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Abstract Recent advances in high-throughput measurement technologies have enabled the analysis of molecular perturbations associated with disease phenotypes at the multi-omic level. Such perturbations can range in scale from fluctuations of individual molecules to entire biological pathways. Data-driven clustering algorithms have long been used to group interactions into interpretable functional modules; however, these modules are typically constrained to a fixed size or statistical cutoff. Furthermore, modules are often analyzed independently of their broader biological context. Consequently, such clustering approaches limit the ability to explore functional module associations with disease phenotypes across multiple scales. Here, we introduce AutoFocus, a data-driven method that hierarchically organizes biomolecules and tests for phenotype enrichment at every level within the hierarchy. As a result, the method allows disease-associated modules to emerge at any scale. We evaluated this approach using two datasets: First, we explored associations of biomolecules from the multi-omic QMDiab dataset (n = 388) with the well-characterized type 2 diabetes phenotype. Secondly, we utilized the ROS/MAP Alzheimer’s disease dataset (n = 500), consisting of high-throughput measurements of brain tissue to explore modules associated with multiple Alzheimer’s Disease-related phenotypes. Our method identifies modules that are multi-omic, span multiple pathways, and vary in size. We provide an interactive tool to explore this hierarchy at different levels and probe enriched modules, empowering users to examine the full hierarchy, delve into biomolecular drivers of disease phenotype within a module, and incorporate functional annotations.

Abstract In-depth multiomic phenotyping provides molecular insights into complex physiological processes and their pathologies. Here, we report on integrating 18 diverse deep molecular phenotyping (omics-) technologies applied to urine, blood, and saliva samples from 391 participants of the multiethnic diabetes Qatar Metabolomics Study of Diabetes (QMDiab). Using 6,304 quantitative molecular traits with 1,221,345 genetic variants, methylation at 470,837 DNA CpG sites, and gene expression of 57,000 transcripts, we determine (1) within-platform partial correlations, (2) between-platform mutual best correlations, and (3) genome-, epigenome-, transcriptome-, and phenome-wide associations. Combined into a molecular network of > 34,000 statistically significant trait-trait links in biofluids, our study portrays “The Molecular Human”. We describe the variances explained by each omics in the phenotypes (age, sex, BMI, and diabetes state), platform complementarity, and the inherent correlation structures of multiomics data. Further, we construct multi-molecular network of diabetes subtypes. Finally, we generated an open-access web interface to “The Molecular Human” ( http://comics.metabolomix.com ), providing interactive data exploration and hypotheses generation possibilities.

Abstract Background Metformin and sodium-glucose-cotransporter-2 inhibitors (SGLT2i) are cornerstone therapies for managing hyperglycemia in diabetes. However, their detailed impacts on metabolic processes, particularly within the citric acid (TCA) cycle and its anaplerotic pathways, remain unclear. This study investigates the tissue-specific metabolic effects of metformin, both as a monotherapy and in combination with SGLT2i, on the TCA cycle and associated anaplerotic reactions in both mice and humans. Methods Metformin-specific metabolic changes were initially identified by comparing metformin-treated diabetic mice (MET) with vehicle-treated db/db mice (VG). These findings were then assessed in two human cohorts (KORA and QBB) and a longitudinal KORA study of metformin-naïve patients with Type 2 Diabetes (T2D). We also compared MET with db/db mice on combination therapy (SGLT2i + MET). Metabolic profiling analyzed 716 metabolites from plasma, liver, and kidney tissues post-treatment, using linear regression and Bonferroni correction for statistical analysis, complemented by pathway analyses to explore the pathophysiological implications. Results Metformin monotherapy significantly upregulated TCA cycle intermediates such as malate, fumarate, and α-ketoglutarate (α-KG) in plasma, and anaplerotic substrates including hepatic glutamate and renal 2-hydroxyglutarate (2-HG) in diabetic mice. Downregulated hepatic taurine was also observed. The addition of SGLT2i, however, reversed these effects, such as downregulating circulating malate and α-KG, and hepatic glutamate and renal 2-HG, but upregulated hepatic taurine. In human T2D patients on metformin therapy, significant systemic alterations in metabolites were observed, including increased malate but decreased citrulline. The bidirectional modulation of TCA cycle intermediates in mice influenced key anaplerotic pathways linked to glutaminolysis, tumorigenesis, immune regulation, and antioxidative responses. Conclusion This study elucidates the specific metabolic consequences of metformin and SGLT2i on the TCA cycle, reflecting potential impacts on the immune system. Metformin shows promise for its anti-inflammatory properties, while the addition of SGLT2i may provide liver protection in conditions like metabolic dysfunction-associated steatotic liver disease (MASLD). These observations underscore the importance of personalized treatment strategies.

Abstract As spaceflight becomes more common with commercial crews, blood-based measures of crew health can guide both astronaut biomedicine and countermeasures. By profiling plasma proteins, metabolites, and extracellular vesicles/particles (EVPs) from the SpaceX Inspiration4 crew, we generated “spaceflight secretome profiles,” which showed significant differences in coagulation, oxidative stress, and brain-enriched proteins. While >93% of differentially abundant proteins (DAPs) in vesicles and metabolites recovered within six months, the majority (73%) of plasma DAPs were still perturbed post-flight. Moreover, these proteomic alterations correlated better with peripheral blood mononuclear cells than whole blood, suggesting that immune cells contribute more DAPs than erythrocytes. Finally, to discern possible mechanisms leading to brain-enriched protein detection and blood-brain barrier (BBB) disruption, we examined protein changes in dissected brains of spaceflight mice, which showed increases in PECAM-1, a marker of BBB integrity. These data highlight how even short-duration spaceflight can disrupt human and murine physiology and identify spaceflight biomarkers that can guide countermeasure development.

Abstract Prostate cancer is the second most common cancer in men and affects 1 in 9 men in the United States. Early screening for prostate cancer often involves monitoring levels of prostate-specific antigen (PSA) and performing digital rectal exams. However, a prostate biopsy is always required for definitive cancer diagnosis. The Early Detection Research Network (EDRN) is a consortium within the National Cancer Institute aimed at improving screening approaches and early detection of cancers. As part of this effort, the Weill Cornell EDRN Prostate Cancer has collected and biobanked specimens from men undergoing a prostate biopsy between 2008 and 2017. In this report, we describe blood metabolomics measurements for a subset of this population. The dataset includes detailed clinical and prospective records for 580 patients who underwent prostate biopsy, 287 of which were subsequentially diagnosed with prostate cancer, combined with profiling of 1,482 metabolites from plasma samples collected at the time of biopsy. We expect this dataset to provide a valuable resource for scientists investigating prostate cancer metabolism.

Abstract Background Acute respiratory distress syndrome (ARDS), a life-threatening condition during critical illness, is a common complication of COVID-19. It can originate from various disease etiologies, including severe infections, major injury, or inhalation of irritants. ARDS poses substantial clinical challenges due to a lack of etiology-specific therapies, multisystem involvement, and heterogeneous, poor patient outcomes. A molecular comparison of ARDS groups holds the potential to reveal common and distinct mechanisms underlying ARDS pathogenesis. Methods We performed a comparative analysis of urine-based metabolomics and proteomics profiles from COVID-19 ARDS patients (n = 42) and bacterial sepsis-induced ARDS patients (n = 17). To this end, we used two different approaches, first we compared the molecular omics profiles between ARDS groups, and second, we correlated clinical manifestations within each group with the omics profiles. Results The comparison of the two ARDS etiologies identified 150 metabolites and 70 proteins that were differentially abundant between the two groups. Based on these findings, we interrogated the interplay of cell adhesion/extracellular matrix molecules, inflammation, and mitochondrial dysfunction in ARDS pathogenesis through a multi-omic network approach. Moreover, we identified a proteomic signature associated with mortality in COVID-19 ARDS patients, which contained several proteins that had previously been implicated in clinical manifestations frequently linked with ARDS pathogenesis. Conclusion In summary, our results provide evidence for significant molecular differences in ARDS patients from different etiologies and a potential synergy of extracellular matrix molecules, inflammation, and mitochondrial dysfunction in ARDS pathogenesis. The proteomic mortality signature should be further investigated in future studies to develop prediction models for COVID-19 patient outcomes.

Variable autoencoders offer an alternative way to interrogate metabolomic data and identify meaningful, non-linear relationships.

Background Acute respiratory distress syndrome (ARDS), a life-threatening condition characterized by hypoxemia and poor lung compliance, is associated with high mortality. ARDS induced by COVID-19 has similar clinical presentations and pathological manifestations as non-COVID-19 ARDS. However, COVID-19 ARDS is associated with a more protracted inflammatory respiratory failure compared to traditional ARDS. Therefore, a comprehensive molecular comparison of ARDS of different etiologies groups may pave the way for more specific clinical interventions. Methods and findings In this study, we compared COVID-19 ARDS (n = 43) and bacterial sepsis-induced (non-COVID-19) ARDS (n = 24) using multi-omic plasma profiles covering 663 metabolites, 1,051 lipids, and 266 proteins. To address both between- and within- ARDS group variabilities we followed two approaches. First, we identified 706 molecules differently abundant between the two ARDS etiologies, revealing more than 40 biological processes differently regulated between the two groups. From these processes, we assembled a cascade of therapeutically relevant pathways downstream of sphingosine metabolism. The analysis suggests a possible overactivation of arginine metabolism involved in long-term sequelae of ARDS and highlights the potential of JAK inhibitors to improve outcomes in bacterial sepsis-induced ARDS. The second part of our study involved the comparison of the two ARDS groups with respect to clinical manifestations. Using a data-driven multi-omic network, we identified signatures of acute kidney injury (AKI) and thrombocytosis within each ARDS group. The AKI-associated network implicated mitochondrial dysregulation which might lead to post-ARDS renal-sequalae. The thrombocytosis-associated network hinted at a synergy between prothrombotic processes, namely IL-17, MAPK, TNF signaling pathways, and cell adhesion molecules. Thus, we speculate that combination therapy targeting two or more of these processes may ameliorate thrombocytosis-mediated hypercoagulation. Conclusion We present a first comprehensive molecular characterization of differences between two ARDS etiologies–COVID-19 and bacterial sepsis. Further investigation into the identified pathways will lead to a better understanding of the pathophysiological processes, potentially enabling novel therapeutic interventions. Author summary Acute respiratory distress syndrome (ARDS) is a critical condition of the lung that can arise after severe infections, traumatic injury, or inhalation of toxins. Patients with ARDS are in a complex disease state and at risk of multiple clinical complications, such as thrombosis, lung fibrosis, acute kidney injury, and increased mortality. Currently, there are substantial challenges in the treatment of ARDS due to the high heterogeneity of this condition across patients. Our study compared metabolomic and proteomic changes induced by two different causes of ARDS—COVID-19 infection and bacterial sepsis. We used blood samples of patients from each ARDS group for molecular profiling and identified several hundred molecules from various biological processes differing between the two groups. Based on these results, we made several new propositions: (1) A role of arginine metabolism in long-term sequelae of ARDS. (2) The potential use of JAK-STAT pathway inhibitors for bacterial sepsis-induced ARDS. (3) ARDS-associated mitochondrial dysfunction as a reason for poor prognosis of acute kidney injury that occurred during ARDS. (4) A synergy between prothrombotic processes as a potential reason for hypercoagulation in ARDS. We hypothesize that combination therapy targeting two or more of these prothrombotic processes may ameliorate hypercoagulation.

Summary: The coronavirus disease-19 (COVID-19) pandemic has ravaged global healthcare with previously unseen levels of morbidity and mortality. In this study, we performed large-scale integrative multi-omics analyses of serum obtained from COVID-19 patients with the goal of uncovering novel pathogenic complexities of this disease and identifying molecular signatures that predict clinical outcomes. We assembled a network of protein-metabolite interactions through targeted metabolomic and proteomic profiling in 330 COVID-19 patients compared to 97 non-COVID, hospitalized controls. Our network identified distinct protein-metabolite cross talk related to immune modulation, energy and nucleotide metabolism, vascular homeostasis, and collagen catabolism. Additionally, our data linked multiple proteins and metabolites to clinical indices associated with long-term mortality and morbidity. Finally, we developed a novel composite outcome measure for COVID-19 disease severity based on metabolomics data. The model predicts severe disease with a concordance index of around 0.69, and shows high predictive power of 0.83–0.93 in two independent datasets.

Correlation network inference is typically based on the significance of the correlation coefficients, but this procedure is not guaranteed to capture biological mechanisms. Here, the authors develop a cutoff selection algorithm that maximizes the overlap between inferred networks and prior knowledge.

Sex and the APOE ε4 genotype are important risk factors for late-onset Alzheimer’s disease. In the current study, the authors investigate how sex and APOE ε4 genotype modify the association between Alzheimer’s disease biomarkers and metabolites in serum.

Deep understanding of the SARS-CoV-2 effects on host molecular pathways is paramount for the discovery of early biomarkers of outcome of coronavirus disease 2019 (COVID-19) and the identification of novel therapeutic targets. In that light, we generated metabolomic data from COVID-19 patient blood using high-throughput targeted nuclear magnetic resonance (NMR) spectroscopy and high-dimensional flow cytometry. We find considerable changes in serum metabolome composition of COVID-19 patients associated with disease severity, and response to tocilizumab treatment. We built a clinically annotated, biologically-interpretable space for precise time-resolved disease monitoring and characterize the temporal dynamics of metabolomic change along the clinical course of COVID-19 patients and in response to therapy. Finally, we leverage joint immuno-metabolic measurements to provide a novel approach for patient stratification and early prediction of severe disease. Our results show that high-dimensional metabolomic and joint immune-metabolic readouts provide rich information content for elucidation of the host’s response to infection and empower discovery of novel metabolic-driven therapies, as well as precise and efficient clinical action.

Multiple studies have investigated the role of blood circulating proteins in COVID-19 disease using the Olink affinity proteomics platform. However, study inclusion criteria and sample collection conditions varied between studies, leading to sometimes incongruent associations. To identify the most robust protein markers of the disease and the underlying pathways that are relevant under all conditions, it is essential to identify proteins that replicate most widely. Here we combined the Olink proteomics profiles of two newly recruited COVID-19 studies (N=68 and N=98) with those of three previously published COVID-19 studies (N=383, N=83, N=57). For these studies, three Olink panels (Inflammation and Cardiovascular II & III) with 253 unique proteins were compared. Case/control analysis revealed thirteen proteins (CCL16, CCL7, CXCL10, CCL8, LGALS9, CXCL11, IL1RN, CCL2, CD274, IL6, IL18, MERTK, IFNγ, and IL18R1) that were differentially expressed in COVID-19 patients in all five studies. Except CCL16, which was higher in controls, all proteins were overexpressed in COVID-19 patients. Pathway analysis revealed concordant trends across all studies with pathways related to cytokine-cytokine interaction, IL18 signaling, fluid shear stress and rheumatoid arthritis. Our results reaffirm previous findings related to a COVID-19 cytokine storm syndrome. Cross-study robustness of COVID-19 specific protein expression profiles support the utility of affinity proteomics as a tool and for the identification of potential therapeutic targets.

High-dimensional omics datasets frequently contain missing data points, which typically occur due to concentrations below the limit of detection (LOD) of the profiling platform. The presence of such missing values significantly limits downstream statistical analysis and result interpretation. Two common techniques to deal with this issue include the removal of samples with missing values and imputation approaches that substitute the missing measurements with reasonable estimates. Both approaches, however, suffer from various shortcomings and pitfalls. In this paper, we present “rox”, a novel statistical model for the analysis of omics data with missing values without the need for imputation. The model directly incorporates missing values as “low” concentrations into the calculation. We show the superiority of rox over common approaches on simulated data and on six metabolomics datasets. Fully leveraging the information contained in LOD-based missing values, rox provides a powerful tool for the statistical analysis of omics data.

Very preterm infants are at high risk for suboptimal nutrition in the first weeks of life leading to insufficient weight gain and complications arising from metabolic imbalances such as insufficient bone mineral accretion. We investigated the use of a novel set of standardized parenteral nutrition (PN; MUC PREPARE) solutions regarding improving nutritional intake, accelerating termination of parenteral feeding, and positively affecting growth in comparison to individually prescribed and compounded PN solutions. We studied the effect of MUC PREPARE on macro- and micronutrient intake, metabolism, and growth in 58 very preterm infants and compared results to a historic reference group of 58 very preterm infants matched for clinical characteristics. Infants receiving MUC PREPARE demonstrated improved macro- and micronutrient intake resulting in balanced electrolyte levels and stable metabolomic profiles. Subsequently, improved energy supply was associated with up to 1.5 weeks earlier termination of parenteral feeding, while simultaneously reaching up to 1.9 times higher weight gain at day 28 in extremely immature infants (

The role of the gut microbiota in hepatic lipid metabolism is controversial and incompletely understood. Here the authors perform multi-omics analyses of altered lipid metabolic processes in germ-free and specific pathogen-free mice, revealing how the gut microbiota affects hepatic fatty acid desaturation and elongation.

Despite the proven clinical antineoplastic activity of histone deacetylase inhibitors (HDACI), their effect has been reported to be lower than expected in B-cell lymphomas. Traditionally considered as “epigenetic drugs”, HDACI modify the acetylation status of an extensive proteome, acting as general lysine deacetylase inhibitors (KDACI), and thus potentially impacting various branches of cellular metabolism. Here, we demonstrate through metabolomic profiling of patient plasma and cell lines that the KDACI panobinostat alters lipid metabolism and downstream survival signaling in diffuse large B-cell lymphomas (DLBCL). Specifically, panobinostat induces metabolic adaptations resulting in newly acquired dependency on the choline pathway and activation of PI3K signaling. This metabolic reprogramming decreased the antineoplastic effect of panobinostat. Conversely, inhibition of these metabolic adaptations resulted in superior anti-lymphoma effect as demonstrated by the combination of panobinostat with a choline pathway inhibitor. In conclusion, our study demonstrates the power of metabolomics in identifying unknown effects of KDACI, and emphasizes the need for a better understanding of these drugs in order to achieve successful clinical implementation.

IgG glycosylation is an important factor in immune function, yet the molecular details of protein glycosylation remain poorly understood. The data-driven approach presented here uses large-scale plasma IgG mass spectrometry measurements to infer new biochemical reactions in the glycosylation pathway.

BACKGROUND:Genome-wide association studies of common diseases or metabolite quantitative traits often identify common variants of small effect size, which may contribute to phenotypes by modulation of gene expression. Thus, there is growing demand for cellular models enabling to assess the impact of gene regulatory variants with moderate effects on gene expression. Mitochondrial fatty acid oxidation is an important energy metabolism pathway. Common noncoding acyl-CoA dehydrogenase short chain (ACADS) gene variants are associated with plasma C4-acylcarnitine levels and allele-specific modulation of ACADS expression may contribute to the observed phenotype. METHODS AND FINDINGS:We assessed ACADS expression and intracellular acylcarnitine levels in human lymphoblastoid cell lines (LCL) genotyped for a common ACADS variant associated with plasma C4-acylcarnitine and found a significant genotype-dependent decrease of ACADS mRNA and protein. Next, we modelled gradual decrease of ACADS expression using a tetracycline-regulated shRNA-knockdown of ACADS in Huh7 hepatocytes, a cell line with high fatty acid oxidation-(FAO)-capacity. Assessing acylcarnitine flux in both models, we found increased C4-acylcarnitine levels with decreased ACADS expression levels. Moreover, assessing time-dependent changes of acylcarnitine levels in shRNA-hepatocytes with altered ACADS expression levels revealed an unexpected effect on long- and medium-chain fatty acid intermediates. CONCLUSIONS:Both, genotyped LCL and regulated shRNA-knockdown are valuable tools to model moderate, gradual gene-regulatory effects of common variants on cellular phenotypes. Decreasing ACADS expression levels modulate short and surprisingly also long/medium chain acylcarnitines, and may contribute to increased plasma acylcarnitine levels.

Investigations into the mechanisms regulating obesity are frantic and novel translational approaches are needed. The raccoon dog (Nyctereutes procyonoides) is a canid species representing a promising model to study metabolic regulation in a species undergoing cycles of seasonal obesity and fasting. To understand the molecular mechanisms of metabolic regulation in seasonal adaptation, we analyzed key central nervous system and peripheral signals regulating food intake and metabolism from raccoon dogs after autumnal fattening and winter fasting. Expressions of neuropeptide Y (NPY), orexin-2 receptor (OX2R), pro-opiomelanocortin (POMC) and leptin receptor (ObRb) were analyzed as examples of orexigenic and anorexigenic signals using qRT-PCR from raccoon dog hypothalamus samples. Plasma metabolic profiles were measured with 1H NMR-spectroscopy and LC-MS. Circulating hormones and cytokines were determined with canine specific antibody assays. Surprisingly, NPY and POMC were not affected by the winter fasting nor autumn fattening and the metabolic profiles showed a remarkable equilibrium, indicating conserved homeostasis. However, OX2R and ObRb expression changes suggested seasonal regulation. Circulating cytokine levels were not increased, demonstrating that the autumn fattening did not induce subacute inflammation. Thus, the raccoon dog developed seasonal regulatory mechanisms to accommodate the autumnal fattening and prolonged fasting making the species unique in coping with the extreme environmental challenges.

Metabolomics studies have seen a steady growth due to the development and implementation of affordable and high-quality metabolomics platforms. In large metabolite panels, measurement values are frequently missing and, if neglected or sub-optimally imputed, can cause biased study results. We provided a publicly available, user-friendly R script to streamline the imputation of missing endogenous, unannotated, and xenobiotic metabolites. We evaluated the multivariate imputation by chained equations (MICE) and k-nearest neighbors (kNN) analyses implemented in our script by simulations using measured metabolites data from the Netherlands Epidemiology of Obesity (NEO) study (n = 599). We simulated missing values in four unique metabolites from different pathways with different correlation structures in three sample sizes (599, 150, 50) with three missing percentages (15%, 30%, 60%), and using two missing mechanisms (completely at random and not at random). Based on the simulations, we found that for MICE, larger sample size was the primary factor decreasing bias and error. For kNN, the primary factor reducing bias and error was the metabolite correlation with its predictor metabolites. MICE provided consistently higher performance measures particularly for larger datasets (n > 50). In conclusion, we presented an imputation workflow in a publicly available R script to impute untargeted metabolomics data. Our simulations provided insight into the effects of sample size, percentage missing, and correlation structure on the accuracy of the two imputation methods.

Glycomics measurements, like all other high-throughput technologies, are subject to technical variation due to fluctuations in the experimental conditions. The removal of this non-biological signal from the data is referred to as normalization. Contrary to other omics data types, a systematic evaluation of normalization options for glycomics data has not been published so far. In this paper, we assess the quality of different normalization strategies for glycomics data with an innovative approach. It has been shown previously that Gaussian Graphical Models (GGMs) inferred from glycomics data are able to identify enzymatic steps in the glycan synthesis pathways in a data-driven fashion. Based on this finding, here, we quantify the quality of a given normalization method according to how well a GGM inferred from the respective normalized data reconstructs known synthesis reactions in the glycosylation pathway. The method therefore exploits a biological measure of goodness. We analyzed 23 different normalization combinations applied to six large-scale glycomics cohorts across three experimental platforms: Liquid Chromatography-ElectroSpray Ionization-Mass Spectrometry (LC-ESI-MS), Ultra High Performance Liquid Chromatography with Fluorescence Detection (UHPLC-FLD), and Matrix Assisted Laser Desorption Ionization-Furier Transform Ion Cyclotron Resonance-Mass Spectrometry (MALDI-FTICR-MS). Based on our results, we recommend normalizing glycan data using the ‘Probabilistic Quotient’ method followed by log-transformation, irrespective of the measurement platform. This recommendation is further supported by an additional analysis, where we ranked normalization methods based on their statistical associations with age, a factor known to associate with glycomics measurements.

BackgroundAround 0.3% of newborns will develop autoimmunity to pancreatic beta cells in childhood and subsequently develop type 1 diabetes before adulthood. Primary prevention of type 1 diabetes will require early intervention in genetically at-risk infants. The objective of this study was to determine to what extent genetic scores (two previous genetic scores and a merged genetic score) can improve the prediction of type 1 diabetes.Methods and findingsThe Environmental Determinants of Diabetes in the Young (TEDDY) study followed genetically at-risk children at 3- to 6-monthly intervals from birth for the development of islet autoantibodies and type 1 diabetes. Infants were enrolled between 1 September 2004 and 28 February 2010 and monitored until 31 May 2016. The risk (positive predictive value) for developing multiple islet autoantibodies (pre-symptomatic type 1 diabetes) and type 1 diabetes was determined in 4,543 children who had no first-degree relatives with type 1 diabetes and either a heterozygous HLA DR3 and DR4-DQ8 risk genotype or a homozygous DR4-DQ8 genotype, and in 3,498 of these children in whom genetic scores were calculated from 41 single nucleotide polymorphisms. In the children with the HLA risk genotypes, risk for developing multiple islet autoantibodies was 5.8% (95% CI 5.0%-6.6%) by age 6 years, and risk for diabetes by age 10 years was 3.7% (95% CI 3.0%-4.4%). Risk for developing multiple islet autoantibodies was 11.0% (95% CI 8.7%-13.3%) in children with a merged genetic score of >14.4 (upper quartile; n = 907) compared to 4.1% (95% CI 3.3%-4.9%, P < 0.001) in children with a genetic score of ≤14.4 (n = 2,591). Risk for developing diabetes by age 10 years was 7.6% (95% CI 5.3%-9.9%) in children with a merged score of >14.4 compared with 2.7% (95% CI 1.9%-3.6%) in children with a score of ≤14.4 (P < 0.001). Of 173 children with multiple islet autoantibodies by age 6 years and 107 children with diabetes by age 10 years, 82 (sensitivity, 47.4%; 95% CI 40.1%-54.8%) and 52 (sensitivity, 48.6%, 95% CI 39.3%-60.0%), respectively, had a score >14.4. Scores were higher in European versus US children (P = 0.003). In children with a merged score of >14.4, risk for multiple islet autoantibodies was similar and consistently >10% in Europe and in the US; risk was greater in males than in females (P = 0.01). Limitations of the study include that the genetic scores were originally developed from case-control studies of clinical diabetes in individuals of mainly European decent. It is, therefore, possible that it may not be suitable to all populations.ConclusionsA type 1 diabetes genetic score identified infants without family history of type 1 diabetes who had a greater than 10% risk for pre-symptomatic type 1 diabetes, and a nearly 2-fold higher risk than children identified by high-risk HLA genotypes alone. This finding extends the possibilities for enrolling children into type 1 diabetes primary prevention trials.

Immunoglobulin G (IgG), a glycoprotein secreted by plasma B-cells, plays a major role in the human adaptive immune response and are associated with a wide range of diseases. Glycosylation of the Fc binding region of IgGs, responsible for the antibody’s effector function, is essential for prompting a proper immune response. This study focuses on the general genetic impact on IgG glycosylation as well as corresponding subclass specificities. To identify genetic loci involved in IgG glycosylation, we performed a genome-wide association study (GWAS) on liquid chromatography electrospray mass spectrometry (LC–ESI-MS)—measured IgG glycopeptides of 1,823 individuals in the Cooperative Health Research in the Augsburg Region (KORA F4) study cohort. In addition, we performed GWAS on subclass-specific ratios of IgG glycans to gain power in identifying genetic factors underlying single enzymatic steps in the glycosylation pathways. We replicated our findings in 1,836 individuals from the Leiden Longevity Study (LLS). We were able to show subclass-specific genetic influences on single IgG glycan structures. The replicated results indicate that, in addition to genes encoding for glycosyltransferases (i.e., ST6GAL1, B4GALT1, FUT8, and MGAT3), other genetic loci have strong influences on the IgG glycosylation patterns. A novel locus on chromosome 1, harboring RUNX3, which encodes for a transcription factor of the runt domain-containing family, is associated with decreased galactosylation. Interestingly, members of the RUNX family are cross-regulated, and RUNX3 is involved in both IgA class switching and B-cell maturation as well as T-cell differentiation and apoptosis. Besides the involvement of glycosyltransferases in IgG glycosylation, we suggest that, due to the impact of variants within RUNX3, potentially mechanisms involved in B-cell activation and T-cell differentiation during the immune response as well as cell migration and invasion involve IgG glycosylation.

The application of metabolomics technology to epidemiological studies is emerging as a new approach to elucidate disease etiology and for biomarker discovery. However, analysis of metabolomics data is complex and there is an urgent need for the standardization of analysis workflow and reporting of study findings. To inform the development of such guidelines, we conducted a survey of 47 cohort representatives from the Consortium of Metabolomics Studies (COMETS) to gain insights into the current strategies and procedures used for analyzing metabolomics data in epidemiological studies worldwide. The results indicated a variety of applied analytical strategies, from biospecimen and data pre-processing and quality control to statistical analysis and reporting of study findings. These strategies included methods commonly used within the metabolomics community and applied in epidemiological research, as well as novel approaches to pre-processing pipelines and data analysis. To help with these discrepancies, we propose use of open-source initiatives such as the online web-based tool COMETS Analytics, which includes helpful tools to guide analytical workflow and the standardized reporting of findings from metabolomics analyses within epidemiological studies. Ultimately, this will improve the quality of statistical analyses, research findings, and study reproducibility.

It is not controversial that study design considerations and challenges must be addressed when investigating the linkage between single omic measurements and human phenotypes. It follows that such considerations are just as critical, if not more so, in the context of multi-omic studies. In this review, we discuss (1) epidemiologic principles of study design, including selection of biospecimen source(s) and the implications of the timing of sample collection, in the context of a multi-omic investigation, and (2) the strengths and limitations of various techniques of data integration across multi-omic data types that may arise in population-based studies utilizing metabolomic data.

Kit-based assays, such as AbsoluteIDQTM p150, are widely used in large cohort studies and provide a standardized method to quantify blood concentrations of phosphatidylcholines (PCs). Many disease-relevant associations of PCs were reported using this method. However, their interpretation is hampered by lack of functionally-relevant information on the detailed fatty acid side-chain compositions as only the total number of carbon atoms and double bonds is identified by the kit. To enable more substantiated interpretations, we characterized these PC sums using the side-chain resolving LipidyzerTM platform, analyzing 223 samples in parallel to the AbsoluteIDQTM. Combining these datasets, we estimated the quantitative composition of PC sums and subsequently tested their replication in an independent cohort. We identified major constituents of 28 PC sums, revealing also various unexpected compositions. As an example, PC 16:0_22:5 accounted for more than 50% of the PC sum with in total 38 carbon atoms and 5 double bonds (PC aa 38:5). For 13 PC sums, we found relatively high abundances of odd-chain fatty acids. In conclusion, our study provides insights in PC compositions in human plasma, facilitating interpretation of existing epidemiological data sets and potentially enabling imputation of PC compositions for future meta-analyses of lipidomics data.

Serum metabolite profiling can be used to identify pathways involved in the pathogenesis of and potential biomarkers for a given disease. Both restless legs syndrome (RLS) and Parkinson`s disease (PD) represent movement disorders for which currently no blood-based biomarkers are available and whose pathogenesis has not been uncovered conclusively. We performed unbiased serum metabolite profiling in search of signature metabolic changes for both diseases.456 metabolites were quantified in serum samples of 1272 general population controls belonging to the KORA cohort, 82 PD cases and 95 RLS cases by liquid-phase chromatography and gas chromatography separation coupled with tandem mass spectrometry. Genetically determined metabotypes were calculated using genome-wide genotyping data for the 1272 general population controls.After stringent quality control, we identified decreased levels of long-chain (polyunsaturated) fatty acids of individuals with PD compared to both RLS (PD vs. RLS: p = 0.0001 to 5.80x10-9) and general population controls (PD vs. KORA: p = 6.09x10-5 to 3.45x10-32). In RLS, inositol metabolites were increased specifically (RLS vs. KORA: p = 1.35x10-6 to 3.96x10-7). The impact of dopaminergic drugs was reflected in changes in the phenylalanine/tyrosine/dopamine metabolism observed in both individuals with RLS and PD.A first discovery approach using serum metabolite profiling in two dopamine-related movement disorders compared to a large general population sample identified significant alterations in the polyunsaturated fatty acid metabolism in PD and implicated the inositol metabolism in RLS. These results provide a starting point for further studies investigating new perspectives on factors involved in the pathogenesis of the two diseases as well as possible points of therapeutic intervention.

Correction to: Nature Communications (2017) 8:1231. doi:10.1038/s41467-017-01525-0

Biological systems consist of multiple organizational levels all densely interacting with each other to ensure function and flexibility of the system. Simultaneous analysis of cross-sectional multi-omics data from large population studies is a powerful tool to comprehensively characterize the underlying molecular mechanisms on a physiological scale. In this study, we systematically analyzed the relationship between fasting serum metabolomics and whole blood transcriptomics data from 712 individuals of the German KORA F4 cohort. Correlation-based analysis identified 1,109 significant associations between 522 transcripts and 114 metabolites summarized in an integrated network, the 'human blood metabolome-transcriptome interface' (BMTI). Bidirectional causality analysis using Mendelian randomization did not yield any statistically significant causal associations between transcripts and metabolites. A knowledge-based interpretation and integration with a genome-scale human metabolic reconstruction revealed systematic signatures of signaling, transport and metabolic processes, i.e. metabolic reactions mainly belonging to lipid, energy and amino acid metabolism. Moreover, the construction of a network based on functional categories illustrated the cross-talk between the biological layers at a pathway level. Using a transcription factor binding site enrichment analysis, this pathway cross-talk was further confirmed at a regulatory level. Finally, we demonstrated how the constructed networks can be used to gain novel insights into molecular mechanisms associated to intermediate clinical traits. Overall, our results demonstrate the utility of a multi-omics integrative approach to understand the molecular mechanisms underlying both normal physiology and disease.

Metabolomics is a relatively new high-throughput technology that aims at measuring all endogenous metabolites within a biological sample in an unbiased fashion. The resulting metabolic profiles may be regarded as functional signatures of the physiological state, and have been shown to comprise effects of genetic regulation as well as environmental factors. This potential to connect genotypic to phenotypic information promises new insights and biomarkers for different research fields, including biomedical and pharmaceutical research. In the statistical analysis of metabolomics data, many techniques from other omics fields can be reused. However recently, a number of tools specific for metabolomics data have been developed as well. The focus of this mini review will be on recent advancements in the analysis of metabolomics data especially by utilizing Gaussian graphical models and independent component analysis.

Metabolomics is a relatively new high-throughput technology that aims at measuring all endogenous metabolites within a biological sample in an unbiased fashion. The resulting metabolic profiles may be regarded as functional signatures of the physiological state, and have been shown to comprise effects of genetic regulation as well as environmental factors. This potential to connect genotypic to phenotypic information promises new insights and biomarkers for different research fields, including biomedical and pharmaceutical research. In the statistical analysis of metabolomics data, many techniques from other omics fields can be reused. However recently, a number of tools specific for metabolomics data have been developed as well. The focus of this mini review will be on recent advancements in the analysis of metabolomics data especially by utilizing Gaussian graphical models and independent component analysis.

ObjectiveTo characterise the influence of the fat free mass on the metabolite profile in serum samples from participants of the population-based KORA (Cooperative Health Research in the Region of Augsburg) S4 study.Subjects and methodsAnalyses were based on metabolite profile from 965 participants of the S4 and 890 weight-stable subjects of its seven-year follow-up study (KORA F4). 190 different serum metabolites were quantified in a targeted approach including amino acids, acylcarnitines, phosphatidylcholines (PCs), sphingomyelins and hexose. Associations between metabolite concentrations and the fat free mass index (FFMI) were analysed using adjusted linear regression models. To draw conclusions on enzymatic reactions, intra-metabolite class ratios were explored. Pairwise relationships among metabolites were investigated and illustrated by means of Gaussian graphical models (GGMs).ResultsWe found 339 significant associations between FFMI and various metabolites in KORA S4. Among the most prominent associations (p-values 4.75 × 10(-16)-8.95 × 10(-06)) with higher FFMI were increasing concentrations of the branched chained amino acids (BCAAs), ratios of BCAAs to glucogenic amino acids, and carnitine concentrations. For various PCs, a decrease in chain length or in saturation of the fatty acid moieties could be observed with increasing FFMI, as well as an overall shift from acyl-alkyl PCs to diacyl PCs. These findings were reproduced in KORA F4. The established GGMs supported the regression results and provided a comprehensive picture of the relationships between metabolites. In a sub-analysis, most of the discovered associations did not exist in obese subjects in contrast to non-obese subjects, possibly indicating derangements in skeletal muscle metabolism.ConclusionA set of serum metabolites strongly associated with FFMI was identified and a network explaining the relationships among metabolites was established. These results offer a novel and more complete picture of the FFMI effects on serum metabolites in a data-driven network.

Recent genome-wide association studies (GWAS) with metabolomics data linked genetic variation in the human genome to differences in individual metabolite levels. A strong relevance of this metabolic individuality for biomedical and pharmaceutical research has been reported. However, a considerable amount of the molecules currently quantified by modern metabolomics techniques are chemically unidentified. The identification of these "unknown metabolites" is still a demanding and intricate task, limiting their usability as functional markers of metabolic processes. As a consequence, previous GWAS largely ignored unknown metabolites as metabolic traits for the analysis. Here we present a systems-level approach that combines genome-wide association analysis and Gaussian graphical modeling with metabolomics to predict the identity of the unknown metabolites. We apply our method to original data of 517 metabolic traits, of which 225 are unknowns, and genotyping information on 655,658 genetic variants, measured in 1,768 human blood samples. We report previously undescribed genotype-metabotype associations for six distinct gene loci (SLC22A2, COMT, CYP3A5, CYP2C18, GBA3, UGT3A1) and one locus not related to any known gene (rs12413935). Overlaying the inferred genetic associations, metabolic networks, and knowledge-based pathway information, we derive testable hypotheses on the biochemical identities of 106 unknown metabolites. As a proof of principle, we experimentally confirm nine concrete predictions. We demonstrate the benefit of our method for the functional interpretation of previous metabolomics biomarker studies on liver detoxification, hypertension, and insulin resistance. Our approach is generic in nature and can be directly transferred to metabolomics data from different experimental platforms.

Metabolomic profiling and the integration of whole-genome genetic association data has proven to be a powerful tool to comprehensively explore gene regulatory networks and to investigate the effects of genetic variation at the molecular level. Serum metabolite concentrations allow a direct readout of biological processes, and association of specific metabolomic signatures with complex diseases such as Alzheimer's disease and cardiovascular and metabolic disorders has been shown. There are well-known correlations between sex and the incidence, prevalence, age of onset, symptoms, and severity of a disease, as well as the reaction to drugs. However, most of the studies published so far did not consider the role of sexual dimorphism and did not analyse their data stratified by gender. This study investigated sex-specific differences of serum metabolite concentrations and their underlying genetic determination. For discovery and replication we used more than 3,300 independent individuals from KORA F3 and F4 with metabolite measurements of 131 metabolites, including amino acids, phosphatidylcholines, sphingomyelins, acylcarnitines, and C6-sugars. A linear regression approach revealed significant concentration differences between males and females for 102 out of 131 metabolites (p-values

Hematopoiesis is an ideal model system for stem cell biology with advanced experimental access. A systems view on the interactions of core transcription factors is important for understanding differentiation mechanisms and dynamics. In this manuscript, we construct a Boolean network to model myeloid differentiation, specifically from common myeloid progenitors to megakaryocytes, erythrocytes, granulocytes and monocytes. By interpreting the hematopoietic literature and translating experimental evidence into Boolean rules, we implement binary dynamics on the resulting 11-factor regulatory network. Our network contains interesting functional modules and a concatenation of mutual antagonistic pairs. The state space of our model is a hierarchical, acyclic graph, typifying the principles of myeloid differentiation. We observe excellent agreement between the steady states of our model and microarray expression profiles of two different studies. Moreover, perturbations of the network topology correctly reproduce reported knockout phenotypes in silico. We predict previously uncharacterized regulatory interactions and alterations of the differentiation process, and line out reprogramming strategies.