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Pàgina inicial > Articles > Articles publicats > Site-directed cysteine coupling of disulfide-containing non-antibody carrier proteins (THIOCAPs) |
Data: | 2023 |
Resum: | The development of a new generation of non-antibody protein drug delivery systems requires site-directed conjugation strategies to produce homogeneous, reproducible and scalable nanomedicines. For that, the genetic addition of cysteine residues into solvent-exposed positions allows the thiol-mediated cysteine coupling of therapeutic drugs into protein-based nanocarriers. However, the high reactivity of unpaired cysteine residues usually reduces protein stability, consequently imposing the use of more methodologically demanding purification procedures. This is especially relevant for disulfide-containing nanocarriers, as previously observed in THIOMABs. Moreover, although many protein scaffolds and targeting ligands are also rich in disulfide bridges, the use of these methodologies over emerging non-antibody carrier proteins has been completely neglected. Here, we report the development of a simple and straightforward procedure for a one-step production and site-directed cysteine conjugation of disulfide-containing non-antibody thiolated carrier proteins (THIOCAPs). This method is validated in a fluorescent C-X-C chemokine receptor 4 (CXCR4)-targeted multivalent nano-carrier containing two intramolecular disulfide bridges and one reactive cysteine residue strategically placed into a solvent-exposed position (THIO-T22-GFP-H6) for drug conjugation and in a humanized alternative intended for clinical applications (T22-HSNBT-H6). Thus, we produce very stable, homogeneous and fully functional antitumoral nanoconjugates (THIO-T22-GFP-H6-MMAE and T22-HSNBT-H6-MMAE) that selectively eliminate target cancer cells via CXCR4-receptor. Altogether, the developed methodology appears as a powerful tool for the rational engineering of emerging non-antibody, cell-targeted protein nanocarriers that contain disulfide bridges together with a solvent-exposed reactive cysteine (THIOCAP). This should pave the way for the development of a new generation of stable, homogeneous and efficient nanomedicines. |
Ajuts: | Instituto de Salud Carlos III PI20/00400 Agencia Estatal de Investigación PID2019-105416RB-I00 Instituto de Salud Carlos III PI21/00150 Agència de Gestió d'Ajuts Universitaris i de Recerca 2021/SGR-01140 Agència de Gestió d'Ajuts Universitaris i de Recerca 2021/SGR-00092 Instituto de Salud Carlos III CB06/01/1031 Ministerio de Sanidad y Consumo CB06/01/0014 Instituto de Salud Carlos III CP19/00028 Instituto de Salud Carlos III FI21/00012 Ministerio de Ciencia, Innovación y Universidades FPU18/04615 |
Nota: | Altres ajuts: CERCA Programme/Generalitat de Catalunya ; Asociación Española Contra el Cáncer (POSTD20070ALBA) ; Icrea Academia award |
Drets: | Tots els drets reservats. |
Llengua: | Anglès |
Document: | Article ; recerca ; Versió acceptada per publicar |
Matèria: | Cysteine coupling ; Disulfide-bonds ; Nanoconjugates ; Nanomedicine ; Protein carriers ; Thiocap ; SDG 3 - Good Health and Well-being |
Publicat a: | Science China Materials, Vol. 66 (October 2023) , p. 4109-4120, ISSN 2199-4501 |
Obra relacionada: | Rueda, Ariana; Mendoza, Julian Ignacio; Alba-Castellon, Lorena; Parladé Molist, Eloi; Voltà-Durán, Eric; Páez, David; Aviñó, Ana; Eritja, Ramon; Vázquez Gómez, Esther; Villaverde Corrales, Antonio; Mangues, Ramon; Unzueta Elorza, Ugutz, 2024, "Raw data to support the manuscript "Site-directed cysteine coupling of disulfide-containing non-antibody carrier proteins (THIOCAPs)"", CORA.Repositori de Dades de Recerca, V1 https://doi.org/10.34810/data1090 |
Disponible a partir de: 2024-10-31 Postprint |
Disponible a partir de: 2024-10-31 Supporting information |