Article Text

  1. S. Telang,
  2. A. Yalcin,
  3. A. Clem,
  4. J. Chesney
  1. Department of Medicine (Medical Oncology), J.G. Brown Cancer Center


Introduction Increased glucose metabolism in neoplastic cells is orchestrated by the oncogenic proteins ras and c-myc. Ectopic expression of oncogenic ras rapidly increases glucose metabolism by activating the synthesis of fructose-2,6-bisphosphate (F2,6BP), an allosteric activator of 6-phosphofructo-1-kinase (PFK-1), the rate-limiting step of glycolysis. The steady-state concentration of F2,6BP depends on the activity of 6-phosphofructo-2-kinase (PFK-2), and an inducible isoform of PFK-2 (iPFK) was recently identified to be overexpressed by human solid tumors relative to adjacent normal tissues. We hypothesize that increased F2,6BP, arising from activation of iPFK-2 by oncogenic ras, is required for the increased flux of glucose carbons into anabolic pathways and is necessary for the anchorage-independent growth of neoplastic cells.

Methods/Results We isolated lung fibroblasts from iPFK-2+/+ and iPFK-2+/- mice and transduced the cells with retroviruses expressing SV40 large T-antigen (LT) and human H-rasV12 mutant. Introduction of LT and activated H-rasV12 into wild-type lung fibroblasts conferred the ability for anchorage-independent growth as soft agar colonies, a hallmark of neoplastic transformation (143.3 ± 29.9 colonies [n = 3], 9 days of culture). In contrast, introduction of LT and HrasV12 into iPFK-2+/- lung fibroblasts was unable to transform the cells and permit anchorage-independent growth (0 colonies [n = 3], 14 days of culture). Ras-transformed fibroblasts isolated from iPFK-2+/- mice were found to secrete significantly more protons than their iPFK-2+/+ counterparts (48 hrs exponential growth: iPFK-2+/-, pH 7.0 vs iPFK-2+/+, pH 7.5). We reasoned that such media acidification could be caused by a perturbation of mitochondrial respiration leading to a proton efflux and/or a compensatory increase in glycolytic flux to lactate. Although we found no significant difference in lactate secretion, we did observe a fully dissipated mitochondrial membrane potential in the ras-transformed fibroblasts isolated from iPFK2+/- mice.

Conclusions The observation that heterozygote genomic deletion of iPFK-2 causes a loss of the mitochondrial membrane potential indicates that iPFK-2 may serve as an essential coupler of glycolysis with respiration. That this metabolic phenotype is not compatible with anchorage-independent growth supports the rationale for molecular targeting of iPFK-2 as an antineoplastic agent. Given the recent observation that iPFK-2+/- mice display no overt pathology, these studies suggest that pharmacologic inhibition of iPFK-2 may yield antineoplastic effects without causing significant toxicity.

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