{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,1]],"date-time":"2026-10-01T09:58:28Z","timestamp":1790848708482,"version":"4.1.0"},"reference-count":55,"publisher":"Oxford University Press (OUP)","issue":"4","license":[{"start":{"date-parts":[[2026,7,31]],"date-time":"2026-07-31T00:00:00Z","timestamp":1785456000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by-nc\/4.0\/"}],"funder":[{"name":"Yantai Science and Technology Innovation Development Plan Projects","award":["2023ZDCX030"],"award-info":[{"award-number":["2023ZDCX030"]}]},{"name":"Shandong Boan biotechnology Co., Ltd."},{"name":"Intramural Joint Program Fund of State Key Laboratory of Microbial Technology","award":["SKLMTIJP-2025-03"],"award-info":[{"award-number":["SKLMTIJP-2025-03"]}],"id":[{"id":"https:\/\/ror.org\/03x08qn04","id-type":"ROR","asserted-by":"crossref"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"published-print":{"date-parts":[[2026,10,1]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                  <jats:sec>\n                    <jats:title>Background<\/jats:title>\n                    <jats:p>Antibody\u2013drug conjugates (ADCs) with homogeneous drug-to-antibody ratios (DARs) are increasingly favored for their improved physicochemical and pharmacokinetic properties; however, strategies to reliably generate homogeneous DAR6 ADCs remain limited.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Methods<\/jats:title>\n                    <jats:p>In this study, we investigate hinge-region cysteine engineering as an approach to constrain conjugation stoichiometry, using a potent biparatopic anti\u2013c-Met antibody as a model. Various biological and physicochemical assays were used for its characterization.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Results<\/jats:title>\n                    <jats:p>We show that substitution of a hinge-region cysteine within a CPPC (Cysteine - Proline - Proline - Cysteine) motif reduces the number of accessible conjugation sites, allowing reproducible generation of homogeneous DAR6 ADCs. During characterization of Ser- or Thr-substituted hinge variants, we observed additional mass species consistent with mucin-type O-linked glycosylation. Site-specific mass spectrometry analyses localized these modifications to a serine residue within hinge-region and identified the core structures of the O-glycans. Analysis of additional antibodies carrying analogous mutations demonstrated that hinge-associated O-glycosylation is not restricted to a single antibody scaffold.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Conclusions<\/jats:title>\n                    <jats:p>These results demonstrate that hinge-region cysteine engineering can serve as a rational and adaptable approach for DAR6 ADC development while highlighting the need for case-by-case evaluation of associated post-translational modifications during antibody engineering.<\/jats:p>\n                  <\/jats:sec>","DOI":"10.1093\/abt\/tbag040","type":"journal-article","created":{"date-parts":[[2026,7,30]],"date-time":"2026-07-30T11:54:34Z","timestamp":1785412474000},"page":"521-534","source":"Crossref","is-referenced-by-count":0,"title":["Hinge-region Cys-to-Ser mutation enables homogeneous DAR6 anti-c-Met biparatopic ADCs with tolerable O-glycosylation"],"prefix":"10.1093","volume":"9","author":[{"given":"Deyong","family":"Song","sequence":"first","affiliation":[{"name":"National Glycoengineering Research Center, Shandong Center of Technology Innovation for Carbohydrate, Shandong Key Laboratory of Carbohydrate and Carbohydrate-Conjugate Drugs, and State Key Laboratory of Microbial Technology, Shandong University , Qingdao 266237 ,","place":["China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Zhenduo","family":"Shen","sequence":"additional","affiliation":[{"name":"National Glycoengineering Research Center, Shandong Center of Technology Innovation for Carbohydrate, Shandong Key Laboratory of Carbohydrate and Carbohydrate-Conjugate Drugs, and State Key Laboratory of Microbial Technology, Shandong University , Qingdao 266237 ,","place":["China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xiaolin","family":"Zhu","sequence":"additional","affiliation":[{"name":"Research and Development Center, Shandong Boan Biotechnology Co., Ltd , Yantai 264670 ,","place":["China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Haifeng","family":"Zhao","sequence":"additional","affiliation":[{"name":"Research and Development Center, Shandong Boan Biotechnology Co., Ltd , Yantai 264670 ,","place":["China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yingdi","family":"Wang","sequence":"additional","affiliation":[{"name":"Research and Development Center, Shandong Boan Biotechnology Co., Ltd , Yantai 264670 ,","place":["China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Junmei","family":"Zhang","sequence":"additional","affiliation":[{"name":"Research and Development Center, Shandong Boan Biotechnology Co., Ltd , Yantai 264670 ,","place":["China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Rui","family":"Li","sequence":"additional","affiliation":[{"name":"Research and Development Center, Shandong Boan Biotechnology Co., Ltd , Yantai 264670 ,","place":["China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hongguang","family":"Xu","sequence":"additional","affiliation":[{"name":"Research and Development Center, Shandong Boan Biotechnology Co., Ltd , Yantai 264670 ,","place":["China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Zengjin","family":"Cheng","sequence":"additional","affiliation":[{"name":"Research and Development Center, Shandong Boan Biotechnology Co., Ltd , Yantai 264670 ,","place":["China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xiaoju","family":"Ma","sequence":"additional","affiliation":[{"name":"Research and Development Center, Shandong Boan Biotechnology Co., Ltd , Yantai 264670 ,","place":["China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Haixia","family":"Lin","sequence":"additional","affiliation":[{"name":"Research and Development Center, Shandong Boan Biotechnology Co., Ltd , Yantai 264670 ,","place":["China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Changlin","family":"Dou","sequence":"additional","affiliation":[{"name":"Research and Development Center, Shandong Boan Biotechnology Co., Ltd , Yantai 264670 ,","place":["China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4905-8321","authenticated-orcid":false,"given":"Min","family":"Xiao","sequence":"additional","affiliation":[{"name":"National Glycoengineering Research Center, Shandong Center of Technology Innovation for Carbohydrate, Shandong Key Laboratory of Carbohydrate and Carbohydrate-Conjugate Drugs, and State Key Laboratory of Microbial Technology, Shandong University , Qingdao 266237 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