ABCMdb

PMID: 9154877

Walker LC, Venglarik CJ, Aubin G, Weatherly MR, McCarty NA, Lesnick B, Ruiz F, Clancy JP, Sorscher EJ
Relationship between airway ion transport and a mild pulmonary disease mutation in CFTR.
Am J Respir Crit Care Med. 1997 May;155(5):1684-9., [PubMed]

Sentences

No. Mutations Sentence Comment

1 ABCC7 p.Ala455Glu We studied airway bioelectric properties in five CF patients with the rare A455E mutation that is associated with mild pulmonary disease. Login to comment 2 ABCC7 p.Arg553* ABCC7 p.Gly542* We also evaluated five patients possessing premature truncation mutations (G542X and R553X) for which an association with mild pulmonary disease has not been as well established. Login to comment 5 ABCC7 p.Ala455Glu Based on our findings in a naturally occurring group of CF patients with an improved pulmonary prognosis (A455E), one can argue that marked clinical benefit might be possible without any improvement whatsoever in airway bioelectric phenotype. Login to comment 6 ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu Moreover, if genetic modifiers exist that influence the severity of a particular CFTR mutation (e.g., A455E), these may be independent of human airway Cl secretion in vivo, since we detected minimal Cl- -secretory responses in patients with A455E. Login to comment 19 ABCC7 p.Arg553* ABCC7 p.Gly542* As a control, we measured airway ion transport in patients with the common R553X and G542X truncation mutations. Login to comment 20 ABCC7 p.Arg553* ABCC7 p.Gly542* As a control, we measured airway ion transport in patients with the common R553X and G542X truncation mutations. Login to comment 24 ABCC7 p.Arg553* ABCC7 p.Gly542* The clinical features of the A455E, R553X, and G542X CF patients we studied are given in Table 1. Login to comment 25 ABCC7 p.Arg553* ABCC7 p.Gly542* The clinical features of the A455E, R553X, and G542X CF patients we studied are given in Table 1. Login to comment 30 ABCC7 p.Gly551Asp ABCC7 p.Arg553* ABCC7 p.Gly542* ABCC7 p.Gly542* ABCC7 p.Gly542* ABCC7 p.Gly542* ABCC7 p.Gly542* Age (yr) Sex Genotype Pseudomonas Colonization % Predicted FVC 96 Predicted FEV, Sweat C1 (meq/L) PS versus PI* 1 121/2 F A455E/621 + 1G-T + 113 106 106 P1 2 111/2 F A455E/621 + 1G-*T - 109 101 97 PI 3 16 M AF508/A455E - 77 63 84 PS 4 16 F AF508/A455E - 123 120 59 PS 5 151/2 F AF508/A455E + 101 94 54 PS 6 21 M G551D/G542X + 56 26 84 PI 7 18 M AF508/G542X + 90 80 80 PI 8 16 M AF508/G542X + 121 110 132 PI 9 16 F R553X/G542X + 68 46 78 PI 10 18 M AF508/G542X + 51 41 100 PI 1685 * PS = pancreatic sufficient; PI = pancreatic insufficient terfly needle filled with a 4% agar/lactated Ringer's solution) was inserted subcutaneously in the dorsal aspect of the distal forearm or hand. Login to comment 31 ABCC7 p.Gly551Asp ABCC7 p.Arg553* ABCC7 p.Gly542* ABCC7 p.Gly542* ABCC7 p.Gly542* ABCC7 p.Gly542* ABCC7 p.Gly542* Age (yr) Sex Genotype Pseudomonas Colonization % Predicted FVC 96 Predicted FEV, Sweat C1 (meq/L) PS versus PI* 1 121/2 F A455E/621 + 1G-T + 113 106 106 P1 2 111/2 F A455E/621 + 1G-*T - 109 101 97 PI 3 16 M AF508/A455E - 77 63 84 PS 4 16 F AF508/A455E - 123 120 59 PS 5 151/2 F AF508/A455E + 101 94 54 PS 6 21 M G551D/G542X + 56 26 84 PI 7 18 M AF508/G542X + 90 80 80 PI 8 16 M AF508/G542X + 121 110 132 PI 9 16 F R553X/G542X + 68 46 78 PI 10 18 M AF508/G542X + 51 41 100 PI 1685 * PS = pancreatic sufficient; PI = pancreatic insufficient terfly needle filled with a 4% agar/lactated Ringer's solution) was inserted subcutaneously in the dorsal aspect of the distal forearm or hand. Login to comment 44 ABCC7 p.Ala455Glu Expression of CFTR in COS-7 (Simian Kidney) Cells Expression of wild type, AF508 or A445E CFTR under the regulatory control of the 17 promoter was studied with the pTM1 vector (gift of Dr. B. Moss, of the National Institutes of Health; the A455E construct in this vector backbone was a generous gift of Dr. M. Welsh of the University of Iowa). Login to comment 45 ABCC7 p.Ala455Glu Expression of CFTR in COS-7 (Simian Kidney) Cells Expression of wild type, AF508 or A445E CFTR under the regulatory control of the 17 promoter was studied with the pTM1 vector (gift of Dr. B. Moss, of the National Institutes of Health; the A455E construct in this vector backbone was a generous gift of Dr. M. Welsh of the University of Iowa). Login to comment 58 ABCC7 p.Ala455Glu ABCC7 p.Arg553* ABCC7 p.Gly542* RESULTS Table 1 summarizes clinical features of the A455E, G542X, and R553X CF patients we studied. Login to comment 59 ABCC7 p.Ala455Glu ABCC7 p.Arg553* ABCC7 p.Gly542* RESULTS Table 1 summarizes clinical features of the A455E, G542X, and R553X CF patients we studied. Login to comment 61 ABCC7 p.Ala455Glu Taken together, there had been a total of 10 hospital admissions for intravenous antibiotic therapy among the A455E patients, and more than 36 admissions among the patients with truncation mutations. Login to comment 62 ABCC7 p.Ala455Glu Taken together, there had been a total of 10 hospital admissions for intravenous antibiotic therapy among the A455E patients, and more than 36 admissions among the patients with truncation mutations. Login to comment 66 ABCC7 p.Ala455Glu ABCC7 p.Arg553* ABCC7 p.Arg553* ABCC7 p.Gly542* ABCC7 p.Gly542* Decisions regard- -10 26t5 mV -39*9 mVtt -57&#b1;5 mVtt -43*9 mVtt Control G542X or R553X A544E AF508 B 50 40 ߦ C A PD 30 ߦ (mV) ߦ 20 ߦ ߦ ߦ 10 ' ߦ ߦ o - t---------------------------- - aj J0 mVtt -27&#b1;8 mVtt Control G542X or R553X A455E AF508 C -40 -30 1686 AMERICAN JOURNAL OF RESPIRATORY AND CRITICAL CARE MEDICINE VOL. 155 1997 A -60 -50 ߦ t 1 ' PD max -40 I (mV) ߦ ߦ ߦ -30 ߦ ߦ s -20 to mV L - - i--- --- - -- -------loses A Amilo,id Amilonde Amiloride Low CI' Low Cl* fenol lomV L l0 Sea --------------^--------- --------t- - --- -- - --------- l0 mV L ------^--- --------Y- -^ t t lOmV L 10Sa. Login to comment 67 ABCC7 p.Ala455Glu ABCC7 p.Arg553* ABCC7 p.Arg553* ABCC7 p.Gly542* ABCC7 p.Gly542* Decisions regard- -10 26t5 mV -39*9 mVtt -57±5 mVtt -43*9 mVtt Control G542X or R553X A544E AF508 B 50 40 • C A PD 30 • (mV) • 20 • •• 10 ' • • o - t----------------------------- aj J0 mVtt -27±8 mVtt Control G542X or R553X A455E AF508 C -40 -30 1686 AMERICAN JOURNAL OF RESPIRATORY AND CRITICAL CARE MEDICINE VOL. 155 1997 A -60 -50 • t 1 ' PD max -40 I (mV) • • • -30 • • s-20 to mV L - - i--- --- - -- -------loses A Amilo,id Amilonde Amiloride Low CI' Low Cl* fenol lomV L l0 Sea --------------^--------- --------t- - --- -- - --------- l0 mV L ------^--- --------Y- -^ t t lOmV L 10Sa. Login to comment 70 ABCC7 p.Ala455Glu ABCC7 p.Gly542* Genotypes of the patients shown include B:A455E/621 + 1 G-*T; C:AF508/A455E; and D:R553X/G542X. Login to comment 71 ABCC7 p.Ala455Glu ABCC7 p.Gly542* Genotypes of the patients shown include B:A455E/621 + 1 G-*T; C:AF508/A455E; and D:R553X/G542X. Login to comment 73 ABCC7 p.Ala455Glu Figure 1B and C show tracings from compound heterozygotes for the A455E mutation. Login to comment 74 ABCC7 p.Ala455Glu Figure 1B and C show tracings from compound heterozygotes for the A455E mutation. Login to comment 76 ABCC7 p.Ala455Glu ABCC7 p.Arg553* ABCC7 p.Gly542* The remainder of patients with the A455E mutation exhibited no response to low Cl- /isoproterenol, as in the tracing in Figure I.C. Figure 1D shows a tracing from a patient homozygous for truncation mutations within CFTR (G542X/R553X). Login to comment 77 ABCC7 p.Ala455Glu ABCC7 p.Arg553* ABCC7 p.Gly542* The remainder of patients with the A455E mutation exhibited no response to low Cl- /isoproterenol, as in the tracing in Figure I.C. Figure 1D shows a tracing from a patient homozygous for truncation mutations within CFTR (G542X/R553X). Login to comment 79 ABCC7 p.Ala455Glu ABCC7 p.Arg553* ABCC7 p.Gly542* Figure 2 shows a summary of the data from patients and healthy controls in the study, and describes the baseline PD during perfusion with lactated Ringer's solution (Figure 2A), -20 APD (mv) ߦ -10 tl ߦ 0ߦ------' ------------------s---------------------------------- --- 10 12t7 mV -2:4 rVtt -4t8 mVtt -5)mVtt Control G542X or R553X A455E AF508 Figure 2. Login to comment 80 ABCC7 p.Ala455Glu ABCC7 p.Arg553* ABCC7 p.Gly542* Figure 2 shows a summary of the data from patients and healthy controls in the study, and describes the baseline PD during perfusion with lactated Ringer's solution (Figure 2A), -20 APD (mv) • -10 tl • 0•------'------------------s---------------------------------- --- 10 12t7 mV -2:4 rVtt -4t8 mVtt -5)mVtt Control G542X or R553X A455E AF508 Figure 2. Login to comment 88 ABCC7 p.Ala455Glu In B, both A455E and AF508 differed from the truncation mutation group (p < 0.01). Login to comment 89 ABCC7 p.Ala455Glu In B, both A455E and AF508 differed from the truncation mutation group (p < 0.01). Login to comment 91 ABCC7 p.Ala455Glu When data from each experimental group are taken together, neither A455E patients nor those with truncation mutations showed evidence of either a Cl- -secretory response (Figure 2C) or a suggestion of normalization of this aspect of the PD measurement. Login to comment 92 ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu When data from each experimental group are taken together, neither A455E patients nor those with truncation mutations showed evidence of either a Cl- -secretory response (Figure 2C) or a suggestion of normalization of this aspect of the PD measurement. Login to comment 93 ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu ABCC7 p.Arg553* ABCC7 p.Gly542* In the A455E patients, the maximal PD during perfusion with lactated Ringer's solution and the change after amiloride were well within the range anticipated for the general CF population (Figure 2A and B; [6-21]). Login to comment 94 ABCC7 p.Ala455Glu ABCC7 p.Arg553* ABCC7 p.Arg553* ABCC7 p.Gly542* In the patients with CFTR truncations G542X or R553X, the magnitude of the amiloride-sensitive PD (Figure 2B) was less than in A455E or AF508 patients, although these measurements were still highly useful in discriminating all CF patient groups from individuals without CF (p < 0.01). Login to comment 95 ABCC7 p.Arg553* In vitro data suggesting that R553X CFTR is capable of downregulating amiloride-sensitive sodium transport during oocyte coexpression have recently been reported (31). Login to comment 97 ABCC7 p.Ala455Glu On the basis of these in vivo experiments, we were interested in studying the in vitro activity of CFTR containing the A455E mutation. Login to comment 98 ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu On the basis of these in vivo experiments, we were interested in studying the in vitro activity of CFTR containing the A455E mutation. Login to comment 99 ABCC7 p.Ala455Glu During in vitro overexpression in heterologous cells, the A455E mutation has been reported to exhibit partial Cl- -channel activity (7). Login to comment 100 ABCC7 p.Ala455Glu The results indicate that cAMP-dependent increases in halide permeability occur after A455E CFTR overexpression in vitro, and are intermediate between halide permeability with wild-type and AF508 CFTR. Login to comment 101 ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu The results indicate that cAMP-dependent increases in halide permeability occur after A455E CFTR overexpression in vitro, and are intermediate between halide permeability with wild-type and AF508 CFTR. Login to comment 102 ABCC7 p.Ala455Glu DISCUSSION The A455E mutation in CFTR has been associated with improved FEV1 and FVC, a reduced incidence of pulmonary colonization by Pseudomonas aeruginosa, better exocrine pancreatic function, residual Cl- -channel activity in vitro, and improved overall prognosis in CF (2-7). Login to comment 104 ABCC7 p.Ala455Glu The incidence of A455E in both populations is believed to be based on a genetic founder effect. Login to comment 106 ABCC7 p.Ala455Glu The incidence of A455E in both populations is believed to be based on a genetic founder effect. Login to comment 107 ABCC7 p.Ala455Glu However, our results suggest that whether it is the A455E mutation itself or (as yet undetermined) interactions with other genes that lead to mild lung disease, the mitigating factor is independent of large improvements in either transepithelial Cl- or Na+ transport, since no such improvements were observed in our experiments. Login to comment 109 ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu However, our results suggest that whether it is the A455E mutation itself or (as yet undetermined) interactions with other genes that lead to mild lung disease, the mitigating factor is independent of large improvements in either transepithelial Cl- or Na+ transport, since no such improvements were observed in our experiments. Login to comment 110 ABCC7 p.Ala455Glu Functional assay of cAMP-mediated halide permeability in COS-7 cells transiently expressing wild-type (open circles), A455E (open triangles), or AF508 CFTR (filled diamonds). Login to comment 111 ABCC7 p.Ala455Glu It.ie not known whether the airway bioelectric measurement is sufficiently sensitive to identify a lower limit of gene transfer necessary to provide benefit in this setting, --A-- A455E (11/53) 0 eF508 (0/27) CFTR (42/42) S S S S S 0 0 S SCl C) er U) 10 h CD 0) SECONDS Figure 3. Login to comment 112 ABCC7 p.Ala455Glu Functional assay of cAMP-mediated halide permeability in COS-7 cells transiently expressing wild-type (open circles), A455E (open triangles), or AF508 CFTR (filled diamonds). Login to comment 115 ABCC7 p.Ala455Glu Approximately 20% of A455E cells (11 of 53) had a detectable cAMP response comparable with that of the wild-type CFTR. Login to comment 116 ABCC7 p.Ala455Glu Each curve is the mean &#b1; SEM of responding (wild-type CFTR or A455E) or total (&#c1;F508) cells. Login to comment 117 ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu Approximately 20% of A455E cells (11 of 53) had a detectable cAMP response comparable with that of the wild-type CFTR. Login to comment 118 ABCC7 p.Ala455Glu Each curve is the mean ± SEM of responding (wild-type CFTR or A455E) or total (ÁF508) cells. Login to comment 119 ABCC7 p.Ala455Glu A455E responses differed from those of AF508 (p < 0.001). Login to comment 122 ABCC7 p.Ala455Glu We also studied the A455E mutation in a recombinant overexpression system used to analyze the function of CFTR mutations (24-27). Login to comment 123 ABCC7 p.Ala455Glu The data presented in Figure 3 indicate that A455E retains residual activity when studied in vitro. Login to comment 124 ABCC7 p.Ala455Glu We also studied the A455E mutation in a recombinant overexpression system used to analyze the function of CFTR mutations (24-27). Login to comment 125 ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu The data presented in Figure 3 indicate that A455E retains residual activity when studied in vitro. Login to comment 127 ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu Together with our in vivo experiments, this result suggests that A455E is active following overexpression, but that the activity in vivo is not detectable by the nasal PD assay. Login to comment 129 ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu First, the nasal PD, the best available method for studying the CF bioelectric phenotype in vivo, may not be sufficiently sensitive to detect clinically important improvements in Cl- secretion in a small number of CF patients with an improved prognosis (i.e, five individuals with the A455E mutation). Login to comment 130 ABCC7 p.Ala455Glu For example, it is possible that detectable A455E C1 - -channel activity in isolated cells or membrane patches depends on a very high level of protein expression that is absent in vivo. Login to comment 131 ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu ABCC7 p.Ala455Glu Alternatively, since the levels of A455E J m m m A LL w y w V z 51 dequench cAMP AMERICAN JOURNAL OF RESPIRATORY AND CRITICAL CARE MEDICINE VOL. 155 1997 mRNA or protein in vivo are not known, it is possible that previous in vitro overexpression studies of A455E (and the results shown in Figure 3) reflect a phenomenon that does not predict activity of the mutation in vivo. Login to comment 132 ABCC7 p.Ala455Glu For example, it is possible that detectable A455E C1 - -channel activity in isolated cells or membrane patches depends on a very high level of protein expression that is absent in vivo. Login to comment 133 ABCC7 p.Ala455Glu Because PD measurements detect Cl- secretion across polarized epithelium, studies of A455E in nonpolarized cells grown on plastic might have quite different results from the in vivo situation. Login to comment