Clinical & Experimental Metastasis

, Volume 22, Issue 5, pp 377–390

Perlecan Knockdown in Metastatic Prostate Cancer Cells Reduces Heparin-binding Growth Factor Responses in vitro and Tumor Growth in vivo

  • Cristiana Savorè
  • Chu Zhang
  • Caroline Muir
  • Riting Liu
  • Jeffrey Wyrwa
  • Jun Shu
  • Haiyen E. Zhau
  • Leland W.K. Chung
  • Daniel D. Carson
  • Mary C. Farach-Carson
Article

Abstract

Perlecan (Pln) is a major heparan sulfate proteoglycan (HSPG) of extracellular matrices and bone marrow stroma. Pln, via glycosaminoglycans in domains I and V, acts as a co-receptor for delivery of heparin binding growth factors (HBGFs) that support cancer growth and vascularization. Specifically, glycosaminoglycans bind HBGFs and activate HBGF receptors, including those for FGF-2 and VEGF-A. The contribution of Pln to prostate cancer growth was tested using a ribozyme approach to knockdown Pln expression levels. Transfection into the androgen-independent, bone targeted prostate cancer line, C4-2B, and efficient stable knockdown of Pln was demonstrated by quantitative PCR, immunohistochemistry and immunoblotting. Three individually isolated subclones with 75–80% knockdown in Pln mRNA, protein expression and secretion into ECM were used to study in vitro growth responses to FGF-2 and VEGF-A. While cells with normal Pln levels responded to both HBGFs, knockdown cells responded poorly. All lines responded to serum growth factors and IGF-I. Anchorage-independent growth assays showed reduced colony size and cohesiveness by all Pln deficient subclones compared to parental C4-2B cells. In vivo effects of Pln knockdown were measured by inoculating knockdown and control ribozyme transfected cell lines into athymic mice. A reduced growth rate, smaller tumor size, diminished vascularization and failure to elevate serum PSA characterized mice bearing Pln knockdown C4-2B cells. Poor vascularization correlated with reduced levels of VEGF-A secreted by Pln knockdown lines. We conclude that Pln is an essential ECM component involved in growth responses of metastatic prostate cancer cells to HBGFs deposited in local and metastatic microenvironment.

Keywords

angiogenesis extracellular matrix heparin binding growth factors perlecan prostate cancer ribozyme 

Abbreviations

: ECM

extracellular matrix

FBS

fetal bovine serum

FGF-2

basic fibroblast growth factor

FGF-BP

fibroblast growth factor binding protein

HBGF(R)

heparin binding growth factor (receptor)

HSPG

heparan sulfate proteoglycan

PBS

phosphate buffered saline

Pln

perlecan

SCID

severe combined immunodeficient

VEGF-A

vascular endothelial growth factor

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Copyright information

© Springer 2005

Authors and Affiliations

  • Cristiana Savorè
    • 1
  • Chu Zhang
    • 1
  • Caroline Muir
    • 1
  • Riting Liu
    • 1
  • Jeffrey Wyrwa
    • 1
  • Jun Shu
    • 2
  • Haiyen E. Zhau
    • 2
  • Leland W.K. Chung
    • 2
  • Daniel D. Carson
    • 1
  • Mary C. Farach-Carson
    • 1
  1. 1.Department of Biological SciencesUniversity of DelawareNewarkUSA
  2. 2.Molecular Urology and Therapeutics Program, Department of Urology and Winship Cancer Institute, School of MedicineEmory UniversityAtlantaUSA

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