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系統識別號 U0026-1803201517285200
論文名稱(中文) 針對癌症治療發展對於整合蛋白αvβ3和/或α5β1具有選擇性和親和力的去整合蛋白
論文名稱(英文) Development of Selective and Potent Integrin αvβ3- and/or α5β1-specific Disintegrins for Cancer Therapy
校院名稱 成功大學
系所名稱(中) 基礎醫學研究所
系所名稱(英) Institute of Basic Medical Sciences
學年度 103
學期 2
出版年 104
研究生(中文) 黃俊豪
研究生(英文) Chun-Hao Huang
學號 S58971497
學位類別 博士
語文別 中文
論文頁數 116頁
口試委員 指導教授-莊偉哲
口試委員-謝奇璋
口試委員-鄭宏祺
口試委員-王淑鶯
口試委員-符文美
口試委員-陳金榜
中文關鍵字 蛇毒蛋白  馬來腹蛇  整合蛋白  癌症治療 
英文關鍵字 Rhodostomin  integrin  disintegrin  cancer therapy 
學科別分類
中文摘要 整合蛋白(integrin)是一群由α和β兩個次單元所組成的異質雙體,其參與許多細胞過程。整合蛋白的配體在與其相互結合時,RGD、NGR和LDV這些胺基酸序列是主要的辨認位置。在含有RGD序列的配體中,其RGD序列周遭區域、協同區域和C端區域可以調節對整合蛋白結合的親和力與專一性。在第一型纖維連結蛋白(fibronectin)之第五(Fn-I5)和第七(Fn-I7)模組中,NGR序列可與整合蛋白α5β1結合,且其自發性去胺基作用會產生isoDGR序列,進而成為一個可以與整合蛋白αvβ3結合的序列,並且在調控纖維連結蛋白的纖維組裝,及調控內皮細胞的黏著和增生都扮演重要的角色。在我的研究中使用含有RGD序列的去整合蛋白,馬來腹蛇的蛇毒蛋白(Rhodostomin,Rho)為一個骨架去探討在去整合蛋白中可能與整合蛋白作用的區域。利用細胞黏著實驗、核磁共振、X光結晶學和分子嵌合模擬去研究整合蛋白與去整合蛋白之間結構和功能的關係。迄今,我們已經成功利用Rho設計出對整合蛋白具有親和力和專一性的抑制劑,包含了對整合蛋白αvβ3具有專一性的48ARLDDL突變株,和對整合蛋白αvβx和α5β1具有專一性的KG突變株。在我的研究中發現RGD的N端區域(殘基39-43)對於整合蛋白αvβ3、αIIbβ3和α5β1的結合扮演重要的角色。細胞黏著實驗顯示馬來腹蛇蛇毒蛋白突變株含有39KKKRT序列有最高的抑制效果,對整合蛋白αvβ3、αIIbβ3和α5β1分別有3.6、4.6和6.2倍的抑制效果提升。相對來說,Rho突變株含有46DD對於整合蛋白αvβ3、αIIbβ3和α5β1分別有7.2、124.5和>582.0倍的抑制效果下降。而RGD的C端區域突變株將54DD置換成54YY對於整合蛋白αvβ3和α5β1的抑制有2.7和3.8倍的提升。野生型Rho、KKKRT和R46E突變株的三維結構已經以X光結晶學解出。而結構的分析顯示,在KKKRT序列中的R42與C端區域的Y67和H68間有交互作用,且在野生型Rho的SRAGK序列沒有發現這樣的交互作用,而KKKRT突變株和整合蛋白嵌合模型中,也顯示R42是與整合蛋白β1和β3次單元中的D216交互作用,而這些作用也沒有在野生型Rho與整合蛋白的嵌合模型中發現。Rho野生型R46側鏈與R46E突變株的E46側鏈方向相反,且在嵌合模型中發現R46可與整合蛋白αIIb次單位的D159形成鹽橋,然而在R46E突變株中E46卻不能形成。這些結果顯示在去整合蛋白中RGD序列附近區域會去影響他們對於整合蛋白之間結合與交互作用。將GNGRG序列放入Rho中去研究其對於整合蛋白的結合,而質譜與核磁共振結果顯示,NGR會轉換成DGR和isoDGR異構物。在細胞黏著的實驗顯示兩個異構物對於整合蛋白αvβ3的IC50約為250 μM,而對於整合蛋白α5β1有較高的抑制能力,isoDGR異構物的IC50為4.5 μM,而isoDGR異構物相較於DGR異構物的抑制能力有6倍增加,且含有isoDGR序列的蛋白對於整合蛋白α5β1是一個較好的配體。流式細胞儀和西方墨點法結果顯示整合蛋白β3降低的A375細胞已經建立,而其黏著實驗顯示細胞無法黏著在纖維蛋白原(fibrinogen)上。細胞爬行實驗顯示Rho、RLD和ARLDDL可以抑制A375細胞的爬行其IC50分別為9.3,3.7和45.1 nM,相對來說Rho、RLD和ARLDDL對整合蛋白β3降低的A375細胞抑制能力為0.5,3.0和>100 μM。這些結果顯示對整合蛋白αvβ3專一性的突變株抑制人類的黑色素細胞瘤是取決於整合蛋白β3的路徑。我們發現Rho和其突變株可以有效抑制管柱形成、細胞爬行和生長,其IC50為5-50 nM。並且也發現可以抑制胰臟癌細胞(AsPC-1、BxPC-3、PANC-1和Mia paca-2)的在vitronectin和fibronectin上的黏著。然而無法抑制黑色素細胞瘤A375細胞在fibronectin的黏著。而Rho和KG抑制胰臟癌和黑色素細胞瘤細胞爬行能力的IC50為40-250 nM。細胞存活實驗也顯示Rho和KG可以抑制細胞生長和藉由Akt/caspase-3途徑誘導細胞凋亡。異種移植動物模型也顯示KG可以抑制胰臟癌腫瘤的生長。這些研究結果可以提供對於癌症治療去設計高度專一性整合蛋白藥物提供一個新的觀點。
英文摘要 Integrins are a family of α/β heterodimeric receptors and modulate many cellular processes. These integrin ligands employ a variant of the RGD, NGR, or LDV motifs as a key element of their major recognition site. RGD-containing ligands are shown to collaborate with specific flanking residues, synergistic site, and C-terminal region to control their integrin binding affinity and specificity. The NGR motif in the 5th and 7th type I repeats of fibronectin is known to bind integrin α5β1, and the spontaneous deamidation of NGR to isoDGR, an integrin αvβ3-binding motif, plays an important role in fibronectin fibril assembly and in regulating endothelial cell adhesion and proliferation. In my study I used rhodostomin (Rho), a disintegrin containing a 48PRGDMP53 motif, as a protein scaffold to investigate the regions of disintegrins involved in integrin interactions. The cell adhesion assay, NMR technique, X-ray crystallography, and molecular docking were used to study structure and functional relationships of disintegrin and integrins. To date, we have successfully used Rho to design potent and selective integrin-specific antagonists, including 48ARLDDL, an integrin αvβ3-specific mutant, and KG, an integrin αvβx-specific mutant. In my study I found the N-terminal region (residues 39-47) adjacent to the RGD motif that plays an important role in interacting with integrins αvβ3, αIIbβ3 and α5β1. The cell adhesion analysis of N-terminal mutants showed that Rho mutant with a 39KKKRT sequence exhibited the highest inhibitory activity with 3.6-, 4.6-, and 6.2-fold increases in inhibiting integrins αIIbβ3, αvβ3, and α5β1 in comparison with those of Rho 48ARGDNP-67NGLYG mutant. We also showed that Rho N-terminal mutant with a 46RR sequence exhibited 3.7- and 3.8-fold increases in inhibiting integrins αvβ3 and α5β1. In contrast, Rho mutant with a 46DD sequence exhibited 7.2-, 124.5-, and >582.0-fold decreases in inhibiting integrins αIIbβ3, αvβ3, and α5β1. The C-terminal mutations of 54DD into YY caused the increases 2.7- and 3.8-fold increases in inhibiting integrins αvβ3 and α5β1. 3D structures of Rho, KKKRT, and R46E mutants were determined by X-ray crystallography. Structural analysis showed that R42 of Rho KKKRT mutant interacted with C-terminal Y67 and H68 residues, which was not found from Rho with a SRAGK sequence. The docking of Rho and its KKKRT mutant into integrins showed that the sidechain of the R42 residue of KKKRT mutant interacted with the residue D216 of β1 and β3. In contrast, these interactions were absent in the Rho-integrin complexes. The sidechain orientation of the R46 residue in Rho and the E46 residue in R46E mutant were different. The docking analysis showed that the sidechain of R46 of Rho, but not E46 of R46E mutant, formed a salt bridge with D159 of αIIb subunit. These results demonstrate that the regions adjacent to the RGD motif in disintegrins affect their function and interactions with integrins. The incorporation of GNGRG amino acid sequence into Rho was used to study its role in integrin recognition. Mass and NMR analyses found that it can be converted into isoDGR and DRG isomers. The cell adhesion analysis showed that two isomers exhibited similar integrin αvβ3 inhibitory activity with the IC50 value of ~250 μM. In contrast, two isomers exhibited higher integrin α5β1 inhibitory activity. The DGR isomer had the IC50 value of 4.5 μM that is 6-fold more active than that of isoDGR isomer, suggesting that integrin ligands with an isoDGR motif are better ligands for integrin α5β1. Flow cytometry and western blot analysis showed that integrin β3-knockdown A375 cells were established. The adhesion analysis showed that the β3-knockdown cells cannot adhere on fibrinogen. The migration analysis showed that Rho, RLD and ALRDDL mutants inhibited A375 cell with the IC50 values of 9.3, 37.4, and 45.1 nM. In contrast, Rho, RLD and ALRDDL mutants inhibited the migration of β3-knockdown cells with the IC50 values of 0.5, 3, and > 100 μM. This result demonstrated that integrin αvβ3-specific mutant inhibited migration of human melanoma cells A375 cells via an integrin β3-dependent pathway. The use of Rho, ARLDDL and KG mutants in inhibiting angiogenesis, pancreatic cancer, and melanoma were evaluated. We found that they effectively inhibited tube formation, migration, and cell growth of endothelial cells with the IC50 values of 5-50 nM. They also inhibited the adhesion of pancreatic cancer cells (AsPC-1, Bx-PC-3, PANC-1, and Mia Paca-2) to vitronectin and fibronectin. In contrast, they cannot inhibit the adhesion of human melanoma A375 cell to fibronectin. We found that Rho and KG inhibited the migration of pancreatic cancer and melanoma cells with the IC50 values of 40-250 nM. The cell viability analysis showed that they also inhibited cell growth and induced cell apoptosis via AKT/caspase-3 pathway. Xenograft animal model showed that KG can inhibit the growth of pancreatic tumor. The results of this will provide new insight into the design of integrin-specific drugs for cancer therapy.
論文目次 CHINESE ABSTRACT I
ABSTRACT III
ACKNOWLEDGMENT V
TABLE OF CONTENTS VI
LIST OF TABLES XI
LIST OF FIGURES XII
ABBREVIATION XIV
CHAPTER 1 INTRODUCTION 1
1.1 RATIONALE 1
1.2 INTEGRINS 1
1.2.1 Overview of integrins 1
1.2.2 Integrin structures 2
1.2.3 Integrins and cancers 3
1.2.4 Integrins and melanoma 4
1.2.5 Integrins and pancreatic cancer 5
1.2.6 Integrins and angiogenesis 5
1.3 INTEGRINS AND ITS LIGANDS 6
1.4 DISINTEGRINS 8
1.4.1 Overview of disintegrins 8
1.4.2 Biomedical applications of disintegrins 9
1.5 RHODOSTOMIN (Rho) 10
1.5.1 Functions of Rhodostomin 10
1.5.2 Structure of Rhodostomin 11
1.5.3 Potent and selecitive Rhodostomin mutants 11
1.6 INTEGRINS AND NGR-COTAINING LIGANDS 12
1.7 STRUCTURE-BASED TECHNIQUE IN PROTEIN DRUG DEVELOPMENT 13
1.7.1 Nuclear magnetic resonance (NMR) 13
1.7.2 X-ray crystallography 13
1.7.3 Molecular docking 14
1.8 SPECIFIC AIMS AND RATIONALES 15
1.8.1 N-terminal region adjacent to RGD loop of disintegrin in integrin recognition 15
1.8.2 Rho and αvβx and α5β1-specific Rho mutant, 39KKART43-46AR47- 48GRGDNP53, in pancreatic cancer therapy 16
1.8.3 Rho and αvβ3-specific Rho mutant 48ARLDDL53 in melanoma 16
1.8.4 Rho and αvβx and α5β1-specific Rho mutant, 39KKART43-46AR47- 48GRGDNP53, in angiogenesis and tumor progression 17
1.8.5 Deamidation of NGR containing ligands 17
CHAPTER 2 MATERIALS AND METHODS 19
2.1 EXPRESSION AND PURIFICATION OF RHO AND ITS MUTANTS 19
2.2 CELL LINES 20
2.3 ANTIBODIES 20
2.4 PLATELET AGGREGATION ASSAY 20
2.5 CELL ADHESION ASSAY 21
2.6 PROTEIN CRYSTALLOGRAPHY 22
2.7 MOLECULAR DOCKING 23
2.8 FLOW CYTOMETRY 25
2.9 MIGRATION AND INVASION ASSAYS 25
2.10 VIABILITY ASSAY 26
2.11 TRANSFECTION OF A375 CELL 26
2.12 CELL APOPTOSIS ASSAY 27
2.13 TUBE FORMATION ASSAY 27
2.14 DEAMIDATION OF RHO MUTANTS 28
CHAPTER 3 RESULTS 29
3.1 EXPRESSION, PURIFICATION AND MASS CHARACTERIZATION OF RHO AND ITS MUTANTS 29
3.2 THE EFFECT OF N-TERMINAL REGION ADJACENT TO RGD LOOP ON INTEGRIN RECOGNITION 29
3.2.1 The effect of N-terminal region (residues 39-43) adjacent to RGD loop on platelet aggregation 30
3.2.2 The effect of N-terminal region (residues 39-43) adjacent to RGD loop on cell adhesion 30
3.2.3 The effect of N-terminal region (residues 46-47) adjacent to RGD loop on platelet aggregation 31
3.2.4 The effect of N-terminal region (residues 46-47) adjacent to RGD loop on cell adhesion 33
3.3 STRUCTURAL ANALYSIS AND DOCKING MODELS OF RHO AND ITS MUTANTS 35
3.3.1 Crystal structures of Rho KKKRT and R46E mutant 35
3.3.2 Structural differences between Rho and KKKRT mutant 36
3.3.3 Interactions in docking model of Rho KKKRT mutant 37
3.3.4 Structural differences between Rho and R46E mutant 38
3.3.5 The differences between the integrin αIIbβ3 complexes of Rho and R46E mutant 38
3.4 EFFECTS OF RHO AND αvβx- AND α5β1-SPECIFIC MUTANT ON HUMAN PANCREATIC CANCER CELL LINES 39
3.4.1 Integrin expression profile of pancreatic cancer cell lines 39
3.4.2 Inhibition of pancreatic cell adhesion by Rho and αvβx- and α5β1-specific mutant 40
3.4.3 Inhibition of pancreatic cell migration by Rho and αvβx- and α5β1-specific mutant 41
3.4.4 Inhibition of cell viability by Rho and αvβx- and α5β1-specific mutant 42
3.4.5 Inhibition of Mia paca-2 cell vibility by Rho and αvβx- and α5β1-specific through inducing apoptosis and reduction akt phosphorylation 43
3.5 EFFECT OF αvβ3-SPECIFIC MUTANT ON MELANOMA 44
3.5.1 Inhibition of melanoma cell adhesion, migration and invasion by integrin αvβ3-specific mutants 44
3.5.2 Involvement of integrin αvβ3 with melanoma cell adhesion, migration and invasion 45
3.6 EFFECTS OF ANGIOGENESIS BY RHO AND ITS INTEGRIN SPECIFIC MUTANTS 46
3.6.1 Inhibition of HUVEC viability by Rho and its mutants 46
3.6.2 Inhibition of serum-induced HUVEC migration by Rho and and its mutants 46
3.6.3 Inhibition of tube formation by Rho and and its mutants 47
3.7 DEAMIDATION OF NGR CONTAINING DISINTEGRINS AND INTEGRIN RECOGNITION 47
CHAPTER 4 DISCUSSION 49
4.1 EFFECT OF N-TERMINAL REGION ADJACENT TO RGD LOOP ON INTEGRINS BINDING 49
4.2 BINDING MECHANISM OF RGD LIGANDS TO INTEGRINS 49
4.2.1 Comparison of synergy region of fibronectin and the N-terminal region adjacent to RGD loop of Rhodostomin 50
4.2.2 Integrin binding site of the N-terminal and C-terminal region adjacent to RGD loop of Rhodostomin 51
4.3 DISINTEGRINS FOR THE TREATMENT OF PANCREATIC CANCER 52
4.4 DISINTEGRINS FOR THE TREATMENT OF MELANOMA 54
4.5 DISINTEGRINS FOR THE INHIBITION OF ANGIOGENESIS 56
4.6 BINDING MECHANISM OF NGR LIGANDS TO INTEGRINS 57
CHAPTER 5 CONCLUSIONS 58
REFERENCES 61
TABLES 70
FIGURES 78
PUBLICATIONS 106
APPENDIX 107
參考文獻 Abdollahi, A., Griggs, D.W., Zieher, H., Roth, A., Lipson, K.E., Saffrich, R., Grone, H.J., Hallahan, D.E., Reisfeld, R.A., Debus, J., et al. (2005). Inhibition of alpha(v)beta3 integrin survival signaling enhances antiangiogenic and antitumor effects of radiotherapy. Clinical cancer research : an official journal of the American Association for Cancer Research 11, 6270-6279.

Au, L.C., Huang, Y.B., Huang, T.F., Teh, G.W., Lin, H.H., and Choo, K.B. (1991). A common precursor for a putative hemorrhagic protein and rhodostomin, a platelet aggregation inhibitor of the venom of Calloselasma rhodostoma: molecular cloning and sequence analysis. Biochem Biophys Res Commun 181, 585-593.

Bhaskar, V., Zhang, D., Fox, M., Seto, P., Wong, M.H., Wales, P.E., Powers, D., Chao, D.T., Dubridge, R.B., and Ramakrishnan, V. (2007). A function blocking anti-mouse integrin alpha5beta1 antibody inhibits angiogenesis and impedes tumor growth in vivo. Journal of translational medicine 5, 61.

Buckley, C.D., Pilling, D., Henriquez, N.V., Parsonage, G., Threlfall, K., Scheel-Toellner, D., Simmons, D.L., Akbar, A.N., Lord, J.M., and Salmon, M. (1999). RGD peptides induce apoptosis by direct caspase-3 activation. Nature 397, 534-539.

Calvete, J.J., Marcinkiewicz, C., Monleon, D., Esteve, V., Celda, B., Juarez, P., and Sanz, L. (2005). Snake venom disintegrins: evolution of structure and function. Toxicon : official journal of the International Society on Toxinology 45, 1063-1074.

Chen, Y.C., Cheng, C.H., Shiu, J.H., Chang, Y.T., Chang, Y.S., Huang, C.H., Lee, J.C., and Chuang, W.J. (2012). Expression in Pichia pastoris and characterization of echistatin, an RGD-containing short disintegrin. Toxicon : official journal of the International Society on Toxinology 60, 1342-1348.

Curnis, F., Longhi, R., Crippa, L., Cattaneo, A., Dondossola, E., Bachi, A., and Corti, A. (2006). Spontaneous formation of L-isoaspartate and gain of function in fibronectin. J Biol Chem 281, 36466-36476.

de Vries, S.J., van Dijk, M., and Bonvin, A.M. (2010). The HADDOCK web server for data-driven biomolecular docking. Nature protocols 5, 883-897.

Dennis, M.S., Carter, P., and Lazarus, R.A. (1993). Binding interactions of kistrin with platelet glycoprotein IIb-IIIa: analysis by site-directed mutagenesis. Proteins 15, 312-321.

Dennis, M.S., Henzel, W.J., Pitti, R.M., Lipari, M.T., Napier, M.A., Deisher, T.A., Bunting, S., and Lazarus, R.A. (1990). Platelet glycoprotein IIb-IIIa protein antagonists from snake venoms: evidence for a family of platelet-aggregation inhibitors. Proc Natl Acad Sci U S A 87, 2471-2475.

Desgrosellier, J.S., and Cheresh, D.A. (2010). Integrins in cancer: biological implications and therapeutic opportunities. Nature reviews Cancer 10, 9-22.

Di Matteo, P., Curnis, F., Longhi, R., Colombo, G., Sacchi, A., Crippa, L., Protti, M.P., Ponzoni, M., Toma, S., and Corti, A. (2006). Immunogenic and structural properties of the Asn-Gly-Arg (NGR) tumor neovasculature-homing motif. Molecular immunology 43, 1509-1518.

Felding-Habermann, B., Fransvea, E., O'Toole, T.E., Manzuk, L., Faha, B., and Hensler, M. (2002). Involvement of tumor cell integrin alpha v beta 3 in hematogenous metastasis of human melanoma cells. Clinical & experimental metastasis 19, 427-436.

Fokas, E., Engenhart-Cabillic, R., Daniilidis, K., Rose, F., and An, H.X. (2007). Metastasis: the seed and soil theory gains identity. Cancer metastasis reviews 26, 705-715.

Friess, H., Langrehr, J.M., Oettle, H., Raedle, J., Niedergethmann, M., Dittrich, C., Hossfeld, D.K., Stoger, H., Neyns, B., Herzog, P., et al. (2006). A randomized multi-center phase II trial of the angiogenesis inhibitor Cilengitide (EMD 121974) and gemcitabine compared with gemcitabine alone in advanced unresectable pancreatic cancer. BMC cancer 6, 285.

Goh, K.L., Yang, J.T., and Hynes, R.O. (1997). Mesodermal defects and cranial neural crest apoptosis in alpha5 integrin-null embryos. Development (Cambridge, England) 124, 4309-4319.

Gould, R.J., Polokoff, M.A., Friedman, P.A., Huang, T.F., Holt, J.C., Cook, J.J., and Niewiarowski, S. (1990). Disintegrins: a family of integrin inhibitory proteins from viper venoms. Proc Soc Exp Biol Med 195, 168-171.

Graells, J., Vinyals, A., Figueras, A., Llorens, A., Moreno, A., Marcoval, J., Gonzalez, F.J., and Fabra, A. (2004). Overproduction of VEGF concomitantly expressed with its receptors promotes growth and survival of melanoma cells through MAPK and PI3K signaling. The Journal of investigative dermatology 123, 1151-1161.

Grzesiak, J.J., and Bouvet, M. (2006). The alpha2beta1 integrin mediates the malignant phenotype on type I collagen in pancreatic cancer cell lines. British journal of cancer 94, 1311-1319.

Grzesiak, J.J., Tran Cao, H.S., Burton, D.W., Kaushal, S., Vargas, F., Clopton, P., Snyder, C.S., Deftos, L.J., Hoffman, R.M., and Bouvet, M. (2011). Knockdown of the beta(1) integrin subunit reduces primary tumor growth and inhibits pancreatic cancer metastasis. Int J Cancer 129, 2905-2915.

Guo, R.T., Chou, L.J., Chen, Y.C., Chen, C.Y., Pari, K., Jen, C.J., Lo, S.J., Huang, S.L., Lee, C.Y., Chang, T.W., et al. (2001). Expression in Pichia pastoris and characterization by circular dichroism and NMR of rhodostomin. Proteins 43, 499-508.

Hailey, S., Adams, E., Penn, R., Wong, A., and McLane, M.A. (2013). Effect of the disintegrin eristostatin on melanoma-natural killer cell interactions. Toxicon : official journal of the International Society on Toxinology 61, 83-93.

Hausner, S.H., Abbey, C.K., Bold, R.J., Gagnon, M.K., Marik, J., Marshall, J.F., Stanecki, C.E., and Sutcliffe, J.L. (2009). Targeted in vivo imaging of integrin alphavbeta6 with an improved radiotracer and its relevance in a pancreatic tumor model. Cancer research 69, 5843-5850.

Hersey, P., Sosman, J., O'Day, S., Richards, J., Bedikian, A., Gonzalez, R., Sharfman, W., Weber, R., Logan, T., Buzoianu, M., et al. (2010). A randomized phase 2 study of etaracizumab, a monoclonal antibody against integrin alpha(v)beta(3), + or - dacarbazine in patients with stage IV metastatic melanoma. Cancer 116, 1526-1534.

Hezel, A.F., Deshpande, V., Zimmerman, S.M., Contino, G., Alagesan, B., O'Dell, M.R., Rivera, L.B., Harper, J., Lonning, S., Brekken, R.A., et al. (2012). TGF-beta and alphavbeta6 integrin act in a common pathway to suppress pancreatic cancer progression. Cancer research 72, 4840-4845.

Hofmann, U.B., Westphal, J.R., Waas, E.T., Becker, J.C., Ruiter, D.J., and van Muijen, G.N. (2000). Coexpression of integrin alpha(v)beta3 and matrix metalloproteinase-2 (MMP-2) coincides with MMP-2 activation: correlation with melanoma progression. The Journal of investigative dermatology 115, 625-632.

Humphries, J.D., Byron, A., and Humphries, M.J. (2006). Integrin ligands at a glance. Journal of cell science 119, 3901-3903.

Hynes, R.O. (2002). Integrins: bidirectional, allosteric signaling machines. Cell 110, 673-687.

Kageshita, T., Hamby, C.V., Hirai, S., Kimura, T., Ono, T., and Ferrone, S. (2000). Alpha(v)beta3 expression on blood vessels and melanoma cells in primary lesions: differential association with tumor progression and clinical prognosis. Cancer immunology, immunotherapy : CII 49, 314-318.

Kim, J.B., Yu, J.H., Ko, E., Lee, K.W., Song, A.K., Park, S.Y., Shin, I., Han, W., and Noh, D.Y. (2010). The alkaloid Berberine inhibits the growth of Anoikis-resistant MCF-7 and MDA-MB-231 breast cancer cell lines by inducing cell cycle arrest. Phytomedicine : international journal of phytotherapy and phytopharmacology 17, 436-440.

Koh, D.C., Armugam, A., and Jeyaseelan, K. (2006). Snake venom components and their applications in biomedicine. Cellular and molecular life sciences : CMLS 63, 3030-3041.

Koivunen, E., Gay, D.A., and Ruoslahti, E. (1993). Selection of peptides binding to the alpha 5 beta 1 integrin from phage display library. J Biol Chem 268, 20205-20210.
Koivunen, E., Wang, B., and Ruoslahti, E. (1994). Isolation of a highly specific ligand for the alpha 5 beta 1 integrin from a phage display library. J Cell Biol 124, 373-380.

Krammer, A., Craig, D., Thomas, W.E., Schulten, K., and Vogel, V. (2002). A structural model for force regulated integrin binding to fibronectin's RGD-synergy site. Matrix biology : journal of the International Society for Matrix Biology 21, 139-147.

Laing, G.D., and Moura-da-Silva, A.M. (2005). Jararhagin and its multiple effects on hemostasis. Toxicon : official journal of the International Society on Toxinology 45, 987-996.

Li, X., Regezi, J., Ross, F.P., Blystone, S., Ilic, D., Leong, S.P., and Ramos, D.M. (2001). Integrin alphavbeta3 mediates K1735 murine melanoma cell motility in vivo and in vitro. Journal of cell science 114, 2665-2672.

Liu, C.Z., Wang, Y.W., Shen, M.C., and Huang, T.F. (1994). Analysis of human platelet glycoprotein IIb-IIIa by fluorescein isothiocyanate-conjugated disintegrins with flow cytometry. Thrombosis and haemostasis 72, 919-925.

Mas-Moruno, C., Rechenmacher, F., and Kessler, H. (2010). Cilengitide: the first anti-angiogenic small molecule drug candidate design, synthesis and clinical evaluation. Anti-cancer agents in medicinal chemistry 10, 753-768.

Minea, R.O., Helchowski, C.M., Zidovetzki, S.J., Costa, F.K., Swenson, S.D., and Markland, F.S., Jr. (2010). Vicrostatin - an anti-invasive multi-integrin targeting chimeric disintegrin with tumor anti-angiogenic and pro-apoptotic activities. PloS one 5, e10929.

Miyamoto, H., Murakami, T., Tsuchida, K., Sugino, H., Miyake, H., and Tashiro, S. (2004). Tumor-stroma interaction of human pancreatic cancer: acquired resistance to anticancer drugs and proliferation regulation is dependent on extracellular matrix proteins. Pancreas 28, 38-44.

Mousa, S.A. (2002). Anti-integrin as novel drug-discovery targets: potential therapeutic and diagnostic implications. Current opinion in chemical biology 6, 534-541.

Mousa, S.A. (2003). Antiplatelet therapies: platelet GPIIb/IIIa antagonists and beyond. Current pharmaceutical design 9, 2317-2322.

Nagae, M., Re, S., Mihara, E., Nogi, T., Sugita, Y., and Takagi, J. (2012). Crystal structure of alpha5beta1 integrin ectodomain: atomic details of the fibronectin receptor. J Cell Biol 197, 131-140.

Nakamura, I., Duong le, T., Rodan, S.B., and Rodan, G.A. (2007). Involvement of alpha(v)beta3 integrins in osteoclast function. Journal of bone and mineral metabolism 25, 337-344.

Ogawa, T., Takayama, K., Takakura, N., Kitano, S., and Ueno, H. (2002). Anti-tumor angiogenesis therapy using soluble receptors: enhanced inhibition of tumor growth when soluble fibroblast growth factor receptor-1 is used with soluble vascular endothelial growth factor receptor. Cancer gene therapy 9, 633-640.

Oliva, I.B., Coelho, R.M., Barcellos, G.G., Saldanha-Gama, R., Wermelinger, L.S., Marcinkiewicz, C., Benedeta Zingali, R., and Barja-Fidalgo, C. (2007). Effect of RGD-disintegrins on melanoma cell growth and metastasis: involvement of the actin cytoskeleton, FAK and c-Fos. Toxicon : official journal of the International Society on Toxinology 50, 1053-1063.

Omura, Y., Chen, Y., Lermand, O., Jones, M., Duvvi, H., and Shimotsuura, Y. (2010). Effects of transcutaneous electrical stimulation (1 pulse/sec) through custom-made disposable surface electrodes covering Omura's ST36 area of both legs on normal cell telomeres, oncogen C-fosAb2, integrin alpha5beta1, chlamydia trachomatis, etc. in breast cancer & alzheimer patients. Acupuncture & electro-therapeutics research 35, 147-185.

Pierschbacher, M.D., and Ruoslahti, E. (1984). Cell attachment activity of fibronectin can be duplicated by small synthetic fragments of the molecule. Nature 309, 30-33.

Ramos, O.H., Kauskot, A., Cominetti, M.R., Bechyne, I., Salla Pontes, C.L., Chareyre, F., Manent, J., Vassy, R., Giovannini, M., Legrand, C., et al. (2008). A novel alpha(v)beta (3)-blocking disintegrin containing the RGD motive, DisBa-01, inhibits bFGF-induced angiogenesis and melanoma metastasis. Clinical & experimental metastasis 25, 53-64.

Redick, S.D., Settles, D.L., Briscoe, G., and Erickson, H.P. (2000). Defining fibronectin's cell adhesion synergy site by site-directed mutagenesis. J Cell Biol 149, 521-527.

Ricono, J.M., Huang, M., Barnes, L.A., Lau, S.K., Weis, S.M., Schlaepfer, D.D., Hanks, S.K., and Cheresh, D.A. (2009). Specific cross-talk between epidermal growth factor receptor and integrin alphavbeta5 promotes carcinoma cell invasion and metastasis. Cancer Res 69, 1383-1391.

Selistre-de-Araujo, H.S., Pontes, C.L., Montenegro, C.F., and Martin, A.C. (2010). Snake venom disintegrins and cell migration. Toxins 2, 2606-2621.

Sheu, J.R., Yen, M.H., Kan, Y.C., Hung, W.C., Chang, P.T., and Luk, H.N. (1997). Inhibition of angiogenesis in vitro and in vivo: comparison of the relative activities of triflavin, an Arg-Gly-Asp-containing peptide and anti-alpha(v)beta3 integrin monoclonal antibody. Biochimica et biophysica acta 1336, 445-454.

Shimizu, T., Matsuoka, Y., and Shirasawa, T. (2005). Biological significance of isoaspartate and its repair system. Biol Pharm Bull 28, 1590-1596.

Sumathipala, R., Xu, C., Seago, J., Mould, A.P., Humphries, M.J., Craig, S.E., Patel, Y., Wijelath, E.S., Sobel, M., and Rahman, S. (2006). The "linker" region (amino acids 38-47) of the disintegrin elegantin is a novel inhibitory domain of integrin alpha5beta1-dependent cell adhesion on fibronectin: evidence for the negative regulation of fibronectin synergy site biological activity. J Biol Chem 281, 37686-37696.

Swenson, S., Costa, F., Ernst, W., Fujii, G., and Markland, F.S. (2005). Contortrostatin, a snake venom disintegrin with anti-angiogenic and anti-tumor activity. Pathophysiology of haemostasis and thrombosis 34, 169-176.

Van Agthoven, J.F., Xiong, J.P., Alonso, J.L., Rui, X., Adair, B.D., Goodman, S.L., and Arnaout, M.A. (2014). Structural basis for pure antagonism of integrin alphaVbeta3 by a high-affinity form of fibronectin. Nature structural & molecular biology 21, 383-388.

van der Walle, C.F., Altroff, H., and Mardon, H.J. (2002). Novel mutant human fibronectin FIII9-10 domain pair with increased conformational stability and biological activity. Protein engineering 15, 1021-1024.

Wang, G.K., and Zhang, W. (2005). The signaling network of tumor invasion. Histology and histopathology 20, 593-602.

Weis, S.M., and Cheresh, D.A. (2011). alphaV integrins in angiogenesis and cancer. Cold Spring Harbor perspectives in medicine 1, a006478.

Xiao, T., Takagi, J., Coller, B.S., Wang, J.H., and Springer, T.A. (2004). Structural basis for allostery in integrins and binding to fibrinogen-mimetic therapeutics. Nature 432, 59-67.

Xiong, J.P., Stehle, T., Diefenbach, B., Zhang, R., Dunker, R., Scott, D.L., Joachimiak, A., Goodman, S.L., and Arnaout, M.A. (2001). Crystal structure of the extracellular segment of integrin alpha Vbeta3. Science 294, 339-345.

Xiong, J.P., Stehle, T., Zhang, R., Joachimiak, A., Frech, M., Goodman, S.L., and Arnaout, M.A. (2002). Crystal structure of the extracellular segment of integrin alpha Vbeta3 in complex with an Arg-Gly-Asp ligand. Science 296, 151-155.

Yang, R.S., Tang, C.H., Chuang, W.J., Huang, T.H., Peng, H.C., Huang, T.F., and Fu, W.M. (2005). Inhibition of tumor formation by snake venom disintegrin. Toxicon : official journal of the International Society on Toxinology 45, 661-669.

Yeh, C.H., Peng, H.C., Yang, R.S., and Huang, T.F. (2001). Rhodostomin, a snake venom disintegrin, inhibits angiogenesis elicited by basic fibroblast growth factor and suppresses tumor growth by a selective alpha(v)beta(3) blockade of endothelial cells. Mol Pharmacol 59, 1333-1342.

Zhu, J., Zhu, J., and Springer, T.A. (2013). Complete integrin headpiece opening in eight steps. The Journal of cell biology 201, 1053-1068.

Chiu-Yueh Chen and Woei-Jer Chuang (2006). Use Rhodostomin to study the integrin recognition sequences to establish a method for preparing amino-acid –type selective isotope labeling of proteins

Jia-Hau Shiu and Woei-Jer Chuang (2012). Structure, dynamics, and function relationships of Rhodostomin mutants and variants: insight into their interactions with integrins

Yao-Tsung Chang and Woei-Jer Chuang (2014). Structure-activity relationships of RGD loop, linker region, and C-terminus of Rhodostomin mutants in the recognition of integrins

Yung-Sheng Chang and Woei-Jer Chuang (2013). Design, structure determination, and biological evaluation of potent integrin α5β1- and/or αvβ3-specific antagonist using the ninth and/or tenth module of fibronectin type III domain
Ching-Ting Liau and Woei-Jer Chuang (2008). The role of the linker region of Rhodostomin and trimucrin in recognizing integrins

Yi-Hsueh Lu and Woei-Jer Chuang (2009). Optimization of the fermentation production in pichia pastoris and the function of Rhodostomin mutants

Chun-Hao Chen and Woei-Jer Chuang (2010). The role of Rhodostomin mutants with an RXD motif in integrin recognition and anti-melanoma tumor cell activity

Ping-Tse Chung and Woei-Jer Chuang (2012). Development of integrins αvβx- and α5β1-specific antagonists using Rhodostomin as a scaffold
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