Site specific discrete PEGylation of (124)I-labeled mCC49 Fab' fragments improves tumor MicroPET/CT imaging in mice
The tumor-associated glycoprotein-72 (TAG-72) antigen is highly overexpressed in various human adenocarcinomas and anti-TAG-72 monoclonal antibodies, and fragments are therefore useful as pharmaceutical targeting vectors. In this study, we investigated the effects of site-specific PEGylation with MW...
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| Veröffentlicht in: | Bioconjugate chemistry 2013-11, Vol.24 (11), p.1945-1954 |
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| creator | Ding, Haiming Carlton, Michelle M Povoski, Stephen P Milum, Keisha Kumar, Krishan Kothandaraman, Shankaran Hinkle, George H Colcher, David Brody, Rich Davis, Paul D Pokora, Alex Phelps, Mitchell Martin, Jr, Edward W Tweedle, Michael F |
| description | The tumor-associated glycoprotein-72 (TAG-72) antigen is highly overexpressed in various human adenocarcinomas and anti-TAG-72 monoclonal antibodies, and fragments are therefore useful as pharmaceutical targeting vectors. In this study, we investigated the effects of site-specific PEGylation with MW 2-4 kDa discrete, branched PEGylation reagents on mCC49 Fab' (MW 50 kDa) via in vitro TAG72 binding, and in vivo blood clearance kinetics, biodistribution, and mouse tumor microPET/CT imaging. mCC49Fab' (Fab'-NEM) was conjugated at a hinge region cysteine with maleimide-dPEG 12-(dPEG24COOH)3 acid (Mal-dPEG-A), maleimide-dPEG12-(dPEG12COOH)3 acid (Mal-dPEG-B), or maleimide-dPEG12-(m-dPEG24)3 (Mal-dPEG-C), and then radiolabeled with iodine-124 ((124)I) in vitro radioligand binding assays and in vivo studies used TAG-72 expressing LS174T human colon carcinoma cells and xenograft mouse tumors. Conjugation of mCC49Fab' with Mal-dPEG-A (Fab'-A) reduced the binding affinity of the non PEGylated Fab' by 30%; however, in vivo, Fab'-A significantly lengthened the blood retention vs Fab'-NEM (47.5 vs 28.1%/ID at 1 h, 25.1 vs 8.4%/ID at 5 h, p < 0.01), showed excellent tumor to background, better microPET/CT images due to higher tumor accumulation, and increased tumor concentration in excised tissues at 72 h by 130% (5.09 ± 0.83 vs 3.83 ± 1.50%ID/g, p < 0.05). Despite the strong similarity of the three PEGylation reagents, PEGylation with Mal-dPEG-B or -C reduced the in vitro binding affinity of Fab'-NEM by 70%, blood retention, microPET/CT imaging tumor signal intensity, and residual 72 h tumor concentration by 49% (3.83 ± 1.50 vs 1.97 ± 0.29%ID/g, p < 0.05) and 63% (3.83 ± 1.50 vs 1.42 ± 0.35%ID/g, p < 0.05), respectively. We conclude that remarkably subtle changes in the structure of the PEGylation reagent can create significantly altered biologic behavior. Further study is warranted of conjugates of the triple branched, negatively charged Mal-dPEG-A. |
| doi_str_mv | 10.1021/bc400375f |
| format | Article |
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In this study, we investigated the effects of site-specific PEGylation with MW 2-4 kDa discrete, branched PEGylation reagents on mCC49 Fab' (MW 50 kDa) via in vitro TAG72 binding, and in vivo blood clearance kinetics, biodistribution, and mouse tumor microPET/CT imaging. mCC49Fab' (Fab'-NEM) was conjugated at a hinge region cysteine with maleimide-dPEG 12-(dPEG24COOH)3 acid (Mal-dPEG-A), maleimide-dPEG12-(dPEG12COOH)3 acid (Mal-dPEG-B), or maleimide-dPEG12-(m-dPEG24)3 (Mal-dPEG-C), and then radiolabeled with iodine-124 ((124)I) in vitro radioligand binding assays and in vivo studies used TAG-72 expressing LS174T human colon carcinoma cells and xenograft mouse tumors. Conjugation of mCC49Fab' with Mal-dPEG-A (Fab'-A) reduced the binding affinity of the non PEGylated Fab' by 30%; however, in vivo, Fab'-A significantly lengthened the blood retention vs Fab'-NEM (47.5 vs 28.1%/ID at 1 h, 25.1 vs 8.4%/ID at 5 h, p < 0.01), showed excellent tumor to background, better microPET/CT images due to higher tumor accumulation, and increased tumor concentration in excised tissues at 72 h by 130% (5.09 ± 0.83 vs 3.83 ± 1.50%ID/g, p < 0.05). Despite the strong similarity of the three PEGylation reagents, PEGylation with Mal-dPEG-B or -C reduced the in vitro binding affinity of Fab'-NEM by 70%, blood retention, microPET/CT imaging tumor signal intensity, and residual 72 h tumor concentration by 49% (3.83 ± 1.50 vs 1.97 ± 0.29%ID/g, p < 0.05) and 63% (3.83 ± 1.50 vs 1.42 ± 0.35%ID/g, p < 0.05), respectively. We conclude that remarkably subtle changes in the structure of the PEGylation reagent can create significantly altered biologic behavior. Further study is warranted of conjugates of the triple branched, negatively charged Mal-dPEG-A.</description><identifier>EISSN: 1520-4812</identifier><identifier>DOI: 10.1021/bc400375f</identifier><identifier>PMID: 24175669</identifier><language>eng</language><publisher>United States</publisher><subject>Animals ; Antigens, Neoplasm - immunology ; Colonic Neoplasms - diagnosis ; Female ; Humans ; Immunoglobulin Fab Fragments - chemistry ; Immunoglobulin Fab Fragments - immunology ; Iodine Radioisotopes - chemistry ; Mice ; Mice, Nude ; Molecular Structure ; Multimodal Imaging - methods ; Neoplasms, Experimental - diagnosis ; Polyethylene Glycols - chemistry ; Positron-Emission Tomography - methods ; Tomography, X-Ray Computed - methods</subject><ispartof>Bioconjugate chemistry, 2013-11, Vol.24 (11), p.1945-1954</ispartof><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>314,776,780,27901,27902</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/24175669$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Ding, Haiming</creatorcontrib><creatorcontrib>Carlton, Michelle M</creatorcontrib><creatorcontrib>Povoski, Stephen P</creatorcontrib><creatorcontrib>Milum, Keisha</creatorcontrib><creatorcontrib>Kumar, Krishan</creatorcontrib><creatorcontrib>Kothandaraman, Shankaran</creatorcontrib><creatorcontrib>Hinkle, George H</creatorcontrib><creatorcontrib>Colcher, David</creatorcontrib><creatorcontrib>Brody, Rich</creatorcontrib><creatorcontrib>Davis, Paul D</creatorcontrib><creatorcontrib>Pokora, Alex</creatorcontrib><creatorcontrib>Phelps, Mitchell</creatorcontrib><creatorcontrib>Martin, Jr, Edward W</creatorcontrib><creatorcontrib>Tweedle, Michael F</creatorcontrib><title>Site specific discrete PEGylation of (124)I-labeled mCC49 Fab' fragments improves tumor MicroPET/CT imaging in mice</title><title>Bioconjugate chemistry</title><addtitle>Bioconjug Chem</addtitle><description>The tumor-associated glycoprotein-72 (TAG-72) antigen is highly overexpressed in various human adenocarcinomas and anti-TAG-72 monoclonal antibodies, and fragments are therefore useful as pharmaceutical targeting vectors. In this study, we investigated the effects of site-specific PEGylation with MW 2-4 kDa discrete, branched PEGylation reagents on mCC49 Fab' (MW 50 kDa) via in vitro TAG72 binding, and in vivo blood clearance kinetics, biodistribution, and mouse tumor microPET/CT imaging. mCC49Fab' (Fab'-NEM) was conjugated at a hinge region cysteine with maleimide-dPEG 12-(dPEG24COOH)3 acid (Mal-dPEG-A), maleimide-dPEG12-(dPEG12COOH)3 acid (Mal-dPEG-B), or maleimide-dPEG12-(m-dPEG24)3 (Mal-dPEG-C), and then radiolabeled with iodine-124 ((124)I) in vitro radioligand binding assays and in vivo studies used TAG-72 expressing LS174T human colon carcinoma cells and xenograft mouse tumors. Conjugation of mCC49Fab' with Mal-dPEG-A (Fab'-A) reduced the binding affinity of the non PEGylated Fab' by 30%; however, in vivo, Fab'-A significantly lengthened the blood retention vs Fab'-NEM (47.5 vs 28.1%/ID at 1 h, 25.1 vs 8.4%/ID at 5 h, p < 0.01), showed excellent tumor to background, better microPET/CT images due to higher tumor accumulation, and increased tumor concentration in excised tissues at 72 h by 130% (5.09 ± 0.83 vs 3.83 ± 1.50%ID/g, p < 0.05). Despite the strong similarity of the three PEGylation reagents, PEGylation with Mal-dPEG-B or -C reduced the in vitro binding affinity of Fab'-NEM by 70%, blood retention, microPET/CT imaging tumor signal intensity, and residual 72 h tumor concentration by 49% (3.83 ± 1.50 vs 1.97 ± 0.29%ID/g, p < 0.05) and 63% (3.83 ± 1.50 vs 1.42 ± 0.35%ID/g, p < 0.05), respectively. We conclude that remarkably subtle changes in the structure of the PEGylation reagent can create significantly altered biologic behavior. Further study is warranted of conjugates of the triple branched, negatively charged Mal-dPEG-A.</description><subject>Animals</subject><subject>Antigens, Neoplasm - immunology</subject><subject>Colonic Neoplasms - diagnosis</subject><subject>Female</subject><subject>Humans</subject><subject>Immunoglobulin Fab Fragments - chemistry</subject><subject>Immunoglobulin Fab Fragments - immunology</subject><subject>Iodine Radioisotopes - chemistry</subject><subject>Mice</subject><subject>Mice, Nude</subject><subject>Molecular Structure</subject><subject>Multimodal Imaging - methods</subject><subject>Neoplasms, Experimental - diagnosis</subject><subject>Polyethylene Glycols - chemistry</subject><subject>Positron-Emission Tomography - methods</subject><subject>Tomography, X-Ray Computed - methods</subject><issn>1520-4812</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2013</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNo1kF1LwzAUhoMgbk4v_AOSO-dFXU6SNumllG0OJg6c1yVNT0akXzatsH9vQb06cM7Dy3NeQu6APQHjsCqsZEyo2F2QOcScRVIDn5HrED4ZYylofkVmXIKKkySdk_DuB6ShQ-udt7T0wfY4bQ7r7bkyg28b2jq6BC4fd1FlCqywpHWWyZRuTPFAXW9ONTZDoL7u-vYbAx3Guu3pq7d9e1gfV9lxOpmTb07UN7T2Fm_IpTNVwNu_uSAfm_Uxe4n2b9td9ryPOuDJEMlUOSkVaCswRsVAmMJoEEmMohDM2FKUWoPC0iobcyM1l4nWCQiH6BQTC7L8zZ3EvkYMQ15P72FVmQbbMeQgJ1YCU8mE3v-hY1FjmXf95Nyf8_-ixA_QJ2Vt</recordid><startdate>20131120</startdate><enddate>20131120</enddate><creator>Ding, Haiming</creator><creator>Carlton, Michelle M</creator><creator>Povoski, Stephen P</creator><creator>Milum, Keisha</creator><creator>Kumar, Krishan</creator><creator>Kothandaraman, Shankaran</creator><creator>Hinkle, George H</creator><creator>Colcher, David</creator><creator>Brody, Rich</creator><creator>Davis, Paul D</creator><creator>Pokora, Alex</creator><creator>Phelps, Mitchell</creator><creator>Martin, Jr, Edward W</creator><creator>Tweedle, Michael F</creator><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>7X8</scope></search><sort><creationdate>20131120</creationdate><title>Site specific discrete PEGylation of (124)I-labeled mCC49 Fab' fragments improves tumor MicroPET/CT imaging in mice</title><author>Ding, Haiming ; Carlton, Michelle M ; Povoski, Stephen P ; Milum, Keisha ; Kumar, Krishan ; Kothandaraman, Shankaran ; Hinkle, George H ; Colcher, David ; Brody, Rich ; Davis, Paul D ; Pokora, Alex ; Phelps, Mitchell ; Martin, Jr, Edward W ; Tweedle, Michael F</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-p126t-497f44718c3e5e7013aba81365e3b30acd3d8817edc7c52a4824688613feef703</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2013</creationdate><topic>Animals</topic><topic>Antigens, Neoplasm - immunology</topic><topic>Colonic Neoplasms - diagnosis</topic><topic>Female</topic><topic>Humans</topic><topic>Immunoglobulin Fab Fragments - chemistry</topic><topic>Immunoglobulin Fab Fragments - immunology</topic><topic>Iodine Radioisotopes - chemistry</topic><topic>Mice</topic><topic>Mice, Nude</topic><topic>Molecular Structure</topic><topic>Multimodal Imaging - methods</topic><topic>Neoplasms, Experimental - diagnosis</topic><topic>Polyethylene Glycols - chemistry</topic><topic>Positron-Emission Tomography - methods</topic><topic>Tomography, X-Ray Computed - methods</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Ding, Haiming</creatorcontrib><creatorcontrib>Carlton, Michelle M</creatorcontrib><creatorcontrib>Povoski, Stephen P</creatorcontrib><creatorcontrib>Milum, Keisha</creatorcontrib><creatorcontrib>Kumar, Krishan</creatorcontrib><creatorcontrib>Kothandaraman, Shankaran</creatorcontrib><creatorcontrib>Hinkle, George H</creatorcontrib><creatorcontrib>Colcher, David</creatorcontrib><creatorcontrib>Brody, Rich</creatorcontrib><creatorcontrib>Davis, Paul D</creatorcontrib><creatorcontrib>Pokora, Alex</creatorcontrib><creatorcontrib>Phelps, Mitchell</creatorcontrib><creatorcontrib>Martin, Jr, Edward W</creatorcontrib><creatorcontrib>Tweedle, Michael F</creatorcontrib><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>MEDLINE - Academic</collection><jtitle>Bioconjugate chemistry</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Ding, Haiming</au><au>Carlton, Michelle M</au><au>Povoski, Stephen P</au><au>Milum, Keisha</au><au>Kumar, Krishan</au><au>Kothandaraman, Shankaran</au><au>Hinkle, George H</au><au>Colcher, David</au><au>Brody, Rich</au><au>Davis, Paul D</au><au>Pokora, Alex</au><au>Phelps, Mitchell</au><au>Martin, Jr, Edward W</au><au>Tweedle, Michael F</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Site specific discrete PEGylation of (124)I-labeled mCC49 Fab' fragments improves tumor MicroPET/CT imaging in mice</atitle><jtitle>Bioconjugate chemistry</jtitle><addtitle>Bioconjug Chem</addtitle><date>2013-11-20</date><risdate>2013</risdate><volume>24</volume><issue>11</issue><spage>1945</spage><epage>1954</epage><pages>1945-1954</pages><eissn>1520-4812</eissn><abstract>The tumor-associated glycoprotein-72 (TAG-72) antigen is highly overexpressed in various human adenocarcinomas and anti-TAG-72 monoclonal antibodies, and fragments are therefore useful as pharmaceutical targeting vectors. In this study, we investigated the effects of site-specific PEGylation with MW 2-4 kDa discrete, branched PEGylation reagents on mCC49 Fab' (MW 50 kDa) via in vitro TAG72 binding, and in vivo blood clearance kinetics, biodistribution, and mouse tumor microPET/CT imaging. mCC49Fab' (Fab'-NEM) was conjugated at a hinge region cysteine with maleimide-dPEG 12-(dPEG24COOH)3 acid (Mal-dPEG-A), maleimide-dPEG12-(dPEG12COOH)3 acid (Mal-dPEG-B), or maleimide-dPEG12-(m-dPEG24)3 (Mal-dPEG-C), and then radiolabeled with iodine-124 ((124)I) in vitro radioligand binding assays and in vivo studies used TAG-72 expressing LS174T human colon carcinoma cells and xenograft mouse tumors. Conjugation of mCC49Fab' with Mal-dPEG-A (Fab'-A) reduced the binding affinity of the non PEGylated Fab' by 30%; however, in vivo, Fab'-A significantly lengthened the blood retention vs Fab'-NEM (47.5 vs 28.1%/ID at 1 h, 25.1 vs 8.4%/ID at 5 h, p < 0.01), showed excellent tumor to background, better microPET/CT images due to higher tumor accumulation, and increased tumor concentration in excised tissues at 72 h by 130% (5.09 ± 0.83 vs 3.83 ± 1.50%ID/g, p < 0.05). Despite the strong similarity of the three PEGylation reagents, PEGylation with Mal-dPEG-B or -C reduced the in vitro binding affinity of Fab'-NEM by 70%, blood retention, microPET/CT imaging tumor signal intensity, and residual 72 h tumor concentration by 49% (3.83 ± 1.50 vs 1.97 ± 0.29%ID/g, p < 0.05) and 63% (3.83 ± 1.50 vs 1.42 ± 0.35%ID/g, p < 0.05), respectively. We conclude that remarkably subtle changes in the structure of the PEGylation reagent can create significantly altered biologic behavior. Further study is warranted of conjugates of the triple branched, negatively charged Mal-dPEG-A.</abstract><cop>United States</cop><pmid>24175669</pmid><doi>10.1021/bc400375f</doi><tpages>10</tpages></addata></record> |
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| subjects | Animals Antigens, Neoplasm - immunology Colonic Neoplasms - diagnosis Female Humans Immunoglobulin Fab Fragments - chemistry Immunoglobulin Fab Fragments - immunology Iodine Radioisotopes - chemistry Mice Mice, Nude Molecular Structure Multimodal Imaging - methods Neoplasms, Experimental - diagnosis Polyethylene Glycols - chemistry Positron-Emission Tomography - methods Tomography, X-Ray Computed - methods |
| title | Site specific discrete PEGylation of (124)I-labeled mCC49 Fab' fragments improves tumor MicroPET/CT imaging in mice |
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