With the expanded availability of next generation sequencing (NGS)-based clinical genetic tests, clinicians seeking to test patients with Mendelian diseases must weigh the superior coverage of ...targeted gene panels with the greater number of genes included in whole exome sequencing (WES) when considering their first-tier testing approach. Here, we use an in silico analysis to predict the analytic sensitivity of WES using pathogenic variants identified on targeted NGS panels as a reference.
Corresponding nucleotide positions for 1533 different alterations classified as pathogenic or likely pathogenic identified on targeted NGS multi-gene panel tests in our laboratory were interrogated in data from 100 randomly-selected clinical WES samples to quantify the sequence coverage at each position. Pathogenic variants represented 91 genes implicated in hereditary cancer, X-linked intellectual disability, primary ciliary dyskinesia, Marfan syndrome/aortic aneurysms, cardiomyopathies and arrhythmias.
When assessing coverage among 100 individual WES samples for each pathogenic variant (153,300 individual assessments), 99.7% (n = 152,798) would likely have been detected on WES. All pathogenic variants had at least some coverage on exome sequencing, with a total of 97.3% (n = 1491) detectable across all 100 individuals. For the remaining 42 pathogenic variants, the number of WES samples with adequate coverage ranged from 35 to 99. Factors such as location in GC-rich, repetitive, or homologous regions likely explain why some of these alterations were not detected across all samples. To validate study findings, a similar analysis was performed against coverage data from 60,706 exomes available through the Exome Aggregation Consortium (ExAC). Results from this validation confirmed that 98.6% (91,743,296/93,062,298) of pathogenic variants demonstrated adequate depth for detection.
Results from this in silico analysis suggest that exome sequencing may achieve a diagnostic yield similar to panel-based testing for Mendelian diseases.
Data sharing through ClinVar offers a unique opportunity to identify interpretation differences between laboratories. As part of a ClinGen initiative, four clinical laboratories (Ambry, GeneDx, ...Partners Healthcare Laboratory for Molecular Medicine, and University of Chicago Genetic Services Laboratory) collaborated to identify the basis of interpretation differences and to investigate if data sharing and reassessment resolve interpretation differences by analyzing a subset of variants.
ClinVar variants with submissions from at least two of the four participating laboratories were compared. For a subset of identified differences, laboratories documented the basis for discordance, shared internal data, independently reassessed with the American College of Medical Genetics and Genomics-Association for Molecular Pathology (ACMG-AMP) guidelines, and then compared interpretations.
At least two of the participating laboratories interpreted 6,169 variants in ClinVar, of which 88.3% were initially concordant. Laboratories reassessed 242/724 initially discordant variants, of which 87.2% (211) were resolved by reassessment with current criteria and/or internal data sharing; 12.8% (31) of reassessed variants remained discordant owing to differences in the application of the ACMG-AMP guidelines.
Participating laboratories increased their overall concordance from 88.3 to 91.7%, indicating that sharing variant interpretations in ClinVar-thereby allowing identification of differences and motivation to resolve those differences-is critical to moving toward more consistent variant interpretations.Genet Med advance online publication 09 March 2017.
Germline pathogenic variants in BRCA1 and BRCA2 predispose to an increased lifetime risk of breast cancer. However, the relevance of germline variants in other genes from multigene hereditary cancer ...testing panels is not well defined.
To determine the risks of breast cancer associated with germline variants in cancer predisposition genes.
A study population of 65 057 patients with breast cancer receiving germline genetic testing of cancer predisposition genes with hereditary cancer multigene panels. Associations between pathogenic variants in non-BRCA1 and non-BRCA2 predisposition genes and breast cancer risk were estimated in a case-control analysis of patients with breast cancer and Exome Aggregation Consortium reference controls. The women underwent testing between March 15, 2012, and June 30, 2016.
Breast cancer risk conferred by pathogenic variants in non-BRCA1 and non-BRCA2 predisposition genes.
The mean (SD) age at diagnosis for the 65 057 women included in the analysis was 48.5 (11.1) years. The frequency of pathogenic variants in 21 panel genes identified in 41 611 consecutively tested white women with breast cancer was estimated at 10.2%. After exclusion of BRCA1, BRCA2, and syndromic breast cancer genes (CDH1, PTEN, and TP53), observed pathogenic variants in 5 of 16 genes were associated with high or moderately increased risks of breast cancer: ATM (OR, 2.78; 95% CI, 2.22-3.62), BARD1 (OR, 2.16; 95% CI, 1.31-3.63), CHEK2 (OR, 1.48; 95% CI, 1.31-1.67), PALB2 (OR, 7.46; 95% CI, 5.12-11.19), and RAD51D (OR, 3.07; 95% CI, 1.21-7.88). Conversely, variants in the BRIP1 and RAD51C ovarian cancer risk genes; the MRE11A, RAD50, and NBN MRN complex genes; the MLH1 and PMS2 mismatch repair genes; and NF1 were not associated with increased risks of breast cancer.
This study establishes several panel genes as high- and moderate-risk breast cancer genes and provides estimates of breast cancer risk associated with pathogenic variants in these genes among individuals qualifying for clinical genetic testing.
Despite the rapid uptake of multigene panel testing (MGPT) for hereditary cancer predisposition, there is limited guidance surrounding indications for testing and genes to include.
To inform the ...clinical approach to hereditary cancer MGPT, we comprehensively evaluated 32 cancer predisposition genes by assessing phenotype-specific pathogenic variant (PV) frequencies, cancer risk associations, and performance of genetic testing criteria in a cohort of 165,000 patients referred for MGPT.
We identified extensive genetic heterogeneity surrounding predisposition to cancer types commonly referred for germline testing (breast, ovarian, colorectal, uterine/endometrial, pancreatic, and melanoma). PV frequencies were highest among patients with ovarian cancer (13.8%) and lowest among patients with melanoma (8.1%). Fewer than half of PVs identified in patients meeting testing criteria for only BRCA1/2 or only Lynch syndrome occurred in the respective genes (33.1% and 46.2%). In addition, 5.8% of patients with PVs in BRCA1/2 and 26.9% of patients with PVs in Lynch syndrome genes did not meet respective testing criteria.
Opportunities to improve upon identification of patients at risk for hereditary cancer predisposition include revising BRCA1/2 and Lynch syndrome testing criteria to include additional clinically actionable genes with overlapping phenotypes and relaxing testing criteria for associated cancers.
Breast cancer is the most commonly diagnosed cancer in women, with 10% of disease attributed to hereditary factors. Although BRCA1 and BRCA2 account for a high percentage of hereditary cases, there ...are more than 25 susceptibility genes that differentially impact the risk for breast cancer. Traditionally, germline testing for breast cancer was performed by Sanger dideoxy terminator sequencing in a reflexive manner, beginning with BRCA1 and BRCA2. The introduction of next-generation sequencing (NGS) has enabled the simultaneous testing of all genes implicated in breast cancer resulting in diagnostic labs offering large, comprehensive gene panels. However, some physicians prefer to only test for those genes in which established surveillance and treatment protocol exists. The NGS based BRCAplus test utilizes a custom tiled PCR based target enrichment design and bioinformatics pipeline coupled with array comparative genomic hybridization (aCGH) to identify mutations in the six high-risk genes: BRCA1, BRCA2, PTEN, TP53, CDH1, and STK11. Validation of the assay with 250 previously characterized samples resulted in 100% detection of 3,025 known variants and analytical specificity of 99.99%. Analysis of the clinical performance of the first 3,000 BRCAplus samples referred for testing revealed an average coverage greater than 9,000X per target base pair resulting in excellent specificity and the sensitivity to detect low level mosaicism and allele-drop out. The unique design of the assay enabled the detection of pathogenic mutations missed by previous testing. With the abundance of NGS diagnostic tests being released, it is essential that clinicians understand the advantages and limitations of different test designs.
Purpose
Genetic predisposition to male breast cancer (MBC) is not well understood. The aim of this study was to better define the predisposition genes contributing to MBC and the utility of germline ...multi-gene panel testing (MGPT) for explaining the etiology of MBCs.
Methods
Clinical histories and molecular results were retrospectively reviewed for 715 MBC patients who underwent MGPT from March 2012 to June 2016.
Results
The detection rate of MGPT was 18.1% for patients tested for variants in 16 breast cancer susceptibility genes and with no prior
BRCA1/2
testing.
BRCA2
and
CHEK2
were the most frequently mutated genes (11.0 and 4.1% of patients with no prior
BRCA1/2
testing, respectively). Pathogenic variants in
BRCA2
odds ratio (OR) = 13.9;
p
= 1.92 × 10
−16
,
CHEK2
(OR = 3.7;
p
= 6.24 × 10
−24
), and
PALB2
(OR = 6.6,
p
= 0.01) were associated with significantly increased risks of MBC. The average age at diagnosis of MBC was similar for patients with (64 years) and without (62 years) pathogenic variants.
CHEK2
1100delC carriers had a significantly lower average age of diagnosis (
n
= 7; 54 years) than all others with pathogenic variants (
p
= 0.03). No significant differences were observed between history of additional primary cancers (non-breast) and family history of male breast cancer for patients with and without pathogenic variants. However, patients with pathogenic variants in
BRCA2
were more likely to have a history of multiple primary breast cancers.
Conclusion
These data suggest that all MBC patients regardless of age of diagnosis, history of multiple primary cancers, or family history of MBC should be offered MGPT.
Li-Fraumeni syndrome (LFS) has traditionally been identified by single-gene testing (SGT) of TP53 triggered by clinical criteria, but the widespread use of multigene panel tests (MGPTs) has upended ...this paradigm. We sought to compare the personal and family cancer histories of TP53-positive result (TP53+) carriers who were identified by either MGPT or SGT.
Of 44 310 individuals who underwent testing of TP53 in a single clinical diagnostic laboratory between 2010 and 2014, 44 086 (40 885 MGPT and 3201 SGT) met study eligibility criteria. Personal cancer histories were available for 38 938 subjects. The frequency of germline TP53 results and various phenotypic manifestations were compared according to test type. All statistical tests were two-sided.
MGPT TP53+ individuals (n = 126) had an older median age at first cancer than SGT TP53+ carriers (n = 96; women: median = 36 vs 28 years, P < .001; and men: median = 40 vs 15 years, P = .004). The median age of breast cancer diagnosis was 40 years in MGPT TP53+ women vs 33 years in SGT TP53+ women (P < .001). In both cohorts, childhood and LFS core cancers, and for women, multiple primary cancers (not multiple breast tumors), were associated with TP53+ results. Established LFS testing criteria were less often met by MGPT TP53+ individuals.
MGPT TP53+ individuals differ in phenotype from those ascertained through SGT and are notably older at cancer diagnosis and less likely to meet LFS clinical criteria. These findings suggest that LFS may have a greater phenotypic spectrum than previously appreciated. This has implications for the counseling of MGPT TP53+ individuals. Prospective follow-up of these individuals and families is needed to re-evaluate cancer risks.
Immunoregulatory functions of VISTA Nowak, Elizabeth C.; Lines, J. Louise; Varn, Frederick S. ...
Immunological reviews,
March 2017, 2017-03-00, 20170301, Volume:
276, Issue:
1
Journal Article
Peer reviewed
Open access
Summary
Utilization of negative checkpoint regulators (NCRs) for cancer immunotherapy has garnered significant interest with the completion of clinical trials demonstrating efficacy. While the ...results of monotherapy treatments are compelling, there is increasing emphasis on combination treatments in an effort to increase response rates to treatment. One of the most recently discovered NCRs is VISTA (V‐domain Ig‐containing Suppressor of T cell Activation). In this review, we describe the functions of this molecule in the context of cancer immunotherapy. We also discuss factors that may influence the use of anti‐VISTA antibody in combination therapy and how genomic analysis may assist in providing indications for treatment.
Many in silico predictors of genetic variant pathogenicity have been previously developed, but there is currently no standard application of these algorithms for variant assessment. Using 4,094 ...ClinVar-curated missense variants in clinically actionable genes, we evaluated the accuracy and yield of benign and deleterious evidence in 5 in silico meta-predictors, as well as agreement of SIFT and PolyPhen2, and report the derived thresholds for the best performing predictor(s). REVEL and BayesDel outperformed all other meta-predictors (CADD, MetaSVM, Eigen), with higher positive predictive value, comparable negative predictive value, higher yield, and greater overall prediction performance. Agreement of SIFT and PolyPhen2 resulted in slightly higher yield but lower overall prediction performance than REVEL or BayesDel. Our results support the use of gene-level rather than generalized thresholds, when gene-level thresholds can be estimated. Our results also support the use of 2-sided thresholds, which allow for uncertainty, rather than a single, binary cut-point for assigning benign and deleterious evidence. The gene-level 2-sided thresholds we derived for REVEL or BayesDel can be used to assess in silico evidence for missense variants in accordance with current classification guidelines.
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