Native American ancestry and breast cancer risk in Colombian and Mexican women: ruling out potential confounding through ancestry-informative markers.

BACKGROUND: Latin American and Hispanic women are less likely to develop breast cancer (BC) than women of European descent. Observational studies have found an inverse relationship between the individual proportion of Native American ancestry and BC risk. Here, we use ancestry-informative markers to rule out potential confounding of this relationship, estimating the confounder-free effect of Native American ancestry on BC risk. METHODS AND STUDY POPULATION: We used the informativeness for assignment measure to select robust instrumental variables for the individual proportion of Native American ancestry. We then conducted separate Mendelian randomization (MR) analyses based on 1401 Colombian women, most of them from the central Andean regions of Cundinamarca and Huila, and 1366 Mexican women from Mexico City, Monterrey and Veracruz, supplemented by sensitivity and stratified analyses. RESULTS: The proportion of Colombian Native American ancestry showed a putatively causal protective effect on BC risk (inverse variance-weighted odds ratio [OR] = 0.974 per 1% increase in ancestry proportion, 95% confidence interval [CI] 0.970-0.978, p = 3.1 × 10-40). The corresponding OR for Mexican Native American ancestry was 0.988 (95% CI 0.987-0.990, p = 1.4 × 10-44). Stratified analyses revealed a stronger association between Native American ancestry and familial BC (Colombian women: OR = 0.958, 95% CI 0.952-0.964; Mexican women: OR = 0.973, 95% CI 0.969-0.978), and stronger protective effects on oestrogen receptor (ER)-positive BC than on ER-negative and triple-negative BC. CONCLUSIONS: The present results point to an unconfounded protective effect of Native American ancestry on BC risk in both Colombian and Mexican women which appears to be stronger for familial and ER-positive BC. These findings provide a rationale for personalised prevention programmes that take genetic ancestry into account, as well as for future admixture mapping studies.

Gallbladder Cancer Risk and Indigenous South American Mapuche Ancestry: Instrumental Variable Analysis Using Ancestry-Informative Markers.

A strong association between the proportion of indigenous South American Mapuche ancestry and the risk of gallbladder cancer (GBC) has been reported in observational studies. Chileans show the highest incidence of GBC worldwide, and the Mapuche are the largest indigenous people in Chile. We set out to assess the confounding-free effect of the individual proportion of Mapuche ancestry on GBC risk and to investigate the mediating effects of gallstone disease and body mass index (BMI) on this association. Genetic markers of Mapuche ancestry were selected based on the informativeness for assignment measure, and then used as instrumental variables in two-sample Mendelian randomization analyses and complementary sensitivity analyses. Results suggested a putatively causal effect of Mapuche ancestry on GBC risk (inverse variance-weighted (IVW) risk increase of 0.8% per 1% increase in Mapuche ancestry proportion, 95% CI 0.4% to 1.2%, p = 6.7 × 10-5) and also on gallstone disease (3.6% IVW risk increase, 95% CI 3.1% to 4.0%), pointing to a mediating effect of gallstones on the association between Mapuche ancestry and GBC. In contrast, the proportion of Mapuche ancestry showed a negative effect on BMI (IVW estimate -0.006 kg/m2, 95% CI -0.009 to -0.003). The results presented here may have significant implications for GBC prevention and are important for future admixture mapping studies. Given that the association between the individual proportion of Mapuche ancestry and GBC risk previously noted in observational studies appears to be free of confounding, primary and secondary prevention strategies that consider genetic ancestry could be particularly efficient.

Development and internal validation of a multifactorial risk prediction model for gallbladder cancer in a high-incidence country.

Since 2006, Chile has been implementing a gallbladder cancer (GBC) prevention program based on prophylactic cholecystectomy for gallstone patients aged 35 to 49 years. The effectiveness of this prevention program has not yet been comprehensively evaluated. We conducted a retrospective study of 473 Chilean GBC patients and 2137 population-based controls to develop and internally validate three GBC risk prediction models. The Baseline Model accounted for gallstones while adjusting for sex and birth year. Enhanced Model I also included the non-genetic risk factors: body mass index, educational level, Mapuche surnames, number of children and family history of GBC. Enhanced Model II further included Mapuche ancestry and the genotype for rs17209837. Multiple Cox regression was applied to assess the predictive performance, quantified by the area under the precision-recall curve (AUC-PRC) and the number of cholecystectomies needed (NCN) to prevent one case of GBC at age 70 years. The AUC-PRC for the Baseline Model (0.44%, 95%CI 0.42-0.46) increased by 0.22 (95%CI 0.15-0.29) when non-genetic factors were included, and by 0.25 (95%CI 0.20-0.30) when incorporating non-genetic and genetic factors. The overall NCN for Chileans with gallstones (115, 95%CI 104-131) decreased to 92 (95%CI 60-128) for Chileans with a higher risk than the median according to Enhanced Model I, and to 80 (95%CI 59-110) according to Enhanced Model II. In conclusion, age, sex and gallstones are strong risk factors for GBC, but consideration of other non-genetic factors and individual genotype data improves risk prediction and may optimize allocation of financial resources and surgical capacity.

Gallstones, Cholecystectomy, and Kidney Cancer: Observational and Mendelian Randomization Results Based on Large Cohorts.

BACKGROUND & AIMS: Gallstones (cholelithiasis) constitute a major health burden with high costs related to surgical removal of the gallbladder (cholecystectomy), generally indicated for symptomatic gallstones. The association between gallstones and cholecystectomy and kidney cancer is controversial. We comprehensively investigated this association, considering age at cholecystectomy and time from cholecystectomy to kidney cancer diagnosis, and assessed the causal effect of gallstones on kidney cancer risk by Mendelian randomization (MR). METHODS: We compared the risk of kidney cancer in cholecystectomized and noncholecystectomized patients (16.6 million in total) from the Swedish nationwide cancer, census, patient, and death registries using hazard ratios (HRs). For 2-sample and multivariable MR, we used summary statistics based on 408,567 UK Biobank participants. RESULTS: During a median follow-up of 13 years, 2627 of 627,870 cholecystectomized Swedish patients developed kidney cancer (HR, 1.17; 95% CI, 1.12-1.22). Kidney cancer risk was particularly increased in the first 6 months after cholecystectomy (HR, 3.79; 95% CI, 3.18-4.52) and in patients cholecystectomized before age 40 years (HR, 1.55; 95% CI, 1.39-1.72). MR results based on 18,417 patients with gallstones and 1788 patients with kidney cancer from the United Kingdom revealed a causal effect of gallstones on kidney cancer risk (9.6% risk increase per doubling in gallstone prevalence; 95% CI, 1.2%-18.8%). CONCLUSIONS: Both observational and causal MR estimates based on large prospective cohorts support an increased risk of kidney cancer in patients with gallstones. Our findings provide solid evidence for the compelling need to diagnostically rule out kidney cancer before and during gallbladder removal, to prioritize kidney cancer screening in patients undergoing cholecystectomy in their 30s, and to investigate the underlying mechanisms linking gallstones and kidney cancer in future studies.

Development of an Application for Electronic Retrieval of Patient and Sample Information in Latin American Regions with a High Incidence of Gallbladder Cancer.

The European-Latin American Consortium towards Eradication of Preventable Gallbladder Cancer, EULAT Eradicate GBC, is collecting high-quality data and samples in four Latin American countries with high gallbladder cancer incidence (Argentina, Bolivia, Chile, and Peru) to build a unique biorepository integrated into a tailored IT platform, to identify, validate, and functionally characterize new risk biomarkers, and to develop prediction models that integrate epidemiological and genetic-molecular risk factors. We decided to develop an application for electronic data collection to facilitate the retrieval of sociodemographic, clinical, lifestyle, dietary, and sample-related information from 15,000 Latin American study participants. The application EULAT eCollect will facilitate the work of study nurses, reduce time spent by participants, limit the use of paper and ink, minimize costs and errors associated with filling out written forms and subsequent digitisation, and support the monitoring of local recruitment rates and data quality. We describe in this article the design and implementation of the EULAT eCollect application, which started with the specification of functional and non-functional requirements, and ended with the implementation and validation of four separate application modules: Socio-Demographic Interview, Sample Information, Case Report Form, and Food-Frequency Questionnaire. We present both general and technical results, and our experience with the free and open-source software, Open Data Kit (ODK), which may be of interest for future related research projects, especially those on personalised cancer prevention carried out in low- and middle-income regions.

NGS allele counts versus called genotypes for testing genetic association.

RNA sequence data are commonly summarized as read counts. By contrast, so far there is no alternative to genotype calling for investigating the relationship between genetic variants determined by next-generation sequencing (NGS) and a phenotype of interest. Here we propose and evaluate the direct analysis of allele counts for genetic association tests. Specifically, we assess the potential advantage of the ratio of alternative allele counts to the total number of reads aligned at a specific position of the genome (coverage) over called genotypes. We simulated association studies based on NGS data from HapMap individuals. Genotype quality scores and allele counts were simulated using NGS data from the Personal Genome Project. Real data from the 1000 Genomes Project was also used to compare the two competing approaches. The average proportions of probability values lower or equal to 0.05 amounted to 0.0496 for called genotypes and 0.0485 for the ratio of alternative allele counts to coverage in the null scenario, and to 0.69 for called genotypes and 0.75 for the ratio of alternative allele counts to coverage in the alternative scenario (9% power increase). The advantage in statistical power of the novel approach increased with decreasing coverage, with decreasing genotype quality and with decreasing allele frequency - 124% power increase for variants with a minor allele frequency lower than 0.05. We provide computer code in R to implement the novel approach, which does not preclude the use of complementary data quality filters before or after identification of the most promising association signals. Author summary: Genetic association tests usually rely on called genotypes. We postulate here that the direct analysis of allele counts from sequence data improves the quality of statistical inference. To evaluate this hypothesis, we investigate simulated and real data using distinct statistical approaches. We demonstrate that association tests based on allele counts rather than called genotypes achieve higher statistical power with controlled type I error rates.

Identification of Circulating lncRNAs Associated with Gallbladder Cancer Risk by Tissue-Based Preselection, Cis-eQTL Validation, and Analysis of Association with Genotype-Based Expression.

Long noncoding RNAs (lncRNAs) play key roles in cell processes and are good candidates for cancer risk prediction. Few studies have investigated the association between individual genotypes and lncRNA expression. Here we integrate three separate datasets with information on lncRNA expression only, both lncRNA expression and genotype, and genotype information only to identify circulating lncRNAs associated with the risk of gallbladder cancer (GBC) using robust linear and logistic regression techniques. In the first dataset, we preselect lncRNAs based on expression changes along the sequence "gallstones → dysplasia → GBC". In the second dataset, we validate associations between genetic variants and serum expression levels of the preselected lncRNAs (cis-lncRNA-eQTLs) and build lncRNA expression prediction models. In the third dataset, we predict serum lncRNA expression based on individual genotypes and assess the association between genotype-based expression and GBC risk. AC084082.3 and LINC00662 showed increasing expression levels (p-value = 0.009), while C22orf34 expression decreased in the sequence from gallstones to GBC (p-value = 0.04). We identified and validated two cis-LINC00662-eQTLs (r2 = 0.26) and three cis-C22orf34-eQTLs (r2 = 0.24). Only LINC00662 showed a genotyped-based serum expression associated with GBC risk (OR = 1.25 per log2 expression unit, 95% CI 1.04-1.52, p-value = 0.02). Our results suggest that preselection of lncRNAs based on tissue samples and exploitation of cis-lncRNA-eQTLs may facilitate the identification of circulating noncoding RNAs linked to cancer risk.

Inbreeding, Native American ancestry and child mortality: linking human selection and paediatric medicine.

The children of related parents show increased risk of early mortality. The Native American genome typically exhibits long stretches of homozygosity, and Latin Americans are highly heterogeneous regarding the individual burden of homozygosity, the proportion and the type of Native American ancestry. We analysed nationwide mortality and genome-wide genotype data from admixed Chileans to investigate the relationship between common causes of child mortality, homozygosity and Native American ancestry. Results from two-stage linear-Poisson regression revealed a strong association between the sum length of runs of homozygosity (SROH) above 1.5 Megabases (Mb) in each genome and mortality due to intracranial non-traumatic haemorrhage of foetus and newborn (5% increased risk of death per Mb in SROH, P = 1 × 10-3) and disorders related to short gestation and low birth weight (P = 3 × 10-4). The major indigenous populations in Chile are Aymara-Quechua in the north of the country and the Mapuche-Huilliche in the south. The individual proportion of Aymara-Quechua ancestry was associated with an increased risk of death due to anencephaly and similar malformations (P = 4 × 10-5), and the risk of death due to Edwards and Patau trisomy syndromes decreased 4% per 1% Aymara-Quechua ancestry proportion (P = 4 × 10-4) and 5% per 1% Mapuche-Huilliche ancestry proportion (P = 2 × 10-3). The present results suggest that short gestation, low birth weight and intracranial non-traumatic haemorrhage mediate the negative effect of inbreeding on human selection. Independent validation of the identified associations between common causes of child death, homozygosity and fine-scale ancestry proportions may inform paediatric medicine.

Ceramic Crowns and Sleep Bruxism: First Results from a Randomized Trial.

Background: This randomized clinical trial was conducted to assess whether sleep bruxism (SB) is associated with an increased rate of technical complications (ceramic defects) in lithium disilicate (LiDi) or zirconia (Z) molar single crowns (SCs). Methods: Adult patients were classified as affected or unaffected by SB based on structured questionnaires, clinical signs, and overnight portable electromyography (BruxOff) and block randomized into four groups according to SB status and crown material (LiDi or Z): LiDi-SB (n = 29), LiDi-no SB (n = 24), Z-SB (n = 23), and Z-no SB (n = 27). Differences in technical complications (main outcome) and survival and success rates (secondary outcomes) one year after crown cementation were assessed using Fisher's exact test with significance level α = 0.05. Results: No technical complications occurred. Restoration survival rates were 100% in the LiDi-SB and LiDi-no SB groups, 95.7% in the Z-SB group, and 96.3% in the Z-no SB group (p > 0.999). Success rates were 96.6% in the LiDi-SB group, 95.8% in the LiDi-no SB group (p > 0.999), 91.3% in the Z-SB group, and 96.3% in the Z-no SB group (p ≥ 0.588). Conclusions: With a limited observation time and sample size, no effect of SB on technical complication, survival, and success rates of molar LiDi and Z SCs was detected.

A New Pipeline for the Normalization and Pooling of Metabolomics Data.

Pooling metabolomics data across studies is often desirable to increase the statistical power of the analysis. However, this can raise methodological challenges as several preanalytical and analytical factors could introduce differences in measured concentrations and variability between datasets. Specifically, different studies may use variable sample types (e.g., serum versus plasma) collected, treated, and stored according to different protocols, and assayed in different laboratories using different instruments. To address these issues, a new pipeline was developed to normalize and pool metabolomics data through a set of sequential steps: (i) exclusions of the least informative observations and metabolites and removal of outliers; imputation of missing data; (ii) identification of the main sources of variability through principal component partial R-square (PC-PR2) analysis; (iii) application of linear mixed models to remove unwanted variability, including samples' originating study and batch, and preserve biological variations while accounting for potential differences in the residual variances across studies. This pipeline was applied to targeted metabolomics data acquired using Biocrates AbsoluteIDQ kits in eight case-control studies nested within the European Prospective Investigation into Cancer and Nutrition (EPIC) cohort. Comprehensive examination of metabolomics measurements indicated that the pipeline improved the comparability of data across the studies. Our pipeline can be adapted to normalize other molecular data, including biomarkers as well as proteomics data, and could be used for pooling molecular datasets, for example in international consortia, to limit biases introduced by inter-study variability. This versatility of the pipeline makes our work of potential interest to molecular epidemiologists.

Survival and success of tooth-implant-supported and solely implant-supported double-crown-retained overdentures: A prospective study over a period of up to 11 years.

OBJECTIVES: To investigate the long-term performance, over a period of up to 11 years, of tooth-implant-supported and solely implant-supported double-crown-retained overdentures (DCRDs) for complete restoration of the jaw. MATERIALS AND METHODS: Between 2002 and 2015, patients were consecutively enrolled in the study at Heidelberg University Hospital's Department of Prosthodontics. Scheduled and unscheduled visits were both documented on standardized forms. Kaplan-Meier curves were used to evaluate survival and success of the DCRDs. Mixed effects Cox regression was used to identify the potential effects of age, sex, jaw, location of implants, and the number of abutment teeth on the success of the DCRDs. RESULTS: One hundred and thirty-nine DCRDs were placed in 126 participants (mean age: 65.6 ± 9.1 years; 76 [60.3%] men). The mean follow-up period was 4.2 years. Fifty-three (38%) DCRDs were solely implant-supported (213 implants) and 86 (62%) were tooth-implant-supported (239 teeth and 199 implants). Cumulative five-year survival was 96.2% in the implant-supported group and 97.7% in the tooth-implant-supported group. Simultaneous consideration of failures and severe complications revealed cumulative five-year success of 88.2% for implant-supported DCRDs and 81.9% for tooth-implant-supported DCRDs, with no significant difference between the two types of restoration. Cox regression analysis revealed a greater risk of major complications (hazard ratio: 4.87, p = .04) for maxillary DCRDs than for mandibular DCRDs. CONCLUSIONS: Although the study design has limitations, the results show that tooth-implant-supported and solely implant-supported DCRDs are both recommendable treatment options. After a mean follow-up of 4.2 years, survival and success were high. However, minor complications were common and required much aftercare. CLINICAL TRIAL REGISTRATION: This study started in 2005 and was approved by the local review board of the University of Heidelberg. In 2005, clinical trial registration was not yet common. For this reason, the study was not registered.

Maxillary implant overdentures on two or four implants. A prospective randomized cross-over clinical trial of implant and denture success and survival.

OBJECTIVES: To determine implant survival and success of four conventionally but initially asynchronously loaded implants in implant-supported overdentures for the edentulous maxilla, in participants with opposing mandibular two-implant overdentures. MATERIAL AND METHODS: Twenty-six participants received four implants in the region of the maxillary canines and molars. After healing, 24 of these participants (mean age: 68.3 years) were randomly allocated to one of two treatment groups, and the adapted overdenture was attached to two unsplinted cylindrical attachments. The other two matrixes remained unattached to the implants for 3 months. After this period, the other two implants were loaded for 3 months (cross-over design). Then, all four implants were loaded for another 3 months. Kaplan-Meier curves were used to evaluate survival and success of implants and dentures. RESULTS: During the active prosthetic study phase, three participants lost one implant. Two participants lost three implants during the recall period. Implant survival after loading was 93.8% after a mean observation period of 3.1 years. Denture survival was 100%, but denture success was 95.8%, due to major prosthetic complications. Most participants preferred four implants to two. CONCLUSIONS: Within the limitations of the study, it can be concluded that maxillary implant overdentures on two or four implants are both recommendable treatment options. Two posterior implants are not superior to two anterior implants under overdentures retained by unsplinted cylindrical attachments. Implant and prosthetic complications and aftercare measures are common but are mostly easy to handle. However, 23 of the 24 participants preferred the 4-implant maxillary overdenture.

Gallstones, Body Mass Index, C-Reactive Protein, and Gallbladder Cancer: Mendelian Randomization Analysis of Chilean and European Genotype Data.

BACKGROUND AND AIMS: Gallbladder cancer (GBC) is a neglected disease with substantial geographical variability: Chile shows the highest incidence worldwide, while GBC is relatively rare in Europe. Here, we investigate the causal effects of risk factors considered in current GBC prevention programs as well as C-reactive protein (CRP) level as a marker of chronic inflammation. APPROACH AND RESULTS: We applied two-sample Mendelian randomization (MR) using publicly available data and our own data from a retrospective Chilean and a prospective European study. Causality was assessed by inverse variance weighted (IVW), MR-Egger regression, and weighted median estimates complemented with sensitivity analyses on potential heterogeneity and pleiotropy, two-step MR, and mediation analysis. We found evidence for a causal effect of gallstone disease on GBC risk in Chileans (P = 9 × 10-5 ) and Europeans (P = 9 × 10-5 ). A genetically elevated body mass index (BMI) increased GBC risk in Chileans (P = 0.03), while higher CRP concentrations increased GBC risk in Europeans (P = 4.1 × 10-6 ). European results suggest causal effects of BMI on gallstone disease (P = 0.008); public Chilean data were not, however, available to enable assessment of the mediation effects among causal GBC risk factors. CONCLUSIONS: Two risk factors considered in the current Chilean program for GBC prevention are causally linked to GBC risk: gallstones and BMI. For Europeans, BMI showed a causal effect on gallstone risk, which was itself causally linked to GBC risk.

Integrative molecular characterisation of gallbladder cancer reveals micro-environment-associated subtypes.

BACKGROUND & AIMS: Gallbladder cancer (GBC) is the most common type of biliary tract cancer, but the molecular mechanisms involved in gallbladder carcinogenesis remain poorly understood. In this study, we applied integrative genomics approaches to characterise GBC and explore molecular subtypes associated with patient survival. METHODS: We profiled the mutational landscape of GBC tumours (whole-exome sequencing on 92, targeted sequencing on 98, in total 190 patients). In a subset (n = 45), we interrogated the matched transcriptomes, DNA methylomes, and somatic copy number alterations. We explored molecular subtypes identified through clustering tumours by genes whose expression was associated with survival in 47 tumours and validated subtypes on 34 publicly available GBC cases. RESULTS: Exome analysis revealed TP53 was the most mutated gene. The overall mutation rate was low (median 0.82 Mut/Mb). APOBEC-mediated mutational signatures were more common in tumours with higher mutational burden. Aflatoxin-related signatures tended to be highly clonal (present in ≥50% of cancer cells). Transcriptome-wide survival association analysis revealed a 95-gene signature that stratified all GBC patients into 3 subtypes that suggested an association with overall survival post-resection. The 2 poor-survival subtypes were associated with adverse clinicopathologic features (advanced stage, pN1, pM1), immunosuppressive micro-environments (myeloid-derived suppressor cell accumulation, extensive desmoplasia, hypoxia) and T cell dysfunction, whereas the good-survival subtype showed the opposite features. CONCLUSION: These data suggest that the tumour micro-environment and immune profiles could play an important role in gallbladder carcinogenesis and should be evaluated in future clinical studies, along with mutational profiles. LAY SUMMARY: Gallbladder cancer is highly fatal, and its causes are poorly understood. We evaluated gallbladder tumours to see if there were differences between tumours in genetic information such as DNA and RNA. We found evidence of aflatoxin exposure in these tumours, and immune cells surrounding the tumours were associated with survival.

Robust Huber-LASSO for improved prediction of protein, metabolite and gene expression levels relying on individual genotype data.

Least absolute shrinkage and selection operator (LASSO) regression is often applied to select the most promising set of single nucleotide polymorphisms (SNPs) associated with a molecular phenotype of interest. While the penalization parameter λ restricts the number of selected SNPs and the potential model overfitting, the least-squares loss function of standard LASSO regression translates into a strong dependence of statistical results on a small number of individuals with phenotypes or genotypes divergent from the majority of the study population-typically comprised of outliers and high-leverage observations. Robust methods have been developed to constrain the influence of divergent observations and generate statistical results that apply to the bulk of study data, but they have rarely been applied to genetic association studies. In this article, we review, for newcomers to the field of robust statistics, a novel version of standard LASSO that utilizes the Huber loss function. We conduct comprehensive simulations and analyze real protein, metabolite, mRNA expression and genotype data to compare the stability of penalization, the cross-iteration concordance of the model, the false-positive and true-positive rates and the prediction accuracy of standard and robust Huber-LASSO. Although the two methods showed controlled false-positive rates ≤2.1% and similar true-positive rates, robust Huber-LASSO outperformed standard LASSO in the accuracy of predicted protein, metabolite and gene expression levels using individual SNP data. The conducted simulations and real-data analyses show that robust Huber-LASSO represents a valuable alternative to standard LASSO in genetic studies of molecular phenotypes.

Epigenome-Wide Analysis of Methylation Changes in the Sequence of Gallstone Disease, Dysplasia, and Gallbladder Cancer.

BACKGROUND AND AIMS: Gallbladder cancer (GBC) is a highly aggressive malignancy of the biliary tract. Most cases of GBC are diagnosed in low-income and middle-income countries, and research into this disease has long been limited. In this study we therefore investigate the epigenetic changes along the model of GBC carcinogenesis represented by the sequence gallstone disease → dysplasia → GBC in Chile, the country with the highest incidence of GBC worldwide. APPROACH AND RESULTS: To perform epigenome-wide methylation profiling, genomic DNA extracted from sections of formalin-fixed, paraffin-embedded gallbladder tissue was analyzed using Illumina Infinium MethylationEPIC BeadChips. Preprocessed, quality-controlled data from 82 samples (gallstones n = 32, low-grade dysplasia n = 13, high-grade dysplasia n = 9, GBC n = 28) were available to identify differentially methylated markers, regions, and pathways as well as changes in copy number variations (CNVs). The number and magnitude of epigenetic changes increased with disease development and predominantly involved the hypermethylation of cytosine-guanine dinucleotide islands and gene promoter regions. The methylation of genes implicated in Wnt signaling, Hedgehog signaling, and tumor suppression increased with tumor grade. CNVs also increased with GBC development and affected cyclin-dependent kinase inhibitor 2A, MDM2 proto-oncogene, tumor protein P53, and cyclin D1 genes. Gains in the targetable Erb-B2 receptor tyrosine kinase 2 gene were detected in 14% of GBC samples. CONCLUSIONS: Our results indicate that GBC carcinogenesis comprises three main methylation stages: early (gallstone disease and low-grade dysplasia), intermediate (high-grade dysplasia), and late (GBC). The identified gradual changes in methylation and CNVs may help to enhance our understanding of the mechanisms underlying this aggressive disease and eventually lead to improved treatment and early diagnosis of GBC.

Long-term chipping and failure rates of implant-supported and combined tooth-implant-supported metal-ceramic and ceramic fixed dental prostheses: A cohort study.

STATEMENT OF PROBLEM: Combined tooth-implant-supported fixed dental prostheses have been associated with an increased risk of long-term failure. Furthermore, high chipping rates have been reported for ceramic fixed dental prostheses. However, clinical data are sparse. PURPOSE: The purpose of this observational cohort study was to evaluate the chipping and failure rates of metal-ceramic and ceramic implant-supported and combined tooth-implant-supported fixed dental prostheses. MATERIAL AND METHODS: Four hundred thirty-four fixed dental prostheses placed in 324 patients (mean age: 60.8 years) were selected from a prospective clinical long-term study comprising 213 implant-supported fixed dental prostheses, 66 implant-supported cantilever fixed dental prostheses, and 155 tooth-implant-supported fixed dental prostheses. Metal-ceramic fixed dental prostheses (n=260) were fabricated with a high noble metal alloy (n=225) or Co-Cr base metal alloy (n=35) frameworks. Ceramic fixed dental prostheses (n=174) were all zirconia based and had monolithic (n=68), completely veneered (n=43), or partially veneered frameworks (n=63). Kaplan-Meier curves were used to estimate the survival probability and the chipping-free survival rate of the fixed dental prostheses. RESULTS: During the observation period of 0.5 to 12.6 years (mean: 4.26 years), 17 fixed dental prostheses failed because of implant failure (n=6), tooth loss (n=5), major chipping (n=5), or abutment screw loosening (n=1). Survival probability was 96% after 5 years and 91% after 10 years. Cox regression analysis showed that age, sex, fixed dental prosthesis location, type of fixed dental prosthesis support, and fixed dental prosthesis material had no significant effect on fixed dental prosthesis failure. Chipping (n=61) was significantly affected by the framework material and type of veneer (P=.001). After 5 years, the greatest incidence of chipping (39%) was observed for zirconia fixed dental prostheses with a complete veneer compared with an 18% incidence of chipping for metal-ceramic fixed dental prostheses with a high noble metal framework. A lower incidence of chipping was observed for zirconia fixed dental prostheses with a partial veneer or monolithic design. CONCLUSIONS: Implant-implant-supported and combined tooth-implant-supported fixed dental prostheses have promising long-term survival rates. Chipping seems to occur less frequently in monolithic or partially veneered fixed dental prostheses than in fixed dental prostheses with complete veneers.

Genotype-Based Gene Expression in Colon Tissue-Prediction Accuracy and Relationship with the Prognosis of Colorectal Cancer Patients.

Colorectal cancer (CRC) survival has environmental and inherited components. The expression of specific genes can be inferred based on individual genotypes-so called expression quantitative trait loci. In this study, we used the PrediXcan method to predict gene expression in normal colon tissue using individual genotype data from 91 CRC patients and examined the correlation ρ between predicted and measured gene expression levels. Out of 5434 predicted genes, 58% showed a negative ρ value and only 16% presented a ρ higher than 0.10. We subsequently investigated the association between genotype-based gene expression in colon tissue for genes with ρ > 0.10 and survival of 4436 CRC patients. We identified an inverse association between the predicted expression of ARID3B and CRC-specific survival for patients with a body mass index greater than or equal to 30 kg/m2 (HR (hazard ratio) = 0.66 for an expression higher vs. lower than the median, p = 0.005). This association was validated using genotype and clinical data from the UK Biobank (HR = 0.74, p = 0.04). In addition to the identification of ARID3B expression in normal colon tissue as a candidate prognostic biomarker for obese CRC patients, our study illustrates the challenges of genotype-based prediction of gene expression, and the advantage of reassessing the prediction accuracy in a subset of the study population using measured gene expression data.

Polymorphisms in the Angiogenesis-Related Genes EFNB2, MMP2 and JAG1 Are Associated with Survival of Colorectal Cancer Patients.

An individual's inherited genetic variation may contribute to the 'angiogenic switch', which is essential for blood supply and tumor growth of microscopic and macroscopic tumors. Polymorphisms in angiogenesis-related genes potentially predispose to colorectal cancer (CRC) or affect the survival of CRC patients. We investigated the association of 392 single nucleotide polymorphisms (SNPs) in 33 angiogenesis-related genes with CRC risk and survival of CRC patients in 1754 CRC cases and 1781 healthy controls within DACHS (Darmkrebs: Chancen der Verhütung durch Screening), a German population-based case-control study. Odds ratios and 95% confidence intervals (CI) were estimated from unconditional logistic regression to test for genetic associations with CRC risk. The Cox proportional hazard model was used to estimate hazard ratios (HR) and 95% CIs for survival. Multiple testing was adjusted for by a false discovery rate. No variant was associated with CRC risk. Variants in EFNB2, MMP2 and JAG1 were significantly associated with overall survival. The association of the EFNB2 tagging SNP rs9520090 (p < 0.0001) was confirmed in two validation datasets (p-values: 0.01 and 0.05). The associations of the tagging SNPs rs6040062 in JAG1 (p-value 0.0003) and rs2241145 in MMP2 (p-value 0.0005) showed the same direction of association with overall survival in the first and second validation sets, respectively, although they did not reach significance (p-values: 0.09 and 0.25, respectively). EFNB2, MMP2 and JAG1 are known for their functional role in angiogenesis and the present study points to novel evidence for the impact of angiogenesis-related genetic variants on the CRC outcome.