Expression of SLC22A1 variants may affect the response of hepatocellular carcinoma and cholangiocarcinoma to sorafenib

Authors

  • Elisa Herraez,

    1. Laboratory of Experimental Hepatology and Drug Targeting (HEVEFARM), Biomedical Research Institute of Salamanca (IBSAL), University of Salamanca, Salamanca, Spain
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  • Elisa Lozano,

    1. Laboratory of Experimental Hepatology and Drug Targeting (HEVEFARM), Biomedical Research Institute of Salamanca (IBSAL), University of Salamanca, Salamanca, Spain
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  • Rocio I.R. Macias,

    1. Laboratory of Experimental Hepatology and Drug Targeting (HEVEFARM), Biomedical Research Institute of Salamanca (IBSAL), University of Salamanca, Salamanca, Spain
    2. National Institute for the Study of Liver and Gastrointestinal Diseases (CIBERehd), Spain
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  • Javier Vaquero,

    1. Laboratory of Experimental Hepatology and Drug Targeting (HEVEFARM), Biomedical Research Institute of Salamanca (IBSAL), University of Salamanca, Salamanca, Spain
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  • Luis Bujanda,

    1. National Institute for the Study of Liver and Gastrointestinal Diseases (CIBERehd), Spain
    2. Department of Liver Diseases, Biodonostia Research Institute (Donostia University Hospital), IKEBASQUE (Basque Foundation for Science), University of Basque Country (UPV/EHU), San Sebastian, Spain
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  • Jesus M. Banales,

    1. National Institute for the Study of Liver and Gastrointestinal Diseases (CIBERehd), Spain
    2. Department of Liver Diseases, Biodonostia Research Institute (Donostia University Hospital), IKEBASQUE (Basque Foundation for Science), University of Basque Country (UPV/EHU), San Sebastian, Spain
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  • Jose J.G. Marin,

    Corresponding author
    1. National Institute for the Study of Liver and Gastrointestinal Diseases (CIBERehd), Spain
    • Laboratory of Experimental Hepatology and Drug Targeting (HEVEFARM), Biomedical Research Institute of Salamanca (IBSAL), University of Salamanca, Salamanca, Spain
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  • Oscar Briz

    1. Laboratory of Experimental Hepatology and Drug Targeting (HEVEFARM), Biomedical Research Institute of Salamanca (IBSAL), University of Salamanca, Salamanca, Spain
    2. National Institute for the Study of Liver and Gastrointestinal Diseases (CIBERehd), Spain
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  • Potential conflict of interest: Nothing to report.

  • Supported in part by the Junta de Castilla y Leon (Grants BIO39/SA27/10, SA023A11-2 and SA070A11-2), Spain; the Ministerio de Ciencia y Tecnologia (Grants SAF2009-08493 and SAF2010-15517), Spain; the Fundacion Investigacion Medica Mutua Madrileña (Call 2009), Spain, and Fundacion Samuel Solorzano Barruso (Grant FS/1-2011), Spain. E.L. was supported by the AP2008-0376 PhD grant from the Junta de Castilla y Leon/Fondo Social Europeo and J.V. by the AP2007-00105 PhD grant from the Ministerio de Educacion (AP2007-00105), Spain. J.M.B. and L.B. received grants from the Spanish Association Against Cancer (AECC), the Basque Department of Industry (Saiotek), and the Carlos III Health Institute, Spain (FIS grant PI12/00380).

Address reprint requests to: Jose J.G. Marin, University of Salamanca, Department of Physiology and Pharmacology, Campus Miguel de Unamuno, E.D. S-09, 37007 - Salamanca, Spain. E-mail: jjgmarin@usal.es; fax: +34-923-294669.

Abstract

Reduced drug uptake is an important mechanism of chemoresistance. Down-regulation of SLC22A1 encoding the organic cation transporter-1 (OCT1) may affect the response of hepatocellular carcinoma (HCC) and cholangiocarcinoma (CGC) to sorafenib, a cationic drug. Here we investigated whether SLC22A1 variants may contribute to sorafenib chemoresistance. Complete sequencing and selective variant identification were carried out to detect single nucleotide polymorphisms (SNPs) in SLC22A1 complementary DNA (cDNA). In HCC and CGC biopsies, in addition to previously described variants, two novel alternative spliced variants and three SNPs were identified. To study their functional consequences, these variants were mimicked by directed mutagenesis and expressed in HCC (Alexander and SK-Hep-1) and CGC (TFK1) cells. The two novel described variants, R61S fs*10 and C88A fs*16, encoded truncated proteins unable to reach the plasma membrane. Both variants abolished OCT1-mediated uptake of tetraethylammonium, a typical OCT1 substrate, and were not able to induce sorafenib sensitivity. In cells expressing functional OCT1 variants, OCT1 inhibition with quinine prevented sorafenib-induced toxicity. Expression of OCT1 variants in Xenopus laevis oocytes and determination of quinine-sensitive sorafenib uptake by high-performance liquid chromatography-dual mass spectrometry confirmed that OCT1 is able to transport sorafenib and that R61S fs*10 and C88A fs*16 abolish this ability. Screening of these SNPs in 23 HCC and 15 CGC biopsies revealed that R61S fs*10 was present in both HCC (17%) and CGC (13%), whereas C88A fs*16 was only found in HCC (17%). Considering all SLC22A1 variants, at least one inactivating SNP was found in 48% HCC and 40% CGC. Conclusion: Development of HCC and CGC is accompanied by the appearance of aberrant OCT1 variants that, together with decreased OCT1 expression, may dramatically affect the ability of sorafenib to reach active intracellular concentrations in these tumors. (Hepatology 2013;53:1065–1073)

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