A double-blind, dose escalation gene transfer trial was conducted in subjects with cystic fibrosis (CF), among whom placebo (saline) or compacted DNA superfused onto the inferior turbinate of right left nostril. The vector consisted single molecules plasmid carrying transmembrane regulator- encoding into nanoparticles, using polyethylene glycol-substituted 30-mer lysine peptides. Entry criteria included age greater than 18 years, FEV1 exceeding 50% predicted, and basal nasal potential...

Background The eye is an excellent candidate for gene therapy as it immune privileged and much of the disease-causing genetics are well understood. Towards this goal, we evaluated efficiency compacted DNA nanoparticles a system non-viral transfer to ocular tissues. examined here have been shown be safe effective in human clinical trial, no theoretical limitation on plasmid size, do not provoke responses, can highly concentrated. Methods Findings Here show that these targeted different...

Charge-neutral DNA nanoparticles have been developed in which single molecules of are compacted to their minimal possible size. We speculated that the small size these may facilitate gene transfer postmitotic cells, permitting nuclear uptake across 25-nm membrane pore. To determine whether can transfect nondividing growth-arrested neuroblastoma and hepatoma cells were transfected with DNA/liposome mixtures encoding luciferase. In both models, robustly by (6,900–360-fold more than naked DNA)....

Mutations in the photoreceptor-specific flippase ABCA4 are associated with Stargardt disease and many other forms of retinal degeneration that currently lack curative therapies. Gene replacement is a logical strategy for ABCA4-associated disease, particularly given current success traditional viral-mediated gene delivery, such as adeno-associated viral (AAV) vectors. However, large size cDNA (6.8 kbp) has hampered progress development genetic treatments. Nonviral DNA nanoparticles (NPs) can...

DNA can be compacted using polyethylene glycol-substituted poly-l-lysine into discrete unimolecular (with respect to DNA) nanoparticles with minor diameter <20 nm that are stable in normal saline for at least 23 months 4°C. We compared the activity of firefly luciferase lungs C57BL/6 mice received 100 μg plasmid 25 μl (shown optimal dose) via intratracheal or intranasal instillation levels animals given naked untreated mice. Mice dosed had peak lung 2 days postinstillation, which declined...

We have evaluated the effect of NaCl concentration on mode binding poly-l-lysine to DNA and resulting structural functional features condensed particles using precipitation, DNase I resistance, electron microscopy, receptor-mediated gene transfer assays. At a high in presence excess DNA, interacted with cooperatively, fully condensing some leaving rest unbound. low concentrations, molecules noncooperative fashion, <i>i.e.</i> they bind randomly whole population molecules. Cooperative...

Chromogranin A (CGA), also referred to as secretory protein I, is an acidic that has been detected in all neuroendocrine cell types examined and often present large amounts relative other secreted proteins. For example, CGA comprises at least 40% of the soluble adrenal chromaffin granule, it appears be major parathyroid granules. complementary DNAs (cDNAs) from bovine pituitary have recently cloned sequenced found nearly identical. region a high degree amino acid sequence identity...

Compacted DNA nanoparticles deliver transgenes efficiently to the lung following intrapulmonary dosing. Here we show that nucleolin, a protein known shuttle between nucleus, cytoplasm, and cell surface, is receptor for at surface. By using surface plasmon resonance (SPR), demonstrate nucleolin binds directly. The presence of on HeLa 16HBEo- cells was confirmed by biotinylation assay immunofluorescence. Rhodamine-labeled colocalize with as well in cytoplasm but not transferrin or markers...

The purpose of the present study was to test therapeutic efficiency and safety compacted-DNA nanoparticle-mediated gene delivery into subretinal space a juvenile mouse model retinitis pigmentosa. Nanoparticles containing opsin promoter wild-type Rds were injected subretinally mice carrying haploinsufficiency mutation in retinal degeneration slow (rds+/−) at postnatal day (P)5 22. Control either with saline, uncompacted naked plasmid DNA gene, or remained untreated. mRNA levels peaked...

Previously we have shown that compacted DNA nanoparticles can drive high levels of transgene expression after subretinal injection in the mouse eye. Here delivered containing a therapeutic gene to retinas model retinitis pigmentosa. Nanoparticles wild-type retinal degeneration slow (Rds) were injected into space rds(+/-) mice on postnatal day 5. Gene was sustained for up four months at times higher than controls with saline or naked DNA. The taken virtually all photoreceptors and mediated...

For effective airway gene therapy of cystic fibrosis (CF), inhaled carriers must first penetrate the hyperviscoelastic sputum covering epithelium. Whether clinically studied can CF remains unknown. Here, we measured diffusion a tested nonviral carrier, composed poly-l-lysine conjugated with 10 kDa polyethylene glycol segment (CK30PEG10k). We found that CK30PEG10k/DNA nanoparticles were trapped in sputum. To improve carrier across sputum, adjuvant regimens consisting N-acetylcysteine (NAC),...

Gene therapy is a critical tool for the treatment of monogenic retinal diseases. However, limited vector capacity current benchmark delivery strategy, adeno-associated virus (AAV), makes development larger alternatives, such as compacted DNA nanoparticles (NPs), critical. Here we conduct side-by-side comparison self-complementary AAV and CK30PEG NPs using matched ITR plasmids. We report that although AAVs are more efficient per genome (vg) than NPs, can drive gene expression on comparable...

Mutations in genes the retinal pigment epithelium (RPE) cause or contribute to debilitating ocular diseases, including Leber's congenital amaurosis (LCA). Genetic therapies, particularly adeno-associated viruses (AAVs), are a popular choice for monogenic diseases; however, limited payload capacity of AAVs combined with large number disease exceeding that make development alternative delivery methods critical. Here, we test ability compacted DNA nanoparticles (NPs) containing plasmid scaffold...

This study demonstrates proof of concept for delivery and expression compacted plasmid DNA in the central nervous system. Plasmid was with polyethylene glycol substituted lysine 30-mer peptides, forming rod-like nanoparticles diameters between 8 11 nm. Here we show that an intracerebral injection can transfect both neurons glia, produce transgene striatum up to weeks, which at least 100-fold greater than injections naked plasmids. Bioluminescent imaging (BLI) injected animals 11th...

Previously it was established that infusion of glial cell line-derived neurotrophic factor (GDNF) protein into grafts embryonic dopamine cells has a effect on the grafted cells. In this study we used nonviral technique to transfer gene encoding for GDNF striatal Plasmid DNA compacted nanoparticles (DNPs) by 10 kDa polyethylene glycol (PEG)-substituted lysine 30-mers (CK 30 PEG10k) and then injected denervated striatum rats with unilateral 6-hydroxydopamine lesions. Sham controls were saline....

Subretinal delivery of polyethylene glycol-substituted lysine peptide (CK30PEG)-compacted DNA nanoparticles results in efficient gene expression retinal cells. This work evaluates the ocular safety compacted nanoparticles. CK30PEG-compacted containing an EGFP plasmid were subretinally injected adult mice (1 microl at 0.3, 1.0 and 3.0 microg/microl). Retinas examined for signs inflammation 1, 2, 4 7 days post-injection. Neither infiltration polymorphonuclear neutrophils or lymphocytes was...

Nanoparticles containing DNA compacted with poly-l-lysine modified on an N-terminal cysteine polyethylene glycol can effectively transfect cells of the airway epithelium when applied by luminal route. To evaluate toxicity these nanoparticles, we administered 10 and 100 μg into nanoparticles suspended in normal saline intranasal route to mice determined pulmonary systemic responses this challenge, compared administration alone, some experiments, naked DNA, Escherichia coli genomic or...

The imprinted H19 gene is highly expressed in human embryos, fetal tissues and nearly completely shut off adults. However, it reexpressed a number of tumors including bladder carcinoma, demonstrating that RNA an oncofetal RNA. Tumors induced by injection carcinoma cell lines express contrast to the cells before injection. These observations support notion positive correlation between expression carcinoma. Loss imprinting IGF‐2 was observed samples

Mutations in the rhodopsin gene cause retinal degeneration and clinical phenotypes including retinitis pigmentosa (RP) congenital stationary night blindness. Effective therapies have been difficult to develop, however, because generating precise levels of expression is critical; overexpression causes toxicity, underexpression would result incomplete rescue. Current delivery strategies routinely use cDNA-based vectors for targeting; inclusion noncoding components genomic DNA (gDNA) such as...