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PEGylated polyplex with optimized PEG shielding enhances gene introduction in lungs by minimizing inflammatory responses.

ABSTRACT: Safety is a critical issue in clinical applications of nonviral gene delivery systems. Safe and effective gene introduction into the lungs was previously achieved using polyplexes from poly(ethyleneglycol) (PEG)-block-polycation [PEG-block-PAsp(DET)] and plasmid DNA (pDNA). Although PEGylated polyplexes appeared to be safe, an excess ratio of polycation to pDNA was needed to obtain sufficient transgene expression, which may cause toxicities shortly after gene introduction. In the present study, we investigated the combined use of two polymers, PEG-block-PAsp(DET) (B) and homo PAsp(DET) (H) across a range of mixing ratios to construct polyplexes. Although transgene expressions following in vitro transfections increased in parallel with increased proportions of H, polyplexes with B/H = 50/50 formulation produced the highest expression level following in vivo intratracheal administration. Higher proportions of H elicited high levels of cytokine induction with significant inflammation as assessed by histopathological examinations. Based on the aggregation behavior of polyplexes in bronchoalveolar lavage fluids (BALFs), we suggested that rapid aggregation of polyplexes in the lung induced acute inflammatory responses, resulting in reduced transgene expression. B/H formulation of polyplex can help to improve gene therapy for the respiratory system because it achieves both effective PEG shielding of polyplexes and functioning of PAsp(DET) polycations to enhance endosomal escape.

SUBMITTER: Uchida S

PROVIDER: S-EPMC3369293 | biostudies-literature | 2012 Jun

REPOSITORIES: biostudies-literature

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PEGylated polyplex with optimized PEG shielding enhances gene introduction in lungs by minimizing inflammatory responses.

Uchida Satoshi S Itaka Keiji K Chen Qixian Q Osada Kensuke K Ishii Takehiko T Shibata Masa-Aki MA Harada-Shiba Mariko M Kataoka Kazunori K

Molecular therapy : the journal of the American Society of Gene Therapy 20120214 6

Safety is a critical issue in clinical applications of nonviral gene delivery systems. Safe and effective gene introduction into the lungs was previously achieved using polyplexes from poly(ethyleneglycol) (PEG)-block-polycation [PEG-block-PAsp(DET)] and plasmid DNA (pDNA). Although PEGylated polyplexes appeared to be safe, an excess ratio of polycation to pDNA was needed to obtain sufficient transgene expression, which may cause toxicities shortly after gene introduction. In the present study, ...[more]

PMID: 22334020

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Project description:A rationally-designed library of ternary siRNA polyplexes was developed and screened for gene silencing efficacy in vitro and in vivo with the goal of overcoming both cell-level and systemic delivery barriers. [2-(dimethylamino)ethyl methacrylate] (DMAEMA) was homopolymerized or copolymerized (50mol% each) with butyl methacrylate (BMA) from a reversible addition - fragmentation chain transfer (RAFT) chain transfer agent, with and without pre-conjugation to polyethylene glycol (PEG). Both single block polymers were tested as core-forming units, and both PEGylated, diblock polymers were screened as corona-forming units. Ternary siRNA polyplexes were assembled with varied amounts and ratios of core-forming polymers to PEGylated corona-forming polymers. The impact of polymer composition/ratio, hydrophobe (BMA) placement, and surface PEGylation density was correlated to important outcomes such as polyplex size, stability, pH-dependent membrane disruptive activity, biocompatibility, and gene silencing efficiency. The lead formulation, DB4-PDB12, was optimally PEGylated not only to ensure colloidal stability (no change in size by DLS between 0 and 24h) and neutral surface charge (0.139mV) but also to maintain higher cell uptake (>90% positive cells) than the most densely PEGylated particles. The DB4-PDB12 polyplexes also incorporated BMA in both the polyplex core- and corona-forming polymers, resulting in robust endosomolysis and in vitro siRNA silencing (~85% protein level knockdown) of the model gene luciferase across multiple cell types. Further, the DB4-PDB12 polyplexes exhibited greater stability, increased blood circulation time, reduced renal clearance, increased tumor biodistribution, and greater silencing of luciferase compared to our previously-optimized, binary parent formulation following intravenous (i.v.) delivery. This polyplex library approach enabled concomitant optimization of the composition and ratio of core- and corona-forming polymers (indirectly tuning PEGylation density) and identification of a ternary nanomedicine optimized to overcome important siRNA delivery barriers in vitro and in vivo.

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PEGylated polyplex with optimized PEG shielding enhances gene introduction in lungs by minimizing inflammatory responses.

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PEGylated polyplex with optimized PEG shielding enhances gene introduction in lungs by minimizing inflammatory responses.

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