PEGylation to Improve Protein Stability During Melt Processing

Parker, Lee D.; Towslee, Jenna Harris; Maia, João M.; Pokorski, Jonathan K.

doi:10.1002/mabi.201500143

Cited by 25 publications

(25 citation statements)

References 47 publications

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“…Previous DSC studies of lysozyme, glycinin, and human growth hormone in the solid-state have indicated protein denaturation over temperature ranges of 180–200 °C. 38,43,44 The higher denaturation temperature observed with lyophilized Q β was likely due to strong intermolecular attractions associated with dimer stability. The DSC study of Q β provided insight into the denaturation process during heating in the solid state and ensured that no major denaturation processes occurred in the processing window of 80–100 °C typically used for melt-encapsulation of proteins with PLGA.…”

Section: Resultsmentioning

confidence: 99%

Biodegradable Viral Nanoparticle/Polymer Implants Prepared via Melt-Processing

et al. 2017

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Viral nanoparticles have been utilized as a platform for vaccine development and are a versatile system for the display of antigenic epitopes for a variety of disease states. However, the induction of a clinically relevant immune response often requires multiple injections over an extended period of time, limiting patient compliance. Polymeric systems to deliver proteinaceous materials have been extensively researched to provide sustained release, which would limit administration to a single dose. Melt-processing is an emerging manufacturing method that has been utilized to create polymeric materials laden with proteins as an alternative to typical solvent-based production methods. Melt-processing is advantageous because it is continuous, solvent-free, and 100% of the therapeutic protein is encapsulated. In this study, we utilized melt-encapsulation to fabricate viral nanoparticle laden polymeric materials that effectively deliver intact particles and generate carrier specific antibodies in vivo. The effects of initial processing and postprocessing on particle integrity and aggregation were studied to develop processing windows for scale-up and the creation of more complex materials. The dispersion of particles within the PLGA matrix was studied, and the effect of additives and loading level on the release profile was determined. Overall, melt-encapsulation was found to be an effective method to produce composite materials that can deliver viral nanoparticles over an extended period and elicit an immune response comparable to typical administration schedules.

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Section: Resultsmentioning

confidence: 99%

Biodegradable Viral Nanoparticle/Polymer Implants Prepared via Melt-Processing

et al. 2017

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“…A2 (A) and (B)), which we have previously assigned to total denaturation of the protein secondary structure (Lee et al, 2015). In order to analyze whether exposure to higher temperatures over time causes thermal denaturation, we incubated lysozyme and GOx in the DSC at 42 °C for 40 min (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…In our previous work, we determined the aggregation state of lysozyme within PLGA after HME and found that the retained enzymatic activity of lysozyme was directly related to the size of protein aggregates observed within the matrix (Lee et al, 2015), indicating that the retained activity is dependent on protein dispersion. Furthermore, the aggregation state directly correlated to the release profile of lysozyme from the HME prepared materials.…”

Section: Introductionmentioning

confidence: 99%

“…This effect can be mitigated via chemical modification of the protein, for example the covalent attachment of a hydrophobic dye or the polymer polyethylene glycol (PEG) (Lee et al, 2015). However, the reaction products can be difficult to separate, require extensive filtration or chromatography to remove unreacted components, and result in substantial purification loss (Freitas and Abrahão-Neto, 2010; Lee et al, 2003).…”

Section: Introductionmentioning

confidence: 99%

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Milling solid proteins to enhance activity after melt-encapsulation

Lee

Maia

Pokorski

2017

International Journal of Pharmaceutics

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Polymeric systems for the immobilization and delivery of proteins have been extensively used for therapeutic and catalytic applications. While most devices have been created via solution based methods, hot melt extrusion (HME) has emerged as an alternative due to the high encapsulation efficiencies and solvent-free nature of the process. HME requires high temperatures and mechanical stresses that can result in protein aggregation and denaturation, but additives and chemical modifications have been explored to mitigate these effects. This study explores the use of solid-state ball milling to decrease protein particle size before encapsulation within poly (lactic-co-glycolic acid) (PLGA) via HME. The impact of milling on particle dispersion, retained enzymatic activity, secondary structure stability, and release was explored for lysozyme, glucose oxidase, and the virus-like particle derived from Qβ to fully understand the impact of milling on protein systems with different sizes and complexities. The results of this study describe the utility of milling to further increase the stability of protein/polymer systems prepared via HME.

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“…Besides, it has been demonstrated that grafting of PEG molecules onto lysozymes increased their thermal stability retaining their activity up to 90%. 40 PEG chains grafted to the protein, therefore, protect it from being denatured by heat, keeping its chemical structure intact, and preserving its biological activity. Lysozyme has similar size and structure to RNase A.…”

Section: B Heat Transfer Distribution Inside the Microchannelmentioning

confidence: 99%

Modelling of electrokinetic phenomena for capture of PEGylated ribonuclease A in a microdevice with insulating structures

et al. 2016

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Synthesis of PEGylated proteins results in a mixture of protein-polyethylene glycol (PEG) conjugates and the unreacted native protein. From a ribonuclease A (RNase A) PEGylation reaction, mono-PEGylated RNase A (mono-PEG RNase A) has proven therapeutic effects against cancer, reason for which there is an interest in isolating it from the rest of the reaction products. Experimental trapping of PEGylated RNase A inside an electrokinetically driven microfluidic device has been previously demonstrated. Now, from a theoretical point of view, we have studied the electrokinetic phenomena involved in the dielectrophoretic streaming of the native RNase A protein and the trapping of the mono-PEG RNase A inside a microfluidic channel. To accomplish this, we used two 3D computational models, a sphere and an ellipse, adapted to each protein. The effect of temperature on parameters related to trapping was also studied. A temperature increase showed to rise the electric and thermal conductivities of the suspending solution, hindering dielectrophoretic trapping. In contrast, the dynamic viscosity of the suspending solution decreased as the temperature rose, favoring the dielectrophoretic manipulation of the proteins. Also, our models were able to predict the magnitude and direction of the velocity of both proteins indicating trapping for the PEGylated conjugate or no trapping for the native protein. In addition, a parametric sweep study revealed the effect of the protein zeta potential on the electrokinetic response of the protein. We believe this work will serve as a tool to improve the design of electrokinetically driven microfluidic channels for the separation and recovery of PEGylated proteins in one single step. Published by AIP Publishing. [http://dx

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PEGylation to Improve Protein Stability During Melt Processing

Cited by 25 publications

References 47 publications

Biodegradable Viral Nanoparticle/Polymer Implants Prepared via Melt-Processing

Biodegradable Viral Nanoparticle/Polymer Implants Prepared via Melt-Processing

Milling solid proteins to enhance activity after melt-encapsulation

Modelling of electrokinetic phenomena for capture of PEGylated ribonuclease A in a microdevice with insulating structures

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