RAS Mutational Status Detection in Tissue, Plasma, and Stool Samples for Colorectal Cancer

Objective. RAS gene testing on tumor tissue biopsies is required for metastatic colorectal cancer (CRC) patients. However, it is infeasible for patients after curative surgery and repeated biopsy. This study is aimed at evaluating the consistency of RAS genes in patient’s plasma, stool, and tumor tissue samples, to explore whether plasma and stool samples can supplement or replace tumor tissue to assess baseline RAS gene status. Methods. Between June 2016 and October 2017, 53 patients with stage I-IV CRC from the Liaoning Cancer Hospital and the Department of Medical Oncology of the First Hospital of China Medical University were enrolled in the study. Patient tissues, peripheral blood, and stool samples were collected, and RAS gene tests were performed. Results. Analysis of the KRAS gene in tissue, plasma, and stool samples from 53 CRC patients detected 25 cases (47%) of KRAS gene mutations in the tissue samples, 20 cases (38%) of KRAS gene mutations in plasma, and 18 (34%) KRAS gene mutations in fecal samples. The overall consistency of KRAS gene status between tissue samples and plasma samples was 77.4% (p≤0.05) and between tissue samples and stool samples was 83% (p≤0.05). In stage IV cases, the agreement of KRAS gene status between tissue and plasma samples was 93.8% (p≤0.05) and 93.8% (p≤0.05) between tissue and stool samples. Conclusion. There was a high overall consistency in KRAS mutational assessment between plasma, stool, and tissue samples. In stage IV patients, the consistency of KRAS gene detection between tissue and stools or plasma was higher.

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Expression of tumor pyruvate kinase M2 isoform in plasma and stool of patients with colorectal cancer or adenomatous polyps

10.21203/rs.2.16791/v1 ◽

2019 ◽

Author(s):

Farideh Rigi ◽

Aliakbar Jannatabad ◽

Azra Izanloo ◽

Reza Roshanravan ◽

Hamid Reza Hashemian ◽

...

Keyword(s):

Colorectal Cancer ◽

Pyruvate Kinase ◽

Sensitivity And Specificity ◽

Adenomatous Polyps ◽

Plasma Samples ◽

Proliferating Cells ◽

Pyruvate Kinase M2 ◽

Specificity And Sensitivity ◽

Tumor Group ◽

Stool Samples

Abstract Background: Tumor pyruvate kinase M2 isoform (tM2-PK), which is an isoform of PK-glycolytic enzyme and appears on the surface of cancerous proliferating cells, has been used as a diagnostic biomarker for colorectal cancer (CRC). The aim of this study was to evaluate the tM2-PK measurement test for the diagnosis of CRCs and adenomatous polyps in plasma and stool samples in an Iranian population. Methods: In this prospective study, a total of 226 stool and 178 plasma samples were received from patients referred to colonoscopy units. tM2-PK enzyme was measured using two separate ScheBo-Biotech-AG ELISA kits for stool and plasma samples. Results: At the cut-off value of 4 U/ml, in tumor group, the sensitivity of fecal tM2-PK test was 100% and the specificity was 68%, and in polyp group, the sensitivity and specificity were 87% and 68%, respectively. At the cut-off value of 15 U/ml in tumor group, the sensitivity of plasma tM2-PK test was 98% and specificity was 74% and in polyp group the sensitivity and specificity were 98% and 74%, respectively. Based on our results, a cut-off range of 4.8-8 U/ml and >8 U/ml could be used to detect polyp and tumor in stool samples, respectively. Similarly, a cut-off range of 19-25 U/ml and >25 U/ml is recommended in plasma samples for polyp and tumor detection, respectively. Conclusions: This study revealed a high specificity and sensitivity of tM2-PK test for stool and plasma samples in patients with CRC and polyps suggesting it as a non-invasive assay to diagnose CRC and adenomatous polyps.

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The association between fecal enterotoxigenic B. fragilis with colorectal cancer

10.21203/rs.2.10895/v3 ◽

2019 ◽

Author(s):

Fakhri Haghi ◽

Elshan Goli ◽

Bahman Mirzaei ◽

Habib Zeighami

Keyword(s):

Colorectal Cancer ◽

Direct Detection ◽

Mucosal Immune Response ◽

Colorectal Disease ◽

Marker Genes ◽

Tissue Samples ◽

Potential Marker ◽

Paired Normal Tissue ◽

Stool Samples ◽

Pcr Targeting

Abstract Background Enterotoxigenic Bacteroides fragilis (ETBF) is an enterotoxin-producing bacterium that possibily has a role in the occurrence and progression of colorectal cancer (CRC) by modulating the mucosal immune response and inducing epithelial cell changes. The aim of this study was to investigate the frequency of ETBF in stool samples of CRC patients and healthy volunteers.Methods A total of 60 stool samples from confirmed CRC patients and 60 stool samples from healthy volunteers with no personal or familial history or diagnosis of colorectal disease were collected. Stool samples were screened for direct detection of B. fragilis using PCR targeting the marker genes of neu and bft. Enterotoxin isotypes bft-1, bft-2 and bft-3 were also detected in B. fragilis positive samples.Results The frequency of B. fragilis among CRC and control cases was 58.3% and 26.6%, respectively (P<0.05). The rate of bft gene in CRC cases was significantly higher than in controls (P<0.05). Also, the presence of bf t gene in CRC patients stage III was significantly higher than stages I and II (P< 0.05). Enterotoxin isotype bft-2 was detected with higher frequency among CRC patients than healthy control (P<0.05).Conclusion Our results show the association between fecal ETBF and CRC, and we suggest that detection of ETBF may be a potential marker for colorectal cancer diagnosis. However, additional investigations on tumor and paired normal tissue samples are required to substantiate this possible correlation.

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Interim analysis of the data of non-interventional study to determine the prevalence of EGFR mutations in advanced NSCLC Russian patients (ORTUS).

Journal of Clinical Oncology ◽

10.1200/jco.2019.37.15_suppl.e13109 ◽

2019 ◽

Vol 37 (15_suppl) ◽

pp. e13109-e13109

Author(s):

Konstantin K. Laktionov ◽

Irina Demidova ◽

Aleksandr Ageev ◽

Valeriy Vladimirovich Breder ◽

Pavel Grigoriev ◽

...

Keyword(s):

Interim Analysis ◽

Advanced Nsclc ◽

Gene Mutations ◽

Stage Iv ◽

Tumor Burden ◽

Egfr Mutations ◽

Plasma Samples ◽

Stage Iv Disease ◽

Treatment Naïve ◽

Nsclc Patients

e13109 Background: There is a little information of correlation between EGFRm rate in cytology and plasma samples in Russian NSCLC population. The interim analysis of the study aimed to evaluate the prevalence and types of EGFR mutations in paired cytology and plasma samples in treatment-naive patients with advanced NSCLC. Methods: ORTUS is a multicenter, non-interventional, prospective study to determine EGFR mutations rate in treatmentnaive Russian patients with advanced NSCLC. ClinicalTrials.gov identifier: NCT02321046. The study enrolled 426 patients in stage IIIB / IV of NSCLC. Interim analysis covered the data of 214 cytology verified patients (mean age - 62.6 (range 32-86) years, 58,4% of men) with EGFRm test results in paired cytology and ctDNA samples. 99.5% cases were adenocarcinoma. The proportion of non-smokers/smokers/exsmokers was 46.3%/36%/17.8% respectively. Stage IV disease was in 81% of cases; 84.1% of patients had symptoms. DNA isolation performed using QIAamp DNA FFPE Tissue Kit for cytology and Qiagen Circulating Nucleic Acid Kit for ctDNA according to the manufacturer’s instructions. EGFR gene mutations were analyzed using THerascreen RGQ EGFR PCR Kit in cytology samples and RGQ Plasma EGFR PCR kit in plasma (Qiagen). Results: EGFRm was identified in 17,8% cytology samples (38/214) that is close to 20,2% (1759/8716) in Russian tissue EGFRm study (Imyanitov et al., 2016). 10,3% of paired ctDNA samples were EGFRm positive. Sensitivity and specificity for ctDNA were 42.1%, and 97.1% respectively. The EGFRm rate was 3,9% and 2,6% in smokers, 5,3% and 0% in ex-smokers and 33,3% and 21,2% in nonsmokers in cytology and plasma samples respectively. EGFRm rate and concordance between cytology and ctDNA are presented in a table. EGFRm in ctDNA were detected more frequently in M1a/b groups (p = 0,028). Conclusions: Cytology samples are appropriate for EGFRm testing in NSCLC patients in comparison with tumor tissue ones. High tumor burden (positive metastatic status) is an important factor for successful mutation analysis in ctDNA. [Table: see text]

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Identifying genomic alterations in small cell lung cancer using the liquid biopsy of bronchial washing fluid.

Journal of Clinical Oncology ◽

10.1200/jco.2020.38.15_suppl.e21098 ◽

2020 ◽

Vol 38 (15_suppl) ◽

pp. e21098-e21098

Author(s):

Jinfang Zhai ◽

Songyan Han ◽

Qinxiang Guo ◽

Binbin Shan ◽

Jing Wang ◽

...

Keyword(s):

Lung Cancer ◽

Small Cell Lung Cancer ◽

Liquid Biopsy ◽

Tumor Tissue ◽

Small Cell ◽

Plasma Samples ◽

Small Cell Lung ◽

Tissue Samples ◽

Bronchial Washing ◽

Treatment Naïve

e21098 Background: With the rapid development of cancer genomics, the precision medicine of small cell lung cancer (SCLC) is emerging. However, there are limitations to the clinical use of tumor tissue and peripheral blood biopsies. The main purpose of this study was to evaluate the potential use of bronchial washing fluid (BWF) in the liquid biopsy of SCLC. Methods: Twenty-one SCLC patients diagnosed in 2019 were enrolled in this study. BWF (separated as supernatant and precipitate), treatment-naive plasma and tumor tissue samples were collected from all of patients and subjected to next-generation sequencing (NGS) using a 1021-gene panel. The concordance rates of genomic profiling using NGS in these four types of samples were evaluated. Results: Of these 21 patients, 20 BWF supernatant (BWFs) samples, 21 BWF precipitate (BWFp) samples, 21 tumor tissue samples and 20 plasma samples were successfully tested. The detectability of somatic mutations was 100% for BWFs, BWFp and tumor tissues, and only one plasma was absent with any mutation. Matched tumor tissue, BWFs, BWFp and plasma samples were subsistent for 19 patients. For these patients, 204 genomic alterations were identified in tissue samples, of which 189 (92.6%), 175 (85.5%) and 163 (79.9%) alterations were detected in the matched BWFs, BWFp and plasma samples, respectively. Moreover, tumor mutation burden (TMB) was also calculated. Compared with the proportion of TMB-H samples in tissue samples counting 61.9% (13/21), 60% (12/20) of BWFs samples and 52.38 % (11/21) of BWFp samples were TMB-H (defined as more than or equal to 9 mutations per megabase), which was a molecular biomarker that can be used in immunotherapy efficacy prediction. The TMBs of BWFs, BWFp and treatment-naive plasma samples all had strong correlation with that of tissue samples. The TMB of BWFs had the strongest correlation (Pearson r = 0.9512, p < 0.0001), and the TMB of treatment-naive plasma had relatively lower correlation (Pearson r = 0.8782, p < 0.0001) compared with those of BWFs (Pearson r = 0.936, p < 0.0001) and BWFp (Pearson r = 0.8782, p < 0.0001). Conclusions: For SCLC patients, the liquid biopsy of BWF showed high potential to identify DNA alterations and calculate TMB grades, which suggested that genomic analysis of BWF liquid biopsy may have clinical value in predicting the effectiveness of targeted therapy and immunotherapy. It can be widely used in routine clinical practice.

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Detection of SNCA and FBN1 Methylation in the Stool as a Biomarker for Colorectal Cancer

Disease Markers ◽

10.1155/2015/657570 ◽

2015 ◽

Vol 2015 ◽

pp. 1-6 ◽

Cited By ~ 13

Author(s):

Wen-han Li ◽

Hao Zhang ◽

Qi Guo ◽

Xuan-di Wu ◽

Zi-sen Xu ◽

...

Keyword(s):

Colorectal Cancer ◽

Methylation Status ◽

Good Alternative ◽

Noninvasive Detection ◽

Healthy Controls ◽

Chain Reaction ◽

Tissue Samples ◽

Specific Polymerase Chain Reaction ◽

Stool Samples ◽

Polymerase Chain

Aim.We examined the methylation status of SNCA and FBN1 genes in patients’ paired tissue and stool samples for detection of colorectal cancer (CRC).Patients and Methods. 89 DNA tissue samples (normal/cancer) and corresponding stool samples were analyzed in our study. In addition, 30 stool samples were collected as healthy controls.Results. The methylation level of those samples was measured by methylation-specific polymerase chain reaction (MSP). The result shows that compared with the paired controls, both SNCA and FBN1 were significantly hypermethylated in CRC patients in tissue samples (P<0.001). In the stool samples, hypermethylated SNCA and FBN1 were detected to be significantly higher than that in normal stool samples (P<0.001). The combined sensitivity of at least one positive among the two markers in stool samples was 84.3%, with a specificity of 93.3%. In addition, our experiment suggested that the positive rates of SNCA and FBN1 in Dukes A stage were significantly higher than that of FOBT (P=0.039;P=0.006, resp.).Conclusion. We concluded that methylation testing of SNCA and FBN1 genes in stool sample may offer a good alternative in a simple, promising, and noninvasive detection of colorectal cancer.

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DNA Methylation Biomarkers for Blood-Based Colorectal Cancer Screening

Clinical Chemistry ◽

10.1373/clinchem.2007.095992 ◽

2008 ◽

Vol 54 (2) ◽

pp. 414-423 ◽

Cited By ~ 298

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.

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Prognosis of the combination therapy results in patients with synchronous metastases of colorectal cancer

Science and Innovations in Medicine ◽

10.35693/2500-1388-2019-4-3-60-64 ◽

2019 ◽

Vol 4 (3) ◽

pp. 60-64

Author(s):

S. V Kozlov ◽

O. I Kaganov ◽

A. A Moryatov ◽

A. M Kozlov ◽

A. P Borisov

Keyword(s):

Colorectal Cancer ◽

Liver Metastases ◽

Cytoreductive Surgery ◽

Thermal Ablation ◽

Combined Treatment ◽

Stage Iv ◽

Radiofrequency Thermal Ablation ◽

Mutational Status ◽

Synchronous Metastases ◽

Metastatic Liver

Objectives - to optimize the indications for radiofrequency thermal ablation in patients with synchronous multiple metastases of colorectal cancer to the liver on the basis of prognostic treatment results. Material and methods. The study group included 78 patients with colorectal cancer with synchronous multiple bilobar liver metastases, who have underwent combined treatment in the period of 2007- 2015, such as cytoreductive surgery removing the primary intestinal tumor in combination with RFA of metastases in the liver, followed by chemotherapy. Results. A computer program for preoperative risk assessment of disease progression was developed and introduced in clinical practice. It is based on the results of the analysis of the factors, predicting the risk of relapse during the first year after cytoreductive surgery with RFA of synchronous multiple CRC liver metastases. Conclusion. The index of metastatic liver damage (the product of the sum of the diameters of metastatic liver lesions by their number), the mutational status of the KRAS gene, CEA values are significant factors in predicting the progression of the disease, which can optimize indications for radiofrequency thermal ablation in the treatment of patients with stage IV CRC with synchronous metastases to the liver.

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Novel Diagnostic Biomarkers in Colorectal Cancer

International Journal of Molecular Sciences ◽

10.3390/ijms23020852 ◽

2022 ◽

Vol 23 (2) ◽

pp. 852

Author(s):

Aneta L. Zygulska ◽

Piotr Pierzchalski

Keyword(s):

Colorectal Cancer ◽

Somatic Mutations ◽

Tumor Tissue ◽

Locally Advanced ◽

Survival Rates ◽

High Sensitivity ◽

Diagnostic Biomarkers ◽

Blood Tests ◽

Stool Samples ◽

Endoscopic Methods

Colorectal cancer (CRC) is still a leading cause of cancer death worldwide. Less than half of cases are diagnosed when the cancer is locally advanced. CRC is a heterogenous disease associated with a number of genetic or somatic mutations. Diagnostic markers are used for risk stratification and early detection, which might prolong overall survival. Nowadays, the widespread use of semi-invasive endoscopic methods and feacal blood tests characterised by suboptimal accuracy of diagnostic results has led to the detection of cases at later stages. New molecular noninvasive tests based on the detection of CRC alterations seem to be more sensitive and specific then the current methods. Therefore, research aiming at identifying molecular markers, such as DNA, RNA and proteins, would improve survival rates and contribute to the development of personalized medicine. The identification of “ideal” diagnostic biomarkers, having high sensitivity and specificity, being safe, cheap and easy to measure, remains a challenge. The purpose of this review is to discuss recent advances in novel diagnostic biomarkers for tumor tissue, blood and stool samples in CRC patients.

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Expression of tumor pyruvate kinase M2 isoform in plasma and stool of patients with colorectal cancer or adenomatous polyps

10.21203/rs.2.16791/v2 ◽

2020 ◽

Author(s):

Farideh Rigi ◽

Aliakbar Jannatabad ◽

Azra Izanloo ◽

Reza Roshanravan ◽

Hamid Reza Hashemian ◽

...

Keyword(s):

Colorectal Cancer ◽

Pyruvate Kinase ◽

Sensitivity And Specificity ◽

Glycolytic Enzyme ◽

Adenomatous Polyps ◽

Plasma Samples ◽

Polyp Detection ◽

Proliferating Cells ◽

Pyruvate Kinase M2 ◽

Stool Samples

Abstract Background: Tumor pyruvate kinase M2 isoform (tM2-PK), which is an isoform of PK-glycolytic enzyme and appears on the surface of cancerous proliferating cells, has been used as a diagnostic biomarker for colorectal cancer (CRC). The aim of this study was to evaluate the tM2-PK measurement test for the diagnosis of CRCs and adenomatous polyps in plasma and stool samples in an Iranian population. Methods: In this prospective study, a total of 226 stool and 178 plasma samples were received from patients referred to colonoscopy units. tM2-PK enzyme was measured using two separate ScheBo-Biotech-AG ELISA kits for stool and plasma samples. Results: According to ROC curves, in the tumor group, at the cut-off value of 4 U/ml, the sensitivity of fecal tM2-PK test was 100% and the specificity was 68%, and in the polyp group, the sensitivity and specificity were 87% and 68%, respectively. For tumor detection in plasma specimens, a cut-off value >25 U/ml has a sensitivity and specificity of 90.9% and 91.3%, respectively. Similarly, for polyp detection, a cut-off value >19 U/ml has a sensitivity of 96.3% and the specificity of 85.5%. Conclusions: Based on our results, a cut-off range of 4.8-8 U/ml and >8 U/ml could be used to detect polyp and tumor in stool samples, respectively. Similarly, a cut-off range of 19-25 U/ml and >25 U/ml is recommended in plasma samples, suggesting tM2-PK test as a non-invasive assay to diagnose CRC and adenomatous polyps.

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