Polyethyleneimine‐based immunopolyplex for targeted gene transfer in human lymphoma celllines

Guillem, Vicent; Tormo, Mar; Revert, Fernando; Benet, Isabel; Garcı́a-Conde, J; Crespo, Antonio; Aliño, Salvador F.

doi:10.1002/jgm.228

Cited by 24 publications

(11 citation statements)

References 45 publications

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“…This antibody-mediated uptake has been successfully used to deliver plasmid DNA and antisense oligonucleotides. [13][14][15][16][17] Recently, Song et al 18 demonstrated that siRNA targeted delivered to human immunodeficiency virus-infected cells and HER2 positive cells by this strategy executed gene silencing. However, whether this promising strategy for siRNA delivery is applicable to other diseases remains unknown.…”

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confidence: 99%

Targeted inhibition of HBV gene expression by single-chain antibody mediated small interfering RNA delivery

et al. 2007

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RNA interference is highly effective at inhibiting HBV gene expression and replication. However, before small interfering RNA (siRNA) can be used in the clinic, it is essential to develop a system to target their delivery. Antibody-mediated delivery is a novel approach for targeting siRNA to appropriate cells. In this report, we asked whether this siRNA delivery strategy would be effective against HBV. Of 5 candidates, a specific siRNA that effectively inhibited HBV gene expression and replication was determined. Two fusion proteins, s-tP and sC-tP, were constructed to contain a single chain of the human variable fragment, scFv, against hepatitis B surface antigen (HBsAg), a truncated protamine (tP), and in the case of sC-tP, a constant region of the chain (C). S-tP and sC-tP were developed to provide targeted delivery of the siRNA, siRNA expressing cassettes (SEC), and siRNA-producing plasmids. Fluorescein isothiocyanate-siRNA, fluorescein isothiocyanate-SEC, and plasmid DNA were specifically delivered into HBsAg-positive cells using the sC-tP fusion protein, and effectively inhibited HBV gene expression and replication. HBV gene expression was also inhibited by siRNA or siRNA-producing plasmids in HBV transgenic mice. Conclusion: Our results describe a potential method for the targeted delivery of siRNA or siRNAproducing plasmids against HBV, using anti-HBsAg fusion proteins. (HEPATOLOGY 2007;46: 84-94.) R ibonucleic acid interference is a naturally occurring mechanism for silencing homologous genes. 1,2 Synthetic and plasmid-based siRNA expression systems are both effective at suppressing HBV infection, in vitro and in vivo, [3][4][5][6][7][8][9] providing the basis for a new therapeutic strategy against HBV. However, the absence of a suitable delivery system presents a major obstacle against their clinical use. [10][11][12] Targeted delivery of small interfering RNA (siRNA) to relevant cell populations should increase the therapeutic index and minimize potential side effects and cellular toxicity.Antibody-mediated delivery is an effective method of targeting siRNA to particular cells. This involves fusing the nucleic acid-binding domain to a Fab fragment or scFv that recognizes a membrane receptor, resulting in a fusion protein that possesses cell recognition and nucleic acid-binding abilities. The fusion protein can then bind nucleic acids and deliver them into target cells through receptor-mediated endocytosis. This antibody-mediated uptake has been successfully used to deliver plasmid DNA and antisense oligonucleotides. [13][14][15][16][17] Song et al. 18 demonstrated that siRNA targeted delivered to human immunodeficiency virus-infected cells and HER2 positive cells by this strategy executed gene silencing. However, whether this promising strategy for siRNA delivery is applicable to other diseases remains unknown. 19 In this report, we provide evidence that antibody-mediated siRNA delivery is effectively targeted to HBV-infected cells.In a previous study, we have obtained 5 human antihepatitis B sur...

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confidence: 99%

Targeted inhibition of HBV gene expression by single-chain antibody mediated small interfering RNA delivery

et al. 2007

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“…The cells were transfected by means of a chemical procedure based on PEI 25 kDa (polyethyleneimine, Sigma, Madrid, Spain) polyplexes (DNA:PEI, 1:1.41) with 5 μg/mL of plasmids, as previously described [25,32]. The transfection percentage with this method lies between 20% and 40% of total cells, as observed using the reporter EGFP gene (data not shown).…”

Section: Methodsmentioning

confidence: 91%

“…The BD OptEIA ELISA kit for mGM-CSF (Pharmingen, BD Biosciences, Madrid, Spain) was used. A time point of 5 days was chosen on the basis of prior experimental results, assessed to study cytokine production over time, using the referred transfection conditions [25,32], in order to guarantee adequate GM-CSF production according to the literature, i.e. , >35 ng/10 6 cells/24 h [28,31].…”

Section: Methodsmentioning

confidence: 99%

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Antitumor Cell-Complex Vaccines Employing Genetically Modified Tumor Cells and Fibroblasts

Miguel

Herrero

Sendra

et al. 2014

Toxins

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The present study evaluates the immune response mediated by vaccination with cell complexes composed of irradiated B16 tumor cells and mouse fibroblasts genetically modified to produce GM-CSF. The animals were vaccinated with free B16 cells or cell complexes. We employed two gene plasmid constructions: one high producer (pMok) and a low producer (p2F). Tumor transplant was performed by injection of B16 tumor cells. Plasma levels of total IgG and its subtypes were measured by ELISA. Tumor volumes were measured and survival curves were obtained. The study resulted in a cell complex vaccine able to stimulate the immune system to produce specific anti-tumor membrane proteins (TMP) IgG. In the groups vaccinated with cells transfected with the low producer plasmid, IgG production was higher when we used free B16 cell rather than cell complexes. Nonspecific autoimmune response caused by cell complex was not greater than that induced by the tumor cells alone. Groups vaccinated with B16 transfected with low producer plasmid reached a tumor growth delay of 92% (p ≤ 0.01). When vaccinated with cell complex, the best group was that transfected with high producer plasmid, reaching a tumor growth inhibition of 56% (p ≤ 0.05). Significant survival (40%) was only observed in the groups vaccinated with free transfected B16 cells.

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“…PEI is an excellent choice for a basic building block in the design of gene carriers because of its high efficiency and abundant amino groups are amenable to conjugation 45,46,[63][64][65][66][67][68] . Further improvements in the transfection efficiency generated by PEI can often be achieved after conjugation of a cell binding ligand to PEI, thereby enabling ligand-receptor mediated endocytosis.…”

Section: Modified Pei Used In Gene Deliverymentioning

confidence: 99%

Application of polymers in nucleic acid delivery

Jiang¹

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Gene therapy and immunotherapy are powerful techniques in the treatment of many life threatening diseases. The major challenge in these therapies is to seek a safe and efficient delivery carrier for gene and antigen materials. Carriers are designed to protect these molecules from degradation, improve their stability and facilitate the delivery of them to the site of action. This research study aims to develop appropriate carriers for small interferencing RNA (siRNA), DNA, antigen and ajuvant respectively. In the case of siRNA, material encompassing mannose, polyethylene glycol (PEG) and polyethylenimine (PEI) was investigated. Two structures were assembled: in one construct, mannose was conjugated to PEI directly (Mannose-PEI-PEG) whilst in a second construct; the mannose was conjugated to PEI via a PEG spacer (PEI-PEGmannose). Confocal microscopy images suggested a faster escape and release of siRNA into the perinuclear region when siRNA was complexed with mannose-PEG-PEI. Mannosylation and PEGylation generated significant toxicity reduction compared to unmodified PEI alone. Real-time polymerase chain reaction (RT-PCR) results showed a significant decrease on mRNA knockdown when using modified PEIs. It was found that PEI-PEG-mannose was a stronger candidate for siRNA delivery because it displays lower toxicity, higher uptake efficiency and higher relative knockdown efficiencies. In the case of pDNA delivery, dextran was introduced to reduce the toxicity generated by PEI. PEI 2000 was more effective than PEI 800 in condensing DNA and inducing transfection when incorporated with dextran. The toxicity of dextran-PEI was greatly reduced when compared to unmodified PEI. Dextran-PEI was able to generate significantly higher transfection efficiencies than PEI alone in the presence of serum. An improved stability of complexes in serum by dextran-PEI was noticed along with a faster release of complexes to the perinuclear region of cells after endocytosis. These observations help to account for the higher efficiency of dextran-PEI in gene transfer. In our final study on vaccines, we utilized cationic polyamidoamine (PAMAM) dendrimer polymers to modify biodegradable particles for enhanced delivery of antigens and adjuvants. Vaccines were formulated by loading CpG oligonucleotide (CpG ODN) and ovalbumin (OVA) into biodegradable microparticles. In one group, OVA and CpG were conjugated together and then loaded into the PLGA microparticles. In other groups, CpG was loaded into the particles and OVA bound to the surface and finally particles were prepared that were loaded with OVA and surface modified with cationic PAMAM dendrimers to electrostatically bind CpG ODN. The microparticles were able to provide sustained release of antigen and adjuvants over 14 day's course. The up regulation of CD86 and H2Kb indicated strong activation of DC and therefore strong induction of CD8+ T-cells. MHC II markers were not as significantly affected. Particles loaded with OVA and surface bound CpG ODN ((OVA)-CpG) showed the highest cytotoxic C...

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Polyethyleneimine‐based immunopolyplex for targeted gene transfer in human lymphoma celllines

Abstract: It is concluded that immunopolyplex is a good non-viral vector for specific and selective nucleic acid delivery. Immunopolyplex design allows easy replacement of the targeting element (antibody) - the streptavidin-polyplex backbone remaining intact - thereby conferring high versatility.

Cited by 24 publications

References 45 publications

Targeted inhibition of HBV gene expression by single-chain antibody mediated small interfering RNA delivery

Targeted inhibition of HBV gene expression by single-chain antibody mediated small interfering RNA delivery

Antitumor Cell-Complex Vaccines Employing Genetically Modified Tumor Cells and Fibroblasts

Application of polymers in nucleic acid delivery

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