Despite the scientific progress in the last decades in terms of therapeutic agents to fight cancer there is still the need of developing safer and more effective drugs. Developing an innovative drug is not only very expensive, but also highly time consuming; furthermore, the number of anticancer agents that fail in clinical trials with high attrition rates mainly caused by unexpected toxicity and lack of efficacy far outweighs those considered effective, which indicates that drug screening processes require further improvements. In this project the application of Fourier transform infrared microspectroscopy is evaluated in order to develop a spectral based model that could be used to describe the drug-cell interaction and also to discriminate between the metabolic modifications due to a particular drug and the inherent cell cycle of a cell. A computational method was built using the FTIR spectra from a highly resistant renal cell carcinoma cell line, Caki-2, in order to discriminate between the phases of the continuous cell cycle that cells undergo while proliferating in vitro. Such model enabled the discrimination of early events of the cell cycle (G0/G1 phase cells) from G2/M phase cells with a prediction accuracy of 90% and 92.9% respectively. On the other hand, when the RMieS-corrected FTIR spectra corresponding to G0/G1, S and G2/M phases were modelled, the algorithm was able to retrieve each stage of proliferation with 82.3%, 71.8% and 84.4% accuracy respectively. Although the average accuracy yielded by the method was relatively low compared with what has previously been reported in the literature, these results emphasize the need to correct the data from physical distortions due to size and prove the principle that it is possible to create a method for identifying different events of the cell cycle based on the data that the FTIR spectroscopy provides, as well as using the scattering profile characteristic of each phase of development. Once the underlying biochemistry of proliferating Caki-2 cells were characterised by FTIR, the cells were treated with 5-Fluorouracil and paclitaxel, two widely used cytotoxic agents known to induce cellular damage at S or G2/M phase of the cell cycle respectively. The FTIR spectra collected were analysed via multivariate and bivariate techniques. Results demonstrated that, after 24 hours of treatment at the IC50 concentration of each drug, Caki-2 cells displayed spectral features consistent with early stages of apoptosis. These spectral characteristics did not appear to be linked either to the drug’s mode of action or the cell’s cycle phase. The cell’s proliferation stage was not the main classification trend among the drug-treated spectra; nevertheless, the cell cycle phase of each drug-treated population was successfully retrieved by an optimized model capable to classify such phases with an average accuracy of 77.98%. Altogether, this study offers a new perspective when analysing FTIR data from single cells as a function of the cell cycle and also when investigating the biochemical response of a cell line to a given anticancer agent.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:626974 |
| Date | January 2014 |
| Creators | Jimenez Hernandez, Melody |
| Contributors | Gardner, Peter |
| Publisher | University of Manchester |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | https://www.research.manchester.ac.uk/portal/en/theses/the-development-of-an-infrared-method-for-the-characterization-of-drugcell-interactions(91b99ca2-691e-4032-9274-b4f2aeb64024).html |
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