Effects of small molecule malic enzyme 2 inhibitors on the viability of human melanoma cell lines

Abstract

Cancer continues to be a leading cause of disease burden and death; new treatments are still urgently required to improve patient outcomes. An emerging area of anti-cancer treatment is the targeting of metabolic features common amongst cancers, such as altered glucose and glutamine metabolism. One facilitator of this altered metabolism is malic enzyme 2 (ME2). MEs are oxidative decarboxylases that catalyse the conversion of L-malate to pyruvate, producing CO2 and reducing either NAD+ or NADP+. Three human ME isoforms have been identified (ME1-ME3), with overexpression of ME1 and ME2 documented in a large variety of cancers. ME2 genetic knockdown in cancer cell lines is associated with decreased NADPH, increased ROS and induction of cellular senescence. Two small molecule ME2 inhibitors, NPD-389 and embonic acid were identified from high through-put screens. NPD-389 displays an IC50 of 4.63 ± 0.36 μM against purified recombinant human ME2 and inhibits malate-dependent respiration within isolated mitochondria. Embonic acid is used in drug formulations to decrease water solubility and displays an IC50 of 1.4 ± 0.4 μM against purified ME2. Embonic acid also induces senescence in the H1299 immortalised cancer cell line. The current study sought to test the effects of NPD-389 and embonic acid on the viability of primary human melanoma cell lines. Four melanoma cell lines were cultured: two with high expression of ME2 and two with low expression of all ME isoforms, as suggested by previous RNA sequencing. RNA sequencing results were generally confirmed by western blotting. Cell number following 72 hours exposure to NPD-389 or embonic acid was quantified via DAPI staining. NPD-389 potently reduced cell viability to a similar degree in all cell lines examined (IC50: 19-63 μM), while embonic acid had minimal effect. The current findings show that NPD-389 is potentially a valuable lead molecule for development of anti-cancer therapy. Further investigation and chemical modification of NPD-389 could result in a potent ME2 inhibitor for use as part of anti-cancer therapy.