Construction and characterization of an anti-asialoglycoprotein receptor single-chain variable-fragment-targeted melittin
Antibody–therapeutic agent conjugation to be delivered specifically to tumor cells is required for many target‐based therapeutic strategies. In the present study, a recombinant immunotoxin was constructed by which melittin was fused to an anti‐asialoglycoprotein receptor (ASGPR) single‐chain variabl...
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| Veröffentlicht in: | Biotechnology and applied biochemistry 2011-11, Vol.58 (6), p.405-411 |
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| creator | Zhao, Xiaorong Yu, Zhen Dai, Wentao Yao, Zhenjiang Zhou, Wei Zhou, Weiping Zhou, Junli Yang, Yonggang Zhu, Yinchang Chen, Sidong Cao, Limin |
| description | Antibody–therapeutic agent conjugation to be delivered specifically to tumor cells is required for many target‐based therapeutic strategies. In the present study, a recombinant immunotoxin was constructed by which melittin was fused to an anti‐asialoglycoprotein receptor (ASGPR) single‐chain variable fragment antibody (C1), and targeting ability and cytolytic efficacy of the fusion protein were studied. Our results suggested that the recombinant 29.4 kDa protein C1M was expressed in Escherichia coli as a soluble style. Binding of C1M to the surface of hepatocellular carcinoma (HCC) cells was confirmed by both immunohistochemistry and flow cytometry assays. C1M kept the hemolytic activity of melittin and exhibited cytolytic capacity to HepG2 cells at a concentration of 1.5 µg/mL, under which erythrocytes would not be lysed. The effects were greatly inhibited by coadministration with asialoorosomucoid, a natural ligand for ASGPR. These results suggested that C1M conferred targeting and ASGPR‐specific cytotoxicity to HCC cells. This work makes it possible to further investigate its antihepatoma efficacy in vivo. |
| doi_str_mv | 10.1002/bab.57 |
| format | Article |
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In the present study, a recombinant immunotoxin was constructed by which melittin was fused to an anti‐asialoglycoprotein receptor (ASGPR) single‐chain variable fragment antibody (C1), and targeting ability and cytolytic efficacy of the fusion protein were studied. Our results suggested that the recombinant 29.4 kDa protein C1M was expressed in Escherichia coli as a soluble style. Binding of C1M to the surface of hepatocellular carcinoma (HCC) cells was confirmed by both immunohistochemistry and flow cytometry assays. C1M kept the hemolytic activity of melittin and exhibited cytolytic capacity to HepG2 cells at a concentration of 1.5 µg/mL, under which erythrocytes would not be lysed. The effects were greatly inhibited by coadministration with asialoorosomucoid, a natural ligand for ASGPR. These results suggested that C1M conferred targeting and ASGPR‐specific cytotoxicity to HCC cells. This work makes it possible to further investigate its antihepatoma efficacy in vivo.</description><identifier>ISSN: 0885-4513</identifier><identifier>EISSN: 1470-8744</identifier><identifier>DOI: 10.1002/bab.57</identifier><identifier>PMID: 22172103</identifier><identifier>CODEN: BABIEC</identifier><language>eng</language><publisher>Hoboken: Wiley Subscription Services, Inc., A Wiley Company</publisher><subject>asialoglycoprotein receptor ; Asialoglycoprotein Receptor - genetics ; Asialoglycoprotein Receptor - immunology ; Asialoglycoproteins - pharmacology ; Base Sequence ; Biological and medical sciences ; Biotechnology ; Carcinoma, Hepatocellular - drug therapy ; Cell Line, Tumor ; Escherichia coli - genetics ; Fundamental and applied biological sciences. Psychology ; HCC ; Hemolytic Agents - pharmacology ; Humans ; Immunotoxins - pharmacology ; Liver Neoplasms - drug therapy ; Melitten - genetics ; Melitten - pharmacology ; melittin ; Molecular Sequence Data ; Orosomucoid - analogs & derivatives ; Orosomucoid - pharmacology ; Recombinant Fusion Proteins - genetics ; Recombinant Fusion Proteins - metabolism ; Recombinant Fusion Proteins - pharmacology ; recombinant immunotoxin ; Single-Chain Antibodies ; single-chain Fv ; targeting</subject><ispartof>Biotechnology and applied biochemistry, 2011-11, Vol.58 (6), p.405-411</ispartof><rights>Copyright © 2011 International Union of Biochemistry and Molecular Biology, Inc.</rights><rights>2015 INIST-CNRS</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c3807-f0bc82cb50b796367c19fc3272986d84ee96ee21cdb4f8132aee51e52f91f7b83</citedby><cites>FETCH-LOGICAL-c3807-f0bc82cb50b796367c19fc3272986d84ee96ee21cdb4f8132aee51e52f91f7b83</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://onlinelibrary.wiley.com/doi/pdf/10.1002%2Fbab.57$$EPDF$$P50$$Gwiley$$H</linktopdf><linktohtml>$$Uhttps://onlinelibrary.wiley.com/doi/full/10.1002%2Fbab.57$$EHTML$$P50$$Gwiley$$H</linktohtml><link.rule.ids>314,776,780,1411,27903,27904,45553,45554</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=25303976$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/22172103$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Zhao, Xiaorong</creatorcontrib><creatorcontrib>Yu, Zhen</creatorcontrib><creatorcontrib>Dai, Wentao</creatorcontrib><creatorcontrib>Yao, Zhenjiang</creatorcontrib><creatorcontrib>Zhou, Wei</creatorcontrib><creatorcontrib>Zhou, Weiping</creatorcontrib><creatorcontrib>Zhou, Junli</creatorcontrib><creatorcontrib>Yang, Yonggang</creatorcontrib><creatorcontrib>Zhu, Yinchang</creatorcontrib><creatorcontrib>Chen, Sidong</creatorcontrib><creatorcontrib>Cao, Limin</creatorcontrib><title>Construction and characterization of an anti-asialoglycoprotein receptor single-chain variable-fragment-targeted melittin</title><title>Biotechnology and applied biochemistry</title><addtitle>Biotechnology and Applied Biochemistry</addtitle><description>Antibody–therapeutic agent conjugation to be delivered specifically to tumor cells is required for many target‐based therapeutic strategies. In the present study, a recombinant immunotoxin was constructed by which melittin was fused to an anti‐asialoglycoprotein receptor (ASGPR) single‐chain variable fragment antibody (C1), and targeting ability and cytolytic efficacy of the fusion protein were studied. Our results suggested that the recombinant 29.4 kDa protein C1M was expressed in Escherichia coli as a soluble style. Binding of C1M to the surface of hepatocellular carcinoma (HCC) cells was confirmed by both immunohistochemistry and flow cytometry assays. C1M kept the hemolytic activity of melittin and exhibited cytolytic capacity to HepG2 cells at a concentration of 1.5 µg/mL, under which erythrocytes would not be lysed. The effects were greatly inhibited by coadministration with asialoorosomucoid, a natural ligand for ASGPR. These results suggested that C1M conferred targeting and ASGPR‐specific cytotoxicity to HCC cells. This work makes it possible to further investigate its antihepatoma efficacy in vivo.</description><subject>asialoglycoprotein receptor</subject><subject>Asialoglycoprotein Receptor - genetics</subject><subject>Asialoglycoprotein Receptor - immunology</subject><subject>Asialoglycoproteins - pharmacology</subject><subject>Base Sequence</subject><subject>Biological and medical sciences</subject><subject>Biotechnology</subject><subject>Carcinoma, Hepatocellular - drug therapy</subject><subject>Cell Line, Tumor</subject><subject>Escherichia coli - genetics</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>HCC</subject><subject>Hemolytic Agents - pharmacology</subject><subject>Humans</subject><subject>Immunotoxins - pharmacology</subject><subject>Liver Neoplasms - drug therapy</subject><subject>Melitten - genetics</subject><subject>Melitten - pharmacology</subject><subject>melittin</subject><subject>Molecular Sequence Data</subject><subject>Orosomucoid - analogs & derivatives</subject><subject>Orosomucoid - pharmacology</subject><subject>Recombinant Fusion Proteins - genetics</subject><subject>Recombinant Fusion Proteins - metabolism</subject><subject>Recombinant Fusion Proteins - pharmacology</subject><subject>recombinant immunotoxin</subject><subject>Single-Chain Antibodies</subject><subject>single-chain Fv</subject><subject>targeting</subject><issn>0885-4513</issn><issn>1470-8744</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2011</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNp10EuPFCEQB3BiNO646kcwfVHjgZVH08Bxd_ahZn3E6HokQFePaDc9AqOOn17cGdeTCQmh8qOq8kfoISVHlBD23Fl3JOQttKCtJFjJtr2NFkQpgVtB-QG6l_MXQoiSit1FB4xRySjhC7RdzjGXtPElzLGxsW_8Z5usL5DCL3tdnIdar6cEbHOw47wat35ep7lAiE0CD-sypyaHuBoB1--1-t2mYF19DsmuJogFF5tWUKBvJhhDKSHeR3cGO2Z4sL8P0cfzsw_LF_jy7cXL5fEl9lwRiQfivGLeCeKk7ngnPdWD50wyrbpetQC6A2DU964dFOXMAggKgg2aDtIpfoie7vrWjb9tIBczhexhHG2EeZONplS3WklS5ZOd9GnOOcFg1ilMNm0NJeZPyqambISs8NG-5cZN0N-wv7FW8HgPbPZ2rCFEH_I_JzjhWnbVPdu5H2GE7X_GmZPjk-uheGdDLvDzxtr01XSSS2E-vbkw7PTV1bvXV6fmPf8NhY2kHg</recordid><startdate>201111</startdate><enddate>201111</enddate><creator>Zhao, Xiaorong</creator><creator>Yu, Zhen</creator><creator>Dai, Wentao</creator><creator>Yao, Zhenjiang</creator><creator>Zhou, Wei</creator><creator>Zhou, Weiping</creator><creator>Zhou, Junli</creator><creator>Yang, Yonggang</creator><creator>Zhu, Yinchang</creator><creator>Chen, Sidong</creator><creator>Cao, Limin</creator><general>Wiley Subscription Services, Inc., A Wiley Company</general><general>Wiley</general><scope>BSCLL</scope><scope>IQODW</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7X8</scope></search><sort><creationdate>201111</creationdate><title>Construction and characterization of an anti-asialoglycoprotein receptor single-chain variable-fragment-targeted melittin</title><author>Zhao, Xiaorong ; Yu, Zhen ; Dai, Wentao ; Yao, Zhenjiang ; Zhou, Wei ; Zhou, Weiping ; Zhou, Junli ; Yang, Yonggang ; Zhu, Yinchang ; Chen, Sidong ; Cao, Limin</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c3807-f0bc82cb50b796367c19fc3272986d84ee96ee21cdb4f8132aee51e52f91f7b83</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2011</creationdate><topic>asialoglycoprotein receptor</topic><topic>Asialoglycoprotein Receptor - genetics</topic><topic>Asialoglycoprotein Receptor - immunology</topic><topic>Asialoglycoproteins - pharmacology</topic><topic>Base Sequence</topic><topic>Biological and medical sciences</topic><topic>Biotechnology</topic><topic>Carcinoma, Hepatocellular - drug therapy</topic><topic>Cell Line, Tumor</topic><topic>Escherichia coli - genetics</topic><topic>Fundamental and applied biological sciences. Psychology</topic><topic>HCC</topic><topic>Hemolytic Agents - pharmacology</topic><topic>Humans</topic><topic>Immunotoxins - pharmacology</topic><topic>Liver Neoplasms - drug therapy</topic><topic>Melitten - genetics</topic><topic>Melitten - pharmacology</topic><topic>melittin</topic><topic>Molecular Sequence Data</topic><topic>Orosomucoid - analogs & derivatives</topic><topic>Orosomucoid - pharmacology</topic><topic>Recombinant Fusion Proteins - genetics</topic><topic>Recombinant Fusion Proteins - metabolism</topic><topic>Recombinant Fusion Proteins - pharmacology</topic><topic>recombinant immunotoxin</topic><topic>Single-Chain Antibodies</topic><topic>single-chain Fv</topic><topic>targeting</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Zhao, Xiaorong</creatorcontrib><creatorcontrib>Yu, Zhen</creatorcontrib><creatorcontrib>Dai, Wentao</creatorcontrib><creatorcontrib>Yao, Zhenjiang</creatorcontrib><creatorcontrib>Zhou, Wei</creatorcontrib><creatorcontrib>Zhou, Weiping</creatorcontrib><creatorcontrib>Zhou, Junli</creatorcontrib><creatorcontrib>Yang, Yonggang</creatorcontrib><creatorcontrib>Zhu, Yinchang</creatorcontrib><creatorcontrib>Chen, Sidong</creatorcontrib><creatorcontrib>Cao, Limin</creatorcontrib><collection>Istex</collection><collection>Pascal-Francis</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>MEDLINE - Academic</collection><jtitle>Biotechnology and applied biochemistry</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Zhao, Xiaorong</au><au>Yu, Zhen</au><au>Dai, Wentao</au><au>Yao, Zhenjiang</au><au>Zhou, Wei</au><au>Zhou, Weiping</au><au>Zhou, Junli</au><au>Yang, Yonggang</au><au>Zhu, Yinchang</au><au>Chen, Sidong</au><au>Cao, Limin</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Construction and characterization of an anti-asialoglycoprotein receptor single-chain variable-fragment-targeted melittin</atitle><jtitle>Biotechnology and applied biochemistry</jtitle><addtitle>Biotechnology and Applied Biochemistry</addtitle><date>2011-11</date><risdate>2011</risdate><volume>58</volume><issue>6</issue><spage>405</spage><epage>411</epage><pages>405-411</pages><issn>0885-4513</issn><eissn>1470-8744</eissn><coden>BABIEC</coden><abstract>Antibody–therapeutic agent conjugation to be delivered specifically to tumor cells is required for many target‐based therapeutic strategies. In the present study, a recombinant immunotoxin was constructed by which melittin was fused to an anti‐asialoglycoprotein receptor (ASGPR) single‐chain variable fragment antibody (C1), and targeting ability and cytolytic efficacy of the fusion protein were studied. Our results suggested that the recombinant 29.4 kDa protein C1M was expressed in Escherichia coli as a soluble style. Binding of C1M to the surface of hepatocellular carcinoma (HCC) cells was confirmed by both immunohistochemistry and flow cytometry assays. C1M kept the hemolytic activity of melittin and exhibited cytolytic capacity to HepG2 cells at a concentration of 1.5 µg/mL, under which erythrocytes would not be lysed. The effects were greatly inhibited by coadministration with asialoorosomucoid, a natural ligand for ASGPR. These results suggested that C1M conferred targeting and ASGPR‐specific cytotoxicity to HCC cells. This work makes it possible to further investigate its antihepatoma efficacy in vivo.</abstract><cop>Hoboken</cop><pub>Wiley Subscription Services, Inc., A Wiley Company</pub><pmid>22172103</pmid><doi>10.1002/bab.57</doi><tpages>7</tpages></addata></record> |
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| subjects | asialoglycoprotein receptor Asialoglycoprotein Receptor - genetics Asialoglycoprotein Receptor - immunology Asialoglycoproteins - pharmacology Base Sequence Biological and medical sciences Biotechnology Carcinoma, Hepatocellular - drug therapy Cell Line, Tumor Escherichia coli - genetics Fundamental and applied biological sciences. Psychology HCC Hemolytic Agents - pharmacology Humans Immunotoxins - pharmacology Liver Neoplasms - drug therapy Melitten - genetics Melitten - pharmacology melittin Molecular Sequence Data Orosomucoid - analogs & derivatives Orosomucoid - pharmacology Recombinant Fusion Proteins - genetics Recombinant Fusion Proteins - metabolism Recombinant Fusion Proteins - pharmacology recombinant immunotoxin Single-Chain Antibodies single-chain Fv targeting |
| title | Construction and characterization of an anti-asialoglycoprotein receptor single-chain variable-fragment-targeted melittin |
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