Involvement of CD40 Targeting miR-224 and miR-486 on the Progression of Pancreatic Ductal Adenocarcinomas

BackgroundGenetic and epigenetic alterations during development of pancreatic ductal adenocarcinomas (PDAC) are well known. Genetic and epigenetic data were correlated with tumor biology to find specific alterations responsible for invasion and metastasis in pancreatic ductal adenocarcinomas.MethodsA total of 16 human PDAC cell lines were used in murine orthotopic PDAC models. By means of standardized dissemination scores, local invasion and metastatic spread were assessed. mRNA and microRNA expression were studied by microarray and TaqMan low-density array. Quantitative real-time–polymerase chain reaction and flow cytometry were used for expression validation.ResultsCD40 was detected as a relevant target gene for differentially expressed miRNAs observed in highly invasive and metastatic PDAC only. A significant overexpression (P < .05) of CD40-related miRNAs miR-224 and miR-486 was detected in highly invasive and metastatic PDAC, whereas CD40 mRNA expression was not significantly altered. Instead, CD40 protein expression at cell surfaces of these highly invasive and metastatic PDAC was significantly reduced (P < .01).ConclusionsEpigenetic alterations with upregulated CD40-targeting miR-224 and miR-486 are related to downregulated CD40 protein expression at cell surfaces in highly invasive and metastatic PDAC. Thus, miRNA-regulated CD40 expression seems to play an important role in progression of PDAC. These data suggest a diagnostic and therapeutic potential for CD40 and/or its targeting miRNAs in PDAC.

[1]  E. Barksdale,et al.  Neuroblastoma-induced inhibition of dendritic cell IL-12 production via abrogation of CD40 expression. , 2005, Journal of pediatric surgery.

[2]  K. Takeda,et al.  Antitumor effect of CD40 ligand: elicitation of local and systemic antitumor responses by IL-12 and B7. , 1998, Journal of immunology.

[3]  George A. Calin,et al.  Mammalian microRNAs: a small world for fine-tuning gene expression , 2006, Mammalian Genome.

[4]  L. French,et al.  Impaired CD40L signaling is a cause of defective IL-12 and TNF-alpha production in Sézary syndrome: circumvention by hexameric soluble CD40L. , 2005, Blood.

[5]  Prasenjit Dey,et al.  Genetics and biology of pancreatic ductal adenocarcinoma , 2006, Genes & development.

[6]  D. Taub,et al.  Role for CD40-CD40 ligand interactions in the immune response to solid tumours. , 2000, Molecular immunology.

[7]  Laura Pelletier,et al.  MicroRNA profiling in hepatocellular tumors is associated with clinical features and oncogene/tumor suppressor gene mutations , 2008, Hepatology.

[8]  G. Tseng,et al.  MicroRNA expression profiling of thyroid tumors: biological significance and diagnostic utility. , 2008, The Journal of clinical endocrinology and metabolism.

[9]  Thomas D. Schmittgen,et al.  Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method. , 2001, Methods.

[10]  R. Schmid,et al.  Pancreatic cancer: basic and clinical aspects. , 2005, Gastroenterology.

[11]  Stijn van Dongen,et al.  miRBase: tools for microRNA genomics , 2007, Nucleic Acids Res..

[12]  T. Golub,et al.  Distinct microRNA expression profiles in acute myeloid leukemia with common translocations , 2008, Proceedings of the National Academy of Sciences.

[13]  H. Horvitz,et al.  MicroRNA expression profiles classify human cancers , 2005, Nature.

[14]  Y. Yatabe,et al.  Reduced Expression of the let-7 MicroRNAs in Human Lung Cancers in Association with Shortened Postoperative Survival , 2004, Cancer Research.

[15]  D. Bartel,et al.  MicroRNA-Directed Cleavage of HOXB8 mRNA , 2004, Science.

[16]  G. Hannon.  RNA interference : RNA , 2002 .

[17]  F. Slack,et al.  Oncomirs — microRNAs with a role in cancer , 2006, Nature Reviews Cancer.

[18]  G. Hannon,et al.  Control of translation and mRNA degradation by miRNAs and siRNAs. , 2006, Genes & development.

[19]  Gregory Y. Lauwers,et al.  Hedgehog is an early and late mediator of pancreatic cancer tumorigenesis , 2003, Nature.

[20]  Z. Yurkovetsky,et al.  Inhibition of CD40 expression and CD40‐mediated dendritic cell function by tumor‐derived IL‐10 , 2002, International journal of cancer.

[21]  R. Flavell,et al.  CD40 and CD154 in cell-mediated immunity. , 1998, Annual review of immunology.

[22]  Y. Benjamini,et al.  More powerful procedures for multiple significance testing. , 1990, Statistics in medicine.

[23]  H. Buhr,et al.  An Orthotopic Nude Mouse Model for Evaluating Pathophysiology and Therapy of Pancreatic Cancer , 2003, Pancreas.

[24]  R. Hruban,et al.  Notch mediates TGF alpha-induced changes in epithelial differentiation during pancreatic tumorigenesis. , 2003, Cancer cell.

[25]  D. Olive,et al.  What is the real role of CD40 in cancer immunotherapy? , 1999, Immunology today.

[26]  Stefano Volinia,et al.  MicroRNA expression abnormalities in pancreatic endocrine and acinar tumors are associated with distinctive pathologic features and clinical behavior. , 2006, Journal of clinical oncology : official journal of the American Society of Clinical Oncology.

[27]  Molly Megraw,et al.  miRGen: a database for the study of animal microRNA genomic organization and function , 2006, Nucleic Acids Res..

[28]  G. Hämmerling,et al.  Vascular targeting of anti-CD40 antibodies and IL-2 into autochthonous tumors enhances immunotherapy in mice. , 2008, The Journal of clinical investigation.

[29]  C. Croce,et al.  MicroRNA expression patterns to differentiate pancreatic adenocarcinoma from normal pancreas and chronic pancreatitis. , 2007, JAMA.

[30]  Gideon Rechavi,et al.  MIR-451 and Imatinib mesylate inhibit tumor growth of Glioblastoma stem cells. , 2008, Biochemical and biophysical research communications.

[31]  W. Filipowicz,et al.  Inhibition of Translational Initiation by Let-7 MicroRNA in Human Cells , 2005, Science.

[32]  R. DePinho,et al.  Genetics and biology of pancreatic ductal adenocarcinoma , 2006, Genes & development.

[33]  George A. Calin,et al.  Expression of microRNAs and protein‐coding genes associated with perineural invasion in prostate cancer , 2008, The Prostate.

[34]  R. Xie,et al.  MicroRNA expression profile of bronchioalveolar stem cells from mouse lung. , 2008, Biochemical and biophysical research communications.

[35]  L. French,et al.  Impaired CD 40 L signaling is a cause of defective IL-12 and TNF-production in Sézary syndrome : circumvention by hexameric soluble CD 40 L , 2004 .

[36]  J. Neoptolemos,et al.  New Techniques and Agents in the Adjuvant Therapy of Pancreatic Cancer , 2002, Acta oncologica.

[37]  Kuo-Bin Li,et al.  Profiling MicroRNA Expression in Hepatocellular Carcinoma Reveals MicroRNA-224 Up-regulation and Apoptosis Inhibitor-5 as a MicroRNA-224-specific Target* , 2008, Journal of Biological Chemistry.

[38]  M. Büchler,et al.  Activating anti-CD40 antibodies induce tumour invasion by cytotoxic T-lymphocytes and inhibition of tumour growth in experimental liver cancer. , 2006, European journal of cancer.

[39]  Michael T. McManus,et al.  The microRNA miR-196 acts upstream of Hoxb8 and Shh in limb development , 2005, Nature.

[40]  Scott E Kern,et al.  Focus on pancreas cancer. , 2002, Cancer cell.

[41]  B. Ehlin‐Henriksson,et al.  A p50 surface antigen restricted to human urinary bladder carcinomas and B lymphocytes , 2004, Cancer Immunology, Immunotherapy.

[42]  C. Croce,et al.  Human microRNA genes are frequently located at fragile sites and genomic regions involved in cancers. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[43]  Wendy Frankel,et al.  MicroRNA-21 is Overexpressed in Pancreatic Cancer and a Potential Predictor of Survival , 2008, Journal of Gastrointestinal Surgery.

[44]  D. Bartel.  MicroRNAs Genomics, Biogenesis, Mechanism, and Function , 2004, Cell.

[45]  B. Reinhart,et al.  The 21-nucleotide let-7 RNA regulates developmental timing in Caenorhabditis elegans , 2000, Nature.

[46]  J. Schmidt,et al.  Transfection with CD40L induces tumour suppression by dendritic cell activation in an orthotopic mouse model of pancreatic adenocarcinoma , 2007, Gut.