Clinically overt brain metastases occur in approximately 10%-15% of patients with breast cancer. At autopsy, cerebral metastases are found in up to 30% of breast cancer patients. The incidence of brain metastasis seems to have increased over the past decade and may be the paradoxical result of effectiveness of drugs on primary breast cancer. Perhaps even more alarming is the growing number of breast cancer patients who die from complications related to brain metastasis, at a time when systemic disease is under good control. In part, this may be due to the fact that chemotherapeutic agents that show efficacy against systemic disease may have poor penetration of the blood-brain barrier (BBB), which means that breast cancer metastasis in the brain may remain untreated and inaccessible to conventional chemotherapeutics. It is therefore important to use model systems representative of brain metastases for experimental studies. Tumor microcirculation and oxygenation play important roles in malignant progression and metastasis, as well as response to various therapies. It is postulated that the initial growth of tumor metastasis, in the absence of established vasculature, results in intratumoral hypoxia. We hypothesize that tumor hypoxia is a major driving force for progression of breast cancer brain metastasis and represents a critical target for therapeutic strategies. However, there is little knowledge about tumor hypoxia during brain metastasis development. Traditionally, pathophysiological and biological studies of tumor models required sacrificing animals at different time points, requiring a large number of animals. In vivo imaging promises greater efficiency since each animal serves as its own control and multiple time points can be examined sequentially. In addition to anatomic information, magnetic resonance imaging (MRI) has been increasingly applied to studying tumor pathophysiology. We have developed an MRI approach based on an interleaved T2*- and T1-weighted sequence, which will provide noninvasive monitoring of both tumor vascular and tissue oxygenation. The presence of the BBB presents a huge challenge for effective delivery of therapeutics to brain. Studies have shown that small brain metastases have an intact BBB, which becomes disrupted when the tumor enlarges. We have also developed a MRI approach to study BBB permeability based on dynamic contrast enhanced (DCE) MRI. We plan to apply these MRI approaches to correlating hypoxia with BBB and tumor aggressiveness of breast cancer brain metastasis on both a temporal and spatial basis. Bioluminescence imaging (BLI), based on in vivo expression of luciferase, the light emitting enzyme of the firefly, is being rapidly adopted in cancer research. By genetically engineering breast cancer cells with a hypoxia reporter system, tumor hypoxia will be monitored noninvasively by the cheaper and faster BLI system. Integration of MRI and BLI will provide temporal and spatial information of tumor hypoxia evolution. Acquired information of temporal and spatial correlation between tumor hypoxia and BBB status by in vivo imaging approaches will facilitate a design for optimum combination treatment of radiation and chemotherapeutics on breast cancer brain metastasis. 2-Methoxyestradiol, currently in Phase I/II trials against primary breast cancer, is a naturally occurring estrogen metabolite. Studies have shown that 2-methoxyestradiol is a radiation sensitizer by reducing tumor hypoxia. Thus, we hypothesize that 2-methoxyestradiol will enhance radiation response by modifying tumor hypoxia. We will apply in vivo imaging to monitoring tumor hypoxia in response to 2-methoxyestradiol and therapeutic efficacy of this novel combination treatment on breast cancer brain metastasis. I believe that this research will enhance our understanding of the chronological pathophysiological changes in developing brain metastases from breast cancer, which will further lay the foundation for clinical novel treatment regimes.