ABSTRACT
Ovarian cancer is the leading cause of death from gynecological malignancy and the fourth leading cause of cancer death among American women, yet little is known about its molecular aetiology. Studies using comparative genomic hybridization (CGH) have revealed several regions of recurrent, abnormal, DNA sequence copy number that may encode genes involved in the genesis or progression of the disease. One region at 3q26 found to be increased in copy number in approximately 40% of ovarian and others cancers contains PIK3CA, which encodes the p110alpha catalytic subunit of phosphatidylinositol 3-kinase (PI3-kinase). The association between PIK3CA copy number and PI3-kinase activity makes PIK3CA a candidate oncogene because a broad range of cancer-related functions have been associated with PI3-kinase mediated signalling. These include proliferation, glucose transport and catabolism, cell adhesion, apoptosis, RAS signalling and oncogenic transformation. In addition, downstream effectors of PI3-kinase, AKT1 and AKT2, have been found to be amplified or activated in human tumours, including ovarian cancer. We show here that PIK3CA is frequently increased in copy number in ovarian cancers, that the increased copy number is associated with increased PIK3CA transcription, p110alpha protein expression and PI3-kinase activity and that treatment with the PI3-kinase inhibitor LY294002 decreases proliferation and increases apoptosis. Our observations suggest PIK3CA is an oncogene that has an important role in ovarian cancer.
Subject(s)
Chromosomes, Human, Pair 3 , Oncogenes , Ovarian Neoplasms/genetics , Phosphatidylinositol 3-Kinases/genetics , Chromones/pharmacology , Enzyme Inhibitors/pharmacology , Female , Humans , In Situ Hybridization, Fluorescence , Morpholines/pharmacology , Phosphatidylinositol 3-Kinases/biosynthesis , Phosphoinositide-3 Kinase Inhibitors , Tumor Cells, CulturedABSTRACT
Tryptases are serine proteases implicated in asthma and are very highly expressed in human mast cells. They fall into two groups, alpha and beta. Although several related tryptase mRNAs are known, it is unclear which if any are transcripts of separate haploid genes. The studies described here investigated the nature and number of human tryptases and sought possibly novel members of the family. To this end, two human bacterial artificial chromosome (BAC) clones containing tryptase genes were identified and mapped to chromosome 16p13.3, of which approximately 2.2 megabases are syntenic with the part of mouse chromosome 17 containing tryptase genes mouse mast cell protease (mMCP)-6 and -7. Sequencing and restriction mapping suggest that the BACs may partially overlap. Sequenced BAC genes correspond to three known beta-tryptases (betaI, betaII, and betaIII), an alpha-like gene, and a pair of novel hybrid genes related partly to alpha/beta-tryptases and partly to orthologs of mMCP-7. betaII and betaIII, betaI and alphaII, as well as the two mMCP-7-like genes, may be alleles at single loci; in total, there are at least three nonallelic tryptase genes in the isolated BAC clones. DNA blotting and restriction analysis suggest that the BACs include most members of the immediate tryptase family. Thus, chromosome 16p13.3 harbors a cluster of known and previously undescribed members of the tryptase gene family.
Subject(s)
Chromosomes, Human, Pair 16 , Mast Cells/enzymology , Serine Endopeptidases/genetics , Amino Acid Sequence , Animals , Base Sequence , Chromosome Mapping , Chromosomes, Artificial, Yeast , Chymases , Deoxyribonucleases, Type II Site-Specific/metabolism , Humans , In Situ Hybridization, Fluorescence , Mice , Molecular Sequence Data , Recombinant Proteins/genetics , Sequence Homology, Amino Acid , Serine Endopeptidases/metabolism , TryptasesABSTRACT
Loss of E-cadherin (CDH1) function is thought to contribute to progression in breast cancer and other solid tumors by increasing proliferation, invasion, and/or metastasis. In some cases, the restoration of CDH1 function may be an important therapeutic option. This possibility will depend on the mechanism by which CDH1 is inactivated. Here we present analyses of CDH1 expression, genetic mutation, and promoter methylation in CDH1 in 10 commonly used breast cancer cell lines. Five cell lines (BT-474, MCF-7, MDA-MB-361, MDA-MB-468, and T-47D) expressed CDH1 and were genetically normal. Five others (SK-BR-3, 600 MPE, MDA-MB-134 IV, CAMA1, and MDA-MB-435) did not express CDH1. Fluorescence in situ hybridization analyses of each of these cell lines showed evidence for the physical deletion of one allele of CDH1, and three cell lines were found to carry homozygous deletions. SK-BR-3 was deleted from exon 12 through the promoter; exon 6 was deleted in MDA-MB-134 IV cells, and 600 MPE cells carried a 21-bp deletion in the splicing acceptor site for exon 9. CAMA1 seemed to have been inactivated through promoter methylation. No explanation was found for the inactivation of CDH1 in MDA-MB-435.
