title: ERK2-regulated TIMP1 induces hyperproliferation of K-Ras(G12D)-transformed pancreatic ductal cells.
abstract: Pancreatic ductal adenocarcinoma (PDAC) commonly contains a mutation in K-Ras(G12D) and is characterized by a desmoplastic reaction composed of deregulated, proliferating cells embedded in an abnormal extracellular matrix (ECM). Our previous observations imply that inhibiting the mitogen-activated protein kinase (MAPK)-extracellular signal-regulated kinase (ERK2) kinase signal pathway reverses a matrix metalloproteinase 1-specific invasive phenotype. Here, we investigated the specific genes downstream of MAPK-ERK2 responsible for the hyperproliferative abilities of human and murine primary ductal epithelial cells (PDCs) within an ECM. Compared with control, DNA synthesis and total cell proliferation was significantly increased in human PDCs harboring the PDAC common p53, Rb/p16(INK4a), and K-Ras (G12D) mutations. Both of these effects were readily reversed following small-molecule inhibition or lentiviral silencing of ERK2. Microarray analysis of PDCs in three-dimensional (3D) culture revealed a unique, MAPK-influenced gene signature downstream of K-Ras (G12D). Unbiased hierarchical analysis permitted filtration of tissue inhibitor of matrix metalloproteinase 1 (TIMP1). Pancreatic cells isolated from Pdx1-Cre; LSL-K-ras(G12D/+)-mutated mice exhibit increased TIMP1 RNA transcription compared to wild-type littermate controls. Analyses of both 3D, in vitro human K-Ras (G12D) PDCs and data mining of publicly annotated human pancreatic data sets correlatively indicate increased levels of TIMP1 RNA. While silencing TIMP1 did not significantly effect PDC proliferation, exogenous addition of human recombinant TIMP1 significantly increased proliferation but only in transformed K-Ras (G12D) PDCs in 3D. Overall, TIMP1 is an upregulated gene product and a proliferative inducer of K-Ras(G12D)-mutated PDCs through the ERK2 signaling pathway.
authors.affiliation: Department of Biochemistry and Molecular Biology, Molecular and Cellular Biology and Genetics Program, Drexel University College of Medicine, Philadelphia, PA, USA.
authors.forename: Gregory P
authors.initials: GP
authors.lastname: Botta
authors.forename: Maximilian
authors.initials: M
authors.lastname: Reichert
authors.forename: Mauricio J
authors.initials: MJ
authors.lastname: Reginato
authors.forename: Steffen
authors.initials: S
authors.lastname: Heeg
authors.forename: Anil K
authors.initials: AK
authors.lastname: Rustgi
authors.forename: Peter I
authors.initials: PI
authors.lastname: Lelkes
mesh_terms: D000818:Animals; D002070:Butadienes / Q000494:pharmacology; D021441:Carcinoma, Pancreatic Ductal; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D004847:Epithelial Cells / Q000378:metabolism; D015972:Gene Expression Regulation, Neoplastic; D055785:Gene Knockdown Techniques; D006801:Humans; D020935:MAP Kinase Signaling System; D051379:Mice; D008822:Mice, Transgenic; D019950:Mitogen-Activated Protein Kinase 1 / Q000037:antagonists & inhibitors / Q000502:physiology*; D020125:Mutation, Missense; D009570:Nitriles / Q000494:pharmacology; D020411:Oligonucleotide Array Sequence Analysis; D010183:Pancreatic Ducts / Q000473:pathology; D010190:Pancreatic Neoplasms; D011518:Proto-Oncogene Proteins / Q000235:genetics*; D016283:Proto-Oncogene Proteins p21(ras); D034741:RNA, Small Interfering / Q000235:genetics; D019715:Tissue Inhibitor of Metalloproteinase-1 / Q000235:genetics* / Q000378:metabolism; D059467:Transcriptome; D018631:ras Proteins / Q000235:genetics*
chemical_list: D002070:Butadienes; C117307:KRAS protein, human; D009570:Nitriles; D011518:Proto-Oncogene Proteins; D034741:RNA, Small Interfering; C560822:TIMP1 protein, human; D019715:Tissue Inhibitor of Metalloproteinase-1; C113580:U 0126; C535150:MAPK1 protein, human; D019950:Mitogen-Activated Protein Kinase 1; D016283:Proto-Oncogene Proteins p21(ras); D018631:ras Proteins
references.citation: Oncogene. 2002 Mar 28;21(14):2245-52
references.pmid: 11948407
references.citation: J Biol Chem. 2003 Oct 10;278(41):40364-72
references.pmid: 12904305
references.citation: Nature. 2010 Mar 18;464(7287):427-30
references.pmid: 20179705
references.citation: Biochem Biophys Res Commun. 2008 Aug 8;372(4):845-9
references.pmid: 18533112
references.citation: FEBS Lett. 1992 Feb 17;298(1):29-32
references.pmid: 1544418
references.citation: Cancer Res. 2007 Mar 1;67(5):2098-106
references.pmid: 17332339
references.citation: Br J Cancer. 1996 Apr;73(8):972-8
references.pmid: 8611434
references.citation: J Pathol. 2002 Jul;197(3):307-13
references.pmid: 12115876
references.citation: J Surg Res. 2002 Jan;102(1):39-44
references.pmid: 11792150
references.citation: Genes Dev. 2007 Nov 1;21(21):2788-803
references.pmid: 17974918
references.citation: Gastroenterology. 2004 Jul;127(1):250-60
references.pmid: 15236190
references.citation: J Cell Biochem. 2007 Jul 1;101(4):887-907
references.pmid: 17266048
references.citation: Dev Biol. 1999 Jul 15;211(2):238-54
references.pmid: 10395785
references.citation: J Proteome Res. 2011 May 6;10(5):2359-76
references.pmid: 21443201
references.citation: Breast Cancer Res Treat. 2009 Sep;117(1):31-44
references.pmid: 18787947
references.citation: J Mammary Gland Biol Neoplasia. 2006 Oct;11(3-4):283-97
references.pmid: 17089203
references.citation: Oncogene. 2000 Jun 8;19(25):2930-42
references.pmid: 10871844
references.citation: Br J Cancer. 2004 Jan 26;90(2):463-70
references.pmid: 14735194
references.citation: Cancer Cell. 2011 May 17;19(5):664-78
references.pmid: 21575865
references.citation: FASEB J. 2008 Feb;22(2):466-76
references.pmid: 17928366
references.citation: Mol Cell. 2010 Apr 9;38(1):114-27
references.pmid: 20385094
references.citation: FEBS J. 2011 Jan;278(1):28-45
references.pmid: 21087458
references.citation: Biochim Biophys Acta. 1992 Nov 15;1171(1):41-55
references.pmid: 1420363
references.citation: J Clin Invest. 2011 Dec;121(12):4572-8
references.pmid: 22133881
references.citation: Semin Cancer Biol. 2010 Jun;20(3):161-8
references.pmid: 20470890
references.citation: Br J Cancer. 2006 Oct 23;95(8):1114-20
references.pmid: 17047657
references.citation: Genes Dev. 2013 Feb 1;27(3):288-300
references.pmid: 23355395
references.citation: J Surg Res. 2002 Jan;102(1):35-8
references.pmid: 11792149
references.citation: J Pathol. 1997 Jul;182(3):347-55
references.pmid: 9349239
references.citation: Int J Mass Spectrom. 2011 Apr;302(1-3):101-109
references.pmid: 21785572
references.citation: Bioorg Med Chem Lett. 2006 Dec 15;16(24):6281-7
references.pmid: 17000106
references.citation: Development. 2002 May;129(10):2447-57
references.pmid: 11973276
references.citation: Mol Cancer Res. 2012 Feb;10(2):183-96
references.pmid: 22160930
references.citation: Blood. 2011 Jun 16;117(24):6479-88
references.pmid: 21521782
references.citation: PLoS Med. 2008 Jun 10;5(6):e123
references.pmid: 18547137
references.citation: PLoS One. 2007 Nov 07;2(11):e1155
references.pmid: 17982507
references.citation: EMBO J. 2006 Sep 6;25(17):3934-42
references.pmid: 16917503
references.citation: Cancer Res. 2003 Dec 15;63(24):8614-22
references.pmid: 14695172
references.citation: Genes Dev. 2006 May 15;20(10):1218-49
references.pmid: 16702400
references.citation: Cancer Gene Ther. 2001 Nov;8(11):869-78
references.pmid: 11773977
references.citation: Mol Cell Biol. 2008 Jan;28(1):511-27
references.pmid: 17967895
references.citation: Nat Rev Cancer. 2002 Mar;2(3):161-74
references.pmid: 11990853
references.citation: Mol Cancer Ther. 2007 Apr;6(4):1186-97
references.pmid: 17406031
references.citation: Cell. 2012 Jan 20;148(1-2):349-61
references.pmid: 22265420
references.citation: Biochim Biophys Acta. 2005 Nov 25;1756(2):97-101
references.pmid: 16169155
references.citation: Hepatology. 2005 Apr;41(4):857-67
references.pmid: 15726641
references.citation: Gastroenterology. 2008 Oct;135(4):1288-300
references.pmid: 18725219
references.citation: Genes Dev. 2003 Dec 15;17(24):3112-26
references.pmid: 14681207
references.citation: J Virol. 2004 Jun;78(11):5934-45
references.pmid: 15140991
references.citation: J Biol Chem. 2003 Mar 21;278(12):10304-13
references.pmid: 12525489
references.citation: Pancreas. 2009 Aug;38(6):613-8
references.pmid: 19629003
references.citation: J Biol. 2006;5(5):14
references.pmid: 16805921
references.citation: Genes Dev. 2001 Dec 15;15(24):3243-8
references.pmid: 11751630
references.citation: Biochem Biophys Res Commun. 1999 Apr 13;257(2):494-9
references.pmid: 10198240
references.citation: Nat Protoc. 2006;1(4):2116-27
references.pmid: 17487203
references.citation: J Biol Chem. 2000 Oct 20;275(42):32664-71
references.pmid: 11032844
references.citation: Clin Cancer Res. 2011 Feb 15;17(4):805-16
references.pmid: 21325298
references.citation: EMBO J. 2010 Mar 17;29(6):1091-104
references.pmid: 20150892