Breast cancer is one of the most common cancers among women. To discover molecular targets that are applicable for development of novel breast cancer therapy, we previously did genome-wide expression profile analysis of 81 breast cancers and found dozens of genes that were highly and commonly up-regulated in breast cancer cells. Among them, we here focused on one gene that encodes PDZ-binding kinase/T-LAK cell-originated protein kinase (PBK/TOPK), including a kinase domain. Northern blot analyses using mRNAs of normal human organs, breast cancer tissues, and cancer cell lines indicated this molecule to be a novel cancer/testis antigen. Reduction of PBK/TOPK expression by small interfering RNA resulted in significant suppression of cell growth probably due to dysfunction in the cytokinetic process. Immunocytochemical analysis with anti-PBK/TOPK antibody implicated a critical role of PBK/TOPK in an early step of mitosis. PBK/ TOPK could phosphorylate histone H3 at Ser 10 in vitro and in vivo, and mediated its growth-promoting effect through histone H3 modification. Because PBK/TOPK is the cancer/ testis antigen and its kinase function is likely to be related to its oncogenic activity, we suggest PBK/TOPK to be a promising molecular target for breast cancer therapy. (Cancer Res 2006; 66(18): 9186-95)

To elucidate the molecular mechanisms of mammary carcinogenesis and discover novel therapeutic targets for breast cancer, we previously carried out genome-wide expression profile analysis of

Breast cancer is the most common cancer among women worldwide; the breast cancer incidence and death rates for Japanese were 102.8 and 17.1 per 100 000 population, respectively, in the year 2006.(1,2) Incidence of breast cancer is increasing in most countries including the USA and Japan, and the increasing rate is much higher in countries where its incidence was previously low.(1,2) It has been known that breast cancer is a hormonedependent disease, and estrogens through an interaction with estrogen receptor (ER) drastically enhance the proliferative and metastatic activity in breast tumor cells.(3,4) However, despite the clinical benefit of interruption of the ER function with synthetic anti-estrogen drugs such as tamoxifen, the precise mechanism of an estrogen/ER signaling pathway in breast cancer progression is not well understood. Therefore, further characterization of the pathophysiologic roles of this pathway and development of novel drugs targeting this pathway should be eagerly expected to provide better management for breast cancer patients.Gene-expression profile analysis can generate a considerable amount of information for characterizing the nature of individual cancers; such information should be applied for extraction of potential molecular targets for improving clinical strategies to treat neoplastic diseases.(5,6) Through the genome-wide expression analysis of a large number of microdissected clinical cancer materials, we have identified dozens of genes that function as oncogenes in the process of development and/or progression of breast cancer, (7)(8)(9)(10)(11) bladder cancer, (12,13) synovial sarcomas, (14,15) testicular seminoma, (19,20) and identified dozens of molecules that were overexpressed in a great majority of breast cancers and were low or undetectably expressed in normal human organs.Among many over-expressed genes in breast cancers, we report in this study identification and characterization of a novel gene, brefeldin A-inhibited guanine nucleotide-exchange protein 3 (BIG3), a novel member of the BIG1/Sec7p subfamily of ADP ribosylation factor-GTP exchange factors (ARF-GEFs), to be a key molecule regulating an estrogen/estrogen receptor (ER) signaling pathway in breast cancer. We also demonstrate an interaction of BIG3 with prohibitin 2/repressor of estrogen receptor activity (PHB2/REA) protein, and that their interaction can enhance the ERα transcriptional activity. Our findings imply BIG3 to be a promising target for development of novel anti-cancer drugs for breast cancer. Materials and MethodsCell lines and clinical samples. Human breast cancer cell lines HCC1937, MCF-7, MDA-MB-231, SK-BR-3, T47D, BT-549, HCC1395, MDA-MB-157, BT-20, MDA-MB-453, ZR-75-1, BT-483, BT-474, HCC1143, HCC1500, HCC1599, and OCUB-F, as well as African green monkey SV40-transfected kidney fibroblast cell line, COS-7, were purchased from American Type Culture Collection (ATCC, Rockville, MD, USA), and cultured under their respective depositors' recommendations. All cells were cultured according to previ...

PurposeWe have previously described a novel pathway controlling drug resistance, epithelial-to-mesenchymal transition (EMT) and stemness in breast cancer cells. Upstream in the pathway, three miRs (miR-106b, miR-93 and miR-25) target EP300, a transcriptional activator of E-cadherin. Upregulation of these miRs leads to the downregulation of EP300 and E-cadherin with initiation of an EMT. However, miRs regulate the expression of many genes, and the contribution to EMT by miR targets other than EP300 cannot be ruled out.MethodsWe used lentiviruses expressing EP300-targeting shRNA to downregulate its expression in MCF-7 cells as well as an EP300-knocked-out colon carcinoma cell line. An EP300-expression plasmid was used to upregulate its expression in basal-like CAL51 and MDA-MB-231 breast cancer cells. Drug resistance was determined by short-term proliferation and long-term colony formation assays. Stemness was determined by tumour sphere formation in both soft agar and liquid cultures as well as by the expression of CD44/CD24/ALDH markers. Gene expression microarray analysis was performed in MCF-7 cells lacking EP300. EP300 expression was analysed by immunohistochemistry in 17 samples of metaplastic breast cancer.ResultsCells lacking EP300 became more resistant to paclitaxel whereas EP300 overexpression increased their sensitivity to the drug. Expression of cancer stem cell markers, as well as tumour sphere formation, was also increased in EP300-depleted cells, and was diminished in EP300-overexpressing cells. The EP300-regulated gene signature highlighted genes associated with adhesion (CEACAM5), cytoskeletal remodelling (CAPN9), stemness (ABCG2), apoptosis (BCL2) and metastasis (TGFB2). Some genes in this signature were also validated in a previously generated EP300-depleted model of breast cancer using minimally transformed mammary epithelial cells. Importantly, two key genes in apoptosis and stemness, BCL2 and ABCG2, were also upregulated in EP300-knockout colon carcinoma cells and their paclitaxel-resistant derivatives. Immunohistochemical analysis demonstrated that EP300 expression was low in metaplastic breast cancer, a rare, but aggressive form of the disease with poor prognosis that is characterized by morphological and physiological features of EMT.ConclusionsEP300 plays a major role in the reprogramming events, leading to a more malignant phenotype with the acquisition of drug resistance and cell plasticity, a characteristic of metaplastic breast cancer.Electronic supplementary materialThe online version of this article (doi:10.1007/s10549-017-4202-z) contains supplementary material, which is available to authorized users.

Through analysis of the detailed genome-wide gene expression profiles of 81 breast tumors, we identified a novel gene, G-patch domain containing 2 (GPATCH2), that was overexpressed in the great majority of breast cancer cases. Treatment of breast cancer cells MCF-7 and T47D with siRNA against GPATCH2 effectively suppressed its expression, and resulted in the growth suppression of cancer cells, suggesting its essential role in breast cancer cell growth. We found an interaction of GPATCH2 protein with hPrp43, an RNA-dependent ATPase. Their interaction could significantly enhance the ATPase activity of hPrp43 and induce a growth-promoting effect on mammalian cells. Because northern blot analyses of normal human organs implied GPATCH2 to be a novel cancer/testis antigen, targeting GPATCH2 or inhibition of the interaction between GPATCH2 and hPrp43 could be a promising novel therapeutic strategy of breast cancer. (Cancer Sci