uu.seUppsala University Publications
Change search
ReferencesLink to record
Permanent link

Direct link
Endogenous Gene and Protein Expression of Drug Transporting Proteins in Cell Lines Routinely used in Drug Discovery Programs
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmacy. (Läkemedelsformulering)
AstraZeneca R&D, Mölndal.
AstraZeneca R&D, Mölndal, Sweden.
Department of Proteomics, The Royal Institute of Technology, Stockholm, Sweden.
Show others and affiliations
2009 (English)In: Drug Metabolism And Disposition, ISSN 0090-9556, E-ISSN 1521-009X, Vol. 37, no 12, 2275-2283 p.Article in journal (Refereed) Published
Abstract [en]

The aim of this study was to investigate the gene and protein expression profiles of important drug transporting proteins in human cell lines commonly used for studies of drug transport mechanisms. Human cell lines used to transiently or stably express single transporters (HeLa, HEK293) and leukaemia cell lines used to study drug resistance by ABC-transporters (HL-60, K562) were investigated, and compared with organotypic cell lines (HepG2, Saos-2, Caco-2 and Caco-2 TC7). For gene expression studies, real-time PCR was used, while monospecific polyclonal antibodies were generated and used to investigate protein expression by immunohistochemistry. Thirty-six transporters were studied for gene expression and nine for protein expression. The antibodies were validated using expression patterns in human tissues. Finally, the function of one ubiquitously expressed transporter, MCT1; SLC16A1 was investigated using 14C-lactic acid as a substrate. In general, the adherent cell lines (HeLa, HEK293) displayed low transporter expression and the expression patterns were barely affected by transfection. The leukaemia cell lines (K562, HL-60) and Saos-2 also had low endogenous transporter expression, while the organotypic cell lines (HepG2 and Caco-2) showed higher expression of some transporters. Comparison of gene and protein expression profiles gave poor correlations, but better agreement was obtained for antibodies with a good validation score, indicating that antibody quality was a significant variable. Importantly, the monocarboxylic acid transporting protein MCT1 was significantly expressed in all, and functional in most of the cell lines, indicating that MCT1 may be a confounding factor when the transport of small anionic drugs is investigated.

Place, publisher, year, edition, pages
2009. Vol. 37, no 12, 2275-2283 p.
Keyword [en]
Cell lines, Caco-2, HEK293, HeLa, Saos-2, HL-60, K562, HepG2, Gene expression, Protein expression, MCT1
National Category
Pharmaceutical Sciences
Research subject
Biopharmaceutics; Pharmaceutics
URN: urn:nbn:se:uu:diva-107571DOI: 10.1124/dmd.109.028654ISI: 000271935200002PubMedID: 19741037OAI: oai:DiVA.org:uu-107571DiVA: diva2:231770
Available from: 2009-08-17 Created: 2009-08-17 Last updated: 2010-06-28Bibliographically approved
In thesis
1. In vitro and in silico prediction of drug-drug interactions with transport proteins
Open this publication in new window or tab >>In vitro and in silico prediction of drug-drug interactions with transport proteins
2009 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Drug transport across cells and cell membranes in the human body is crucial for the pharmacological effect of drugs. Active transport governed by transport proteins plays an important role in this process. A vast number of transport proteins with a wide tissue distribution have been identified during the last 15 years. Several important examples of their role in drug disposition and drug-drug interactions have been described to date. Investigation of drug-drug interactions at the transport protein level are therefore of increasing interest to the academic, industrial and regulatory research communities.

The gene expression of transport proteins involved in drug transport was investigated in the jejunum, liver, kidney and colon to better understand their influence on the ADMET properties of drugs. In addition, the gene and protein expression of transport proteins in cell lines, widely used for predictions of drug transport and metabolism, was examined.

The substrate and inhibitor heterogeneity of many transport proteins makes it difficult to foresee whether the transport proteins will cause drug-drug interactions. Therefore, in vitro assays for OCT1 and OATP1B1, among the highest expressed transport proteins in human liver, were developed to allow investigation of the inhibitory patterns of these proteins. These assays were used to investigate two data sets, consisting of 191 and 135 registered drugs and drug-like molecules for the inhibition of OCT1 and OATP1B1, respectively. Numerous new inhibitors of the transport proteins were identified in the data sets and the properties governing inhibition were determined. Further, antidepressant drugs and statins displayed strong inhibition of OCT1 and OATP1B1, respectively. The inhibition data was used to develop predictive in silico models for each of the two transport proteins.

The highly polymorphic nature of some transport proteins has been shown to affect drug response and may lead to an increased risk of drug-drug interactions, and therefore, the OCT1 in vitro assay was used to study the effect of common genetic variants of OCT1 on drug inhibition and drug-drug interactions. The results indicated that OCT1 variants with reduced function were more susceptible to inhibition. Further, a drug-drug interaction of potential clinical significance in the genetic OCT1 variant M420del was proposed.

In summary, gene expression of transport proteins was investigated in human tissues and cell lines. In vitro assays for two of the highest expressed liver transport proteins were used to identify previously unknown SLC transport protein inhibitors and to develop predictive in silico models, which may detect previously known drug-drug interactions and enable new ones to be identified at the transport protein level. In addition, the effect of genetic variation on inhibition of the OCT1 was investigated.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2009. 65 p.
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Pharmacy, ISSN 1651-6192 ; 104
Solute carrier, SLC transporter, OCT1, Organic cation transporter 1, SLC22A1, OATP1B1, SLCO1B1, Organic anion transporting peptide 1B1, Drug transport, Active transport, Genetic polymorphism, Cell lines, Gene expression, Multivariate data analysis, OPLS
National Category
Pharmaceutical Sciences Pharmaceutical Sciences
Research subject
Pharmaceutics; Biopharmaceutics
urn:nbn:se:uu:diva-107492 (URN)978-91-554-7589-5 (ISBN)
Public defence
2009-10-02, B21, Biomedical Center, BMC, Husargatan 3, Uppsala, 13:15 (English)
Available from: 2009-09-10 Created: 2009-08-13 Last updated: 2009-09-16Bibliographically approved

Open Access in DiVA

No full text

Other links

Publisher's full textPubMed

Search in DiVA

By author/editor
Ahlin, Gustav
By organisation
Department of Pharmacy
In the same journal
Drug Metabolism And Disposition
Pharmaceutical Sciences

Search outside of DiVA

GoogleGoogle Scholar
The number of downloads is the sum of all downloads of full texts. It may include eg previous versions that are now no longer available

Altmetric score

Total: 308 hits
ReferencesLink to record
Permanent link

Direct link