Leptin, acylcarnitine metabolites and development of adiposity in the Rhea mother-child cohort in Crete, Greece.

OBJECTIVE: This study aims to investigate relations of serum leptin at age 4 with development of adiposity and linear growth during 3 years of follow-up among 75 Greek children and to identify serum metabolites associated with leptin at age 4 and to characterize their associations with adiposity gain and linear growth. METHODS: Linear regression models that accounted for maternal age, education and gestational weight gain and child's age and sex were used to examine associations of leptin and leptin-associated metabolites measured at age 4 with indicators of adiposity and linear growth at age 7. RESULTS: Each 1-unit increment in natural log-(ln)-transformed leptin corresponded with 0.33 (95% CI: 0.10, 0.55) units greater body mass index-for-age z-score gain during follow-up. Likewise, higher levels of the leptin-associated metabolites methylmalonyl-carnitine and glutaconyl-carnitine corresponded with 0.14 (95% CI: 0.01, 0.27) and 0.07 (95% CI: -0.01, 0.16) units higher body mass index-for-age z-score gain, respectively. These relationships did not differ by sex or baseline weight status and were independent of linear growth. CONCLUSIONS: These findings suggest that leptin, methylmalonyl-carnitine and possibly glutaconyl-carnitine are associated with weight gain during early childhood. Future studies are warranted to confirm these findings in other populations.

Systematic integration of molecular profiles identifies miR-22 as a regulator of lipid and folate metabolism in breast cancer cells.

Dysregulated microRNA (miRNA) mediate malignant phenotypes, including metabolic reprogramming. By performing an integrative analysis of miRNA and metabolome data for the NCI-60 cell line panel, we identified an miRNA cluster strongly associated with both c-Myc expression and global metabolic variation. Within this cluster the cancer-associated and cardioprotective miR-22 was shown to repress fatty acid synthesis and elongation in tumour cells by targeting ATP citrate lyase and fatty acid elongase 6, as well as impairing mitochondrial one-carbon metabolism by suppression of methylene tetrahydrofolate dehydrogenase/cyclohydrolase. Across several data sets, expression of these target genes were associated with poorer outcomes in breast cancer patients. Importantly, a beneficial effect of miR-22 on clinical outcomes in breast cancer was shown to depend on the expression levels of the identified target genes, demonstrating the relevance of miRNA/mRNA interactions to disease progression in vivo. Our systematic analysis establishes miR-22 as a novel regulator of tumour cell metabolism, a function that could contribute to the role of this miRNA in cellular differentiation and cancer development. Moreover, we provide a paradigmatic example of effect modification in outcome analysis as a consequence of miRNA-directed gene targeting, a phenomenon that could be exploited to improve patient prognosis and treatment.

Inadequate fine-tuning of protein synthesis and failure of amino acid homeostasis following inhibition of the ATPase VCP/p97.

The cellular mechanisms that control protein degradation may constitute a non-oncogenic cancer cell vulnerability and, therefore, a therapeutic target. Although this proposition is supported by the clinical success of proteasome inhibitors in some malignancies, most cancers are resistant to proteasome inhibition. The ATPase valosin-containing protein (VCP; p97) is an essential regulator of protein degradation in multiple pathways and has emerged as a target for cancer therapy. We found that pharmacological depletion of VCP enzymatic activity with mechanistically different inhibitors robustly induced proteotoxic stress in solid cancer and multiple myeloma cells, including cells that were insensitive, adapted, or clinically resistant to proteasome inhibition. VCP inhibition had an impact on two key regulators of protein synthesis, eukaryotic initiation factor 2α (eIF2α) and mechanistic target of rapamycin complex 1 (mTORC1), and attenuated global protein synthesis. However, a block on protein translation that was itself cytotoxic alleviated stress signaling and reduced cell death triggered by VCP inhibition. Some of the proteotoxic effects of VCP depletion depended on the eIF2α phosphatase, protein phosphatase 1 regulatory subunit 15A (PPP1R15A)/PP1c, but not on mTORC1, although there appeared to be cross-talk between them. Thus, cancer cell death following VCP inhibition was linked to inadequate fine-tuning of protein synthesis and activity of PPP1R15A/PP1c. VCP inhibitors also perturbed intracellular amino acid levels, activated eukaryotic translation initiation factor 2α kinase 4 (EIF2AK4), and enhanced cellular dependence on amino acid supplies, consistent with a failure of amino acid homeostasis. Many of the observed effects of VCP inhibition differed from the effects triggered by proteasome inhibition or by protein misfolding. Thus, depletion of VCP enzymatic activity triggers cancer cell death in part through inadequate regulation of protein synthesis and amino acid metabolism. The data provide novel insights into the maintenance of intracellular proteostasis by VCP and may have implications for the development of anti-cancer therapies.

Integrated histopathological and urinary metabonomic investigation of the pathogenesis of microcystin-LR toxicosis.

Patterns of change of endogenous metabolites may closely reflect systemic and organ-specific toxic changes. The authors examined the metabolic effects of the cyanobacterial (blue-green algal) toxin microcystin-LR by (1)H-nuclear magnetic resonance (NMR) analysis of urinary endogenous metabolites. Rats were treated with a single sublethal dose, either 20 or 80 µg/kg intraperitoneally, and sacrificed at 2 or 7 days post dosing. Changes in the high-dose, 2-day sacrifice group included centrilobular hepatic necrosis and congestion, accompanied in some animals by regeneration and neovascularization. By 7 days, animals had recovered, the necrotizing process had ended, and the centrilobular areas had been replaced by regenerative, usually hypertrophic hepatocytes. There was considerable interanimal variation in the histologic process and severity, which correlated with the changes in patterns of endogenous metabolites in the urine, thus providing additional validation of the biomarker and biochemical changes. Similarity of the shape of the metabolic trajectories suggests that the mechanisms of toxic effects and recovery are similar among the individual animals, albeit that the magnitude and timing are different for the individual animals. Initial decreases in urinary citrate, 2-oxoglutarate, succinate, and hippurate concentrations were accompanied by a temporary increase in betaine and taurine, then creatine from 24 to 48 hours. Further changes were an increase in guanidinoacetate, dimethylglycine, urocanic acid, and bile acids. As a tool, urine can be repeatedly and noninvasively sampled and metabonomics utilized to study the onset and recovery after toxicity, thus identifying time points of maximal effect. This can help to employ histopathological examination in a guided and effective fashion.

A metabolic phenotyping approach to understanding relationships between metabolic syndrome and breast tumour responses to chemotherapy.

PURPOSE: Breast cancer is associated with adverse outcomes in patients with the metabolic syndrome phenotype. To study this further, we examined the relationship between serum metabolite levels and the components of metabolic syndrome with treatment outcomes in breast cancer. METHODS: A total of 88 women with measurable breast cancer were studied; their serum metabolites as assessed by (1)H nuclear magnetic resonance spectroscopy, blood pressure, lipids, glucose, body mass index and waist circumference were recorded and correlated with treatment response. RESULTS: We identified metabolic syndrome in approximately half of our cohort (42 patients) and observed a significant trend (P = 0.03) of increased incidence of metabolic syndrome in partial response (33.3%), stable disease (42.9%) and progressive disease groups (66.1%). High blood sugar predicted a poor response (P < 0.001). Logistic regression of metabonomic data demonstrated that high lactate (P = 0.03) and low alanine (P = 0.01) combined with high glucose (P = 0.01) were associated with disease progression. CONCLUSIONS: Metabolic syndrome is commonly observed in metastatic breast cancer and these patients have poorer outcomes. These data, which support our previous findings, suggest that high blood glucose as part of metabolic syndrome is associated with a poor response in breast cancer. They also validate new therapeutic approaches that focus on metabolism.

Biomarker discovery for drug development and translational medicine using metabonomics.

There exists at present an urgent desire for better biomarkers, especially in the context of pharmaceutical drug development and in the detection and management of disease. Many researchers in the area of biomarker discovery and development have turned to the "-omics" sciences as a way of addressing these needs. Metabolic profiling, or metabonomics, defines the metabolic phenotype and offers a source of novel biomarkers that have better potential to translate effectively. This review will discuss the broad philosophy and motivations behind metabonomics, and illustrate the case with applications relevant to pharmaceutical development and patient management. Particular focus will be paid to the potential of metabonomics to contribute to biomarker discovery in toxicology and cancer research.

Curve-fitting method for direct quantitation of compounds in complex biological mixtures using 1H NMR: application in metabonomic toxicology studies.

A new software tool has been developed that provides automated measurement of signal intensities in NMR spectra of complex mixtures without using data reduction procedures. The algorithm finds best-fit transformations between signals in reference compound spectra and the corresponding signals in analyte spectra. Unlike other algorithms, it is insensitive to variation in chemical shift and can even be used for relative quantitation of compounds whose identities have not yet been established. Additionally, the parameters of the transformation provide information and error metrics that may assist in the streamlining of quality control. The approach presented is general in scope but has been tested by application to peak quantitation in NMR spectra of biofluids. Replicate NMR measurements of solutions of biologically important compounds at various concentrations were made. Further NMR data were collected on urine samples from human, rat, and mouse, which were "spiked" with reference compound solutions at known concentrations. Finally, existing data from an independent toxicology project involving several hundred samples were analyzed, and the consistency of the measurements for metabolites that give multiple NMR signals was assessed. The results of all these tests give confidence that the technique can be used in automated quantitation of compounds in large NMR data sets with minimal operator intervention.