Mitochondrial uncoupler and retinoic acid synergistically induce differentiation and inhibit proliferation in neuroblastoma.

Neuroblastoma is a leading cause of death in childhood cancer cases. Unlike adult malignancies, which typically develop from aged cells through accumulated damage and mutagenesis, neuroblastoma originates from neural crest cells with disrupted differentiation. This distinct feature provides novel therapeutic opportunities beyond conventional cytotoxic methods. Previously, we reported that the mitochondrial uncoupler NEN (niclosamide ethanolamine) activated mitochondria respiration to reprogram the epigenome, promoting neuronal differentiation. In the current study, we further combine NEN with retinoic acid (RA) to promote neural differentiation both in vitro and in vivo. The treatment increased the expression of RA signaling and neuron differentiation-related genes, resulting in a global shift in the transcriptome towards a more favorable prognosis. Overall, these results suggest that the combination of a mitochondrial uncoupler and the differentiation agent RA is a promising therapeutic strategy for neuroblastoma.

Epigenetic priming targets tumor heterogeneity to shift transcriptomic phenotype of pancreatic ductal adenocarcinoma towards a Vitamin D susceptible state.

As a highly heterogeneous tumor, pancreatic ductal adenocarcinoma (PDAC) exhibits non-uniform responses to therapies across subtypes. Overcoming therapeutic resistance stemming from this heterogeneity remains a significant challenge. Here, we report that Vitamin D-resistant PDAC cells hijacked Vitamin D signaling to promote tumor progression, whereas epigenetic priming with glyceryl triacetate (GTA) and 5-Aza-2'-deoxycytidine (5-Aza) overcame Vitamin D resistance and shifted the transcriptomic phenotype of PDAC toward a Vitamin D-susceptible state. Increasing overall H3K27 acetylation with GTA and reducing overall DNA methylation with 5-Aza not only elevated the Vitamin D receptor (VDR) expression but also reprogrammed the Vitamin D-responsive genes. Consequently, Vitamin D inhibited cell viability and migration in the epigenetically primed PDAC cells by activating genes involved in apoptosis as well as genes involved in negative regulation of cell proliferation and migration, while the opposite effect of Vitamin D was observed in unprimed cells. Studies in genetically engineered mouse PDAC cells further validated the effects of epigenetic priming for enhancing the anti-tumor activity of Vitamin D. Using gain- and loss-of-function experiments, we further demonstrated that VDR expression was necessary but not sufficient for activating the favorable transcriptomic phenotype in respond to Vitamin D treatment in PDAC, highlighting that both the VDR and Vitamin D-responsive genes were prerequisites for Vitamin D response. These data reveal a previously undefined mechanism in which epigenetic state orchestrates the expression of both VDR and Vitamin D-responsive genes and determines the therapeutic response to Vitamin D in PDAC.

Mitochondrial Uncoupling Inhibits Reductive Carboxylation in Cancer Cells.

When the electron transport chain (ETC) function is impaired, cancer cells rely on reductive carboxylation (RC) to convert α-ketoglutarate (αKG) to citrate for macromolecular synthesis, thereby promoting tumor growth. Currently, there is no viable therapy to inhibit RC for cancer treatment. In this study, we demonstrate that the mitochondrial uncoupler treatment effectively inhibits RC in cancer cells. Mitochondrial uncoupler treatment activates the ETC and increases the NAD+/NADH ratio. Using U-13C-glutamine and 1-13C-glutamine tracers, we show that mitochondrial uncoupling accelerates the oxidative tricarboxylic acid (TCA) cycle and blocks RC under hypoxia, in von Hippel-Lindau (VHL) tumor suppressor-deficient kidney cancer cells, or under anchorage-independent growth condition. Together, these data demonstrate that mitochondrial uncoupling redirects α-KG from RC back to the oxidative TCA cycle, highlighting that the NAD+/NADH ratio is one key switch that determines the metabolic fate of α-KG. Inhibiting RC could be a key mechanism by which mitochondrial uncouplers inhibit tumor growth. IMPLICATIONS: Mitochondrial uncoupling is a novel strategy to target RC in cancer.