ABSTRACT
Since a few years, the acoustic sensing of whole cell is the focus of increasing interest for monitoring the cytoskeletal cellular response to morphological modulators. We aimed at illustrating the potentialities of the quartz crystal microbalance with dissipation (QCM-D) technique for the real-time detection of the earliest morphological changes that occur at the cell-substrate interface during programmed cell death. Human breast cancer cells (MCF-7) grown on serum protein-coated gold sensors were placed in dynamic conditions under a continuous medium flow. The mass and viscoelasticity changes of the cells were tracked by monitoring the frequency and dissipation shifts during the first 4h of cell exposure to staurosporine, a well-known apoptosis inducer. We have identified a QCM-D signature characteristic of morphological modifications and cell detachment from the sensing surface that are related to the pro-apoptotic treatment. In particular, for low staurosporine doses below 1 µM, we showed that recording the dissipation shift allows to detect an early cell response which is undetectable after the same duration by the classical analytical techniques in cell biology. Furthermore, this sensing method allows quantifying the efficiency of the drug effect in less than 4h without requiring labeling and without interfering in the system, thus preventing any loss of information. In the actual context of targeted cancer therapy development, we believe that these results bring new insights in favor of the use of the non invasive QCM-D technique for quickly probing the cancer cell sensitivity to death inducer drugs.
Subject(s)
Apoptosis/drug effects , Biosensing Techniques/instrumentation , Breast Neoplasms/drug therapy , Breast Neoplasms/physiopathology , Drug Screening Assays, Antitumor/instrumentation , Micro-Electrical-Mechanical Systems/instrumentation , Staurosporine/administration & dosage , Antineoplastic Agents/administration & dosage , Breast Neoplasms/diagnosis , Computer Systems , Dose-Response Relationship, Drug , Drug Screening Assays, Antitumor/methods , Equipment Design , Equipment Failure Analysis , Humans , MCF-7 Cells , TransducersABSTRACT
BACKGROUND: Statins have long been used as anti-hypercholesterolemia drugs, but numerous lines of evidence suggest that they may also bear anti-tumour potential. We have recently demonstrated that it was possible to isolate cancer cells adapted to growth in the continuous presence of lovastatin. These cells grew more slowly than the statin-sensitive cells of origin. In the present study, we compared the ability of both statin-sensitive and statin-resistant cells to give rise to tumours in Nude mice. METHODS: HGT-1 human gastric cancer cells and L50 statin-resistant derivatives were injected subcutaneously into Nude mice and tumour growth was recorded. At the end of the experiment, tumours were recovered and marker proteins were analyzed by western blotting, RT-PCR and immunohistochemistry. RESULTS: L50 tumours grew more slowly, showed a strong decrease in cyclin B1, over-expressed collagen IV, and had reduced laminin 332, VEGF and CD34 levels, which, collectively, may have restricted cell division, cell adhesion and neoangiogenesis. CONCLUSIONS: Taken together, these results showed that statin-resistant cells developed into smaller tumours than statin-sensitive cells. This may be reflective of the cancer restricting activity of statins in humans, as suggested from several retrospective studies with subjects undergoing statin therapy for several years.
Subject(s)
Hydroxymethylglutaryl-CoA Reductase Inhibitors/pharmacology , Lovastatin/pharmacology , Stomach Neoplasms/prevention & control , Animals , Blotting, Western , Cell Adhesion/drug effects , Cell Adhesion Molecules/metabolism , Cell Division/drug effects , Collagen/metabolism , Cyclin B1/metabolism , Drug Resistance, Neoplasm , Humans , Immunohistochemistry , Mice , Mice, Nude , Real-Time Polymerase Chain Reaction , Stomach Neoplasms/metabolism , Stomach Neoplasms/pathology , Tumor Cells, Cultured , Vascular Endothelial Growth Factor A/metabolism , KalininABSTRACT
Statins are lipid-lowering drugs that may help limit cancer occurrence in humans. They drive blockage of the mevalonate pathway, trigger cancer cell apoptosis in vitro and reduce tumour incidence in animals. We have shown in the present study that statins induced apoptosis in HGT-1 human gastric cancer cells, and this was prevented by intermediates of the cholesterol synthetic pathway. In addition, similarly to what we have reported previously for caspase 2 [Logette, Le Jossic-Corcos, Masson, Solier, Sequeira-Legrand, Dugail, Lemaire-Ewing, Desoche, Solary and Corcos (2005) Mol. Cell. Biol. 25, 9621-9631], caspase 7 may also be induced by statins and is under the positive control of SREBP (sterol-regulatory-element-binding protein)-1 and -2, major activators of cholesterol and fatty acid synthesis genes, in HGT-1 cells. Knocking down these proteins strongly reduced caspase 7 mRNA and protein expression, and chromatin immunoprecipitation analyses showed that the proximal promoter region of the CASP7 gene could bind either SREBP-1 or -2. Strikingly, cells selected to grow in the continuous presence of statins showed increased expression of caspase 7 mRNA and protein, which was maintained in the absence of statins for several weeks, suggesting that high expression of this caspase might participate in adaptation to blunting of the mevalonate pathway in this model. Taken together, our results show that caspase 7, as an SREBP-1/2 target, can be induced under mevalonate-restricting conditions, which might help overcome its shortage.