The CFTR Mutation c.3453G > C (D1152H) Confers an Anion Selectivity Defect in Primary Airway Tissue that Can be Rescued by Ivacaftor

Abstract

The Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) gene variant, c.3453G > C (D1152H), is associated with mild Cystic Fibrosis (CF) disease, though there is considerable clinical variability ranging from no detectable symptoms to lung disease with early acquisition of Pseudomonas aeruginosa. The approval extension of ivacaftor, the first CFTR modulator drug approved, to include D1152H was based on a positive drug response of defective CFTR-D1152H chloride channel function when expressed in FRT cells. Functional analyses of primary human nasal epithelial cells (HNE) from an individual homozygous for D1152H now revealed that while CFTR-D1152H demonstrated normal, wild-type level chloride conductance, its bicarbonate-selective conductance was impaired. Treatment with ivacaftor increased this bicarbonate-selective conductance. Extensive genetic, protein and functional analysis of the nasal cells of this D1152H/D1152H patient revealed a 90% reduction of CFTR transcripts due to the homozygous presence of the 5T polymorphism in the poly-T tract forming a complex allele with D1152H. Thus, we confirm previous observation in patient-derived tissue that 10% normal CFTR transcripts confer normal, wild-type level chloride channel activity. Together, this study highlights the benefit of patient-derived tissues to study the functional expression and pharmacological modulation of CF-causing mutations, in order to understand pathogenesis and therapeutic responses.

Preclinical Studies of a Rare CF-Causing Mutation in the Second Nucleotide Binding Domain (c.3700A>G) Show Robust Functional Rescue in Primary Nasal Cultures by Novel CFTR Modulators

“…I1234_R1239del/W1282X nasal epithelial cells were obtained through Dr. Carlos Milla at the CF Center at Stanford University, and I1234_R1239del homozygous nasal cells were obtained from family members at The Hospital for Sick Children (after obtaining informed consent). The subsequent nasal epithelial cell culture was performed as previously described [ 24 , 25 , 26 ]. Cells were seeded on collagen coated transwell inserts and once confluent, the cells were cultured for 14 days at an air liquid interface (ALI) with basal differentiation media (PneumaCult TM ALI, StemCell Tech., Vancouver, BC, Canada).…”

Personalized Medicine Based on Nasal Epithelial Cells: Comparative Studies with Rectal Biopsies and Intestinal Organoids

“…Recently, different cellular models have been developed to determine the response of CFTR mutants to modulators through a personalized CFTR pharmacotherapy approach [ 11 ]. The most common cellular models include primary human nasal (pHNE) and bronchial (pHBE) epithelial cells and intestinal organoids obtained from rectal biopsies [ 16 , 17 , 18 , 19 , 20 , 21 , 22 , 23 , 24 , 25 , 26 , 27 ]. Although pHBEs are the gold standard to test for CFTR modulator efficacy since they can recapitulate the in vivo morphology and key processes happening in lungs [ 28 ], intestinal organoids are the most advanced model in CF research, incorporating many physiological relevant tissue features [ 29 ].…”

Phenotyping Rare CFTR Mutations Reveal Functional Expression Defects Restored by TRIKAFTATM

“…On the other hand, the clinical response size is highly variable even in patients homozygous for F508del [ 7 ]. Therefore, we and others are examining the utility of modulator testing on patient-derived nasal cultures or rectal organoids to predict clinical response size [ 18 , 36 , 37 , 38 , 39 , 40 , 41 , 42 , 43 , 44 , 45 , 46 , 47 ].…”

“…After18h, the cells were treated with DMSO (0.1%), 3 µM VX-809, and 3 µM VX-661 + 3 µM VX-445 (Selleck Chemicals, Houston, TX, USA) for 24 h at 37 °C. Then, the cells were loaded with a blue membrane potential dye dissolved in chloride free buffer [ 18 ] for 35 min at 37 °C. The plate was then read in a fluorescence plate reader (SpectraMax i3; Molecular Devices, San Jose, CA, USA) at 37 °C.…”