Frequency of F508del Variant in Patients with Cystic Fibrosis from Paraguay

Abstract Cystic fibrosis (CF) is an autosomal recessive disorder and is caused by variants in the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) gene. We aimed to study the frequency of the F508del variant, the most common variant worldwide, in patients with CF from Paraguay. The frequency of the F508del variant in Paraguayan patients with a clinical diagnosis of CF was assessed using a polymerase chain reaction followed by the sequencing of the PCR products. 43 of the 86 patients (50%) were homozygous for the F508del variant, 28 were heterozygous (32.56%), and the remaining 15 (17.44%) were non-carriers. In terms of alleles, there were 114 mutated (114/172 or 66.28%) and 58 did not correspond to this variant (58/172 or 33.72%). This is the first study of the frequency of the F508del variant in patients with CF in Paraguay. This information is of utmost relevance when planning and offering treatments from health services.

Identificación de la mutación ΔF508 en afectados con íleo meconial

Objetivo: Identificar la mutación ΔF508 en pacientes con íleo meconial. Ambiente: En el Instituto de Investigaciones Genéticas de la Facultad de Medicina de la Universidad del Zulia. Maracaibo. Métodos: Se estudiaron diez pacientes con ileo meconial. La detección de la mutación ΔF508 se realizó a partir de amplificación por reacción en cadena de la polimerasa de un segmento del gen de fibrosis quística de 98 pares de bases que contiene el codón que codifica a la fenilalanina en la posición 508 y el cual está ausente en los que tienen la mutación. Resultados: Se detectó la mutación ΔF508 en ambos alelos del gen de la fibrosis quística en tres pacientes, en un solo alelo en cinco y en dos no se identificó el alelo ΔF508 en su patrón molecular. Conclusión: El íleo meconial fue el marcador que sugirió el diagnóstico de fibrosis quística y permitió el asesoramiento genético de las familias al confirmar la presencia de la mutación ΔF508.

Objective: To perform ΔF508 mutation in patients with meconium ileus. Setting: In the Genetic Research Institute of the Faculty of Medicine. University of Zulia. Maracaibo. Methods: We studied 10 patients with meconium ileus. Detection of the mutation was performed from the amplification of a 98 pair of bases cystic fibrosis gene segment which contains the codon that encodes fenilalanine in the 508 position by polymerase chain reaction. This amplified product is absent in those who have the mutation. Results: The ΔF508 mutation was detected in both alleles of the cystic fibrosis gene in 3 patients, 5 were heterozygous for this mutation and in two patients were undetectable. Conclusion: Meconium ileus was the marker that suggested the diagnosis of cystic fibrosis and allowed the genetic counseling in this family to confirm the presence of the ΔF508 mutation.

Prevalência da mutação ΔF508 no gene cystic fibrosis transmembrane conductance regulator em pacientes com fibrose cística em um centro de referência no Brasil / Prevalence of ΔF508 mutation in the cystic fibrosis transmembrane conductance regulator gene among cystic fibrosis patients from a Brazilian referral center

OBJETIVO: Verificar a presença da mutação ΔF508 no gene cystic fibrosis transmembrane conductance regulator na população de pacientes com fibrose cística, diagnosticados pelo teste de sódio e cloro no suor, em acompanhamento no Ambulatório de Pneumologia Pediátrica da Universidade Estadual de Campinas, centro de referência no tratamento da fibrose cística. MÉTODOS: Foram analisadas 167 amostras de DNA de pacientes com fibrose cística. O genótipo dos pacientes foi determinado pela técnica de reação da polimerase e realizado cálculo para a frequência dos alelos e genótipos da mutação ΔF508. RESULTADOS: A frequência genotípica encontrada foi, respectivamente, para os genótipos -/-, ΔF508/- e ΔF508/ΔF508: 43,7% (73 pacientes), 32,9% (55 pacientes) e 23,4% (39 pacientes). Do total de 334 alelos analisados, foi observada a frequência de 201 (60,18%) alelos para a ausência da mutação ΔF508 e de 133 (39,82%) para a presença da mutação ΔF508. O cálculo do equilíbrio de Hardy-Weinberg foi realizado, e obtivemos o valor de qui-quadrado = 16,34 (p < 0,001). A população analisada está fora do equilíbrio. Os valores esperados são, para os respectivos genótipos -/-, ΔF508/- e ΔF508/ΔF508: 32,22% (60,48 pacientes), 47,93% (80,04 pacientes) e 15,86% (26,48 pacientes). CONCLUSÕES: Na população analisada, a mutação ΔF508 se mostrou menos prevalente em relação ao alelo sem a mutação. A frequência encontrada neste estudo foi semelhante à de outras regiões do Brasil e do mundo, principalmente devido à origem predominantemente caucasoide da população incluída no estudo.

OBJECTIVE: To verify the presence of ΔF508 mutation in the cystic fibrosis transmembrane conductance regulator gene among patients with cystic fibrosis diagnosed by the sweat test for sodium and chlorine and followed at the Pediatric Pneumology Outpatient Clinic of Universidade Estadual de Campinas, Brazil, a referral center for the treatment of cystic fibrosis. METHODS: The study analyzed 167 DNA samples from cystic fibrosis patients. Patients' genotype was determined by polymerase chain reaction, and allele and genotype frequencies of ΔF508 mutation were calculated. RESULTS: The genotype frequencies found for -/-, ΔF508/-, and ΔF508/ΔF508 genotypes were respectively: 43.7% (73 patients), 32.9% (55 patients), and 23.4% (39 patients). Of the 334 alleles analyzed, we observed a frequency of 201 (60.18%) alleles for the absence of ΔF508 mutation and of 133 (39.82%) for the presence of ΔF508 mutation. Hardy-Weinberg equilibrium was calculated, obtaining a chi-square value = 16.34 (p < 0.001). The study population was out of equilibrium. The expected values for -/-, ΔF508/-, and ΔF508/ΔF508 genotypes were respectively: 32.22% (60.48 patients), 47.93% (80.04 patients), and 15.86% (26.48 patients). CONCLUSIONS: In the analyzed population, ΔF508 mutation was less prevalent than the allele without this mutation. The frequency observed in this study was similar to that from other areas in Brazil and in the world, mainly due to the predominantly Caucasian origin of the population included in the study.

Utilidade da concentração da elastase-1 fecal monoclonal na avaliação da função pancreática nos pacientes com fibrose cística / Use of monoclonal faecal elastase-1 concentration for pancreatic status assessment in cystic fibrosis patients

OBJETIVO: Avaliar a concentração da elastase-1 (EL-1) fecal em pacientes pediátricos com fibrose cística, portadores da mutação ∆F508. MÉTODOS: Estudo transversal com amostras colhidas consecutivamente de 51 pacientes com idade entre 4 meses e 17 anos (média 9,11±4,74), sendo 32 (62,8 por cento) pacientes do sexo masculino. Houve coleta de dados clínico-demográficos e do tipo de mutação. A insuficiência pancreática exócrina foi definida pela atividade da EL-1 fecal < 200 µg/g. A quantificação da EL-1 foi realizada pelo método ELISA monoclonal (ScheBo Biotech AG, Germany). A suplementação pancreática foi utilizada em 46 (90,2 por cento) pacientes. RESULTADOS: Quarenta e um (80,4 por cento) pacientes apresentaram insuficiência pancreática (EL-1 fecal < 100 µg/g), sendo 17 (41,5 por cento) homozigotos, 14 heterozigotos (34,1 por cento) e 10 sem ∆F508 (24,4 por cento). Ao considerar a mutação, houve associação estatisticamente significativa entre os homozigotos e a concentração da EL-1 fecal < 100 µg/g (p = 0,010). Todos os pacientes considerados insuficientes pancreáticos (n = 41) pelo teste utilizavam suplemento pancreático. Dez (19,6 por cento) apresentaram EL-1 fecal > 200 µg/g, e 5/10 (50 por cento) utilizavam enzimas. CONCLUSÕES: A atividade de EL-1 fecal < 100 µg/g, indicativa de insuficiência pancreática, apresentou-se em 17/17 (100 por cento) dos homozigotos, conforme o esperado, sendo menos frequente nos heterozigotos para ∆F508 e nos pacientes com ausência dessa mutação. Não houve relação entre a concentração da EL-1 fecal com idade e sexo dos pacientes. O teste foi padronizado, é de fácil execução e poderá ser utilizado para avaliação da função pancreática dos pacientes com fibrose cística.

OBJECTIVE: To assess the concentration of faecal elastase-1 (EL-1) in pediatric patients with cystic fibrosis with mutation ∆F508. METHODS: Cross-sectional study with samples collected consecutively from 51 patients aged 4 months to 17 years old (mean 9.11±4.74); 32 (62.8 percent) patients were male. Clinical-demographic data were collected, as well as data on the type of mutation. Exocrine pancreatic insufficiency was established by the activity of faecal EL-1 < 200 µg/g. EL-1 was quantified through the monoclonal ELISA method (ScheBo Biotech AG, Germany). Pancreatic supplements were used in 46 (90.2 percent) patients. RESULTS: Forty-one (80.4 percent) patients presented with pancreatic insufficiency (EL-1 fecal < 100 µg/g): 17 (41.5 percent) were homozygous, 14 were heterozygous (34.1 percent) and 10 were non-∆F508 (24.4 percent). Regarding the mutation, there was a statistically significant association of homozygosity with faecal EL-1 concentration < 100 µg/g (p = 0.010). All patients considered to be pancreatic insufficient (n = 41) by the test were using pancreatic supplements. Ten (19.6 percent) presented faecal EL-1 > 200 µg/g, and 5/10 (50 percent) used enzymes. CONCLUSIONS: The activity of faecal EL-1 < 100 µg/g, indicating pancreatic insufficiency, was observed in 17/17 (100 percent) of homozygous patients, as expected, and was less frequent in patients who were heterozygous for ∆F508 and in patients without the mutation. There was no association of faecal EL-1 concentration with age and sex of patients. The test was standardized, is easy to execute, and can be used to assess the pancreatic status of patients with cystic fibrosis.

Frequency of 8 CFTR gene mutations in cystic fibrosis patients in Minas Gerais, Brazil, diagnosed by neonatal screening

The nature and frequency of cystic fibrosis mutations in Brazil is not uniform due to the highly varied ethnic composition of the population. The average frequency of the F508del mutation has been reported to be 48.6 percent. Other common mutations in Brazil are G542X, R1162X, and N1303K. The aim of this study was to analyze the frequency of 8 mutations (F508del, G542X, R1162X, N1303K, W1282X, G85E, 3120+1G>A, and 711+1G>T) in a sample of 111 newborn patients with cystic fibrosis diagnosed by the Cystic Fibrosis Neonatal Screening Program of Minas Gerais State. The mutations were tested by allele-specific oligonucleotide PCR with specially designed primers. An allele frequency of 48.2 percent was observed for the F508del mutation, and allele frequencies of 5.41, 4.50, 4.05, and 3.60 percent were found for the R1162X, G542X, 3120+1G>A, and G85E mutations, respectively. The genotypes obtained were in Hardy-Weinberg equilibrium. These data demonstrate that the 8-mutation panel studied here has extensive coverage (68 percent) for the cystic fibrosis mutations in Minas Gerais. These data improve our knowledge of cystic fibrosis in Brazil, particularly in this region. In addition, this investigation contributed to the establishment of a sensitive and population-specific mutation panel, which can be helpful for molecular diagnosis of cystic fibrosis.

p.F508del in a heterogeneous cystic fibrosis population from Minas Gerais, Brazil

Cystic fibrosis (CF) is the most common autosomal recessive disease of the Caucasian population. Among the various CF mutations, p.F508del is the most frequent, accounting for two-thirds of the global CF chromosomes, although showing great variability among populations. We have studied 115 unrelated CF patients from a mixed population of Minas Gerais (Brazil). To evaluate part of the DNA sequence of the cystic fibrosis transmembrane conductance regulator (CFTR) gene, blood DNA was obtained and PCR was performed using two pairs of primers that anneal to exons 10 and 24 of the CFTR gene. The PCR product was then submitted to automatic sequencing using the ABI PRISM 310 Genetic Analyzer. The p.F508del mutation was found in 50 (21.7 percent) of 230 unrelated CF alleles. Fifteen (13.0 percent) patients were homozygous for this mutation, while 20 (17.4 percent) were heterozygous; the remaining 80 (69.6 percent) patients did not carry the p.F508del mutation. Exon 24 sequence had no change in 75 (65.2 percent) patients, 21 (18.3 percent) had the sequence variation 4521G/A, 11 (9.6 percent) had a not yet described sequence variation 4407T/A and 8 (7.0 percent) patients had both sequence variations (4521G/A and 4407T/A). The polymorphism 4407T/A results in an amino acid modification from aspartic acid to glutamic acid, which will probably have no function effect in CFTR. This low p.F508del prevalence can be due to the variable ethnic origin of this population from Minas Gerais, which may have a high diversity of CF rare mutations.

Frecuencia de la mutación F508del en estudiantes de la Facultad de Medicina de la Universidad del Rosario, Bogotá, Colombia / Carrier frequency of F508del mutation of cystic fibrosis in medical students from Universidad del Rosario, Bogotá, Colombia

Introducción: La fibrosis quística es la enfermedad letal autosómica recesiva más frecuente en caucásicos, donde su incidencia es de 1 en 2000 nacidos vivos. Se debe a mutaciones en el gen CFTR, de las cuales la más frecuente es la F508del presente en 66% de los enfermos y en 1 de cada 25 personas sanas de origen caucásico. Objetivo: Identificar la tasa de portadores de la mutación F508del en una muestra de estudiantes de la Facultad de Medicina de la Universidad del Rosario.Materiales y métodos: Se determinó la presencia de la mutación F508del mediante PCR y análisis de heterodúplex en 110 estudiantes de IV y VII semestre de la Facultad de Medicina. Resultados: Se obtuvieron 4 heterocigotos para la mutación F508del, es decir, una frecuencia de portadores de 1 en 27 estudiantes. Conclusiones: La frecuencia de portadores de esta mutación en la población analizada es considerable, e indica que hay un alto número de personas en riesgo de heredar la enfermedad y que deben recibir asesoramiento genético. Es necesario aumentar el tamaño de la muestra para obtener datos representativos de la población colombiana.

Introduction: Cystic fibrosis (CF) is the most frequent autosomical recessive disorder in Caucasian population with an incidence of 1 in 2000 newborns. The disease is caused by mutations in the cftr gene, but the most common mutation is F508del, which accounts for 66% of CF chromosomes worldwide and a carrier frequency for Caucasian population of 1 in 25. Objective: To determine the carrier frequency of the F508del mutation in 110 unrelated, healthy students from the Facultad de Medicina, Universidad del Rosario. Methods: The presence of F508del mutation using PCR and heteroduplex analysis was determined. Results: Only four heterozygotes for F508del mutation were discovered. This represents a carrier frequency of 1 in 27 students. Conclusions: This estimated frequency of F508del carriers is higher than expected, encouraging further screening in normal control individuals from different regions of Colombia.

Cystic Fibrosis In India : Report Of 30 Cases And Frequency Of Delta F508 Mutation.

Cystic fibrosis (CF) is the most common potentially lethal disorder of Caucassians. Its frequency in India is not known. We diagnosed 30 cases of CF based on sweat chloride testing. The clinical profile and frequency of delta F508 mutation was studied. The frequency of F508 mutation was 27% (16 chromosomes) which is much lower as compared to that patients in the Western world. Eight patients carried other mutations, three of them were new mutations.