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Acute and chronic hypoxia exists within the microenvironment of solid tumours and drives therapy resistance, genetic instability and metastasis. Despite its importance in solid tumour progression, very little is known regarding the functional consequences of hypoxia-mediated changes in the expression of DNA repair proteins. I studied the relationship between hypoxia and DNA repair using a prolonged chronic hypoxic gas treatment model in a variety of human tumour cell lines to mimic the dynamic state of proliferation and DNA repair in cells distant from the tumour blood vasculature. I observed decreased expression of homologous recombination (HR) and base excision repair (BER) proteins due to a novel mechanism involving decreased protein synthesis. Error-free HR was suppressed 3-fold under 0.2% O2 as measured by the DR-GFP reporter system and functional BER was impaired as assessed with a functional glycosylase assay. This decrease in protein expression and function resulted in increased sensitivity to the DNA damaging agents MMC, cisplatin, H2O2 and MMS. Additionally, chronically hypoxic cells were relatively radiosensitive (OER = 1.37) when compared to acutely hypoxic or anoxic cells (OER = 1.96 - 2.61). As HR defects are synthetically lethal with poly(ADP-ribose) polymerase 1 (PARP1) inhibition, I evaluated the sensitivity of repair-defective hypoxic cells to PARP inhibition. I observed increased clonogenic killing in HR-deficient hypoxic cells following inhibition or depletion of PARP1. PARP-inhibited hypoxic cells accumulated γH2AX foci consistent with an accumulation of collapsed replication forks. Additionally, tumour xenografts exposed to PARP1 inhibition showed increased γH2AX and cleaved caspase-3 expression in hypoxic subregions with suppressed RAD51 protein expression and decreased ex vivo clonogenic survival. I conclude that persistent down-regulation of DNA repair components by the microenvironment could result in faulty DNA repair with significant implications for therapeutic response and genetic instability in human cancers. Specifically, hypoxic cells may be sensitized to PARP inhibitors and other agents targeting repair pathways down-regulated by hypoxia as a consequence of microenvironment-mediated “contextual synthetic lethality”.

Approved for public release; distribution is unlimited / The National Aeronautic and Space Administration (NASA) and its prime contractors currently use a software tool called RMAT (the Reliability and Maintainability Assessment Tool) for the forecasting of Orbital Replacement Unit (ORU) failure rates and associated maintenance demands for the International Space Station (ISS). This thesis introduces a new model: CMAM (the Comparative Maintenance Analysis Tool), which was developed to replicate some of the basic functionality of RMAT in order to provide a comparative look at RMAT results. The CMAM program, developed in Visual Basic.net and dynamically linked to a Microsoft ACCESS database, focuses on a representative set of critical Orbital Replacement Units (ORUs that represent key items that require both internal and external maintenance in both pressurized and un-pressurized storage) and generated failure rate data for each critical ORU. The results of the CMAM model are then compared with the failure rates generated by RMAT program for the same set of critical ORUs. These two independently developed sets of data are then analyzed against historic failure rates for these ISS parts. / Major, United States Army