Comparison of action of the anti-neoplastic drug lonidamine on drug-sensitive and drug-resistant human breast cancer cells: 31P and 13C nuclear magnetic resonance studies.

Lonidamine (LND) is a relatively new anti-cancer drug, and several clinical trials have indicated that it may be effective in combinations with other therapeutic modalities. LND is classified within the metabolic inhibitor agents. Multidrug resistance (MDR) phenomenon is often associated with increased energy requirements, and enhanced glycolysis rate. These studies were performed to delineate the mechanism of action of LND on MDR human breast cancer cells, and to investigate whether LND as a single agent, or in combination with another anti-metabolism drug, 2-deoxyglucose (2-DG), may be useful against MDR tumors. The effects of LND on intact perfused drug-sensitive (WT) and 33-fold resistant to Adriamycin (Adr) MCF-7 cells, embedded in alginate micro capsules, were continuously monitored by 31P and 13C nuclear magnetic resonance (NMR) spectroscopy. 31P NMR studies showed that LND induced intracellular acidification and depletion of NTP in both WT and Adr cells. However, pH and NTP levels decreased less in the Adr cells than in the WT cells (p < 0.05 for both parameters). 13C NMR demonstrated that LND inhibited lactate transport, and lactate signals were elevated in both cell lines. However, the intracellular lactate levels increased to a greater extent in the WT than in the Adr cells (p < 0.05). There were major differences in the effects of LND on metabolism between sensitive and resistant cells. While LND enhanced glucose uptake in the WT cells, and its administration was followed by continuous increase of lactate signal, both processes were not affected by LND in the Adr cells. 2-DG is a glucose analogue that inhibits both cellular uptake and utilization of glucose, leading to cell starvation. Combined treatment with LND and 2-DG yielded at best additive, but not synergistic, cellular toxicity, and the metabolic effects of LND were attenuated by 2-DG. These results showed that the principal mechanism of action of LND is inhibition of lactate transport leading to intracellular lactate accumulation and acidification in both WT and Adr cells. The Adr cells were only 2-fold resistant to LND (compared to the WT cells), and since cellular uptake of alkaloid chemotherapy is improved in acidic environment, LND may have a role in the treatment protocols of MDR tumors, especially when given as the initial means for induction of intracellular acidification.

Mechanism of action of the antineoplastic drug lonidamine: 31P and 13C nuclear magnetic resonance studies.

The mechanism of action of the antineoplastic drug lonidamine (LND) on MCF-7 human breast cancer cells was studied with the use of 31P and 13C nuclear magnetic resonance (NMR) spectroscopy. The cells were embedded in alginate microcapsules, perfused with growth media and LND at physiological conditions in the NMR tube, and continuously monitored in vivo for the effects of LND. 31P NMR demonstrated intracellular acidification after LND perfusion concomitant with ATP depletion and changes in phospholipid metabolites. 13C NMR showed marked LND-induced accumulation of lactate, and spectra of the perfusate disclosed that LND inhibited lactate transport. Kinetic 13C NMR also furnished information on LND effects on glucose metabolism; LND decreased initial glucose uptake and lactate formation, although the final intracellular glucose levels were higher compared with those in controls. Combined administration of LND and the metabolic inhibitor 2-deoxyglucose yielded additive but not synergistic cytotoxicity and enabled assessment of hexokinase activity. Overall, the results indicate that the major metabolic changes induced by LND are inhibition of lactate transport and its accumulation, which lead to intracellular acidification.

Early biochemical events occurring after infection of Bacillus cereus 569-SP1 with bacteriophage GSW.

After infection of Bacillus cereus 569-SP1 with the 5-hydroxymethyluracil-containing phage GSW, new dTTPase, dUTPase, and dUMP-hydroxymethylase activities appear. No significant changes in activities of other pyrimidine ribonucleoside or 2'-deoxyribonucleoside triphosphate nucleotidohydrolases were detected. dUTP and dUMP inhibit the dTTPase activity, whereas dTTP failed to inhibit dUTPase activity. The K(m) value for the substrate dUTP is 10(-4) M and for dTTP is 4.85 x 10(-4) M. Thymidylate synthetase activity is inhibited only when cells are infected during the late lag or very early log phases of growth; when cells are infected with phage during mid-log, thymidylate synthetase activity is unaffected. The data support the suggestion that, although phage GSW may inhibit an otherwise expected increase in activity of thymidylate synthetase, it fails to affect the already existing activity. The data presented do not allow discrimination as to whether the phage specifies inhibition of de novo synthesis of thymidylate synthetase or the increase in activity of already existing but not fully expressed enzyme.