Analysis of missense variation in human BRCA1 in the context of interspecific sequence variation

Abstract

Introduction: Interpretation of results from mutation screening of tumour suppressor genes known to harbour high risk susceptibility mutations, such as APC, BRCA1, BRCA2, MLH1, MSH2, TP53, and PTEN, is becoming an increasingly important part of clinical practice. Interpretation of truncating mutations, gene rearrangements, and obvious splice junction mutations, is generally straightforward. However, classification of missense variants often presents a difficult problem. From a series of 20 000 full sequence tests of BRCA1 carried out at Myriad Genetic Laboratories, a total of 314 different missense changes and eight in-frame deletions were observed. Before this study, only 21 of these missense changes were classified as deleterious or suspected deleterious and 14 as neutral or of little clinical significance. Methods: We have used a combination of a multiple sequence alignment of orthologous BRCA1 sequences and a measure of the chemical difference between the amino acids present at individual residues in the sequence alignment to classify missense variants and in-frame deletions detected during mutation screening of BRCA1. Results: In the present analysis we were able to classify an additional 50 missense variants and two inframe deletions as probably deleterious and 92 missense variants as probably neutral. Thus we have tentatively classified about 50% of the unclassified missense variants observed during clinical testing of BRCA1. Discussion: An internal test of the analysis is consistent with our classification of the variants designated probably deleterious; however, we must stress that this classification is tentative and does not have sufficient independent confirmation to serve as a clinically applicable stand alone method.

Comprehensive Mutation Screening in 55 Probands with Type 1 Primary Hyperoxaluria Shows Feasibility of a Gene-Based Diagnosis

“…Recently, a scheme developed for the BRCA1 gene has become a model for classification of missense variants, taking into account the scoring system of chemical differences between amino acids that initially was developed by Grantham (16) and sequence conservation data that were taken from multiple sequence alignment (20). Since such an approach had not yet been instituted for PH1, we applied a similar strategy for classifying both our newly discovered and all previously described AGXT missense variants.…”

Clinical Value of NPHS2 Analysis in Early- and Adult-Onset Steroid-Resistant Nephrotic Syndrome

“…The high percentage of NPHS2 missense variants represents a diagnostic challenge because in some cases it is difficult to differentiate between a disease-causing mutation and a neutral variant. We describe here an in silico sequence variant classification strategy for the NPHS2 amino acid substitutions based on the combination of different approaches previously reported for other genes (30,41,42), which takes into account (1) the analysis of control chromosomes, (2) the cosegregation with the disease in a family, (3) the biophysical and biochemical difference between wild-type and mutant amino acids (28), (4) the evolutionary conservation of the amino acid residue in orthologs (29,43,44), and (5) software that uses in silico predictors of pathogenic effect [Polyphen (45), SIFT (46)] and splice site [Neural Network (47)]. The accuracy of this in silico analysis was tested using previously described and classified podocin-amino acid substitutions.…”

Methods to Classify BRCA1 Variants of Uncertain Clinical Significance: The More the Merrier

“…Recently, several other methods based on sequence comparisons or biophysical methods have been developed raising the possibility of a comprehensive crossvalidated approach to classify these alleles. [28][29][30] It is prudent to exercise caution before applying the results from these different methods to the clinic. Importantly, the field still needs common standards for assessing sensitivity and specificity.…”