Characterization of antiangiogenic and antitumor activities of kringle fragments derived from human apolipoprotein(a)

Abstract

[ Part I ] Angiogenesis, the formation of new capillaries from pre-existing vessels, is essential for tumor growth and metastasis. Apolipoprotein(a) [apo(a)] contains tandemly repeated kringle domains that are closely related to plasminogen kringle 4, followed by a single kringle 5-like domain and inactive protease-like domains. Recently, the anti-angiogenic activities of apo(a) have been demonstrated both in vitro and in vivo. However, its effects on tumor angiogenesis and the underlying mechanisms involved have not been fully elucidated. To evaluate the anti-angiogenic and anti-tumor activities of the apo(a) kringle domains and to elucidate their mechanism of action, the last three kringle domains of apo(a), KIV-9, KIV-10, and KV, were expressed in Escherichia coll. The resultant recombinant protein, termed rhLK68, exhibited a dose-dependent inhibition of bFGF-stimulated human umbilical vein endothelial cell proliferation and migration in vitro, and inhibited the neovascularization in chick chorioallantoic membranes in vivo. To explore the molecular mechanism of rhLK68-mediated inhibition of angiogenesis, the effects - of rhLK68 on extracellular signal regulated kinase 1 and 2 (ERK1/2) and phosphatidylinositol 3-kinase/Akt signaling pathways were determined. Treatment with rhLK68 inhibited ERK1/2 phosphorylation but did not influence Akt activation. Interestingly, an inhibitor of protein tyrosine phosphatase, sodium orthovanadate, dose-dependently reversed both rhLK68-induced dephosphorylation of ERK1/2 and decreased migration of endothelial cells, whereas rhLK68 showed no significant effects on mitogen-activated protein kinase kinases (MEKs) phosphorylation. These results indicate that inhibition of endothelial cell migration and angiogenesis by rhLK68 may be achieved by interfering with ERK1/2 activation via a protein tyrosine phosphatase-dependent pathway. Furthermore, systemic administration of rhLK68 suppressed human lung (A549) and colon (HCT-15) tumor ...