A novel peptide (GX1) homing to gastric cancer vasculature inhibits angiogenesis and cooperates with TNF alpha in anti-tumor therapy.
BACKGROUND
The discovery of the importance of angiogenesis in tumor growth has emphasized the need to find specific vascular targets for tumor-targeted therapies. Previously, using phage display technology, we identified the peptide GX1 as having the ability to target the gastric cancer vasculature. The present study investigated the bioactivities of GX1, as well as its potential ability to cooperate with recombinant mutant human tumor necrosis factor alpha (rmhTNFalpha), in gastric cancer therapy.


RESULTS
Tetrazolium salt (MTT) assay showed that GX1 could inhibit cell proliferation of both human umbilical vein endothelial cells (HUVEC) (44%) and HUVEC with tumor endothelium characteristics, generated by culturing in tumor-conditioned medium (co-HUVEC) (62%). Flow-cytometry (FCM) and western blot assays showed that GX1 increased the rate of apoptosis from 11% to 31% (p < 0.01) by up-regulating caspase 3 expression level. A chorioallantoic membrane assay indicated that GX1 could suppress neovascularization in vivo, with the microvessel count decreasing from 21 to 11 (p < 0.05). When GX1 was fused to rmhTNFalpha, GX1-rmhTNFalpha selectively concentrated in the gastric cancer vasculature, as shown by enzyme-linked immunosorbent assay, immunofluorescence and emission-computed tomography. In vitro MTT and FCM assays showed that, compared to rmhTNFalpha alone, GX1-rmhTNFalpha was more effective at suppressing co-HUVEC proliferation (45% vs. 61%, p < 0.05) and inducing apoptosis (11% vs. 23%, p < 0.05). In a tumor formation test, GX1-rmhTNFalpha more effectively inhibited tumor growth than rmhTNFalpha (tumor volume: 271 mm3 vs. 134 mm3, p < 0.05), with less systemic toxicity as measured by body weight (20.57 g vs. 19.30 g, p < 0.05). These therapeutic effects may be mediated by selectively enhanced tumor vascular permeability, as indicated by Evan's blue assay.


CONCLUSIONS
GX1 had both homing activity and the ability to inhibit vascular endothelial cell proliferation in vitro and neovascularization in vivo. Furthermore, when GX1 was conjugated to rmhTNFalpha, the fusion protein was selectively delivered to targeted tumor sites, significantly improving the anti-tumor activity of rmhTNFalpha and decreasing systemic toxicity. These results demonstrate the potential of GX1 as a homing peptide in vascular targeted therapy for gastric cancer.
Chen
Bei
B
State Key Laboratory of Cancer Biology and Xijing Hospital of Digestive Diseases, the Fourth Military Medical University, Xi'an, Shaanxi, PR China. bzcheese@hotmail.com
Cao
Shanshan
S
Zhang
Yingqi
Y
Wang
Xin
X
Liu
Jie
J
Hui
Xiaoli
X
Wan
Yi
Y
Du
Wenqi
W
Wang
Li
L
Wu
Kaichun
K
Fan
Daiming
D
D020533:Angiogenesis Inhibitors / Q000627:therapeutic use*; D000818:Animals; D000970:Antineoplastic Agents / Q000627:therapeutic use*; D017209:Apoptosis / Q000187:drug effects; D002478:Cells, Cultured; D018450:Disease Progression; D042783:Endothelial Cells / Q000166:cytology / Q000187:drug effects; D006801:Humans; D051379:Mice; D008819:Mice, Nude; D009842:Oligopeptides / Q000627:therapeutic use*; D010456:Peptides, Cyclic; D013274:Stomach Neoplasms / Q000098:blood supply* / Q000188:drug therapy* / Q000473:pathology; D014409:Tumor Necrosis Factor-alpha / Q000737:chemistry / Q000378:metabolism / Q000627:therapeutic use*
D020533:Angiogenesis Inhibitors; D000970:Antineoplastic Agents; D009842:Oligopeptides; D010456:Peptides, Cyclic; D014409:Tumor Necrosis Factor-alpha; C534327:cyclo(cysteinyl-glycyl-aspargyl-seryl-aspargyl-prolyl-lysyl-seryl-cysteine)