Potent anti-tumor and prolonged survival effects of E. coli-derived non-glycosylated kringle domain of tissue-type plasminogen activator

Abstract

Abstract. The two-kringle domain of tissue-type plasminogen activator (TK1-2) has been identified as a novel angiogenesis inhibitor. In the previous study, purified Pichia-derived TK1-2 has been shown to suppress in vivo growth of human lung and colon cancer cells. Here, we demonstrate that E. coliderived non-glycosylated TK1-2 suppresses tumor growth more potently than Pichia-derived TK1-2 and prolongs the survival of tumor bearing mice. The recombinant TK1-2 prepared through E. coli expression, His-tag affinity chromatography and in vitro refolding was injected intraperitoneally once daily into nude mice 7 days after subcutaneous implantation with PC14 lung cancer cells (n=10). Measurement of tumor volumes indicated that low-dose TK1-2 treatment (10 mg/kg) suppressed tumor growth by approximately 85.2% (p<0.01), while high-dose TK1-2 treatment (50 mg/kg) even more potently inhibited tumor growth (>93.8%) (p<0.005). Treatment of TK1-2 also prolonged the survival of tumor-bearing mice in a dose-dependent fashion. In an independent HCT116 xenograft model, E. coli-derived TK1-2 was more effective in suppressing tumor growth than Pichia-derived TK1-2. Immunohistochemical analysis of tumor tissue also revealed that the expression of VEGF, SMA-·, TNF-· and angiogenin was less positive in the E. coli-derived TK1-2-treated group than in the Pichia-derived TK1-2-treated group. These results suggest that E. coli-derived refolded, non-glycosylated TK1-2 can be used more effectively as an anti-cancer agent.

Inhibition of Pathological Retinal Neovascularization by a Small Peptide Derived from Human Tissue-Type Plasminogen Kringle 2

“…In previous studies, Kim et al reported the recombinant protein TK1-2 and the t-PA kringle 2 domain alone inhibited angiogenesis in vitro and in vivo (Kim et al,2003;Carroll et al, 2005;Kang et al, 2006;Kim et al, 2008). Kim also reported a peptide derived from t-PA kringle 2 (named TP-7) that inhibited angiogenesis and corneal neovascularization (Kim et al, 2017).…”

A Novel Peptide Derived From Tissue‐Type Plasminogen Activator Potently Inhibits Angiogenesis and Corneal Neovascularization

“…In previous studies, we reported that the recombinant protein TK1‐2 inhibits angiogenesis and tumor growth in vitro and in vivo and also stated that its anti‐angiogenic activity is mediated in part by the interaction with integrin α2β1 [Kim et al, , ; Kang et al, ]. Notably, the t‐PA kringle 2 domain alone can elicit the anti‐angiogenic activity [Carroll et al, ].…”

“…Previously, we reported that the recombinant protein TK1‐2 comprising two kringle domains (kringle 1 and 2) of tissue‐type plasminogen activator (t‐PA) has anti‐angiogenic activities in vitro and in vivo and that it suppresses the growth of several types of tumor in vivo [Kim et al, ; Shim et al, ; Kang et al, ; Oh et al, ]. We also found that TK1‐2 potently inhibits endothelial cell migration in part by interfering with integrin α2β1 function [Kim et al, ].…”

The Kringle Domain of Tissue-Type Plasminogen Activator Inhibits Extracellular Matrix-Induced Adhesion and Migration of Endothelial Cells

“…TK1-2 has been identified as a novel angiogenesis inhibitor through in vitro and in vivo studies, [3][4][5] but its mechanism of action has not yet been studied. In this study, we found that TK1-2 inhibits ECM-induced cell adhesion and the formation of focal adhesions and actin stress fibers.…”

“…3) In addition, it exhibits potent inhibition of in vivo angiogenesis in chick chorioallantoic membrane, and tumor growth in lung and colon cancer xenografted murine models. [3][4][5] Recently, we reported that TK1-2 inhibits the differentiation and migration of endothelial progenitor cells (EPCs), and its contribution to tumor angiogenesis, 6) but its mechanism of action remains unclear.…”