Characterization of the organic cation transporter SLC22A16: A doxorubicin importer

doi:10.1016/j.bbrc.2005.05.174

Biochemical and Biophysical Research Communications

Volume 333, Issue 3, 5 August 2005, Pages 754-762

https://doi.org/10.1016/j.bbrc.2005.05.174 Get rights and content

Abstract

Specific efflux transporters, such as P-glycoprotein, have been shown to confer drug resistance by decreasing the intracellular accumulation of anticancer drugs. Understanding influx transporters, as well as efflux transporters, is essential to overcome this resistance. We report the expression profile and pharmacological characterization of an organic cation transporter, SLC22A16. The results of our experiments indicate that SLC22A16 is a mediator of doxorubicin uptake in cancer cells. Quantitative real-time RT-PCR analyses show that SLC22A16 is expressed in primary samples taken from patients with acute leukemia. Xenopus oocytes injected with SLC22A16 cRNA import doxorubicin, a widely used anticancer drug for hematological malignancies, in a saturable and dose-dependent manner. The apparent K_m value for doxorubicin import was 5.2 ± 0.4 μM. In cytotoxic assays, stable transfectants of leukemic Jurkat cells overexpressing SLC22A16 cells became significantly more sensitive to doxorubicin (2 μM) treatment. Characterization of SLC22A16 will help in designing novel therapies targeting hematological malignancies.

Section snippets

Materials and methods

Northern blot analysis of SLC22A16 mRNA in normal tissues. Multiple tissue Northern blots containing 2 μg of human mRNA were purchased from BD Biosciences Clontech (Palo Alto, CA, USA). The ³²P-labeled fragment of the coding region of SLC22A16 (ApaI–HindIII, 1339 bp) was used as a probe. In Northern blot analyses, filters were hybridized in a buffer containing 50% formamide, 5× SSC, 5× Denhardt’s solution, and 1% SDS at 42 °C overnight. The filter was washed with 0.2× SSC and with 0.1% SDS at 55 °C

Expression of SLC22A16 mRNA in normal tissues and cancer cell lines

To characterize the physiological role of SLC22A16, we analyzed its expression profile in normal tissues and in various cancer cell lines. Northern blot analysis showed a single band at 2.4 kb in fetal liver, whereas no band could be seen in adult liver. Among the adult tissues, a single strong band was seen in bone marrow and testis (Fig. 1). Real-time quantitative RT-PCR was performed to measure the expression levels of SLC22A16 in human cancer cell lines derived from the liver, bile duct,

Discussion

Earlier work from our laboratory showed that the human organic anion transporter, LST-2, mediates the cellular influx of methotrexate, which is an organic anion, in several cancer cells [8] . These results suggested the possibility that further ion transporters are capable of mediating cellular influx of anticancer drugs. At the same time, we also isolated an organic cation transporter (OCT), SLC22A16. We postulated that ionized, or maybe non-ionized, anticancer drugs might be transported by

Acknowledgments

The authors thank Drs. M.M. Gottesman, S.V. Ambudkar, G. Szakacs, J. Paterson, S. Abe, S. Takahashi, S. Fujimaki, T. Funato, T. Fujiwara, K. Ishizawa, and J. Kameoka for discussion, and Ms. C. Suzuki and E. Shibuya for their excellent technical help.

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Citation Excerpt :
Additional results from this study further propose a model in which MATE1 would act in concert with OCT2 to regulate intracellular levels of AGM [49]. SLC22A16 encodes for the carnitine transporter 2 (CT2) which is strictly expressed in adult human testes [56], bone marrow, and fetal liver [57]. Initially, CT2 was characterized in Xenopus laevis oocytes as a bidirectional transporter of 14[C]-l-carnitine [56].
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Characterization of the organic cation transporter SLC22A16: A doxorubicin importer

Abstract

Section snippets

Materials and methods

Expression of SLC22A16 mRNA in normal tissues and cancer cell lines

Discussion

Acknowledgments

Cancer Cell

Blood

Gastroenterology

J. Biol. Chem.

Exp. Hematol.

J. Biol. Chem.

BBRC

J. Biol. Chem.

J. Biol. Chem.

FEBS

Biochem. Biophys. Res. Commun.

FEBS

Blood

J. Biol. Chem.

Blood

Biochem. Pharmacol.

Biochem. Pharmacol.

The human ATP-binding cassette (ABC) transporter superfamily

Genome Res.

Expression of a full-length cDNA for the human “MDR1” gene confers resistance to colchicine, doxorubicin, and vinblastine

Proc. Natl. Acad. Sci. USA

P-glycoprotein and multidrug resistance protein activities in relation to treatment outcome in acute myeloid leukemia

Clin. Cancer Res.