Abstract 473: Generation of a mouse model of breast cancer brain metastasis

Tumor metastasis to the brain is a common complication of cancer, affecting 500,000 patients each year. Despite aggressive treatment, survival at one year following diagnosis remains a dismal ∼20%. Due to poor penetration of most chemotherapeutics into the brain, brain metastasis may occur even while systemic disease is under control. Unfortunately, the incidence of breast cancer brain metastasis is increasing, as the increased survival of patients achieved with current therapeutic advances has led to an ‘unmasking’ of brain metastases. Given the dire clinical situation, there is a clear need to develop appropriate animal models for studying the mechanism of brain metastasis and for testing potential therapeutics. The goal of this project is to develop a mouse model of breast cancer brain metastasis in an immune-competent system. To accomplish this, we are delivering luciferase-expressing 4T1 cells into the brain of syngeneic Balb/c mice by intra-arterial injection through the internal carotid to mimic the vascular delivery in the native metastatic process. The growth of cells is analyzed non-invasively by Xenogen In Vivo Imaging System (IVIS) and selective animals are imaged by MRI. Tumors are harvested and cultured or fixed and analyzed by immunohistochemistry. We first determined the optimal number of injected cells to result in 100% tumor take with host survival of at least 25 days. We then performed serial rounds of culture of isolated experimental brain metastases, and re-injection into animals to produce sublines with enhanced brain metastatic potential and increased selectivity for brain. Four rounds of selection resulted in the BR1, BR2, BR3, and BR4 4T1 sublines. The series exhibits increasing proliferation rate in vitro as measured by MTS assay and increasing aggressive growth in vivo as evidenced by decreasing survival time post-injection despite a reduction in number of injected tumor cells. Both micro-and macro-metastatic tumor cell clusters are observed, with most of the macro-metastases occurring in the frontal and parietal cortex. These metastases are characterized by a high proliferation rate as evidenced by Ki67 staining, high Cox2 expression, and significant reactive gliosis in the surrounding brain tissue as evidenced by GFAP staining. We have also observed that, in vitro, BR4 4T1 cells passaged through the brain four times exhibit increased ability to invade through extracellular matrix compared to parental cells. We are currently per