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Trypanosoma brucei is the causative agent of Human African Trypanosomiasis (also known as sleeping sickness), a disease causing serious neurological disorders and fatal if left untreated. Due to its lethal pathogenicity, a variety of treatments have been developed over the years, but which have some important limitations such as acute toxicity and parasite resistance. Metabolomics is an innovative tool used to better understand the parasite's cellular metabolism, and identify new potential targets, modes of action and resistance mechanisms. The metabolomic approach is mainly associated with robust analytical techniques, such as NMR and Mass Spectrometry. Applying these tools to the trypanosome parasite is, thus, useful for providing new insights into the sleeping sickness pathology and guidance towards innovative treatments. The present review aims to comprehensively describe the T. brucei biology and identify targets for new or commercialized antitrypanosomal drugs. Recent metabolomic applications to provide a deeper knowledge about the mechanisms of action of drugs or potential drugs against T. brucei are highlighted. Additionally, the advantages of metabolomics, alone or combined with other methods, are discussed. Compared to other parasites, only few studies employing metabolomics have to date been reported on Trypanosoma brucei. Published metabolic studies, treatments and modes of action are discussed. The main interest is to evaluate the metabolomics contribution to the understanding of T. brucei's metabolism.

Metabolomics is a reliable omics tool to study the metabolome, the assortment of metabolites that provide energy, signaling or building blocks essential for biological systems survival. • Malaria is a deadly disease, especially severe when caused by Plasmodium falciparum, that still affects over 200 million people yearly.1 • The metabolome closely reflects the state of the biological system, and if applied to the P. falciparum, it can be used to characterize antimalarial mechanisms of action or study resistance. • The parasite’s intracellular nature, in red blood cells in suspension, introduces significant hurdles to metabolomics extraction methods.

Arginase 1 (Arg1) is a key hydrolase involved in the urea cycle and in the immune system. Arg1 is upregulated in various types of cancers and plays crucial roles in the regulation of tumor growth and metastasis. Thus, Arg1 is an increasingly attractive target for cancer therapy. In this thesis, we aimed to find new scaffolds. A reliable enzymatic evaluation method of chemical libraries such as that of 5-bromo-2-amino-thiazole was set up. A molecular modeling approach (SBDD) led to a better understanding of the possibilities and limitations in the Arg1 inhibitors design for the study of larger scaffolds. A Fragment-Based approach (FBDD) was then developed. A screening of coordinating motif libraries found two hits: 2-furanylboronic acid and cyclohexylboronic acid. Finally, we attempt to optimize these two hits and discussed about the challenges of obtaining such derivatives., (BIFA - Sciences biomédicales et pharmaceutiques) -- UCL, 2022

Malaria is a parasitic disease that remains a global health issue, responsible for a significant death and morbidity toll. Various factors have impacted the use and delayed the development of antimalarial therapies, such as the associated financial cost and parasitic resistance. In order to discover new drugs and validate parasitic targets, a powerful omics tool, metabolomics, emerged as a reliable approach. However, as a fairly recent method in malaria, new findings are timely and original practices emerge frequently. This review aims to discuss recent research towards the development of new metabolomic methods in the context of uncovering antiplasmodial mechanisms of action in vitro and to point out innovative metabolic pathways that can revitalize the antimalarial pipeline.

Neovascular age-related macular degeneration (nAMD) is the leading cause of blindness in aging populations. Here, we applied metabolomics to human sera of patients with nAMD during an active (exudative) phase of the pathology and found higher lactate levels and a shift in the lipoprotein profile (increased VLDL-LDL/HDL ratio). Similar metabolomics changes were detected in the sera of mice subjected to laser-induced choroidal neovascularization (CNV). In this experimental model, we provide evidence for two sites of lactate production: first, a local one in the injured eye, and second a systemic site associated with the recruitment of bone marrow-derived inflammatory cells. Mechanistically, lactate promotes the angiogenic response and M2-like macrophage accumulation in the eyes. The therapeutic potential of our findings is demonstrated by the pharmacological control of lactate levels through pyruvate dehydrogenase kinase (PDK) inhibition by dichloroacetic acid (DCA). Mice treated with DCA exhibited normalized lactate levels and lipoprotein profiles, and inhibited CNV formation. Collectively, our findings implicate the key role of the PDK/lactate axis in AMD pathogenesis and reveal that the regulation of PDK activity has potential therapeutic value in this ocular disease. The results indicate that the lipoprotein profile is a traceable pattern that is worth considering for patient follow-up. KEY MESSAGES: Lactate and lipoprotein profile are associated with the active phase of AMD and CNV development. Lactate is a relevant and functional metabolite correlated with AMD progression. Modulating lactate through pyruvate dehydrogenase kinase led to a decrease of CNV progression. Pyruvate dehydrogenase kinase is a new therapeutic target for neovascular AMD.

Along with Mass spectrometry data, metabolomics studies are commonly based on proton 1D 1H-NMR spectral multivariate data sources, typically characterized by a high dimensionality. Very powerful and complex pre-processing and analytical tools are adapted to these data, well-mastered and have demonstrated their potential. Nevertheless, when the studied medium is very rich in metabolites, some limitations quickly occur: spectral peaks overlapping, multiplicity of peaks for a same product, difficulty of peak interpretation… 2D spectral data sources (as COSY, for COrrelation SpectroscopY) can overcome these limitations by construction via the additional information contained in correlation cross peaks. If this information is really relevant, 2D peaks help identify and quantify with more accuracy and certainty metabolites of interest. But COSY spectra are not yet widely used and there is not an agreed upon best practice for handling, pre-processing and assessment of 2D NMR spectra in metabolomics studies. In this thesis, a whole process is proposed to promote a more massive use of 2D data in such studies. First, for handling, grouping in a global object all the individual spectra stemming from a design and for pre-processing via the novel Global Peak List (GPL) and Vectorization workflows. Then, for assessing their quality in terms of signal capture via the Metabolomic Informative Content (MIC) concept, which is based on unsupervised clustering quality measures. MIC indexes are proposed to compare 1D and 2D spectral matrices, and different 2D data sets pre-processed with various scenarii. Finally, the biomarker issue is deeply considered (using PCA, PLS, OPLS, sPLS and ensemble trees approaches) and the orthogonality and sparsity issues are addressed. A novel sparse algorithm is proposed: L-sOPLS which can combine orthogonality, sparsity and efficient predictions. This process was convincingly applied on six real experimental designs., (SC - Sciences) -- UCL, 2019

Introduction The pre-processing of analytical data in metabolomics must be considered as a whole to allow the construction of a global and unique object for any further simultaneous data analysis or multivariate statistical modelling. For 1D 1H-NMR metabolomics experiments, best practices for data pre-processing are well defined, but not yet for 2D experiments (for instance COSY in this paper). Objective By considering the added value of a second dimension, the objective is to propose two workflows dedicated to 2D NMR data handling and preparation (the Global Peak List and Vectorization approaches) and to compare them (with respect to each other and with 1D standards). This will allow to detect which methodology is the best in terms of amount of metabolomic content and to explore the advantages of the selected workflow in distinguishing among treatment groups and identifying relevant biomarkers. Therefore, this paper explores both the necessity of novel 2D pre-processing workflows, the evaluation of their quality and the evaluation of their performance in the subsequent determination of accurate (2D) biomarkers. Methods To select the more informative data source, MIC (Metabolomic Informative Content) indexes are used, based on clustering and inertia measures of quality. Then, to highlight biomarkers or critical spectral zones, the PLS-DA model is used, along with more advanced sparse algorithms (sPLS and L-sOPLS). Results Results are discussed according to two different experimental designs (one which is unsupervised and based on human urine samples, and the other which is controlled and based on spiked serum media). MIC indexes are shown, leading to the choice of the more relevant workflow to use thereafter. Finally, biomarkers are provided for each case and the predictive power of each candidate model is assessed with cross-validated measures of RMSEP. Conclusion In conclusion, it is shown that no solution can be universally the best in every case, but that 2D exp

In the analysis of biological samples, control over experimental design and data acquisition procedures alone cannot ensure well-conditioned 1H NMR spectra with maximal information recovery for data analysis. A third major element affects the accuracy and robustness of results: the data pre-processing/ pre-treatment for which not enough attention is usually devoted, in particular in metabolomic studies. The usual approach is to use proprietary software provided by the analytical instruments' manufacturers to conduct the entire pre-processing strategy. This widespread practice has a number of advantages such as a user-friendly interface with graphical facilities, but it involves non-negligible drawbacks: a lack of methodological information and automation, a dependency of subjective human choices, only standard processing possibilities and an absence of objective quality criteria to evaluate pre-processing quality. This paper introduces PepsNMR to meet these needs, an R package dedicated to the whole processing chain prior to multivariate data analysis, including, among other tools, solvent signal suppression, internal calibration, phase, baseline and misalignment corrections, bucketing and normalisation. Methodological aspects are discussed and the package is compared to the gold standard procedure with two metabolomic case studies. The use of PepsNMR on these data shows better information recovery and predictive power based on objective and quantitative quality criteria. Other key assets of the package are workflow processing speed, reproducibility, reporting and flexibility, graphical outputs and documented routines.

Kidney injury is a common complication of severe disease. Here, we report that injuries of the zebrafish embryonal kidney are rapidly repaired by a migratory response in 2-, but not in 1-day-old embryos. Gene expression profiles between these two developmental stages identify cxcl12a and myca as candidates involved in the repair process. Zebrafish embryos with cxcl12a, cxcr4b, or myca deficiency display repair abnormalities, confirming their role in response to injury. In mice with a kidney-specific knockout, Cxcl12 and Myc gene deletions suppress mitochondrial metabolism and glycolysis, and delay the recovery after ischemia/reperfusion injury. Probing these observations in zebrafish reveal that inhibition of glycolysis slows fast migrating cells and delays the repair after injury, but does not affect the slow cell movements during kidney development. Our findings demonstrate that Cxcl12 and Myc facilitate glycolysis to promote fast migratory responses during development and repair, and potentially also during tumor invasion and metastasis.

Utilisation de la métabonomique en spectroscopie RMN pour l'identification de biomarqueurs d'exposition à l'acide Aristolochique. Développement de modèles expérimentaux chez des rats mâles. Analyse d'échantillons urinaires provenant de patients croates potentiellement touchés par la Néphropathie endémique des Balkans, Doctorat en Sciences biomédicales et pharmaceutiques (Médecine), info:eu-repo/semantics/nonPublished

In the analysis of complex biological samples, control over experimental design and data acquisition procedures cannot ensure alone well-conditioned 1H-NMR spectra with maximal information recovery for data analysis. A third major element affects the accuracy and robustness of the results: the data pre-processing/pre-treatment for which not enough attention is usually devoted, in particular in the metabolomic studies. The usual approach is to use proprietary software provided by the analytical instruments’ manufacturers to conduct the whole pre-processing strategies. This widespread practice has a number of advantages such as a user-friendly interface with graphical facilities, yet it involve non-negligible drawbacks: a lack of methodological information and automation, a dependency of subjective human choices, only basic treatment possibilities and an absence of objective quality criteria to evaluate the pre-processing quality. This paper introduces PepsNMR to meet the needs, a R package dedicated to the whole processing chain prior to multivariate data analysis including, among other tools, solvent signal suppression, internal calibration, phase, baseline and misalignment corrections, bucketing and normalisation. Methodological aspects are discussed and the library is compared to the gold standard procedure with two metabolomic case studies. The use of PepsNMR on these data shows better information recovery and predictive power based on objective and quantitative quality criteria. Other key assets of the library are workflow processing speed, reproducibility, reporting and flexibility, graphical outputs and documented routines.

Introduction: Pre-analytical processing significantly affects tissue metabolomes. Since most frozen kidney samples are stored after embedding, standardization of cryoprotective medium removal before metabolomics is essential. Objectives: We used rodent and human kidney samples to develop an easy and robust pre-analytical procedure compatible with 1H-nuclear magnetic resonance (NMR)-based metabolomics. Methods: In mice, renal ischemia was induced for 30 min, followed by 48-h reperfusion (I/R, n = 6). Right kidneys were transversally cut in two fragments, and snap-frozen in liquid nitrogen (LN2) or in Optimal Cutting Temperature® (OCT) fixative. In man, double kidney biopsies were simultaneously obtained before transplantation (n = 15), and snap-frozen in LN2 or OCT. Results: 1H-NMR spectrum of pure OCT highlighted two major peaks, i.e. from 3.4 to 4.2 ppm (47.2%) and from 1.2 to 2.2 ppm (42.5%). 1H-NMR spectra of mouse OCT kidneys were biased at 3.7. By contrast, 1H-NMR analyses of mouse OCT kidneys iteratively rinsed in saline significantly discriminated sham versus I/R groups, with Q² at 0.695 (to be compared with Q² at 0.866 for LN2 sham vs. I/R kidneys). Discriminant metabolites were analogous in both OCT and LN2 kidneys, with a correlation coefficient of 0.83. In man, iteratively rinsing OCT kidneys in saline eliminated the spectral 3.7-peak, thereby making metabolomes of OCT kidneys interpretable and similar to LN2 samples, with a correlation coefficient of 0.73. Conclusion: NMR metabolomics using OCT-frozen kidney samples is valuable in mouse and man, following standardized OCT removal. This may help use residual biobanked human tissues to better understand renal pathophysiology.

The “Journées Franco-Belges de Pharmacochimie” is a recognized two-day annual meeting on Medicinal Chemistry that is renowned for the advanced science presented, conviviality, and outstanding opportunities for senior and young scientists to exchange knowledge. Abstracts of plenary lectures, oral communications, and posters presented during the meeting are collected in this report., SCOPUS: ar.j, info:eu-repo/semantics/published

A new series of N-aryl-N′-3,4-dihydro-2,2-dimethyl-2H-1-benzopyran-4-yl)ureas bearing an alkoxycarbonylamino group at the 6-position were synthesized and examined as putative anticancer agents targeting sirtuins in glioma cells. On the basis of computational docking combined to in vitro sirtuin 1/2 inhibition assays, we selected compound 18 [R/S-N-3-cyanophenyl-N′-(6-tert-butoxycarbonylamino-3,4-dihydro-2,2-dimethyl-2H-1-benzopyran-4-yl)urea] which displays a potent antiproliferative activity on various glioma cell types, assessed by quantitative videomicroscopy, eventually triggering senescence. The impact on normal glial cells was lower with a selectivity index of >10. Furthermore, human U373 and Hs683 glioblastoma cell lines served to demonstrate the inhibitory activity of 18 against histone deacetylase (HDAC) class III sirtuins 1 and 2 (SIRT1/2) by quantifying acetylation levels of histone and non-histone proteins. The translational potential of 18 was validated by an NCI-60 cell line screen and validation of growth inhibition of drug resistant cancer cell models. Eventually, the anticancer potential of 18 was validated in 3D glioblastoma spheroids and in vivo by zebrafish xenografts. In summary, compound 18 is the first representative of a new class of SIRT inhibitors opening new perspectives in the medicinal chemistry of HDAC inhibitors., SCOPUS: ar.j, info:eu-repo/semantics/published

Preoperative radiotherapy (RT) is a mainstay in the management of rectal cancer, a tumor characterized by desmoplastic stroma containing cancer-associated fibroblasts (CAF). Although CAFs are abundantly present, the effects of RT to CAF and its impact on cancer cells are unknown. We evaluated the damage responses of CAF to RT and investigated changes in colorectal cancer cell growth, transcriptome, metabolome, and kinome in response to paracrine signals emerging from irradiated CAF. RT to CAF induced DNA damage, p53 activation, cell-cycle arrest, and secretion of paracrine mediators, including insulin-like growth factor-1 (IGF1). Subsequently, RT-activated CAFs promoted survival of colorectal cancer cells, as well as a metabolic switch favoring glutamine consumption through IGF1 receptor (IGF1R) activation. RT followed by IGF1R neutralization in orthotopic colorectal cancer models reduced the number of mice with organ metastases. Activation of the downstream IGF1R mediator mTOR was significantly higher in matched (intrapatient) samples and in unmatched (interpatient) samples from rectal cancer patients after neoadjuvant chemoradiotherapy. Taken together, our data support the notion that paracrine IGF1/ IGF1R signaling initiated by RT-activated CAF worsens colorectal cancer progression, establishing a preclinical rationale to target this activation loop to further improve clinical responses and patient survival. Significance: These findings reveal that paracrine IGF1/IGF1R signaling promotes colorectal cancer progression, establishing a preclinical rationale to target this activation loop., SCOPUS: ar.j, info:eu-repo/semantics/published

The synthesis of 2,2-dimethylchromans bearing a 3/4-chloro/cyano-substituted phenylureido or phenylthioureido moiety at the 4-position and an alkoxycarbonylamino ('carbamate') group at the 6-position is described. These new analogs of the potassium channel opener (±)-cromakalim were further tested on rat pancreatic islets as putative inhibitors of insulin release and on rat aorta rings as putative vasorelaxants. All compounds inhibited insulin secretion and induced a myorelaxant activity. Compound 14o [R/S-N-3-cyanophenyl-N'-(6-tert-butoxycarbonylamino-3,4-dihydro-2,2-dimethyl-2H-1-benzopyran-4-yl)urea; BPDZ 711] emerged as the most potent inhibitor of the glucose-sensitive insulin releasing process (IC50 = 0.24 μM) and displayed selectivity towards the pancreatic endocrine tissue. Radioisotopic, fluorimetric and pharmacological investigations were performed on rat pancreatic islet and rat vascular smooth muscle cells in order to decipher its mechanism of action. Our findings suggest that the mechanism of action of 14o is rather unspecific. The compound behaves as a KATP channel opener, a Ca2+ entry blocker, and promotes an intracellular calcium translocation., SCOPUS: ar.j, info:eu-repo/semantics/published

A series of 1,3-benzothiazoles (series I) and 4,5,6,7-tetrahydro-1,3-benzothiazoles (series II) bearing an urea or a thiourea moiety at the 2-position were synthesized and tested as myorelaxants and inhibitors of insulin secretion. Several compounds (i.e. 13u and 13v) from series I showed a marked myorelaxant activity. Benzothiazoles bearing a strong electron withdrawing group (NO2, CN) at the 6-position and an alkyl group linked to the urea or the thiourea function at the 2-position were found to be the most potent compounds. The weak vasorelaxant activity of series II compounds evidenced the necessity of the presence of a complete aromatic heterocyclic system. The myorelaxant activity of some active compounds was reduced when measured on aorta rings precontracted by 80 mM KCl or by 30 mM KCl in the presence of 10 μM glibenclamide, suggesting the involvement of KATP channels in the vasorelaxant effect. Some compounds of series I tested on rat pancreatic islets provoked a marked inhibition of insulin secretion, among which 13a exhibited a clear tissue selectivity for pancreatic β-cells., SCOPUS: ar.j, info:eu-repo/semantics/published

Myoferlin is a multiple C2-domain-containing protein that regulates membrane repair, tyrosine kinase receptor function and endocytosis in myoblasts and endothelial cells. Recently it has been reported as overexpressed in several cancers and shown to contribute to proliferation, migration and invasion of cancer cells. We have previously demonstrated that myoferlin regulates epidermal growth factor receptor activity in breast cancer. In the current study, we report a consistent overexpression of myoferlin in triple-negative breast cancer cells (TNBC) over cells originating from other breast cancer subtypes. Using a combination of proteomics, metabolomics and electron microscopy, we demonstrate that myoferlin depletion results in marked alteration of endosomal system and metabolism. Mechanistically, myoferlin depletion caused impaired vesicle traffic that led to a misbalance of saturated/unsaturated fatty acids. This provoked mitochondrial dysfunction in TNBC cells. As a consequence of the major metabolic stress, TNBC cells rapidly triggered AMP activated protein kinase-mediated metabolic reprogramming to glycolysis. This reduced their ability to balance between oxidative phosphorylation and glycolysis, rendering TNBC cells metabolically inflexible, and more sensitive to metabolic drug targeting in vitro. In line with this, our in vivo findings demonstrated a significantly reduced capacity of myoferlin-deficient TNBC cells to metastasise to lungs. The significance of this observation was further supported by clinical data, showing that TNBC patients whose tumors overexpress myoferlin have worst distant metastasis-free and overall survivals. This novel insight into myoferlin function establishes an important link between vesicle traffic, cancer metabolism and progression, offering new diagnostic and therapeutic concepts to develop treatments for TNBC patients.Oncogene advance online publication, 24 October 2016; doi:10.1038/onc.2016.369., SCOPUS: ar.j, info:eu-repo/semantics/published