Several techniques exist for the molecular diagnosis of hypercoagulable states, included but not limited to these techniques are restricted fragment length polymorphism, polymerase chain reaction, and array technology. Due to the heterogeneity of mutations, it is currently not practical to test all genes that are responsible for defects in the coagulation pathway. This article gives a brief background on hypercoagulable states and the laboratory’s role in diagnosing them.
Pendred syndrome (PDS) and DFNB4 comprise a phenotypic spectrum of sensorineural hearing loss disorders that typically result from biallelic mutations of the SLC26A4 gene. Although PDS and DFNB4 are recessively inherited, sequencing of the coding regions and splice sites of SLC26A4 in individuals suspected to be affected with these conditions often fails to identify two mutations. We investigated the potential contribution of large SLC26A4 deletions and duplications to sensorineural hearing loss (SNHL) by screening 107 probands with one known SLC26A4 mutation by Multiplex Ligation-dependent Probe Amplification (MLPA). A heterozygous deletion, spanning exons 4-6, was detected in only one individual, accounting for approximately 1% of the missing mutations in our cohort. This low frequency is consistent with previously published MLPA results. We also examined the potential involvement of digenic inheritance in PDS/DFNB4 by sequencing the coding regions of FOXI1 and KCNJ10. Of the 29 probands who were sequenced, three carried nonsynonymous variants including one novel sequence change in FOXI1 and two polymorphisms in KCNJ10. We performed a review of prior studies and, in conjunction with our current data, conclude that the frequency of FOXI1 (1.4%) and KCNJ10 (3.6%) variants in PDS/DFNB4 individuals is low. Our results, in combination with previously published reports, indicate that large SLC26A4 deletions and duplications as well as mutations of FOXI1 and KCNJ10 play limited roles in the pathogenesis of SNHL and suggest that other genetic factors likely contribute to the phenotype.
We assessed the frequency and clinicopathologic significance of 19 genes currently identified as significantly mutated in myeloid neoplasms, RUNX1, ASXL1, TET2, CEBPA, IDH1, IDH2, DNMT3A, FLT3, NPM1, TP53, NRAS, EZH2, CBL, U2AF1, SF3B1, SRSF2, JAK2, CSF3R, and SETBP1, across 93 cases of acute myeloid leukemia (AML) using capture target enrichment and next-generation sequencing. Of these cases, 79% showed at least one nonsynonymous mutation, and cases of AML with recurrent genetic abnormalities showed a lower frequency of mutations versus AML with myelodysplasia-related changes (P<0.001). Mutational analysis further demonstrated that TP53 mutations are associated with complex karyotype AML, whereas ASXL1 and U2AF1 mutations are associated with AML with myelodysplasia-related changes. Furthermore, U2AF1 mutations were specifically associated with trilineage morphologic dysplasia. Univariate analysis demonstrated that U2AF1 and TP53 mutations are associated with absence of clinical remission, poor overall survival (OS), and poor disease-free survival (DFS; P<0.0001), whereas TET2 and ASXL1 mutations are associated with poor OS (P<0.03). In multivariate analysis, U2AF1 and TP53 mutations retained independent prognostic significance in OS and DFS, respectively. Our results demonstrate unique relationships between mutations in AML, clinicopathologic prognosis, subtype categorization, and morphologic dysplasia.
The Tissue of Origin Frozen (TOO-FRZ) assay from Pathwork Diagnostics has been cleared by the Food and Drug Administration as a diagnostic study for malignancies of unknown primary. The goal of this study was to evaluate the performance of TOO-FRZ on a diverse collection of malignancies. We collected a diverse set of 49 malignancies. We classified each case into 1 of 4 groups: common morphology from a tissue type included in the TOO-FRZ assay (n=29), uncommon morphology from a tissue type included in the TOO-FRZ assay (n=10), tumor from a tissue type not included in the TOO-FRZ assay (n=3), and malignancies of unknown primary (n=7). We found strong diagnostic performance for common morphologies from tissue types on the TOO-FRZ [overall accuracy=26 of 29 (90%, 95% CI, 73% to 97%)], with perfect performance in all tissue types except gastric (0 of 2) and pancreatic (1 of 2) tissues. There was a significant decline in performance for uncommon morphologies from tissue types included in the TOO-FRZ assay [6 of 10 (60%) cases with an indeterminate result, 1 of 10 (10%) cases with an incorrect prediction, and 3 of 10 (30%) with a correct prediction] and for tumors from tissue types not included in the assay (incorrect prediction in 2 of 3 cases). For the 7 malignancies of unknown primary in our study set, the TOO-FRZ provided a likely clinically useful result in only 2 of 7 cases. These results provide an insight into the strengths and limitations of this molecular assay for the surgical pathologist, and our findings suggest future directions for research in this area.
Background: Hearing loss is a clinically and genetically heterogeneous condition with major medical and social consequences. It affects up to 8% of the general population. Objective: This review recapitulates the principles of auditory physiology and the molecular basis of hearing loss, outlines the main types of non-syndromic and syndromic deafness by mode of inheritance, and provides an overview of current clinically available genetic testing. Methods: This paper reviews the literature on auditory physiology and on genes, associated with hearing loss, for which genetic testing is presently offered. Results/conclusion: The advent of molecular diagnostic assays for hereditary hearing loss permits earlier detection of the underlying causes, facilitates appropriate interventions, and is expected to generate the data necessary for more specific genotype–phenotype correlations.
The Bayer transcription-mediated amplification (TMA) and the Roche PCR Amplicor version 2.0 molecular assays for the qualitative detection of hepatitis C virus were compared for cost, hands-on time, assay duration, and complexity. The TMA assay compares well to PCR and may be especially useful for laboratories with large numbers of test requests.
Craniosynostosis is characterized by premature fusion of one or more cranial sutures and is associated with mutations in fibroblast growth factor receptor (FGFR) genes. Here we describe a novel mutation (1084+1G>A) in the FGFR2 gene of a patient with isolated bicoronal synostosis. We detected two isoforms that result from the mutation and are characterized, respectively, by exon skipping and the use of a cryptic splice site. Interestingly, the alternatively spliced forms of FGFR2 appear to induce fusion of the cranial sutures suggesting that the mutation acts via a gain-of-function mechanism rather than a loss of protein functionality.
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