scholarly journals High sensitivity of both sequencing and real-time PCR analysis of KRAS mutations in colorectal cancer tissue

2009 ◽  
Vol 14 (8) ◽  
pp. 2122-2131 ◽  
Author(s):  
Jolien Tol ◽  
Jeroen R. Dijkstra ◽  
Marianne E. Vink-Börger ◽  
Iris D. Nagtegaal ◽  
Cornelis J.A. Punt ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Real Time ◽  
Real Time Pcr ◽  
High Sensitivity ◽  
Pcr Analysis ◽  
Cancer Tissue ◽  
Kras Mutations ◽  
Colorectal Cancer Tissue

Detection of Epstein-Barr virus and Human Cytomegalovirus in Colorectal Cancer Tissue by Real Time PCR

2021 ◽  
Vol 24 (05) ◽  
Author(s):  
Bashar A. Abdulhassan ◽  
Asmaa B. Al-obaidi ◽  
Noora M. Kareem ◽  
Haidar A. Shamran
Keyword(s):  
Colorectal Cancer ◽  
Real Time ◽  
Human Cytomegalovirus ◽  
Real Time Pcr ◽  
Epstein Barr Virus ◽  
Cancer Tissue ◽  
Colorectal Cancer Tissue ◽  
Barr Virus ◽  
Epstein Barr

scholarly journals DNA Methylation Biomarkers for Blood-Based Colorectal Cancer Screening

2008 ◽  
Vol 54 (2) ◽  
pp. 414-423 ◽  
Author(s):  
Catherine Lofton-Day ◽  
Fabian Model ◽  
Theo DeVos ◽  
Reimo Tetzner ◽  
Juergen Distler ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Real Time ◽  
Restriction Enzyme ◽  
Real Time Pcr ◽  
Selection Process ◽  
Pcr Analysis ◽  
Plasma Samples ◽  
Tissue Samples ◽  
High Performing ◽  
Healthy Control

Abstract Background: Sensitive, specific blood-based tests are difficult to develop unless steps are taken to maximize performance characteristics at every stage of marker discovery and development. We describe a sieving strategy for identifying high-performing marker assays that detect colorectal cancer (CRC)-specific methylated DNA in plasma. Methods: We first used restriction enzyme–based discovery methods to identify marker candidates with obviously different methylation patterns in CRC tissue and nonpathologic tissue. We then used a selection process incorporating microarrays and/or real-time PCR analysis of tissue samples to further test marker candidates for maximum methylation in CRC tissue and minimum amplification in tissues from both healthy individuals and patients with other diseases. Real-time assays of 3 selected markers were validated with plasma samples from 133 CRC patients and 179 healthy control individuals in the same age range. Results: Restriction enzyme–based testing identified 56 candidate markers. This group was reduced to 6 with microarray and real-time PCR testing. Three markers, TMEFF2, NGFR, and SEPT9, were tested with plasma samples. TMEFF2 methylation was detected in 65% [95% confidence interval, 56%–73%] of plasma samples from CRC patients and not detected in 69% (62%–76%) of the controls. The corresponding results for NGFR were 51% (42%–60%) and 84% (77%–89%); for SEPT9, the values were 69% (60%–77%) and 86% (80%–91%). Conclusions: The stringent criteria applied at all steps of the selection and validation process enabled successful identification and ranking of blood-based marker candidates.

scholarly journals Correlation between KRAS, NRAS and BRAF mutations and tumor localizations in patients with primary and metastatic colorectal cancer

2021 ◽  
Author(s):  
Aleksandra Bożyk ◽  
Paweł Krawczyk ◽  
Katarzyna Reszka ◽  
Kinga Krukowska ◽  
Agnieszka Kolak ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Real Time ◽  
Real Time Pcr ◽  
Ascending Colon ◽  
Braf Mutations ◽  
Kras Mutations ◽  
Formalin Fixed Paraffin ◽  
Ffpe Tissues ◽  
Roche Diagnostics ◽  
Formalin Fixed

IntroductionDetection of abnormalities in the KRAS, NRAS and BRAF genes is extremely important for proper qualification of colorectal cancer (CRC) patients for therapy with anti-EGFR monoclonal antibodies. However, data about prevalence of mutations in these genes, in different localizations of CRC tumors, is limited.Material and methodsWe examined the frequency of mutations in the KRAS, NRAS and BRAF genes in 500 Caucasian CRC patients (200 women and 300 men, median age – 66 years). DNA was isolated from formalin- fixed, paraffin embeded (FFPE) tissues using Qiagen QIAmp DNA FFPE-kit. Analysis of mutations was carried out using KRAS/BRAF, NRAS and BRAF Mutation Analysis Kit for Real-Time PCR (EntroGen) using the Cobas 480 real-time PCR apparatus (Roche Diagnostics)ResultsKRAS mutations have been detected in 190 patients (38%), NRAS mutations in 20 patients (4%), whereas BRAF mutations in 24 patients (4,8%). There were no associations between age of CRC patients and frequency of KRAS, NRAS and BRAF genes mutations. These mutations were significantly more often diagnosed in women (55.5%) than in man (41%, p<0.005). Tumors of rectum and sigmoideum were the most often observed in both groups of CRC patients – with and without KRAS, NRAS and BRAF genes mutations. However, transverse colon, ascending colon and cecum cancers were the most often affected by mutationsConclusionsOur study showed that the occurrence of mutations in the KRAS, NRAS and BRAF genes is not accidental and depends on the location of CRC tumors.

KRAS aKtive, an Italian network for assessment of KRAS mutations in colorectal cancer patients: Results on 7,432 cases.

2012 ◽  
Vol 30 (15_suppl) ◽  
pp. e14042-e14042
Author(s):  
Antonio Marchetti ◽  
Carmine Pinto ◽  
Gian Luigi Taddei ◽  
Claudio Clemente ◽  
Giancarlo Troncone ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Real Time ◽  
Cancer Patients ◽  
Real Time Pcr ◽  
Accurate Estimation ◽  
Kras Mutations ◽  
Detection Techniques ◽  
Mutational Status ◽  
Colorectal Cancer Patients ◽  
Large Survey

e14042 Background: The KRAS aKtive program was started on March 2009, promoted by the Italian Association of Medical Oncology (AIOM) and the Italian Society of Surgical Pathology and Cytopathologyy (SIAPEC) to support the activity of oncologists and pathologists involved in the management of metastatic colorectal cancer patients who need the assessment of the mutational status of the KRAS gene. Methods: The program was specifically devised to facilitate the exchange of biologic material, clinicopathological data and diagnostic reports within a network of oncologist, pathologists and pathology/molecular biology reference laboratories throughout Italy, connected through the site www.kras-aKtive.it. KRAS mutation analysis was performed by Sanger sequencing (SS), real time PCR or other techniques, including pyrosequencing and hybridization strip assays. Data were collected in a common database. Results: The KRAS aKtive program has involved 478 oncologists, 144 pathologists, and 24 reference laboratories. A total of 7,432 KRAS mutations analyses were performed. The tests were informative in 7,265 cases (98%). The vast majority of tests (5,626 cases, 77.4%) were conducted by SS. In 529 (7.3%) cases a real-time PCR assay was used, other detection techniques were used in 1,110 (15.3%) cases. KRAS mutations at codons 12-13 were detected in 2,874 cases (39,6%). The frequency of mutations detected by real-time PCR or other techniques (45%, and 43%, respectively) was significantly higher (p=0.002, and p=0.008%, respectively) than that observed by SS (38%). The percentage of cases evaluated by non-SS-based methods has increased during the first three years of the program. Conclusions: The results of this large survey allow an accurate estimation of the actual prevalence of KRAS mutations and their types in caucasian colorectal cancer patients. Our data indicate that the frequency of mutations detected by non-SS-based methods is higher than that obtained by SS.

Analysis of KRAS mutations in circulating tumor DNA and colorectal cancer tissue

2020 ◽  
pp. 1-8
Author(s):  
Yankui Liu ◽  
Longhai Li ◽  
Yu Tian ◽  
Xiao Zhu ◽  
Aijuan Sun ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Circulating Tumor Dna ◽  
Cancer Tissue ◽  
Kras Mutations ◽  
Colorectal Cancer Tissue ◽  
Tumor Dna

scholarly journals Detection of Microsatellite Instability in Colorectal Cancer Patients With a Plasma-Based Real-Time PCR Analysis

2021 ◽  
Vol 12 ◽  
Author(s):  
Namjoo Kim ◽  
Sung Min Kim ◽  
Beom Jae Lee ◽  
Byung il Choi ◽  
Hee Sook Yoon ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Microsatellite Instability ◽  
Real Time ◽  
Real Time Pcr ◽  
Predictive Value ◽  
Checkpoint Inhibitors ◽  
Concordance Rate ◽  
Pcr Analysis ◽  
Status Assessment ◽  
Colorectal Cancer Patients

A microsatellite instability (MSI) test is crucial for screening for HNPCC (Hereditary nonpolyposis colorectal cancer; Lynch syndrome) and optimization of colorectal cancer (CRC) treatment. Mismatch repair (MMR) deficiency is a predictor for good response of immune checkpoint inhibitors in various malignancies. In this study, we evaluated the results of a newly developed plasma-based real-time PCR kit for the detection of MSI in CRC patients. We assessed a peptide nucleotide acid (PNA) probe-mediated real-time PCR test (U-TOP MSI Detection Kit Plus) that determines MSI status by using amplicon melting analysis of five markers (NR21, NR24, NR27, BAT25, and BAT26) from plasma. Eighty-four CRC patients (46 dMMR and 38 pMMR) with colorectal cancer were analyzed. The concordance rate of MSI status assessment between the plasma kit and IHC was 63.0% in dMMR patients (29/46), but in the pMMR evaluation, a 100% (38/38) concordance rate was observed. In the evaluation of the performance of a custom tissue U-TOP MSI Detection Kit and plasma kit in 28 patients, sensitivity, specificity, PPV (positive predictive value) and NPV (negative predictive value) of plasma kit were 68.4, 100, 100, and 44.4%, respectively, with the tissue U-TOP MSI Detection Kit. Our results demonstrate the feasibility of a non-invasive and rapid plasma-based real-time PCR kit (U-TOP MSI Detection Kit Plus) for the detection of MSI in colorectal cancer.

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