Abstract B083: NUAK1 regulates breast cancer cell bioenergetics

Abstract

Abstract NUAK1 is a serine/threonine kinase member of AMPK catalytic subunit family, and localizes in the cytosolic and nuclear subcellular compartments. NUAK1 expression is associated with cell migration, invasion, and metastasis in breast and other types of cancer, which correlate with poor patient prognosis. In addition, NUAK1 has been involved in cancer cell proliferation, and cancer cell survival under metabolic and oxidative stress. Consistent with a role in cancer cell metabolism, NUAK1 was shown to maintain ATP levels in hepatocarcinoma cancer cells, but the underlying mechanism and its association with NUAK1 subcellular localization are unclear. Because NUAK1 expression correlates with poor prognosis in breast cancer patients, we studied the role of the cytosolic NUAK1 in the regulation of breast cancer cell bioenergetics. We initially measured ATP levels by using a luminescence-based assay kit. By overexpression of NUAK1 or its inhibition with the specific inhibitor HTH-01-015, we determined that NUAK1 increases ATP levels in breast cancer cells. Overexpression of a nuclear localization-deficient mutant indicated that the effect on the ATP levels is mainly associated with the cytosolic NUAK1. To address the source of the ATP increment, we analyzed energy metabolism by using a Seahorse XF analyzer. Interestingly, NUAK1 inhibition with HTH-01-015 decreased both mitochondrial respiration and glycolysis. According with the association of cytosolic NUAK1 with the increment of ATP levels, overexpression of the nuclear localization-deficient NUAK1 mutant significantly increased mitochondrial respiration and glycolysis. As expected, NUAK1 overexpression decreased the mitochondrial membrane potential, analyzed by in vivo TMRE (tetramethyl rhodamine ethyl ester)-based fluorescent microscopy. Complementary, NUAK1 inhibition increased the mitochondrial membrane potential and dramatically altered the mitochondrial morphology. Because mitochondrial dysfunction induces oxidative stress by generating excesses of reactive oxygen species, we analyzed whether this stress was associated with the altered mitochondrial morphology. The effect of NUAK1 inhibition on the mitochondrial morphology was partially reversed by treatment with the antioxidant N-acetylcysteine or with TROLOX. Altogether, our results suggest that cytosolic NUAK1 participates in ATP production in breast cancer cells through maintaining properly glycolysis and mitochondrial function. How this metabolic NUAK1 function associates with tumor progression remains to be determined. The maintenance of the cellular bioenergetics could be another means by which NUAK1 promotes metastasis in breast cancer. Funded by Fondecyt 1160731 (AFC), Fondecyt 1180983 and IMII P09-016-F (AAE) Citation Format: Emilia Escalona, Marcelo Muñoz, Roxana Pincheira, Alvaro A. Elorza, Ariel F. Castro. NUAK1 regulates breast cancer cell bioenergetics [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr B083. doi:10.1158/1535-7163.TARG-19-B083