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Metformin and phenformin deplete tricarboxylic acid cycle and glycolytic intermediates during cell transformation and NTPs in cancer stem cells.

ABSTRACT: Metformin, a first-line diabetes drug linked to cancer prevention in retrospective clinical analyses, inhibits cellular transformation and selectively kills breast cancer stem cells (CSCs). Although a few metabolic effects of metformin and the related biguanide phenformin have been investigated in established cancer cell lines, the global metabolic impact of biguanides during the process of neoplastic transformation and in CSCs is unknown. Here, we use LC/MS/MS metabolomics (>200 metabolites) to assess metabolic changes induced by metformin and phenformin in an Src-inducible model of cellular transformation and in mammosphere-derived breast CSCs. Although phenformin is the more potent biguanide in both systems, the metabolic profiles of these drugs are remarkably similar, although not identical. During the process of cellular transformation, biguanide treatment prevents the boost in glycolytic intermediates at a specific stage of the pathway and coordinately decreases tricarboxylic acid (TCA) cycle intermediates. In contrast, in breast CSCs, biguanides have a modest effect on glycolytic and TCA cycle intermediates, but they strongly deplete nucleotide triphosphates and may impede nucleotide synthesis. These metabolic profiles are consistent with the idea that biguanides inhibit mitochondrial complex 1, but they indicate that their metabolic effects differ depending on the stage of cellular transformation.

SUBMITTER: Janzer A

PROVIDER: S-EPMC4115496 | biostudies-literature | 2014 Jul

REPOSITORIES: biostudies-literature

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Metformin and phenformin deplete tricarboxylic acid cycle and glycolytic intermediates during cell transformation and NTPs in cancer stem cells.

Janzer Andreas A German Natalie J NJ Gonzalez-Herrera Karina N KN Asara John M JM Haigis Marcia C MC Struhl Kevin K

Proceedings of the National Academy of Sciences of the United States of America 20140707 29

Metformin, a first-line diabetes drug linked to cancer prevention in retrospective clinical analyses, inhibits cellular transformation and selectively kills breast cancer stem cells (CSCs). Although a few metabolic effects of metformin and the related biguanide phenformin have been investigated in established cancer cell lines, the global metabolic impact of biguanides during the process of neoplastic transformation and in CSCs is unknown. Here, we use LC/MS/MS metabolomics (>200 metabolites) to ...[more]

PMID: 25002509

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Project description:The tricarboxylic acid (TCA) cycle is a central part of carbon and energy metabolism, also connecting to glycolysis, amino acid, and lipid metabolism. The quantitation of the TCA cycle intermediate within one method is lucrative due to the interest in central carbon metabolism profiling in cells and tissues. In addition, TCA cycle intermediates in serum have been discovered to correspond as biomarkers to various underlying pathological conditions. In this work, an Liquid Chromatography-Mass Spectrometry/Mass Spectrometry-based quantification method is developed and validated, which takes advantage of fast, specific, sensitive, and cost-efficient precipitation extraction. Chromatographic separation is achieved while using Atlantis dC18 2.1 mm × 100 mm, particle size 3-μm of Waters column with a gradient elution mobile phase while using formic acid in water (0.1% v/v) and acetonitrile. Linearity was clearly seen over a calibration range of: 6.25 to 6400 ng/mL (r2 > 0.980) for malic acid; 11.72 to 12,000 ng/mL (r2 > 0.980) for cis-aconitic acid and L-aspartic acid; 29.30 to 30,000 ng/mL (r2 > 0.980) for isocitric acid, l-serine, and l-glutamic acid; 122.07 to 125,000 ng/mL (r2 > 0.980) for citric acid, glycine, oxo-glutaric acid, l-alanine, and l-glutamine; 527.34 to 540,000 ng/mL (r2 > 0.980) for l-lactic acid; 976.56 to 1,000,000 ng/mL (r2 > 0.980) for d-glucose; 23.44 to 24,000 ng/mL (r2 > 0.980) for fumaric acid and succinic acid; and, 244.14 to 250,000 ng/mL (r2 > 0.980) for pyruvic acid. Validation was carried out, as per European Medicines Agency (EMA) "guidelines on bioanalytical method validation", for linearity, precision, accuracy, limit of detection (LOD), limit of quantification (LLOQ), recovery, matrix effect, and stability. The recoveries from serum and tissue were 79-119% and 77-223%, respectively. Using this method, we measured TCA intermediates in serum, plasma (NIST 1950 SRM), and in mouse liver samples. The concentration found in NIST SRM 1950 (n = 6) of glycine (246.4 µmol/L), l-alanine (302.4 µmol/L), and serine (92.9 µmol/L).

Dataset Information

Metformin and phenformin deplete tricarboxylic acid cycle and glycolytic intermediates during cell transformation and NTPs in cancer stem cells.

Publications

Metformin and phenformin deplete tricarboxylic acid cycle and glycolytic intermediates during cell transformation and NTPs in cancer stem cells.

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