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Judah Folkman - One of the best experts on this subject based on the ideXlab platform.
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linking antibody fc domain to Endostatin significantly improves Endostatin half life and efficacy
Clinical Cancer Research, 2008Co-Authors: Robert Tjin Tham M Sjin, Judah Folkman, Shahla Movahedi, Bissan Ahmed, Elke Pravda, Kinming Lo, Stephen D Gillies, Kashi JavaherianAbstract:Purpose: The half-life of the antiangiogenic molecule Endostatin that has been used in clinical trial is short (∼2 h). In addition, ∼50% of the clinical grade Endostatin molecules lack four amino acids at their NH 2 termini. Lack of these amino acids gives rise to a molecule that is devoid of zinc, resulting in no antitumor activity. Our goal was to develop a new version of Endostatin that does not show such deficiency. Experimental Design: A recombinant human Endostatin conjugated to the Fc domain of IgG was constructed and expressed in mammalian cell culture. The presence of Fc has been shown by previous investigators to play a major role in increasing the half-life of the molecule. Fc-Endostatin was tested in tumor-bearing mice, and its half-life was compared with the clinical grade Endostatin. Results: The antitumor dose of Fc-Endostatin was found to be ∼100 times less than the clinical grade Endostatin. The half-life of Fc-Endostatin in the circulation was found to be weeks rather than hours, as observed for Endostatin alone. In addition, a U-shaped curve was observed for antitumor activity of Endostatin as a function of Endostatin concentration delivered to the animals. Conclusion: Fc-Endostatin is a superior molecule to the original clinical Endostatin. Due to its long half-life, the amount of protein required is substantially reduced compared with the clinically tested Endostatin. Furthermore, in view of the U-shaped curve of efficacy observed for Endostatin, we estimate that the requirement for Fc-Endostatin is ∼700-fold less than Endostatin alone. The half-life of Endostatin is similar to that of vascular endothelial growth factor–Trap and Avastin, two other antiangiogenic reagents. We conclude that a new clinical trial of Endostatin, incorporating Fc, may benefit cancer patients.
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continuous administration of Endostatin by intraperitoneally implanted osmotic pump improves the efficacy and potency of therapy in a mouse xenograft tumor model
Cancer Research, 2001Co-Authors: Oliver Kisker, Evelyn Flynn, Christian Becker, D Prox, Michael Fannon, Robert J Damato, William E Fogler, Elizabeth N Allred, Judah FolkmanAbstract:In the first Phase I clinical trials of Endostatin as an antiangiogenic therapy for cancer, the protein was administered as an i.v. bolus for ∼20–30 min each day. This protocol was based on experimental studies in which animals were treated by s.c. bolus once a day. However, it was not clear in the previous studies whether this schedule could be maximized further. Therefore, we developed experimental models involving continuous administration of Endostatin to determine the potency and efficacy of this approach. Endostatin was administered to tumor-bearing mice either s.c. or i.p. in single bolus doses. The efficacy of these regimens was compared with Endostatin administered continuously via an i.p. implanted mini-osmotic pump. Our results show that Endostatin remains stable and active in mini-osmotic pumps for at least 7 days. We show that Endostatin injected i.p. is rapidly cleared within 2 h, whereas Endostatin administered continuously via mini-osmotic pump maintains systemic concentrations of 200–300 ng/ml for the duration of administration. Furthermore, continuous i.p. administration of Endostatin results in more effective tumor suppression at significantly reduced doses (5-fold), compared with bolus administration. Additional experiments using a human pancreatic cancer model in severe combined immunodeficient mice showed that there was a significant decrease in the microvessel density between the treatment groups and the control group. These data show that continuous administration of human Endostatin results in sustained systemic concentrations of the protein leading to: ( a ) increased efficacy manifested as increased tumor regression; and ( b ) an 8–10-fold decrease in the dose required to achieve the same antitumor effect as the single daily bolus administration of Endostatin. On the basis of this approach, an additional clinical trial has been designed and initiated and is under way in two countries.
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continuous administration of Endostatin by intraperitoneally implanted osmotic pump improves the efficacy and potency of therapy in a mouse xenograft tumor model
Cancer Research, 2001Co-Authors: Oliver Kisker, Evelyn Flynn, D Prox, Michael Fannon, Robert J Damato, William E Fogler, Elizabeth N Allred, Christian M Becker, Bkl Sim, Judah FolkmanAbstract:In the first Phase I clinical trials of Endostatin as an antiangiogenic therapy for cancer, the protein was administered as an i.v. bolus for approximately 20-30 min each day. This protocol was based on experimental studies in which animals were treated by s.c. bolus once a day. However, it was not clear in the previous studies whether this schedule could be maximized further. Therefore, we developed experimental models involving continuous administration of Endostatin to determine the potency and efficacy of this approach. Endostatin was administered to tumor-bearing mice either s.c. or i.p. in single bolus doses. The efficacy of these regimens was compared with Endostatin administered continuously via an i.p. implanted mini-osmotic pump. Our results show that Endostatin remains stable and active in mini-osmotic pumps for at least 7 days. We show that Endostatin injected i.p. is rapidly cleared within 2 h, whereas Endostatin administered continuously via mini-osmotic pump maintains systemic concentrations of 200-300 ng/ml for the duration of administration. Furthermore, continuous i.p. administration of Endostatin results in more effective tumor suppression at significantly reduced doses (5-fold), compared with bolus administration. Additional experiments using a human pancreatic cancer model in severe combined immunodeficient mice showed that there was a significant decrease in the microvessel density between the treatment groups and the control group. These data show that continuous administration of human Endostatin results in sustained systemic concentrations of the protein leading to: (a) increased efficacy manifested as increased tumor regression; and (b) an 8-10-fold decrease in the dose required to achieve the same antitumor effect as the single daily bolus administration of Endostatin. On the basis of this approach, an additional clinical trial has been designed and initiated and is under way in two countries.
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the generation of Endostatin is mediated by elastase
Cancer Research, 1999Co-Authors: Marsha A Moses, Dmitri Wiederschain, Jack L Arbiser, Judah FolkmanAbstract:Endostatin, a potent inhibitor of angiogenesis and tumor growth, is a COOH-terminal fragment of collagen XVIII derived through cleavage of an Ala-His linkage by an as yet unidentified Endostatin-processing enzyme. Endostatin was originally isolated from the conditioned medium of hemangioendothelioma (EOMA) cells. By investigating the processing of collagen XVIII to Endostatin by EOMA cells, we show here that the generation of Endostatin can be mediated by an elastase activity. We also show that several members of the elastase family can act as an Endostatin-processing enzyme by specifically cleaving the Ala-His linkage and releasing Endostatin from a precursor molecule. We further suggest that the generation of Endostatin from collagen XVIII is at least a two-step process, involving a metal-dependent early step and an elastase activity-dependent final step.
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antiangiogenic therapy of experimental cancer does not induce acquired drug resistance
Nature, 1997Co-Authors: Thomas Boehm, Judah Folkman, Timothy Browder, Michael S OreillyAbstract:Acquired drug resistance is a major problem in the treatment of cancer. Of the more than 500,000 annual deaths from cancer in the United States1, many follow the development of resistance to chemotherapy. The emergence of resistance depends in part on the genetic instability, heterogeneity and high mutational rate of tumour cells2. In contrast, endothelial cells are genetically stable, homogenous and have a low mutational rate. Therefore, antiangiogenic therapy directed against a tumour's endothelial cells should, in principle, induce little or no drug resistance. Endostatin3, a potent angiogenesis inhibitor, was administered to mice bearing Lewis lung carcinoma, T241 fibrosarcoma or B16F10 melanoma. Treatment was stopped when tumours had regressed. Tumours were then allowed to re-grow and Endostatin therapy was resumed. After 6, 4 or 2 treatment cycles, respectively, no tumours recurred after discontinuation of therapy. These experiments show that drug resistance does not develop in three tumour types treated with a potent angiogenesis inhibitor. An unexpected finding is that repeated cycles of antiangiogenic therapy are followed by prolonged tumour dormancy without further therapy.
Rupert Timpl - One of the best experts on this subject based on the ideXlab platform.
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the minimal active domain of Endostatin is a heparin binding motif that mediates inhibition of tumor vascularization
Cancer Research, 2004Co-Authors: Annakarin Olsson, Irja Johansson, Helena Akerud, Barbro Einarsson, Rolf Christofferson, Takako Sasaki, Rupert Timpl, Lena ClaessonwelshAbstract:Endostatin constitutes the COOH-terminal 20,000 Da proteolytic fragment of collagen XVIII and has been shown to possess antiangiogenic and antitumorigenic properties. In the present study, we have investigated the role of the heparin-binding sites in the in vivo mechanism of action of Endostatin. The majority of the heparin binding is mediated by arginines 155/158/184/270 in Endostatin, but there is also a minor site constituted by arginines 193/194. Using Endostatin mutants lacking either of these two sites, we show that inhibition of fibroblast growth factor-2-induced angiogenesis in the chicken chorioallantoic membrane requires both heparin-binding sites. In contrast, inhibition of vascular endothelial growth factor-A-induced chorioallantoic membrane angiogenesis by Endostatin was only dependent on the minor heparin-binding site (R193/194). These arginines were also required for Endostatin to inhibit fibroblast growth factor-2- and vascular endothelial growth factor-A-induced chemotaxis of primary endothelial cells. Moreover, we show that a synthetic peptide corresponding to amino acids 180-199 of human Endostatin (which covers the minor heparin-binding site) inhibits endothelial cell chemotaxis and reduces tumor vascularization in vivo. Substitution of arginine residues 193/194 for alanine attenuates the antiangiogenic effects of the peptide. These data show an essential role for heparin binding in the antiangiogenic action of Endostatin.
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Endostatin reduces vascularization blood flow and growth in a rat gliosarcoma
Neuro-oncology, 2002Co-Authors: Dag R Sorensen, Takako Sasaki, Rupert Timpl, Kim Lee B Sim, Bjorn R Olsen, Tracyann Read, Torsten Porwol, Per Ole Iversen, Haakon B Benestad, Rolf BjerkvigAbstract:Endostatin, the 20-kDa C-terminal fragment of collagen XVIII, has previously been shown to inhibit growth and induce regression of different experimental tumors in rodents. In this study, we show that recombinant murine and human Endostatin, produced in 293 EBNA cells and yeast, respectively, inhibit ectotopic as well as orthotopic growing BT4Cn gliosarcomas in BD-IX rats. In rats in which s.c. gliomas were grown for a total of 29 days, systemic treatment with recombinant murine Endostatin induced about 50% reduction of intratumoral blood flow and tumor size after only 10 days of therapy. In contrast, the blood flow to irrelevant organs was unaffected by Endostatin, indicating its specificity of action. Tumors were not observed to increase in size or regrow after cessation of therapy. Furthermore, Endostatin-treated rats with i.c. tumors had significantly longer survival time than did untreated controls. In the treated rats, Endostatin therapy resulted in a reduced tumor blood vessel volume and an increased tumor cell density with an increased apoptotic index within a given tumor volume, as verified by flow cytometry and by staining with deoxynucleotidyltransferase-mediated dUTP nick-end labeling. This work verifies the general anti-angiogenic and antitumor effects of Endostatin and indicates that the protein may also be considered as a treatment strategy for malignant brain tumors.
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the angiogenesis inhibitor Endostatin impairs blood vessel maturation during wound healing
The FASEB Journal, 2000Co-Authors: Wilhelm Bloch, Takako Sasaki, Rupert Timpl, Katharina Huggel, Richard Grose, Philippe Bugnon, Klaus Addicks, Sabine WernerAbstract:Endostatin is a cleavage product of collagen XVIII that strongly inhibits tumor angiogenesis. To determine if Endostatin affects other angiogenic processes, we generated full-thickness excisional wounds on the back of mice that were systemically treated with recombinant murine Endostatin. No macroscopic abnormalities of the wound healing process were observed. Histological analysis revealed normal wound contraction and re-epithelialization, but a slight reduction in granulation tissue formation and reduced matrix deposition at the wound edge. The blood vessel density in the wounds of Endostatin-treated mice was not affected. However, ultrastructural analysis demonstrated severe abnormalities in blood vessel maturation. The wound vessels in the Endostatin-treated mice were narrowed or closed with an irregular luminal surface, resulting in a severe reduction in the number of functional vessels and extravasation of erythrocytes. Endostatin treatment did not affect the expression level and localization of collagen XVIII mRNA and protein. Furthermore, the angiogenesis regulators vascular endothelial growth factor, angiopoietin-1, and angiopoietin-2 were normally expressed in the wounds of Endostatin-treated mice. However, expression of the major wound matrix proteins fibronectin and collagens I and III was significantly reduced. This reduction is likely to explain the reduced density of the wound matrix. Our results demonstrate that Endostatin treatment reduces the number of functional blood vessels and the matrix density in the granulation tissue, but does not significantly affect the overall wound healing process.
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Endostatins derived from collagens xv and xviii differ in structural and binding properties tissue distribution and anti angiogenic activity
Journal of Molecular Biology, 2000Co-Authors: Takako Sasaki, Lena Claessonwelsh, Helena Larsson, Dominic Tisi, Erhard Hohenester, Rupert TimplAbstract:Endostatin is a fragment of the C-terminal domain NC1 of collagen XVIII that inhibits angiogenesis and tumor growth. We report the characterization of a collagen XV Endostatin analogue and its parent NC1 domain, obtained by recombinant expression in mammalian cells. Both NC1 domains contain a trimerization domain, a hinge region that is more sensitive to proteolysis in collagen XVIII and the Endostatin domain. Unlike Endostatin-XVIII, Endostatin-XV does not bind zinc or heparin, which is explained by the crystal structure of Endostatin-XV. The collagen XV and XVIII fragments inhibited chorioallantoic membrane angiogenesis induced by basic fibroblast growth factor (FGF-2) or vascular endothelial growth factor (VEGF), but there are striking differences depending on which cytokine is used and whether free Endostatins or NC1 domains are applied. The collagen XV and XVIII fragments showed a similar binding repertoire for extracellular matrix proteins. Differences were found in the immunohistological localization in vessel walls and basement membrane zones. Together, these data indentify Endostatin-XV as an angiogenesis inhibitor, which differs from Endostatin-XVIII in several important functional details.
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Endostatin induced tyrosine kinase signaling through the shb adaptor protein regulates endothelial cell apoptosis
Blood, 2000Co-Authors: Johan Dixelius, Takako Sasaki, Rupert Timpl, Helena Larsson, Kristina Holmqvist, Ake Engstrom, Michael Welsh, Lena ClaessonwelshAbstract:Endostatin, which corresponds to the C-terminal fragment of collagen XVIII, is a potent inhibitor of angiogenesis. Fibroblast growth factor-2 (FGF-2)-induced angiogenesis in the chicken chorioallantoic membrane was inhibited by Endostatin, but not by an Endostatin mutant R158/270A, lacking heparin-binding ability. Endostatin was internalized by endothelial cells, but not by mouse fibroblasts. Treatment of murine brain endothelial (IBE) cells with Endostatin reduced the proportion of cells in S phase, whereas growth-arrested IBE cells in collagen gels treated with Endostatin displayed enhanced tubular morphogenesis. IBE cells overexpressing Shb, an adaptor protein implicated in angiostatin-induced apoptosis, displayed elevated apoptosis and decreased tubular morphogenesis in collagen gels in response to Endostatin when added together with FGF-2. Induction of apoptosis was dependent on the heparin-binding ability of Endostatin and the expression of Shb with a functional Src homology 2 (SH2)-domain. Endostatin treatment for 10 minutes or 24 hours induced tyrosine phosphorylation of Shb and formation of multiprotein complexes. An Shb SH2 domain fusion protein precipitated a 125-kd phosphotyrosyl protein in Endostatin-treated cells. The 125-kd component either contained intrinsic tyrosine kinase activity or occurred in complex with a tyrosine kinase. In conclusion, our data show that Endostatin induces tyrosine kinase activity and enhanced apoptosis in FGF-treated endothelial cells.
Takako Sasaki - One of the best experts on this subject based on the ideXlab platform.
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the minimal active domain of Endostatin is a heparin binding motif that mediates inhibition of tumor vascularization
Cancer Research, 2004Co-Authors: Annakarin Olsson, Irja Johansson, Helena Akerud, Barbro Einarsson, Rolf Christofferson, Takako Sasaki, Rupert Timpl, Lena ClaessonwelshAbstract:Endostatin constitutes the COOH-terminal 20,000 Da proteolytic fragment of collagen XVIII and has been shown to possess antiangiogenic and antitumorigenic properties. In the present study, we have investigated the role of the heparin-binding sites in the in vivo mechanism of action of Endostatin. The majority of the heparin binding is mediated by arginines 155/158/184/270 in Endostatin, but there is also a minor site constituted by arginines 193/194. Using Endostatin mutants lacking either of these two sites, we show that inhibition of fibroblast growth factor-2-induced angiogenesis in the chicken chorioallantoic membrane requires both heparin-binding sites. In contrast, inhibition of vascular endothelial growth factor-A-induced chorioallantoic membrane angiogenesis by Endostatin was only dependent on the minor heparin-binding site (R193/194). These arginines were also required for Endostatin to inhibit fibroblast growth factor-2- and vascular endothelial growth factor-A-induced chemotaxis of primary endothelial cells. Moreover, we show that a synthetic peptide corresponding to amino acids 180-199 of human Endostatin (which covers the minor heparin-binding site) inhibits endothelial cell chemotaxis and reduces tumor vascularization in vivo. Substitution of arginine residues 193/194 for alanine attenuates the antiangiogenic effects of the peptide. These data show an essential role for heparin binding in the antiangiogenic action of Endostatin.
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Endostatin reduces vascularization blood flow and growth in a rat gliosarcoma
Neuro-oncology, 2002Co-Authors: Dag R Sorensen, Takako Sasaki, Rupert Timpl, Kim Lee B Sim, Bjorn R Olsen, Tracyann Read, Torsten Porwol, Per Ole Iversen, Haakon B Benestad, Rolf BjerkvigAbstract:Endostatin, the 20-kDa C-terminal fragment of collagen XVIII, has previously been shown to inhibit growth and induce regression of different experimental tumors in rodents. In this study, we show that recombinant murine and human Endostatin, produced in 293 EBNA cells and yeast, respectively, inhibit ectotopic as well as orthotopic growing BT4Cn gliosarcomas in BD-IX rats. In rats in which s.c. gliomas were grown for a total of 29 days, systemic treatment with recombinant murine Endostatin induced about 50% reduction of intratumoral blood flow and tumor size after only 10 days of therapy. In contrast, the blood flow to irrelevant organs was unaffected by Endostatin, indicating its specificity of action. Tumors were not observed to increase in size or regrow after cessation of therapy. Furthermore, Endostatin-treated rats with i.c. tumors had significantly longer survival time than did untreated controls. In the treated rats, Endostatin therapy resulted in a reduced tumor blood vessel volume and an increased tumor cell density with an increased apoptotic index within a given tumor volume, as verified by flow cytometry and by staining with deoxynucleotidyltransferase-mediated dUTP nick-end labeling. This work verifies the general anti-angiogenic and antitumor effects of Endostatin and indicates that the protein may also be considered as a treatment strategy for malignant brain tumors.
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the angiogenesis inhibitor Endostatin impairs blood vessel maturation during wound healing
The FASEB Journal, 2000Co-Authors: Wilhelm Bloch, Takako Sasaki, Rupert Timpl, Katharina Huggel, Richard Grose, Philippe Bugnon, Klaus Addicks, Sabine WernerAbstract:Endostatin is a cleavage product of collagen XVIII that strongly inhibits tumor angiogenesis. To determine if Endostatin affects other angiogenic processes, we generated full-thickness excisional wounds on the back of mice that were systemically treated with recombinant murine Endostatin. No macroscopic abnormalities of the wound healing process were observed. Histological analysis revealed normal wound contraction and re-epithelialization, but a slight reduction in granulation tissue formation and reduced matrix deposition at the wound edge. The blood vessel density in the wounds of Endostatin-treated mice was not affected. However, ultrastructural analysis demonstrated severe abnormalities in blood vessel maturation. The wound vessels in the Endostatin-treated mice were narrowed or closed with an irregular luminal surface, resulting in a severe reduction in the number of functional vessels and extravasation of erythrocytes. Endostatin treatment did not affect the expression level and localization of collagen XVIII mRNA and protein. Furthermore, the angiogenesis regulators vascular endothelial growth factor, angiopoietin-1, and angiopoietin-2 were normally expressed in the wounds of Endostatin-treated mice. However, expression of the major wound matrix proteins fibronectin and collagens I and III was significantly reduced. This reduction is likely to explain the reduced density of the wound matrix. Our results demonstrate that Endostatin treatment reduces the number of functional blood vessels and the matrix density in the granulation tissue, but does not significantly affect the overall wound healing process.
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Endostatins derived from collagens xv and xviii differ in structural and binding properties tissue distribution and anti angiogenic activity
Journal of Molecular Biology, 2000Co-Authors: Takako Sasaki, Lena Claessonwelsh, Helena Larsson, Dominic Tisi, Erhard Hohenester, Rupert TimplAbstract:Endostatin is a fragment of the C-terminal domain NC1 of collagen XVIII that inhibits angiogenesis and tumor growth. We report the characterization of a collagen XV Endostatin analogue and its parent NC1 domain, obtained by recombinant expression in mammalian cells. Both NC1 domains contain a trimerization domain, a hinge region that is more sensitive to proteolysis in collagen XVIII and the Endostatin domain. Unlike Endostatin-XVIII, Endostatin-XV does not bind zinc or heparin, which is explained by the crystal structure of Endostatin-XV. The collagen XV and XVIII fragments inhibited chorioallantoic membrane angiogenesis induced by basic fibroblast growth factor (FGF-2) or vascular endothelial growth factor (VEGF), but there are striking differences depending on which cytokine is used and whether free Endostatins or NC1 domains are applied. The collagen XV and XVIII fragments showed a similar binding repertoire for extracellular matrix proteins. Differences were found in the immunohistological localization in vessel walls and basement membrane zones. Together, these data indentify Endostatin-XV as an angiogenesis inhibitor, which differs from Endostatin-XVIII in several important functional details.
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Endostatin induced tyrosine kinase signaling through the shb adaptor protein regulates endothelial cell apoptosis
Blood, 2000Co-Authors: Johan Dixelius, Takako Sasaki, Rupert Timpl, Helena Larsson, Kristina Holmqvist, Ake Engstrom, Michael Welsh, Lena ClaessonwelshAbstract:Endostatin, which corresponds to the C-terminal fragment of collagen XVIII, is a potent inhibitor of angiogenesis. Fibroblast growth factor-2 (FGF-2)-induced angiogenesis in the chicken chorioallantoic membrane was inhibited by Endostatin, but not by an Endostatin mutant R158/270A, lacking heparin-binding ability. Endostatin was internalized by endothelial cells, but not by mouse fibroblasts. Treatment of murine brain endothelial (IBE) cells with Endostatin reduced the proportion of cells in S phase, whereas growth-arrested IBE cells in collagen gels treated with Endostatin displayed enhanced tubular morphogenesis. IBE cells overexpressing Shb, an adaptor protein implicated in angiostatin-induced apoptosis, displayed elevated apoptosis and decreased tubular morphogenesis in collagen gels in response to Endostatin when added together with FGF-2. Induction of apoptosis was dependent on the heparin-binding ability of Endostatin and the expression of Shb with a functional Src homology 2 (SH2)-domain. Endostatin treatment for 10 minutes or 24 hours induced tyrosine phosphorylation of Shb and formation of multiprotein complexes. An Shb SH2 domain fusion protein precipitated a 125-kd phosphotyrosyl protein in Endostatin-treated cells. The 125-kd component either contained intrinsic tyrosine kinase activity or occurred in complex with a tyrosine kinase. In conclusion, our data show that Endostatin induces tyrosine kinase activity and enhanced apoptosis in FGF-treated endothelial cells.
Fengshan Wang - One of the best experts on this subject based on the ideXlab platform.
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Tat PTD-Endostatin-RGD: A novel protein with anti-angiogenesis effect in retina via eye drops.
Biochimica et Biophysica Acta, 2016Co-Authors: Yan Li, Juzheng Sheng, Xinke Zhang, Danyang Feng, Zhiwei Li, Aijun Wang, Lian Li, Xu Zhang, Fengshan WangAbstract:Abstract Background Diabetic retinopathy is a leading cause of blindness. The objective was to design a novel fusion protein, Tat PTD-Endostatin-RGD, to treat retinal neovascularization via eye drops instead of traditional intravitreal injection trepapeutical methods. Method The anti-angiogenesis ability was evaluated in vitro by chick embryo chorioallantoic membrane assay, wound healing assay and tube formation assay. Corneal barrier and blood-retina barrier were constructed in vitro to investigate the penetration ability of Tat PTD-Endostatin-RGD. Western blot was used to detect the integrin αvβ3 expression level in rat retina microvascular endothelial cells which was stimulated by S-nitroso-N-acetylpenicillamine. The binding affinity of Tat PTD-Endostatin-RGD to integrin αvβ3 was investigated by evaluating the penetration ability on blood-retina barriers treated with S-nitroso-N-acetylpenicillamine. The pharmacodynamics and efficacy analysis were further carried out in the oxygen-induced retinopathy model in vivo. In addition, the pharmacokinetic profile via eye drops was studied on a C57BL/6 mice model. Result Tat PTD-Endostatin-RGD showed high anti-angiogenesis activity and high ability to penetrate these two barriers in vitro. The Western blot results indicated S-nitroso-N-acetylpenicillamine upregulated the expression level of integrin αvβ3 in a dose-dependent manner. Tat PTD-Endostatin-RGD showed a high affinity to rat retina microvascular endothelial cells treated with S-nitroso-N-acetylpenicillamine. The results showed that Tat PTD-Endostatin-RGD could inhibit abnormal angiogenesis in retina via eye drops. Conclusion Tat PTD-Endostatin-RGD showed high penetration ability through ocular barriers, bound specifically to integrin αvβ3 and effectively inhibited the abnormal angiogenesis. General significance Tat PTD-Endostatin-RGD represents a potent novel drug applied via eye drops for fundus oculi neovascularization diseases.
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tat ptd Endostatin a novel anti angiogenesis protein with ocular barrier permeability via eye drops
Biochimica et Biophysica Acta, 2015Co-Authors: Xinke Zhang, Yanna Cheng, Haining Tan, Fengshan WangAbstract:Abstract Background Endostatin, a specific inhibitor of endothelial cell proliferation and angiogenesis, has been proved to have effects on ocular neovascular diseases by intraocular injection. In order to increase its permeability to ocular barriers and make it effective on fundus oculi angiogenesis diseases via non-invasive administration (eye drops), Endostatin was fused to Tat PTD via a genetic engineering method. Methods Most of the Tat PTD– Endostatin was expressed as inclusion bodies in Escherichia coli, so pure and active Tat PTD–Endostatin was prepared by a series of operations, including inclusion body denaturation, refolding and chromatography. The anti-angiogenesis activity of Tat PTD–Endostatin was investigated by cell proliferation experiments and chick embryo chorioallantoic membrane assay. In addition, its translocating ability and concrete entry mechanism into cells were also investigated by fluorescence microscope and flow cytometry. The penetrating ability to ocular barriers was also studied by immunohistochemistry. A mouse choroidal neovascularization model was established to investigate the pharmacodynamics of Tat PTD–Endostatin. Results The obtained Tat PTD–Endostatin had excellent anti-angiogenesis activity and was superior to Es in cellular translocating. Macropinocytosis may be the dominant route of entry of Tat PTD–Endostatin into cells. Tat PTD–Endostatin could cross ocular barriers and arrive at the retina after eye-drop administration. In addition, it displayed inhibitory effects on choroidal neovascularization via eye drops. Conclusions Tat PTD–Endostatin possessed excellent ocular penetrating ability and anti-angiogenesis effects. General significance Tat PTD is a promising ocular delivery tool, and Tat PTD–Endostatin is a potential drug for curing fundus oculi angiogenesis diseases.
Zhu Jiayong - One of the best experts on this subject based on the ideXlab platform.
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antitumor activities of liver targeting peptide modified recombinant human Endostatin in balb c nu mice with hepatocellular carcinoma
Scientific Reports, 2017Co-Authors: Bao Dongmei, Zhang Jingjing, Jin Xiaobao, Zhu JiayongAbstract:In our previous study, a liver-targeting peptide CSP I-plus modified recombinant human Endostatin (rEndostatin, endostar) (rES-CSP) was constructed and showed potent antiangiogenic capability and could specifically bind to human hepatocellular carcinoma cells to make a direct inhibition in vitro. In this study, the biological activities of rES-CSP in vivo were evaluated by subcutaneous and orthotopic xenograft nude mice model of human hepatocellular carcinoma cells HepG2. We found that rES-CSP significantly decreased tumor volume to 54.9% in the nude mice with subcutaneous xenograft compared with the control. In orthotopic xenograft model, rES-CSP not only decreased tumor volume (to 39.6% compared with the control) and tumor weight, it also increased its biodistribution in the liver tissue and hepatoma tissue. Moreover, lower microvessel density (MVD) and higher apoptotic index (AI) were also observed in the tumor tissues. It had no significant side-effects on the heart, liver, spleen, lung and kidney of mice. Results indicated CSP I-plus modified Endostar may be a potential candidate for a targeting therapy on hepatocellular carcinoma.
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Antitumor activities of Liver-targeting peptide modified Recombinant human Endostatin in BALB/c-nu mice with Hepatocellular carcinoma
Nature Publishing Group, 2017Co-Authors: Ma Yan, Bao Dongmei, Zhang Jingjing, Jin Xiaobao, Wang Jie, Wang Yan, Zhu JiayongAbstract:Abstract In our previous study, a liver-targeting peptide CSP I-plus modified recombinant human Endostatin (rEndostatin, endostar) (rES-CSP) was constructed and showed potent antiangiogenic capability and could specifically bind to human hepatocellular carcinoma cells to make a direct inhibition in vitro. In this study, the biological activities of rES-CSP in vivo were evaluated by subcutaneous and orthotopic xenograft nude mice model of human hepatocellular carcinoma cells HepG2. We found that rES-CSP significantly decreased tumor volume to 54.9% in the nude mice with subcutaneous xenograft compared with the control. In orthotopic xenograft model, rES-CSP not only decreased tumor volume (to 39.6% compared with the control) and tumor weight, it also increased its biodistribution in the liver tissue and hepatoma tissue. Moreover, lower microvessel density (MVD) and higher apoptotic index (AI) were also observed in the tumor tissues. It had no significant side-effects on the heart, liver, spleen, lung and kidney of mice. Results indicated CSP I-plus modified Endostar may be a potential candidate for a targeting therapy on hepatocellular carcinoma