Towards the identification of new markers of pancreatic cancer by gene expression analysis

The poor prognosis of pancreatic cancer has remained unchanged for many years, with a 5-year survival of less than 5 %. Current methods for diagnosing pancreatic cancer are inadequate at identifying small tumors that can be resected by surgery. Characterization of gene expression patterns in pancreatic cancer provided a list of genes that are specifically overexpressed in cancer cells. These genes are putative novel markers for the diagnosis and the prognosis of pancreatic cancer and for the development of targeted therapies. Gene expression analysis should lead to the discovery of molecular markers for early detection of pancreatic cancer that could benefit patients at high risk of developing pancreatic cancer.     

CD4 lymphopenia as a risk factor for febrile neutropenia and early death after cytotoxic chemotherapy in adult patients with cancer

BACKGROUND: Lymphopenia is frequently observed in patients with cancer and correlates with the risk of febrile neutropenia and early death after chemotherapy. The phenotype of the depleted lymphocyte populations was investigated in the current study. METHODS: Peripheral blood lymphocyte subsets (CD3, CD4, CD8, CD19, CD56) were quantified on Day 1 using fluorescence-activated cell sorting in a prospective study of 213 patients with cancer treated with chemotherapy in a single oncology ward during 12 months. Correlations between lymphocyte phenotype, clinical characteristics, and the risk of febrile neutropenia and early death within 31 days after chemotherapy were investigated in univariate and multivariate analyses. RESULTS: Total lymphocyte count and CD3, CD4, and CD8 lymphocyte subsets were significantly lower in patients who experienced febrile neutropenia. Total lymphocyte count and CD3, CD4, CD8, CD19, and CD56 lymphocyte subsets were significantly lower in patients who died within 31 days after chemotherapy. Using logistic regression, CD4 lymphopenia (< 450/muL; odds ratio [OR] = 2.9, 95% confidence interval [CI] = 1.5-5.9) and the dose of chemotherapy (OR = 3,9, 95% CI = 2.0-7.8) were both identified as independent risk factors for febrile neutropenia. Fifty-four percent of patients with both risk factors experienced febrile neutropenia. CD4 lymphocyte count < 450/muL was also an independent risk factor for early death (OR = 7.7, 95% CI = 1.7-35). Thirteen percent of patients with a CD4 lymphocyte count 相似文献   

Molecular classifiers for gastric cancer and nonmalignant diseases of the gastric mucosa

High incidence of gastric cancer-related death is mainly due to diagnosis at an advanced stage in addition to the lack of adequate neoadjuvant therapy. Hence, new tools aimed at early diagnosis would have a positive impact in the outcome of the disease. Using cDNA arrays having 376 genes either identified previously as altered in gastric tumors or known to be altered in human cancer, we determined expression signature of 99 tissue fragments representing normal gastric mucosa, gastritis, intestinal metaplasia, and adenocarcinomas. We first validated the array by identifying molecular markers that are associated with intestinal metaplasia, considered as a transition stage of gastric adenocarcinomas of the intestinal type as well as markers that are associated with diffuse type of gastric adenocarcinomas. Next, we applied Fisher's linear discriminant analysis in an exhaustive search of trios of genes that could be used to build classifiers for class distinction. Many classifiers could distinguish between normal and tumor samples, whereas, for the distinction of gastritis from tumor and for metaplasia from tumor, fewer classifiers were identified. Statistical validations showed that trios that discriminate between normal and tumor samples are powerful classifiers to distinguish between tumor and nontumor samples. More relevant, it was possible to identify samples of intestinal metaplasia that have expression signature resembling that of an adenocarcinoma and can now be used for follow-up of patients to determine their potential as a prognostic test for malignant transformation.     

Evidence of antiangiogenic and antimetastatic activities of the recombinant disintegrin domain of metargidin

Metargidin, a transmembrane protein of the adamalysin family, and integrins, e.g., alpha5beta1 and alphav, are preferentially expressed on endothelial cells on angiogenesis. Furthermore, metargidin interacts with these integrins via its disintegrin domain. In this study, recombinant human disintegrin domain (RDD) was produced in Escherichia coli by subcloning its cDNA into the pGEX-2T vector, and the effect of purified RDD on different steps of angiogenesis was evaluated. At concentrations of 2-10 micro g/ml, RDD exhibited inhibitory activities in a variety of in vitro functional assays, including endothelial cell proliferation and adhesion on the integrin substrates fibronectin, vitronectin, and fibrinogen. RDD (10 micro g/ml) totally abrogated endothelial cell migration and blocked most capillary formation in a three-dimensional fibrin gel. To test RDD efficacy in vivo, the RDD gene inserted into pBi vector containing a tetracycline-inducible promoter was electrotransferred into nude mouse muscle. RDD was successfully synthesized by muscle cells in vivo as shown by immunolabeling and Western blotting. In addition, 78% less MDA-MB-231 tumor growth, associated with strong inhibition of tumor angiogenesis, was observed in athymic mice bearing electrotransferred RDD. Moreover, in the presence of RDD, 74% fewer B16F10 melanoma lung metastases were found in C57BL/6 mice. Taken together, these results identified this RDD as a potent intrinsic inhibitor of angiogenesis, tumor growth, and metastasis, making it a promising tool for use in anticancer treatment.     

Antitumor activity of a kinesin inhibitor

Several members of the kinesin family of microtubule motor proteins play essential roles in mitotic spindle function and are potential targets for the discovery of novel antimitotic cancer therapies. KSP, also known as HsEg5, is a kinesin that plays an essential role in formation of a bipolar mitotic spindle and is required for cell cycle progression through mitosis. We identified a potent inhibitor of KSP, CK0106023, which causes mitotic arrest and growth inhibition in several human tumor cell lines. Here we show that CK0106023 is an allosteric inhibitor of KSP motor domain ATPase with a Ki of 12 nM. Among five kinesins tested, CK0106023 was specific for KSP. In tumor-bearing mice, CK0106023 exhibited antitumor activity comparable to or exceeding that of paclitaxel and caused the formation of monopolar mitotic figures identical to those produced in cultured cells. KSP was most abundant in proliferating human tissues and was absent from cultured postmitotic neurons. These findings are the first to demonstrate the feasibility of targeting mitotic kinesins for the treatment of cancer.     

P-cadherin is up-regulated by the antiestrogen ICI 182,780 and promotes invasion of human breast cancer cells

P-cadherin expression in breast carcinomas has been associated with tumors of high histologic grade and lacking estrogen receptor-alpha, suggesting a link between these proteins. In the MCF-7/AZ breast cancer cell line, blocking estrogen receptor-alpha signaling with the antiestrogen ICI 182,780 induced an increase of P-cadherin, which coincided with induction of in vitro invasion. Retroviral transduction of MCF-7/AZ cells, as well as HEK 293T cells, showed the proinvasive activity of P-cadherin, which requires the juxtamembrane domain of its cytoplasmic tail. This study establishes a direct link between P-cadherin expression and the lack of estrogen receptor-alpha signaling in breast cancer cells and suggests a role for P-cadherin in invasion, through its interaction with proteins bound to the juxtamembrane domain.     

Prognostic value of von Willebrand factor, CD34, CD31, and vascular endothelial growth factor expression in women with uterine leiomyosarcomas

BACKGROUND AND OBJECTIVES: To compare uterine leiomyosarcomas (LMS) and leiomyomas (LM) with normal myometrium in terms of microvessel density (MVD), and to correlate this parameter with vascular endothelial growth factor (VEGF) expression and clinical/pathological parameters. METHODS: An immunohistochemical technique, using antibodies against von Willebrand factor (FvW), CD34, CD31, and VEGF, was applied to formalin-fixed paraffin-embedded samples of 32 normal myometria, 32 uterine LM, and 12 LMS. MVD was calculated by a digital image analyzer. RESULTS: Using anti-FvW, mean +/- SD MVD in myometrium, LM, and LMS was 107.0 +/- 53.6, 66.2 +/- 55.4, and 64.4 +/- 44.2, respectively (P = 0.001). MVD was lower in LMS (P = 0.021) and in LM (P = 0.0004) than in normal myometrium. Using anti-CD34, mean +/- SD MVD in myometrium, LM, and LMS was 187.6 +/- 91.2, 106.1 +/- 55.5, and 114.2 +/- 98.8, respectively (P = 0.001). MVD was lower in LMS (P = 0.012) and LM (P = 0.0004) than in normal myometrium. No such differences were found using anti-CD31 and anti-VEGF. No correlation was found between MVD and VEGF expression. In women with uterine LMS, low MVD (assessed with anti-FvW) correlated with recurrence (P = 0.04) and poor overall survival (P = 0.03). CONCLUSIONS: Uterine smooth muscle tumors exhibit a lower MVD than normal myometrium, as assessed using anti-FvW or anti-CD34 antibodies. A reduced MVD, as assessed by FvW staining, has prognostic value in uterine LMS.     

A role for human MUC4 mucin gene, the ErbB2 ligand, as a target of TGF-beta in pancreatic carcinogenesis

MUC4: encodes a large transmembrane mucin that is overexpressed in pancreatic adenocarcinomas. The molecular mechanisms responsible for that altered pattern of expression are unknown. TGF-beta, a pleiotropic cytokine, regulates numerous genes involved in pancreatic carcinogenesis via activation of the Smads proteins and MUC4 promoter is rich in Smad-binding elements. Our aim was to study whether the regulation of MUC4 expression by TGF-beta in pancreatic cancer cells was strictly dependent on Smad4 activity. Three pancreatic cancer cell lines, CAPAN-1 (MUC4+/Smad4-), CAPAN-2 (MUC4+/Smad4+) and PANC-1 (MUC4-/Smad4+), were used. By RT-PCR, transfection assays and immunohistochemistry, we show that (i) both MUC4 mRNA and apomucin expression are upregulated by TGF-beta, (ii) Smad2 positively cooperates with Smad4 to activate the promoter, (iii) activation of Smad4 by exogenous TGF-beta induces Smad4 binding to the promoter, (iv) Smad7 and c-ski both inhibit activation by Smad4. When Smad4 is mutated and inactive, TGF-beta activates MUC4 expression via MAPK, PI3K and PKA signaling pathways. Absence of expression in PANC-1 cells is due to histone deacetylation. Altogether, these results indicate that upregulation of MUC4 by TGF-beta is restricted to well-differentiated pancreatic cancer cells, and point out a novel mechanism for TGF-beta as a key molecule in targeting MUC4 overexpression in pancreatic adenocarcinomas.     

Oncology studies using siRNA libraries: the dawn of RNAi-based genomics

High-throughput, human cell-based applications of RNA-mediated interference (RNAi) have emerged in recent years as perhaps the most powerful of a 'second wave' of functional genomics technologies. The available reagents and methodologies for RNAi screening studies now enable a wide range of different scopes and scales of investigation, from single-parameter assays applied to focused subsets of genes, to comprehensive genome-wide surveys based on rich, multiparameter readouts. As such, RNAi-based screens are offering important new avenues for the discovery and validation of novel therapeutic targets for several disease areas, including oncology. By enabling a 'clean' determination of gene function, that is the creation of direct causal links between gene and phenotype in human cells, RNAi investigations promise levels of pathophysiological relevance, efficiency, and range of applicability never before possible on this scale. The field of oncology, with its many assays using readily transfectable cell lines, has offered particularly fertile ground for showcasing the potential of RNAi-based genomics. However, like any other technology before it, RNAi is not without its own challenges, limitations, and caveats. Many of these issues stem directly from the choice of silencing reagent to be used in such studies, and the design of the overall screening strategy. Here, we discuss the basic design issues, potential advantages, and technical challenges of large-scale RNAi screens based on the use of chemically synthesized siRNA libraries.