ABCMdb

PMID: 11401894

Clancy JP, Bebok Z, Ruiz F, King C, Jones J, Walker L, Greer H, Hong J, Wing L, Macaluso M, Lyrene R, Sorscher EJ, Bedwell DM
Evidence that systemic gentamicin suppresses premature stop mutations in patients with cystic fibrosis.
Am J Respir Crit Care Med. 2001 Jun;163(7):1683-92., [PubMed]

Sentences

No. Mutations Sentence Comment

61 ABCC7 p.Gly542* ABCC7 p.Gly542* ABCC7 p.Gly542* ABCC7 p.Gly542* Nasal cells from a G542X/ ⌬F508 patient (A, B, C) and a ⌬F508/⌬F508 patient (E, F ) in increasing concentrations of gentamicin: (A) G542X/⌬F508 0 ␮gm/mlgentamicin;(B)G542X/ ⌬F508 10 ␮gm/ml gentamicin; (C) G542X/⌬F508 100 ␮gm/ ml gentamicin; (D) G542X/⌬F508 no gentamicin, no primary antibody (control); (E) ⌬F508/⌬F508, no gentamicin; (F) ⌬F508/⌬F508, 100 ␮gm/ml gentamicin. Login to comment 86 ABCC7 p.Arg553* ABCC7 p.Gly542* ABCC7 p.Gly542* ABCC7 p.Gly542* STUDY PATIENT DEMOGRAPHICS* Age/Sex Premature Stop CF Subjects Control CF Subjects Genotype P.a FEV1 (% pred) Age/Sex Genotype P.a FEV1 (% pred ) 18/M G542X/⌬F508 (ϩ) 52 23/F ⌬F508/⌬F508 (ϩ) 38 18/M G542X/⌬F508 (ϩ) 76 18/F ⌬F508/⌬F508 (ϩ) 29 15/F W128X/⌬F508 (ϩ) 105 30/M ⌬F508/G55ID (ϩ) 19 28/M G542X/⌬F508 (ϩ) 25 26/F ⌬F508/⌬F508 (ϩ) 53 13/F R553X/621G-T (ϩ) 40 28/F ⌬F508/⌬F508 (ϩ) 34 18.4 yr (5.77)† 59.6 (31.5)† 25 yr (4.69)† 34.6 (11.8)† * Age (yr, mean Ϯ SD), sex (M ϭ male, F ϭ female), bacterial colonization (P.a ϭ Pseudomonas aeruginosa), FEV1 (forced expiratory volume in 1 s, %pred. Login to comment 90 ABCC7 p.Gly542* (A) Cells from a G542X/⌬F508 patient (open symbols) and a ⌬F508/⌬F508 patient (closed symbols) were isolated, grown, and loaded with SPQ as described in METHODS. Login to comment 94 ABCC7 p.Gly542* A positive response was obtained for 5/44 G542X/⌬F508 cells treated with 10 ␮gm/ml gentamicin and 42 of 44 cells treated with 100 ␮gm/ml gentamicin, and the means of these responding cells are plotted (Ϯ S.E.M.). Login to comment 95 ABCC7 p.Gly542* No responding cells were identified in the G542X/⌬F508 cells without gentamicin treatment (n ϭ 40), or any of the ⌬F508/⌬F508 cells (n Ͼ 80 cells tested in each condition). Login to comment 97 ABCC7 p.Gly542* (B) The percent of positive cells in each condition are shown for the ⌬F508/⌬F508 (left) and G542X/ ⌬F508 cells (right) in the increasing concentrations of gentamicin. Login to comment 112 ABCC7 p.Gly542* To investigate the effects of gentamicin on CFTR biosynthesis in human tissues, nasal cells from a G542X/⌬F508 patient with CF were grown on glass coverslips and exposed to medium with gentamicin at 0, 10, and 100 ␮g/ml for 16 h followed by immunohistochemical staining for surface-localized CFTR. Login to comment 114 ABCC7 p.Gly542* Figures 1A-1C identify CFTR antigen at the cell membrane from G542X/⌬F508 nasal cells with increasing gentamicin concentrations. Login to comment 118 ABCC7 p.Gly542* In a parallel set of experiments, nasal cells isolated from a G542X/⌬F508 patient with CF were grown on glass coverslips and studied for evidence of halide transport, using the fluorescent dye SPQ. Login to comment 120 ABCC7 p.Gly542* Figure 2 shows that G542X nasal cells exposed to gentamicin at 10 and 100 ␮g/ml had increased halide efflux in response to stimulation with a cAMP-elevating cocktail, compared with cells with no gentamicin exposure. Login to comment 122 ABCC7 p.Arg553* ABCC7 p.Gly542* Similar (immunocytochemical and functional) results have been observed for airway cells derived from other patients with premature stop mutations (G542X/⌬F508 and R553X/⌬F508 ([23] and our unpublished observations). Login to comment 179 ABCC7 p.Trp1282* ABCC7 p.Trp1282* Previous in vitro studies using the IB3-1 human CF bronchial airway cell line (genotype W1282X/⌬F508) demonstrated that a relatively short incubation (18-24 h) with the aminoglycoside G-418 at concentrations of 100-200 ␮g/ml restored chromosomal W1282X mRNA levels to that of the control ⌬F508 allele, induced surface-localized CFTR protein production, and conferred on cells a new, cAMP-activated [Cl- ] current (13). Login to comment 180 ABCC7 p.Trp1282* ABCC7 p.Arg553* ABCC7 p.Gly542* ABCC7 p.Arg1162* Similar observations were made with transient expression systems for other CF-associated premature stop mutations, including G542X, R553X, and R1162X mutations, in addition to W1282X (12, 13). Login to comment 196 ABCC7 p.Arg117His ABCC7 p.Ala455Glu ABCC7 p.Arg334Trp Reports indicate that some CF patients with uncommon alleles (i.e., conduction mutants such as R117H or R334W, or the uncommon A455E allele) may have abnormally elevated sweat [Cl- ] values diagnostic of CF but lower than those of the general CF population (9, 25-27). Login to comment