Reduction of IL‐2 fragmentation during manufacturing of a novel immunocytokine by DoE process optimization
Interleukin‐2 (IL‐2) is a potent molecule in cancer therapy. Clinical application, however, is limited due to its strong side effects during the treatment. We developed an IL‐2 variant (IL‐2v) immunocytokine to circumvent the drawbacks of the current IL‐2 therapy. During the production of the IL‐2v...
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Veröffentlicht in: | Biotechnology and bioengineering 2019-10, Vol.116 (10), p.2503-2513 |
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description | Interleukin‐2 (IL‐2) is a potent molecule in cancer therapy. Clinical application, however, is limited due to its strong side effects during the treatment. We developed an IL‐2 variant (IL‐2v) immunocytokine to circumvent the drawbacks of the current IL‐2 therapy. During the production of the IL‐2v immunocytokine in Chinese hamster ovary (CHO) cells, molecules with fragmented IL‐2v and therefore reduced cytokine activity can be observed. To control product fragmentation different production process conditions were investigated. By shifting temperature or pH after the cell growth phase to lower values, fragmented species can be reduced from 10% to 12% to about 4%. However, with the adopted process conditions, the effective titer is decreased concomitantly. Moreover, fermentation length and inoculation cell density are parameters to adjust fragmentation and effective titer. A suitable method for efficient process optimization is the design of experiment approach. With this procedure, novel optimal values for temperature, pH value, harvest day, and inoculation cell densities were proposed and tested subsequently. In comparison to the former process, the improved process reduces fragmentation by 66% while keeping the effective titer comparable. In summary, these findings will help to control fragmentation in CHO production processes of different IL‐2v or IL‐2 containing therapeutic proteins.
During manufacturing of a novel IL‐2 variant (IL‐2v) immunocytokine in CHO cells molecules with fragmented IL‐2v moieties were observed. The decrease of the process parameters temperature, pH value, fermentation duration, and inoculation cell density were found to lower fragmentation but also product titer. Using a DoE optimization approach to balance fragmentation and titer, fragmented molecules were reduced by 66% in the improved process compared to the former process while keeping the effective titer comparable. |
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During manufacturing of a novel IL‐2 variant (IL‐2v) immunocytokine in CHO cells molecules with fragmented IL‐2v moieties were observed. The decrease of the process parameters temperature, pH value, fermentation duration, and inoculation cell density were found to lower fragmentation but also product titer. Using a DoE optimization approach to balance fragmentation and titer, fragmented molecules were reduced by 66% in the improved process compared to the former process while keeping the effective titer comparable.</description><identifier>ISSN: 0006-3592</identifier><identifier>EISSN: 1097-0290</identifier><identifier>DOI: 10.1002/bit.27090</identifier><identifier>PMID: 31180133</identifier><language>eng</language><publisher>United States: Wiley Subscription Services, Inc</publisher><subject>Cell density ; CHO cells ; Design of experiments ; Design optimization ; DoE ; Fermentation ; Fragmentation ; Inoculation ; Interleukins ; interleukin‐2 ; Menopause ; pH effects ; proteolytic cleavage ; Side effects ; Therapy</subject><ispartof>Biotechnology and bioengineering, 2019-10, Vol.116 (10), p.2503-2513</ispartof><rights>2019 Wiley Periodicals, Inc.</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c3900-647f437f69e7755bd2282e955f7555247283a0de558b4198f8eb8a63d9020af23</citedby><cites>FETCH-LOGICAL-c3900-647f437f69e7755bd2282e955f7555247283a0de558b4198f8eb8a63d9020af23</cites><orcidid>0000-0003-0093-7189 ; 0000-0003-0046-6452</orcidid></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://onlinelibrary.wiley.com/doi/pdf/10.1002%2Fbit.27090$$EPDF$$P50$$Gwiley$$H</linktopdf><linktohtml>$$Uhttps://onlinelibrary.wiley.com/doi/full/10.1002%2Fbit.27090$$EHTML$$P50$$Gwiley$$H</linktohtml><link.rule.ids>314,780,784,1417,27924,27925,45574,45575</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/31180133$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Schneider, Alina</creatorcontrib><creatorcontrib>Gorr, Ingo H.</creatorcontrib><creatorcontrib>Larraillet, Vincent</creatorcontrib><creatorcontrib>Frensing, Timo</creatorcontrib><creatorcontrib>Popp, Oliver</creatorcontrib><title>Reduction of IL‐2 fragmentation during manufacturing of a novel immunocytokine by DoE process optimization</title><title>Biotechnology and bioengineering</title><addtitle>Biotechnol Bioeng</addtitle><description>Interleukin‐2 (IL‐2) is a potent molecule in cancer therapy. Clinical application, however, is limited due to its strong side effects during the treatment. We developed an IL‐2 variant (IL‐2v) immunocytokine to circumvent the drawbacks of the current IL‐2 therapy. During the production of the IL‐2v immunocytokine in Chinese hamster ovary (CHO) cells, molecules with fragmented IL‐2v and therefore reduced cytokine activity can be observed. To control product fragmentation different production process conditions were investigated. By shifting temperature or pH after the cell growth phase to lower values, fragmented species can be reduced from 10% to 12% to about 4%. However, with the adopted process conditions, the effective titer is decreased concomitantly. Moreover, fermentation length and inoculation cell density are parameters to adjust fragmentation and effective titer. A suitable method for efficient process optimization is the design of experiment approach. With this procedure, novel optimal values for temperature, pH value, harvest day, and inoculation cell densities were proposed and tested subsequently. In comparison to the former process, the improved process reduces fragmentation by 66% while keeping the effective titer comparable. In summary, these findings will help to control fragmentation in CHO production processes of different IL‐2v or IL‐2 containing therapeutic proteins.
During manufacturing of a novel IL‐2 variant (IL‐2v) immunocytokine in CHO cells molecules with fragmented IL‐2v moieties were observed. The decrease of the process parameters temperature, pH value, fermentation duration, and inoculation cell density were found to lower fragmentation but also product titer. Using a DoE optimization approach to balance fragmentation and titer, fragmented molecules were reduced by 66% in the improved process compared to the former process while keeping the effective titer comparable.</description><subject>Cell density</subject><subject>CHO cells</subject><subject>Design of experiments</subject><subject>Design optimization</subject><subject>DoE</subject><subject>Fermentation</subject><subject>Fragmentation</subject><subject>Inoculation</subject><subject>Interleukins</subject><subject>interleukin‐2</subject><subject>Menopause</subject><subject>pH effects</subject><subject>proteolytic cleavage</subject><subject>Side effects</subject><subject>Therapy</subject><issn>0006-3592</issn><issn>1097-0290</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2019</creationdate><recordtype>article</recordtype><recordid>eNp1kc1O3DAURi1UBFNg0ReoLHVDF4FrO_5bFkphpJGQEKwjJ7GRaWJP46TVsOoj9Bn7JDUT6AKJ1dV3dXR0dT-EPhA4IQD0tPbjCZWgYQctCGhZANXwDi0AQBSMa7qP3qf0kKNUQuyhfUaIAsLYAnU3tp2a0ceAo8PL1d_ffyh2g7nvbRjNdt9Ogw_3uDdhcqYZ55Rhg0P8aTvs-34KsdmM8bsPFtcb_DVe4PUQG5sSjuvR9_5xqzpEu850yR49zwN09-3i9vyqWF1fLs-_rIqGaYBClNKVTDqhrZSc1y2lilrNucuJ01JSxQy0lnNVl0Qrp2ytjGCtBgrGUXaAjmdvPuLHZNNY9T41tutMsHFKFaWcCC25lhn99Ap9iNMQ8nWZUiUpqRYiU59nqhliSoN11XrwvRk2FYHqqYIqV1BtK8jsx2fjVPe2_U--_DwDpzPwy3d287apOlvezsp_N4SQMQ</recordid><startdate>201910</startdate><enddate>201910</enddate><creator>Schneider, Alina</creator><creator>Gorr, Ingo H.</creator><creator>Larraillet, Vincent</creator><creator>Frensing, Timo</creator><creator>Popp, Oliver</creator><general>Wiley Subscription Services, Inc</general><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7QF</scope><scope>7QO</scope><scope>7QQ</scope><scope>7SC</scope><scope>7SE</scope><scope>7SP</scope><scope>7SR</scope><scope>7T7</scope><scope>7TA</scope><scope>7TB</scope><scope>7U5</scope><scope>8BQ</scope><scope>8FD</scope><scope>C1K</scope><scope>F28</scope><scope>FR3</scope><scope>H8D</scope><scope>H8G</scope><scope>JG9</scope><scope>JQ2</scope><scope>KR7</scope><scope>L7M</scope><scope>L~C</scope><scope>L~D</scope><scope>P64</scope><scope>7X8</scope><orcidid>https://orcid.org/0000-0003-0093-7189</orcidid><orcidid>https://orcid.org/0000-0003-0046-6452</orcidid></search><sort><creationdate>201910</creationdate><title>Reduction of IL‐2 fragmentation during manufacturing of a novel immunocytokine by DoE process optimization</title><author>Schneider, Alina ; Gorr, Ingo H. ; Larraillet, Vincent ; Frensing, Timo ; Popp, Oliver</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c3900-647f437f69e7755bd2282e955f7555247283a0de558b4198f8eb8a63d9020af23</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2019</creationdate><topic>Cell density</topic><topic>CHO cells</topic><topic>Design of experiments</topic><topic>Design optimization</topic><topic>DoE</topic><topic>Fermentation</topic><topic>Fragmentation</topic><topic>Inoculation</topic><topic>Interleukins</topic><topic>interleukin‐2</topic><topic>Menopause</topic><topic>pH effects</topic><topic>proteolytic cleavage</topic><topic>Side effects</topic><topic>Therapy</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Schneider, Alina</creatorcontrib><creatorcontrib>Gorr, Ingo H.</creatorcontrib><creatorcontrib>Larraillet, Vincent</creatorcontrib><creatorcontrib>Frensing, Timo</creatorcontrib><creatorcontrib>Popp, Oliver</creatorcontrib><collection>PubMed</collection><collection>CrossRef</collection><collection>Aluminium Industry Abstracts</collection><collection>Biotechnology Research Abstracts</collection><collection>Ceramic Abstracts</collection><collection>Computer and Information Systems Abstracts</collection><collection>Corrosion Abstracts</collection><collection>Electronics & Communications Abstracts</collection><collection>Engineered Materials Abstracts</collection><collection>Industrial and Applied Microbiology Abstracts (Microbiology A)</collection><collection>Materials Business File</collection><collection>Mechanical & Transportation Engineering Abstracts</collection><collection>Solid State and Superconductivity Abstracts</collection><collection>METADEX</collection><collection>Technology Research Database</collection><collection>Environmental Sciences and Pollution Management</collection><collection>ANTE: Abstracts in New Technology & Engineering</collection><collection>Engineering Research Database</collection><collection>Aerospace Database</collection><collection>Copper Technical Reference Library</collection><collection>Materials Research Database</collection><collection>ProQuest Computer Science Collection</collection><collection>Civil Engineering Abstracts</collection><collection>Advanced Technologies Database with Aerospace</collection><collection>Computer and Information Systems Abstracts Academic</collection><collection>Computer and Information Systems Abstracts Professional</collection><collection>Biotechnology and BioEngineering Abstracts</collection><collection>MEDLINE - Academic</collection><jtitle>Biotechnology and bioengineering</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Schneider, Alina</au><au>Gorr, Ingo H.</au><au>Larraillet, Vincent</au><au>Frensing, Timo</au><au>Popp, Oliver</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Reduction of IL‐2 fragmentation during manufacturing of a novel immunocytokine by DoE process optimization</atitle><jtitle>Biotechnology and bioengineering</jtitle><addtitle>Biotechnol Bioeng</addtitle><date>2019-10</date><risdate>2019</risdate><volume>116</volume><issue>10</issue><spage>2503</spage><epage>2513</epage><pages>2503-2513</pages><issn>0006-3592</issn><eissn>1097-0290</eissn><abstract>Interleukin‐2 (IL‐2) is a potent molecule in cancer therapy. Clinical application, however, is limited due to its strong side effects during the treatment. We developed an IL‐2 variant (IL‐2v) immunocytokine to circumvent the drawbacks of the current IL‐2 therapy. During the production of the IL‐2v immunocytokine in Chinese hamster ovary (CHO) cells, molecules with fragmented IL‐2v and therefore reduced cytokine activity can be observed. To control product fragmentation different production process conditions were investigated. By shifting temperature or pH after the cell growth phase to lower values, fragmented species can be reduced from 10% to 12% to about 4%. However, with the adopted process conditions, the effective titer is decreased concomitantly. Moreover, fermentation length and inoculation cell density are parameters to adjust fragmentation and effective titer. A suitable method for efficient process optimization is the design of experiment approach. With this procedure, novel optimal values for temperature, pH value, harvest day, and inoculation cell densities were proposed and tested subsequently. In comparison to the former process, the improved process reduces fragmentation by 66% while keeping the effective titer comparable. In summary, these findings will help to control fragmentation in CHO production processes of different IL‐2v or IL‐2 containing therapeutic proteins.
During manufacturing of a novel IL‐2 variant (IL‐2v) immunocytokine in CHO cells molecules with fragmented IL‐2v moieties were observed. The decrease of the process parameters temperature, pH value, fermentation duration, and inoculation cell density were found to lower fragmentation but also product titer. Using a DoE optimization approach to balance fragmentation and titer, fragmented molecules were reduced by 66% in the improved process compared to the former process while keeping the effective titer comparable.</abstract><cop>United States</cop><pub>Wiley Subscription Services, Inc</pub><pmid>31180133</pmid><doi>10.1002/bit.27090</doi><tpages>11</tpages><orcidid>https://orcid.org/0000-0003-0093-7189</orcidid><orcidid>https://orcid.org/0000-0003-0046-6452</orcidid></addata></record> |
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subjects | Cell density CHO cells Design of experiments Design optimization DoE Fermentation Fragmentation Inoculation Interleukins interleukin‐2 Menopause pH effects proteolytic cleavage Side effects Therapy |
title | Reduction of IL‐2 fragmentation during manufacturing of a novel immunocytokine by DoE process optimization |
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