Effect of cholesterol on nanoparticle translocation across a lipid bilayer.

Nanoparticles (NPs) have attracted significant attention as carriers for the delivery of drugs, genes, and macromolecules for biomedical and therapeutic applications. These technologies require NPs to be delivered to the interior of the cell. However, this translocation is unlikely because of the presence of a cell membrane composed of phospholipids, cholesterol, proteins, and glycans. The cell membrane composition can influence its rigidity; thus, membrane composition is a crucial factor in determining the translocation of NPs across the cell membrane. Here, we focus on cholesterol, which is an essential component of biological cell membranes, and investigate NP translocation across membranes containing cholesterol under an applied electric field using a coarse-grained molecular dynamics simulation. We found that NPs could translocate directly across cholesterol-containing membranes without irreversible membrane disruption. This unique translocation was induced by two key phenomena. Before NP translocation, a phospholipid-rich/cholesterol-poor domain was formed at the NP-membrane contact interface. Second, a smaller transmembrane pore was formed in the cholesterol-containing membrane during membrane crossing of the NP. Our findings imply that the delivery of NPs to the cell interior across the cholesterol-containing membrane can be achieved by appropriately controlling the strength of the applied electric field, depending on the cholesterol content in the membrane.

Enhancement of cell membrane permeability by using charged nanoparticles and a weak external electric field.

Because the cell membrane is the main barrier of intracellular delivery, it is important to facilitate and control the translocation of extracellular compounds across it. Our earlier molecular dynamics simulations suggested that charged nanoparticles under a weak external electric field can enhance the permeability of the cell membrane without disrupting it. However, this membrane permeabilization approach has not been tested experimentally. This study investigated the membrane crossing of a model compound (dextran with a Mw of 3000-5000) using charged nanoparticles and a weak external electric field. A model bilayer lipid membrane was prepared by using a droplet contact method. The permeability of the membrane was evaluated using the electrophysiological technique. Even when the applied electric field was below the critical strength for membrane breakdown, dextran was able to cross the membrane without causing membrane breakdown. These results indicate that adding nanomaterials under a weak electric field may enhance the translocation of delivery compounds across the cell membrane with less damage, suggesting a new strategy for intracellular delivery systems.

Viltolarsen in Japanese Duchenne muscular dystrophy patients: A phase 1/2 study.

OBJECTIVE: The novel morpholino antisense oligonucleotide viltolarsen targets exon 53 of the dystrophin gene, and could be an effective treatment for patients with Duchenne muscular dystrophy (DMD). We investigated viltolarsen's ability to induce dystrophin expression and examined its safety in DMD patients. METHODS: In this open-label, multicenter, parallel-group, phase 1/2, exploratory study, 16 ambulant and nonambulant males aged 5-12 years with DMD received viltolarsen 40 or 80 mg/kg/week via intravenous infusion for 24 weeks. Primary endpoints were dystrophin expression and exon 53 skipping levels. RESULTS: In western blot analysis, mean changes in dystrophin expression (% normal) from baseline to Weeks 12 and 24 were - 1.21 (P = 0.5136) and 1.46 (P = 0.1636), respectively, in the 40 mg/kg group, and 0.76 (P = 0.2367) and 4.81 (P = 0.0536), respectively, in the 80 mg/kg group. The increase in mean dystrophin level at Weeks 12 and 24 was significant in the 80 mg/kg group (2.78%; P = 0.0364). Patients receiving 80 mg/kg showed a higher mean exon 53 skipping level (42.4%) than those receiving 40 mg/kg (21.8%). All adverse events were judged to be mild or moderate in intensity and none led to study discontinuation. INTERPRETATION: Treatment with viltolarsen 40 or 80 mg/kg elicited an increasing trend in dystrophin expression and exon 53 skipping levels, and was safe and well tolerated. The decline in motor function appeared less marked in patients with higher dystrophin levels; this may warrant further investigation. This study supports the potential clinical benefit of viltolarsen.