Integration of genomic analysis and in vivo transfection to identify sprouty 2 as a candidate tumor suppressor in liver cancer

Hepatocellular carcinoma (HCC) is 1 of the leading causes of cancer‐related deaths worldwide, yet the molecular genetics underlying this malignancy are still poorly understood. In our study, we applied statistical methods to correlate human HCC gene expression data obtained from complementary DNA (cDNA) microarrays and corresponding DNA copy number variation data obtained from array‐based comparative genomic hybridization. We have thus identified 76 genes that are up‐regulated and show frequent DNA copy number gain, and 37 genes that are down‐regulated and show frequent DNA copy loss in human HCC samples. Among these down‐regulated genes is Sprouty2 (Spry2), a known inhibitor of receptor tyrosine kinases. We investigated the potential role of Spry2 in HCC by expressing dominant negative Spry2 (Spry2Y55F) and activated β‐catenin (ΔN90‐β‐catenin) in the mouse liver through hydrodynamic injection and sleeping beauty–mediated somatic integration. When stably expressed in mouse hepatocytes, Spry2Y55F cooperates with ΔN90‐β‐catenin to confer a neoplastic phenotype in mice. Tumor cells show high levels of expression of phospho‐extracellular signal‐regulated kinase (ERK), as well as deregulation of genes involved in cell proliferation, apoptosis, and angiogenesis. Conclusion: We identified a set of candidate oncogenes and tumor suppressor genes for human HCC. Our study provides evidence that inhibition of Spry activity cooperates with other oncogenes to promote liver cancer in mouse models, and Spry2 may function as a candidate tumor suppressor for HCC development in vivo. In addition, we demonstrate that the integration of genomic analysis and in vivo transfection is a powerful tool to identify genes that are important during hepatic carcinogenesis. (HEPATOLOGY 2008.)

[1]  M. Kay,et al.  Distinct pathways of genomic progression to benign and malignant tumors of the liver , 2007, Proceedings of the National Academy of Sciences.

[2]  R. Jaenisch,et al.  Sprouty-2 regulates oncogenic K-ras in lung development and tumorigenesis. , 2007, Genes & development.

[3]  G. Guy,et al.  Sprouty and cancer: the first terms report. , 2006, Cancer letters.

[4]  S. Thorgeirsson,et al.  Ubiquitous activation of Ras and Jak/Stat pathways in human HCC. , 2006, Gastroenterology.

[5]  S. So,et al.  Sprouty 2, an inhibitor of mitogen-activated protein kinase signaling, is down-regulated in hepatocellular carcinoma. , 2006, Cancer research.

[6]  J. Licht,et al.  Sprouty proteins: multifaceted negative-feedback regulators of receptor tyrosine kinase signaling. , 2006, Trends in cell biology.

[7]  Rui Li,et al.  Array-based comparative genomic hybridization reveals recurrent chromosomal aberrations and Jab1 as a potential target for 8q gain in hepatocellular carcinoma. , 2005, Carcinogenesis.

[8]  Corey M. Carlson,et al.  Somatic integration of an oncogene-harboring Sleeping Beauty transposon models liver tumor development in the mouse. , 2005, Proceedings of the National Academy of Sciences of the United States of America.

[9]  Philip M. Long,et al.  Molecular changes from dysplastic nodule to hepatocellular carcinoma through gene expression profiling , 2005, Hepatology.

[10]  A. Bell,et al.  Epidermal growth factor receptor: a novel target of the Wnt/beta-catenin pathway in liver. , 2005, Gastroenterology.

[11]  Rui Li,et al.  An integrated data analysis approach to characterize genes highly expressed in hepatocellular carcinoma , 2005, Oncogene.

[12]  G. Martin,et al.  Sprouty2, a mouse deafness gene, regulates cell fate decisions in the auditory sensory epithelium by antagonizing FGF signaling. , 2005, Developmental cell.

[13]  C. Robson,et al.  Epigenetic inactivation of the human sprouty2 (hSPRY2) homologue in prostate cancer , 2005, Oncogene.

[14]  C. Cohen,et al.  Survivin expression in hepatocellular carcinoma: correlation with proliferation, prognostic parameters, and outcome , 2004, Modern Pathology.

[15]  M. Rijn,et al.  Novel endothelial cell markers in hepatocellular carcinoma , 2004, Modern Pathology.

[16]  S. Thorgeirsson,et al.  Classification and prediction of survival in hepatocellular carcinoma by gene expression profiling , 2004, Hepatology.

[17]  S. Thorgeirsson,et al.  Disregulation of E-cadherin in transgenic mouse models of liver cancer , 2004, Laboratory Investigation.

[18]  T. Putti,et al.  The Ras/Mitogen-Activated Protein Kinase Pathway Inhibitor and Likely Tumor Suppressor Proteins, Sprouty 1 and Sprouty 2 Are Deregulated in Breast Cancer , 2004, Cancer Research.

[19]  D. Bar-Sagi,et al.  Modulation of signalling by Sprouty: a developing story , 2004, Nature Reviews Molecular Cell Biology.

[20]  J. Licht,et al.  Tyrosine phosphorylation of Sprouty proteins regulates their ability to inhibit growth factor signaling: a dual feedback loop. , 2004, Molecular biology of the cell.

[21]  M. Taketo,et al.  Hepatocarcinogenesis in Mice with β-Catenin and Ha-Ras Gene Mutations , 2004, Cancer Research.

[22]  M. Taketo,et al.  Hepatocarcinogenesis in mice with beta-catenin and Ha-ras gene mutations. , 2004, Cancer research.

[23]  M. Miyazaki,et al.  Angiopoietins and Tie‐2 expression in angiogenesis and proliferation of human hepatocellular carcinoma , 2003, Hepatology.

[24]  X. Wang,et al.  Predicting hepatitis B virus–positive metastatic hepatocellular carcinomas using gene expression profiling and supervised machine learning , 2003, Nature Medicine.

[25]  Danielle Hulsman,et al.  Genome-wide retroviral insertional tagging of genes involved in cancer in Cdkn2a-deficient mice , 2002, Nature Genetics.

[26]  Takeshi Suzuki,et al.  New genes involved in cancer identified by retroviral tagging , 2002, Nature Genetics.

[27]  David I. Smith,et al.  Mutational spectrum of β-catenin, AXIN1, and AXIN2 in hepatocellular carcinomas and hepatoblastomas , 2002, Oncogene.

[28]  D. Botstein,et al.  Gene expression patterns in human liver cancers. , 2002, Molecular biology of the cell.

[29]  M. Feitelson,et al.  Genetic mechanisms of hepatocarcinogenesis , 2002, Oncogene.

[30]  M. Taketo,et al.  Lack of tumorigenesis in the mouse liver after adenovirus-mediated expression of a dominant stable mutant of beta-catenin. , 2002, Cancer research.

[31]  A. Yoshimura,et al.  Identification of a Dominant Negative Mutant of Sprouty That Potentiates Fibroblast Growth Factor- but Not Epidermal Growth Factor-induced ERK Activation* , 2001, The Journal of Biological Chemistry.

[32]  D. Parkin,et al.  Global cancer statistics in the year 2000. , 2001, The Lancet. Oncology.

[33]  I. Ng,et al.  Expression of p27KIP1 and p21WAF1/CIP1 in primary hepatocellular carcinoma: Clinicopathologic correlation and survival analysis , 2001 .

[34]  I. Ng,et al.  Expression of p27(KIP1) and p21(WAF1/CIP1) in primary hepatocellular carcinoma: clinicopathologic correlation and survival analysis. , 2001, Human pathology.

[35]  Y. Jeng,et al.  Beta-catenin mutations are associated with a subset of low-stage hepatocellular carcinoma negative for hepatitis B virus and with favorable prognosis. , 2000, The American journal of pathology.

[36]  M. Kay,et al.  Somatic integration and long-term transgene expression in normal and haemophilic mice using a DNA transposon system , 2000, Nature Genetics.

[37]  C. Challen,et al.  Infrequent point mutations in codons 12 and 61 of ras oncogenes in human hepatocellular carcinomas. , 1992, Journal of hepatology.

[38]  S. Hirohashi,et al.  Low Incidence of Point Mutation of c‐Ki‐ras and N‐ras Oncogenes in Human Hepatocellular Carcinoma , 1989, Japanese journal of cancer research : Gann.