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Next-generation sequencing (NGS) based population screening holds great promise for disease prevention and earlier diagnosis, but associated sequencing costs remain prohibitive. We developed double batched sequencing (DoBSeq) and tested it on neonatal blood spot DNA in an explorative (n = 100) and a validation (n = 100) cohort selected from a nation-wide childhood cancer whole genome sequencing (WGS) study. Each cohort was enriched for loss-of-function/pathogenic variants in cancer predisposition syndrome genes. Using a commercial 113-gene panel benchmarked against individual WGS data, we demonstrated replicable detection of deleterious and pathogenic variants, with positive and negative predictive values of 100% (95%CI 0.91-1.00 & 95%CI 0.98-1.00, respectively). Pathogenic variants were detected in RB1, TP53, BRCA2, APC, and 19 other genes. Analyses of larger batches (n = 24, 48, 72 & 96) indicate that DoBSeq is highly scalable and thus a promising, cost-effective method for realizing the potential of population-scale NGS-based screening.

ImportanceSpermine oxidase (SMOX) catalyzes the oxidation of spermine to spermidine. Observational studies have reported SMOX as a source of induced reactive oxygen species associated with cancer, implying that inhibition of SMOX could be a target for chemoprevention.ObjectiveTo test the causality of SMOX levels with cancer risk using a Mendelian randomization analysis.Design, Setting, and ParticipantsWe performed a GWAS of spermidine/spermine ratio, from blood of 534 infants, to identify genetic variants associated with regulation of SMOX activity. In two additional data sets of 262 newborns and 508 adults, we quantified SMOX gene expression using RNA-sequencing and performed expression and methylation QTL lookups. We then did a Mendelian randomization analysis by testing the association between the SMOX genetic instrument and various cancer types using GWAS summary statistics.Main Outcomes and MeasuresNeuroblastoma, gastric, lung, breast, prostate, and colorectal cancers.ResultsThe GWAS of spermidine/spermine ratio identified a genome-wide significant locus (P=1.34×10−49) explaining 32% of the variance. The lead SNP rs1741315 was also associated with SMOX gene expression in newborns (P=8.48×10−28) and adults (P=2.748×10−8) explaining 37% and 6% of the variance, respectively. rs1741315 was not associated with neuroblastoma (OR=0.95; 95% CI:0.88, 1.03; P=0.18), gastric (OR=0.99; 95% CI:0.95, 1.03; P=0.54), lung (OR=0.97; 95% CI:0.94, 1.00; P=0.08), breast (OR=0.99; 95% CI:0.96, 1.02; P=0.47), prostate (OR=0.98; 95% CI:0.96, 1.00; P=0.05) nor colorectal cancer (OR=1.03; 95% CI:0.99, 1.07; P=0.10). A PheWAS of rs1741315 did not reveal any associations with risk factors of the cancers tested.Conclusions and RelevanceGenetic variation in the SMOX gene was strongly associated with SMOX activity in newborns, and less strongly in adults. Genetic down-regulation of SMOX was not significantly associated with lower odds of neuroblastoma, gastric, lung, breast, prostate and colorectal cancer. Further studies are needed to understand the effect of SMOX inhibition in relation to cancer risk.ARTICLE SUMMARYQuestionIs SMOX causally associated with risk of cancer?FindingsIn this Mendelian randomization study, genetically lower levels of SMOX were not associated with decreased risk of neuroblastoma, gastric, lung, breast, prostate and colorectal cancer.MeaningThese findings do not support a causal association between SMOX activity and risk of cancer, suggesting that ongoing efforts to identify SMOX inhibitors for chemoprevention may not be successful.STRENGHTS AND LIMITATIONS-Previous studies which examined SMOX activity and cancer risk were susceptible to recall bias, confounding and reverse causation, none of which are concerns of this Mendelian randomization study.-Our genetic instrument explained a sizeable proportion of the variance of SMOX activity-We used summary statistics from the largest meta-analyses of primary cancer GWAS to date.-Elevated SMOX levels in cancer could also be due to environmental factors not captured by genetics.-Our genetic instrument was developed based on normal range SMOX activity data, thus additional genetic variants might play a role in aberrant expression of this enzyme.

N-acetyltransferase 2 (NAT2) is the main enzyme metabolizing isoniazid and genotype-based treatment has been studied for years without becoming common practice. To investigate whether genotype-based isoniazid treatment is feasible in Greenland, we sequenced the coding sequence of NAT2 and determined the NAT2 enzyme-activity by caffeine test. No additional genetic variants were identified in the coding sequence of NAT2, so that genotype status in 260 study participants could be assessed by a well-established 7-SNP panel. Studying the enzyme activity by the ratio of the two caffeine metabolites AFMU and 1X in 260 participants showed a high rate of slow phenotypes with intermediate or rapid genotype. These misclassifications were mainly observed in urine samples with pH3, we observed a moderate level of discrepancies (19 of the 116 individuals with intermediate or rapid genotype status having a slow phenotype). Further investigation showed that drinking coffee and not tea or cola was the most important factor for high levels of both metabolites. The concordance between phenotype and genotype status with regard to slow metabolism supported the recommendation of lower isoniazid doses in individuals with slow genotype status in order to avoid liver injury, a frequent side effect. The phenotypical variation observed for individuals with intermediate or rapid genotype status warrants further research before increased dosing of isoniazid can be recommended., EXCLI Journal;Vol. 17 2018

Prior studies suggest dental caries traits in children and adolescents are partially heritable, but there has been no large-scale consortium genome-wide association study (GWAS) to date. We therefore performed GWAS for caries in participants aged 2.5-18.0 years from 9 contributing centers. Phenotype definitions were created for the presence or absence of treated or untreated caries, stratified by primary and permanent dentition. All studies tested for association between caries and genotype dosage (imputed to Haplotype Reference Consortium or 1000 Genomes phase 1 version 3 panels) accounting for population stratification. Fixed–effects meta-analysis was performed weighted by inverse standard error. Analysis included up to 19,003 individuals (7,530 affected) for primary teeth and 13,353 individuals (5,875 affected) for permanent teeth. Evidence for association with caries status was observed at rs1594318-C for primary teeth (intronic within ALLC, Odds Ratio (OR) 0.85, Effect Allele Frequency (EAF) 0.60, p 4.13e-8) and rs7738851-A (intronic within NEDD9, OR 1.28, EAF 0.85, p 1.63e-8) for permanent teeth. Consortium-wide estimated heritability of caries was low (h2 of 1% [95% CI: 0%:7%] and 6% [95% CI 0%:13%] for primary and permanent dentitions, respectively) compared to corresponding within-study estimates (h2 of 28%, [95% CI: 9%:48%] and 17% [95% CI:2%:31%]) or previously published estimates. This study was designed to identify common genetic variants with modest effects which are consistent across different populations. We found few single variants associated with caries status under these assumptions. Phenotypic heterogeneity between cohorts and limited statistical power will have contributed; these findings could also reflect complexity not captured by our study design, such as genetic effects which are conditional on environmental exposure.Author summaryDental caries (tooth decay) is a common disease in children. Previous studies suggest genetic factors alter caries risk, but to date there is a gap of knowledge in identifying which specific genetic variants are responsible. We undertook analysis in a consortium including around 19,000 children and investigated whether any of 8 million common genetic variants were associated with risk of caries in primary (milk) or permanent teeth. If identified, these variants are used as ‘tags’ to highlight genes which may be involved in a disease. We identified variants in two loci associated with caries status; in the primary (rs1594318) and permanent dentition (rs7738851). The former is intronic in ALLC, a gene with poorly understood function. The latter is an intronic variant within NEDD9, a gene which has several known functions including a role in development of craniofacial structures. To gain a more comprehensive understanding of genetic effects which influence caries larger studies and a better understanding of environmental modifiers or interactions with genetic effects are required.

Background— Inuit have lived for thousands of years in an extremely cold environment on a diet dominated by marine-derived fat. To investigate how this selective pressure has affected the genetic regulation of fatty acid metabolism, we assessed 233 serum metabolic phenotypes in a population-based sample of 1570 Greenlanders. Methods and Results— Using array-based and targeted genotyping, we found that rs80356779, a p.Pro479Leu variant in CPT1A , was strongly associated with markers of n -3 fatty acid metabolism, including degree of unsaturation ( P =1.16×10 − 34 ), levels of polyunsaturated fatty acids, n -3 fatty acids, and docosahexaoenic acid relative to total fatty acid levels ( P =2.35×10 − 15 , P =4.02×10 − 19 , and P =7.92×10 − 27 ). The derived allele (L479) occurred at a frequency of 76.2% in our sample while being absent in most other populations, and we found strong signatures of positive selection at the locus. Furthermore, we found that each copy of L479 reduced height by an average of 2.1 cm ( P =1.04×10 − 9 ). In exome sequencing data from a sister population, the Nunavik Inuit, we found no other likely causal candidate variant than rs80356779. Conclusion— Our study shows that a common CPT1A missense mutation is strongly associated with a range of metabolic phenotypes and reduced height in Greenlanders. These findings are important from a public health perspective and highlight the usefulness of complex trait genetic studies in isolated populations.

Antisense oligonucleotides that recruit RNase H and thereby cleave complementary messenger RNAs are being developed as therapeutics. Dose-dependent hepatic changes associated with hepatocyte necrosis and increases in serum alanine-aminotransferase levels have been observed after treatment with certain oligonucleotides. Although general mechanisms for drug-induced hepatic injury are known, the characteristics of oligonucleotides that determine their hepatotoxic potential are not well understood. Here, we present a comprehensive analysis of the hepatotoxic potential of locked nucleic acid-modified oligonucleotides in mice. We developed a random forests classifier, in which oligonucleotides are regarded as being composed of dinucleotide units, which distinguished between 206 oligonucleotides with high and low hepatotoxic potential with 80% accuracy as estimated by out-of-bag validation. In a validation set, 17 out of 23 oligonucleotides were correctly predicted (74% accuracy). In isolation, some dinucleotide units increase, and others decrease, the hepatotoxic potential of the oligonucleotides within which they are found. However, a complex interplay between all parts of an oligonucleotide can influence the hepatotoxic potential. Using the classifier, we demonstrate how an oligonucleotide with otherwise high hepatotoxic potential can be efficiently redesigned to abate hepatotoxic potential. These insights establish analysis of sequence and modification patterns as a powerful tool in the preclinical discovery process for oligonucleotide-based medicines.