235 Reducing MUC5AC or MUC5B Expression Using Antisense Oligonucleotides As a Therapeutic Approach for Cystic Fibrosis and Other Muco-Obstructive Diseases

likely to work when applied to larger super-exon vectors that would be designed to rescue most CFTR mutations and help determine the relevance of CF mice and CF mouse organoids for preclinical testing of CFTR superexon therapeutic strategies.Methods: We tested six constructs for restoration of CFTR function in W1282X mouse intestinal organoids.The constructs contained human super-exon 23-27 or mouse super-exon 23-27, combined with a human CFTR or rabbit beta globin splice acceptor and a bovine growth hormone (BGH) or a simian virus 40 (SV40) polyadenylation signal sequence.CFTR function restoration was assessed using a forskolin-induced swelling (FIS) assay.To create a CF mouse with a human super-exon, we introduced donor sequences containing human CFTR super-exon 23-27 into mouse intron 22 upstream of a W1282X mutation.Results: In W1282X mutant mouse intestinal organoids, two of the six constructs fully restored CFTR function: human super-exon 23-27 and mouse super-exon 23-27 with a rabbit beta globin splice acceptor with a SV40 polyadenylation sequence.We are continuing to assess the other four constructs' potential to restore CFTR function.To test whether a super-exon would ameliorate CF disease, we have established a line of mice with human super-exon 23-27 integrated into mouse Cftr intron 22, but because the W1282X mutation in one of the homology arms of the construct did not integrate into mouse exon 23, we are creating the W1282X mutation in the human super-exon mouse line.Once completed, we will assess whether the presence of the super-exon alleviates CF manifestations in the CF mouse.Conclusions: For at least one super exon construct, a human and a mouse super-exon 23-27 can fully restore CFTR function in W1282X mouse intestinal organoids.We have established a line of mice with human superexon 23-27 and will assess the ability of this to ameliorate CF manifestations.