video call away – moving 3,000 miles did not feel so far because you were always so close. Grandpa, thank you for inspiring my resilience and determination. Dad, thank you for trying to give advice even when my questions were beyond your expertise and your unwavering confidence in me. The work described in Chapter 2 was performed with guidance from published work: Sauna ZE, Nandigama K, Ambudkar S V. Multidrug resistance protein 4 (ABCC4)-mediated ATP hydrolysis: Effect of transport substrates and characterization of the post-hydrolysis transition state. J Biol Chem. 2004;279(47):48855-48864. doi:10.1074/jbc.M408849200. I appreciate the helpful advice of the Ambudkar laboratory at the National Cancer Institute while setting up this assay. Finally, I would like to thank everyone in the Kroetz and Giacomini labs for their support in and out of the laboratory. Without you, I would still be looking for the MilliQ water (hidden in the fish room). v Table of Contents Abstract ......................................................................................................................................... iii Acknowledgements ...................................................................................................................... iv List of Tables .............................................................................................................................. viii List of Figures ............................................................................................................................... ix Chapter 1 : Introduction .............................................................................................................. 1 ABC Transporters .................................................................................................................................. 1 Human ABC Transporters .................................................................................................................... 1 Functional Characteristics .................................................................................................................... 5 ABC Transporter Associated Diseases................................................................................................. 6 Multidrug Resistance-Associated Protein 4 (MRP4) ........................................................................... 8 Tissue Distribution ............................................................................................................................... 8 Interaction with Drugs .......................................................................................................................... 8 ATP Binding and Hydrolysis ............................................................................................................. 11 MRP4 Homology Modeling ............................................................................................................... 13 ABCC4 Genetic Variants .................................................................................................................... 15 Goals and Rationale for Project.......................................................................................................... 19 References ............................................................................................................................................. 21 Chapter 2 : Effect of MRP4 G487E Variant on ATPase Activity .......................................... 26 Materials and Methods ........................................................................................................................ 26 Sf21 Cell Culture ................................................................................................................................ 26 Sf21 Cell Infection and Harvest ......................................................................................................... 27 Membrane Isolation............................................................................................................................ 27 vi Western Blotting ................................................................................................................................ 29 ATPase Activity Assay ...................................................................................................................... 30 Homology Template Search and Selection. ....................................................................................... 32 Model Building and Quality Assessment ........................................................................................... 32 Results .................................................................................................................................................... 33 Assay Development............................................................................................................................ 33 MRP4 Specific ATPase Activity ........................................................................................................ 35 Substrate-Dependent MRP4 ATPase Activity ................................................................................... 37 Proposed Reaction Mechanism .......................................................................................................... 39 Structural Observations ...................................................................................................................... 40 Discussion .............................................................................................................................................. 46 Assay Optimization
Load more

Recommended publications
  • Multidrug Transporter MRP4/ABCC4 As a Key Determinant of Pancreatic
    www.nature.com/scientificreports OPEN Multidrug transporter MRP4/ ABCC4 as a key determinant of pancreatic cancer aggressiveness A. Sahores1, A. Carozzo1, M. May1, N. Gómez1, N. Di Siervi1, M. De Sousa Serro1, A. Yanef1, A. Rodríguez‑González2, M. Abba3, C. Shayo2 & C. Davio1* Recent fndings show that MRP4 is critical for pancreatic ductal adenocarcinoma (PDAC) cell proliferation. Nevertheless, the signifcance of MRP4 protein levels and function in PDAC progression is still unclear. The aim of this study was to determine the role of MRP4 in PDAC tumor aggressiveness. Bioinformatic studies revealed that PDAC samples show higher MRP4 transcript levels compared to normal adjacent pancreatic tissue and circulating tumor cells express higher levels of MRP4 than primary tumors. Also, high levels of MRP4 are typical of high-grade PDAC cell lines and associate with an epithelial-mesenchymal phenotype. Moreover, PDAC patients with high levels of MRP4 depict dysregulation of pathways associated with migration, chemotaxis and cell adhesion. Silencing MRP4 in PANC1 cells reduced tumorigenicity and tumor growth and impaired cell migration. Transcriptomic analysis revealed that MRP4 silencing alters PANC1 gene expression, mainly dysregulating pathways related to cell-to-cell interactions and focal adhesion. Contrarily, MRP4 overexpression signifcantly increased BxPC-3 growth rate, produced a switch in the expression of EMT markers, and enhanced experimental metastatic incidence. Altogether, our results indicate that MRP4 is associated with a more aggressive phenotype in PDAC, boosting pancreatic tumorigenesis and metastatic capacity, which could fnally determine a fast tumor progression in PDAC patients. Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal human malignancies, due to its late diag- nosis, inherent resistance to treatment and early dissemination 1.
    [Show full text]
  • Quantitative Evaluation of Drug Resistance Profile of Cells Expressing Wild-Type Or Genetic Polymorphic Variants of the Human ABC Transporter ABCC4
    Article Quantitative Evaluation of Drug Resistance Profile of Cells Expressing Wild-Type or Genetic Polymorphic Variants of the Human ABC Transporter ABCC4 Megumi Tsukamoto 1, Shiori Sato 2, Kazuhiro Satake 1, Mizuki Miyake 1 and Hiroshi Nakagawa 2,* 1 Department of Applied Biological Chemistry, Graduate School of Bioscience and Biotechnology, Chubu University, 1200 Matsumoto-cho, Kasugai 487-8501, Japan; [email protected] (M.T.); [email protected] (K.S.); [email protected] (M.M.) 2 Department of Applied Biological Chemistry, College of Bioscience and Biotechnology, Chubu University, 1200 Matsumoto-cho, Kasugai, Aichi 487-8501, Japan; [email protected] * Correspondence: [email protected]; Tel.: +81-568-51-9606; Fax: +81-568-52-6594 Received: 14 April 2017; Accepted: 26 June 2017; Published: 4 July 2017 Abstract: Broad-spectrum resistance in cancer cells is often caused by the overexpression of ABC transporters; which varies across individuals because of genetic single-nucleotide polymorphisms (SNPs). In the present study; we focused on human ABCC4 and established cells expressing the wild-type (WT) or SNP variants of human ABCC4 using the Flp-In™ system (Invitrogen, Life Technologies Corp, Carlsbad, CA, USA) based on Flp recombinase-mediated transfection to quantitatively evaluate the effects of nonsynonymous SNPs on the drug resistance profiles of cells. The mRNA levels of the cells expressing each ABCC4 variant were comparable. 3-(4,5-Dimethyl-2- thiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay clearly indicated that the EC50 values of azathioprine against cells expressing ABCC4 (WT) were 1.4–1.7-fold higher than those against cells expressing SNP variants of ABCC4 (M184K; N297S; K304N or E757K).
    [Show full text]
  • Interindividual Differences in the Expression of ATP-Binding
    Supplemental material to this article can be found at: http://dmd.aspetjournals.org/content/suppl/2018/02/02/dmd.117.079061.DC1 1521-009X/46/5/628–635$35.00 https://doi.org/10.1124/dmd.117.079061 DRUG METABOLISM AND DISPOSITION Drug Metab Dispos 46:628–635, May 2018 Copyright ª 2018 by The American Society for Pharmacology and Experimental Therapeutics Special Section on Transporters in Drug Disposition and Pharmacokinetic Prediction Interindividual Differences in the Expression of ATP-Binding Cassette and Solute Carrier Family Transporters in Human Skin: DNA Methylation Regulates Transcriptional Activity of the Human ABCC3 Gene s Tomoki Takechi, Takeshi Hirota, Tatsuya Sakai, Natsumi Maeda, Daisuke Kobayashi, and Ichiro Ieiri Downloaded from Department of Clinical Pharmacokinetics, Graduate School of Pharmaceutical Sciences, Kyushu University, Fukuoka, Japan (T.T., T.H., T.S., N.M., I.I.); Drug Development Research Laboratories, Kyoto R&D Center, Maruho Co., Ltd., Kyoto, Japan (T.T.); and Department of Clinical Pharmacy and Pharmaceutical Care, Graduate School of Pharmaceutical Sciences, Kyushu University, Fukuoka, Japan (D.K.) Received October 19, 2017; accepted January 30, 2018 dmd.aspetjournals.org ABSTRACT The identification of drug transporters expressed in human skin and levels. ABCC3 expression levels negatively correlated with the methylation interindividual differences in gene expression is important for understanding status of the CpG island (CGI) located approximately 10 kilobase pairs the role of drug transporters in human skin. In the present study, we upstream of ABCC3 (Rs: 20.323, P < 0.05). The reporter gene assay revealed evaluated the expression of ATP-binding cassette (ABC) and solute carrier a significant increase in transcriptional activity in the presence of CGI.
    [Show full text]
  • Regulation of MRP4 Expression by Circhipk3 Via Sponging Mir-124-3P/Mir-4524-5P in Hepatocellular Carcinoma
    biomedicines Article Regulation of MRP4 Expression by circHIPK3 via Sponging miR-124-3p/miR-4524-5p in Hepatocellular Carcinoma Haihong Hu †, Yu Wang †, Zhiyuan Qin, Wen Sun, Yanhong Chen, Jiaqi Wang, Yingying Wang, Jing Nie, Lu Chen, Sheng Cai, Lushan Yu * and Su Zeng * Cancer Center of Zhejiang University, Zhejiang Province Key Laboratory of Anti-Cancer Drug Research, Institute of Drug Metabolism and Pharmaceutical Analysis, College of Pharmaceutical Sciences, Zhejiang University, Hangzhou 310058, China; [email protected] (H.H.); [email protected] (Y.W.); [email protected] (Z.Q.); [email protected] (W.S.); [email protected] (Y.C.); [email protected] (J.W.); [email protected] (Y.W.); [email protected] (J.N.); [email protected] (L.C.); [email protected] (S.C.) * Correspondence: [email protected] (L.Y.); [email protected] (S.Z.); Tel.: +86-571-8820-8407 (L.Y.); +86-571-8820-8405 (S.Z.) † These authors contributed equally to this work. Abstract: Multidrug resistance-associated protein 4 (MRP4), a member of the adenosine triphosphate (ATP) binding cassette transporter family, pumps various molecules out of the cell and is involved in cell communication and drug distribution. Several studies have reported the role of miRNAs in downregulating the expression of MRP4. However, regulation of MRP4 by circular RNA (circRNA) Citation: Hu, H.; Wang, Y.; Qin, Z.; is yet to be elucidated. In this study, MRP4 was significantly upregulated in hepatocellular carcinoma Sun, W.; Chen, Y.; Wang, J.; Wang, Y.; (HCC) tissues compared to the adjacent noncancerous tissues.
    [Show full text]
  • Contribution of Abcc4-Mediated Gastric Transport to the Absorption and Efficacy of Dasatinib
    Published OnlineFirst June 21, 2013; DOI: 10.1158/1078-0432.CCR-13-0980 Clinical Cancer Cancer Therapy: Preclinical Research Contribution of Abcc4-Mediated Gastric Transport to the Absorption and Efficacy of Dasatinib Brian D. Furmanski1, Shuiying Hu1, Ken-ichi Fujita1, Lie Li1, Alice A. Gibson1, Laura J. Janke2, Richard T. Williams3, John D. Schuetz1, Alex Sparreboom1, and Sharyn D. Baker1 Abstract Purpose: Several oral multikinase inhibitors are known to interact in vitro with the human ATP-binding cassette transporter ABCC4 (MRP4), but the in vivo relevance of this interaction remains poorly understood. We hypothesized that host ABCC4 activity may influence the pharmacokinetic profile of dasatinib and subsequently affect its antitumor properties. Experimental Design: Transport of dasatinib was studied in cells transfected with human ABCC4 or the ortholog mouse transporter, Abcc4. Pharmacokinetic studies were done in wild-type and Abcc4-null mice. þ The influence of Abcc4 deficiency on dasatinib efficacy was evaluated in a model of Ph acute lymphoblastic leukemia by injection of luciferase-positive, p185(BCR-ABL)-expressing Arf(À/À) pre-B cells. Results: Dasatinib accumulation was significantly changed in cells overexpressing ABCC4 or Abcc4 compared with control cells (P < 0.001). Deficiency of Abcc4 in vivo was associated with a 1.75-fold decrease in systemic exposure to oral dasatinib, but had no influence on the pharmacokinetics of intravenous dasatinib. Abcc4 was found to be highly expressed in the stomach, and dasatinib efflux from isolated mouse stomachs ex vivo was impaired by Abcc4 deficiency (P < 0.01), without any detectable changes in gastric pH. Abcc4-null mice receiving dasatinib had an increase in leukemic burden, based on bioluminescence imaging, and decreased overall survival compared with wild-type mice (P ¼ 0.048).
    [Show full text]
  • Pleiotropic Roles of ABC Transporters in Breast Cancer
    International Journal of Molecular Sciences Review Pleiotropic Roles of ABC Transporters in Breast Cancer Ji He 1 , Erika Fortunati 1, Dong-Xu Liu 2 and Yan Li 1,2,3,* 1 School of Science, Auckland University of Technology, Auckland 1010, New Zealand; [email protected] (J.H.); [email protected] (E.F.) 2 The Centre for Biomedical and Chemical Sciences, School of Science, Faculty of Health and Environmental Sciences, Auckland University of Technology, Auckland 1010, New Zealand; [email protected] 3 School of Public Health and Interprofessional Studies, Auckland University of Technology, Auckland 0627, New Zealand * Correspondence: [email protected]; Tel.: +64-9921-9999 (ext. 7109) Abstract: Chemotherapeutics are the mainstay treatment for metastatic breast cancers. However, the chemotherapeutic failure caused by multidrug resistance (MDR) remains a pivotal obstacle to effective chemotherapies of breast cancer. Although in vitro evidence suggests that the overexpression of ATP-Binding Cassette (ABC) transporters confers resistance to cytotoxic and molecularly targeted chemotherapies by reducing the intracellular accumulation of active moieties, the clinical trials that target ABCB1 to reverse drug resistance have been disappointing. Nevertheless, studies indicate that ABC transporters may contribute to breast cancer development and metastasis independent of their efflux function. A broader and more clarified understanding of the functions and roles of ABC transporters in breast cancer biology will potentially contribute to stratifying patients for precision regimens and promote the development of novel therapies. Herein, we summarise the current knowledge relating to the mechanisms, functions and regulations of ABC transporters, with a focus on the roles of ABC transporters in breast cancer chemoresistance, progression and metastasis.
    [Show full text]
  • Role of Genetic Variation in ABC Transporters in Breast Cancer Prognosis and Therapy Response
    International Journal of Molecular Sciences Article Role of Genetic Variation in ABC Transporters in Breast Cancer Prognosis and Therapy Response Viktor Hlaváˇc 1,2 , Radka Václavíková 1,2, Veronika Brynychová 1,2, Renata Koževnikovová 3, Katerina Kopeˇcková 4, David Vrána 5 , Jiˇrí Gatˇek 6 and Pavel Souˇcek 1,2,* 1 Toxicogenomics Unit, National Institute of Public Health, 100 42 Prague, Czech Republic; [email protected] (V.H.); [email protected] (R.V.); [email protected] (V.B.) 2 Biomedical Center, Faculty of Medicine in Pilsen, Charles University, 323 00 Pilsen, Czech Republic 3 Department of Oncosurgery, Medicon Services, 140 00 Prague, Czech Republic; [email protected] 4 Department of Oncology, Second Faculty of Medicine, Charles University and Motol University Hospital, 150 06 Prague, Czech Republic; [email protected] 5 Department of Oncology, Medical School and Teaching Hospital, Palacky University, 779 00 Olomouc, Czech Republic; [email protected] 6 Department of Surgery, EUC Hospital and University of Tomas Bata in Zlin, 760 01 Zlin, Czech Republic; [email protected] * Correspondence: [email protected]; Tel.: +420-267-082-711 Received: 19 November 2020; Accepted: 11 December 2020; Published: 15 December 2020 Abstract: Breast cancer is the most common cancer in women in the world. The role of germline genetic variability in ATP-binding cassette (ABC) transporters in cancer chemoresistance and prognosis still needs to be elucidated. We used next-generation sequencing to assess associations of germline variants in coding and regulatory sequences of all human ABC genes with response of the patients to the neoadjuvant cytotoxic chemotherapy and disease-free survival (n = 105).
    [Show full text]
  • The MRP4/ABCC4 Gene Encodes a Novel Apical Organic Anion Transporter in Human Kidney Proximal Tubules: Putative Efflux Pump for Urinary Camp and Cgmp
    J Am Soc Nephrol 13: 595–603, 2002 The MRP4/ABCC4 Gene Encodes a Novel Apical Organic Anion Transporter in Human Kidney Proximal Tubules: Putative Efflux Pump for Urinary cAMP and cGMP RE´ MON A. M. H. VAN AUBEL,* PASCAL H. E. SMEETS,* JANNY G. P. PETERS,* RENE´ J. M. BINDELS,† and FRANS G. M. RUSSEL* Departments of *Pharmacology and Toxicology and †Cell Physiology, Nijmegen Center for Molecular Life Sciences, Nijmegen, The Netherlands. Abstract. The cyclic nucleotides cAMP and cGMP play key dependent transport of [3H]cAMP and [3H]cGMP. Both roles in cellular signaling and the extracellular regulation of probenecid and dipyridamole are potent MRP4 inhibitors. 3 3 fluid balance. In the kidney, cAMP is excreted across the apical ATP-dependent [ H]methotrexate and [ H]estradiol-17␤-D- proximal tubular membrane into urine, where it reduces phos- glucuronide transport by MRP4 and interactions with the phate reabsorption through a dipyridamole-sensitive mecha- anionic conjugates S-(2,4-dinitrophenyl)-glutathione, nism that is not fully understood. It has long been known that N-acetyl-(2,4-dinitrophenyl)-cysteine, ␣-naphthyl-␤-D- this cAMP efflux pathway is dependent on ATP and is glucuronide, and p-nitrophenyl-␤-D-glucuronide are also inhibited by probenecid. However, its identity and whether demonstrated. In kidneys of rats deficient in the apical cGMP shares the same transporter have not been estab- anionic conjugate efflux pump Mrp2, Mrp4 expression is lished. Here the expression, localization, and functional maintained at the same level. It is concluded that MRP4 is properties of human multidrug resistance protein 4 (MRP4) a novel apical organic anion transporter and the putative are reported.
    [Show full text]
  • Overexpression of MRP4 (ABCC4) and MRP5 (ABCC5) Confer Resistance to the Nucleoside Analogs Cytarabine and Troxacitabine, but No
    Adema et al. SpringerPlus 2014, 3:732 http://www.springerplus.com/content/3/1/732 a SpringerOpen Journal RESEARCH Open Access Overexpression of MRP4 (ABCC4) and MRP5 (ABCC5) confer resistance to the nucleoside analogs cytarabine and troxacitabine, but not gemcitabine Auke D Adema1, Karijn Floor1, Kees Smid1, Richard J Honeywell1, George L Scheffer2, Gerrit Jansen3 and Godefridus J Peters1* Abstract We aimed to determine whether the multidrug-resistance-proteins MRP4 (ABCC4) and MRP5 (ABCC5) confer resistance to the antimetabolites cytarabine (Ara-C), gemcitabine (GEM), and the L-nucleoside analog troxacitabine. For this purpose we used HEK293 and the transfected HEK/MRP4 (59-fold increased MRP4) or HEK/MRP5i (991-fold increased MRP5) as model systems and tested the cells for drug sensitivity using a proliferation test. Drug accumulation was performed by using radioactive Ara-C, and for GEM and troxacitabine with HPLC with tandem-MS or UV detection. At 4-hr exposure HEK/MRP4 cells were 2-4-fold resistant to troxacitabine, ara-C and 9-(2-phosphonylmethoxyethyl)adenine (PMEA), and HEK/MRP5i to ara-C and PMEA, but none to GEM. The inhibitors probenecid and indomethacin reversed resistance. After 4-hr exposure ara-C-nucleotides were 2-3-fold lower in MRP4/5 cells, in which they decreased more rapidly after washing with drug-free medium (DFM). Trocacitabine accumulation was similar in the 3 cell lines, but after the DFM period troxacitabine decreased 2-4-fold faster in MRP4/5 cells. Troxacitabine-nucleotides were about 25% lower in MRP4/5 cells and decreased rapidly in MRP4, but not in MRP5 cells.
    [Show full text]
  • Identification of ABCG2 As an Exporter of Uremic Toxin Indoxyl Sulfate in Mice and As a Crucial Factor Influencing CKD Progressi
    www.nature.com/scientificreports OPEN Identifcation of ABCG2 as an Exporter of Uremic Toxin Indoxyl Sulfate in Mice and as a Crucial Received: 6 March 2018 Accepted: 6 July 2018 Factor Infuencing CKD Progression Published: xx xx xxxx T. Takada1, T. Yamamoto1, H. Matsuo2, J. K. Tan1, K. Ooyama3, M. Sakiyama2, H. Miyata1, Y. Yamanashi1, Y. Toyoda 1, T. Higashino2, A. Nakayama2, A. Nakashima4, N. Shinomiya2, K. Ichida5, H. Ooyama6, S. Fujimori7 & H. Suzuki1 Chronic kidney disease (CKD) patients accumulate uremic toxins in the body, potentially require dialysis, and can eventually develop cardiovascular disease. CKD incidence has increased worldwide, and preventing CKD progression is one of the most important goals in clinical treatment. In this study, we conducted a series of in vitro and in vivo experiments and employed a metabolomics approach to investigate CKD. Our results demonstrated that ATP-binding cassette transporter subfamily G member 2 (ABCG2) is a major transporter of the uremic toxin indoxyl sulfate. ABCG2 regulates the pathophysiological excretion of indoxyl sulfate and strongly afects CKD survival rates. Our study is the frst to report ABCG2 as a physiological exporter of indoxyl sulfate and identify ABCG2 as a crucial factor infuencing CKD progression, consistent with the observed association between ABCG2 function and age of dialysis onset in humans. The above fndings provided valuable knowledge on the complex regulatory mechanisms that regulate the transport of uremic toxins in our body and serve as a basis for preventive and individualized treatment of CKD. Chronic kidney disease (CKD) is a disease characterized by chronically impaired kidney function and is attrib- uted to various causes.
    [Show full text]
  • Substrate Overlap Between Mrp4 and Abcg2/Bcrp Affects Purine Analogue Drug Cytotoxicity and Tissue Distribution
    Research Article Substrate Overlap between Mrp4 and Abcg2/Bcrp Affects Purine Analogue Drug Cytotoxicity and Tissue Distribution Kazumasa Takenaka,1 Jessica A. Morgan,1 George L. Scheffer,2,3 Masashi Adachi,1 Clinton F. Stewart,1 Daxi Sun,1 Markos Leggas,1 Karin F.K. Ejendal,4 Christine A. Hrycyna,4 and John D. Schuetz1 1Department of Pharmaceutical Sciences, St. Jude Children’s Research Hospital, Memphis, Tennessee; 2Department of Pathology, VU Medical Center and 3Division of Molecular Biology, The Netherlands Cancer Institute, Amsterdam, the Netherlands; and 4Department of Chemistry and the Purdue Cancer Center, Purdue University, West Lafayette, Indiana Abstract interactions amongABC transporters is to evaluate both substrate The use of probe substrates and combinations of ATP-binding and transporter tissue distribution in knockout (KO) animals. We cassette (ABC) transporter knockout (KO) animals may speculated that if two transporters showed similar tissue facilitate the identification of common substrates between distribution and patterns of up-regulation in each others absence, apparently unrelated ABC transporters.An unexpectedly low then it was likely they shared similar endogenous or drug concentration of the purine nucleotide analogue, 9-(2-(phos- substrates. Mrp4 (also known as Abcc4) and Abcg2 have similar phonomethoxy)ethyl)-adenine (PMEA), and up-regulation of patterns of tissue distribution (3, 4). Mrp4 was first identified as a Abcg2 in some tissues of the Mrp4 KO mouse prompted us to transporter of nucleoside monophosphates [primarily purine evaluate the possibility that Abcg2 might transport purine- nucleoside monophosphates; e.g., 9-(2-(phosphonomethoxy)ethyl)- adenine (PMEA); ref. 5], but more recent studies have indicated derived drugs.Abcg2 transported and conferred resistance to PMEA.Moreover, a specific Abcg2 inhibitor, fumitremorgin C, that Mrp4 transports substrates in common with Abcg2, most both increased PMEA accumulation and reversed Abcg2- recently camptothecin analogues (6, 7).
    [Show full text]
  • ABC. See ATP-Binding Cassette (ABC) ABC Transporters, 12, 107
    325 Index a amidotransferases, 263 ABC. see ATP-binding cassette (ABC) – γ-glutamyl transpeptidase, 263 ABC transporters, 12, 107 amine hypothesis, 21 – effects on drug disposition, 112 amino acidpolyamine-organocation – mutations, 113 superfamily, 16 – neurological disorders, role in, 180–181 amino acid/polyamine/organocation – PET imaging, 181–183 transporter (APC), 262 – PET tracers amino acid transporters, 254 –– applications, 184–185 γ-aminobutyric acid (GABA), 24, 69 –– designing, challenges in, 184 – neurotransmission, 85 – pharmacochaperones and, 113, 114 – signaling, 73, 85 – proteins, 7 aminophospholipid, 204 – role of, 108 ammonia channel transporter (Amt) family, 7 – superfamily, 2 amyloid precursor protein (APP), 180 – from targets to antitargets, 111–113 anaesthetics, 1 absorption, disposition, metabolism and anion exchanger, 144 excretion (ADME), 253 anoctamin 1 (ANO1), 231, 233, 234 – SLC transporters, importance in, 253 – activators of, 240 acetylcholine, 41 – biophysical properties of, 234, 235 ADME. see absorption, disposition, metabolism – calcium activated chloride channel, 232 and excretion (ADME) – and cancer, 236–238 age-related insensitivity, 40 – characterization of, 232 alanine-serine-cysteine transporter 1 – as contributor to renal cyst growth, 245 (ASCT1), 260 – cystic fibrosis-related diabetes (CFRD), 232, alanine-serine-cysteine transporter 2 245 (ASCT2), 253, 260 – discovery of, 232, 233 Alisma orientalis, 212, 213 – effect on motility of human cancer cells, 237 allosteric effect – expression and physiological
    [Show full text]