Targeted Extended Cystic Fibrosis Mutation Testing On Known And At-risk Patients And Relatives
blood; cftr mutations; early-diagnosis; experience; Genetics & Heredity; immunoreactive trypsinogen; midlands; newborn; outcomes; Research & Experimental Medicine; vas-deferens; west; wisconsin
This paper reports mathematically derived residual risks of being a carrier or being affected with cystic fibrosis following various screening scenarios to assist in interpreting test results and advising patients. While parental screening with 23 American College of Medical Genetics (ACMG) cystic fibrosis mutations defines the 64% of affected U. S. Caucasian fetuses with two detectable mutations, newborn screening for elevated immunoreactive trypsinogen (IRT) and sweat chloride identifies an additional 36% of affected newborns with zero or one detected mutation. The relatives of these affected newborns with less than two detectable mutations have higher posterior (after) 23 mutation-negative test risks of carrying undetected mutations. These calculations emphasize how knowledge of the mutations in the related affected patient substantially improves upon the quality of after-test advice to patients. Furthermore, negative tests of the partner without a family history and/or more extensive cystic fibrosis transmembrane conductance regulator (CFTR) gene testing also increases the likelihood that a negative report is truly negative. When a newborn patient with zero or one detected CFTR mutation has an inconclusive sweat test result, the sweat test should be repeated before ordering additional often unnecessary CFTR gene sequencing. Given the same composite mutation panel test accuracy, a higher proportion of reported test results would be correct during parental screening than when testing at-risk fetuses or symptomatic newborns. Prenatal and newborn screening would be enhanced substantially by medical professionals offering copies of all positive parental and newborn test reports to the parents to share with their relatives. These principles are likely to be applicable to other genetic diseases as the most common mutation frequencies are reported.
Lebo R V; Omlor G J
Genetic Testing
2007
2007-12
Journal Article or Conference Abstract Publication
<a href="http://doi.org/10.1089/gte.2007.0050" target="_blank" rel="noreferrer noopener">10.1089/gte.2007.0050</a>
Variable Penetrance And Expressivity Of The Splice Altering 5t Sequence In The Cystic Fibrosis Gene
cftr gene; congenital bilateral absence; disease; Genetics & Heredity; males; messenger-rna; mutations; polymorphism; repeat; Research & Experimental Medicine; risk; vas-deferens
This manuscript reviews the frequencies, symptoms, testing, and reporting of genotypes with the 5T poly-thymidine tract which reduces splicing efficiency in the cystic fibrosis transmembrane conductance regulator ( CFTR) gene in congenital bilateral absence of the vas deferens (CBAVD) patients and in patients and fetuses with cystic fibrosis-like symptoms. The 5T sequence has not been included in the American College of Medical Genetics (ACMG) CFTR mutation panel recommended for screening pregnant women for an increased fetal risk of cystic fibrosis (CF; MIM 219700) because finding this allele would raise concern for possible CFTR gene-related symptoms in many fetuses, even though only a fraction inheriting 5T and another major CFTR mutation would develop CF-like symptoms. In contrast, 40-80% of the symptomatic patients with CBAVD (MIM 277180) are compound heterozygotes for the 5T sequence. This submission provides template report summaries for CBAVD patient results for the 5T allele when tested along with the 23 most common ACMG mutation panel. If CBAVD patients were also tested with the remaining 16 most common reported mutations in CBAVD, the derived proportion of patients with at least one CFTR mutant allele is predicted to increase from 63% to 97%. Testing for the 5T sequence in symptomatic patients and reflex 5T testing in fetuses found to carry a major CF allele are discussed because finding the 5T sequence in these patients lowers the risk of typical severe symptoms. Additional reflex testing for the number of TG repeats adjacent to a 5T allele further modifies the predicted long-term severity of disease symptoms in patients and fetuses that are compound heterozygotes for a major CF mutation and the 5T sequence. Even though patient advice can be modified currently based upon the adjacent TG-repeat number, the final most accurate risk frequencies with different 5T + TG-repeat alleles are likely to become available only after a larger patient population is completed with multiple well-defined clinical and mutation categories.
Lebo R V; Grody W W
Genetic Testing
2007
2007-03
Journal Article or Conference Abstract Publication
<a href="http://doi.org/10.1089/gte.2006.9997" target="_blank" rel="noreferrer noopener">10.1089/gte.2006.9997</a>
Testing And Reporting Acmg Cystic Fibrosis Mutation Panel Results
congenital bilateral absence; Genetics & Heredity; nonpaternity; population; Research & Experimental Medicine; vas-deferens
Testing strategies and summary reports for pregnant patients and symptomatic patients being tested for cystic fibrosis (CF; MIM 219700) were developed based upon calculated after (posterior) test risk tables incorporating patient and family histories, ethnicities, and prior testing status. This manuscript defines the proportion of all mutations detected by the American College of Medical Genetics (ACMG)-recommended 23-mutation cystic fibrosis transmembrane conductance regulator (CFTR) gene core panel when testing all patient categories with severe symptoms, including pregnant couples with no family history as well as CF patients, their partners, and other family members. Reference tables incorporate prior and posterior test risks sufficient to complete > 99% of all tested cases and to report the results according to HIPAA guidelines. These tables were calculated based on the assumption that all patient samples have been collected, labeled, analyzed, and reported correctly, including the patient's reported relationship to a known affected or carrier relative, even though the template letter states that the likelihood is about 99% that each reported result is accurate. Pedigrees and tables with the prior (before; a priori) test risks of patients offered CF screening with a family history of a CF patient and/or a known carrier patient are provided for ready reference with each risk frequency, dependent upon the assumption that the patient's pedigree reflects familial relationships correctly. Comparison of tables emphasizes the value of asking the tested partner to ask a relative known to have CF or who tested positive for a CF mutation to donate a sample as a DNA test control and/or to obtain a copy of a prior molecular test result and/or extracted DNA sample. These tables posterior test risks also indicate that when one partner with no family history tests negative for the 23 mutation panel, no further prenatal testing is indicated.
Lebo R V; Grody W W
Genetic Testing
2007
2007-03
Journal Article or Conference Abstract Publication
<a href="http://doi.org/10.1089/gte.2006.9996" target="_blank" rel="noreferrer noopener">10.1089/gte.2006.9996</a>
One Multiplex Control For 29 Cystic Fibrosis Mutations
gene; Genetics & Heredity; Research & Experimental Medicine
A simple approach is described to synthesize and clone an inexhaustible supply of any homozygous and/ or heterozygous controls diluted with yeast genomic DNA to mimic human genome equivalents for use throughout the entire multiplex mutation assay. As a proof of principle, the 25 cystic fibrosis mutation panel selected by the American College of Medical Genetics and four additional mutant sequences were prepared as a single control mixture. The 29 CFTR mutations were incorporated into 17 gene fragments by PCR amplification of targeted sequences using mutagenic primers on normal human genomic DNA template. Flanking primers selected to bind beyond all published PCR primer sites amplified controls for most assay platforms. The 17 synthesized 433-933-bp CFTR fragments each with one to four homozygous mutant sequences were cloned into nine plasmid vectors at the multiple cloning site and bidirectionally sequenced. Miniplasmid preps from these nine clones were mixed and diluted with genomic yeast DNA to mimic the final nucleotide molar ratio of two CFTR genes in 6 x 10(9) bp total human genomic DNA. This mixture was added to control PCR reactions prior to amplification as the only positive control sample. In this fashion <200 multiplex clinical PCR analyses of > 4,000 clinical patient samples have been controlled simultaneously for PCR amplification and substrate specificity for 29 tested mutations without cross contamination. This clinically validated multiplex cystic fibrosis control can be modified readily for different test formats and provides a robust means to control for all mutations instead of rotating human genomic controls each with a fraction of the mutations. This approach allows scores of additional mutation controls from any gene loci to be added to the same mixture annually.
Lebo R V; Bixler M; Galehouse D
Genetic Testing
2007
2007
Journal Article or Conference Abstract Publication
<a href="http://doi.org/10.1089/gte.2007.9994" target="_blank" rel="noreferrer noopener">10.1089/gte.2007.9994</a>