The synthesis and biodistribution of [11C]metformin as a PET probe to study hepatobiliary transport mediated by the multi-drug and toxin extrusion transporter 1 (MATE1) in vivo

doi:10.1016/j.bmc.2013.10.041

Bioorganic & Medicinal Chemistry

Volume 21, Issue 24, 15 December 2013, Pages 7584-7590

https://doi.org/10.1016/j.bmc.2013.10.041 Get rights and content

Abstract

In order to develop a new positron emission tomography (PET) probe to study hepatobiliary transport mediated by the multi-drug and toxin extrusion transporter 1 (MATE1), ¹¹C-labelled metformin was synthesized and then evaluated as a PET probe. [¹¹C]Metformin ([¹¹C]4) was synthesized in three steps, from [¹¹C]methyl iodide. Evaluation by small animal PET of [¹¹C]4 showed that there was increased concentrations of [¹¹C]4 in the livers of mice pre-treated with pyrimethamine, a potential inhibitor of MATEs, inhibiting the hepatobiliary excretion of metformin. Radiometabolite analysis showed that [¹¹C]4 was not degraded in vivo during the PET scan. Biodistribution studies were undertaken and the organ distributions were extrapolated into a standard human model. In conclusion, [¹¹C]4 may be useful as a PET probe to non-invasively study the in vivo function of hepatobiliary transport and drug–drug interactions, mediated by MATE1 in future clinical investigations.

Graphical abstract

Introduction

Membrane transporters are found in eukaryotes and prokaryotes and are involved in both the uptake and excretion of nutrients and xenobiotics from cells. There are over 400 known transporters which belong to the two principle membrane transporter superfamilies; the ATP-binding cassette (ABC) and solute carrier (SLC) superfamilies. In humans, membrane transporters are widely distributed throughout the body, and have a role in drug disposition and in drug–drug interactions.¹

Metformin is an orally bioavailable biguanide widely used as an insulin sensitizer in the treatment of type II diabetes, and its main pharmacological effect is to decrease the fasting blood sugar levels in patients.² The principle site of action of metformin is the liver, where it brings about the inhibition of glucose-6-phosphatase and phosphoenolpyruvate carboxykinase, the enzymes involved in gluconeogenesis.³ Metformin is well tolerated, does not undergo metabolism and is excreted in an unchanged form in the bile and urine.⁴ In the liver, metformin undergoes carrier mediated transport and is transported into the hepatocytes via the organic cation transporter 1 (OCT1),⁵ which is a member of the SLC22A transporter family, and excreted from the hepatocytes and into the bile canaliculi via the multi-drug and toxin extrusion transporter 1 (MATE1),⁶ which is a member of the SLC47A transporter family. In the kidney, metformin is transported into the proximal tubule via the organic cation transporter 2 (OCT2)⁷ and excreted into the urine via the multi-drug and toxin extrusion transporters MATE1 and MATE2-K.⁶

Recently, owing to the possible clinical significance of canalicular transporters, such as MATE1, in drug–drug interactions, draft guidelines were issued by the US Food and Drug Administration⁸ and the European Medicines Agency.⁹ Deficiency of canalicular transporters, for example due to pharmacogenomics or drug–drug interactions, can lead to an increased concentration of substrates/drugs within organs but have negligible effects on the plasma concentration of the substrates/drugs. Moreover, there are severe limitations in the current methodologies for the direct and separate measurements of influx and canalicular clearances in conventional clinical studies using non-radiolabeled drugs.¹⁰

Positron emission tomography (PET) is a highly sensitive and quantitative imaging modality which allows the non-invasive imaging of an administered positron emitting probe in a living organism with spatiotemporal resolution and we have reported the use of PET to study membrane transporter function.¹¹ In order to study the hepatobiliary transport of metformin mediated by OCT1 and MATE1 for future clinical studies, we successfully synthesized [¹¹C]metformin ([¹¹C]4) and investigated the potential of [¹¹C]4 as a PET probe, by undertaking a drug–drug interaction study with pyrimethamine, a potential inhibitor of the hepatobiliary transport of metformin meditated by MATE1, in mice in vivo.

Section snippets

Radiochemistry

The synthesis of [¹¹C]4 was carried out in 3 steps and the route is shown in Scheme 1. The route is based on that devised by Jacobson et al.¹² for the synthesis of ¹¹C-labeled guanidines, however [¹¹C]dimethylamine ([¹¹C]2) was used in place of [¹¹C]cyanogen bromide. [¹¹C]2 was synthesized according to the method of Jacobson and Mishani,¹³ by reacting methylamine with [¹¹C]methyl iodide ([¹¹C]1) at 45 °C for 5 min, and was not purified. Subsequently, [¹¹C]dimethylcyanamide ([¹¹C]3) was

Conclusion

We have shown that the effects of the inhibition of the hepatobiliary transport of [¹¹C]4, by pyrimethamine, in mice can be studied non-invasively using PET. Furthermore, we showed that [¹¹C]4 is metabolically stable. The organ distributions of [¹¹C]4 were also subsequently extrapolated into a standard human model, and the calculated effective dose was comparable to other ¹¹C-labeled PET tracers. Therefore, it may be possible to safely administer [¹¹C]4 to humans. Thus, [¹¹C]4 may be a useful

Materials

Metformin, compound 4, was purchased from Alexis Biochemicals (San Diego, USA); pyrimethamine, from MP Biomedicals (Santa Ana, USA); cyanogen bromide, from Wako Pure Chemical Industries, Ltd., (Tokyo, Japan); and 2 M methylamine in THF, from Sigma Aldrich Japan (Tokyo, Japan). All other reagents together with solvents including HPLC grade solvents were purchase from Nacali Tesque, Inc., (Kyoto, Japan) and used as obtained. HPLC grade H₂O was obtained using a Millipore Elix water system.

General radiochemistry

Acknowledgements

This study was supported by the New Energy and Industrial Technology Development Organization, NEDO, as part of the “Establishment of Evolutional Drug Development with the Use of Microdose Clinical Trial” project. In addition, partial support was also obtained from the Japanese Ministry of Education, Culture, Sports, Science and Technology via a Grant-in-Aid for Scientific Research (S), Kankenhi 24229002 (Y.S.). We thank Mr. Masahiro Kurahashi, Sumitomo Heavy Industry Accelerator Service, Ltd.,

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^†: Present address: Mochida Pharmaceutical Co., Ltd., 722 Jinbajiuenohara, Gotenba, Shizuoka 412–8524, Japan.

^‡: Present address: RIKEN Program for Drug Discovery and Medical Technology Platforms, 2–1 Hirosawa, Wako, Saitama 351–0198, Japan.

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The synthesis and biodistribution of [11C]metformin as a PET probe to study hepatobiliary transport mediated by the multi-drug and toxin extrusion transporter 1 (MATE1) in vivo

Abstract

Graphical abstract

Introduction

Section snippets

Radiochemistry

Conclusion

Materials

General radiochemistry

Acknowledgements

Biochem. Pharmacol.

Appl. Radiat. Isot.

Nat. Rev. Drug Disc.

Ann. Intern. Med.

Clin. Sci.

J. Pharmacol. Exp. Ther.

Pharm. Res.

J. Pharmacol.

Mol. Pharmacol.

J. Am. Chem. Soc.

The synthesis and biodistribution of [¹¹C]metformin as a PET probe to study hepatobiliary transport mediated by the multi-drug and toxin extrusion transporter 1 (MATE1) in vivo