Inter-laboratory proficiency testing scheme for tumour next-generation sequencing in Ontario: A pilot study

Background A pilot inter-laboratory proficiency scheme for 5 Ontario clinical laboratories testing tumour samples for the Ontario-wide Cancer Targeted Nucleic Acid Evaluation (OCTANE) study was undertaken to assess proficiency in the identification and reporting of next-generation sequencing (NGS) test results in solid tumour testing from archival formalin-fixed, paraffin-embedded (FFPE) tissue. Methods One laboratory served as the reference centre and provided samples to 4 participating laboratories. An analyte-based approach was applied: each participating laboratory received 10 FFPE tissue specimens profiled at the reference centre, with tumour site and histology provided. Laboratories performed testing per their standard NGS tumour test protocols. Items returned for assessment included genes and variants that would be typically reported in routine clinical testing and variant call format (VCF) files to allow for assessment of NGS technical quality. Results Two main aspects were assessed: Technical quality and accuracy of identification of exonic variants Site-specific reporting practices Technical assessment included evaluation of exonic variant identification, quality assessment of the VCF files to evaluate base calling, variant allele frequency, and depth of coverage for all exonic variants. Concordance at 100% was observed from all sites in the technical identification of 98 exonic variants across the 10 cases. Variability between laboratories in the choice of variants considered clinically reportable was significant. Of the 38 variants reported as clinically relevant by at least 1 site, only 3 variants were concordantly reported by all participating centres as clinically relevant. Conclusions Although excellent technical concordance for NGS tumour profiling was observed across participating institutions, differences in the reporting of clinically relevant variants were observed, highlighting reporting as a gap where consensus on the part of Ontario laboratories is needed

Genetic association of CDC2 with cerebrospinal fluid tau in Alzheimer's disease

We have recently reported that a polymorphism in the cell division cycle (CDC2) gene, designated Ex6 + 7I/D, is associated with Alzheimer's disease (AD). The CDC2 gene is located on chromosome 10q21.1 close to the marker D10S1225 linked to AD. Active cdc2 accumulates in neurons containing neurofibrillary tangles (NFT), a process that can precede the formation of NFT. Therefore, CDC2 is a promising candidate susceptibility gene for AD. We investigated the possible effects of the CDC2 polymorphism on cerebrospinal fluid (CSF) biomarkers in AD patients. CDC2 genotypes were evaluated in relation to CSF protein levels of total tau, phospho-tau and beta-amyloid (1-42) in AD patients and control individuals, and in relation to the amount of senile plaques and NFT in the frontal cortex and in the hippocampus in patients with autopsy-proven AD and controls. The CDC2 Ex6 + 7I allele was associated with a gene dose-dependent increase of CSF total tau levels (F-2,F- 626 = 7.0, p = 0.001) and the homozygous CDC2Ex6 +7II genotype was significantly more frequent among AD patients compared to controls (p = 0.006, OR = 1.57, 95% CI 1.13-2.17). Our results provide further evidence for an involvement of cdc2 in the pathogenesis of AD. Copyright (C) 2005 S. Karger AG, Basel

OCTANE (Ontario-Wide Cancer Targeted Nucleic Acid Evaluation): A Platform for Intraprovincial, National, and International Clinical Data-Sharing

Cancer is a genetic disease resulting from germline or somatic genetic aberrations. Rapid progress in the field of genomics in recent years is allowing for increased characterization and understanding of the various forms of the disease. The Ontario-wide Cancer Targeted Nucleic Acid Evaluation (octane) clinical trial, open at cancer centres across Ontario, aims to increase access to genomic sequencing of tumours and to facilitate the collection of clinical data related to enrolled patients and their clinical outcomes. The study is designed to assess the clinical utility of next-generation sequencing (ngs) in cancer patient care, including enhancement of treatment options available to patients. A core aim of the study is to encourage collaboration between cancer hospitals within Ontario while also increasing international collaboration in terms of sharing the newly generated data. The single-payer provincial health care system in Ontario provides a unique opportunity to develop a province-wide registry of ngs testing and a repository of genomically characterized, clinically annotated samples. It also provides an important opportunity to use province-wide real-world data to evaluate outcomes and the cost of ngs for patients with advanced cancer. The octane study is attempting to translate knowledge to help deliver precision oncology in a Canadian environment. In this article, we discuss the background to the study and its implementation, current status, and future directions

OCTANE (ontario-wide cancer targeted nucleic acid evaluation): A platform for intraprovincial, national, and international clinical data-sharing

Cancer is a genetic disease resulting from germline or somatic genetic aberrations. Rapid progress in the field of genomics in recent years is allowing for increased characterization and understanding of the various forms of the disease. The Ontario-wide Cancer Targeted Nucleic Acid Evaluation (octane) clinical trial, open at cancer centres across Ontario, aims to increase access to genomic sequencing of tumours and to facilitate the collection of clinical data related to enrolled patients and their clinical outcomes. The study is designed to assess the clinical utility of next-generation sequencing (ngs) in cancer patient care, including enhancement of treatment options available to patients. A core aim of the study is to encourage collaboration between cancer hospitals within Ontario while also increasing international collaboration in terms of sharing the newly generated data. The single-payer provincial health care system in Ontario provides a unique opportunity to develop a province-wide registry of ngs testing and a repository of genomically characterized, clinically annotated samples. It also provides an important opportunity to use province-wide real-world data to evaluate outcomes and the cost of ngs for patients with advanced cancer. The octane study is attempting to translate knowledge to help deliver precision oncology in a Canadian environment. In this article, we discuss the background to the study and its implementation, current status, and future directions

Evaluation of gene validity for CPVT and short QT syndrome in sudden arrhythmic death

Aims Catecholaminergic polymorphic ventricular tachycardia (CPVT) and short QT syndrome (SQTS) are inherited arrhythmogenic disorders that can cause sudden death. Numerous genes have been reported to cause these conditions, but evidence supporting these gene–disease relationships varies considerably. To ensure appropriate utilization of genetic information for CPVT and SQTS patients, we applied an evidence-based reappraisal of previously reported genes. Methods and results Three teams independently curated all published evidence for 11 CPVT and 9 SQTS implicated genes using the ClinGen gene curation framework. The results were reviewed by a Channelopathy Expert Panel who provided the final classifications. Seven genes had definitive to moderate evidence for disease causation in CPVT, with either autosomal dominant (RYR2, CALM1, CALM2, CALM3) or autosomal recessive (CASQ2, TRDN, TECRL) inheritance. Three of the four disputed genes for CPVT (KCNJ2, PKP2, SCN5A) were deemed by the Expert Panel to be reported for phenotypes that were not representative of CPVT, while reported variants in a fourth gene (ANK2) were too common in the population to be disease-causing. For SQTS, only one gene (KCNH2) was classified as definitive, with three others (KCNQ1, KCNJ2, SLC4A3) having strong to moderate evidence. The majority of genetic evidence for SQTS genes was derived from very few variants (five in KCNJ2, two in KCNH2, one in KCNQ1/SLC4A3). Conclusions Seven CPVT and four SQTS genes have valid evidence for disease causation and should be included in genetic testing panels. Additional genes associated with conditions that may mimic clinical features of CPVT/SQTS have potential utility for differential diagnosis

Analysis of the 10q23 chromosomal region and the PTEN gene in human sporadic breast carcinoma

We examined a panel of sporadic breast carcinomas for loss of heterozygosity (LOH) in a 10-cM interval on chromosome 10 known to encompass the PTEN gene. We detected allele loss in 27 of 70 breast tumour DNAs. Fifteen of these showed loss limited to a subregion of the area studied. The most commonly deleted region was flanked by D10S215 and D10S541 and encompasses the PTEN locus. We used a combination of denaturing gradient gel electrophoresis and single-strand conformation polymorphism analyses to investigate the presence of PTEN mutations in tumours with LOH in this region. We did not detect mutations of PTEN in any of these tumours. Our data show that, in sporadic breast carcinoma, loss of heterozygosity of the PTEN locus is frequent, but mutation of PTEN is not. These results are consistent with loss of another unidentified tumour suppressor in this region in sporadic breast carcinoma. © 1999 Cancer Research Campaig

Advances in genetics: widening our understanding of prostate cancer

Prostate cancer is a leading cause of cancer-related death in Western men. Our understanding of the genetic alterations associated with disease predisposition, development, progression, and therapy response is rapidly improving, at least in part, owing to the development of next-generation sequencing technologies. Large advances have been made in our understanding of the genetics of prostate cancer through the application of whole-exome sequencing, and this review summarises recent advances in this field and discusses how exome sequencing could be used clinically to promote personalised medicine for prostate cancer patients.</ns4:p

EMT transcription factors snail and slug directly contribute to cisplatin resistance in ovarian cancer

<p>Abstract</p> <p>Background</p> <p>The epithelial to mesenchymal transition (EMT) is a molecular process through which an epithelial cell undergoes transdifferentiation into a mesenchymal phenotype. The role of EMT in embryogenesis is well-characterized and increasing evidence suggests that elements of the transition may be important in other processes, including metastasis and drug resistance in various different cancers.</p> <p>Methods</p> <p>Agilent 4 × 44 K whole human genome arrays and selected reaction monitoring mass spectrometry were used to investigate mRNA and protein expression in A2780 cisplatin sensitive and resistant cell lines. Invasion and migration were assessed using Boyden chamber assays. Gene knockdown of <it>snail </it>and <it>slug </it>was done using targeted siRNA. Clinical relevance of the EMT pathway was assessed in a cohort of primary ovarian tumours using data from Affymetrix GeneChip Human Genome U133 plus 2.0 arrays.</p> <p>Results</p> <p>Morphological and phenotypic hallmarks of EMT were identified in the chemoresistant cells. Subsequent gene expression profiling revealed upregulation of EMT-related transcription factors including <it>snail, slug, twist2 </it>and <it>zeb2</it>. Proteomic analysis demonstrated up regulation of Snail and Slug as well as the mesenchymal marker Vimentin, and down regulation of E-cadherin, an epithelial marker. By reducing expression of <it>snail </it>and <it>slug</it>, the mesenchymal phenotype was largely reversed and cells were resensitized to cisplatin. Finally, gene expression data from primary tumours mirrored the finding that an EMT-like pathway is activated in resistant tumours relative to sensitive tumours, suggesting that the involvement of this transition may not be limited to <it>in vitro </it>drug effects.</p> <p>Conclusions</p> <p>This work strongly suggests that genes associated with EMT may play a significant role in cisplatin resistance in ovarian cancer, therefore potentially leading to the development of predictive biomarkers of drug response or novel therapeutic strategies for overcoming drug resistance.</p

PTEN deficiency: a role in mammary carcinogenesis

The PTEN gene is often mutated in primary human tumors and cell lines, but the low rate of somatic PTEN mutation in human breast cancer has led to debate over the role of this tumor suppressor in this disease. The involvement of PTEN in human mammary oncogenesis has been implicated from studies showing that germline PTEN mutation in Cowden disease predisposes to breast cancer, the frequent loss of heterozygosity at the PTEN locus, and reduced PTEN protein levels in sporadic breast cancers. To assay the potential contribution of PTEN loss in breast tumor promotion, Li et al. [1] crossed Pten heterozygous mice with mouse mammary tumor virus-Wnt-1 transgenic (Wnt-1 TG, Pten+/-) mice. Mammary ductal carcinoma developed earlier in Wnt-1 TG, Pten+/- mice than in mice bearing either genetic change alone, and showed frequent loss of the remaining wild-type PTEN allele. These data indicate a role for PTEN in breast tumorigenesis in an in vivo model