Michala Gylling Rolver:
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Tumor microenvironmental acidosis arises as a consequence of increased glycolytic and oxidative metabolism and poor tumor vascularization. It has been shown to increase genomic instability, induce profound metabolic reprogramming, enhance motility, EMT, and invasion, and ultimately increase metastasis. However, the mechanisms linking extracellular acidosis to carcinogenesis remain unknown. The focus of the present PhD study was to investigate the effects of chronic acidosis on intracellular pH (pHi) regulation, metabolism and stemness in pancreatic and breast cancer cells and elucidate key underlying mechanisms.
In Paper I, we summarized the current knowledge on the tumor microenvironment and discussed the reciprocal interactions between cancer cells, stromal cells and pH-regulatory transporters. In Paper II, we employed cancer cells adapted to growth under chronic acidosis, mimicking the acidic tumor microenvironment, to establish that an acidic extracellular pH (pHe) upregulates net acid extruders, and increases pHi when measured at physiological pHe, but not at acidic pHe. Metabolic profiling, shotgun lipidomics, and immunofluorescence- and biochemical assays demonstrated that the acid-adapted cancer cells exhibit enhanced metabolic flexibility and increased lipid droplet-, triacylglycerol-, and peroxisome and
mitochondrial content. We demonstrated an increased activity of, and dependence on, peroxisome proliferator-activated receptor-a (PPARa) in the acid-adapted cells, indicating an increased need for fatty acid metabolism.
In Paper III, we therefore studied the interplay between mitochondria, iron- and fatty acid metabolism and ferroptosis. Using advanced microscopy-based analyses, western blotting, and metabolic profiling, we observed shared and cell-type specific alterations to mitochondrial morphology dynamics, the electron transport chain and iron metabolism. Interestingly, we found that pharmacological induction of ferroptosis was ineffective against acid-adapted MDA-MB-231 cells, whereas Panc-1 cells were equally susceptible irrespective of acid exposure, indicating a potential therapeutic liability.
Increased fatty acid metabolism has been linked to stemness, and in Paper IV, we assessed whether prolonged acid exposure induced a stem-cell like phenotype in pancreatic cancer cell lines. GSEA plots indicated an enrichment of genes related to stemness which was confirmed by qPCR and Western blotting of selected targets. Pancreatosphere assays demonstrated an increased self-renewal capacity, and flow cytometry indicated an increased fraction of cancer stem cells (CSCs) in acid-adapted cells. Overall, our study suggests that extracellular acidosis selects for CSC traits, thus contributing to disease aggressiveness.
Collectively, the work of this PhD demonstrates that acid-adaptation causes profound metabolic alterations in cancer cells, resulting in distinct phenotypical traits which could potentially be exploited therapeutically.