Mutation of human plasminogen kringle 1-5 enhances anti-angiogenic action via increased interaction with integrin αvβ3

Po Chiao Chang, Yu Jia Chang, Hua Lin Wu, Chin Wei Chang, Chung I. Lin, Wei Chih Wang, Guey Yueh Shi

Research output: Contribution to journalArticle

6 Citations (Scopus)

Abstract

Angiogenesis plays a primary role in tumor growth and metastasis. Angiostatin, a proteolytic fragment containing the first four kringle domains of human plasminogen, can inhibit angiogenesis. The anti-angiogenic activities of kringle 1-5 (K1-5) and kringle 5 fragments of plasminogen are greater than angiostatin in inhibiting angiogenesis and angiogenesis-dependent tumor growth. To further optimize kringle fragment anti-angiogenic activities, mutations were created at the potential glycosylation sites Asn-289 and Thr-346 and the Lys binding site, Leu-532, at kringle 5, including K1-5N289A (replacing Asn by Ala at residue 289), K1-5T346A, K 1-5L532R, K1-5N289A/T346A, K1-5T346A/L532R, K1-5N289A/L532R, and K1-5N289A/T346A/L532R. Wild-type and mutant K1-5 proteins were expressed successfully by the Pichia pastoris expression system. Native K1-5 from proteolytic cleavage and wild-type K1-5 have similar activity in inhibiting basic fibroblast growth factor-induced endothelial cell proliferation. Among these mutated proteins, K1-5N289A/T346A/L532R exhibited the greatest effect in inhibiting endothelial cell proliferation and in inducing endothelial cell apoptosis. Integrin αvβ3-mediated adhesion of K1-5N289A/T346A/L532R to endothelial cells was more greatly enhanced when compared to wild type K1-5. Furthermore, K1-5N289A/ T346A/L532R was most potent in inhibiting basic fibroblast growth factor-induced angiogenesis in Matrigel assay in vivo. Angiogenesis-dependent tumor growth was inhibited by systemically injected K1-5N289A/T346A/L532R into mice. These results demonstrate that alteration of glycosylation and Lys binding properties could increase the anti-angiogenic action of K1-5, possibly via enhanced interaction with integrin αvβ 3 in endothelial cells.

Original languageEnglish
Pages (from-to)729-738
Number of pages10
JournalThrombosis and Haemostasis
Volume99
Issue number4
DOIs
Publication statusPublished - Apr 2008
Externally publishedYes

Keywords

  • Angiogenesis and inhibitors
  • Endothelial cells
  • Integrins
  • Plasminogen (-fragments)

ASJC Scopus subject areas

  • Hematology

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