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Purpose: Improved prognostic stratification of patients with stage II and III colorectal cancer is warranted for postoperative clinical decision making. This study was conducted to develop a clinically feasible and robust prognostic classifier for these patients independent of adjuvant treatment.Experimental Design: Global gene expression profiles from altogether 387 stage II and III colorectal cancer tissue samples from three independent patient series were included in the study. ColoGuidePro, a seven-gene prognostic classifier, was developed from a selected Norwegian learning series (n = 95; no adjuvant treatment) using lasso-penalized multivariate survival modeling with cross-validation.Results: The expression signature significantly stratified patients in a consecutive Norwegian test series, in which patients were treated according to current standards [HR, 2.9 (1.1–7.5); P = 0.03; n = 77] and an external validation series [HR, 3.7 (2.0–6.8); P < 0.001; n = 215] according to survival. ColoGuidePro was also an independent predictor of prognosis in multivariate models including tumor stage in both series (HR, ≥3.1; P ≤ 0.03). In the validation series, which consisted of patients from other populations (United States and Australia), 5-year relapse-free survival was significantly predicted for stage III patients only (P < 0.001; n = 107). Here, prognostic stratification was independent of adjuvant treatment (P = 0.001).Conclusions: We present ColoGuidePro, a prognostic classifier developed for patients with stage II and III colorectal cancer. The test is suitable for transfer to clinical use and has best prognostic prediction potential for stage III patients. Clin Cancer Res; 18(21); 6001–10. ©2012 AACR.

Supplemental Materials, Figures S1-6, Tables S1-4. Fig. S1. Flow diagram for inclusion of patients in the study. Fig. S2. RNA secondary structure analysis of the 5'UTR RCC2 mutation compared to wild type. Fig. S3. Vector construction overview. Fig. S4. Morphological changes in actin fiber patterns following depletion of RCC2 protein in HCT15 cells. Fig. S5. Subgroup analyses of in situ protein expression of RCC2 for a consecutive CRC series. Fig. S6. Representative electropherograms from fragment analysis of the 5'UTR RCC2 mutation. Table S1. Statistics for relevant parameters for the test series (n = 37). Table S2. Statistics for relevant parameters for the validation series (n = 122). Table S3. Statistics for relevant parameters for the validation series (n = 60, R0 patients). Table S4. Primer and PCR details.

PDF file - 292K, The Supplementary Material contains Supplementary Methods, Supplementary Tables S1 and S2, and Supplementary Figures S1-S4 Table S1. Genes in the prognostic expression signature Table S2. Cox proportional hazards analyses for patients in the learning series Figure S1. Cross-validated partial likelihood and number of active predictors in the learning series as a function of the penalty parameter Figure S2. Expression in the learning series of the genes in the 7-gene prognostic signature. Figure S3. Survival curves for stage III patients in the validation series. Figure S4. Survival curves for stage III patients in the validation series aged 75 years or older

Purpose: Colorectal cancer has high incidence and mortality worldwide. Patients with microsatellite instable (MSI) tumors have significantly better prognosis than patients with microsatellite stable (MSS) tumors. Considerable variation in disease outcome remains a challenge within each subgroup, and our purpose was to identify biomarkers that improve prediction of colorectal cancer prognosis.Experimental Design: Mutation analyses of 42 MSI target genes were performed in two independent MSI tumor series (n = 209). Markers that were significantly associated with prognosis in the test series were assessed in the validation series, followed by functional and genetic explorations. The clinical potential was further investigated by immunohistochemistry in a population-based colorectal cancer series (n = 903).Results: We identified the cell-cycle gene regulator of chromosome condensation 2 (RCC2) as a cancer biomarker. We found a mutation in the 5′ UTR region of RCC2 that in univariate and multivariate analyses was significantly associated with improved outcome in the MSI group. This mutation caused reduction of protein expression in dual luciferase gene reporter assays. siRNA knockdown in MSI colon cancer cells (HCT15) caused reduced cell proliferation, cell-cycle arrest, and increased apoptosis. Massive parallel sequencing revealed few RCC2 mutations in MSS tumors. However, weak RCC2 protein expression was significantly associated with poor prognosis, independent of clinical high-risk parameters, and stratifies clinically important patient subgroups with MSS tumors, including elderly patients (>75 years), stage II patients, and those with rectal cancer.Conclusions: Impaired RCC2 affects functional and clinical endpoints of colorectal cancer. High-risk patients with either MSI or MSS tumors can be identified with cost-effective routine RCC2 assays. Clin Cancer Res; 21(16); 3759–70. ©2015 AACR.

The extensive and rapid development of the human brain during the first years of life complicates the postmortem diagnosis of brain edema in infancy. The aim of this study was to describe brain water content, the brain weight/body weight ratio, and the brain weight/head circumference ratio throughout the first years of life. Furthermore, we examined the relationship between these parameters and rs2075575 in the AQP4 gene. Our hypothesis was that dysregulated water homeostasis might be a risk factor for sudden infant death syndrome (SIDS), which may be reflected by increased water content in the brain. The study included 90 subjects with sudden unexpected death

Violent shaking is believed to be a common mechanism of injury in pediatric abusive head trauma. Typical intracranial injuries include subdural and retinal hemorrhages. Using a laboratory surrogate model we conducted experiments evaluating the head motion patterns that may occur in violent shaking. An anthropomorphic test device (ATD; Q0 dummy) matching an infant of 3.5 kg was assembled. The head interior was equipped with accelerometers enabling assessment of three-axial accelerations. Fifteen volunteers were asked to shake the surrogate vigorously holding a firm grip around the torso. We observed the volunteers performing manual shaking of the surrogate at a median duration of 15.5 sec (range 5–54 sec). Typical acceleration/deceleration patterns were produced after 2–3 shakes with a steady-state shaking motion at a pace of 4–6 cycles (back and forth) per second. Mean peak sagittal tangential accelerations at the vertex were 45.7g (range 14.2–105.1g). The acceleration component in the orthogonal direction, the radial acceleration, fluctuated around a negative mean of more than 4g showing that the surrogate head was continuously subjected to centripetal forces caused by rotations. This surrogate experiment showed that violent shaking may induce high peak tangential accelerations and concomitantly a continuous high-magnitude centripetal force. We hypothesize that the latter component may cause increased pressure in the subdural compartment in the cranial roof and may cause constant compression of the brain and possibly increased stretching or shearing of the bridging veins. This may contribute to the mechanism accountable for subdural hematoma in abusive head trauma.

Aim The aim of this study was to investigate if a range of known rare and common genetic variants in the Toll‐like receptor 4 (TLR4)/myeloid differentiation primary response 88 (MyD88) pathway were present or overrepresented in sudden infant death syndrome (SIDS) compared to controls. Methods Genetic variations in the genes encoding TLR4, MyD88 and Interleukin‐1 receptor‐associated kinase 4 were analysed. The subjects investigated included 158 SIDS cases with a median age of 15.25 weeks (2–47 weeks), 80 cases of infectious death with a median age of 24.9 weeks (0–285 weeks) and 199 adult controls with a median age of 50 years (11–86 years). The cases were collected in the years 1988–2017, and the autopsies were performed at the Department of Forensic Sciences at Oslo University Hospital, Oslo, Norway. Results The results showed that none of the genetic variants selected from the MyD88 pathway were associated with neither SIDS nor infectious death. Most of the rare genetic variants were homozygote for the common allele in all groups, while the rest revealed allelic variation. Conclusion The genetic variations investigated in this study did not appear to be involved in the pathogenesis of SIDS.

Several studies have indicated that a vulnerability in the development and regulation of brain function is involved in sudden infant death syndrome (SIDS). The aim of this study was to investigate the genes encoding the brain aquaporins (AQPs) AQP1 and AQP9 in SIDS. The hypothesis was that specific variants of these genes are part of the genetic vulnerability predisposing infants to sudden unexpected death. The study included 168 SIDS cases with a median age of 15.5 (range 2–52) weeks and 372 adolescent/adult deceased controls with a median age of 44 (range 11–91) years. In the AQP1 gene, the rs17159702 CC/CT genotypes were found to be associated with SIDS (p = 0.02). In the AQP9 gene, the combination of a TT genotype of rs8042354, rs2292711 and rs13329178 was more frequent in SIDS cases than in controls (p = 0.03). In the SIDS group, an association was found between genetic variations in the AQP1 gene and maternal smoking and between the 3xTT combination in the AQP9 gene and being found lifeless in a prone position. In conclusion, this study adds further evidence to the involvement of brain aquaporins in SIDS, suggesting that specific variants of AQP genes constitute a genetic predisposition, making the infant vulnerable to sudden death together with external risk factors and probably other genetic factors. Supplementary Information The online version contains supplementary material available at 10.1007/s00414-020-02493-9.

Aquaporin 4 (AQP4) is the main membrane water channel in the brain involved in regulating water homeostasis. The water distribution in neural tissue is often dysregulated after hypoxic neural injury. Previous research has indicated that victims of sudden infant death syndrome (SIDS) and sudden unexplained death in childhood (SUDC) have an underlying brain dysfunction that impairs their critical arousal response to hypoxic stress during sleep. The aim of this study was to determine the expression levels of AQP4 in the hippocampus in SIDS/SUDC cases and controls, and compare the findings with AQP4 genotypes that previously have been shown to be associated with SIDS. Immunochemical staining and morphometry were used to evaluate the density of AQP4-positive astrocytes in 30 SIDS/SUDC cases and 26 controls. AQP4-positive cells were counted in grids covering three layers in the hippocampus, which revealed that their count in any of the layers did not differ significantly between cases and controls. A decline in AQP4 expression was observed for infants older than 12 weeks. The AQP4 expression was lower in infants and children with the rs2075575 CT/TT genotype than in those with the CC genotype. This study indicates that AQP4 expression may be influenced by both age and genotype in infants. The role of AQP4 in the pathogenesis of SIDS remains to be elucidated.

This report details the proceedings and conclusions from the 3rd International Congress on Unexplained Deaths in Infants and Children, held November 26–27, 2018 at the Radcliffe Institute at Harvard University. The Congress was motivated by the increasing rejection of the diagnosis Sudden Infant Death Syndrome (SIDS) in the medical examiner community, leading to falsely depressed reported SIDS rates and undermining the validity and reliability of the diagnosis, which remains a leading cause of infant and child mortality. We describe the diagnostic shift away from SIDS and the practical issues contributing to it. The Congress was attended by major figures and opinion leaders in this area from countries significantly engaged in this problem. Four categories (International Classification of Diseases (ICD)-11 categories of MH11, MH12, MH14, PB00-PB0Z) were recommended for classification, and explicit definitions and guidance were provided for death certifiers. SIDS was reframed as unexplained sudden death in infancy or SIDS/MH11 to emphasize that either term signifies the lack of explanation following a rigorous investigation. A distinct category for children over the age of 1 was recommended (MH12). Definitions and exclusions were provided for the alternative categories of accidental asphyxia and undetermined. As recommended, unexplained sudden death in infancy or SIDS on a death certificate will code a unique, trackable entity, accurately reflecting the inability to determine a definitive explanation, while satisfying surveillance needs and reliable identification for research efforts. The conclusions will be submitted to the World Health Organization for inclusion in the upcoming ICD-11.

Background Genetic predispositions in cases suffering sudden unexpected infant death have been a research focus worldwide during the past decade. Despite large efforts, there is still uncertainty concerning the molecular pathogenesis of these deaths. With genetic technology in constant development, the possibility of an alternative approach into this research field has become available, like mRNA expression studies. Methods In this study, we investigated mRNA gene expression in 14 cases who died suddenly and unexpectedly from infection without a history of severe illness prior to death. The control group included eight accidents, two cases of natural death, one undetermined, one case of medical malpractice, and two homicides. The study included tissue from liver, heart, and brain using Illumina whole-genome gene expression assay. Results From the array, 19 genes showed altered expression in the infectious deaths compared to controls. Tissue from the heart showed 15 genes with altered mRNA expression compared to the control group. Conclusions Downregulation of KCNE5 in heart tissue from cases of infectious death was of particular interest. Variants of KCNE5 are associated with Brugada syndrome and sudden death and could be responsible for the fatal outcome in the group of infectious death. Impact KCNE5 is downregulated in tissue from the heart in cases of infectious death in infancy.This study provides knowledge about the gene expression profile in cases of infectious death.Variants of a gene known to give increased risk of cardiac arrhythmia is downregulated in cases of infectious death in infancy.The results could give us better knowledge as to why some infants do not survive an infection.This study provides a candidate gene for future studies.

Background: During the pandemic outbreak of the 2009 swine influenza (A(H1N1) pdm09), 32 fatal cases occurred in Norway and 19 of these were included in this study. Objectives: We characterised pulmonary changes in these fatal Norwegian cases. Patients and Methods: Upon hospitalisation, detailed clinical information and specimens from the upper and lower respiratory pathways were collected. At post-mortem, lung tissue was collected, formalin-fixed and paraffin-embedded. Immunohistochemical and light microscopic examination was performed to visualise the local expression of the A(H1N1) pdm09 virus. Reverse transcription-polymerase chain reaction (RT-PCR) and pyrosequencing of the non-fixed specimens allowed the identification of mutations in the influenza virus surface glycoprotein (haemagglutinin gene) particularly at position 222. Results and Conclusions: The overall course of illness lasted from 2 to 40 days (median 9 days). Diffused alveolar damage (DAD) was evident in 11 cases, 4 of which had no apparent underlying illness. Obesity was prominent in 12 cases, where three individuals were classified as otherwise healthy. The HA D222G mutation was detected in six cases, 3 of which had no underlying illness. Immunohistochemistry showed the A(H1N1)pdm09 virus to be prominent at the site of inflammation both in close proximity to and inside alveolar structures in the lung tissue. In addition to a possible role for the HA D222G mutation, our findings indicate that host factors and underlying conditions in the infected individuals are fundamental for disease outcome in many cases. This study increases our understanding of determinants for the clinical outcome of pandemic influenza, which could guide future treatment. publishedVersion

Measurement of vitreous humor potassium (K(+)) has since the 1960s been recognized as an adjunct for estimation of time since death. In 1991 we introduced hypoxanthine (Hx) as a new marker. Furthermore we demonstrated that time since death estimation was more accurate when ambient temperature was included in the calculations, both for K(+) and for Hx. In this paper we present a refined method. The subjects consist of 132 cases with known time of death and ambient temperature. One sample from each subject was used in the calculations. Vitreous humor Hx levels were available in all subjects, while K(+) was measured in 106 of the subjects, due to insufficient volume of vitreous humor. Linear regression analysis was applied to model the correlation between vitreous humor Hx and K(+), taking the interactions with temperature into consideration. The diagrams published in 1991, which also included ambient temperature, estimated median time since death with range between the 10th and 90th percentile, whereas the linear regression analysis presented in this paper estimates mean time since death with a corresponding 95% interval of confidence. We conclude that time since death may be estimated with relatively high precision applying vitreous humor Hx and K(+) concentrations combined with ambient temperature.

A large number of studies have tried to uncover a genetic predisposition for sudden infant death syndrome (SIDS), but there is still uncertainty concerning the pathogenesis of these deaths. The purpose of this study was to investigate mRNA gene expression in SIDS cases and controls, in order to uncover genes that are differentially expressed in the two groups. Tissue from brain, heart, and liver from 15 SIDS cases and 15 controls were included in the study, and mRNA expression was determined using the Illumina whole genome gene expression DASL HT assay. Seventeen genes showed significantly altered expression compared to controls, after correction for multiple testing. Three genes involved in the immune system were of particular interest, including the downregulation of MyD88 in tissue from SIDS brains, as well as the downregulation of the genes encoding CCL3 and UNC13 in the liver. These findings indicate that there is an altered expression of genes involved in the inflammatory process in a proportion of SIDS cases, which further strengthen the hypothesis that impaired immune response play a role in this syndrome.

Introduction Postmortem evaluations of cerebral edema typically involve examinations of macroscopic features such as the presence of pressure signs and compression of the ventricles. Global massive edema is easily detectable in an autopsy, but less-extensive edema may be difficult to diagnose. Aim The aim of this study was to compare measurements of brain water contents, postmortem CT radiodensity and brain weight to skull size in edematous and nonedematous brains in order to develop an objective method for postmortem evaluations of brain edema. Method Fifty-four subjects autopsied at Oslo University Hospital underwent a standard forensic postmortem examination, including a computed axial tomography (CT) scan, measurement of brain weight, and macroscopic evaluation of the brain. CT images were used to roughly measure the inner skull circumference. The water content of the brain was determined by excising samples of approximately 1 g of brain tissue from eight different areas of the brain surface, drying them, and measuring their percentage water content. Results The main finding was a significant relationship between brain weight and inner skull circumference, with the ratio between these two parameters being significantly higher in cases with severe postmortem brain edema than in cases with very little or no brain edema. The water content did not differ significantly between the edema and nonedema cases. There were no significant changes in radiodensity. Conclusion This indicates that the brain-weight-to-inner-skull-circumference ratio may serve as a good marker for severe brain edema in postmortem diagnostics, whereas measurements of water content can be misleading.

We report a case of a woman who experienced intrauterine fetal death at full term pregnancy, and then died suddenly soon after learning about the death of her fetus. At autopsy, previously undiagnosed neurofibromatosis and an adrenal gland pheochromocytoma were discovered in the mother. Genetic screening also revealed a novel KCNH2mutation in both fetus and mother indicating type 2 congenital long-QT syndrome (LQTS). A catecholamine surge was suspected as the precipitating event of fetal cardiac arrhythmia and sudden fetal death, while the addition of emotional stress provoked a lethal cardiac event in the mother. This case illustrates the potential for lethal interactions between two occult diseases (pheochromocytoma, LQTS).

Mild infection may trigger sudden death in the vulnerable infant by cytokine interactions with a compromised medullary serotonergic (5-HT) system, leading to disrupted cardiorespiratory regulation and sleep-related sudden death. The cytokine interleukin (IL)-6 is elevated in the cerebrospinal fluid in SIDS. We tested the hypothesis that the expression of IL-6 receptors (IL-6R) and/or gp130 (involved in IL-6R signaling) is altered in the medullary 5-HT system in SIDS. Immunohistochemistry of IL-6R and gp130 was performed on medullae from 25 SIDS infants, 20 infectious deaths, and 14 controls using a semi-quantitative grading system. In the SIDS cases, mean IL-6R intensity grade in the arcuate nucleus (major component of medullary 5-HT system) was significantly higher than in the control group (2.00 +/- 0.07 vs. 1.77 +/- 0.08, P = 0.04), with no other differences in IL-6R or gp130 expression at any other site. Arcuate 5-HT neurons expressed IL-6R, indicating a site of IL-6/5-HT interaction. In SIDS, IL-6R expression is abnormal in the arcuate nucleus, the putative human homolog of rodent ventral medullary chemosensitivity sites involving 5-HT. Aberrant interactions between IL-6 and the arcuate nucleus may contribute to impaired responses to hypercapnia generated by infection (hyper-metabolism) combined with rebreathing.

Gastrointestinal cancers include malignancies arising in the esophagus, liver and bile ducts, gallbladder, pancreas, stomach, small intestine, colon and rectum. Their incidence and mortality differ significantly, however, together they account for approximately one-fifth of the cancer incidence and nearly one-fourth of the cancer related deaths in the US.1 Colorectal and gastric cancers are the most common gastrointestinal tumors worldwide, accounting for ∼2.2 million new cases and an estimated 1.3 million deaths annually.2 When these malignancies are detected at an early, localized stage, the 5-year survival is 90% and 63%, respectively.3 However, more than 60% of the patients with colorectal cancer and 75% of the patients with gastric cancer have regional or distant metastases at the time of diagnosis, resulting in a significant drop in survival.3 Cholangiocarinoma and pancreatic cancer are less prevalent.2,4 However, these malignancies are often “clinically silent” and thus diagnosed at an advanced stage with a corresponding poor prognosis.2,4 Early detection, particularly of colorectal and gastric cancer, may significantly reduce the number of gastrointestinal cancer deaths, and identification of suitable biomarkers for this purpose is therefore warranted. In humans, DNA methylation occurs predominantly at the 5′ position of cytosine, within CpG dinucleotides. Approximately 70% of the human genes have a CpG island—an enrichment of such dinucleotides—in the promoter region.5 In normal cells these islands are usually unmethylated, but several of them become hypermethylated in cancer and this is associated with repressed or lost gene expression.6 Using genome-wide analyzes, DNA methylation was recently shown to be more frequent than genetic changes in colorectal cancer.7 In addition to being frequent, aberrant DNA methylation has been shown to be an early event in tumorigenesis.8 Finally, several examples of genes with promoter DNA hypermethylation have been detected in various bodily fluids from cancer patients, including bile, feces, plasma and urine,9–13 indicating that methylation biomarkers may be useful for non- or minimally invasive cancer diagnostics. Although cancer is a highly heterogeneous disease, comprising more than 200 different conditions, it is believed that tumors arising in the gastrointestinal tract share several molecular alterations. Gut derived adenocarcinomas have indeed been found to display similar copy-number changes, forming tissue-specific clusters when various cancer types were clustered according to amplification activated oncogenes.14,15 Gastrointestinal tumors further show extremely high frequencies of transition mutations at CpG dinucleotides,16 and several genes, including APC, MLH1, MGMT, p16INK4a, CDH1, SFRP1, RASSF1A and VIM have been shown to be epigenetically dysregulated across malignancies of the gastrointestinal tract.17–21 With increased focus on cross-tumor analysis, exemplified by The Cancer Genome Atlas (TCGA) Pan-Cancer project, more shared abnormalities among the gastrointestinal cancers are expected to be uncovered. In the present study we have investigated whether the recently identified methylation biomarkers for cholangiocarcinoma (CDO1, DCLK1, ZSCAN18 and SFRP1)22 were methylated also in other malignancies of the gastrointestinal tract. The promoter methylation frequency of these genes, in addition to ZNF331, previously shown to be methylated across several gastrointestinal cancer cell lines,22 was analyzed in tissue samples from colorectal-, gastric- and pancreatic cancer patients.

Colorectal cancer is a global health challenge with high incidence rate and mortality. The patients' prognosis is strongly associated with disease stage and currently there is a need for improved prognostic and predictive biomarkers. In this study, novel colorectal cancer-specific transcript structures were nominated from whole transcriptome sequencing of seven colorectal cancer cell lines, two primary colorectal carcinomas with corresponding normal colonic mucosa and 16 normal tissues. The nominated transcripts were combined with gene level outlier expression analyses in a cohort of 505 colorectal cancers to identify biomarkers with capacity to stratify colorectal cancer subgroups. The transcriptome sequencing data and outlier expression analysis revealed 11 novel colorectal cancer-specific exon-exon junctions, of which 3 were located in the gene VNN1. The junctions within VNN1 were further characterized using rapid amplification of cDNA ends (RACE) and the prevalence of the subsequently characterized novel transcript, VNN1-AB, was investigated by real-time RT-PCR in 291 samples of miscellaneous origins. VNN1-AB was not present in any of the 43 normal colorectal tissue samples investigated, but in 5 of the 6 polyps, and 102 of the 136 (75%) colorectal cancers. We have identified a novel transcript of the VNN1 gene, with an organ-confined complete specificity for colorectal neoplasia.