Validation of computational determination of microsatellite status using whole exome sequencing data from colorectal cancer patients.

BACKGROUND: Microsatellite instability (MSI), resulting from a defective mismatch repair system, occurs in approximately 15% of sporadic colorectal cancers (CRC). Since MSI is associated with a poor response to 5-fluorouracile based chemotherapy and is a positive predictive marker of immunotherapy, it is routine practice to evaluate the MSI status of resected tumors in CRC patients. MSIsensor is a novel computational tool for determining MSI status using Next Generation Sequencing. However, it is not widely used in the clinic and has not been independently validated in exome data from CRC. To facilitate clinical implementation of computational determination of MSI status, we compared MSIsensor to current gold standard methods for MSI testing. METHODS: MSI status was determined for 130 CRC patients (UICC stage I-IV) using immunohistochemistry, PCR based microsatellite stability testing and by applying MSIsensor to exome sequenced tumors and paired germline DNA. Furthermore, we investigated correlation between MSI status, mutational load and mutational signatures. RESULTS: Eighteen out of 130 (13.8%) patients were microsatellite instable. We found a 100% agreement between MSIsensor and gold standard methods for MSI testing. All MSI tumors were hypermutated. In addition, two microsatellite stable (MSS) tumors were hypermutated, which was explained by a dominant POLE signature and pathogenic POLE mutations (p.Pro286Arg and p.Ser459Phe). CONCLUSION: MSIsensor is a robust tool, which can be used to determine MSI status of tumor samples from exome sequenced CRC patients.

Prognostic value of preoperative D-dimer and carcinoembryonic antigen levels in patients undergoing intended curative resection for colorectal cancer: a prospective cohort study.

PURPOSE: Carcinoembryonic antigen (CEA) has limited value as an isolated predictor for survival among colorectal cancer (CRC) patients. D-dimer (DD) is a strong predictor of survival among metastatic CRC patients, but the prognostic value in non-metastatic CRC patients remains controversial. We examined the prognostic value of preoperative DD levels in relation to CEA levels in non-metastatic, resectable CRC patients. METHODS: Between October 2003 and November 2005, 166 patients were included. We used the Kaplan-Meier method to compute 5-year mortality rates, stratified by preoperative DD and CEA levels. Adjusted Cox regression analysis was used to compute mortality rate ratios (MRRs) during postoperative years 0-1 and 1-5 based on the preoperative CEA and DD levels. RESULTS: The cumulative 5-year mortality rate was 15 % (95 % confidence interval [CI], 9-25 %) in patients with normal DD and CEA levels, 30 % (CI, 16-53 %) in patients with isolated elevated CEA levels, 37 % (CI, 25-53 %) in patients with isolated elevated DD levels, and 60 % (CI, 37-83 %) in patients with elevated CEA and DD levels. Elevated CEA was associated with an approximately ten-fold increase in mortality within the first postoperative year (adjusted MRR 9.8, CI 2.5-38.3); this association was lost during postoperative years 1-5 (adjusted MRR 1.1, CI 0.5-2.7). Elevated DD was associated with a greater than two-fold increase in mortality during postoperative years 0-1 (adjusted MRR 2.8, CI 0.7-11.0) and 1-5 (adjusted MRR 2.2, CI 1.1-4.8). CONCLUSION: DD is a strong predictor of survival among non-metastatic curatively resected CRC patients, particularly in combination with CEA.

Novel DNA methylation biomarkers show high sensitivity and specificity for blood-based detection of colorectal cancer-a clinical biomarker discovery and validation study.

BACKGROUND: Early detection plays an essential role to reduce colorectal cancer (CRC) mortality. While current screening methods suffer from poor compliance, liquid biopsy-based strategies for cancer detection is rapidly gaining promise. Here, we describe the development of TriMeth, a minimal-invasive blood-based test for detection of early-stage colorectal cancer. The test is based on assessment of three tumour-specific DNA methylation markers in circulating cell-free DNA. RESULTS: A thorough multi-step biomarker discovery study based on DNA methylation profiles of more than 5000 tumours and blood cell populations identified CRC-specific DNA methylation markers. The DNA methylation patterns of biomarker candidates were validated by bisulfite sequencing and methylation-specific droplet digital PCR in CRC tumour tissue and peripheral blood leucocytes. The three best performing markers were first applied to plasma from 113 primarily early-stage CRC patients and 87 age- and gender-matched colonoscopy-verified controls. Based on this, the test scoring algorithm was locked, and then TriMeth was validated in an independent cohort comprising 143 CRC patients and 91 controls. Three DNA methylation markers, C9orf50, KCNQ5, and CLIP4, were identified, each capable of discriminating plasma from colorectal cancer patients and healthy individuals (areas under the curve 0.86, 0.91, and 0.88). When combined in the TriMeth test, an average sensitivity of 85% (218/256) was observed (stage I: 80% (33/41), stage II: 85% (121/143), stage III: 89% (49/55), and stage IV: 88% (15/17)) at 99% (176/178) specificity in two independent plasma cohorts. CONCLUSION: TriMeth enables detection of early-stage colorectal cancer with high sensitivity and specificity. The reported results underline the potential utility of DNA methylation-based detection of circulating tumour DNA in the clinical management of colorectal cancer.

The prognostic efficacy of cell-free DNA hypermethylation in colorectal cancer.

Epigenetic alterations in colorectal cancer (CRC) cause important differences in the underlying tumor biology and aggressiveness. DNA hypermethylation is central for the development of CRC but the prognostic impact remains elusive. We aimed to assess the association between cell-free hypermethylated DNA and stage and survival in colorectal cancer (CRC). We analyzed pre-treatment plasma samples from 193 patients with CRC. Thirty gene-promoter regions were analyzed using methylation specific PCR. We compared the median number (range) of hypermethylated promoter regions with CRC stage, and constructed a multivariable Cox-regression model adjusted for stage, to evaluate the added prognostic information. The median number of hypermethylated promoter regions was nine (0-28) in patients with distant metastasis compared to five (0-19) in patients without metastatic disease (p < 0.0001). The majority of the hypermethylated promoter regions inferred a poor prognosis. Cox-regression analysis adjusted for patient age, sex, pre-treatment CEA-levels, and disease stage, showed that RARB (HR = 1.99, 95% CI [1.07, 3.72]) and RASSF1A (HR = 3.35, 95% CI [1.76, 6.38]) hypermethylation inferred a significant effect on survival. The risk of metastasis increase with the number of cell-free hypermethylated promoter regions. The presence of RARB and RASSF1A hypermethylation indicated aggressive disease, regardless of stage at the time of diagnosis.

[Wandering spleen in a patient hospitalized with acute abdomen].

An 11-year-old girl was hospitalized with a two-day history of lower abdominal pain. The patient had a localized peritoneal reaction in the right lower quadrant, an elevated C-reactive protein level, and an increased white blood cell count. On the suspicion of acute appendicitis a diagnostic laparoscopy was performed. This revealed a misplaced spleen in the right side of the pelvis with a partly torsion of a long vascular pedicle. The splenic tissue was vital without any sign of ischaemia. However, splenopexy was not possible, and an uncomplicated elective splenectomy was performed.

Hypermethylated DNA, a circulating biomarker for colorectal cancer detection.

BACKGROUND: Colorectal cancer (CRC) is one of the most common cancers in the western world. Screening is an efficient method of reducing cancer-related mortality. Molecular biomarkers for cancer in general and CRC in particular have been proposed, and hypermethylated DNA from stool or blood samples are already implemented as biomarkers for CRC screening. We aimed to evaluate the performance of proven hypermethylated DNA promoter regions as plasma based biomarkers for CRC detection. METHODS: We conducted a cross-sectional case-control study of 193 CRC patients and 102 colonoscopy-verified healthy controls. Using methylation specific polymerase chain reaction, we evaluated 30 DNA promoter regions previously found to be CRC specific. We used multivariable logistic regression with stepwise backwards selection, and subsequent leave-pair-out cross validation, to calculate the optimism corrected area under the receiver operating characteristics curve (AUC) for all stage as well as early stage CRC. RESULTS: None of the individual DNA promoter regions provided an overall sensitivity above 30% at a reasonable specificity. However, seven hypermethylated promoter regions (ALX4, BMP3, NPTX2, RARB, SDC2, SEPT9, and VIM) along with the covariates sex and age yielded an optimism corrected AUC of 0.86 for all stage CRC and 0.85 for early stage CRC. Overall sensitivity for CRC detection was 90.7% at 72.5% specificity using a cut point value of 0.5. CONCLUSIONS: Individual hypermethylated DNA promoter regions have limited value as CRC screening markers. However, a panel of seven hypermethylated promoter regions show great promise as a model for CRC detection.

[From accident to trauma center--the lapse of time for patients with severe head injuries]. / Fra ulykke til traumecenter--tidsforløb for patienter med alvorlig hovedskade.

INTRODUCTION: Early neurosurgical intervention and specialised neurointensive care have been shown to decrease morbidity and mortality in cases of severe head injury. This makes quick or direct transfer to a trauma centre essential. The aim of this study was to investigate the time from the time of the accident required for secondarily transferred patients with head injury to arrive at the trauma centre in Aarhus. MATERIALS AND METHODS: This was a descriptive study based on consecutive data on patients secondarily transferred to Aarhus Trauma Centre in 2003. Only patients with head injury admitted to the neurosurgical intensive care unit were included. The time of the accident was defined as the time of dispatch of the ambulance. RESULTS: A total of 89 patients were transferred secondarily to the trauma centre in Aarhus 2003; 43 of these had head injury. The median Glasgow Coma Score was 6.5 (3-15). The median time from accident to arrival at the trauma centre was 3 hours and 50 minutes (44 minutes to 20 hours, 4 minutes), and 42% of the patients arrived later than 4 hours after the injury. The distance from the primary hospital to the trauma centre was between 1.9 and 172 kilometers, and there was no association between distance and time. DISCUSSION: The time from accident to arrival at the trauma centre was long, considering the severity of the injuries and the short distances involved. Direct transfer from the site of accident to the trauma centre would almost certainly improve the time. This study gives a reference value for the Danish trauma system.