ABSTRACT
The clinical application of gene silencing is hindered by poor stability and low delivery efficiency of naked oligonucleotides. Here, we present the in vitro and in vivo behaviors of a rationally designed, ternary, self-assembled nanoparticle complex, consisting of an anionic copolymer, cationic DOTAP liposome, and antisense oligonucleotide (AON). The multifunctional copolymers are based on backbone poly(propylacrylic acid) (PPAA), a pH-sensitive hydrophobic polymer, with grafted poly(alkylene oxides) (PAOs) varying in extent of grafting and PAO chemistry. The nanoparticle complexes with PPAA-g-PAO copolymers enhance antisense gene silencing effects in A2780 human ovarian cancer cells. A greater amount of AON is delivered to ovarian tumor xenografts using the ternary copolymer-stabilized delivery system, compared to a binary DOTAP/AON complex, following intraperitoneal injection in mice. Further, intratumoral injection of the nanoparticle complexes containing 1 mol% grafted PAO reduced tumoral bcl-2 expression by up to 60%. The data for complexes across the set of PAO polymers support a strong role for the hydrophilic-lipophilic balance of the graft copolymer in achieving serum stability and cellular uptake. Based upon these results, we anticipate that this novel nanoparticle delivery system can be extended to the delivery of plasmid DNA, siRNA, or aptamers for preclinical and clinical development.
Subject(s)
Acrylic Resins/chemistry , Liposomes/chemistry , Oligonucleotides, Antisense/administration & dosage , Polyethylene Glycols/chemistry , 1,2-Dipalmitoylphosphatidylcholine/chemistry , Acrylamides , Animals , Cations/chemistry , Cell Line, Tumor , Drug Carriers , Drug Delivery Systems , Female , Gene Silencing/drug effects , Genes, bcl-2/genetics , Genetic Therapy/methods , Hemolysis/drug effects , Humans , Mice , Mice, Nude , Nanoparticles , Neoplasms/therapy , Oligonucleotides, Antisense/pharmacokinetics , Oligonucleotides, Antisense/pharmacology , Tissue Distribution , Xenograft Model Antitumor AssaysABSTRACT
The study was aimed to prepare and evaluate gabapentin loaded albumin nanoparticles and to find out their effectiveness in treating epilepsy. Albumin nanoparticles of gabapentin were prepared by pH-coacervation method. The drug was administered into animals as free drug, gabapentin bound with nanoparticles, and gabapentin bound with nanoparticles coated with polysorbate 80. The polysorbate 80 coated nanoparticles increased the gabapentin concentration in the brain about 3 fold in comparison with the free drug. Moreover, the polysorbate 80 coated nanoparticles significantly reduced the duration of all phases of convulsion in both maximal electroshock induced and pentylenetetrazole induced convulsion models in comparison with free drug and drug bound with nanoparticle formulations, which indicates the ability of polysorbate 80 coated nanoparticles to enhance the gabapentin concentration in the brain.
Subject(s)
Amines , Anticonvulsants , Cyclohexanecarboxylic Acids , Drug Carriers , Nanoparticles , Polysorbates , Serum Albumin, Bovine , gamma-Aminobutyric Acid , Amines/administration & dosage , Amines/chemistry , Amines/pharmacokinetics , Animals , Anticonvulsants/administration & dosage , Anticonvulsants/chemistry , Anticonvulsants/pharmacokinetics , Brain/metabolism , Cyclohexanecarboxylic Acids/administration & dosage , Cyclohexanecarboxylic Acids/chemistry , Cyclohexanecarboxylic Acids/pharmacokinetics , Disease Models, Animal , Drug Carriers/administration & dosage , Drug Carriers/chemistry , Drug Carriers/pharmacokinetics , Drug Compounding , Electroshock , Gabapentin , Male , Nanoparticles/administration & dosage , Nanoparticles/chemistry , Pentylenetetrazole , Polysorbates/administration & dosage , Polysorbates/chemistry , Polysorbates/pharmacokinetics , Rats, Wistar , Seizures/drug therapy , Serum Albumin, Bovine/administration & dosage , Serum Albumin, Bovine/chemistry , Serum Albumin, Bovine/pharmacokinetics , gamma-Aminobutyric Acid/administration & dosage , gamma-Aminobutyric Acid/chemistry , gamma-Aminobutyric Acid/pharmacokineticsABSTRACT
The advancement of gene-based therapeutics to the clinic is limited by the ability to deliver physiologically relevant doses of nucleic acids to target tissues safely and effectively. Over the last couple of decades, researchers have successfully employed polymer and lipid based nanoassemblies to deliver nucleic acids for the treatment of a variety of diseases. Results of phase I/II clinical studies to evaluate the efficacy and biosafety of these gene delivery vehicles have been encouraging, which has promoted the design of more efficient and biocompatible systems. Research has focused on designing carriers to achieve biocompatibility, stability in the circulatory system, biodistribution to target the disease site, and intracellular delivery, all of which enhance the resulting therapeutic effect. The family of poly(alkylene oxide) (PAO) polymers includes random, block, and branched structures, among which the ABA type triblocks copolymers of ethylene oxide (EO) and propylene oxide (PO) (commercially known as Pluronic) have received the greatest consideration. In this Account, we highlight examples of polycation-PAO conjugates, liposome-PAO formulations, and PAO micelles for nucleic acid delivery. Among the various polymer design considerations, which include molecular weight of polymer, molecular weight of blocks, and length of blocks, the overall hydrophobic-lipophilic balance (HLB) is a critical parameter in defining the behavior of the polymer conjugates for gene delivery. We discuss the effects of varying this parameter in the context of improving gene delivery processes, such as serum stability and association with cell membranes. Other innovative macromolecular modifications discussed in this category include our work to enhance the serum stability and efficiency of lipoplexes using PAO graft copolymers, the development of a PAO gel-based carrier for sustained and stimuli responsive delivery, and the development of biodegradable PAO-based amphiphilic block copolymers.
Subject(s)
Nucleic Acids/metabolism , Polymers/chemistry , Animals , COS Cells , Chlorocebus aethiops , Liposomes/chemistry , Mice , Micelles , NIH 3T3 Cells , Nucleic Acids/genetics , Poloxamer/chemistry , Polyethylene Glycols/chemistry , TransfectionABSTRACT
Antisense technology holds tremendous potential in the research and clinical settings. However, successful delivery of antisense oligodeoxynucleotides (ODNs) to the intracellular site of action requires the passage of many barriers, including survival against extracellular serum nucleases and escape from endolysosomal degradation. Previous work has shown that the effectiveness of antisense delivery by the cationic liposome, dioleoyl-3-trimethylammonium-propane (DOTAP), is enhanced substantially by the incorporation of a pH-sensitive polymer, poly (propylacrylic acid) (PPAA), in serum-free media. To improve this system for application in serum-containing media conditions, PPAA was modified in this work by grafting onto it either poly(ethylene oxide) (PEO) or a more hydrophobic analog, poly (oxyalkylene amine), known as Jeffamine. The ternary formulation of DOTAP/ODN/PPAA-g-Jeffamine resulted in 8-fold increased uptake of fluorescently-labeled ODNs compared to DOTAP/ODN/PPAA and ~80% silencing of green fluorescent protein (GFP) expression in CHO-d1EGFP cells treated in the presence of 10% FBS-containing media. In contrast, the carrier systems that contained PPAA or PPAA-g-PEO failed to display any significant antisense activity in the presence of serum, even though all of the delivery systems displayed moderate to high levels of antisense activity in serum-free conditions. The results reveal that the carrier system with the Jeffamine graft copolymer effectively mediates specific gene silencing in the presence of serum, while the system with the PEO graft copolymer fails to do so. While the pH-dependent lytic functionality of PPAA was found to be lost upon grafting with PEO or Jeffamine, the hydrophobicity of the latter was sufficient to mediate cellular internalization and endosomal escape. Thus, the PPAA-g-Jeffamine copolymers hold substantial promise as agents for controlled therapeutic delivery of antisense oligonucleotides.
