Inhibiting inflammatory processes or eliminating inflammation represents a logical role in the suppression and treatment strategy of cancer. Several studies have shown that anti-inflammatory drugs (NSAIDs) have promise as anticancer agents while reducing metastases and mortality. NSAIDs are seriously limited by side effects and their toxicity, which can become cumulative with their long-term administration for chemoprevention. The huge development in nanotechnology allows the drugs to exhibit novel and significantly improved properties compared to the large particles of the respective bulk compound, leading to more targeted therapy and reduced dosage. The overall aim of this thesis is to add to our understanding of cancer prevention and treatment through studying the genotoxicity mechanisms of NSAIDs agents in lymphocytes. In this study, the genotoxicity mechanisms of NSAID in bulk and nanoparticles forms a strategy to prevent and minimise the damage in human lymphocytes. Aspirin nano (ASP N) caused a significant decrease in deoxyribonucleic acid (DNA) damage compared to aspirin bulk (ASP B). Also, ibuprofen nano (IBU N) showed a significant reduction in DNA damage compared to ibuprofen bulk (IBU B). Micronuclei (MNi) decreased after ASP N, ASP B and IBU N in prostate cancer patients and healthy individuals, and the ibuprofen bulk showed a significant increase of MNi formation in lymphocytes from healthy and prostate cancer patients when compared to untreated lymphocytes from prostate cancer patients. In order to study the geno-protective properties of these drugs, the protective effect of NSAIDs and the quantification of the DNA repair capacity in lymphocytes was studied. ASP N was found to increase the DNA repair capacity and reduced the reactive oxygen species (ROS) formation significantly more than ASP B. Finally, the role of NSAIDs on some key regulatory signal transduction pathways in isolated lymphocyte cells was investigated by studying their effect on ataxia-telangiectasia-mutated kinase (ATM) and ataxia-telangiectasia and Rad3-related kinase (ATR) mRNA. ATM mRNA significantly increased after treatment with ASP B, ASP N and IBU N. ATR expression also increased after treatment with IBU B and IBU N, but was only significant with IBU N. These findings indicate that a reduction in particle size had an impact on the reactivity of the drug, further emphasising the potential of nanoparticles as improvement to current treatment options.

Thesis (Ph.D. in Molecular Biology) -- University of Colorado at Denver and Health Sciences Center, 2006. / Typescript. Includes bibliographical references (leaves 119-130). Free to UCDHSC affiliates. Online version available via ProQuest Digital Dissertations;

Abasic sites (Ap site, from apurinic/apyrimidinic) are one of the most common lesions generated in DNA by spontaneous base loss or DNA repair processes. There are two equilibrating forms of an Ap site - ring-open aldehyde and cyclic hemiacetal. Ring- opened aldehydes are reactive electrophilic groups capable of formation covalent adduct with nucleophilic sites in DNA. DNA interstrand cross-link (ICL) resulting from the Ap sites is formed spontaneously as a covalent bond between ring-open aldehyde and amin group of adenin residue in the opposite strand of double stranded DNA. ICLs block DNA replication and transcription. The formation of Ap site derived ICL is relatively long process taking several hours. We assume that the ring-opening of an abasic site is the rate-limiting step in the formation of the thermodynamic ICL. However, formation, stability and DNA repair of Ap-ICL are still poorly understood processes. Here, I have set up mechanistic in vitro experiments to reveal and calculate the probability of Ap-ICl formation in vivo. In more detail, I study the rates of formation of Ap-ICLs in the sequence context of neighbouring nucleotides of freshly formed covalent bond of ICL. I focus on sequence preference, the influence of AT/ GC rich regions and the length of oligonucleotides. I have...

The homologous recombination (HR) pathway contributes to genome integrity by mediating double strand break (DSB) repair using a homologous DNA sequence as a template. In mammals Rad51 and Brca2 are molecules central to this process. Little is known about HR repair in Dictyostelium. However, research previously conducted on DSB repair using this organism has shown that DSB repair pathways are highly conserved when compared to humans. This encouraged study of HR in this organism. In this study, through a bioinformatics search I have identified putative orthologues of most human HR proteins and most interestingly of BRCA2, which cannot be found in other lower eukaryotes used as models for DSB repair, such as the budding yeast S.cerevisiae. Brcp, the Dictyostelium BRCA2 ortholog, shows similar domain structure when compared to BRCA2-related proteins identified in other organisms. To verify the implication of HR proteins in DSB repair, I developed a method to monitor recruitment of DNA repair proteins on chromatin upon DSB induction. Findings of this study suggest that both Brcp and Rad51 get recruited to chromatin upon DSB induction and are therefore implicated in DSB repair in Dictyostelium. To further study Brcp function and based on findings suggesting that disruption of brcp might be lethal, I developed a novel system for specific and conditional depletion of endogenous Dictyostelium proteins. Utilizing this system, I conducted phenotypic studies in a strain depleted of Brcp to examine its role in DNA repair. Overall this study shows that the HR pathway in Dictyostelium shows great similarity to vertebrates, making Dictyostelium an appealing model for the study of DSB repair and specifically HR.