Involvement of human and canine MRP1 and MRP4 in benzylpenicillin transport

Autoři: Xiaofen Zhao aff001;  Yangfang Li aff001;  Kun Du aff001;  Yuqin Wu aff001;  Ling Liu aff001;  Shan Cui aff001;  Yan Zhang aff001;  Jin Gao aff001;  Richard F. Keep aff002;  Jianming Xiang aff002
Působiště autorů: Department of Neonate, Kunming Children’s Hospital, Kunming, China aff001;  Department of Neurosurgery, University of Michigan Medical School, Ann Arbor, Michigan, United States of America aff002
Vyšlo v časopise: PLoS ONE 14(11)
Kategorie: Research Article
doi: 10.1371/journal.pone.0225702


The blood-brain barrier (BBB) is a dynamic and complex interface between blood and the central nervous system (CNS). It protects the brain by preventing toxic substances from entering the brain but also limits the entry of therapeutic agents. ATP-binding cassette (ABC) efflux transporters are critical for the functional barrier and present a formidable impediment to brain delivery of therapeutic agents including antibiotics. The aim of this study was to investigate the possible involvement of multidrug resistance-associated protein 1 and 4 (MRP1 and MRP4), two ABC transporters, in benzylpenicillin efflux transport using wild-type (WT) MDCKII cells and cells overexpressing those human transporters, as well as non-selective and selective inhibitors. We found that inhibiting MRP1 or MRP4 significantly increased [3H]benzylpenicillin uptake in MDCKII-WT, -MRP1 or –MRP4 cells. Similar results were also found in HepG2 cells, which highly express MRP1 and MRP4, and hCMEC/D3 cells which express MRP1. The results indicate that human and canine MRP1 and MRP4 are involved in benzylpenicillin efflux transport. They could be potential therapeutic targets for improving the efficacy of benzylpenicillin for treating CNS infections since both MRP1 and MRP4 express at human blood-brain barrier.

Klíčová slova:

Antibiotic resistance – Antibiotics – Cancer treatment – Central nervous system – Endothelial cells – RNA synthesis – Toxic agents – Blood-brain barrier


1. Abbott NJ, Patabendige AA, Dolman DE, Yusof SR, Begley DJ. Structure and function of the blood-brain barrier. [Review] [168 refs].

2. Obermeier B, Verma A, Ransohoff RM. The blood-brain barrier. Handbook of clinical neurology. 2016;133:39–59. Epub 2016/04/27. doi: 10.1016/B978-0-444-63432-0.00003-7 27112670.

3. Serlin Y, Shelef I, Knyazer B, Friedman A. Anatomy and physiology of the blood-brain barrier. Seminars in cell & developmental biology. 2015;38:2–6. Epub 2015/02/15. doi: 10.1016/j.semcdb.2015.01.002 25681530; PubMed Central PMCID: PMC4397166.

4. Hartz AM, Bauer B. ABC transporters in the CNS—an inventory. Curr Pharm Biotechnol. 2011;12(4):656–73. Epub 2010/12/02. doi: 10.2174/138920111795164020 21118088.

5. Loscher W, Potschka H. Blood-brain barrier active efflux transporters: ATP-binding cassette gene family. NeuroRx: the journal of the American Society for Experimental NeuroTherapeutics. 2005;2(1):86–98. Epub 2005/02/18. doi: 10.1602/neurorx.2.1.86 15717060; PubMed Central PMCID: PMC539326.

6. Mahringer A, Fricker G. ABC transporters at the blood-brain barrier. Expert opinion on drug metabolism & toxicology. 2016;12(5):499–508. Epub 2016/03/22. doi: 10.1517/17425255.2016.1168804 26998936.

7. Agarwal S, Hartz AM, Elmquist WF, Bauer B. Breast cancer resistance protein and P-glycoprotein in brain cancer: two gatekeepers team up. Current pharmaceutical design. 2011;17(26):2793–802. Epub 2011/08/11. doi: 10.2174/138161211797440186 21827403; PubMed Central PMCID: PMC3269897.

8. Borst P, Schinkel AH. P-glycoprotein ABCB1: a major player in drug handling by mammals. The Journal of clinical investigation. 2013;123(10):4131–3. Epub 2013/10/03. doi: 10.1172/JCI70430 24084745; PubMed Central PMCID: PMC3784548.

9. Eilers M, Roy U, Mondal D. MRP (ABCC) transporters-mediated efflux of anti-HIV drugs, saquinavir and zidovudine, from human endothelial cells. Experimental biology and medicine (Maywood, NJ). 2008;233(9):1149–60. Epub 2008/06/07. doi: 10.3181/0802-rm-59 18535159; PubMed Central PMCID: PMC2575034.

10. Iorio AL, da Ros M, Fantappie O, Lucchesi M, Facchini L, Stival A, et al. Blood-Brain Barrier and Breast Cancer Resistance Protein: a limit to the therapy of CNS tumors and neurodegenerative diseases. Anti-cancer agents in medicinal chemistry. 2015. Epub 2015/11/21. doi: 10.2174/1871520616666151120121928 26584727.

11. Kim RB. Drugs as P-glycoprotein substrates, inhibitors, and inducers. Drug metabolism reviews. 2002;34(1–2):47–54. Epub 2002/05/09. doi: 10.1081/dmr-120001389 11996011.

12. Lagas JS, van Waterschoot RA, Sparidans RW, Wagenaar E, Beijnen JH, Schinkel AH. Breast cancer resistance protein and P-glycoprotein limit sorafenib brain accumulation. Molecular cancer therapeutics. 2010;9(2):319–26. Epub 2010/01/28. doi: 10.1158/1535-7163.MCT-09-0663 20103600.

13. Potschka H, Fedrowitz M, Loscher W. Multidrug resistance protein MRP2 contributes to blood-brain barrier function and restricts antiepileptic drug activity. The Journal of pharmacology and experimental therapeutics. 2003;306(1):124–31. Epub 2003/03/29. doi: 10.1124/jpet.103.049858 12663688.

14. Shen S, Zhang W. ABC transporters and drug efflux at the blood-brain barrier. Rev Neurosci. 2010;21(1):29–53. Epub 2010/05/13 06:00. doi: 10.1515/revneuro.2010.21.1.29 20458886.

15. Acharya P, Tran TT, Polli JW, Ayrton A, Ellens H, Bentz J. P-Glycoprotein (P-gp) expressed in a confluent monolayer of hMDR1-MDCKII cells has more than one efflux pathway with cooperative binding sites. Biochemistry. 2006;45(51):15505–19. doi: 10.1021/bi060593b 17176072.

16. Azad TD, Pan J, Connolly ID, Remington A, Wilson CM, Grant GA. Therapeutic strategies to improve drug delivery across the blood-brain barrier. Neurosurgical focus. 2015;38(3):E9. Epub 2015/03/03. doi: 10.3171/2014.12.FOCUS14758 25727231; PubMed Central PMCID: PMC4493051.

17. Fricker G, Miller DS. Modulation of drug transporters at the blood-brain barrier. [Review] [61 refs].

18. Hartz AM, Bauer B. Regulation of ABC transporters at the blood-brain barrier: new targets for CNS therapy. Molecular interventions. 2010;10(5):293–304. Epub 2010/11/04. doi: 10.1124/mi.10.5.6 21045243.

19. Miller DS. Regulation of ABC transporters blood-brain barrier: the good, the bad, and the ugly. Advances in cancer research. 2015;125:43–70. Epub 2015/02/03. doi: 10.1016/bs.acr.2014.10.002 25640266.

20. Roberts JA, Webb S, Paterson D, Ho KM, Lipman J. A systematic review on clinical benefits of continuous administration of beta-lactam antibiotics. Critical care medicine. 2009;37(6):2071–8. Epub 2009/04/23. doi: 10.1097/CCM.0b013e3181a0054d 19384201.

21. Tomasz A. The mechanism of the irreversible antimicrobial effects of penicillins: how the beta-lactam antibiotics kill and lyse bacteria. Annual review of microbiology. 1979;33:113–37. Epub 1979/01/01. doi: 10.1146/annurev.mi.33.100179.000553 40528.

22. Ramos SR, Feferbaum R, Manissadjian A, Vaz FA. [Neonatal bacterial meningitis: etiological agents in 109 cases during a 10 year period]. Arquivos de neuro-psiquiatria. 1992;50(3):289–94. Epub 1992/09/01. doi: 10.1590/s0004-282x1992000300005 1308405.

23. Zhu ML, Mai JY, Zhu JH, Lin ZL. [Clinical analysis of 31 cases of neonatal purulent meningitis caused by Escherichia coli]. Zhongguo dang dai er ke za zhi = Chinese journal of contemporary pediatrics. 2012;14(12):910–2. Epub 2012/12/14. 23234776.

24. Norrby SR. Problems in evaluation of adverse reactions to beta-lactam antibiotics. Reviews of infectious diseases. 1986;8 Suppl 3:S358–70. Epub 1986/07/01. doi: 10.1093/clinids/8.supplement_3.s358 3529328.

25. Schliamser SE. Neurotoxicity of beta-lactam antibiotics. Experimental kinetic and neurophysiological studies. Scandinavian journal of infectious diseases Supplementum. 1988;55:1–61. Epub 1988/01/01. doi: 10.3109/inf.1988.20.suppl-55.01 3241957.

26. Rousselle C, Clair P, Temsamani J, Scherrmann JM. Improved brain delivery of benzylpenicillin with a peptide-vector-mediated strategy. Journal of drug targeting. 2002;10(4):309–15. Epub 2002/08/08. doi: 10.1080/10611860290031886 12164379.

27. Ambudkar SV, Kimchi-Sarfaty C, Sauna ZE, Gottesman MM. P-glycoprotein: from genomics to mechanism. Oncogene. 2003;22(47):7468–85. Epub 2003/10/25. doi: 10.1038/sj.onc.1206948 14576852.

28. Poelarends GJ, Mazurkiewicz P, Putman M, Cool RH, Veen HW, Konings WN. An ABC-type multidrug transporter of Lactococcus lactis possesses an exceptionally broad substrate specificity. Drug resistance updates: reviews and commentaries in antimicrobial and anticancer chemotherapy. 2000;3(6):330–4. Epub 2001/08/11. doi: 10.1054/drup.2000.0173 11498401.

29. Seelig A. How does P-glycoprotein recognize its substrates? International journal of clinical pharmacology and therapeutics. 1998;36(1):50–4. Epub 1998/02/26. 9476149.

30. Siarheyeva A, Lopez JJ, Glaubitz C. Localization of multidrug transporter substrates within model membranes. Biochemistry. 2006;45(19):6203–11. Epub 2006/05/10. doi: 10.1021/bi0524870 16681393.

31. Choi MK, Kim H, Han YH, Song IS, Shim CK. Involvement of Mrp2/MRP2 in the species different excretion route of benzylpenicillin between rat and human. Xenobiotica. 2009;39(2):171–81. Epub 2009/03/04. doi: 10.1080/00498250802642256 19255943.

32. Li Y, Wu Q, Li C, Liu L, Du K, Shen J, et al. Role of Human Breast Cancer Related Protein versus P-Glycoprotein as an Efflux Transporter for Benzylpenicillin: Potential Importance at the Blood-Brain Barrier. PloS one. 2016;11(6):e0157576. Epub 2016/06/15. doi: 10.1371/journal.pone.0157576 27300692; PubMed Central PMCID: PMC4907523.

33. Helms HC, Hersom M, Kuhlmann LB, Badolo L, Nielsen CU, Brodin B. An electrically tight in vitro blood-brain barrier model displays net brain-to-blood efflux of substrates for the ABC transporters, P-gp, Bcrp and Mrp-1. The AAPS journal. 2014;16(5):1046–55. Epub 2014/06/18. doi: 10.1208/s12248-014-9628-1 24934296; PubMed Central PMCID: PMC4147044.

34. Tivnan A, Zakaria Z, O'Leary C, Kogel D, Pokorny JL, Sarkaria JN, et al. Inhibition of multidrug resistance protein 1 (MRP1) improves chemotherapy drug response in primary and recurrent glioblastoma multiforme. Frontiers in neuroscience. 2015;9:218. Epub 2015/07/03. doi: 10.3389/fnins.2015.00218 26136652; PubMed Central PMCID: PMC4468867.

35. Cheung L, Flemming CL, Watt F, Masada N, Yu DM, Huynh T, et al. High-throughput screening identifies Ceefourin 1 and Ceefourin 2 as highly selective inhibitors of multidrug resistance protein 4 (MRP4). Biochemical pharmacology. 2014;91(1):97–108. Epub 2014/06/29. doi: 10.1016/j.bcp.2014.05.023 24973542.

36. Usuki F, Fujimura M, Yamashita A. Endoplasmic reticulum stress preconditioning modifies intracellular mercury content by upregulating membrane transporters. Scientific reports. 2017;7(1):12390. Epub 2017/09/30. doi: 10.1038/s41598-017-09435-3 28959040; PubMed Central PMCID: PMC5620048.

37. Sindac JA, Barraza SJ, Dobry CJ, Xiang J, Blakely PK, Irani DN, et al. Optimization of novel indole-2-carboxamide inhibitors of neurotropic alphavirus replication. Journal of medicinal chemistry. 2013;56(22):9222–41. Epub 2013/10/25. doi: 10.1021/jm401330r 24151954; PubMed Central PMCID: PMC3895407.

38. Kuteykin-Teplyakov K, Luna-Tortos C, Ambroziak K, Loscher W. Differences in the expression of endogenous efflux transporters in MDR1-transfected versus wildtype cell lines affect P-glycoprotein mediated drug transport. British journal of pharmacology. 2010;160(6):1453–63. Epub 2010/07/02. doi: 10.1111/j.1476-5381.2010.00801.x 20590635; PubMed Central PMCID: PMC2938816.

39. Pascolo L, Fernetti C, Pirulli D, Bogoni S, Garcia-Mediavilla MV, Spano A, et al. Detection of MRP1 mRNA in human tumors and tumor cell lines by in situ RT-PCR. Biochemical and biophysical research communications. 2000;275(2):466–71. Epub 2000/08/31. doi: 10.1006/bbrc.2000.3339 10964688.

40. Rigalli JP, Ciriaci N, Arias A, Ceballos MP, Villanueva SS, Luquita MG, et al. Regulation of multidrug resistance proteins by genistein in a hepatocarcinoma cell line: impact on sorafenib cytotoxicity. PloS one. 2015;10(3):e0119502. Epub 2015/03/18. doi: 10.1371/journal.pone.0119502 25781341; PubMed Central PMCID: PMC4364073.

41. Dauchy S, Miller F, Couraud PO, Weaver RJ, Weksler B, Romero IA, et al. Expression and transcriptional regulation of ABC transporters and cytochromes P450 in hCMEC/D3 human cerebral microvascular endothelial cells. Biochemical pharmacology. 2009;77(5):897–909. Epub 2008/12/02. doi: 10.1016/j.bcp.2008.11.001 19041851.

42. Ohtsuki S, Ikeda C, Uchida Y, Sakamoto Y, Miller F, Glacial F, et al. Quantitative targeted absolute proteomic analysis of transporters, receptors and junction proteins for validation of human cerebral microvascular endothelial cell line hCMEC/D3 as a human blood-brain barrier model. Molecular pharmaceutics. 2013;10(1):289–96. Epub 2012/11/10. doi: 10.1021/mp3004308 23137377.

43. Schliamser SE, Cars O, Norrby SR. Neurotoxicity of beta-lactam antibiotics: predisposing factors and pathogenesis. J Antimicrob Chemother. 1991;27(4):405–25. Epub 1991/04/01. doi: 10.1093/jac/27.4.405 1856121.

44. Oberoi RK, Parrish KE, Sio TT, Mittapalli RK, Elmquist WF, Sarkaria JN. Strategies to improve delivery of anticancer drugs across the blood-brain barrier to treat glioblastoma. Neuro-oncology. 2016;18(1):27–36. Epub 2015/09/12. doi: 10.1093/neuonc/nov164 26359209; PubMed Central PMCID: PMC4677418.

45. Pardridge WM. Blood-brain barrier endogenous transporters as therapeutic targets: a new model for small molecule CNS drug discovery. Expert opinion on therapeutic targets. 2015;19(8):1059–72. Epub 2015/05/06. doi: 10.1517/14728222.2015.1042364 25936389.

Článek vyšel v časopise


2019 Číslo 11