Développement de nouvelles approches pour la synthèse des flavaglines

We have developed two novel methods to prepare functionalized cyclopentenones and also discovered new acid-catalyzed rearrangements of 1-styryl propargylic alcohols. This work also explored the limitations of a method for the synthesis of flavaglines reported by Bayer researchers. Moreover, the re-synthesis of some flavaglines and the synthesis of an original fluorescence probe allowed some advances in the characterization of the mechanism of action and therapeutic potential of flavaglines for their anti-cancer effects (especially in the resistance to B6RAF inhibitors), the regulation of TRPM6 channel and the infection by Chikungunya virus.Nous avons développé deux méthodes originales de synthèse de cyclopenténones fonctionnalisées et découvert de nouveaux réarrangements d’alcools 1-styryl propargyliques catalysés par des acides.Ce travail de thèse a aussi permis de mettre en évidence les limitations d’une méthode de synthèse des flavaglines développée par des chercheurs de la compagnie Bayer. De plus, la resynthèse de certaines flavaglines et la synthèse d’une sonde de fluorescence originale ont permis de mieux caractériser le mode d’action et le potentiel thérapeutique des flavaglines pour leurs effets anticancéreux (notamment dans la résistance aux inhibiteurs de B-RAF), la régulation du canal TRPM6 et l’infection par le virus du Chikungunya

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LYON1-BU Santé Odontologie (693882213) / SudocPARIS-BIUM (751062103) / SudocSudocFranceF

Novel carbocationic rearrangements of 1-styrylpropargyl alcohols

The dehydration and subsequent cyclization reactions of 1-styrylpropargyl alcohols was examined. In the course of these studies, numerous scaffolds were synthesized, including a furan, a cyclopentenone, an acyclic enone and even a naphthalenone. The diversity of these structural motifs lies in novel cascades of reactions originating from a common carbocationic manifold

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LYON1-BU Santé Odontologie (693882213) / SudocSudocFranceF

Multiresolution image parametrization for improving texture classification

In the paper an innovative alternative to automatic image parametrization on multiple resolutions, based on texture description with specialized association rules, and image evalu- ation with machine learning methods is presented. The algorithm ArTex for parameterizing textures with association rules belonging to structural parametrization algorithms was de- veloped. In order to improve the classification accuracy a multi-resolution approach is used. The algorithm ARes for finding more informative resolutions based on the SIFT algorithm is described. The presented algorithms are evaluated on several public domains and the results are compared to other well-known parametrization algorithms belonging to statistical and spectral parametrization algorithms. Significant improvement of classification results was observed when combining parametrization attributes at several image resolutions for most parametrization algorithms. Our results show that multi-resolution image parametrization should be considered when improvement of classification accuracy in textural domains is required. These resolutions have to be selected carefully and may depend on the domain itself

Flavaglines Stimulate Transient Receptor Potential Melastatin Type 6 (TRPM6) Channel Activity.

Magnesium (Mg2+) is essential for enzymatic activity, brain function and muscle contraction. Blood Mg2+ concentrations are tightly regulated between 0.7 and 1.1 mM by Mg2+ (re)absorption in kidney and intestine. The apical entry of Mg2+ in (re)absorbing epithelial cells is mediated by the transient receptor potential melastatin type 6 (TRPM6) ion channel. Here, flavaglines are described as a novel class of stimulatory compounds for TRPM6 activity. Flavaglines are a group of natural and synthetic compounds that target the ubiquitously expressed prohibitins and thereby affect cellular signaling. By whole-cell patch clamp analyses, it was demonstrated that nanomolar concentrations of flavaglines increases TRPM6 activity by ∼2 fold. The stimulatory effects were dependent on the presence of the alpha-kinase domain of TRPM6, but did not require its phosphotransferase activity. Interestingly, it was observed that two natural occurring TRPM6 mutants with impaired insulin-sensitivity, TRPM6-p.Val1393Ile and TRPM6-p.Lys1584Glu, are not sensitive to flavagline stimulation. In conclusion, we have identified flavaglines as potent activators of TRPM6 activity. Our results suggest that flavaglines stimulate TRPM6 via the insulin receptor signaling pathway

Synthetic flavagline (FL3) binds PHB1 and PHB2 and increases PHB1 leves in H9c2 cells.

<p><b>A.</b> Whole-cell extracts of the H9c2 line (input) were either incubated with the beads Affi-Gel 10 conjugated with FL3 or blocked with ethanolamine (unconjugated Affi-Gel) [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0141826#pone.0141826.ref007" target="_blank">7</a>,<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0141826#pone.0141826.ref017" target="_blank">17</a>]. The bound and eluted proteins (Affi-Gel-FL3) and output proteins (output Affi-Gel-FL3) were analyzed by western blotting using antibodies against PHB1 and PHB2 (<i>n</i> = 3). <b>B and C.</b> Representative western blot analyses and histogram based quantification of total PHB1 levels in the cell lysates by. FL3 alone increased PHB1 protein levels within 10h as compare to non-treated cells (NT). However PHB1 level was lower in the present of both FL3 and doxorubicin (n = 3). <b>D and E.</b> Representative western blot analyses and histogram based quantification of nuclear PHB1 levels by. Doxorubicin accumulates PHB1 in the nucleus but lowers in the cytoplasm that was reduced by preconditioning with FL3. * Indicates p<0.05 as compare to control, ** indicates p<0.05 as compare to the doxorubicin alone group.</p

The mitochondrial STAT-3 phosphorylation is correlated with PHB1 translocation to mitochondria by FL3 in cardiomyocytes.

<p><b>A and B.</b> Representative western blot analyses and histogram based quantification of mitochondrial STAT3 activation by phosphorylation. STAT3 was phosphorylated by FL3 in mitochondrial fraction. <b>C and D.</b> Representative western blot analyses and <b>h</b>istogram based quantification of nuclear STAT3 activation by phosphorylation. STAT3 activation was only detected in nucleus after FL3 treatment. <b>E and F.</b> The western blot and histogram show quantitative analyses of nuclear phosphorylated STAT3 levels upon treatment of H9c2 cells with control (DMSO), FL3 (100 nM), doxorubicin (1μM), and FL3 + doxorubicin. Doxorubicin elevated phosphorylated STAT3 levels, which were reduced by FL3 (<i>n</i> = 3; *p < 0.05, compared to vehicle; **p < 0.05, compared to doxorubicin treatment).</p

The flavagline FL3 induces translocation of PHB1 to mitochondria in cardiomyocytes.

<p><b>A.</b> H9c2 cells were incubated with FL3 (100 nM) and analyzed by confocal microscopy. The cells were co-labeled with the anti-PHB1 antibody (green staining), mitotracker (red staining), and (DAPI; blue staining). The latter two dyes stained mitochondria and the nucleus, respectively. Merged confocal images show that FL3 induced the translocation of PHB1 to mitochondria (white arrows show PHB1 and mitotracker co-localization). <b>B.</b> The histogram shows quantitative analyses of co-localization of PHB1 and Mito Tracker in each cell by confocal analyses (n = 6). <b>C.</b> Representative illustration of PHB1 levels in mitochondrial and nuclear fractions upon FL3 treatment. In the mitochondrial fraction PHB1 accumulation by FL3 occurred within 20 min. PHB1 was initially increased in nucleus and rapidly reduced within 20 min. <b>D and E.</b> The histogram shows quantitative analyses of mitochondrial and nuclear PHB1 levels upon treatment of H9c2 cells with FL3 (100 nM). * Indicates p<0.05 as compare to vehicle (n = 3).</p