Polymer Conjugation
Following the initial positive results of conjugation of polymers to proteins based on simple conjugation methods, the chemistry has continued to experience advances with the development of new selective approaches of conjugation, some based on the exploitation of enzymes as tools to couple polymers and proteins or on genetic methods (e.g., thiol conjugation on an inserted cysteine or genetic code expansion with Amber codon technique). Among different techniques PEGylation has become the gold standard in this field. Protein PEGylation is primarily studied for prolonging the half-life of proteins by decreasing their body elimination, either through a reduced kidney clearance or a reduced liver clearance. Furthermore, PEGylation can also reduce the immunogenicity of non-human proteins by shielding the protein to the immune system.
In this field we investigated several polymers beside PEG, for example polyoxazolines, hyaluronic acid, polysialic acid, polyglutamic acids, etc.
To further extend the potentialities of polymer conjugation we thoroughly investigated the enzymatic conjugation approaches based on Transglutaminase. In nature, this enzyme catalyses the transfer of the acyl donor, the side chain of a methionine in a protein, to the amino donor, the side chain of a lysine in a second protein, thus forming a cross-link. The amino donor can be substituted by an amino group of a polymer for generating easily and rapidly homogenous conjugates in which the polymer is linked to single glutamines. We also demonstrated that transglutaminase can targeted the lysines of a protein when the acyl donor is the polymer, generating lysine coupled polymer-protein conjugates with the great advantage of selectivity of the transglutaminase.
Key Publications
2021 - Development of Vaccines
Dalla Pietà A, Carpanese D, Grigoletto A, Tosi A, Dalla Santa S, Pedersen GK, Christensen D, Meléndez-Alafort L, Barbieri V, De Benedictis P, Pasut G, Montagner IM, Rosato A. Hyaluronan is a natural and effective immunological adjuvant for protein-based vaccines. Cell Mol Immunol. 2021;18(5):1197-1210
2021 - Cystic Fibrosis Treatment
Delfino D, Mori G, Rivetti C, Grigoletto A, Bizzotto G, Cavozzi C, Malatesta M, Cavazzini D, Pasut G, Percudani R. Actin-Resistant DNase1L2 as a Potential Therapeutics for CF Lung Disease. Biomolecules. 2021;11(3):410. doi: 10.3390/biom11030410.
2020 - New Polymers for Protein Conjugation
Maso K, Grigoletto A, Raccagni L, Bellini M, Marigo I, Ingangi V, Suzuki A, Hirai M, Kamiya M, Yoshioka H, Pasut G. Poly(L-glutamic acid)-co-poly(ethylene glycol) block copolymers for protein conjugation. J Control Release. 2020;324:228-237. doi: 10.1016/j.jconrel.2020.05.015
2017 - PEGylation via Immobilized Transglutaminase
Grigoletto A, Mero A, Yoshioka H, Schiavon O, Pasut G. Covalent immobilisation of transglutaminase: stability and applications in protein pegylation. J Drug Target. 2017;25:856-864. doi: 10.1080/1061186X.2017.1363211
2016 - Enzymatic PEGylation (TGase)
Spolaore B, Raboni S, Satwekar AA, Grigoletto A, Mero A, Montagner IM, Rosato A, Pasut G, Fontana A. Site-specific transglutaminase-mediated conjugation of interferon α-2b at glutamine or lysine residues. Bioconjug Chem. 2016;27(11):2695-2706. doi: 10.1021/acs.bioconjchem.6b00468.
2016 - Inverse Transglutaminase PEGylation
Mero A, Grigoletto A, Maso K, Yoshioka H, Rosato A, Pasut G. Site-selective enzymatic chemistry for polymer conjugation to protein lysine residues: pegylation of G-CSF at lysine-41. Pol Chem. 2016;7:6545-6553. doi: 10.1039/C6PY01616B
2016 - Chemical and Enzymatic PEGylation
Grigoletto A, Mero A, Zanusso I, Schiavon O, Pasut G. Chemical and Enzymatic Site Specific pegylation of hGH: The stability and in vivo activity of PEG-N-terminal-hGH and PEG-Gln141-hGH conjugates. Macromol Biosci. 2016;16(1):50-6. doi: 10.1002/mabi.201500282.
2014 - Hyaluronic Acid-Salmon Calcitonin
Mero A, Campisi M, Favero M, Barbera C, Secchieri C, Dayer JM, Goldring MB, Goldring SR, Pasut G. A hyaluronic acid-salmon calcitonin conjugate for the local treatment of osteoarthritis: Chondro-protective effect in a rabbit model of early OA. J Control Release. 2014;187:30-38. doi: 10.1016/j.jconrel.2014.05.008
2013 - Hyaluronic Acid as Protein Carrier
Mero A, Pasqualin M, Campisi M, Renier D, Pasut G. Conjugation of hyaluronan to proteins. Carbohydr. Polym. 2013;92:2163-2170. doi: 10.1016/j.carbpol.2012.11.090.
2013 - Enzymatic PEGylation (TGase)
Da Silva Freitas D, Mero A, Pasut G. Chemical and enzymatic site specific pegylation of hGH. Bioconjug Chem. 2013:24;456-463. doi: 10.1021/bc300594y
2012 - Poly(2-ethyl 2-oxazoline)
Mero A, Fang Z, Pasut G, Veronese FM, Viegas TX. Selective conjugation of poly (2-ethyl 2-oxazoline) to granulocyte colony stimulating factor. J Control Release. 2012;159:353-361. doi: 10.1016/j.jconrel.2012.02.025
2011 - Enzymatic PEGylation (TGase)
Mero A, Schiavon M, Veronese FM, Pasut G. A new method to increase selectivity of transglutaminase mediated PEGylation of salmon calcitonin and human growth hormone. J Control Release. 2011;154:27-34. doi: 10.1016/j.jconrel.2011.04.024
