Metabolomics perspectives of the ecotoxicological risks of polycyclic aromatic hydrocarbons: A scoping review.

Polycyclic Aromatic Hydrocarbons (PAHs) represent persistent environmental pollutants ubiquitously distributed in the environment. Their presence alongside various other contaminants gives rise to intricate interactions, culminating in profound deleterious consequences. The combination effects of different PAH mixtures on biota remains a relatively unexplored domain. Recent studies have harnessed the exceptional sensitivity of metabolomic techniques to unveil the significant ecotoxicological perils of PAH pollution confronting both human populations and ecosystems. This article furnishes a comprehensive overview of current literature focused on the metabolic repercussions stemming from exposure to complex mixtures of PAHs or PAH-pollution sources using metabolomics approaches. These insights are obtained through a wide range of models, including in vitro assessments, animal studies, investigations on human subjects, botanical specimens, and soil environments. The findings underscore that PAH mixtures induce cellular stress responses and systemic effects, leading to metabolic dysregulations in amino acids, carbohydrates, lipids, and other key metabolites (e.g., organic acids, purines), with specific variations observed based on the organism and PAH compounds involved. Additionally, the ecological consequences of PAH pollutants on plant and soil microbial responses are emphasized, revealing significant changes in stress-related metabolites and nutrient cycling in soil ecosystems. The complex interplay of various PAHs and their metabolic effects on several models, as elucidated through metabolomics, highlight the urgency of further research and the need for comprehensive strategies to mitigate the risks posed by these widespread environmental pollutants.

Identification of key neuronal mechanisms triggered by dimethyl fumarate in SH-SY5Y human neuroblastoma cells through a metabolomic approach.

Dimethyl fumarate (DMF) is an old drug used for psoriasis treatment that has recently been repurposed to treat relapse-remitting multiple sclerosis, mostly due to its neuro- and immunomodulatory actions. However, mining of a pharmacovigilance database recently ranked DMF as the second pharmaceutical most associated with cognitive adverse events. To our best knowledge, the signaling mechanisms underlying its therapeutic and neurotoxic outcomes remain mostly undisclosed. This work thus represents the first-hand assessment of DMF-induced metabolic changes in undifferentiated SH-SY5Y human neuroblastoma cells, through an untargeted metabolomic approach using gas chromatography-mass spectrometry (GC-MS). The endometabolome was analyzed following 24 h and 96 h of exposure to two pharmacologically relevant DMF concentrations (0.1 and 10 µM). None of these conditions significantly reduced metabolic activity (MTT reduction assay). Our data showed that 24 h-exposure to DMF at both concentrations tested mainly affected metabolic pathways involved in mitochondrial activity (e.g., citric acid cycle, de novo triacylglycerol biosynthesis), and the synthesis of catecholamines and serotonin by changing the levels of their respective precursors, namely phenylalanine (0.68-fold decrease for 10 µM DMF vs vehicle), and tryptophan (1.36-fold increase for 0.1 µM DMF vs vehicle). Interestingly, taurine, whose levels can be modulated via Nrf2 signaling (DMF's primary target), emerged as a key mediator of DMF's neuronal action, displaying a 3.86-fold increase and 0.27-fold decrease for 10 µM DMF at 24 h and 96 h, respectively. A 96 h-exposure to DMF seemed to mainly trigger pathways associated with glucose production (e.g., gluconeogenesis, glucose-alanine cycle, malate-aspartate shuttle), possibly related to the metabolism of DMF into monomethyl fumarate and its further conversion into glucose via activation of the citric acid cycle. Overall, our data contribute to improving the understanding of the events associated with neuronal exposure to DMF.

Metabolomics Reveals Tyrosine Kinase Inhibitor Resistance-Associated Metabolic Events in Human Metastatic Renal Cancer Cells.

The development of resistance to tyrosine kinase inhibitors (TKIs) is a major cause of treatment failure in metastatic renal cell carcinoma (mRCC). A deeper understanding of the metabolic mechanisms associated with TKI resistance is critical for refining therapeutic strategies. In this study, we established resistance to sunitinib and pazopanib by exposing a parental Caki-1 cell line to increasing concentrations of sunitinib and pazopanib. The intracellular and extracellular metabolome of sunitinib- and pazopanib-resistant mRCC cells were investigated using a nuclear magnetic resonance (NMR)-based metabolomics approach. Data analysis included multivariate and univariate methods, as well as pathway and network analyses. Distinct metabolic signatures in sunitinib- and pazopanib-resistant RCC cells were found for the first time in this study. A common metabolic reprogramming pattern was observed in amino acid, glycerophospholipid, and nicotinate and nicotinamide metabolism. Sunitinib-resistant cells exhibited marked alterations in metabolites involved in antioxidant defence mechanisms, while pazopanib-resistant cells showed alterations in metabolites associated with energy pathways. Sunitinib-resistant RCC cells demonstrated an increased ability to proliferate, whereas pazopanib-resistant cells appeared to restructure their energy metabolism and undergo alterations in pathways associated with cell death. These findings provide potential targets for novel therapeutic strategies to overcome TKI resistance in mRCC through metabolic regulation.

Metabolomic Signatures of Treatment Response in Bladder Cancer.

Bladder cancer (BC) stands as one of the most prevalent urological malignancies, with over 500 thousand newly diagnosed cases annually. Treatment decisions in BC depend on factors like the risk of recurrence, the type of tumor, and the stage of the disease. While standard therapeutic approaches encompass transurethral resection of the bladder tumor, radical cystectomy, and chemo- or immunotherapy, these methods exhibit limited efficacy in mitigating the aggressive and recurrent nature of bladder tumors. To overcome this challenge, it is crucial to develop innovative methods for monitoring and predicting treatment responses among patients with BC. Metabolomics is gaining recognition as a promising approach for discovering biomarkers. It has the potential to reveal metabolic disruptions that precisely reflect how BC patients respond to particular treatments, providing a revolutionary method to improve accuracy in monitoring and predicting outcomes. In this article, we present a comprehensive review of studies employing metabolomics approaches to investigate the metabolic responses associated with different treatment modalities for BC. The review encompasses an exploration of various models, samples, and analytical techniques applied in this context. Special emphasis is placed on the reported changes in metabolite levels derived from these studies, highlighting their potential as biomarkers for personalized medicine in BC.

A Clinically Relevant Dosage of Mitoxantrone Disrupts the Glutathione and Lipid Metabolic Pathways of the CD-1 Mice Brain: A Metabolomics Study.

Long-term cognitive dysfunction, or "chemobrain", has been observed in cancer patients treated with chemotherapy. Mitoxantrone (MTX) is a topoisomerase II inhibitor that binds and intercalates with DNA, being used in the treatment of several cancers and multiple sclerosis. Although MTX can induce chemobrain, its neurotoxic mechanisms are poorly studied. This work aimed to identify the adverse outcome pathways (AOPs) activated in the brain upon the use of a clinically relevant cumulative dose of MTX. Three-month-old male CD-1 mice were given a biweekly intraperitoneal administration of MTX over the course of three weeks until reaching a total cumulative dose of 6 mg/kg. Controls were given sterile saline in the same schedule. Two weeks after the last administration, the mice were euthanized and their brains removed. The left brain hemisphere was used for targeted profiling of the metabolism of glutathione and the right hemisphere for an untargeted metabolomics approach. The obtained results revealed that MTX treatment reduced the availability of cysteine (Cys), cysteinylglycine (CysGly), and reduced glutathione (GSH) suggesting that MTX disrupts glutathione metabolism. The untargeted approach revealed metabolic circuits of phosphatidylethanolamine, catecholamines, unsaturated fatty acids biosynthesis, and glycerolipids as relevant players in AOPs of MTX in our in vivo model. As far as we know, our study was the first to perform such a broad profiling study on pathways that could put patients given MTX at risk of cognitive deficits.

Comprehensive Metabolomics and Lipidomics Profiling of Prostate Cancer Tissue Reveals Metabolic Dysregulations Associated with Disease Development.

Prostate cancer (PCa) is a global health problem that affects millions of men every year. In the past decade, metabolomics and related subareas, such as lipidomics, have demonstrated an enormous potential to identify novel mechanisms underlying PCa development and progression, providing a good basis for the development of new and more effective therapies and diagnostics. In this study, a multiplatform metabolomics and lipidomics approach, combining untargeted mass spectrometry (MS) and nuclear magnetic resonance (NMR)-based techniques, was applied to PCa tissues to investigate dysregulations associated with PCa development, in a cohort of 40 patients submitted to radical prostatectomy for PCa. Results revealed significant alterations in the levels of 26 metabolites and 21 phospholipid species in PCa tissue compared with adjacent nonmalignant tissue, suggesting dysregulation in 13 metabolic pathways associated with PCa development. The most affected metabolic pathways were amino acid metabolism, nicotinate and nicotinamide metabolism, purine metabolism, and glycerophospholipid metabolism. A clear interconnection between metabolites and phospholipid species participating in these pathways was observed through correlation analysis. Overall, these dysregulations may reflect the reprogramming of metabolic responses to produce high levels of cellular building blocks required for rapid PCa cell proliferation.

Cardiotoxicity of cyclophosphamide's metabolites: an in vitro metabolomics approach in AC16 human cardiomyocytes.

Cyclophosphamide is a widely used anticancer and immunosuppressive prodrug that unfortunately causes severe adverse effects, including cardiotoxicity. Although the exact cardiotoxic mechanisms are not completely understood, a link between cyclophosphamide's pharmacologically active metabolites, namely 4-hydroxycyclophosphamide and acrolein, and the toxicity observed after the administration of high doses of the prodrug is likely. Therefore, the objective of this study is to shed light on the cardiotoxic mechanisms of cyclophosphamide and its main biotransformation products, through classic and metabolomics studies. Human cardiac proliferative and differentiated AC16 cells were exposed to several concentrations of the three compounds, determining their basic cytotoxic profile and preparing the next study, using subtoxic and toxic concentrations for morphological and biochemical studies. Finally, metabolomics studies were applied to cardiac cells exposed to subtoxic concentrations of the aforementioned compounds to determine early markers of damage. The cytotoxicity, morphological and biochemical assays showed that 4-hydroxycyclophosphamide and acrolein induced marked cardiotoxicity at µM concentrations (lower than 5 µM), being significantly lower than the ones observed for cyclophosphamide (higher than 2500 µM). Acrolein led to increased levels of ATP and total glutathione on proliferative cells at 25 µM, while no meaningful changes were observed in differentiated cells. Higher levels of carbohydrates and decreased levels of fatty acids and monoacylglycerols indicated a metabolic cardiac shift after exposure to cyclophosphamide's metabolites, as well as a compromise of precursor amino acids used in the synthesis of glutathione, seen in proliferative cells' metabolome. Overall, differences in cytotoxic mechanisms were observed for the two different cellular states used and for the three molecules, which should be taken into consideration in the study of cyclophosphamide cardiotoxic mechanisms.

Sunitinib versus Pazopanib Dilemma in Renal Cell Carcinoma: New Insights into the In Vitro Metabolic Impact, Efficacy, and Safety.

Sunitinib and pazopanib are tyrosine kinase inhibitors (TKIs) used as first-line therapy for metastatic renal cell carcinoma (RCC). Although these TKIs are associated with similar survival outcomes, some differences have been reported in their safety profiles. In this work, traditional toxicological endpoints (cell viability and growth, oxidative stress, and nuclear morphology) and 1H NMR spectroscopy-based metabolomics analysis were used to provide new insights into the cytotoxicity and metabolic mechanisms underlying sunitinib and pazopanib treatments. Tumoral (Caki-1) and non-tumoral (HK-2) human renal cells were exposed to clinically relevant concentrations of sunitinib (2 µM) or pazopanib (50 µM). Sunitinib showed selectivity for cancer cells, inhibiting proliferation, and inducing apoptotic death of Caki-1 cells, whereas pazopanib had a similar cytotoxic effect in both tumoral and non-tumoral cells. 1H-NMR metabolomics unveiled a higher impact of sunitinib on the levels of intracellular metabolites of Caki-1 cells (seven dysregulated metabolites), suggesting dysregulations on amino acid, glutathione and glycerophospholipid metabolisms. In contrast, pazopanib had a higher impact on the levels of extracellular metabolites of Caki-1 cells (seven dysregulated metabolites in culture medium), unveiling alterations on amino acid and energetic metabolisms. In HK-2 cells, sunitinib caused only a minor increase in intracellular isoleucine levels, whereas pazopanib induced several alterations on the intracellular (three dysregulated metabolites) and extracellular (three dysregulated metabolites) compartments suggesting changes on amino acid, glycerophospholipid, and energy metabolisms. Our results demonstrate that these TKIs elicit distinct cellular and metabolic responses, with sunitinib showing better in vitro efficacy against target RCC cells and lesser nephrotoxic potential than pazopanib.

Urinary Volatilomics Unveils a Candidate Biomarker Panel for Noninvasive Detection of Clear Cell Renal Cell Carcinoma.

Clear cell renal cell carcinoma (ccRCC) is the most common type of kidney cancer usually associated with asymptomatic development and risk of systemic progression. Hence, reliable molecular biomarkers of ccRCC are needed to provide early and minimally invasive detection. In this study, urinary volatilome profiling of patients diagnosed with ccRCC (n = 75), and cancer-free controls (n = 75), was performed to investigate the presence of a volatile signature characteristic of ccRCC. Volatile organic compounds (VOCs) in general, and more specifically volatile carbonyl compounds (VCCs), present in urine were extracted by headspace solid-phase microextraction coupled with gas chromatography-mass spectrometry (HS-SPME-GC-MS). Supervised multivariate models showed a good discriminatory power of ccRCC patients from controls in urine. Overall, 22 volatile metabolites were found significantly altered between the two groups, including aldehydes, ketones, aromatic hydrocarbons, and terpenoids. A candidate six-biomarker panel, comprising octanal, 3-methylbutanal, benzaldehyde, 2-furaldehyde, 4-heptanone, and p-cresol, depicted the best performance for ccRCC detection with 83% sensitivity, 79% specificity, and 81% accuracy. Moreover, the ccRCC urinary volatilome signature suggested dysregulation of energy metabolism and overexpression of enzymes associated with carcinogenesis. These findings provide the molecular basis for the fine-tuning of gas-sensing materials for application in the development of a bioelectronic sensor.

S-(+)-Pentedrone and R-(+)-methylone as the most oxidative and cytotoxic enantiomers to dopaminergic SH-SY5Y cells: Role of MRP1 and P-gp in cathinones enantioselectivity.

Cathinone derivatives are the most representative group within new drugs market, which have been described as neurotoxic. Since cathinones, as pentedrone and methylone, are available as racemates, it is our aim to study the neuronal cytotoxicity induced by each enantiomer. Therefore, a dopaminergic SH-SY5Y cell line was used to evaluate the hypothesis of enantioselectivity of pentedrone and methylone enantiomers on cytotoxicity, oxidative stress, and membrane efflux transport (confirmed by in silico studies). Our study demonstrated enantioselectivity of these cathinones, being the S-(+)-pentedrone and R-(+)-methylone the most oxidative enantiomers and also the most cytotoxic, suggesting the oxidative stress as main cytotoxic mechanism, as previously described in in vitro studies. Additionally, the efflux transporter multidrug resistance associated protein 1 (MRP1) seems to play, together with GSH, a selective protective role against the cytotoxicity induced by R-(-)-pentedrone enantiomer. It was also observed an enantioselectivity in the binding to P-glycoprotein (P-gp), another efflux protein, being the R-(-)-pentedrone and S-(-)-methylone the most transported enantiomeric compounds. These results were confirmed, in silico, by docking studies, revealing that R-(-)-pentedrone is the enantiomer with highest affinity to MRP1 and S-(-)-methylone and R-(-)-pentedrone are the enantiomers with highest affinity to P-gp. In conclusion, our data demonstrated that pentedrone and methylone present enantioselectivity in their cytotoxicity, which seems to involve different oxidative reactivity as well as different affinity to the P-gp and MRP1 that together with GSH play a protective role.

In vivo toxicometabolomics reveals multi-organ and urine metabolic changes in mice upon acute exposure to human-relevant doses of 3,4-methylenedioxypyrovalerone (MDPV).

3,4-Methylenedioxypyrovalerone (MDPV) is consumed worldwide, despite its potential to cause toxicity in several organs and even death. There is a recognized need to clarify the biological pathways through which MDPV elicits general and target-organ toxicity. In this work, a comprehensive untargeted GC-MS-based metabolomics analysis was performed, aiming to detect metabolic changes in putative target organs (brain, heart, kidneys and liver) but also in urine of mice after acute exposure to human-relevant doses of MDPV. Male CD-1 mice received binge intraperitoneal administrations of saline or MDPV (2.5 mg/kg or 5 mg/kg) every 2 h, for a total of three injections. Twenty-four hours after the first administration, target organs, urine and blood samples were collected for metabolomics, biochemical and histological analysis. Hepatic and renal tissues of MDPV-treated mice showed moderate histopathological changes but no significant differences were found in plasma and tissue biochemical markers of organ injury. In contrast, the multivariate analysis significantly discriminated the organs and urine of MDPV-treated mice from the control (except for the lowest dose in the brain), allowing the identification of a panoply of metabolites. Those levels were significantly deviated in relation to physiological conditions and showed an organ specific response towards the drug. Kidneys and liver showed the greatest metabolic changes. Metabolites related with energetic metabolism, antioxidant defenses and inflammatory response were significantly changed in the liver of MDPV-dosed animals, while the kidneys seem to have developed an adaptive response against oxidative stress caused by MDPV. On the other hand, the dysregulation of metabolites that contribute to metabolic acidosis was also observed in this organ. The heart showed an increase of fatty acid biosynthesis, possibly as an adaptation to maintain the cardiac energy homeostasis. In the brain, changes in 3-hydroxybutyric acid levels may reflect the activation of a neurotoxic pathway. However, the increase in metabolites with neuroprotective properties seems to counteract this change. Metabolic profiling of urine from MDPV-treated mice suggested that glutathione-dependent antioxidant pathways may be particularly involved in the compensatory mechanism to counteract oxidative stress induced by MDPV. Overall, this study reports, for the first time, the metabolic profile of liver, kidneys, heart, brain, and urine of MDPV-dosed mice, providing unique insights into the biological pathways of toxicity. Our findings also underline the value of toxicometabolomics as a robust and sensitive tool for detecting adaptive/toxic cellular responses upon exposure to a physiologically relevant dose of a toxic agent, earlier than conventional toxicity tests.

3,4-Methylenedioxymethamphetamine Hepatotoxicity under the Heat Stress Condition: Novel Insights from in Vitro Metabolomic Studies.

Hyperthermia has been extensively reported as a life-threatening consequence of 3,4-methylenedioxymethamphetamine (MDMA, ecstasy) abuse. In this work, we used a sensitive untargeted metabolomic approach based on gas chromatography-mass spectrometry to evaluate the impact of hyperthermia on the hepatic metabolic changes caused by MDMA. For this purpose, primary mouse hepatocytes were exposed to subtoxic (LC01 and LC10) and toxic (LC30) concentrations of MDMA for 24 h, at 37 or 40.5 °C (simulating body temperature increase after MDMA consumption), and alterations on both intracellular metabolome and extracellular volatilome were evaluated. Multivariate analysis showed that metabolic patterns clearly discriminate MDMA treated cells from control cells, both in normothermic and hyperthermic conditions. The metabolic signature was found to be largely common to MDMA subtoxic and toxic concentrations, although with evident differences in the magnitude of response, with metabolic changes significantly more pronounced at 40.5 °C. Discriminant metabolites associated with MDMA-induced hepatotoxicity are mostly involved in the amino acid metabolism, aminoacyl tRNA biosynthesis, glutathione metabolism, tricarboxylic acid cycle, and pyruvate metabolism. Moreover, our metabolomic findings were corroborated by classical toxicity parameters, demonstrating the high sensitivity of this omic approach to assess molecular-level effects. Overall, this study indicates that MDMA triggers significant metabolic alterations on hepatic cells, even at low concentrations, that are clearly exacerbated at high temperatures. These findings provide new metabolic pieces to solve the puzzle of MDMA's hepatotoxicity mechanism and emphasize the increased risks of MDMA abuse due to the thermogenic action of the drug.

Enantioselectivity on the absorption of methylone and pentedrone using Caco-2 cell line: Development and validation of an UHPLC method for cathinones quantification.

Synthetic cathinones, such as methylone and pentedrone, are psychoactive derivatives of cathinone, sold in the internet as "plant food" or "bath salts". However, the level at which these compounds and their enantiomers cross the intestinal barrier has not been yet determined. Thus, the present study aimed to analyze the enantioselectivity on the permeability of these drugs through the intestinal barrier by using the Caco-2 cell line, a widely used in vitro model for drug permeability studies. To achieve this goal, an UHPLC-UV method was developed and validated to quantify both synthetic cathinones. The developed UHPLC-UV method revealed high selectivity and a linearity from 1 to 500 µM with correlation coefficients always higher than 0.999. The method has an accuracy that ranged between 89 and 107%, inter-day and intra-day precisions with coefficients of variation below 10%, limits of detection and quantification of 0.31 µM and 0.93 µM for methylone and 0.17 µM and 0.52 µM for pentedrone, respectively. In Caco-2 cells, a differentiated passage of the enantiomers across monolayer was observed for both cathinones. For pentedrone, the difference was observed after the first hour, being R-(-)-pentedrone the most permeable compound. Regarding methylone, the difference was noted after one hour and 30 min, with S-(-)-methylone being the most absorbed enantiomer. In conclusion, a fully validated method was successfully applied for studying the permeability of methylone and pentedrone enantiomers in an in vitro model of human intestine, which allowed to discover, for the first time, the enantioselectivity in drug permeability of this class of drugs.

Diet aid or aid to die: an update on 2,4-dinitrophenol (2,4-DNP) use as a weight-loss product.

During the last decades, we have witnessed unparalleled changes in human eating habits and lifestyle, intensely influenced by cultural and social pressures. Sports practice became strongly implemented in daily routines, and visits to the gym peaked, driven by the indulgence in intensive 'weight-loss programs'. The pledge of boasting a healthy and beautiful body instigates the use of very attractive 'fat burners', which are purportedly advertised as safe products, easily available in the market and expected to quickly reduce body weight. In this context, the slimming properties of 2,4-dinitrophenol (2,4-DNP) galvanised its use as a weight-loss product, despite the drug ban for human consumption in many countries since 1938, due to its adverse effects. The main symptoms associated with 2,4-DNP intoxication, including hyperthermia, tachycardia, decreased blood pressure, and acute renal failure, motivated a worldwide warning, issued by the Interpol Anti-Doping Unit in 2015, reinforcing its hazard. Information on the effects of 2,4-DNP mainly derive from the intoxication cases reported by emergency care units, for which there is no specific antidote or treatment. This review provides a comprehensive update on 2,4-DNP use, legislation and epidemiology, chemistry and analytical methodologies for drug determination in commercial products and biological samples, pharmacokinetics and pharmacodynamics, toxicological effects, and intoxication diagnosis and management.

Identification of a biomarker panel for improvement of prostate cancer diagnosis by volatile metabolic profiling of urine.

BACKGROUND: The lack of sensitive and specific biomarkers for the early detection of prostate cancer (PCa) is a major hurdle to improve patient management. METHODS: A metabolomics approach based on GC-MS was used to investigate the performance of volatile organic compounds (VOCs) in general and, more specifically, volatile carbonyl compounds (VCCs) present in urine as potential markers for PCa detection. RESULTS: Results showed that PCa patients (n = 40) can be differentiated from cancer-free subjects (n = 42) based on their urinary volatile profile in both VOCs and VCCs models, unveiling significant differences in the levels of several metabolites. The models constructed were further validated using an external validation set (n = 18 PCa and n = 18 controls) to evaluate sensitivity, specificity and accuracy of the urinary volatile profile to discriminate PCa from controls. The VOCs model disclosed 78% sensitivity, 94% specificity and 86% accuracy, whereas the VCCs model achieved the same sensitivity, a specificity of 100% and an accuracy of 89%. Our findings unveil a panel of 6 volatile compounds significantly altered in PCa patients' urine samples that was able to identify PCa, with a sensitivity of 89%, specificity of 83%, and accuracy of 86%. CONCLUSIONS: It is disclosed a biomarker panel with potential to be used as a non-invasive diagnostic tool for PCa.

Occupational exposure to formaldehyde and early biomarkers of cancer risk, immunotoxicity and susceptibility.

Formaldehyde (FA) is a high-volume production chemical manufactured worldwide to which many people are exposed to both environmentally and occupationally. FA was recently reclassified as a human carcinogen. Several epidemiological studies have revealed an increased risk of cancer development among workers exposed to FA. Although FA genotoxicity was confirmed in a variety of experimental systems, data from human studies are conflicting. The aim of the present study was to evaluate the occupational exposure to FA in a multistage approach relating the exposure with different biomarkers (dose and effect) and individual susceptibility. Air monitoring was performed to estimate the level of exposure to FA during shift work. Eighty-five workers from hospital anatomy-pathology laboratories exposed to FA and 87 controls were tested for cytogenetic alterations in lymphocytes (micronucleus, MN; sister-chromatid exchange, SCE) and T-cell receptor (TCR) mutation assay. The frequency of MN in exfoliated buccal cells, a first contact tissue was also assessed. Percentages of different lymphocyte subpopulations were selected as immunotoxicity biomarkers. The level of formic acid in urine was investigated as a potential biomarker of internal dose. The effects of polymorphic genes of xenobiotic metabolising enzymes and DNA repair enzymes on the endpoints studied were determined. The mean level of FA exposure was 0.38 ±â€¯0.03 ppm. MN (in lymphocytes and buccal cells) and SCE were significantly increased in FA-exposed workers compared to controls. MN frequency positively correlated with FA levels of exposure and duration. Significant alterations in the percentage of T cytotoxic lymphocytes, NK cells and B lymphocytes were found between groups. Polymorphisms in CYP2E1, GSTP1 and FANCA genes were associated with increased genetic damage in FA-exposed subjects. The obtained information may provide new important data to be used by health and safety care programs and by governmental agencies responsible for setting the acceptable levels for occupational exposure to FA.

NMR-based metabolomics studies of human prostate cancer tissue.

INTRODUCTION: Prostate cancer (PCa) is one of the most prevalent cancers in men worldwide. Serum prostate-specific antigen (PSA) remains the most used biomarker in the detection and management of patients with PCa, in spite of the problems related with its low specificity, false positive rate and overdiagnosis. Furthermore, PSA is unable to discriminate indolent from aggressive PCa, which can lead to overtreatment. Early diagnosed and treated PCa can have a good prognosis and is potentially curable. Therefore, the discovery of new biomarkers able to detect clinically significant aggressive PCa is urgently needed. METHODS: This revision was based on an electronic literature search, using Pubmed, with Nuclear Magnetic Resonance (NMR), tissue and prostate cancer as keywords. All metabolomic studies performed in PCa tissues by NMR spectroscopy, from 2007 until March 2018, were included in this review. RESULTS: In the context of cancer, metabolomics allows the analysis of the entire metabolic profile of cancer cells. Several metabolic alterations occur in cancer cells to sustain their abnormal rates of proliferation. NMR proved to be a suitable methodology for the evaluation of these metabolic alterations in PCa tissues, allowing to unveil alterations in citrate, spermine, choline, choline-related compounds, lactate, alanine and glutamate. CONCLUSION: The study of the metabolic alterations associated with PCa progression, accomplished by the analysis of PCa tissue by NMR, offers a promising approach for elucidating biochemical pathways affected by PCa and also for discovering new clinical biomarkers. The main metabolomic alterations associated with PCa development and promising biomarker metabolites for diagnosis of PCa were outlined.

GC-MS metabolomics reveals disturbed metabolic pathways in primary mouse hepatocytes exposed to subtoxic levels of 3,4-methylenedioxymethamphetamine (MDMA).

3,4-Methylenedioxymethamphetamine (MDMA, ecstasy) is a well-known hepatotoxic drug. Although its toxicity has been thoroughly studied at high concentrations, there is still insufficient knowledge on possible alterations of cell function at subtoxic concentrations, which are in fact more representative concentrations of intoxication scenarios. In this study, a gas chromatography-mass spectrometry (GC-MS) metabolomics approach was used to investigate the metabolic changes in primary mouse hepatocytes (PMH) exposed to two subtoxic concentrations of MDMA (LC01 and LC10) for 24 h. Metabolomic profiling of both intracellular metabolites and volatile metabolites in the extracellular medium of PMH was performed. Multivariate analysis showed that the metabolic pattern of cells exposed to MDMA discriminates from the controls in a concentration-dependent manner. Exposure to LC10 MDMA induces a significant increase in some intracellular metabolites, including oleic acid and palmitic acid, and a decrease in glutamate, aspartate, 5-oxoproline, fumarate, malate, phosphoric acid, α-ketoglutarate and citrate. Extracellular metabolites such as acetophenone, formaldehyde, pivalic acid, glyoxal and 2-butanone were found significantly increased after exposure to MDMA, compared to controls, whereas 4-methylheptane, 2,4-dimethyl-1-heptene, nonanal, among others, were found significantly decreased. The panel of discriminatory metabolites is mainly involved in tricarboxylic acid (TCA) cycle, fatty acid metabolism, glutamate metabolism, antioxidant defenses and possibly changes in the liver enzyme machinery. Overall, these results highlight the potential of the intra- and extracellular metabolome to study alterations triggered by subtoxic concentrations of MDMA in hepatic cell functions, which represents a more realistic appraisal of early toxicity events posed by exposure to this drug. In addition, these results also revealed some metabolites that may be used as potential biomarkers indicative of early events in the hepatotoxicity induced by MDMA.

GC-MS metabolomics-based approach for the identification of a potential VOC-biomarker panel in the urine of renal cell carcinoma patients.

The analysis of volatile organic compounds (VOCs) emanating from biological samples appears as one of the most promising approaches in metabolomics for the study of diseases, namely cancer. In fact, it offers advantages, such as non-invasiveness and robustness for high-throughput applications. The purpose of this work was to study the urinary volatile metabolic profile of patients with renal cell carcinoma (RCC) (n = 30) and controls (n = 37) with the aim of identifying a potential specific urinary volatile pattern as a non-invasive strategy to detect RCC. Moreover, the effect of some confounding factors such as age, gender, smoking habits and body mass index was evaluated as well as the ability of urinary VOCs to discriminate RCC subtypes and stages. A headspace solid-phase microextraction/gas chromatography-mass spectrometry-based method was performed, followed by multivariate data analysis. A variable selection method was applied to reduce the impact of potential redundant and noisy chromatographic variables, and all models were validated by Monte Carlo cross-validation and permutation tests. Regarding the effect of RCC on the urine VOCs composition, a panel of 21 VOCs descriptive of RCC was defined, capable of discriminating RCC patients from controls in principal component analysis. Discriminant VOCs were further individually validated in two independent samples sets (nine RCC patients and 12 controls, seven RCC patients with diabetes mellitus type 2) by univariate statistical analysis. Two VOCs were found consistently and significantly altered between RCC and controls (2-oxopropanal and, according to identification using NIST14, 2,5,8-trimethyl-1,2,3,4-tetrahydronaphthalene-1-ol), strongly suggesting enhanced potential as RCC biomarkers. Gender, smoking habits and body mass index showed negligible and age-only minimal effects on the urinary VOCs, compared to the deviations resultant from the disease. Moreover, in this cohort, the urinary volatilome did not show ability to discriminate RCC stages and histological subtypes. The results validated the value of urinary volatilome for the detection of RCC and advanced with the identification of potential RCC urinary biomarkers.

Metabolomic approaches in the discovery of potential urinary biomarkers of drug-induced liver injury (DILI).

Drug-induced liver injury (DILI) is a major safety issue during drug development, as well as the most common cause for the withdrawal of drugs from the pharmaceutical market. The identification of DILI biomarkers is a labor-intensive area. Conventional biomarkers are not specific and often only appear at significant levels when liver damage is substantial. Therefore, new biomarkers for early identification of hepatotoxicity during the drug discovery process are needed, thus resulting in lower development costs and safer drugs. In this sense, metabolomics has been increasingly playing an important role in the discovery of biomarkers of liver damage, although the characterization of the mechanisms of toxicity induced by xenobiotics remains a huge challenge. These new-generation biomarkers will offer obvious benefits for the pharmaceutical industry, regulatory agencies, as well as a personalized clinical follow-up of patients, upon validation and translation into clinical practice or approval for routine use. This review describes the current status of the metabolomics applied to the early diagnosis and prognosis of DILI and in the discovery of new potential urinary biomarkers of liver injury.