Optimized Method for Untargeted Metabolomics Analysis of MDA-MB-231 Breast Cancer Cells

Abstract

Cancer cells often have dysregulated metabolism, which is largely characterized by the Warburg effect—an increase in glycolytic activity at the expense of oxidative phosphorylation—and increased glutamine utilization. Modern metabolomics tools offer an efficient means to investigate metabolism in cancer cells. Currently, a number of protocols have been described for harvesting adherent cells for metabolomics analysis, but the techniques vary greatly and they lack specificity to particular cancer cell lines with diverse metabolic and structural features. Here we present an optimized method for untargeted metabolomics characterization of MDA-MB-231 triple negative breast cancer cells, which are commonly used to study metastatic breast cancer. We found that an approach that extracted all metabolites in a single step within the culture dish optimally detected both polar and non-polar metabolite classes with higher relative abundance than methods that involved removal of cells from the dish. We show that this method is highly suited to diverse applications, including the characterization of central metabolic flux by stable isotope labelling and differential analysis of cells subjected to specific pharmacological interventions.

Gd-GLU toward NMR imaging: synthesis, characterization and breast cell uptake assay

“…Glucose was only present when cells were incubated with Glu-Gd, added in the PBS solution. Peterson et al (2016), demonstrated that c231 incorporates glucose into the glycolytic pathways (GP) and shows that the GP was highly active at 60 min Glu incubation (Peterson et al, 2016). Our data showed similar time Gd-Glu uptakes, at 30 and 50 min, in comparison to Peterson and cols information.…”

Impact of chemotherapy on metabolic reprogramming: Characterization of the metabolic profile of breast cancer MDA-MB-231 cells using 1 H HR-MAS NMR spectroscopy

“…Glutamate and glutamine denote a group of glucogenic amino acids that features many biological functions. These are considered to be very important for cell function maintenance and promotion, being glutamine an important metabolic precursor in rapidly-dividing cells [23]. Cell demands for glutamine and glutamate are enormous during severe illness.…”

Metabolomic Approaches to Study Chemical Exposure-Related Metabolism Alterations in Mammalian Cell Cultures

“…The obtention of samples from extracellular media is straightforward and requires a single centrifugation step of culture supernatants to remove cells and debris. On the contrary, numerous protocols aimed at the extraction of metabolites from biological matrices have been developed for NMR and MS analyses, some of which have been specifically tuned for cell cultures [ 49 , 51 , 52 , 56 , 102 , 103 , 104 , 105 , 106 , 107 , 108 , 109 , 110 ]. These protocols are quite similar in principle for both analytical platforms but suit individual requirements such as avoiding high concentrations of protonated solvent molecules as concerns NMR profiling.…”