{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,5,20]],"date-time":"2026-05-20T19:54:46Z","timestamp":1779306886315,"version":"3.51.4"},"reference-count":48,"publisher":"Wiley","issue":"1","license":[{"start":{"date-parts":[[2025,1,20]],"date-time":"2025-01-20T00:00:00Z","timestamp":1737331200000},"content-version":"am","delay-in-days":19,"URL":"http:\/\/creativecommons.org\/licenses\/by-nc\/4.0\/"},{"start":{"date-parts":[[2025,1,20]],"date-time":"2025-01-20T00:00:00Z","timestamp":1737331200000},"content-version":"vor","delay-in-days":19,"URL":"http:\/\/creativecommons.org\/licenses\/by-nc\/4.0\/"}],"funder":[{"DOI":"10.13039\/100000001","name":"National Science Foundation","doi-asserted-by":"publisher","award":["1624698"],"award-info":[{"award-number":["1624698"]}],"id":[{"id":"10.13039\/100000001","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000001","name":"National Science Foundation","doi-asserted-by":"publisher","award":["2100502"],"award-info":[{"award-number":["2100502"]}],"id":[{"id":"10.13039\/100000001","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Biotechnology Journal"],"published-print":{"date-parts":[[2025,1]]},"abstract":"<jats:title>ABSTRACT<\/jats:title><jats:p>The implementation of site\u2010specific integration (SSI) systems in Chinese hamster ovary (CHO) cells for the production of monoclonal antibodies (mAbs) can alleviate concerns associated with production instability and reduce cell line development timelines. SSI cell line performance is driven by the interaction between genomic integration location, clonal background, and the transgene expression cassette, requiring optimization of all three parameters to maximize productivity. Systematic comparison of these parameters has been hindered by SSI platforms involving low\u2010throughput enrichment strategies, such as cell sorting. This study presents a recombinase\u2010mediated cassette exchange (RMCE)\u2010capable SSI system that uses only chemical selection to enrich for transgene\u2010expressing RMCE pools in less than one month. The system was used to compare eight mAb expression cassettes containing two novel genetic regulatory elements, the <jats:italic>Azin1<\/jats:italic> CpG island and the Piggybac transposase 5\u2019 terminal repeat, in various orientations to improve the expression of two therapeutic mAbs from two genomic loci. Similar patterns of productivity and mRNA expression were observed across sites and mAbs, and the best performing cassette universally increased mAb productivity by 7\u2010 to 11\u2010fold. This flexible system allows for higher\u2010throughput comparison of expression cassettes from a consistent clonal and transcriptional background to optimize RMCE\u2010derived cell lines for industrial production of mAbs.<\/jats:p>","DOI":"10.1002\/biot.202400520","type":"journal-article","created":{"date-parts":[[2025,1,21]],"date-time":"2025-01-21T01:40:24Z","timestamp":1737423624000},"source":"Crossref","is-referenced-by-count":3,"title":["A Flexible Hybrid Site\u2010Specific Integration\u2010Based Expression System in CHO Cells for Higher\u2010Throughput Evaluation of Monoclonal Antibody Expression Cassettes"],"prefix":"10.1002","volume":"20","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-9071-9363","authenticated-orcid":false,"given":"Alana C.","family":"Szkodny","sequence":"first","affiliation":[{"name":"Department of Chemical and Biomolecular Engineering University of Delaware  Newark Delaware USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0908-2981","authenticated-orcid":false,"given":"Kelvin H.","family":"Lee","sequence":"additional","affiliation":[{"name":"Department of Chemical and Biomolecular Engineering University of Delaware  Newark Delaware USA"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"311","published-online":{"date-parts":[[2025,1,20]]},"reference":[{"key":"e_1_2_10_2_1","doi-asserted-by":"publisher","DOI":"10.1002\/btpr.436"},{"key":"e_1_2_10_3_1","doi-asserted-by":"publisher","DOI":"10.1007\/978-3-319-97110-0"},{"key":"e_1_2_10_4_1","doi-asserted-by":"publisher","DOI":"10.1002\/bit.10517"},{"key":"e_1_2_10_5_1","doi-asserted-by":"publisher","DOI":"10.1002\/bit.23189"},{"key":"e_1_2_10_6_1","doi-asserted-by":"publisher","DOI":"10.1038\/nbt1026"},{"key":"e_1_2_10_7_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.coche.2018.09.011"},{"key":"e_1_2_10_8_1","doi-asserted-by":"publisher","DOI":"10.1002\/biot.201500082"},{"key":"e_1_2_10_9_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.gene.2012.11.016"},{"key":"e_1_2_10_10_1","doi-asserted-by":"publisher","DOI":"10.1021\/acssynbio.8b00140"},{"key":"e_1_2_10_11_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.tibtech.2019.02.007"},{"key":"e_1_2_10_12_1","doi-asserted-by":"publisher","DOI":"10.1080\/07388551.2020.1768043"},{"key":"e_1_2_10_13_1","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gky216"},{"key":"e_1_2_10_14_1","doi-asserted-by":"publisher","DOI":"10.1002\/bit.28390"},{"key":"e_1_2_10_15_1","doi-asserted-by":"publisher","DOI":"10.1002\/btpr.3140"},{"key":"e_1_2_10_16_1","doi-asserted-by":"publisher","DOI":"10.1002\/btpr.2175"},{"key":"e_1_2_10_17_1","doi-asserted-by":"publisher","DOI":"10.1021\/acssynbio.0c00519"},{"key":"e_1_2_10_18_1","doi-asserted-by":"publisher","DOI":"10.1002\/btpr.3254"},{"key":"e_1_2_10_19_1","doi-asserted-by":"publisher","DOI":"10.1021\/acssynbio.0c00643"},{"key":"e_1_2_10_20_1","doi-asserted-by":"publisher","DOI":"10.1021\/acssynbio.8b00453"},{"key":"e_1_2_10_21_1","doi-asserted-by":"publisher","DOI":"10.1021\/acssynbio.0c00322"},{"key":"e_1_2_10_22_1","doi-asserted-by":"publisher","DOI":"10.1002\/btpr.2967"},{"key":"e_1_2_10_23_1","doi-asserted-by":"publisher","DOI":"10.1002\/btpr.3137"},{"key":"e_1_2_10_24_1","unstructured":"G.Chen R.Babb andJ. P.Fandl \u201cEnhanced Expression and Stability Regions (United States Patent US7771997B2) \u201d Google Patents 2010 https:\/\/patents.google.com\/patent\/US7771997B2\/en."},{"key":"e_1_2_10_25_1","unstructured":"C. K. D.Ng Y. G.Crawford A.Shen et\u00a0al. \u201cTargeted Integration of Nucleic Acids (World Intellectual Property Organization Patent WO2019126634A2) \u201d Google Patents 2019 https:\/\/patents.google.com\/patent\/WO2019126634A2\/en."},{"key":"e_1_2_10_26_1","unstructured":"Y.Shen D.Burakov G.Chen andJ. P.Fandl \u201cCHO Integration Sites and Uses Thereof (United States Patent US9816110B2) \u201d Google Patents 2017 https:\/\/patents.google.com\/patent\/US9816110B2\/en."},{"key":"e_1_2_10_27_1","doi-asserted-by":"publisher","DOI":"10.1042\/BJ20060874"},{"key":"e_1_2_10_28_1","doi-asserted-by":"publisher","DOI":"10.1002\/biot.202000057"},{"key":"e_1_2_10_29_1","doi-asserted-by":"publisher","DOI":"10.1038\/s41467\u2010019\u201013897\u20106"},{"key":"e_1_2_10_30_1","doi-asserted-by":"publisher","DOI":"10.1002\/bit.28393"},{"key":"e_1_2_10_31_1","doi-asserted-by":"publisher","DOI":"10.1002\/bit.27432"},{"key":"e_1_2_10_32_1","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.273.52.34970"},{"key":"e_1_2_10_33_1","doi-asserted-by":"publisher","DOI":"10.1016\/S1097\u20102765(03)00444\u20101"},{"key":"e_1_2_10_34_1","doi-asserted-by":"publisher","DOI":"10.1002\/bit.26268"},{"key":"e_1_2_10_35_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.jmb.2009.05.012"},{"key":"e_1_2_10_36_1","doi-asserted-by":"publisher","DOI":"10.1038\/nbt1362"},{"key":"e_1_2_10_37_1","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.94.8.3789"},{"key":"e_1_2_10_38_1","unstructured":"W. C.HilliardandK. H.Lee \"Chinese Hamster Ovary Cell Genomic Hotspots for Recombinant Protein Production (World Intellectual Property Organization Patent WO2024073464) \" WIPO 2024 https:\/\/patents.google.com\/patent\/WO2024073464A2\/en."},{"key":"e_1_2_10_39_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.ymben.2012.12.003"},{"key":"e_1_2_10_40_1","doi-asserted-by":"publisher","DOI":"10.1002\/bit.22126"},{"key":"e_1_2_10_41_1","doi-asserted-by":"publisher","DOI":"10.1002\/bit.22769"},{"key":"e_1_2_10_42_1","doi-asserted-by":"publisher","DOI":"10.1021\/bp049780w"},{"key":"e_1_2_10_43_1","doi-asserted-by":"publisher","DOI":"10.1093\/protein\/gzm038"},{"key":"e_1_2_10_44_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.jbiotec.2013.03.019"},{"key":"e_1_2_10_45_1","doi-asserted-by":"publisher","DOI":"10.1002\/bit.22946"},{"key":"e_1_2_10_46_1","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pone.0008432"},{"key":"e_1_2_10_47_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.jbc.2021.100838"},{"key":"e_1_2_10_48_1","doi-asserted-by":"publisher","DOI":"10.1002\/biot.201700175"},{"key":"e_1_2_10_49_1","doi-asserted-by":"publisher","DOI":"10.1002\/bit.26388"}],"container-title":["Biotechnology Journal"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/analyticalsciencejournals.onlinelibrary.wiley.com\/doi\/pdf\/10.1002\/biot.202400520","content-type":"application\/pdf","content-version":"vor","intended-application":"syndication"},{"URL":"https:\/\/analyticalsciencejournals.onlinelibrary.wiley.com\/doi\/pdf\/10.1002\/biot.202400520","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,8]],"date-time":"2025-10-08T14:02:41Z","timestamp":1759932161000},"score":1,"resource":{"primary":{"URL":"https:\/\/analyticalsciencejournals.onlinelibrary.wiley.com\/doi\/10.1002\/biot.202400520"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,1]]},"references-count":48,"journal-issue":{"issue":"1","published-print":{"date-parts":[[2025,1]]}},"alternative-id":["10.1002\/biot.202400520"],"URL":"https:\/\/doi.org\/10.1002\/biot.202400520","archive":["Portico"],"relation":{},"ISSN":["1860-6768","1860-7314"],"issn-type":[{"value":"1860-6768","type":"print"},{"value":"1860-7314","type":"electronic"}],"subject":[],"published":{"date-parts":[[2025,1]]},"article-number":"e202400520"}}