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Enhancement of anti-TNFα monoclonal antibody production in CHO cells through the use of UCOE and DHFR elements in vector construction and the optimization of cell culture media
Recently, there has been a high demand for anti-tumor necrosis factor-α monoclonal antibodies (mAbTNFα) in the treatment of rheumatoid arthritis and other autoimmune diseases. Thus, efficient strategies and stable high-producing cell lines need to be established to increase antibody production. In t...
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| Published in: | Preparative biochemistry & biotechnology 2022, Vol.52 (4), p.452-470 |
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| container_title | Preparative biochemistry & biotechnology |
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| creator | Doan, Chinh Chung Ho, Nguyen Quynh Chi Nguyen, Thi Thuy Nguyen, Thi Phuong Thao Do, Dang Giap Hoang, Nghia Son Le, Thanh Long |
| description | Recently, there has been a high demand for anti-tumor necrosis factor-α monoclonal antibodies (mAbTNFα) in the treatment of rheumatoid arthritis and other autoimmune diseases. Thus, efficient strategies and stable high-producing cell lines need to be established to increase antibody production. In this study, we describe an efficient approach to establish a mAbTNFα high-producing clone through the optimization of expression vectors and cell culture media. The ubiquitous chromatin opening element (UCOE) and dihydrofolate reductase (DHFR)-based vectors encoding mAbTNFα were introduced into the CHO-DG44 cells using lipofection. Clones were obtained by selecting transfected cells with G418, amplifying them by treatment with methotrexate, and isolating them by limiting dilution. Different media formulated with commercial feeds and media were also screened to develop an improved medium. The antibody produced by the selected clone was purified, characterized, and compared to standard adalimumab. Using our established protocol, a cell clone obtained from stable mAbTNFα-expressing cell pools showed a 3.8-fold higher antibody titer compared to stable cell pools. Furthermore, the highest antibody yield of selected clones cultured in fed-batch mode using improved medium was 2450 ± 30 µg/mL, which was 13.2-fold higher than that of stable cell pool cultivated in batch mode using a basal medium. The purified antibody had primary chemical and biological characteristics similar to those of adalimumab. Therefore, the use of UCOE and DHFR vectors in combination with the optimization of cell culture media may help in establishing stable and high-producing CHO cell lines for therapeutic antibody production. |
| doi_str_mv | 10.1080/10826068.2021.1963981 |
| format | article |
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Thus, efficient strategies and stable high-producing cell lines need to be established to increase antibody production. In this study, we describe an efficient approach to establish a mAbTNFα high-producing clone through the optimization of expression vectors and cell culture media. The ubiquitous chromatin opening element (UCOE) and dihydrofolate reductase (DHFR)-based vectors encoding mAbTNFα were introduced into the CHO-DG44 cells using lipofection. Clones were obtained by selecting transfected cells with G418, amplifying them by treatment with methotrexate, and isolating them by limiting dilution. Different media formulated with commercial feeds and media were also screened to develop an improved medium. The antibody produced by the selected clone was purified, characterized, and compared to standard adalimumab. Using our established protocol, a cell clone obtained from stable mAbTNFα-expressing cell pools showed a 3.8-fold higher antibody titer compared to stable cell pools. Furthermore, the highest antibody yield of selected clones cultured in fed-batch mode using improved medium was 2450 ± 30 µg/mL, which was 13.2-fold higher than that of stable cell pool cultivated in batch mode using a basal medium. The purified antibody had primary chemical and biological characteristics similar to those of adalimumab. Therefore, the use of UCOE and DHFR vectors in combination with the optimization of cell culture media may help in establishing stable and high-producing CHO cell lines for therapeutic antibody production.</description><identifier>ISSN: 1082-6068</identifier><identifier>EISSN: 1532-2297</identifier><identifier>DOI: 10.1080/10826068.2021.1963981</identifier><identifier>PMID: 34427158</identifier><language>eng</language><publisher>England: Taylor & Francis</publisher><subject>Adalimumab ; Animals ; anti-TNFα monoclonal antibody ; Antibodies ; Antibodies, Monoclonal - genetics ; Antibody Formation ; Arthritis ; Autoimmune diseases ; Batch culture ; Cell culture ; Cell Culture Techniques ; CHO Cells ; CHO-DG44 cell line ; Chromatin ; Cricetinae ; Cricetulus ; Culture media ; DHFR ; Dihydrofolate reductase ; Dilution ; Expression vectors ; gene amplification ; Media ; medium optimization ; Methotrexate ; Monoclonal antibodies ; Optimization ; Reductases ; Rheumatoid arthritis ; Tetrahydrofolate Dehydrogenase - genetics ; Tetrahydrofolate Dehydrogenase - metabolism ; TNF inhibitors ; Tumor Necrosis Factor-alpha - genetics ; Tumor necrosis factor-α ; UCOE element</subject><ispartof>Preparative biochemistry & biotechnology, 2022, Vol.52 (4), p.452-470</ispartof><rights>2021 Taylor & Francis Group, LLC 2021</rights><rights>2021 Taylor & Francis Group, LLC</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c394t-d8fc0914acdac0cf0ca925e5c2f88485aa3cc4bcb1f7f38bd4970909915d237a3</citedby><cites>FETCH-LOGICAL-c394t-d8fc0914acdac0cf0ca925e5c2f88485aa3cc4bcb1f7f38bd4970909915d237a3</cites><orcidid>0000-0003-4732-3643 ; 0000-0001-8743-0094 ; 0000-0001-7853-0990 ; 0000-0002-5002-6327 ; 0000-0002-3267-243X ; 0000-0002-2052-4560 ; 0000-0002-5807-2264</orcidid></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>315,786,790,4043,27956,27957,27958</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/34427158$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Doan, Chinh Chung</creatorcontrib><creatorcontrib>Ho, Nguyen Quynh Chi</creatorcontrib><creatorcontrib>Nguyen, Thi Thuy</creatorcontrib><creatorcontrib>Nguyen, Thi Phuong Thao</creatorcontrib><creatorcontrib>Do, Dang Giap</creatorcontrib><creatorcontrib>Hoang, Nghia Son</creatorcontrib><creatorcontrib>Le, Thanh Long</creatorcontrib><title>Enhancement of anti-TNFα monoclonal antibody production in CHO cells through the use of UCOE and DHFR elements in vector construction and the optimization of cell culture media</title><title>Preparative biochemistry & biotechnology</title><addtitle>Prep Biochem Biotechnol</addtitle><description>Recently, there has been a high demand for anti-tumor necrosis factor-α monoclonal antibodies (mAbTNFα) in the treatment of rheumatoid arthritis and other autoimmune diseases. Thus, efficient strategies and stable high-producing cell lines need to be established to increase antibody production. In this study, we describe an efficient approach to establish a mAbTNFα high-producing clone through the optimization of expression vectors and cell culture media. The ubiquitous chromatin opening element (UCOE) and dihydrofolate reductase (DHFR)-based vectors encoding mAbTNFα were introduced into the CHO-DG44 cells using lipofection. Clones were obtained by selecting transfected cells with G418, amplifying them by treatment with methotrexate, and isolating them by limiting dilution. Different media formulated with commercial feeds and media were also screened to develop an improved medium. The antibody produced by the selected clone was purified, characterized, and compared to standard adalimumab. Using our established protocol, a cell clone obtained from stable mAbTNFα-expressing cell pools showed a 3.8-fold higher antibody titer compared to stable cell pools. Furthermore, the highest antibody yield of selected clones cultured in fed-batch mode using improved medium was 2450 ± 30 µg/mL, which was 13.2-fold higher than that of stable cell pool cultivated in batch mode using a basal medium. The purified antibody had primary chemical and biological characteristics similar to those of adalimumab. Therefore, the use of UCOE and DHFR vectors in combination with the optimization of cell culture media may help in establishing stable and high-producing CHO cell lines for therapeutic antibody production.</description><subject>Adalimumab</subject><subject>Animals</subject><subject>anti-TNFα monoclonal antibody</subject><subject>Antibodies</subject><subject>Antibodies, Monoclonal - genetics</subject><subject>Antibody Formation</subject><subject>Arthritis</subject><subject>Autoimmune diseases</subject><subject>Batch culture</subject><subject>Cell culture</subject><subject>Cell Culture Techniques</subject><subject>CHO Cells</subject><subject>CHO-DG44 cell line</subject><subject>Chromatin</subject><subject>Cricetinae</subject><subject>Cricetulus</subject><subject>Culture media</subject><subject>DHFR</subject><subject>Dihydrofolate reductase</subject><subject>Dilution</subject><subject>Expression vectors</subject><subject>gene amplification</subject><subject>Media</subject><subject>medium optimization</subject><subject>Methotrexate</subject><subject>Monoclonal antibodies</subject><subject>Optimization</subject><subject>Reductases</subject><subject>Rheumatoid arthritis</subject><subject>Tetrahydrofolate Dehydrogenase - genetics</subject><subject>Tetrahydrofolate Dehydrogenase - metabolism</subject><subject>TNF inhibitors</subject><subject>Tumor Necrosis Factor-alpha - genetics</subject><subject>Tumor necrosis factor-α</subject><subject>UCOE element</subject><issn>1082-6068</issn><issn>1532-2297</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2022</creationdate><recordtype>article</recordtype><recordid>eNp9kc1uEzEUhUcIREvhEUCW2LCZ4L-ZsXegkBCkikioXVsej01czdjB9hSFt-qL8EzYScqCBRtf6-o759r3VNVrBBcIMvg-H7iFLVtgiNEC8ZZwhp5Ul6ghuMaYd0_zPTN1gS6qFzHeQYh4h9jz6oJQijvUsMvqYeV20ik9aZeAN0C6ZOubr-vfD2DyzqvROzkeu70fDmAf_DCrZL0D1oHlZguUHscI0i74-fsuVw3mqIvT7XK7ysIBfNqsvwE9HkfEIrvXKvkAlHcxhbNbAYvY75Od7C95bGaXYg_UPKY5aDDpwcqX1TMjx6hfnetVdbte3Sw39fX285flx-taEU5TPTCjIEdUqkEqqAxUkuNGNwobxihrpCRK0V71yHSGsH6gvIMcco6aAZNOkqvq3ck3__nHrGMSk43lOdJpP0eBm5YiwiAhGX37D3rn55AXl6mWZseW0CZTzYlSwccYtBH7YCcZDgJBUTIVj5mKkqk4Z5p1b87uc5838Ff1GGIGPpwA64wPk_zpwziIJA-jDybkdG0U5P8z_gCzI7N7</recordid><startdate>2022</startdate><enddate>2022</enddate><creator>Doan, Chinh Chung</creator><creator>Ho, Nguyen Quynh Chi</creator><creator>Nguyen, Thi Thuy</creator><creator>Nguyen, Thi Phuong Thao</creator><creator>Do, Dang Giap</creator><creator>Hoang, Nghia Son</creator><creator>Le, Thanh Long</creator><general>Taylor & Francis</general><general>Taylor & Francis Ltd</general><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>7QL</scope><scope>7QO</scope><scope>7QR</scope><scope>7T7</scope><scope>7TK</scope><scope>7TM</scope><scope>8FD</scope><scope>C1K</scope><scope>FR3</scope><scope>M7N</scope><scope>P64</scope><scope>7X8</scope><orcidid>https://orcid.org/0000-0003-4732-3643</orcidid><orcidid>https://orcid.org/0000-0001-8743-0094</orcidid><orcidid>https://orcid.org/0000-0001-7853-0990</orcidid><orcidid>https://orcid.org/0000-0002-5002-6327</orcidid><orcidid>https://orcid.org/0000-0002-3267-243X</orcidid><orcidid>https://orcid.org/0000-0002-2052-4560</orcidid><orcidid>https://orcid.org/0000-0002-5807-2264</orcidid></search><sort><creationdate>2022</creationdate><title>Enhancement of anti-TNFα monoclonal antibody production in CHO cells through the use of UCOE and DHFR elements in vector construction and the optimization of cell culture media</title><author>Doan, Chinh Chung ; 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Thus, efficient strategies and stable high-producing cell lines need to be established to increase antibody production. In this study, we describe an efficient approach to establish a mAbTNFα high-producing clone through the optimization of expression vectors and cell culture media. The ubiquitous chromatin opening element (UCOE) and dihydrofolate reductase (DHFR)-based vectors encoding mAbTNFα were introduced into the CHO-DG44 cells using lipofection. Clones were obtained by selecting transfected cells with G418, amplifying them by treatment with methotrexate, and isolating them by limiting dilution. Different media formulated with commercial feeds and media were also screened to develop an improved medium. The antibody produced by the selected clone was purified, characterized, and compared to standard adalimumab. Using our established protocol, a cell clone obtained from stable mAbTNFα-expressing cell pools showed a 3.8-fold higher antibody titer compared to stable cell pools. Furthermore, the highest antibody yield of selected clones cultured in fed-batch mode using improved medium was 2450 ± 30 µg/mL, which was 13.2-fold higher than that of stable cell pool cultivated in batch mode using a basal medium. The purified antibody had primary chemical and biological characteristics similar to those of adalimumab. Therefore, the use of UCOE and DHFR vectors in combination with the optimization of cell culture media may help in establishing stable and high-producing CHO cell lines for therapeutic antibody production.</abstract><cop>England</cop><pub>Taylor & Francis</pub><pmid>34427158</pmid><doi>10.1080/10826068.2021.1963981</doi><tpages>19</tpages><orcidid>https://orcid.org/0000-0003-4732-3643</orcidid><orcidid>https://orcid.org/0000-0001-8743-0094</orcidid><orcidid>https://orcid.org/0000-0001-7853-0990</orcidid><orcidid>https://orcid.org/0000-0002-5002-6327</orcidid><orcidid>https://orcid.org/0000-0002-3267-243X</orcidid><orcidid>https://orcid.org/0000-0002-2052-4560</orcidid><orcidid>https://orcid.org/0000-0002-5807-2264</orcidid></addata></record> |
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| subjects | Adalimumab Animals anti-TNFα monoclonal antibody Antibodies Antibodies, Monoclonal - genetics Antibody Formation Arthritis Autoimmune diseases Batch culture Cell culture Cell Culture Techniques CHO Cells CHO-DG44 cell line Chromatin Cricetinae Cricetulus Culture media DHFR Dihydrofolate reductase Dilution Expression vectors gene amplification Media medium optimization Methotrexate Monoclonal antibodies Optimization Reductases Rheumatoid arthritis Tetrahydrofolate Dehydrogenase - genetics Tetrahydrofolate Dehydrogenase - metabolism TNF inhibitors Tumor Necrosis Factor-alpha - genetics Tumor necrosis factor-α UCOE element |
| title | Enhancement of anti-TNFα monoclonal antibody production in CHO cells through the use of UCOE and DHFR elements in vector construction and the optimization of cell culture media |
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