Effect of PEGylation on the Solution Conformation of Antibody Fragments

Lu, Youming; Harding, Stephen E.; Turner, Alison; Smith, Bryan; Athwal, D. S.; Grossmann, J. Günter; Davis, Kenneth G.; Rowe, Arthur J.

doi:10.1002/jps.21170

Cited by 59 publications

(75 citation statements)

References 49 publications

(77 reference statements)

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“…The size of PEGylated proteins is dominated by the presence of the PEG that is conjugated to the protein . PEG is a random coil polymer with an extended conformation in solution .…”

Section: Resultsmentioning

confidence: 99%

Comparative Study of In Situ Loaded Antibody and PEG‐Fab NIPAAM Gels

Awwad

Al-Shohani

Khaw

et al. 2017

Macromolecular Bioscience

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Hydrogels can potentially prolong the release of a therapeutic protein, especially to treat blinding conditions. One challenge is to ensure that the protein and hydrogel are intimately mixed by better protein entanglement within the hydrogel. N-isopropylacrylamide (NIPAAM) gels are optimized with poly(ethylene glycol) diacrylate (PEDGA) crosslinker in the presence of either bevacizumab or PEG conjugated ranibizumab (PEG -Fab ). The release profiles of the hydrogels are evaluated using an outflow model of the eye, which is previously validated for human clearance of proteins. Release kinetics of in situ loaded bevacizumab-NIPAAM gels displays a prolonged bimodal release profile in phosphate buffered saline compared to bevacizumab loaded into a preformed NIPAAM gel. Bevacizumab release in simulated vitreous from in situ loaded gels is similar to bevacizumab control indicating that diffusion through the vitreous rather than from the gel is rate limiting. Ranibizumab is site-specifically PEGylated by disulfide rebridging conjugation. Prolonged and continuous release is observed with the in situ loaded PEG -Fab -NIPAAM gels compared to PEG -Fab injection (control). Compared to an unmodified protein, there is better mixing due to PEG entanglement and compatibility of PEG -Fab within the NIPAAM-PEDGA hydrogel. These encouraging results suggest that the extended release of PEGylated proteins in the vitreous can be achieved using injectable hydrogels.

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“…The size of PEGylated proteins is dominated by the presence of the PEG that is conjugated to the protein . PEG is a random coil polymer with an extended conformation in solution .…”

Section: Resultsmentioning

confidence: 99%

Comparative Study of In Situ Loaded Antibody and PEG‐Fab NIPAAM Gels

Awwad

Al-Shohani

Khaw

et al. 2017

Macromolecular Bioscience

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“…In general, due to the wide range of sample concentrations and the sensitivity to small fractions of aggregates, AUC is becoming a more widely used tool in the characterization of biopharmaceuticals. AUC-SV is used more and more as an orthogonal tool for the analysis of soluble protein aggregates within the pharmaceutical industry 37,141,147,153 and adequate crosscorrelation between AUC and SEC has been reported. 141,143 However, although better precision of AUC has been achieved lately by equipment improvements 148 reproducibility and precision can still be considered lower than for SEC.…”

Section: Analytical Ultracentrifugationmentioning

confidence: 99%

Protein aggregation: Pathways, induction factors and analysis

Mahler

Frieß

Grauschopf

et al. 2009

Journal of Pharmaceutical Sciences

764

635

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“…Based on prior work[61–63], we hypothesized that in the active tetrameric complex the PEG would be stretched slightly from its normal random coil configuration. As a guide we considered Flory’s Radius (R F = aN 0.6 ) where a = monomer unit length and N = # monomers or degree of polymerization, to estimate the random coil length of various PEGs.…”

Section: Resultsmentioning

confidence: 99%

Fibroblast growth factor 2 dimer with superagonist in vitro activity improves granulation tissue formation during wound healing

et al. 2016

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Site-specific chemical dimerization of fibroblast growth factor 2 (FGF2) with the optimal linker length resulted in a FGF2 homodimer with improved granulation tissue formation and blood vessel formation at exceptionally low concentrations. Homodimers of FGF2 were synthesized through site-specific linkages to both ends of different molecular weight poly(ethylene glycols) (PEGs). The optimal linker length was determined by screening dimer-induced metabolic activity of human dermal fibroblasts and found to be that closest to the inter-cysteine distance, 70 Å, corresponding to 2 kDa PEG. A straightforward analysis of the kinetics of second ligand binding as a function of tether length showed that, as the polymerization index (the number of monomer repeat units in the polymer, N) of the tether decreases, the mean time for second ligand capture decreases as ~N3/2, leading to an enhancement of the number of doubly bound ligands in steady-state for a given (tethered) ligand concentration. FGF2-PEG2k-FGF2 induced greater fibroblast metabolic activity than FGF2 alone, all other dimers, and all monoconjugates, at each concentration tested, with the greatest difference observed at low (0.1 ng/mL) concentration. FGF2-PEG2k-FGF2 further exhibited superior activity compared to FGF2 for both metabolic activity and migration in human umbilical vein endothelial cells, as well as improved angiogenesis in a coculture model in vitro. Efficacy in an in vivo wound healing model was assessed in diabetic mice. FGF2-PEG2k-FGF2 increased granulation tissue and blood vessel density in the wound bed compared to FGF2. The results suggest that this rationally designed construct may be useful for improving the fibroblast matrix formation and angiogenesis in chronic wound healing.

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Effect of PEGylation on the Solution Conformation of Antibody Fragments

Cited by 59 publications

References 49 publications

Comparative Study of In Situ Loaded Antibody and PEG‐Fab NIPAAM Gels

Comparative Study of In Situ Loaded Antibody and PEG‐Fab NIPAAM Gels

Protein aggregation: Pathways, induction factors and analysis

Fibroblast growth factor 2 dimer with superagonist in vitro activity improves granulation tissue formation during wound healing

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