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A novel peptide (GX1) homing to gastric cancer vasculature inhibits angiogenesis and cooperates with TNF alpha in anti-tumor therapy.

Chen B, Cao S, Zhang Y, Wang X, Liu J, Hui X, Wan Y, Du W, Wang L, Wu K, Fan D - BMC Cell Biol. (2009)

Bottom Line: 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).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.

View Article: PubMed Central - HTML - PubMed

Affiliation: 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

ABSTRACT

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.

Conclusion: 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.

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GX1 inhibits angiogenesis in CAM assay. (A-C) GX1 at 20 μg (50 μM) hampered neovascularization of fertilized eggs. Attenuated and tortuous microvessels are shown in the CAM, with fewer angiogenic vessels contacting the filter disks. (D) Number of microvessels contacting the disks. Bars, SD. * p < 0.05.
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Figure 2: GX1 inhibits angiogenesis in CAM assay. (A-C) GX1 at 20 μg (50 μM) hampered neovascularization of fertilized eggs. Attenuated and tortuous microvessels are shown in the CAM, with fewer angiogenic vessels contacting the filter disks. (D) Number of microvessels contacting the disks. Bars, SD. * p < 0.05.

Mentions: Since GX1 could repress vascular endothelial cell proliferation in vitro, we carried out CAM assays to see if the peptide could inhibit angiogenesis in vivo. Disruption of angiogenesis was observed in GX1-treated chicken embryos, with attenuated and tortuous microvessels in the CAM and fewer angiogenic vessels contacting the disk, when compared to the PBS control group. No significant differences existed between the Pep 2 and PBS control groups, with both showing well-developed and leaf vein-like vascular nets (Figure 2).


A novel peptide (GX1) homing to gastric cancer vasculature inhibits angiogenesis and cooperates with TNF alpha in anti-tumor therapy.

Chen B, Cao S, Zhang Y, Wang X, Liu J, Hui X, Wan Y, Du W, Wang L, Wu K, Fan D - BMC Cell Biol. (2009)

GX1 inhibits angiogenesis in CAM assay. (A-C) GX1 at 20 μg (50 μM) hampered neovascularization of fertilized eggs. Attenuated and tortuous microvessels are shown in the CAM, with fewer angiogenic vessels contacting the filter disks. (D) Number of microvessels contacting the disks. Bars, SD. * p < 0.05.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2746182&req=5

Figure 2: GX1 inhibits angiogenesis in CAM assay. (A-C) GX1 at 20 μg (50 μM) hampered neovascularization of fertilized eggs. Attenuated and tortuous microvessels are shown in the CAM, with fewer angiogenic vessels contacting the filter disks. (D) Number of microvessels contacting the disks. Bars, SD. * p < 0.05.
Mentions: Since GX1 could repress vascular endothelial cell proliferation in vitro, we carried out CAM assays to see if the peptide could inhibit angiogenesis in vivo. Disruption of angiogenesis was observed in GX1-treated chicken embryos, with attenuated and tortuous microvessels in the CAM and fewer angiogenic vessels contacting the disk, when compared to the PBS control group. No significant differences existed between the Pep 2 and PBS control groups, with both showing well-developed and leaf vein-like vascular nets (Figure 2).

Bottom Line: 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).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.

View Article: PubMed Central - HTML - PubMed

Affiliation: 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

ABSTRACT

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.

Conclusion: 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.

Show MeSH
Related in: MedlinePlus