O1085: Transamniotic Delivery of Cystic Fibrosis (CF) Transmembrane Conductance Regulator (CFTR) Mrna: A Potential Novel Strategy for the Perinatal Management of Cf-associated Meconium Ileus

Background: Mutations of the cystic fibrosis transmembrane conductance regulator (CFTR), a cell membrane protein serving as chloride/anion channel in numerous epithelial cells, constitute the central component of the pathophysiology of cystic fibrosis (CF) and chief cause of neonatal meconium ileus associated with this disease. Postnatal exogenous mRNA-based therapies for CF have been widely studied experimentally, having already reached clinical trials as inhalation therapies for its respiratory complications. It has been previously shown that the transamniotic route can be an effective means of delivering encapsulated exogenous mRNA to the fetus, as a variant of the transamniotic nucleic acid therapy (TRANAT) principle. In this study, we sought to determine whether TRANAT could be a viable alternative for the administration of CFTR mRNA to the fetal bowel as a potential strategy for the perinatal management of meconium ileus associated with CF.

Methods: After IACUC approval, pregnant Sprague Dawley dams (n=9) underwent midline laparotomy followed by volume-matched (50μL) intra-amniotic injections in all their fetuses (n=109) of either a suspension of a human CFTR mRNA encapsulated by a lipid- and synthetic cationic polymer-based composite, lipopolyplex (mRNA group; n=98), or of a suspension of the same encapsulation composite but free of mRNA (control group; n=11), on gestational day 17 (E17; term=E21-22). At daily time points until term for the mRNA group and at E21 for the control group, samples from fetal small bowel and lung were procured, along with maternal serum, for screening for the presence of the exogenous mRNA-encoded CFTR protein via quantification by ELISA. Statistical analysis was by nonparametric Wilcoxon rank sum test with Bonferroni-adjustment.

Results: Overall fetal survival was 85.3% (93/109), with no significant differences between the groups or time points (p>0.05 for all). When controlled by mRNA-free injections, human CFTR was detected in the small bowel from the mRNA group at all time points (p < 0.001 for all; figure 1A). The amount of intestinal CFTR produced increased over time, being highest at E20 (11840 ng/mL), i.e. three days after injection. Human CFTR was detected in the lungs from both groups, with even significantly decreased amounts at E18-19 in the mRNA group vs. control (p < 0.012 for both; figure 1B), suggesting the possibility of some degree of interspecies homology at least at this anatomical site. When controlled by mRNA-free injections, no human CFTR was detected in any maternal serum sample (p>0.02 for all).

Conclusion: Encapsulated exogenous mRNA encoding for the cystic fibrosis transmembrane conductance regulator protein can be incorporated and translated by fetal small bowel cells after simple intra-amniotic injection in a healthy rat model. Transamniotic mRNA delivery could become a novel strategy for the perinatal prevention or management of meconium ileus associated with cystic fibrosis.