A New Combination Therapy Utilizing Mitochondria-Targeting Small-Molecule Ligands and Clinical Inhibitors against Melanoma

Mitochondria are believed to be a potential drug target in cancer therapies because of their critical and multiple biofunctions in supplying energy and regulating signaling pathways for cell cycle and proliferation. It has been known that mitochondrial DNA (mtDNA) contains many guanine-rich sequences, and some of them may fold into stable G-quadruplex (G4) structures <i>in vitro</i>. The stabilization of mtDNA G4s with potent small-molecule ligands in cancer cells may potentially interrupt mitochondrial metabolism such as impairing the oxidative phosphorylation system (OXPHOS) in ATP synthesis to cause energy deficiency. Therefore, mtDNA G4s have been an emerging drug target for chemical biology and anticancer study. Nonetheless, the development of potent ligands specifically targeting mitochondria and interacting with mtDNA G4s in living cells remains a challenge. This largely limits the feasibility to understand the mechanism of actions targeting mitochondria and mtDNA G4s for drug discovery. Herein, we designed and synthesized several new mitochondria-targeting small molecules that bind to mtDNA G4s in melanoma cancer cells (A375) to cause mitochondrial metabolism alternation. Among the ligands, <b>B1N</b> was found to be the most potent one to downregulate the expression of some critical mitochondrial genes and proteins, inhibit ATP synthesis, and substantially induce metabolism reprogramming to upregulate glycolysis. Moreover, the combination therapy study of 1.75 μM <b>B1N</b> with a clinical BRAF inhibitor (Vemurafenib, 0.2 μM) showed synergistic effects (CI = 0.67) against A375 cells. This new combined treatment significantly downregulates ATP production and glycolysis and induces acute senescence. The present study demonstrates an innovative and effective combination therapy strategy utilizing mitochondrion-targeting ligands and clinical inhibitors against melanoma.