TY - JOUR
T1 - Targeted locus amplification reveals heterogeneity between and within CFTR genotypes and association with CFTR function in patient-derived intestinal organoids
AU - Lefferts, J. W.
AU - Boersma, V.
AU - Nieuwenhuijze, N. D.A.
AU - Suen, S. W.F.
AU - Hajo, K.
AU - Collantes, N. Sanchez
AU - Vermeulen, C.
AU - Groeneweg, T.
AU - Hagemeijer, M. C.
AU - de Jonge, H. R.
AU - van der Ent, C. K.
AU - Splinter, E.
AU - Beekman, J. M.
N1 - Funding Information:
This work was funded by Health Holland grant no. 171114 and the Dutch CF Foundation (NCFS) as part of the HIT-CF2 program.
Publisher Copyright:
© 2023
PY - 2023/5
Y1 - 2023/5
N2 - Background: Cystic fibrosis (CF) disease severity can be highly variable, even between people with CF (pwCF) with similar genotypes. Here we use patient-derived intestinal organoids to study the influence of genetic variation within the cystic fibrosis transmembrane conductance regulator (CFTR) gene on CFTR function. Methods: Organoids of F508del/class I, F508del/S1251N and pwCF with only one detected CF-causing mutation were cultured. Allele-specific CFTR variation was investigated using targeted locus amplification (TLA), CFTR function was measured using the forskolin-induced swelling assay and mRNA levels were quantified using RT-qPCR. Results: We were able to distinguish CFTR genotypes based on TLA data. Additionally, we observed heterogeneity within genotypes, which we were able to link to CFTR function for S1251N alleles. Conclusions: Our results indicate that the paired analysis of CFTR intragenic variation and CFTR function can gain insights in the underlying CFTR defect for individuals where the disease phenotype does not match the CFTR mutations detected during diagnosis.
AB - Background: Cystic fibrosis (CF) disease severity can be highly variable, even between people with CF (pwCF) with similar genotypes. Here we use patient-derived intestinal organoids to study the influence of genetic variation within the cystic fibrosis transmembrane conductance regulator (CFTR) gene on CFTR function. Methods: Organoids of F508del/class I, F508del/S1251N and pwCF with only one detected CF-causing mutation were cultured. Allele-specific CFTR variation was investigated using targeted locus amplification (TLA), CFTR function was measured using the forskolin-induced swelling assay and mRNA levels were quantified using RT-qPCR. Results: We were able to distinguish CFTR genotypes based on TLA data. Additionally, we observed heterogeneity within genotypes, which we were able to link to CFTR function for S1251N alleles. Conclusions: Our results indicate that the paired analysis of CFTR intragenic variation and CFTR function can gain insights in the underlying CFTR defect for individuals where the disease phenotype does not match the CFTR mutations detected during diagnosis.
UR - http://www.scopus.com/inward/record.url?scp=85153600055&partnerID=8YFLogxK
U2 - 10.1016/j.jcf.2023.04.003
DO - 10.1016/j.jcf.2023.04.003
M3 - Article
C2 - 37100706
AN - SCOPUS:85153600055
SN - 1569-1993
VL - 22
SP - 538
EP - 547
JO - Journal of Cystic Fibrosis
JF - Journal of Cystic Fibrosis
IS - 3
ER -