title: High STAT1 mRNA levels but not its tyrosine phosphorylation are associated with macrophage infiltration and bad prognosis in breast cancer.
abstract: BACKGROUND
  STAT1 has been attributed a function as tumor suppressor. However, in breast cancer data from microarray analysis indicated a predictive value of high mRNA expression levels of STAT1 and STAT1 target genes belonging to the interferon-related signature for a poor response to therapy. To clarify this issue we have determined STAT1 expression levels and activation by different methods, and investigated their association with tumor infiltration by immune cells. Additionally, we evaluated the interrelationship of these parameters and their significance for predicting disease outcome.
  
  
  METHODS
  Expression of STAT1, its target genes SOCS1, IRF1, CXCL9, CXCL10, CXCL11, IFIT1, IFITM1, MX1 and genes characteristic for immune cell infiltration (CD68, CD163, PD-L1, PD-L2, PD-1, CD45, IFN-γ, FOXP3) was determined by RT-PCR in two independent cohorts comprising 132 breast cancer patients. For a subset of patients, protein levels of total as well as serine and tyrosine-phosphorylated STAT1 were ascertained by immunohistochemistry or immunoblotting and protein levels of CXCL10 by ELISA.
  
  
  RESULTS
  mRNA expression levels of STAT1 and STAT1 target genes, as well as protein levels of total and serine-phosphorylated STAT1 correlated with each other in neoplastic tissue. However, there was no association between tumor levels of STAT1 mRNA and tyrosine-phosphorylated STAT1 and between CXCL10 serum levels and CXCL10 expression in the tumor. Tumors with increased STAT1 mRNA amounts exhibited elevated expression of genes characteristic for tumor-associated macrophages and immunosuppressive T lymphocytes. Survival analysis revealed an association of high STAT1 mRNA levels and bad prognosis in both cohorts. A similar prognostically relevant correlation with unfavorable outcome was evident for CXCL10, MX1, CD68, CD163, IFN-γ, and PD-L2 expression in at least one collective. By contrast, activation of STAT1 as assessed by the level of STAT1-Y701 phosphorylation was linked to positive outcome. In multivariate Cox regression, the predictive power of STAT1 mRNA expression was lost when including expression of CXCL10, MX1 and CD68 as confounders.
  
  
  CONCLUSIONS
  Our study confirms distinct prognostic relevance of STAT1 expression levels and STAT1 tyrosine phosphorylation in breast cancer patients and identifies an association of high STAT1 levels with elevated expression of STAT1 target genes and markers for infiltrating immune cells.
authors.forename: Piotr
authors.initials: P
authors.lastname: Tymoszuk
authors.forename: Pornpimol
authors.initials: P
authors.lastname: Charoentong
authors.forename: Hubert
authors.initials: H
authors.lastname: Hackl
authors.forename: Rita
authors.initials: R
authors.lastname: Spilka
authors.forename: Elisabeth
authors.initials: E
authors.lastname: Müller-Holzner
authors.forename: Zlatko
authors.initials: Z
authors.lastname: Trajanoski
authors.forename: Peter
authors.initials: P
authors.lastname: Obrist
authors.forename: Françoise
authors.initials: F
authors.lastname: Revillion
authors.forename: Jean-Philippe
authors.initials: JP
authors.lastname: Peyrat
authors.forename: Heidi
authors.initials: H
authors.lastname: Fiegl
authors.affiliation: Division of Medical Biochemistry, Biocenter, Innsbruck Medical University, Innrain 80-82, 6020 Innsbruck, Austria. wolfgang.doppler@i-med.ac.at.
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Doppler
mesh_terms: D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D001943:Breast Neoplasms / Q000235:genetics* / Q000473:pathology; D005260:Female; D015972:Gene Expression Regulation, Neoplastic / Q000235:genetics; D006801:Humans; D008875:Middle Aged; D010766:Phosphorylation / Q000235:genetics; D011379:Prognosis; D012333:RNA, Messenger / Q000096:biosynthesis* / Q000235:genetics; D050794:STAT1 Transcription Factor / Q000096:biosynthesis* / Q000235:genetics; D015398:Signal Transduction / Q000235:genetics; D014443:Tyrosine / Q000235:genetics
chemical_list: D012333:RNA, Messenger; D050794:STAT1 Transcription Factor; C494079:STAT1 protein, human; D014443:Tyrosine
references.citation: Biochimie. 2010 May;92(5):425-44
references.pmid: 20159032
references.citation: PLoS One. 2009 Jun 08;4(6):e5821
references.pmid: 19503789
references.citation: Clin Dev Immunol. 2012;2012:656340
references.pmid: 22611421
references.citation: Oncogene. 2000 May 15;19(21):2628-37
references.pmid: 10851062
references.citation: Proc Natl Acad Sci U S A. 2009 Jun 9;106(23):9373-8
references.pmid: 19478064
references.citation: Bioinformatics. 2002 Jan;18(1):207-8
references.pmid: 11836235
references.citation: J Interferon Cytokine Res. 2011 Jan;31(1):33-40
references.pmid: 21166594
references.citation: Immunity. 2013 Feb 21;38(2):250-62
references.pmid: 23352233
references.citation: Breast Cancer Res. 2007;9(4):212
references.pmid: 17705880
references.citation: FASEB J. 2009 Nov;23(11):3990-9
references.pmid: 19641143
references.citation: J Clin Invest. 2011 Jan;121(1):308-17
references.pmid: 21183794
references.citation: Nat Rev Immunol. 2011 Sep 09;11(10):685-92
references.pmid: 21904387
references.citation: J Gastroenterol. 2007 Dec;42(12):947-56
references.pmid: 18085351
references.citation: BMC Cancer. 2007 Jul 25;7:136
references.pmid: 17651480
references.citation: J Clin Oncol. 2010 Jan 1;28(1):105-13
references.pmid: 19917869
references.citation: Autoimmun Rev. 2009 Mar;8(5):379-83
references.pmid: 19105984
references.citation: Breast Cancer Res. 2012 Jan 20;14(1):R16
references.pmid: 22264274
references.citation: Proc Natl Acad Sci U S A. 2008 Nov 25;105(47):18490-5
references.pmid: 19001271
references.citation: Nat Rev Immunol. 2006 Nov;6(11):836-48
references.pmid: 17063185
references.citation: Cytokine. 2010 Apr;50(1):1-14
references.pmid: 20036577
references.citation: Cell Rep. 2013 Aug 15;4(3):437-44
references.pmid: 23933255
references.citation: Clin Exp Immunol. 2003 Sep;133(3):448-53
references.pmid: 12930373
references.citation: Endocrinology. 2009 Jun;150(6):2924-33
references.pmid: 19246539
references.citation: Cell. 2007 Jun 29;129(7):1275-86
references.pmid: 17570480
references.citation: EMBO J. 1996 Nov 15;15(22):6262-8
references.pmid: 8947049
references.citation: Proc Natl Acad Sci U S A. 2009 Jun 9;106(23):9368-72
references.pmid: 19474293
references.citation: Cancer Res. 2006 Oct 1;66(19):9509-18
references.pmid: 17018607
references.citation: Nat Rev Mol Cell Biol. 2002 Sep;3(9):651-62
references.pmid: 12209125
references.citation: Cancer Discov. 2011 Jun;1(1):54-67
references.pmid: 22039576
references.citation: Clin Cancer Res. 2002 Oct;8(10):3065-74
references.pmid: 12374673
references.citation: Am J Obstet Gynecol. 2004 Nov;191(5):1598-605
references.pmid: 15547530
references.citation: Oncotarget. 2011 Dec;2(12):1043-54
references.pmid: 22185785
references.citation: Breast Cancer Res. 2011;13(6):R126
references.pmid: 22151962
references.citation: J Immunol. 2009 Nov 15;183(10):6767-77
references.pmid: 19846878
references.citation: Cell Signal. 2007 Mar;19(3):454-65
references.pmid: 17085014
references.citation: J Clin Oncol. 2011 May 20;29(15):1949-55
references.pmid: 21483002
references.citation: Neoplasia. 2010 Nov;12(11):899-905
references.pmid: 21076615
references.citation: Clin Breast Cancer. 2013 Aug;13(4):264-70
references.pmid: 23829892
references.citation: JAKSTAT. 2013 Apr 1;2(2):e23353
references.pmid: 24058806
references.citation: Trends Immunol. 2002 Feb;23(2):96-101
references.pmid: 11929133
references.citation: Nat Rev Cancer. 2012 Mar 22;12(4):252-64
references.pmid: 22437870
references.citation: Cell Cycle. 2011 Mar 1;10(5):794-804
references.pmid: 21311224
references.citation: BMC Cancer. 2012 Jul 23;12:306
references.pmid: 22824040
references.citation: Breast Cancer Res Treat. 2009 Mar;114(1):155-67
references.pmid: 18401703
references.citation: Cancer Res. 2012 Sep 15;72(18):4724-32
references.pmid: 22805310
references.citation: Proc Natl Acad Sci U S A. 2012 Feb 21;109(8):2796-801
references.pmid: 21825174
references.citation: Eur J Immunol. 2013 Oct;43(10):2718-29
references.pmid: 23843024
references.citation: Int J Radiat Biol. 2009 May;85(5):421-31
references.pmid: 19437244
references.citation: J Immunol. 2008 Feb 15;180(4):2158-64
references.pmid: 18250422
references.citation: J Biol Chem. 2011 Oct 7;286(40):34743-51
references.pmid: 21859714
references.citation: J Biol Chem. 2013 Apr 26;288(17):11676-88
references.pmid: 23486482
references.citation: Curr Opin Immunol. 2012 Apr;24(2):207-12
references.pmid: 22236695
references.citation: Nat Rev Immunol. 2007 Jun;7(6):454-65
references.pmid: 17525754
references.citation: Clin Cancer Res. 2012 Jun 1;18(11):3015-21
references.pmid: 22615451
references.citation: J Leukoc Biol. 2014 Jun;95(6):853-66
references.pmid: 24295829
references.citation: J Viral Hepat. 2001 Sep;8(5):341-8
references.pmid: 11555191