title: Inhibition of oesophageal squamous cell carcinoma progression by in vivo targeting of hyaluronan synthesis.
abstract: BACKGROUND
  Oesophageal cancer is a highly aggressive tumour entity with at present poor prognosis. Therefore, novel treatment options are urgently needed. Hyaluronan (HA) is a polysaccharide present in the matrix of human oesophageal squamous cell carcinoma (ESCC). Importantly, in vitro ESCC cells critically depend on HA synthesis to maintain the proliferative phenotype. The aim of the present study is (1) to study HA-synthase (HAS) expression and regulation in human ESCC, and (2) to translate the in vitro results into a mouse xenograft model of human ESCC to study the effects of systemic versus tumour targeted HAS inhibition on proliferation and distribution of tumour-bound and stromal hyaluronan.
  
  
  METHODS
  mRNA expression was investigated in human ESCC biopsies by semiquantitative real-time RT PCR. Furthermore, human ESCC were xenografted into NMRI nu/nu mice. The effects on tumour progression and morphology of 4-methylumbelliferone (4-MU), an inhibitor of HA-synthesis, and of lentiviral knock down of HA-synthase 3 (HAS3), the main HAS isoform in the human ESCC tissues and the human ESCC cell line used in this study, were determined. Tumour progression was monitored by calliper measurements and by flat-panel detector volume computed tomography (fpVCT). HA content, cellular composition and proliferation (Ki67) were determined histologically.
  
  
  RESULTS
  mRNA of HAS isoform 3 (HAS3) was upregulated in human ESCC biopsies and HAS3 mRNA was positively correlated to expression of the epidermal growth factor (EGF) receptor. EGF was also proven to be a strong inductor of HAS3 mRNA expression in vitro. During the course of seven weeks, 4-MU inhibited progression of xenograft tumours. Interestingly, remodelling of the tumour into a more differentiated phenotype and inhibition of cell proliferation were observed. Lentiviral knockdown of HAS3 in human ESCC cells prior to xenografting mimicked all effects of 4-MU treatment suggesting that hyaluronan produced by ESCC is accountable for major changes in tumour environment in vivo.
  
  
  CONCLUSIONS
  Systemic inhibition of HA-synthesis and knockdown of tumour cell HAS3 cause decreased ESCC progression accompanied by tumour stroma remodelling and may therefore be used in novel approaches to ESCC therapy.
authors.affiliation: Institut für Pharmakologie und Klinische Pharmakologie, Universitätsklinikum Düsseldorf, Heinrich-Heine Universität Düsseldorf, Düsseldorf, Germany.
authors.forename: Sören
authors.initials: S
authors.lastname: Twarock
authors.forename: Till
authors.initials: T
authors.lastname: Freudenberger
authors.forename: Eva
authors.initials: E
authors.lastname: Poscher
authors.forename: Guang
authors.initials: G
authors.lastname: Dai
authors.forename: Katharina
authors.initials: K
authors.lastname: Jannasch
authors.forename: Christian
authors.initials: C
authors.lastname: Dullin
authors.forename: Frauke
authors.initials: F
authors.lastname: Alves
authors.forename: Klaus
authors.initials: K
authors.lastname: Prenzel
authors.forename: Wolfram T
authors.initials: WT
authors.lastname: Knoefel
authors.forename: Nikolas H
authors.initials: NH
authors.lastname: Stoecklein
authors.forename: Rashmin C
authors.initials: RC
authors.lastname: Savani
authors.forename: Bernhard
authors.initials: B
authors.lastname: Homey
authors.forename: Jens W
authors.initials: JW
authors.lastname: Fischer
mesh_terms: D000818:Animals; D002294:Carcinoma, Squamous Cell / Q000378:metabolism* / Q000473:pathology / Q000517:prevention & control; D002460:Cell Line; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D018450:Disease Progression; D066246:ErbB Receptors / Q000235:genetics / Q000378:metabolism; D004938:Esophageal Neoplasms / Q000378:metabolism* / Q000473:pathology / Q000517:prevention & control; D015972:Gene Expression Regulation, Neoplastic; D014453:Glucuronosyltransferase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D000076002:Hyaluronan Synthases; D006820:Hyaluronic Acid / Q000096:biosynthesis*; D006923:Hymecromone / Q000031:analogs & derivatives / Q000494:pharmacology; D007150:Immunohistochemistry; D051379:Mice; D008815:Mice, Inbred Strains; D008819:Mice, Nude; D034622:RNA Interference; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D023041:Xenograft Model Antitumor Assays
chemical_list: D012333:RNA, Messenger; D006923:Hymecromone; D006820:Hyaluronic Acid; D014453:Glucuronosyltransferase; C105458:HAS3 protein, human; D000076002:Hyaluronan Synthases; D066246:ErbB Receptors