Masamitsu Onda

Amplification, up‐regulation and over‐expression of DVL‐1, the human counterpart of the Drosophila disheveled gene, in primary breast cancers

Wnt proteins form a family of highly conserved, secreted signaling molecules that regulate cell‐to‐cell interactions during embryogenesis. Wnt genes and Wnt signaling are also implicated in cancer. It has been shown that Wnt proteins bind to receptors of the frizzled family on the cell surface. Through several cytoplasmic relay components including DVL‐1, the human counterpart of the Drosophila disheveled gene, the signal is transduced to β‐catenin, which then enters the nucleus and forms a complex with T‐cell factor (TCP) to activate transcription of Wnt target genes. We describe here the amplification of DVL‐1 in 13 of 24 primary breast cancers examined, and increased expression of this gene in 11 of those tumors in comparison to corresponding non‐cancerous breast tissues. Immunohistochemical staining demonstrated that DVL‐1 protein was prominent in the cytoplasm of cancer cells, but not in normal epithelial cells of the mammary duct or in myoepithelial cells. These data indicate that amplification and increased expression of the DVL‐1 gene may play some role in human breast carcinogenesis through derangement of the Wnt signaling pathway.

Upregulation and Overexpression of Human X-box Binding Protein 1 (hXBP-1) Gene in Primary Breast Cancers

BackgroundHuman X-box binding protein 1 (hXBP-1) is a transcription factor essential for hepatocyte growth as well as for plasma cell differentiation. hXBP-1 also binds to cis-elements of human T cell leukemia virus and human major histocompatibility complex genes. In order to clarify the role of XBP-1 in breast cancer, here we investigated the expression of XBP-1 in 11 primary breast cancers and 5 breast cancer cell lines.Materials and MethodsThe study population consisted of eleven patients who were underwent surgery for breast cancer from 2000 to 2002. Five breast cancer cell lines (MDA-MB-453, CRL1500, YMB-1-E, MCF7 and HBL100) were analyzed for XBP-1 expression. Reverse transcription Polymerase chain reaction was performed on 6 primary breast cancers. Then we investigated XBP-1 expression by immunohistochemically on archived paraffin-embedded sections.ResultshXBP-1 mRNA expression was increased in all 11 primary breast cancers we examined, as well as 5 breast cancer cell lines, but hardly detectable in non-cancerous breast tissue. Immunohistochemical staining demonstrated that hXBP-1 protein stained strongly in the cytoplasm of cancer cells but was unreactive in the normal breast ductal epithelial and myoepithelial cells.ConclusionsThese data indicate that increased expression of the hXBP-1 gene may play some role in human breast carcinogenesis through impairment of cell differentiation regulation.

Up-regulation of CC chemokine, CCL3L1, and receptors, CCR3, CCR5 in human glioblastoma that promotes cell growth

Human CC ligand 3-like protein 1 (CCL3L1), a member of the CC chemokine family, that induces MCP1 and RANTES, exhibits a variety of proinflammatory activities including chemotaxis, and functional and proliferative activation of leukocytes, lymphocytes and macrophages. Its signal is transmitted through transmembrane receptors, CC chemokine receptors, CCR1, CCR3 and CCR5. To examine gene expression of chemokine, CCL3L1, and its receptors, CCR1, CCR3 and CCR5, we analyzed tumor tissues from 21 patients with several types of primary gliomas. CCL3L1, CCR3 and CCR5 gene exhibited over-expression in 70% (7/10), 60% (6/10), and 60% (6/10) of glioblastoma, in comparison with lower frequencies seen in lower-grade gliomas. Transfection of CCL3L1-expression vector to glioblastoma cell line enhanced proliferation of the tumor cells. These data suggest that increased expression of the CCL3L1, CCR3 and CCR5 chemokine-receptors system is involved in brain tumorigenesis, especially in the progression of glioblastoma.

Identification of histological markers for malignant glioma by genome-wide expression analysis: dynein, α-PIX and sorcin

Glioblastoma multiforme (GBM), the most malignant class of glial neoplasm (grade IV in WHO criteria), carries the worst clinical prognosis among primary brain tumors in adults. To identify a set of genes involved in the tumorigenesis of GBM, we evaluated expression profiles of GBM tissues from 11 patients using a cDNA microarray representing 25,344 human genes. By comparing the profiles with those of normal brain tissue, we identified a number of differentially expressed genes: 54 with increased expression and 45 with reduced expression in GBMs. Semi-quantitative RT-PCR experiments with 6 of those genes confirmed higher expression of DNCH2, ARHGEF6, NPM1 and SRI and lower expression of NRGN and TM4SF2 in GBM tumors. Immunohistochemical staining for 3 of the respective gene products, dynein (product of DNCH2), α-PIX (product of ARHGEF6), and sorcin (product of SRI) indicated that this technique might be useful for histological grading of glial tumors. To establish criteria for this diagnostic approach, we scored glial tumor tissues of different histological grades according to the staining results; the scores were significantly higher in anaplastic astrocytomas and GBMs than in diffuse astrocytomas or normal brain tissues. These findings indicated that levels of these three proteins might serve as histological markers for malignant glioma classification.

Expression profiling to predict postoperative prognosis for estrogen receptor-negative breast cancers by analysis of 25,344 genes on a cDNA microarray

Estrogen receptor (ER) status is an essential determinant of clinical and biological behavior of human breast cancers. While ER‐positive breast cancers respond well to adjuvant hormone therapy, ER‐negative tumors are generally resistant. To date, no attempts have succeeded in finding molecular markers for classifying ER‐negative breast cancers with respect to postoperative prognosis. To identify a set of prognostic markers for this type of cancer, we used a cDNA microarray consisting of 25,344 human genes to investigate expression profiles of ten primary breast cancers from patients who had died of breast cancer within 5 years after surgery (5y‐D) and 10 from patients who had survived disease‐free for more than 5 years (5y‐S). Sets of genes characterizing each group were identified by Mann‐Whitney and random‐permutation tests. We documented 71 genes with higher expression in the 5y‐D group than in the 5y‐S group, and 15 with higher expression in the 5y‐S group than in the 5y‐D group. Semi‐quantitative RT‐PCR experiments were carried out to confirm the results of the microarray analysis. We established a scoring system for predicting postoperative prognosis of ER‐negative breast cancers on the basis of aberrant gene expression. The list of genes reported here provides valuable information with regard to progression of breast cancer and is a source of possible target molecules for development of novel drugs to treat patients with ER‐negative breast cancers.

Up-regulation of transcriptional factor E2F1 in papillary and anaplastic thyroid cancers

AbstractExpression of genes in the Rb-E2F signaling pathway is controlled by E2F transcriptional factors originally defined as molecules that bind to the promoter of E2 adenovirus. The E2F gene family consists of six members and is designated E2F1-6. The Rb-E2F signaling pathway is among the main regulators of the cell cycle, hence its importance in differentiation and oncogenesis. We document here up-regulation of E2F1, but not other members of the E2F gene family, in 15 of 18 primary papillary thyroid cancers examined (83%) in comparison to corresponding noncancerous thyroid tissues and in all of 11 anaplastic thyroid cancer (ATC) cell lines (100%). The E2F4gene, however, was down-regulated in 12 of the papillary thyroid cancers (67%). Immunohistochemical analysis with antibody to E2F1 revealed prominent intracellular E2F1 protein in most of the primary papillary cancers (16 of 18; 89%) but was not detectable in normal thyroid tissues. These data indicated that increased expression of the E2F1 gene might play a significant role in human thyroid carcinogenesis through derangement of the Rb-E2F signaling pathway.

Overexpressed in anaplastic thyroid carcinoma-1 (OEATC-1) as a novel gene responsible for anaplastic thyroid carcinoma

Anaplastic thyroid carcinoma (ATC) is one of the most fulminant human malignancies. However, the molecular carcinogenic mechanisms of ATC are understood poorly. Recently, the authors performed a cyclic DNA (cDNA) microarray analysis with 11 anaplastic thyroid carcinoma cell lines (ACLs) and discovered several novel responsible genes for ACLs and ATC. From the extended list, they focused on hypothetical and anonymous genes and investigated a novel gene, named the overexpressed in anaplastic thyroid carcinoma‐1 (OEATC‐1) gene.

Amplification, up-regulation and over-expression of C3G (CRK SH3 domain-binding guanine nucleotide-releasing factor) in non-small cell lung cancers

AbstractThe Ras-CRK-Rap1 cellular signal-transduction system is regulated by guanine nucleotide exchange factors (GEFs). Transcription of C3G on chromosome 9q34 and a key member of the GEF gene family is activated by the CRK-adaptor protein; the C3G product is a CRK SH3 domain-binding guanine nucleotide-releasing factor. We document here the amplification of C3G in five of 18 primary non-small cell lung cancers examined and its increased expression in 18 of 28 tumors in comparison to corresponding non-cancerous lung tissues. Immunohistochemical staining revealed prominent C3G protein in the cytoplasm of cancer cells, associated with faint staining at the nucleolar membrane, but C3G was not detectable in normal bronchial mucoepithelial cells or in broncholoalveolar cells of the bronchial/bronchiolar ducts or alveoli. These data indicate that amplification and increased expression of the C3G gene may play some role in human lung carcinogenesis through derangement of the CRK-Rap1 signaling pathway.

Down-regulation of members of glycolipid-enriched membrane raft gene family, MAL and BENE, in cervical squamous cell cancers

Persistent human papillomavirus infections cause infected epithelial cells to lose cellular polarity leading to cell transformation. Glycolipid‐enriched membrane (GEM) rafts are implicated in polarized sorting of apical membrane proteins in epithelial cells and even in signal transduction. The MAL and BENE are essential component of the GEM rafts machinery for apical sorting of membrane proteins. In this study we demonstrated down‐regulation of MAL and BENE mRNA in over two‐thirds of primary cervical squamous cell cancers (14 and 15 of 20 cases, for MAL and BENE, respectively) when compared to corresponding non‐cancerous uterine squamous cells. Allelic loss or hyper‐methylation was not accompanied by MAL or BENE mRNA down‐expression in human primary cervical cancers in microsatellite allelic analysis and HpaII‐PCR‐based methylation analysis of the MAL and BENE genomic region. In addition, we note down‐regulation of these genes in established cervical cancer cell lines. These results suggest that down‐regulation of MAL and BENE genes, which are essential components of the cellular polarized sorting system, play an important role in human cervical squamous cell cancer development.

Genetic Prognostic Index Influences Patient Outcome for Node-Positive Breast Cancer

PurposeTo establish a novel genetic prognostic index among node-positive breast cancer patients.MethodsUsing a cDNA microarray, the gene expression profiles of 20 primary breast cancers that had metastasis to four or more axillary lymph nodes were examined. Ten patients survived disease-free for more than 5 years (5S), while ten patients died of breast cancer within 5 years of surgery (5D).ResultsA set of genes characterizing each group was identified. Sixteen genes were underexpressed in 5D compared to 5S, and 15 genes were underexpressed in 5S in comparison to 5D. The prognostic index (PI) was established, which could predict the postoperative outcome with five genes that were commonly underexpressed in the 5D group; these genes encoded granulin (GRN), heat shock 90 kDa protein 1 beta (HSPCB), large tumor suppressor homolog 1 (LATS1), valosin-containing protein (VCP), and LIM-and-SH3 protein1 (LASP1).ConclusionThese five genes might play an important role in deciding the behavior of node-positive breast cancer. The PI system could thus predict the prognosis of node-positive breast cancer, and might therefore be able to provide valuable information for the prognosis of breast cancer patients.