Improvements to radiation therapy treatment outcomes rely, in part, on consideration of patient specific radiosensitivity. Therefore an assay which quantifies radiation-induced biochemical changes, and subsequently characterizes radiation responses in tumour and normal tissue is required. This work investigates the use of a single cell Raman spectroscopic technique to identify radiation-induced responses in human lung (H460), breast (MCF-7, MDA-MB-231) and prostate (LNCaP) cancer cell lines each selected to create a panel of cells varying in tissue of origin and radiation sensitivity. Cells were cultured in vitro and exposed to clinically relevant (< 10 Gy) doses of radiation. Cell populations receiving 30 and 50 Gy were also studied in order to further elucidate dose-dependent trends and give additional information about the ability of Raman spectroscopy to identify radiation-induced biochemical changes occurring in the cell populations. It was found that using Raman spectroscopy to identify a radiation-induced response in human cells cultured in vitro is governed by the subtlety of the radiation-induced response inherent to a specific cell line. Also the type of biochemical changes occurring in response to exposure to radiation will be dependent on the specific cell line. This work suggests that effective use of single cell Raman spectroscopy to monitor a patient's response to radiation early on in treatment (where the disease has been exposed to 10 Gy or less) is possible. However, a detailed knowledge of the biochemical changes associated with the unique radiation response for that particular disease is required. / Graduate / 0541 / 0752 / 0992 / hardersj@uvic.ca

In this study the direct effect of ionising radiation on DNA plus additives has been studied using both ESR spectroscopy and plasmid DMA (for strand break analysis). The primary radicals were identified as the thymine radical-anion, T', and guanine radical-cation, G'*'. Under normal conditions these were formed in approximately equal yields as defined by careful computer simulations. Certain additives such as oxygen, nitroimidazoles, silver ions and the rest of the nuclear complement (i.e. RNA and histone proteins) , were added to study their effects on the relative yields of T" and G". In all cases, they were shown to capture electrons in competition with T" and have little or no effect on the yield of G"*". In the case of oxygen and nitroimidazoles the effect of reducing the yield of T" radicals was looked at using strand break analyses. Essentially this was found to protect the DNA. Since both single and double strand breaks were found at significant levels when G+ and T~ were the only detectable initial radicals, one must conclude that these radicals are responsible for strand breaks. Fran the relatively high number of double strand breaks found, we deduce that G'*' and T" centres must be close togetlier (in a range of ca. 10-50 A), and that both may give rise to strand breaks, by as yet undefined pathways. In a separate study (Chapter 4), the reaction between superoxide ions, O2"/ and dimethyl formamide has been investigated by ESR spectroscopy. Strong evidence in favour of addition of O2" at the C=0 group to give a relatively stable peroxy radical intermediate has been obtained. This has implications for the mechanism of action of O2" formed both as a result of radiation damage and by other means. Appendix I describes a study of various simple aldehyde and ketone radical-cations, using ESR spectroscopy. Interpretations of these spectra are given, together with structural implications. Appendix II is a paper on work carried out on the ESR spectra of hydroxyl radicals in aqueous glasses. This work was done in collaboration with H. Riederer and J. Hiittermann.

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