Different behavior of branched and linear polyethylenimine for gene deliveryin vitro andin vivo

Wightman, Lionel; Kircheis, Ralf; Rössler, Vanessa; Carotta, Sebastian; Ruzicka, Regina; Kursa, Malgorzata; Wagner, Ernst

doi:10.1002/jgm.187

Cited by 655 publications

(544 citation statements)

References 28 publications

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“…An exception to this is delivery to the lung, where it has been shown that large aggregating PEI/DNA particles passively target the lung following tail-vein injection. 23,24 For receptor-specific endocytosis, ligands such as epidermal growth factor (EGF) 25 and RGD peptide 26 have been utilized to increase cell specificity and overall uptake in vitro and in vivo. Though targeting often increases transfection, some experiments show that uptake that is both specific and efficient only occurs at a narrow window of charge ratios or not at all.…”

Section: Designing Polymeric Nanoparticles For Increased Uptake and Lmentioning

confidence: 99%

A Combinatorial Polymer Library Approach Yields Insight into Nonviral Gene Delivery

Green¹,

2008

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CONSPECTUSThe potential of gene therapy to benefit human health is tremendous because almost all human diseases have a genetic component, from untreatable monogenic disorders to cancer and heart disease. Unfortunately, a method for gene therapy that is both effective and safe has remained elusive. It has been said that "there are only three problems in gene therapy -delivery, delivery, and delivery." (quote from I. M. Verma in Jaroff, L. TIME, 1999; Jan 11).This Account describes an alternative strategy to viral gene delivery: the design of biodegradable polymers that are able to deliver DNA like a synthetic virus. Using high-throughput synthesis and screening techniques, we have created libraries of over 2000 structurally unique poly(β-amino esters) (PBAEs). PBAEs are formed by the conjugate addition of amines to diacrylates. These biomaterials are promising for nonviral gene delivery due to their ability to condense plasmid DNA into small and stable nanoparticles and their ability to promote cellular uptake and endosomal escape. Our laboratory has iteratively improved PBAE nanoparticles through polymer end modifications and nanoparticle coatings. Lead PBAEs have high gene delivery efficacy and low cytotoxicity both in vitro and in vivo.Certain polymer structural characteristics are important for effective gene delivery. The best PBAEs are linear polymers of ~10 kDa that contain hydroxyl side chains and primary amine end groups. These polymers bind DNA to form nanoparticles that are small (<200 nm) and stable and have near-neutral ζpotential in the presence of serum-containing media. Lead PBAEs also contain tertiary amines that can buffer the low pH environment of endosomes and facilitate escape of polymer/DNA particles into the cytoplasm.Diamine end-modified 1,4-butanediol diacrylate-co-5-amino-1-pentanol polymers (C32) bind DNA more tightly and form smaller nanoparticles than other PBAEs. These nanoparticles also have higher cellular uptake and the best gene expression of all gene delivery polymers in the library. These polymers are more effective for gene delivery than top commercially available nonviral vectors including jet-PEI and Lipofectamine 2000 and are comparable to adenovirus for in vitro gene delivery to human primary cells. In vivo, these PBAE/ DNA particles are promising as cancer therapeutics. This Account summarizes the results of our laboratory in using a combinatorial polymer library approach to elucidate polymer structure/function relationships and enable the development of polymeric gene delivery nanoparticles with viral-like efficacy. Nonviral gene delivery systems make use of physical methods or a synthetic chemical vector or both to deliver the gene of interest. Limitations with viral approaches, including small cargo capacity, resistance to repeated infection, difficulty in production and quality control, and low safety, can be potentially overcome with a nonviral approach. Originally, nonviral gene delivery was simply delivery of naked plasmid DNA. To improve transfection, physical str...

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Section: Designing Polymeric Nanoparticles For Increased Uptake and Lmentioning

confidence: 99%

A Combinatorial Polymer Library Approach Yields Insight into Nonviral Gene Delivery

Green¹,

2008

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“…We still do not have reasonable explanation for this discrepancy as is sometimes observed for non-viral vector systems such as polyethylenimine. 35 Presumably, primary liver cells and lined HepG2 cells may have different cellular metabolism. Furthermore, in vitro cell culture condition has quite different circumstances from in vivo.…”

Section: Gene Therapymentioning

confidence: 99%

Polyion complex micelles as vectors in gene therapy – pharmacokinetics and in vivo gene transfer

et al. 2002

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“…Linear PEI appears to assist these transport processes, whereas branched PEI and most nonviral delivery systems do not. 3,30 Other methods of assisting plasmid transport to the nucleus include the use of nuclear localization signal peptides. 31 It is possible that in the case of ODS, SPM directs the complex toward the cell nucleus.…”

Section: Intracellular Traffickingmentioning

confidence: 99%

“…The cultured cells were transfected with labeled plasmid (pGeneGrip) and unlabeled ODS or transfected with unlabeled plasmid and labeled ODS according to the transfection protocol above. At the appropriate time points (30,60,90,120, and 240 min), the cell culture was washed 3 times with sterile PBS and the confocal analysis was performed on the sample of living cells in sterile PBS. The images described in Figure 6 represent the behavior observed for 450% of the cells in every well (four wells for each treatment).…”

Section: Confocal Fluorescence Imaging Microscopymentioning

confidence: 99%

Novel dextran–spermine conjugates as transfecting agents: comparing water-soluble and micellar polymers

et al. 2004

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Recently, a novel cationic polymer, dextran-spermine (D-SPM) was developed for gene delivery. An efficient transfection was obtained using this polycation for a variety of genes and cell lines in serum-free or serum-poor medium. However, transfection using the water-soluble D-SPMbased polyplexes decreased with increasing serum concentration in cell culture in a concentration-dependent manner, reaching 95% inhibition at 50% serum in the cell growth medium. In order to overcome this obstacle, oleyl derivatives of D-SPM (which form micelles in aqueous phase) were synthesized at 1, 10, and 20 mol% of oleyl moiety to polymer e-NH 2 to form N-oleyl-D-SPM (ODS). Polyplexes based on ODS transfected well in medium containing 50% serum. Comparison with polyplexes based on well-established polymers (branched and linear polyethyleneimine) and with DOTAP/Cholesterol lipoplexes showed that regarding b-galactosidase transgene expression level and cytotoxicity in tissue culture, the D-SPM and ODS compare well with the above polyplexes and lipoplexes. Intracellular trafficking using FITC-labeled ODS and Rhodamine-labeled pGeneGrip plasmid cloned with hBMP2 monitored by confocal microscopy revealed that during the transfection process the fluorescent-labeled polymer concentrates in the Golgi apparatus and around the nucleus, while the cell cytoplasm was free of fluorescent particles, suggesting that the polyplexes move in the cell toward the nucleus by vesicular transport through the cytoplasm and not by a random diffusion. We found that the plasmids penetrate the cell nucleus without the polymer. Preliminary results in zebra fish and mice demonstrate the potential of ODS to serve as an efficient nonviral vector for in vivo transfection. Gene Therapy (2005) 12, 494-503.

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Different behavior of branched and linear polyethylenimine for gene deliveryin vitro andin vivo

Cited by 655 publications

References 28 publications

A Combinatorial Polymer Library Approach Yields Insight into Nonviral Gene Delivery

A Combinatorial Polymer Library Approach Yields Insight into Nonviral Gene Delivery

Polyion complex micelles as vectors in gene therapy – pharmacokinetics and in vivo gene transfer

Novel dextran–spermine conjugates as transfecting agents: comparing water-soluble and micellar polymers

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