Comprehensive mutation screening in a cystic fibrosis center

Objectives and Background. The identities of a cystic fibrosis (CF) patient 's CFTR mutations can influence therapeutic strategies, but because >800 CF TR mutations exist, cost-effective, comprehensive screening requires a mult istage approach. Single-strand conformation polymorphism and heteroduplex a nalysis (SSCP/HA) can be an important part of mutation detection, but must be calibrated within each laboratory. The sensitivity of a combined commerc ial-SSCP/HA approach to genotyping in a large, ethnically diverse US center CF population has not been established. Study Design. We screened all 27 CFTR exons in 10 human participants who ha d an unequivocal CF diagnosis including a positive sweat chloride test and at least 1 unknown allele after commercial testing for the 70 most common m utations by SSCP/HA. These participants were compared with 7 participants w ho had negative sweat tests but at least 1 other CF-like symptom meriting c omplete genotyping. Results. For the 10 CF participants, we detected 11 of 16 unknown alleles ( 69%) and all 4 of the known alleles (100%), for an overall rate of 75% inpa tients not fully genotyped by conventional 70 mutation screen. For 7 partic ipants with negative sweat tests, we confirmed 1 identified mutation in 14 alleles and detected 3 additional mutations. Mutations detected in both gro ups included 7 missense mutations (S13F, P67L, G98R, S492F, G970D, L1093P, N1303K) and 9 deletion, frameshift, nonsense or splicing mutations (R75X, G 542X, Delta F508, 451-458 Delta8 bp, 5T, 663 DeltaT, exon 13 frameshift, 12 61+1G-->A and 3272-26A-->G). Three of these mutations were novel (G970D, L1 093P, and 451-458 Delta8 bp(1)). Thirteen other changes were detected, incl uding the novel changes 1812-3 ins T, 4096-278 ins T, 4096-265 ins TG, and 4096-180 T-->G. Conclusion. When combined with the 70 mutation Genzyme test, SSCP/HA analys is allows for detection of >95% of the mutations in an ethnically heterogen eous CF center population. We discuss 5 possible explanations that could ac count for the few remaining undetected mutations.