Interactions between tumor cells and the microenvironment are crucial to tumor formation and metastasis. The central nervous system serves as a “sanctuary” site for metastasis, resulting in poor prognosis in diagnosed patients. The incidence of brain metastasis is increasing; however, little is known about interactions between the brain and metastatic cells. Brain pathology was examined in an experimental model system of brain metastasis, using a subline of MDA-MB-231 human breast cancer cells. The results were compared with an analysis of sixteen resected human brain metastases of breast cancer. Experimental metastases formed preferentially in specific brain regions, with a distribution similar to clinical cases. In both the 231-BR model, and in human specimens, Ki67 expression indicated that metastases were highly proliferative (~50%). Little apoptosis was observed in either set of tumors. In the model system, metastases elicited a brain inflammatory response, with extensive reactive gliosis surrounding metastases. Similarly, large numbers of glial cells were found within the inner tumor mass of human brain metastases. In vitro co-cultures demonstrated that glia induced a ~5-fold increase in metastatic cell proliferation (P < 0.001), suggesting that brain tissue secretes factors conducive to tumor cell growth. Molecules used to signal between tumor cells and the surrounding glia could provide a new avenue of therapeutic targets for brain metastases.

Brain metastases of breast cancer seem to be increasing in incidence as systemic therapy improves. Metastatic disease in the brain is associated with high morbidity and mortality. We present the first gene expression analysis of laser-captured epithelial cells from resected human brain metastases of breast cancer compared with unlinked primary breast tumors. The tumors were matched for histology, tumor-node-metastasis stage, and hormone receptor status. Most differentially expressed genes were down-regulated in the brain metastases, which included, surprisingly, many genes associated with metastasis. Quantitative real-time PCR analysis confirmed statistically significant differences or strong trends in the expression of six genes: BMP1, PEDF, LAMγ3, SIAH, STHMN3, and TSPD2. Hexokinase 2 (HK2) was also of interest because of its increased expression in brain metastases. HK2 is important in glucose metabolism and apoptosis. In agreement with our microarray results, HK2 levels (both mRNA and protein) were elevated in a brain metastatic derivative (231-BR) of the human breast carcinoma cell line MDA-MB-231 relative to the parental cell line (231-P) in vitro. Knockdown of HK2 expression in 231-BR cells using short hairpin RNA reduced cell proliferation when cultures were maintained in glucoselimiting conditions. Finally, HK2 expression was analyzed in a cohort of 123 resected brain metastases of breast cancer. High HK2 expression was significantly associated with poor patient survival after craniotomy (P = 0.028). The data suggest that HK2 overexpression is associated with metastasis to the brain in breast cancer and it may be a therapeutic target. (Mol Cancer Res 2009;7(9):1438-45)