Synthesis and in vitro characterization of an ABC triblock copolymer for siRNA delivery

Tatiana Segura, Jeffrey A. Hubbell

Research output: Contribution to journalArticle

60 Citations (Scopus)

Abstract

The ability to specifically down-regulate gene expression using the RNAi pathway in mammalian cells has tremendous potential in therapy and in basic science. However, delivery systems capable of efficient and biocompatible delivery of siRNA to target cells are not yet satisfactory. Here, we report the synthesis and in vitro characterization of ABC triblock copolymers that self-assemble with siRNA based on electrostatics and with each other by hydrophobic interactions. The ABC triblock copolymer is based on poly(ethylene glycol) (PEG), poly(propylene sulfide) (PPS), and a positively charged peptide (PEG-PPS-peptide). The diblock copolymer PEG45-PPS5,10 was synthesized using anionic polymerization of propylene sulfide upon a PEG macroinitiator, and the peptide domain was coupled to the PPS terminus using a disulfide exchange reaction with an N-terminal cysteine residue on the peptide. The peptides were designed to interact electrostatically with siRNA, selecting the TAT peptide domain of HIV (RKKRRQRRR) and an oligolysine (Lys9). The resulting triblock copolymers were able to self-assemble with siRNA as demonstrated by dynamic light scattering and gel electrophoresis. Complex size was found to be dependent on the amount of polymer used (charge ratio) and the length of the hydrophobic PPS block, achieving sizes ranging from 171 nm to 601 nm. Cell internalization and gene expression down-regulation studies showed that the triblock copolymers are able to transport siRNA inside the cell and mediate gene expression down-regulation, with the amount of internalization and gene transfer affected by charge ratio, PPS length, and the presence of serum. The proposed triblock was able to mediate gene expression down-regulation of GAPDH, achieving up to 90.5% ± 0.02% down-regulation.

Original languageEnglish
Pages (from-to)736-745
Number of pages10
JournalBioconjugate Chemistry
Volume18
Issue number3
DOIs
StatePublished - May 1 2007
Externally publishedYes

Fingerprint

Small Interfering RNA
Polyethylene glycols
Block copolymers
Down-Regulation
Peptides
Gene Expression Regulation
Polypropylenes
tat peptide (49-57), Human immunodeficiency virus 1
Gene expression
Ethylene Glycol
RNA Interference
Static Electricity
Hydrophobic and Hydrophilic Interactions
Polymerization
Disulfides
Cysteine
Electrophoresis
Gene transfer
Polymers
Gels

ASJC Scopus subject areas

  • Chemistry(all)
  • Organic Chemistry
  • Clinical Biochemistry
  • Biochemistry, Genetics and Molecular Biology(all)
  • Biochemistry

Cite this

Synthesis and in vitro characterization of an ABC triblock copolymer for siRNA delivery. / Segura, Tatiana; Hubbell, Jeffrey A.

In: Bioconjugate Chemistry, Vol. 18, No. 3, 01.05.2007, p. 736-745.

Research output: Contribution to journalArticle

Segura, Tatiana ; Hubbell, Jeffrey A. / Synthesis and in vitro characterization of an ABC triblock copolymer for siRNA delivery. In: Bioconjugate Chemistry. 2007 ; Vol. 18, No. 3. pp. 736-745.
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