Metformin selectively targets redox control of complex I energy transduction

Amy R Cameron,Lisa Logie,Kashyap Patel,Stefan Erhardt,Sandra Bacon,Paul Middleton,Jean Harthill,Calum Forteath,Josh T Coats,Calum Kerr,Heather Curry,Derek Stewart,Kei Sakamoto,Peter Repiščák,Martin J Paterson,Ilmo Hassinen,Gordon Mcdougall,Graham Rena

doi:10.1016/j.redox.2017.08.018

Abstract

Many guanide-containing drugs are antihyperglycaemic but most exhibit toxicity, to the extent that only the biguanide metformin has enjoyed sustained clinical use. Here, we have isolated unique mitochondrial redox control properties of metformin that are likely to account for this difference. In primary hepatocytes and H4IIE hepatoma cells we found that antihyperglycaemic diguanides DG5-DG10 and the biguanide phenformin were up to 1000-fold more potent than metformin on cell signalling responses, gluconeogenic promoter expression and hepatocyte glucose production. Each drug inhibited cellular oxygen consumption similarly but there were marked differences in other respects. All diguanides and phenformin but not metformin inhibited NADH oxidation in submitochondrial particles, indicative of complex I inhibition, which also corresponded closely with dehydrogenase activity in living cells measured by WST-1. Consistent with these findings, in isolated mitochondria, DG8 but not metformin caused the NADH/NAD+ couple to become more reduced over time and mitochondrial deterioration ensued, suggesting direct inhibition of complex I and mitochondrial toxicity of DG8. In contrast, metformin exerted a selective oxidation of the mitochondrial NADH/NAD+ couple, without triggering mitochondrial deterioration. Together, our results suggest that metformin suppresses energy transduction by selectively inducing a state in complex I where redox and proton transfer domains are no longer efficiently coupled.

Highlights

We have been using analogues of metformin to study how cellular effects of metformin relate to its chemical properties, which include a strongly hydrophilic character, metal-binding and a pKa within the physiological pH range [1,2,3,4,5,6]
We found that two diguanides, 1,10-Bis(guanidino)decane and 1,6-Bis(guanidino)hexane (DG6 in this study) had effects on both AMPK and S6 phosphorylation at 25 μM, whereas 1,4Bis(guanidino)butane was ineffective (Fig. 1c, quantified in supplementary Fig. 1)
Previous studies established that alkylguanidines with chain lengths comparable with synthalin act as even more potent mitochondrial inhibitors than diguanides or biguanides [13] and we tested dodecylguanidine (AG12), the most potent of these, and found that it too activated AMPK and suppressed S6 phosphorylation (Fig. 1c), demonstrating that the second guanidine group is dispensable for effects of diguanides on these signalling pathways

Summary

Introduction

We have been using analogues of metformin to study how cellular effects of metformin relate to its chemical properties, which include a strongly hydrophilic character, metal-binding and a pKa within the physiological pH range [1,2,3,4,5,6] This collection of properties is not shared by other guanidine-containing compounds many do have mitochondrial and antihyperglycaemic effects. For mGPD, it has been argued elsewhere that effects on glucose production might require additional inhibition of the alternative redox malate/aspartate shuttle [28] None of these models readily explain the suppression of weight gain that is one of the key clinical hallmarks of metformin treatment [29]

Methods

Results

Discussion

Conclusion