Two Multiplex Assays That Simultaneously Identify 22 Possible Mutation Sites in the KRAS, BRAF, NRAS and PIK3CA Genes

Recently a number of randomized trials have shown that patients with advanced colorectal cancer do not benefit from therapies targeting the epidermal growth factor receptor when their tumors harbor mutations in the KRAS, BRAF and PIK3CA genes. We developed two multiplex assays that simultaneously screen 22 nucleotides in the KRAS, NRAS, BRAF and PIK3CA genes for mutations. The assays were validated on 294 tumor DNA samples from patients with advanced colorectal cancer. In these samples 119 KRAS codon 12 and 13 mutations had been identified by sequence analysis, 126 tumors were wild-type for KRAS and the analysis failed in 49 of the 294 samples due to poor DNA quality. The two mutation assays detected 130 KRAS mutations, among which were 3 codon 61 mutations, and in addition 32 PIK3CA, 13 BRAF and 6 NRAS mutations. In 19 tumors a KRAS mutation was found together with a mutation in the PIK3CA gene. One tumor was mutant for both PIK3CA and BRAF. In summary, the mutations assays identified 161 tumors with a mutation, 120 were wild-type and the analysis failed in 13. The material cost of the 2 mutation assays was calculated to be 8-fold lower than the cost of sequencing required to obtain the same data. In addition, the mutation assays are less labor intensive. We conclude that the performance of the two multiplex mutation assays was superior to direct sequencing. In addition, these assays are cheaper and easier to interpret. The assays may also be of use for selection of patients with other tumor types

Prospects for K+→π+ννˉK^+ \to \pi^+ \nu \bar{ \nu } at CERN in NA62

The NA62 experiment will begin taking data in 2015. Its primary purpose is a 10% measurement of the branching ratio of the ultrarare kaon decay $K^+ \to \pi^+ \nu \bar{ \nu }$, using the decay in flight of kaons in an unseparated beam with momentum 75 GeV/c.The detector and analysis technique are described here.Comment: 8 pages for proceedings of 50 Years of CP

Telomerase reverse transcriptase promoter mutations in bladder cancer: High frequency across stages, detection in urine, and lack of association with outcome

Background Hotspot mutations in the promoter of the gene coding for telomerase reverse transcriptase (TERT) have been described and proposed to activate gene expression. Objectives To investigate TERT mutation frequency, spectrum, association with expression and clinical outcome, and potential for detection of recurrences in urine in patients with urothelial bladder cancer (UBC). D

Incidence of Sarcoma Histotypes and Molecular Subtypes in a Prospective Epidemiological Study with Central Pathology Review and Molecular Testing

International audienceBACKGROUND: The exact overall incidence of sarcoma and sarcoma subtypes is not known. The objective of the present population-based study was to determine this incidence in a European region (Rhone-Alpes) of six million inhabitants, based on a central pathological review of the cases. METHODOLOGY/PRINCIPAL FINDINGS: From March 2005 to February 2007, pathology reports and tumor blocks were prospectively collected from the 158 pathologists of the Rhone-Alpes region. All diagnosed or suspected cases of sarcoma were collected, reviewed centrally, examined for molecular alterations and classified according to the 2002 World Health Organization classification. Of the 1287 patients screened during the study period, 748 met the criteria for inclusion in the study. The overall crude and world age-standardized incidence rates were respectively 6.2 and 4.8 per 100,000/year. Incidence rates for soft tissue, visceral and bone sarcomas were respectively 3.6, 2.0 and 0.6 per 100,000. The most frequent histological subtypes were gastrointestinal stromal tumor (18%; 1.1/100,000), unclassified sarcoma (16%; 1/100,000), liposarcoma (15%; 0.9/100,000) and leiomyosarcoma (11%; 0.7/100,000). CONCLUSIONS/SIGNIFICANCE: The observed incidence of sarcomas was higher than expected. This study is the first detailed investigation of the crude incidence of histological and molecular subtypes of sarcomas

BRAF V600E Mutations Are Common in Pleomorphic Xanthoastrocytoma: Diagnostic and Therapeutic Implications

Pleomorphic xanthoastrocytoma (PXA) is low-grade glial neoplasm principally affecting children and young adults. Approximately 40% of PXA are reported to recur within 10 years of primary resection. Upon recurrence, patients receive radiation therapy and conventional chemotherapeutics designed for high-grade gliomas. Genetic changes that can be targeted by selective therapeutics have not been extensively evaluated in PXA and ancillary diagnostic tests to help discriminate PXA from other pleomorphic and often more aggressive astrocytic malignancies are limited. In this study, we apply the SNaPshot multiplexed targeted sequencing platform in the analysis of brain tumors to interrogate 60 genetic loci that are frequently mutated in 15 cancer genes. In our analysis we detect BRAF V600E mutations in 12 of 20 (60%) WHO grade II PXA, in 1 of 6 (17%) PXA with anaplasia and in 1 glioblastoma arising in a PXA. Phospho-ERK was detected in all tumors independent of the BRAF mutation status. BRAF duplication was not detected in any of the PXA cases. BRAF V600E mutations were identified in only 2 of 71 (2.8%) glioblastoma (GBM) analyzed, including 1 of 9 (11.1%) giant cell GBM (gcGBM). The finding that BRAF V600E mutations are common in the majority of PXA has important therapeutic implications and may help in differentiating less aggressive PXAs from lethal gcGBMs and GBMs

BRAF V600E Mutations Are Common in Pleomorphic Xanthoastrocytoma: Diagnostic and Therapeutic Implications

Pleomorphic xanthoastrocytoma (PXA) is low-grade glial neoplasm principally affecting children and young adults. Approximately 40% of PXA are reported to recur within 10 years of primary resection. Upon recurrence, patients receive radiation therapy and conventional chemotherapeutics designed for high-grade gliomas. Genetic changes that can be targeted by selective therapeutics have not been extensively evaluated in PXA and ancillary diagnostic tests to help discriminate PXA from other pleomorphic and often more aggressive astrocytic malignancies are limited. In this study, we apply the SNaPshot multiplexed targeted sequencing platform in the analysis of brain tumors to interrogate 60 genetic loci that are frequently mutated in 15 cancer genes. In our analysis we detect BRAF V600E mutations in 12 of 20 (60%) WHO grade II PXA, in 1 of 6 (17%) PXA with anaplasia and in 1 glioblastoma arising in a PXA. Phospho-ERK was detected in all tumors independent of the BRAF mutation status. BRAF duplication was not detected in any of the PXA cases. BRAF V600E mutations were identified in only 2 of 71 (2.8%) glioblastoma (GBM) analyzed, including 1 of 9 (11.1%) giant cell GBM (gcGBM). The finding that BRAF V600E mutations are common in the majority of PXA has important therapeutic implications and may help in differentiating less aggressive PXAs from lethal gcGBMs and GBMs

FGFR3, HRAS, KRAS, NRAS and PIK3CA Mutations in Bladder Cancer and Their Potential as Biomarkers for Surveillance and Therapy

Background: Fifty percent of patients with muscle-invasive bladder cancer (MI-BC) die from their disease and current chemotherapy treatment only marginally increases survival. Novel therapies targeting receptor tyrosine kinases or activated oncogenes may improve outcome. Hence, it is necessary to stratify patients based on mutations in relevant oncogenes. Patients with non-muscle-invasive bladder cancer (NMI-BC) have excellent survival, however two-thirds develop recurrences. Tumor specific mutations can be used to detect recurrences in urine assays, presenting a more patient-friendly diagnostic procedure than cystoscopy. Methodology/Principal Findings: To address these issues, we developed a mutation assay for the simultaneous detection of 19 possible mutations in the HRAS, KRAS, and NRAS genes. With this assay and mutation assays for the FGFR3 and PIK3CA oncogenes, we screened primary bladder tumors of 257 patients and 184 recurrences from 54 patients. Additionally, in primary tumors p53 expression was obtained by immunohistochemistry. Of primary tumors 64% were mutant for FGFR3, 11% for RAS, 24% for PIK3CA, and 26% for p53. FGFR3 mutations were mutually exclusive with RAS mutations (p = 0.001) and co-occurred with PIK3CA mutations (p = 0.016). P53 overexpression was mutually exclusive with PIK3CA and FGFR3 mutations (p≤0.029). Mutations in the RAS and PIK3CA genes were not predictors for recurrence-free, progression-free and disease-specific survival. In patients presenting with NMI-BC grade 3 and MI-BC, 33 and 36% of the primary tumors were mutant. In patients with low-grade NMI-BC, 88% of the primary tumors carried a mutation and 88% of the recurrences were mutant. Conclusions/Significance: The mutation assays present a companion diagnostic to define patients for targeted therapies. In addition, the assays are a potential biomarker to detect recurrences during surveillance. We showed that 88% of patients presenting with low-grade NMI-BC are eligible for such a follow-up. This may contribute to a reduction in the number of cystoscopical examinations

Neutral pion form factor measurement at NA62

The NA62 experiment at CERN collected a large sample of charged kaon decays with a highly efficient trigger for decays into electrons in 2007. The kaon beam represents a source of tagged neutral pion decays in vacuum. A measurement of the electromagnetic transition form factor slope of the neutral pion in the time-like region from 1.05 × 106 fully reconstructed π0 Dalitz decay is presented. The limits on dark photon production in π0 decays from the earlier kaon experiment at CERN, NA48/2, are also reported