Synthesis of a biotin-functionalized biguanide for the identification of the tumor growth inhibition mechanism of metformin
Metformin is widely used to reduce high blood sugar levels in type 2 diabetes patients. In recent studies, researchers observed an inverse association between metformin use and cancer incidence in different tissues. Inhibition of the mitochondrial respiratory complex I has been proposed to be the cause of the antineoplastic activity of metformin, but the mechanism by which metformin inhibits the complex I and consequently stops proliferation of cancer cells was not fully identified and understood.  In this thesis, we present the functionalization of a biguanide. A metformin - biotin conjugate was synthesized and used to identify the mechanism of action of metformin and its target in mitochondria. Mitochondrial extracts were put in contact with the biotinylated biguanide and were passed over affinity columns containing immobilized streptavidin. The proteins eluted after the washing steps were analyzed by LC-MS/MS and their sequences analyzed using the BLAST algorithm (NCBI). We isolated and identified by mass spectrometry (MS) an 8 KDa protein that specifically and directly binds metformin. This protein has been identified as the ATP synthase-subunit e. Our results suggest that metformin inhibits cancer cell proliferation through the direct inhibition of the ATP synthase of the mitochondrial respiratory chain. The research presented in this thesis has been performed in the Department of Chemistry, in collaboration with the Department of Biochemistry and Molecular Medicine, at the University of Montreal.
Biguanide ATP synthase Metformin Biotin Streptavidin Affinity chromatography Anticancer Complex I Mitochondria respiratory chain