{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,6,5]],"date-time":"2026-06-05T13:51:38Z","timestamp":1780667498073,"version":"3.54.1"},"reference-count":21,"publisher":"Wiley","issue":"2","license":[{"start":{"date-parts":[[2024,9,1]],"date-time":"2024-09-01T00:00:00Z","timestamp":1725148800000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":["onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["Proteins"],"published-print":{"date-parts":[[2025,2]]},"abstract":"<jats:title>ABSTRACT<\/jats:title><jats:p>Inhibition of CD95\/Fas activation is currently under clinical investigation as a therapy for glioblastoma multiforme and preclinical studies suggest that disruption of the CD95\u2013CD95L interaction could also be a strategy to treat inflammatory and neurodegenerative disorders. Besides neutralizing anti\u2010CD95L\/FasL antibodies, mainly CD95ed\u2010Fc, a dimeric Fc fusion protein of the extracellular domain of CD95 (CD95ed), is used to prevent CD95 activation. In view of the fact that full CD95 activation requires CD95L\u2010induced CD95 trimerization and clustering of the resulting liganded CD95 trimers, we investigated whether fusion proteins of the extracellular domain of CD95 with a higher valency than CD95ed\u2010Fc have an improved CD95L\u2010neutralization capacity. We evaluated an IgG1(N297A)\u2010based tetravalent CD95ed fusion protein which was obtained by replacing the variable domains of IgG1(N297A) with CD95ed (CD95ed\u2010IgG1(N297A)) and a hexavalent variant obtained by fusion of CD95ed with a TNC\u2010Fc(DANA) scaffold (CD95ed\u2010TNC\u2010Fc(DANA)) promoting hexamerization. The established N297A and DANA mutations were used to minimize Fc\u03b3R binding of the constructs under maintenance of neonatal Fc receptor (FcRn) binding. Size exclusion high\u2010performance liquid chromatography indicated effective assembly of CD95ed\u2010IgG1(N297A). More important, CD95ed\u2010IgG1(N297A) was much more efficient than CD95ed\u2010Fc in protecting cells from cell death induction by human and murine CD95L. Surprisingly, despite its hexavalent structure, CD95ed\u2010TNC\u2010Fc(DANA) displayed an at best minor improvement of the capacity to neutralize CD95L suggesting that besides valency, other factors, such as spatial organization and agility of the CD95ed domains, play also a role in neutralization of CD95L trimers by CD95ed fusion proteins. More studies are now required to evaluate the superior CD95L\u2010neutralizing capacity of CD95ed\u2010IgG1(N297A) in vivo.<\/jats:p>","DOI":"10.1002\/prot.26741","type":"journal-article","created":{"date-parts":[[2024,9,2]],"date-time":"2024-09-02T05:29:34Z","timestamp":1725254974000},"page":"441-451","update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":2,"title":["A Novel Tetravalent <scp>CD95<\/scp>\/Fas Fusion Protein With Superior <scp>CD95L<\/scp>\/<scp>FasL<\/scp> Antagonism"],"prefix":"10.1002","volume":"93","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-6369-4973","authenticated-orcid":false,"given":"Isabell","family":"Lang","sequence":"first","affiliation":[{"name":"Department of Internal Medicine II, Division of Molecular Internal Medicine University Hospital W\u00fcrzburg  Wurzburg Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Oliver","family":"Paulus","sequence":"additional","affiliation":[{"name":"Department of Internal Medicine II, Division of Molecular Internal Medicine University Hospital W\u00fcrzburg  Wurzburg Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Olena","family":"Zaitseva","sequence":"additional","affiliation":[{"name":"Department of Internal Medicine II, Division of Molecular Internal Medicine University Hospital W\u00fcrzburg  Wurzburg Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Harald","family":"Wajant","sequence":"additional","affiliation":[{"name":"Department of Internal Medicine II, Division of Molecular Internal Medicine University Hospital W\u00fcrzburg  Wurzburg 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