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Inhibiteurs à visée thérapeutique de la phosphomannose isomérase de Candida albicans et du facteur de motilité autocrine : études cinétiques, structurales, mécanistiques et diagnostiques

Abstract : Phosphoglucose isomerase (PGI) and type I phosphomannose isomerase (PMI), a zinc metalloenzyme, catalyze the reversible isomerization of β-D-fructose 6-phosphate (F6P) to α-D-glucose 6-phosphate (G6P) and β-D-mannose 6-phosphate (M6P), respectively. These two enzymes are potential therapeutic targets. Human PGI (hPGI) often called as AMF-PGI (autocrine motility factor-PGI), in addition to its intracellular glycolytic activity, stimulates cell migration in vitro and metastasis in vivo. Inhibition of its extracellular activity is obviously interesting in oncology. On the other hand, Candida albicans is the main yeast involved in human pathology. During recent years, resistance of this pathogenic fungus to conventional antifungal drugs appeared. Consequently, research is moving towards new therapeutic targets, including C. albicans PMI (CaPMI) that plays an important role in the biosynthesis of mannosylated structures required for pathogen survival. The reactions catalyzed by these two enzymes involve the same high energy intermediate (HEI) type 1,2-cis-enediolate, except that it is coordinated to the zinc active site in the case of PMI. Overexpression and purification of both CaPMI and hPGI were performed in our laboratory. A small chemical library was created from the synthon 5-phospho-D-arabinono-1,4-lactone (5-PAL) by modulating the head part of the HEI. A zinc binding group (ZBG) was introduced in several compounds in order to selectively inhibit the CaPMI enzyme. Moreover, two compounds with a terminal amine function were designed to selectively inhibit hPGI by targeting a glutamate residue of the enzyme (Glu357). All these molecules were first tested on rabbit muscle PGI and PMI from E. coli, and later on CaPMI and hPGI. None of these compounds are good inhibitors of CaPMI. However, a series of strong inhibitors of hPGI, and therefore potentially anti-metastatic drugs, was discovered. High-resolved 3D structures of the two enzymes complexed with inhibitors have been successfully obtained. Beyond the therapeutic, mechanistic and structural aspects, an electrochemical biosensor based on one of the synthesized inhibitors was carried out for the detection of hPGI, which is a validated biomarker of metastatic cancers. This biosensor demonstrated a detection limit of 43 fM in phosphate buffer (PBS).
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Lama Ahmad. Inhibiteurs à visée thérapeutique de la phosphomannose isomérase de Candida albicans et du facteur de motilité autocrine : études cinétiques, structurales, mécanistiques et diagnostiques. Chimie thérapeutique. Université Paris-Saclay, 2017. Français. ⟨NNT : 2017SACLS459⟩. ⟨tel-02117107v2⟩

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