Microliter-scale in vitro aerosol-cell delivery of inhalable antisense oligonucleotide (ASO) induces "burst-like" pharmacokinetics

Unlike traditional submerged cell cultures, nebulized drug delivery to air-liquid interface (ALI)-cultured lung cells provides a clinically relevant route for in vitro testing of inhalable drugs. Current nebulizer systems deliver non-physiologically large drug volumes. Here, we introduce the Cloud MAX technology, characterize its accuracy of dosimetry, and apply it to toxicity, efficacy, and biokinetics or permeability testing of a locked nucleic acid-modified antisense oligonucleotide (ASO). On average the Cloud MAX delivers 44% of 3-20 μL aerosolized liquid to ALI-cultured cells, the coefficient of variation (CV) of repeated drug dosing is <11%-rel. and as little as 1 μL of drug/insert (=0.25 μL/cm 2 ) can be delivered to ALI-cultured cells. As proof of concept for dose-controlled in vitro drug testing, we demonstrated target engagement of a LIM and Calponin Homology domain 1 (LIMCH1)-specific ASO in primary lung fibroblasts from Idiopathic Pulmonary Fibrosis (IPF) patients "shielded" by an A549 alveolar epithelial layer mimicking inhalation therapy. LIMCH1-ASO was effective in LIMCH1-silencing even at the lowest dose (0.64 nmol/cm 2 ). Nebulization did neither impair LIMCH1-ASO efficacy nor induce cytotoxicity for the highest tested dose (3.21 nmol/cm 2 ). The permeability (biokinetics) of nebulized ASO through the A549 cell barrier was 14-fold faster ("burst-like") than for larger volume (pipetted) ASO application. Computational modeling revealed that this well-known "burst-like" pharmaco-/biokinetics profile is due to small-volume drug application, not due to nebulizer-induced changes on barrier tightness (apparent permeability P app ). Thus, the Cloud MAX is a substance-efficient platform for in vitro pharmacokinetics/-dynamics testing of inhalable drugs for targeted pulmonary therapy.

Abstract from PubMed Central (PMID 42999459, PMC13743522). This entry was added automatically by our daily literature monitor because it matches the topics we follow; read the full paper at the original source.