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The effect of substitution at C-2 of D-glucose 6-phosphate on the rate of dehydrogenation by glucose 6-phosphate dehydrogenase (from yeast and from rat liver).

ABSTRACT: 1. The deoxyfluoro-d-glucopyranose 6-phosphates are substrates for both yeast and rat liver glucose 6-phosphate dehydrogenase. 2. The V(max.) values (relative to d-glucose 6-phosphate) were determined for a series of d-glucose 6-phosphate derivatives substituted at C-2. The V(max.) values decreased with increasing electronegativity of the C-2 substituent. This is consistent with a mechanism involving hydride-ion transfer. 3. 2-Deoxy-d-arabino-hexose 6-phosphate (2-deoxy-d-glucose 6-phosphate) showed substrate inhibition with the yeast enzyme but not with the rat liver enzyme. 4. 2-Amino-2-deoxy-d-glucose 6-phosphate (d-glucosamine 6-phosphate) was a substrate for the yeast enzyme but a competitive inhibitor for the rat liver enzyme. 5. Lineweaver-Burk plots for the d-glucose 6-phosphate derivatives with yeast glucose 6-phosphate dehydrogenase were biphasic.

SUBMITTER: Bessell EM

PROVIDER: S-EPMC1177441 | biostudies-other | 1973 Jan

REPOSITORIES: biostudies-other

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Project description:BackgroundGlucose-6-phospate dehydrogenase (G6PD) is the limiting enzyme of the pentose phosphate pathway (PPP) correlated to cancer progression and drug resistance. We previously showed that G6PD inhibition leads to Endoplasmic Reticulum (ER) stress often associated to autophagy deregulation. The latter can be induced by target-based agents such as Lapatinib, an anti-HER2 tyrosine kinase inhibitor (TKI) largely used in breast cancer treatment.MethodsHere we investigate whether G6PD inhibition causes autophagy alteration, which can potentiate Lapatinib effect on cancer cells. Immunofluorescence and flow cytometry for LC3B and lysosomes tracker were used to study autophagy in cells treated with lapatinib and/or G6PD inhibitors (polydatin). Immunoblots for LC3B and p62 were performed to confirm autophagy flux analyses together with puncta and colocalization studies. We generated a cell line overexpressing G6PD and performed synergism studies on cell growth inhibition induced by Lapatinib and Polydatin using the median effect by Chou-Talay. Synergism studies were additionally validated with apoptosis analysis by annexin V/PI staining in the presence or absence of autophagy blockers.ResultsWe found that the inhibition of G6PD induced endoplasmic reticulum stress, which was responsible for the deregulation of autophagy flux. Indeed, G6PD blockade caused a consistent increase of autophagosomes formation independently from mTOR status. Cells engineered to overexpress G6PD became resilient to autophagy and resistant to lapatinib. On the other hand, G6PD inhibition synergistically increased lapatinib-induced cytotoxic effect on cancer cells, while autophagy blockade abolished this effect. Finally, in silico studies showed a significant correlation between G6PD expression and tumour relapse/resistance in patients.ConclusionsThese results point out that autophagy and PPP are crucial players in TKI resistance, and highlight a peculiar vulnerability of breast cancer cells, where impairment of metabolic pathways and autophagy could be used to reinforce TKI efficacy in cancer treatment.

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The effect of substitution at C-2 of D-glucose 6-phosphate on the rate of dehydrogenation by glucose 6-phosphate dehydrogenase (from yeast and from rat liver).

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