Krystal Brown

A study of over 35,000 women with breast cancer tested with a 25-gene panel of hereditary cancer genes

As panel testing becomes more common in clinical practice, it is important to understand the prevalence and trends associated with the pathogenic variants (PVs) identified. This is especially true for genetically heterogeneous cancers, such as breast cancer (BC), in which PVs in different genes may be associated with various risks and cancer subtypes. The authors evaluated the outcomes of genetic testing among women who had a personal history of BC.

An independent validation of a gene expression signature to differentiate malignant melanoma from benign melanocytic nevi

Recently, a 23‐gene signature was developed to produce a melanoma diagnostic score capable of differentiating malignant and benign melanocytic lesions. The primary objective of this study was to independently assess the ability of the gene signature to differentiate melanoma from benign nevi in clinically relevant lesions.

Clinical Variant Classification: A Comparison of Public Databases and a Commercial Testing Laboratory

BACKGROUND There is a growing move to consult public databases following receipt of a genetic test result from a clinical laboratory; however, the well-documented limitations of these databases call into question how often clinicians will encounter discordant variant classifications that may introduce uncertainty into patient management. Here, we evaluate discordance in BRCA1 and BRCA2 variant classifications between a single commercial testing laboratory and a public database commonly consulted in clinical practice. MATERIALS AND METHODS BRCA1 and BRCA2 variant classifications were obtained from ClinVar and compared with the classifications from a reference laboratory. Full concordance and discordance were determined for variants whose ClinVar entries were of the same pathogenicity (pathogenic, benign, or uncertain). Variants with conflicting ClinVar classifications were considered partially concordant if ≥1 of the listed classifications agreed with the reference laboratory classification. RESULTS Four thousand two hundred and fifty unique BRCA1 and BRCA2 variants were available for analysis. Overall, 73.2% of classifications were fully concordant and 12.3% were partially concordant. The remaining 14.5% of variants had discordant classifications, most of which had a definitive classification (pathogenic or benign) from the reference laboratory compared with an uncertain classification in ClinVar (14.0%). CONCLUSION Here, we show that discrepant classifications between a public database and single reference laboratory potentially account for 26.7% of variants in BRCA1 and BRCA2. The time and expertise required of clinicians to research these discordant classifications call into question the practicality of checking all test results against a database and suggest that discordant classifications should be interpreted with these limitations in mind. IMPLICATIONS FOR PRACTICE With the increasing use of clinical genetic testing for hereditary cancer risk, accurate variant classification is vital to ensuring appropriate medical management. There is a growing move to consult public databases following receipt of a genetic test result from a clinical laboratory; however, we show that up to 26.7% of variants in BRCA1 and BRCA2 have discordant classifications between ClinVar and a reference laboratory. The findings presented in this paper serve as a note of caution regarding the utility of database consultation.

Detection of somatic variants in peripheral blood lymphocytes using a next generation sequencing multigene pan cancer panel

Next Generation Sequencing (NGS) multigene panels, which are routinely used to assess hereditary cancer risk, can detect both inherited germline variants and somatic variants in cancer-risk genes. We evaluated the frequency and distribution of likely somatic Pathogenic and Likely Pathogenic variants (PVs) detected in >220,000 individuals who underwent clinical testing with a 25-gene panel between September 2013 and March 2016. Likely somatic PVs are defined as variants with NGS read frequencies from 10% to 30%. Overall, 137 (0.06%) individuals were identified as carrying likely somatic PVs, most commonly in TP53 (73), CHEK2 (27), and ATM (20). Among this group, a second PV with a NGS read frequency consistent with a germline variant within the same gene or a different gene on the panel was detected in 21 individuals (15.3%), which is similar to the detection rate in our general testing population. Likely somatic PVs accounted for 38.8% of all PVs in TP53. In comparison, likely somatic PVs accounted for <1% of PVs in most other genes. Likely somatic PVs were more frequently identified in older individuals (p < 0.001). Additional studies are ongoing to further investigate the incidence and clinical implications of somatic variants, enabling the appropriate medical management for these patients.

Impact of Payer Constraints on Access to Genetic Testing

BACKGROUND With increased demand for hereditary cancer genetic testing, some large national health-care insurance payers (LNHPs) have implemented policies to minimize inappropriate testing by mandating consultation with a geneticist or genetic counselor (GC). We hypothesized such a restriction would reduce access and appropriate testing. METHODS Test cancellation rates (ie, tests ordered that did not result in a reported test result), mutation-positive rates, and turnaround times for comprehensive BRCA1/2 testing for a study LNHP that implemented a GC-mandate policy were determined over the 12 months before and after policy implementation (excluding a 4-month transition period). Cancellation rates were evaluated based on the reason for cancellation, National Comprehensive Cancer Network testing criteria, and self-identified ancestry. A control LNHP was evaluated over the same period for comparison. RESULTS The study LNHP cancellation rate increased from 13.3% to 42.1% ( P < .001) after policy implementation. This increase was also observed when only individuals who met National Comprehensive Cancer Network criteria for hereditary breast and ovarian cancer testing were considered (9.5% to 37.7%; P < .001). Cancellation rates increased after policy introduction for all ancestries; however, this was more pronounced among individuals of African or Latin American ancestry, for whom cancellation rates rose to 48.9% and 49.6%, respectively, compared with 33.9% for individuals of European ancestry. Over this same time period, control LNHP cancellation rates decreased or stayed the same for all subgroups. CONCLUSION These findings demonstrate that a GC-mandate policy implemented by a LNHP substantially decreased access to appropriate genetic testing, disproportionately impacting minority populations without any evidence that inappropriate testing was decreased.

Assessment of in silico protein sequence analysis in the clinical classification of variants in cancer risk genes

Missense variants represent a significant proportion of variants identified in clinical genetic testing. In the absence of strong clinical or functional evidence, the American College of Medical Genetics recommends that these findings be classified as variants of uncertain significance (VUS). VUSs may be reclassified to better inform patient care when new evidence is available. It is critical that the methods used for reclassification are robust in order to prevent inappropriate medical management strategies and unnecessary, life-altering surgeries. In an effort to provide evidence for classification, several in silico algorithms have been developed that attempt to predict the functional impact of missense variants through amino acid sequence conservation analysis. We report an analysis comparing internally derived, evidence-based classifications with the results obtained from six commonly used algorithms. We compiled a dataset of 1118 variants in BRCA1, BRCA2, MLH1, and MSH2 previously classified by our laboratory’s evidence-based variant classification program. We compared internally derived classifications with those obtained from the following in silico tools: Align-GVGD, CONDEL, Grantham Analysis, MAPP-MMR, PolyPhen-2, and SIFT. Despite being based on similar underlying principles, all algorithms displayed marked divergence in accuracy, specificity, and sensitivity. Overall, accuracy ranged from 58.7 to 90.8% while the Matthews Correlation Coefficient ranged from 0.26–0.65. CONDEL, a weighted average of multiple algorithms, did not perform significantly better than its individual components evaluated here. These results suggest that the in silico algorithms evaluated here do not provide reliable evidence regarding the clinical significance of missense variants in genes associated with hereditary cancer.

Increased Identification of Candidates for High-Risk Breast Cancer Screening Through Expanded Genetic Testing

PURPOSE Breast MRI screening is recommended for women with a >20% lifetime risk for breast cancer on the basis of estimates derived from risk models dependent largely on family history. Alternatively, a >20% lifetime risk can be established through genetic testing of BRCA1 and BRCA2, as well as a growing selection of other genes associated with inherited breast cancer risk. The aim of this study was to quantify the impact of testing for genes other than BRCA1/2 and the extent to which mutation carriers in these genes would have been identified as candidates for enhanced screening on the basis of family history alone. METHODS Women were tested with a 25-gene hereditary cancer panel including BRCA1/2 and 7 additional genes known to be associated with a >20% lifetime risk for breast cancer (ATM, CHEK2, PALB2, TP53, PTEN, CDH1, and STK11). Women found to carry pathogenic variants (PVs) were evaluated with the Claus model to assess whether they would have been found to be at >20% lifetime risk on the basis of family history. RESULTS In total, 9,751 PVs in the selected breast cancer risk genes were identified in 9,641 women. BRCA1/2 accounted for 59.1% of the PVs, and 38.8% were in ATM, CHEK2, or PALB2. Only 24.7% of all women with PVs found in any gene reached the >20% lifetime risk threshold using the Claus model. CONCLUSIONS Expanding genetic testing beyond BRCA1/2 significantly increases the number of women who are candidates for breast MRI and other risk reduction measures, most of whom would not have been identified through family history assessment.

Community Practice Implementation of a Self-administered Version of PREMM1,2,6 to Assess Risk for Lynch Syndrome

BACKGROUND & AIMS: Lynch syndrome is a genetic disorder that greatly increases risk for colorectal and other cancers, although it is underdiagnosed. Prediction of MLH1, MSH2, and MSH6 (PREMM1,2,6) is a web‐based tool that analyzes individuals’ personal/family histories of cancer to quantify their likelihood of carrying a germline mutation associated with Lynch syndrome. We investigated the feasibility of systematic risk assessment for Lynch syndrome in a community gastroenterology practice using a patient‐completed version of PREMM1,2,6. METHODS: PREMM1,2,6 was adapted into a computer tablet version designed for self‐administration by patients. Individuals presenting to a community gastroenterology office and endoscopy facility in California completed the PREMM1,2,6 assessment before their visit (n = 3134). The total study duration (8 months) comprised a 2‐month initiation period (May 1–June 30, 2013) and a 6‐month study period (July 1–December 31, 2013). Genetic counseling and germline analysis for mutations in genes associated with Lynch syndrome (MLH1, MSH2, MSH6, PMS2, and EPCAM) were offered to individuals with PREMM1,2,6 scores of 5% or higher. Patients and providers completed surveys to evaluate the feasibility and satisfaction with the process. RESULTS: Of the 3134 individuals assessed by PREMM1,2,6 during the 6‐month study period, 177 individuals (5.6%) had scores of 5% or higher. Of these, 146 individuals underwent genetic testing, along with 28 additional participants recruited nonconsecutively during the initiation period. Mutations associated with Lynch syndrome were detected in 3 of the 146 individuals (2.1%) with PREMM1,2,6 scores of 5% or higher who underwent germline testing, and 3 of the 28 patients (10.7%) recruited during study initiation with PREMM1,2,6 scores of 5% or higher. Of the participants who underwent genetic analysis, 98.6% stated that they understood the information provided to them. All of the surveyed providers stated that they were satisfied with the incorporation of PREMM1,2,6 into their clinical practice, and that they would continue using it to assess risk for Lynch syndrome. CONCLUSIONS: A patient self‐administered version of the PREMM1,2,6 Lynch syndrome risk assessment model can be used systematically in community‐based gastroenterology and endoscopy practices.

The influence of a gene-expression signature on the treatment of diagnostically challenging melanocytic lesions

Aim: The effect of a gene-expression-based test on treatment of melanocytic neoplasms by dermatologists was evaluated. Patients & methods: Pathologists submitted diagnostically challenging melanocytic neoplasms to a clinical laboratory for testing accompanied by pretest surveys documenting the intended treatment recommendations. The actual treatment rendered by dermatologists was then documented after testing. Changes between the pretest recommendations and actual treatment were analyzed. Results: In 71.4% (55/77) of cases, there was a change from pretest recommendations to actual treatment. The majority of changes were consistent with the test result. There was an 80.5% (33/41) reduction in the number of biopsy site re-excisions performed for cases with a benign test result. Conclusion: The actual treatment of diagnostically challenging melanocytic neoplasms is influenced by the test.

Analytical Validation of a Cell Cycle Progression Signature Used as a Prognostic Marker in Prostate Cancer

Background: Prostate cancer is the most common cancer in men in the developed world. Appropriate clinical care requires accurate prognostic information to determine whether definitive treatment or conservative management is most appropriate for a given patient. We previously demonstrated that a gene expression signature, which measures the RNA expression of 31 cell cycle progression (CCP) genes and generates a CCP score, is a robust predictor of patient outcome in cohorts of conservatively managed patients diagnosed by needle biopsy or transurethral resection of the prostate. Methods: These current studies represent the analytical validation of this gene signature, for the testing of either formalin-fixed paraffin-embedded (FFPE) prostate resection tissue (radical prostatectomy, RP) or FFPE prostate needle biopsy samples. Results: The measured standard deviation (SD) of the signature was determined to be 0.1 score units, representing 1.6% of the range of scores observed within previous clinical validation studies. Individual amplicons for all genes within the signature had a SD <1 CT, with a median SD of 0.52 CT’s. We observed the median amplification efficiency for all genes was 92.6%. The linear range of the signature was over a ~260-fold range of RNA concentrations. We observed that 100% of RP samples and 99.8% of needle biopsy samples produced sufficient RNA for testing, when RNA was extracted from 7,525 recent prostate samples. Finally, RNA samples were able to reproduce similar CCP scores when stored for up to 8 weeks. Conclusion: These studies indicate that this prognostic gene signature is robust and reproducible, and is analytically validated for use on FFPE prostate biopsy radical prostatectomy samples.