Key words: cytokine; IL-8; protein arrays; breast cancer; estrogen receptor; tumor progressionCytokines, cell communication mediators, play important roles in a wide range of physiological processes, such as cell growth, differentiation, apoptosis, wound healing and homeostasis. 1 A wealth of evidences suggests that cytokines, growth factors, chemokines, angiogenic factors and proteases are involved in cancer progression. 2-5 A number of autocrine and paracrine cytokine loops influence tumor development. 6 -9 Cytokines not only provide defense against cancer cells, but also promote cancer cell growth at every stage of cancer development. 10 Alteration of cytokine levels is associated with cancer progression, 11,12 response to chemotherapy 13,14 and metastatic status. 15,16 Therefore, determination of these factors in human breast cancer cells will shed light on the development of human breast cancer, identify new molecular targets for developing effective agents against breast cancer, discover biomarkers to aid in diagnosis and to predict clinical outcome, and eventually provide guideline for individual therapy. Furthermore, despite recent improvements in curing breast cancer patients, breast cancer still remains a serious threat to women's lives. Therefore, it is imperative to identify new targets for the development of effective treatment strategy. Because of the limitation of technology, however, previous studies only measured single or few cytokines at once. This greatly limits our understanding of the roles of those factors in breast cancer and the potential application in clinical diagnosis and prediction of clinical outcome.We recently developed several novel cytokine antibody array systems, which allow investigators to detect multiple cytokine expression levels simultaneously with high specificity and sensitivity. [17][18][19][20] By using human cytokine antibody array technology, we have begun to analyze differential cytokine expression levels in various human cancers, including human breast cancer, with the goal to identify key factor(s) involved in human breast cancer progression. Human cytokine antibody arrays indicate that expression of IL-8 is linked to ER status, metastasis status and vimentin status. Tumor biology experiments suggest that IL-8 is involved in breast cancer cell invasion and angiogenesis. Furthermore, ER␣ directly downregulates IL-8 expression. Our results suggest a potential new pathway involved in ER␣ inaction and upregulation of IL-8, leading to the invasiveness and angiogenesis of human breast cancer.

Tissue factor (TF) gene expression is rapidly induced in epithelial cells by phorbol 12-myristate 13-acetate and serum. We have shown that this induction is mediated by a novel serum response region (SRR) (؊111 to ؉14 bp) within the human TF promoter. In this study, we characterized cis-acting genetic elements within the SRR that regulated basal and inducible expression of the TF gene in HeLa cells. Gel mobility shift assays using oligonucleotides spanning the entire SRR identified three 12-base pair (bp) motifs within subregions 1, 2, and 3 that bound constitutively expressed Sp1 and inducibly expressed EGR-1. Analysis of protein binding to these 12-bp motifs by competition with Sp1 and EGR-1 sites, mutation, and antibody supershift experiments indicated that they each contained distinct EGR-1 and Sp1 sites that overlapped by 6 bp. Functional studies using HeLa cells transfected with plasmids containing the wild-type TF promoter (؊111 to ؉14 bp) or derivatives containing mutations in the three Sp1 and/or EGR-1 sites examined basal and inducible expression. The Sp1 sites mediated basal promoter activity, and both Sp1 and EGR-1 sites were required for maximal induction of the TF promoter by phorbol 12-myristate 13-acetate or serum. These data indicated that TF gene expression in HeLa cells was regulated by both Sp1 and EGR-1.