10 results on '"Flynn, Nicholas"'
Search Results
2. Effect of zwitterionic betaine surfactant on interfacial behavior of bovine serum albumin (BSA)
- Author
-
Erfani, Amir, Khosharay, Shahin, Flynn, Nicholas H., Ramsey, Joshua D., and Aichele, Clint P.
- Published
- 2020
- Full Text
- View/download PDF
3. Increasing protein stability by association with zwitterionic amphiphile cocamidopropyl betaine
- Author
-
Erfani, Amir, Flynn, Nicholas H., Ramsey, Joshua D., and Aichele, Clint P.
- Published
- 2019
- Full Text
- View/download PDF
4. Effect of cationic grafted copolymer structure on the encapsulation of bovine serum albumin.
- Author
-
Flynn, Nicholas, Topal, Ç. Özge, Hikkaduwa Koralege, Rangika S., Hartson, Steve, Ranjan, Ashish, Liu, Jing, Pope, Carey, and Ramsey, Joshua D.
- Subjects
- *
CATIONIC polymers , *GRAFT copolymers , *ENCAPSULATION (Catalysis) , *SERUM albumin , *POLYMER structure - Abstract
The aim of the present study was to evaluate a library of poly- l -lysine (PLL)-graft (g)-polyethylene glycol (PEG) copolymers for the ability to encapsulate effectively a model protein, bovine serum albumin (BSA), and to characterize the stability and protein function of the resulting nanoparticle. A library of nine grafted copolymers was produced by varying PLL molecular weight and PEG grafting ratio. Electrostatic self-assembly of the protein and the grafted copolymer drove encapsulation. The formation of protein/polymer nanoparticles with a core/shell structure was confirmed using PAGE, dynamic light scattering, and electron microscopy. Encapsulation of the BSA into nanoparticles was strongly dependent on the copolymer-to-protein mass ratio, PEG grafting ratio, and PLL molecular weight. A copolymer-to-protein mass ratio of 7:1 and higher was generally required for high levels of encapsulation, and under these conditions, no loss of protein activity was observed. Copolymer characteristics also influenced nanoparticle resistance to polyanions and protease degradation. The results indicate that a copolymer of 15–30 kDa PLL, with a PEG grafting ratio of 10:1, is most promising for protein delivery. [ABSTRACT FROM AUTHOR]
- Published
- 2016
- Full Text
- View/download PDF
5. Cell-penetrating peptides transport therapeutics into cells.
- Author
-
Ramsey, Joshua D. and Flynn, Nicholas H.
- Subjects
- *
CELL-penetrating peptides , *CELL membranes , *NANOPARTICLES , *CHIMERIC proteins , *DRUG delivery systems , *TARGETED drug delivery , *IN vitro studies - Abstract
Nearly 30 years ago, certain small, relatively nontoxic peptides were discovered to be capable of traversing the cell membrane. These cell-penetrating peptides, as they are now called, have been shown to not only be capable of crossing the cell membrane themselves but can also carry many different therapeutic agents into cells, including small molecules, plasmid DNA, siRNA, therapeutic proteins, viruses, imaging agents, and other various nanoparticles. Many cell-penetrating peptides have been derived from natural proteins, but several other cell-penetrating peptides have been developed that are either chimeric or completely synthetic. How cell-penetrating peptides are internalized into cells has been a topic of debate, with some peptides seemingly entering cells through an endocytic mechanism and others by directly penetrating the cell membrane. Although the entry mechanism is still not entirely understood, it seems to be dependent on the peptide type, the peptide concentration, the cargo the peptide transports, and the cell type tested. With new intracellular disease targets being discovered, cell-penetrating peptides offer an exciting approach for delivering drugs to these intracellular targets. There are hundreds of cell-penetrating peptides being studied for drug delivery, and ongoing studies are demonstrating their success both in vitro and in vivo. [ABSTRACT FROM AUTHOR]
- Published
- 2015
- Full Text
- View/download PDF
6. Polyionic complexes of butyrylcholinesterase and poly-l-lysine-g-poly(ethylene glycol): Comparative kinetics of catalysis and inhibition and in vitro inactivation by proteases and heat.
- Author
-
Hester, Kirstin, Liu, Jing, Flynn, Nicholas, Sultatos, Lester G., Geng, Liyi, Brimijoin, Stephen, Ramsey, Joshua D., Hartson, Steven, Ranjan, Ashish, and Pope, Carey
- Subjects
- *
POLYIONS , *BUTYRYLCHOLINESTERASE , *POLYETHYLENE glycol , *PROTEOLYTIC enzymes , *CATALYSIS - Abstract
We previously reported that recombinant human butyrylcholinesterase (rhBChE) complexed with a series of copolymers of poly- l -lysine (PLL) with grafted (polyethylene) glycol (PEG) (i.e., PLL-g-PEG) showed reduced catalytic activity but relatively similar concentration-dependent inactivation of the organophosphorus inhibitor paraoxon. Herein, we compared the kinetics of catalysis (using butyrylthiocholine as the substrate) and inhibition (using four different inhibitors) of free and copolymer-complexed rhBChE. Using scanning electron microscopy, polyionic complexes of rhBChE with three different PLL-g-PEG copolymers (based on PLL size) appeared as spheroid-shaped particles with relatively similar particle sizes (median diameter = 35 nm). Relatively similar particle sizes were also noted using dynamic light scattering (mean = 26–35 nm). The three copolymer-complexed enzymes exhibited reduced k cat (30–33% reduction), but no significant changes in K m . Inhibitory potency (as reflected by the bimolecular rate constant, k i ) was similar among the free and copolymer-complexed enzymes when paraoxon was the inhibitor, whereas statistically significant reductions in k i (16–60%) were noted with the other inhibitors. Sensitivity to inactivation by proteases and heat was also compared. Copolymer-complexed enzymes showed lesser time-dependent inactivation by the proteases trypsin and pronase and by heat compared to the free enzyme. Understanding the unique properties of PLL-g-PEG-BChE complexes may lead to enhanced approaches for use of BChE and other protein bioscavengers. [ABSTRACT FROM AUTHOR]
- Published
- 2017
- Full Text
- View/download PDF
7. Identification of Excipients for Stabilizing Fiberless Adenovirus as Biopharmaceuticals.
- Author
-
Kupgan, Grit, Choudhari, Shyamal P., Flynn, Nicholas H., Nigatu, Adane, Vupputuri, Sravanthi, Picking, Wendy L., Picking, William D., and Ramsey, Joshua D.
- Subjects
- *
ADENOVIRUSES , *STABILIZING agents , *EXCIPIENTS , *BIOPHARMACEUTICS , *GENE therapy , *THERAPEUTICS - Abstract
Reducing the promiscuous tropism of native adenovirus by using fiberless adenovirus is advantageous toward its use as a gene therapy vector or vaccine component. The removal of the fiber protein on native adenovirus abrogates several undesirable interactions; however, this approach decreases the particle's physical stability. To create stable fiberless adenovirus for pharmaceutical use, the effects of temperature and pH on the particle's stability profile must be addressed. Our results indicate that the stability of fiberless adenovirus is increased when it is stored in mildly acidic conditions around pH 6. The stability of fiberless adenovirus can be further enhanced by using excipients. Excipient screening results indicate that the nonionic surfactant Pluronic F-68 and the amino acid glycine are potential stabilizers because of their ability to increase the thermal transition temperature of the virus particle and promote retention of biological activity after exposure to prolonged thermal stress. Our data indicate that the instability of fiberless adenovirus can be ameliorated by storing the virus in the appropriate environment, and it should be possible to further optimize the virus so that it can be used as a biopharmaceutical. [ABSTRACT FROM AUTHOR]
- Published
- 2017
- Full Text
- View/download PDF
8. In vitro characterization of cationic copolymer-complexed recombinant human butyrylcholinesterase.
- Author
-
Pope, Carey, Uchea, Chibuzor, Flynn, Nicholas, Poindexter, Kirstin, Geng, Liyi, Brimijoin, W. Stephen, Hartson, Steve, Ranjan, Ashish, Ramsey, Joshua D., and Liu, Jing
- Subjects
- *
CHOLINESTERASES , *BUTYRYLCHOLINESTERASE , *COPOLYMERS , *ORGANOPHOSPHORUS compounds , *MOLECULAR weights - Abstract
Effective use of exogenous human BChE as a bioscavenger for organophosphorus toxicants (OPs) is hindered by its limited availability and rapid clearance. Complexes made from recombinant human BChE (rhBChE) and copolymers may be useful in addressing these problems. We used in vitro approaches to compare enzyme activity, sensitivity to inhibition, stability and bioscavenging capacity of free enzyme and copolymer-rhBChE complexes (C-BCs) based on one of nine different copolymers, from combinations of three molecular weights (MW) of poly- l -lysine (PLL; high MW, 30–70 kDa; medium MW, 15–30 kDa; low MW, 4–15 kDa) and three grafting ratios of poly(ethylene glycol) (PEG; 2:1, 10:1, 20:1). Retarded protein migration into acrylamide gels stained for BChE activity was noted with all copolymers as the copolymer-to-protein ratio was increased. BChE activity of C-BCs was lower relative to free enzyme, with the 2:1 grafting ratio showing generally greater reduction. Free enzyme and C-BCs showed relatively similar in vitro sensitivity to inhibition by paraoxon, but use of the 20:1 grafting ratio led to lower potencies. Through these screening assays we selected three C-BCs (high, medium and low MW; 10:1 grafting) for further characterizations. BChE activity was higher in C-BCs made with the medium and low compared to high MW-based copolymer. C-BCs generally showed higher stability than free enzyme when maintained for long periods at 37 °C or following incubation with chymotrypsin. Free enzyme and C-BCs were similarly effective at inactivating paraoxon in vitro . While these results are promising for further development, additional studies are needed to evaluate in vivo performance. [ABSTRACT FROM AUTHOR]
- Published
- 2015
- Full Text
- View/download PDF
9. Effects of cell-penetrating peptides on transduction efficiency of PEGylated adenovirus.
- Author
-
Nigatu, Adane S., Vupputuri, Sravanthi, Flynn, Nicholas, and Ramsey, Joshua D.
- Subjects
- *
CELL-penetrating peptides , *CELLULAR signal transduction , *POLYETHYLENE glycol , *ADENOVIRUSES , *GENE delivery techniques , *IMMUNOGENETICS , *COXSACKIEVIRUSES - Abstract
Background Adenovirus (Ad) is one of the viral vectors most widely used for gene delivery. The virus, however, has serious shortcomings such as immunogenicity, promiscuous tropism, and the inability to efficiently infect certain types of cells. The goal of this study was to improve the ability of an Ad-based vector to efficiently transform cells that lack the native coxsackie-adenovirus receptor (CAR − ) by modifying the virus with CPP-PEG conjugates. Methods The vector was produced by PEGylating Ad, which packages a lacZ reporter gene, and then conjugating CPPs to form CPP-PEG-Ad particles. The study compared the effectiveness of four different CPPs: Pen, Tat, Pep1, and pArg. The effects of CPP amount per virus, degree of PEGylation, and PEG molecular weight on transduction efficiency were studied on CAR − NIH/3T3 cells. Results CPP-PEG-Ad particles transduced CAR − cells significantly better than unmodified Ad. Pen, the most effective CPP, produced an 80-fold improvement in transduction compared to the unmodified virus. The Pen peptide utilized a combination of electrostatic and hydrophobic interactions with the cell membrane to maximize cellular association while the other CPPs used only electrostatic or hydrophobic interactions but not both. Lastly, higher degrees of PEGylation, which prompted PEG to adopt a “brush” conformation, resulted in more efficient CPP-PEG-Ad particles because of both better conjugation of CPPs to the PEGylated virus and better exposure of the conjugated CPPs on the surface of the particle. Conclusions CPP-PEG-Ad particles efficiently deliver genes to cells that Ad alone would not efficiently infect, thereby extending potential gene therapy treatments to a much broader range of cell types and diseases. [ABSTRACT FROM AUTHOR]
- Published
- 2015
- Full Text
- View/download PDF
10. Evaluation of cell-penetrating peptide/adenovirus particles for transduction of CAR-negative cells.
- Author
-
Nigatu, Adane S., Vupputuri, Sravanthi, Flynn, Nicholas, Neely, Brian J., and Ramsey, Joshua D.
- Subjects
- *
CELL-penetrating peptides , *ADENOVIRUSES , *CELLULAR signal transduction , *GENE therapy , *ELECTROSTATICS , *COXSACKIEVIRUSES , *CLINICAL trials - Abstract
Adenovirus (Ad) is a promising gene therapy vector, and is used currently in more than 23% of clinical gene therapy trials. The viral vector, however, has drawbacks such as immunogenicity, promiscuous tropism, and the inability to infect certain types of cells. The focus of this work was to develop an improved vector through electrostatic formation of a complex between negatively charged Ad and positively charged cell-penetrating peptides (CPPs), including Tat, Penetratin, polyarginine, and Pep1. The resulting complexes were demonstrated to be capable of transducing cells that lack the coxsackie-adenovirus receptor (CAR), and are otherwise difficult to infect with native Ad. The transduction efficiency of the complexes was optimized by varying the multiplicity of infection, complex formation time, and ratio of CPPs to Ad, which improved the transduction efficiency of CPP/Ad on CAR-negative cells more than 100-fold compared with unmodified Ad. The size of the CPP/Ad complex was initially less than 300 nm, but stability studies performed in the presence of serum indicate that the complex aggregates with serum after an extended period of time. The results of the current study indicate that electrostatic modification of Ad with CPPs provides a relevant platform for developing effective Ad-based gene therapy vectors. © 2013 Wiley Periodicals, Inc. and the American Pharmacists Association J Pharm Sci 102:1981-1993, 2013 [ABSTRACT FROM AUTHOR]
- Published
- 2013
- Full Text
- View/download PDF
Catalog
Discovery Service for Jio Institute Digital Library
For full access to our library's resources, please sign in.