scholarly journals Design and validation of a next generation sequencing assay for hereditaryBRCA1andBRCA2mutation testing

PeerJ ◽  
2016 ◽  
Vol 4 ◽  
pp. e2162 ◽  
Author(s):  
Hyunseok P. Kang ◽  
Jared R. Maguire ◽  
Clement S. Chu ◽  
Imran S. Haque ◽  
Henry Lai ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Copy Number Variants ◽  
Next Generation ◽  
Single Nucleotide Variants ◽  
Cancer Syndrome ◽  
Cancer Screen ◽  
Inherited Cancer ◽  
Increased Risk ◽  
Large Rearrangements ◽  
Generation Sequencing

Hereditary breast and ovarian cancer syndrome, caused by a germline pathogenic variant in theBRCA1orBRCA2(BRCA1/2) genes, is characterized by an increased risk for breast, ovarian, pancreatic and other cancers. Identification of those who have aBRCA1/2mutation is important so that they can take advantage of genetic counseling, screening, and potentially life-saving prevention strategies. We describe the design and analytic validation of the Counsyl Inherited Cancer Screen, a next-generation-sequencing-based test to detect pathogenic variation in theBRCA1andBRCA2genes. We demonstrate that the test is capable of detecting single-nucleotide variants (SNVs), short insertions and deletions (indels), and copy-number variants (CNVs, also known as large rearrangements) with zero errors over a 114-sample validation set consisting of samples from cell lines and deidentified patient samples, including 36 samples withBRCA1/2pathogenic germline mutations.

scholarly journals Design and validation of a next generation sequencing assay for hereditary breast and ovarian cancer

2015 ◽  
Author(s):  
Hyunseok P. Kang ◽  
Jared R Maguire ◽  
Clement S Chu ◽  
Imran S. Haque ◽  
Henry Lai ◽  
...  
Keyword(s):  
Ovarian Cancer ◽  
Next Generation Sequencing ◽  
Next Generation ◽  
Single Nucleotide Variants ◽  
Cancer Syndrome ◽  
Breast And Ovarian Cancer ◽  
Cancer Screen ◽  
Inherited Cancer ◽  
Increased Risk ◽  
Generation Sequencing

Hereditary breast and ovarian cancer syndrome, caused by a germline deleterious variant in the BRCA1 or BRCA2 genes, is characterized by an increased risk for breast, ovarian, pancreatic and other cancers. Identification of those who have a BRCA1/2 mutation is important so that they can take advantage of genetic counseling, screening, and potentially life-saving prevention strategies. We describe the design and analytic validation of the Counsyl Inherited Cancer Screen, a next-generation-sequencing-based test to detect pathogenic variation in the BRCA1 and BRCA2 genes. We demonstrate that the test is capable of detecting single-nucleotide variants (SNVs), short insertions and deletions (indels), and copy-number variants (CNVs, also known as large rearrangements) with zero errors over a 96-sample validation set consisting of samples from cell lines and deidentified patient samples, including the well-characterized NA12878 sample from HapMap/1000 Genomes.

scholarly journals Design and validation of a next generation sequencing assay for hereditary breast and ovarian cancer

2015 ◽  
Author(s):  
Hyunseok P. Kang ◽  
Jared R Maguire ◽  
Clement S Chu ◽  
Imran S. Haque ◽  
Henry Lai ◽  
...  
Keyword(s):  
Ovarian Cancer ◽  
Next Generation Sequencing ◽  
Next Generation ◽  
Single Nucleotide Variants ◽  
Cancer Syndrome ◽  
Breast And Ovarian Cancer ◽  
Cancer Screen ◽  
Inherited Cancer ◽  
Increased Risk ◽  
Generation Sequencing

Hereditary breast and ovarian cancer syndrome, caused by a germline deleterious variant in the BRCA1 or BRCA2 genes, is characterized by an increased risk for breast, ovarian, pancreatic and other cancers. Identification of those who have a BRCA1/2 mutation is important so that they can take advantage of genetic counseling, screening, and potentially life-saving prevention strategies. We describe the design and analytic validation of the Counsyl Inherited Cancer Screen, a next-generation-sequencing-based test to detect pathogenic variation in the BRCA1 and BRCA2 genes. We demonstrate that the test is capable of detecting single-nucleotide variants (SNVs), short insertions and deletions (indels), and copy-number variants (CNVs, also known as large rearrangements) with zero errors over a 96-sample validation set consisting of samples from cell lines and deidentified patient samples, including the well-characterized NA12878 sample from HapMap/1000 Genomes.

scholarly journals Development and Validation of a Next-Generation Sequencing Panel for Syndromic and Nonsyndromic Hearing Loss

2020 ◽  
Vol 5 (3) ◽  
pp. 467-479 ◽  
Author(s):  
Malinda Butz ◽  
Amber McDonald ◽  
Patrick A Lundquist ◽  
Melanie Meyer ◽  
Sean Harrington ◽  
...  
Keyword(s):  
Hearing Loss ◽  
Next Generation Sequencing ◽  
Copy Number Variants ◽  
Analytical Sensitivity ◽  
Nonsyndromic Hearing Loss ◽  
Next Generation ◽  
Single Nucleotide Variants ◽  
Small Indels ◽  
Syndromic Hearing Loss ◽  
Generation Sequencing

Abstract Background Deafness and hearing loss are common conditions that can be seen independently or as part of a syndrome and are often mediated by genetic causes. We sought to develop and validate a hereditary hearing loss panel (HHLP) to detect single nucleotide variants (SNVs), insertions and deletions (indels), and copy number variants (CNVs) in 166 genes related to nonsyndromic and syndromic hearing loss. Methods We developed a custom-capture next-generation sequencing (NGS) reagent to detect all coding regions, ±10 flanking bp, for the 166 genes related to nonsyndromic and syndromic hearing loss. Our validation consisted of testing 52 samples to establish accuracy, reproducibility, and analytical sensitivity. In addition to NGS, supplementary methods, including multiplex ligation-dependent probe amplification, long-range PCR, and Sanger sequencing, were used to ensure coverage of regions that had high complexity or homology. Results We observed 100% positive and negative percentage agreement for detection of SNVs (n = 362), small indels (1–22 bp, n = 25), and CNVs (gains, n = 8; losses, n = 17). Finally, we showed that this assay was able to detect variants with a variant allele frequency ≥20% for SNVs and indels and ≥30% to 35% for CNVs. Conclusions We validated an HHLP that detects SNVs, indels, and CNVs in 166 genes related to syndromic and nonsyndromic hearing loss. The results of this assay can be utilized to confirm a diagnosis of hearing loss and related syndromic disorders associated with known causal genes.

Identifying the distant metastasis genes by next-generation sequencing panel in nasopharyngeal carcinoma.

2020 ◽  
Vol 38 (15_suppl) ◽  
pp. e18547-e18547
Author(s):  
Man Hu ◽  
Jinming Yu ◽  
Zihan Zhou ◽  
Peifeng Li ◽  
Xianbin Zhang ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Nasopharyngeal Carcinoma ◽  
Distant Metastasis ◽  
Copy Number Variants ◽  
Intensity Modulated Radiotherapy ◽  
Next Generation ◽  
Single Nucleotide Variants ◽  
Chi Square ◽  
Exact Test ◽  
Generation Sequencing

e18547 Background: Even though local and regional controls have been substantially improved in nasopharyngeal carcinoma (NPC) in the contemporary era of intensity-modulated radiotherapy with extensive use of combined chemotherapy, the distant metastasis becomes the major cause of treatment failure and cancer-related death. To date, the genes contributed to metastasis of NPC is still unclear. The aim of this study was to identify the genes which lead to distant metastasis. Methods: A total of forty primary nonkeratinizing NPC patients were diagnosed at Shandong Cancer Hospital in this study. The formaldehyde-fixed paraffin embedded (FFPE) taken from primary sites or metastatic lymph nodes were performed next-generation sequencing (NGS) panel (Shanghai OrigiMed Co., Ltd.) to determine variated genes, such as single nucleotide variants (SNV), copy number variants (CNV) and rearrangement. These patients were followed up until Febr. 8, 2020. The genes related to distant metastasis were identified by logistic regression. Moreover, this study compared the frequency of mutated gene between our data and Catalog of Somatic Mutations in Cancer (COSMIC) database by the Chi-square test or Fisher’s exact test. Results: The study included 31 men and 9 women. The median age of the patients at diagnosis was 47 years (range 15–71 years). With the median follow-up of 10.6 months (range 16.8–72.3 months), 7 patients had distant metastasis and 1 undergone recurrent. Notably, EMSY and MCL1 variants were contributed to NPC distant metastasis (OR = 31, P = 0.049). The top eight SNV of genes in our study were CYLD, KMT2D, BAP1, EP300, TP53, ATM, NFKBIA and SPEN. When compared to COSMIC database, the mutant frequencies of CYLD, EP300 and BAP1 in our study were significantly higher than that of COSMIC database. However, the mutant frequencies of IDH2 and KMT2C were significantly lower than COSMIC database. Conclusions: This is the first study which suggests that EMSY and MCL1 variants were involved in the metastasis of NPC. The study identified 5 genes, which mutation frequency is significantly different from the COSMIC database. The study provided a molecular basis for a comprehensive understanding of, and exploring targeted therapies for nasopharyngeal carcinoma.

Influence of Next-Generation Sequencing on Advancements in the Diagnosis of Major Psychiatric Diseases - A Review

2020 ◽  
Author(s):  
Maheen Nisar
Keyword(s):  
Next Generation Sequencing ◽  
Mental Disorders ◽  
Attention Deficit Disorder ◽  
Autism Spectrum ◽  
Next Generation ◽  
Single Nucleotide Variants ◽  
New Genes ◽  
Depth Analysis ◽  
Pubmed Search ◽  
Generation Sequencing

Rapid progress is being made in the development of next-generation sequencing (NGS) technologies, allowing repeated findings of new genes and a more in-depth analysis of genetic polymorphisms behind the pathogenesis of a disease. In a field such as psychiatry, characteristic of vague and highly variable somatic manifestations, these technologies have brought great advances towards diagnosing various psychiatric and mental disorders, identifying high-risk individuals and towards more effective corresponding treatment. Psychiatry has the difficult task of diagnosing and treating mental disorders without being able to invariably and definitively establish the properties of its illness. This calls for diagnostic technologies that go beyond the traditional ways of gene manipulation to more advanced methods mainly focusing on new gene polymorphism discoveries, one of them being NGS. This enables the identification of hundreds of common and rare genetic variations contributing to behavioral and psychological conditions. Clinical NGS has been useful to detect copy number and single nucleotide variants and to identify structural rearrangements that have been challenging for standard bioinformatics algorithms. The main objective of this article is to review the recent applications of NGS in the diagnosis of major psychiatric disorders, and hence gauge the extent of its impact in the field. A comprehensive PubMed search was conducted and papers published from 2013-2018 were included, using the keywords, “schizophrenia” or “bipolar disorder” or “depressive disorder” or “attention deficit disorder” or “autism spectrum disorder” and “next-generation sequencing”

scholarly journals Next generation sequencing of RB1gene for the molecular diagnosis of ethnic minority with retinoblastoma in Yunnan

2020 ◽  
Author(s):  
Huaiyu Gu ◽  
Zhen Zhang ◽  
Yi-shuang Xiao ◽  
Ru Shen ◽  
Hong-chao Jiang ◽  
...  
Keyword(s):  
Genetic Testing ◽  
Next Generation Sequencing ◽  
Low Income ◽  
Eastern China ◽  
Low Income Countries ◽  
Next Generation ◽  
Single Nucleotide Variants ◽  
Bioinformatics Pipeline ◽  
Generation Sequencing

Abstract Background: Retinoblastoma is a rare intraocular malignancy and typically initiated by inactivating biallelic mutations of RB1 gene. Each year, ~8,000 children worldwide are diagnosed for retinoblastoma. In high-income countries, patient survival is over 95% while low-income countries is ~30%.If disease is diagnosed early and treated in centers specializing in retinoblastoma, the survival might exceed 95% and many eyes could be safely treated and support a lifetime of good vision. In China, approximate 1,100 newly diagnosed cases are expected annually and 28 hospitals covering 25 provinces established centers classified by expertise and resources for better treatment options and follow-up. Comparing with other province of eastern China, Yunnan province is remote geographically. This might result that healthcare staff have low awareness of the role of genetic testing in management and screening in families.Methods: The patients with retinoblastoma were selected in Yunnan. DNA from blood was used for targeted gene sequencing. Then, an in-house bioinformatics pipeline was done to detect both single nucleotide variants and small insertions/deletions. The pathogenic mutations were identified and further confirmed by conventional methods and cosegregation in families.Results: Using our approach, targeted next generation sequencing was used to detect the mutation of these 12 probands. Bioinformatic predictions showed that nine mutations were found in our study and four were novel pathogenic variants in these nine mutations.Conclusions: It’s the first report to describe RB1 mutations in Yunnan children with retinoblastoma. This study would improve role of genetic testing for management and family screening.

scholarly journals PATH-03. CLINICAL UTILITY OF NEXT GENERATION SEQUENCING IN IDH-WILDTYPE GLIOBLASTOMA: THE DANA-FARBER CANCER INSTITUTE EXPERIENCE

2020 ◽  
Vol 22 (Supplement_2) ◽  
pp. ii164-ii164
Author(s):  
Mary Jane Lim-Fat ◽  
Gilbert Youssef ◽  
Mehdi Touat ◽  
Bryan Iorgulescu ◽  
Eleanor Woodward ◽  
...  
Keyword(s):  
Clinical Trial ◽  
Clinical Trials ◽  
Next Generation Sequencing ◽  
Immune Checkpoint Blockade ◽  
Next Generation ◽  
Single Nucleotide Variants ◽  
Dana Farber Cancer Institute ◽  
Clinical Records ◽  
Ngs Data ◽  
Generation Sequencing

Abstract BACKGROUND Comprehensive next generation sequencing (NGS) is available through many academic institutions and commercial entities, and is incorporated in practice guidelines for glioblastoma (GBM). We retrospective evaluated the practice patterns and utility of incorporating NGS data into routine care of GBM patients at a clinical trials-focused academic center. METHODS We identified 1,011 consecutive adult patients with histologically confirmed GBM with OncoPanel testing, a targeted exome NGS platform of 447 cancer-associated genes at Dana Farber Cancer Institute (DFCI), from 2013-2019. We selected and retrospectively reviewed clinical records of all IDH-wildtype GBM patients treated at DFCI. RESULTS We identified 557 GBM IDH-wildtype patients, of which 227 were male (40.7%). OncoPanel testing revealed 833 single nucleotide variants and indels in 44 therapeutically relevant genes (Tier 1 or 2 mutations) including PIK3CA (n=51), BRAF (n=9), FGFR1 (n=8), MSH2 (n=4), MSH6 (n=2) and MLH1 (n=1). Copy number analysis revealed 509 alterations in 18 therapeutically relevant genes including EGFR amplification (n= 186), PDGFRA amplification (N=39) and CDKN2A/2B homozygous loss (N=223). Median overall survival was 17.5 months for the whole cohort. Seventy-four therapeutic clinical trials accrued 144 patients in the upfront setting (25.9%) and 203 patients (36.4%) at recurrence. Altogether, NGS data for 107 patients (19.2%) were utilized for clinical trial enrollment or targeted therapy indications. High mutational burden (>17mutations/Mb) was identified in 11/464 samples (2.4%); of whom 3/11 received immune checkpoint blockade. Four patients received compassionate use therapy targeting EGFRvIII (rindopepimut, n=2), CKD4/6 (abemaciclib, n=1) and BRAFV600E (dabrafenib/trametinib, n=1). CONCLUSION While NGS has greatly improved diagnosis and molecular classification, we highlight that NGS remains underutilized in selecting therapy in GBM, even in a setting where clinical trials and off-label therapies are relatively accessible. Continued efforts to develop better targeted therapies and efficient clinical trial design are required to maximize the potential benefits of genomically-stratified data.

scholarly journals Complexities of Next-Generation Sequencing in Solid Tumors: Case Studies

2020 ◽  
Vol 18 (9) ◽  
pp. 1150-1155
Author(s):  
Alexandra O. Sokolova ◽  
Brian H. Shirts ◽  
Eric Q. Konnick ◽  
Ginger J. Tsai ◽  
Bernardo H.L. Goulart ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Case Studies ◽  
Checkpoint Inhibitors ◽  
Hereditary Cancer Syndrome ◽  
Next Generation ◽  
Cancer Syndrome ◽  
Expert Review ◽  
Tumor Boards ◽  
Tumor Sequencing ◽  
Generation Sequencing

With the promise and potential of clinical next-generation sequencing for tumor and germline testing to impact treatment and outcomes of patients with cancer, there are also risks of oversimplification, misinterpretation, and missed opportunities. These issues risk limiting clinical benefit and, at worst, perpetuating false conclusions that could lead to inappropriate treatment selection, avoidable toxicity, and harm to patients. This report presents 5 case studies illustrating challenges and opportunities in clinical next-generation sequencing interpretation and clinical application in solid tumor oncologic care. First is a case that dissects the origin of an ATM mutation as originating from a hematopoietic clone rather than the tumor. Second is a case illustrating the potential for tumor sequencing to suggest germline variants associated with a hereditary cancer syndrome. Third are 2 cases showing the potential for variant reclassification of a germline variant of uncertain significance when considered alongside family history and tumor sequencing results. Finally, we describe a case illustrating challenges with using microsatellite instability for predicting tumor response to immune checkpoint inhibitors. The common theme of the case studies is the importance of examining clinical context alongside expert review and interpretation, which together highlight an expanding role for contextual examination and multidisciplinary expert review through molecular tumor boards.

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