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Optimizing siRNA-nanocarriers for systemic delivery by monitoring their behavior in blood

Kevin Buyens UGent (2010)
abstract
In conclusion, this thesis deals with the requirements that need to be met by siRNA/liposome complexes that are intended for intravenous application. However, the rational design of such siRNA/liposome complexes is hampered by the lack of methods and techniques that can check whether these complexes meet the requirements for systemic application. Indeed, up until now the available analytical methods to characterize siRNA/carrier complexes were not compatible with serum, plasma and whole blood. Therefore in this thesis novel methods are developed that enable an adequate characterization of siRNA/liposome complexes in biologically relevant media. The first developed method allows us to quantify the dissociation of siRNA/carrier complexes in both buffer and whole human serum. With the aid of this method we come to the conclusion that highly pegylated siRNA/liposome complexes should not be prepared through simple mixing of siRNA with preformed highly pegylated liposomes. This observation urged us to develop pegylated siRNA/liposome complexes that encapsulate the siRNA in their aqueous core. We found that an efficient encapsulation of siRNA in the aqueous core of highly pegylated liposomes can be obtained when the siRNA is added as a concentrated solution to a dry lipid film containing the liposomal constituents. Another observation that was made through use of this novel siRNA release assay is that siRNA complexes with bioresponsive cationic polymers disassemble in presence of serum, and the rate whereby they do so is correlated with a loss of gene knockdown capacity of the complex. Not only the dissociation of siRNA carrier complexes in blood but also the size of siRNA carrier complexes in blood is crucial for the further development of siRNA carrier complexes. Therefore, we established a novel method that allows sizing of nanoparticles in whole blood, even after in vivo intravenous injection of the particles. This has enabled us to study the difference in aggregation behavior of non-pegylated and highly pegylated liposomes in whole blood. While non-pegylated liposomes massively aggregate in blood, we have demonstrated that high pegylation degrees can indeed limit aggregation to a minimum. Finally also sizing of cell-derived microparticles was demonstrated to be possible, opening up possibilities to use this method for future diagnostic applications.
Please use this url to cite or link to this publication:
author
promoter
UGent and UGent
organization
alternative title
Optimalisatie van nanoscopische siRNA dragers voor systemische toediening door het nagaan van hun gedrag in bloed
year
type
dissertation
publication status
published
subject
keyword
systemic delivery, siRNA, nanocarriers
pages
166 pages
publisher
Ghent University. Faculty of Pharmaceutical Sciences
place of publication
Ghent, Belgium
defense location
Gent : Farmaceutisch Instituut (auditorium I)
defense date
2010-11-25 18:00
language
English
UGent publication?
yes
classification
D1
additional info
dissertation consists of copyrighted materials
copyright statement
I have transferred the copyright for this publication to the publisher
id
1081426
handle
http://hdl.handle.net/1854/LU-1081426
date created
2010-12-01 14:47:17
date last changed
2017-01-16 10:37:57
@phdthesis{1081426,
  abstract     = {In conclusion, this thesis deals with the requirements that need to be met by siRNA/liposome complexes that are intended for intravenous application. However, the rational design of such siRNA/liposome complexes is hampered by the lack of methods and techniques that can check whether these complexes meet the requirements for systemic application. Indeed, up until now the available analytical methods to characterize siRNA/carrier complexes were not compatible with serum, plasma and whole blood. Therefore in this thesis novel methods are developed that enable an adequate characterization of siRNA/liposome complexes in biologically relevant media. The first developed method allows us to quantify the dissociation of siRNA/carrier complexes in both buffer and whole human serum. With the aid of this method we come to the conclusion that highly pegylated siRNA/liposome complexes should not be prepared through simple mixing of siRNA with preformed highly pegylated liposomes. This observation urged us to develop pegylated siRNA/liposome complexes that encapsulate the siRNA in their aqueous core. We found that an efficient encapsulation of siRNA in the aqueous core of highly pegylated liposomes can be obtained when the siRNA is added as a concentrated solution to a dry lipid film containing the liposomal constituents. Another observation that was made through use of this novel siRNA release assay is that siRNA complexes with bioresponsive cationic polymers disassemble in presence of serum, and the rate whereby they do so is correlated with a loss of gene knockdown capacity of the complex. Not only the dissociation of siRNA carrier complexes in blood but also the size of siRNA carrier complexes in blood is crucial for the further development of siRNA carrier complexes. Therefore, we established a novel method that allows sizing of nanoparticles in whole blood, even after in vivo intravenous injection of the particles. This has enabled us to study the difference in aggregation behavior of non-pegylated and highly pegylated liposomes in whole blood. While non-pegylated liposomes massively aggregate in blood, we have demonstrated that high pegylation degrees can indeed limit aggregation to a minimum. Finally also sizing of cell-derived microparticles was demonstrated to be possible, opening up possibilities to use this method for future diagnostic applications.},
  author       = {Buyens, Kevin},
  keyword      = {systemic delivery,siRNA,nanocarriers},
  language     = {eng},
  pages        = {166},
  publisher    = {Ghent University. Faculty of Pharmaceutical Sciences},
  school       = {Ghent University},
  title        = {Optimizing siRNA-nanocarriers for systemic delivery by monitoring their behavior in blood},
  year         = {2010},
}

Chicago
Buyens, Kevin. 2010. “Optimizing siRNA-nanocarriers for Systemic Delivery by Monitoring Their Behavior in Blood”. Ghent, Belgium: Ghent University. Faculty of Pharmaceutical Sciences.
APA
Buyens, K. (2010). Optimizing siRNA-nanocarriers for systemic delivery by monitoring their behavior in blood. Ghent University. Faculty of Pharmaceutical Sciences, Ghent, Belgium.
Vancouver
1.
Buyens K. Optimizing siRNA-nanocarriers for systemic delivery by monitoring their behavior in blood. [Ghent, Belgium]: Ghent University. Faculty of Pharmaceutical Sciences; 2010.
MLA
Buyens, Kevin. “Optimizing siRNA-nanocarriers for Systemic Delivery by Monitoring Their Behavior in Blood.” 2010 : n. pag. Print.