Alkali-metal-catalyzed synthesis of isoureas from alcohols and carbodiimides

International audienceCurrent synthetic methods for the formation of isoureas rely on the addition of the alcohol (R'OH) to the corresponding carbodiimide (RN=C=NR). State-of-the-art catalysts rely on transition metal and actinide complexes. Copper (CuCl, CuCl$_2$, Cu$_2$O) and zinc (ZnCl$_2$) salts are reported to act as Lewis acids , able to enhance the electrophilicity of the carbodiimide reagent. Recently, in 2016, Eisen described new actinide complexes (U[N(SiMe$_3$)$_2$]$_3$ and [(Me$_3$Si)$_2$N]2An[κ$^2$ (N,C)-CH$_2$Si(CH$_3$)$_2$N(SiMe$_3$)] (An = Th or U)) able to catalyze the formation of isoureas under mild conditions. Capitalizing on our knowledge on the chemical reactivity of guanidine bases and alkali metal like Lewis acid, we have recently designed the first alkali metal catalysts able to facilitate the addition of alcohols to carbodiimides. The role and influence of the alkali metal has been investigated by controlling the Lewis acidity of the alkali metal with exogenous ligands. Experimental studies, combined with DFT calculations, offer a new vision on the active role of alkali metal cations in catalysis

Formation of low-valent Fe0^0 and FeI^I species in Fe-catalyzed cross-coupling chemistry: key role of ate-FeII^{II} intermediates

International audienceAte-iron(lI) species such as [Ar$_3$Fe$^{II}$-(Ar = aryl) are key intermediates in Fe-catalyzed cross-coupling reactions between aryl Grignard reagents (ArMgX) and organic electrophiles.They can be active species in the catalytic cycle, or lead to Feo and Fel oxidation states. These low oxidation states were shown to be catalytically active in some cases, but they mostly lead to unwished organic byproducts. This works relates a study of the evolution of [Ar$_3$Fe$^{II}$]-complexes towards Feo and Fel oxidation states, through $^1$H NMR, EPR and $^{57}$Fe-Môssbauer spectroscopies, as weil as DFT calculations, so as to discuss the role of both steric parameters and spin states in the reduction processes. Such mechanistic insights give a betler understanding of iron-catalyzed C-C bond formation reactions, and can be exploited in the design of new ligands in order to selectively obtain a sole iron oxidation state in a catalytic process

Prions activate a p38 MAPK synaptotoxic signaling pathway.

Synaptic degeneration is one of the earliest pathological correlates of prion disease, and it is a major determinant of the progression of clinical symptoms. However, the cellular and molecular mechanisms underlying prion synaptotoxicity are poorly understood. Previously, we described an experimental system in which treatment of cultured hippocampal neurons with purified PrPSc, the infectious form of the prion protein, induces rapid retraction of dendritic spines, an effect that is entirely dependent on expression of endogenous PrPC by the target neurons. Here, we use this system to dissect pharmacologically the underlying cellular and molecular mechanisms. We show that PrPSc initiates a stepwise synaptotoxic signaling cascade that includes activation of NMDA receptors, calcium influx, stimulation of p38 MAPK and several downstream kinases, and collapse of the actin cytoskeleton within dendritic spines. Synaptic degeneration is restricted to excitatory synapses, spares presynaptic structures, and results in decrements in functional synaptic transmission. Pharmacological inhibition of any one of the steps in the signaling cascade, as well as expression of a dominant-negative form of p38 MAPK, block PrPSc-induced spine degeneration. Moreover, p38 MAPK inhibitors actually reverse the degenerative process after it has already begun. We also show that, while PrPC mediates the synaptotoxic effects of both PrPSc and the Alzheimer's Aβ peptide in this system, the two species activate distinct signaling pathways. Taken together, our results provide powerful insights into the biology of prion neurotoxicity, they identify new, druggable therapeutic targets, and they allow comparison of prion synaptotoxic pathways with those involved in other neurodegenerative diseases

Formation of low-valent Fe0^0 and FeI^I species in Fe-catalyzed cross-coupling chemistry: key role of ate-FeII^{II} intermediates

International audienceAte-iron(lI) species such as [Ar$_3$Fe$^{II}$-(Ar = aryl) are key intermediates in Fe-catalyzed cross-coupling reactions between aryl Grignard reagents (ArMgX) and organic electrophiles.They can be active species in the catalytic cycle, or lead to Feo and Fel oxidation states. These low oxidation states were shown to be catalytically active in some cases, but they mostly lead to unwished organic byproducts. This works relates a study of the evolution of [Ar$_3$Fe$^{II}$]-complexes towards Feo and Fel oxidation states, through $^1$H NMR, EPR and $^{57}$Fe-Môssbauer spectroscopies, as weil as DFT calculations, so as to discuss the role of both steric parameters and spin states in the reduction processes. Such mechanistic insights give a betler understanding of iron-catalyzed C-C bond formation reactions, and can be exploited in the design of new ligands in order to selectively obtain a sole iron oxidation state in a catalytic process

A Neuronal Culture System to Detect Prion Synaptotoxicity

<div><p>Synaptic pathology is an early feature of prion as well as other neurodegenerative diseases. Although the self-templating process by which prions propagate is well established, the mechanisms by which prions cause synaptotoxicity are poorly understood, due largely to the absence of experimentally tractable cell culture models. Here, we report that exposure of cultured hippocampal neurons to PrP^Sc, the infectious isoform of the prion protein, results in rapid retraction of dendritic spines. This effect is entirely dependent on expression of the cellular prion protein, PrP^C, by target neurons, and on the presence of a nine-amino acid, polybasic region at the N-terminus of the PrP^C molecule. Both protease-resistant and protease-sensitive forms of PrP^Sc cause dendritic loss. This system provides new insights into the mechanisms responsible for prion neurotoxicity, and it provides a platform for characterizing different pathogenic forms of PrP^Sc and testing potential therapeutic agents.</p></div

PK-digested PrP^Sc causes dendritic spine loss.

<p>(<b>A</b>) Silver stain and Western blot (using anti-PrP antibody D18) of a PrP^Sc sample and a mock-purified control sample, after digestion with PK. Lane M, molecular size markers in kDa. Hippocampal neurons from wild-type (WT) mice (<b>B, C</b>) and PrP knockout (<i>Prn-p</i>^0/0) mice (<b>D, E</b>) were treated for 24 hr with 4.4 μg/ml of purified, PK-treated PrP^Sc (<b>C, E</b>), or with an equivalent amount of mock-purified sample (<b>B, D</b>). Neurons were then fixed and stained with Alexa 488-phalloidin. Scale bar in panel E = 20 μm (applicable to panels B-D). Pooled measurements of spine number (<b>F</b>) and area (<b>G</b>) were collected from 20–24 cells from 3 independent experiments. ***p<0.001 by Student’s t-test; N.S., not significantly different.</p

Purified PrP^Sc, prepared using pronase E, causes PrP^C-dependent spine loss.

<p>(<b>A</b>) Silver stain and Western blot analysis (using anti-PrP antibody IPC1) of PrP^Sc purified from scrapie-infected brains using pronase E, and mock-purified material from uninfected brains. Lane M, molecular size markers in kDa. Hippocampal neurons from wild-type (WT) mice (<b>B, C</b>) and PrP knockout (<i>Prn-p</i>^0/0) mice (<b>D, E</b>) were treated for 24 hr with 4.4 μg/ml of purified PrP^Sc (<b>C, E</b>), or with an equivalent amount of material mock-purified from uninfected brains (<b>B, D</b>). Neurons were then fixed and stained with Alexa 488-phalloidin. Scale bar in panel E = 20 μm (applicable to panels B-D). Pooled measurements of spine number (<b>F</b>) and area (<b>G</b>) were collected from 16–18 cells from 3 independent experiments. ***p<0.001 or *p<0.05 by Student’s t-test; N.S., not significantly different.</p

Purified PrP^Sc, prepared without proteases, causes PrP^C-dependent spine loss.

<p>(<b>A</b>) Silver stain and Western blot analysis (using anti-PrP antibody D18) of PrP^Sc purified from scrapie-infected brains without proteases, and mock-purified material from uninfected brains. Lane M, molecular size markers in kDa. Hippocampal neurons from wild-type (WT) mice (<b>B, C</b>) and PrP knockout (<i>Prn-p</i>^0/0) mice (<b>D, E</b>) were treated for 24 hr with 4.4 μg/ml of purified PrP^Sc (<b>C, E</b>), or with an equivalent amount of material mock-purified from uninfected brains (<b>B, D</b>). Neurons were then fixed and stained with Alexa 488-phalloidin. Scale bar in panel E = 20 μm (applicable to panels B-D). Pooled measurements of spine number (<b>F</b>) and area (<b>G</b>) were collected from 22–25 cells from 4 independent experiments. ***p<0.001 by Student’s t-test; N.S., not significantly different.</p

The N-terminal domain of PrP^C is essential for PrP^Sc-induced dendritic spine loss.

<p>Hippocampal neurons from Tg(Δ23–111) mice (<b>A-D</b>) and Tg(Δ23–31) mice (<b>E-H</b>) (both on the <i>Prn-p</i>^0/0 background) were treated for 24 hr with 4.4 μg/ml of PrP^Sc purified without proteases (<b>B, F</b>), or with an equivalent amount of mock-purified material from uninfected brains (<b>A, E</b>). Neurons were then fixed and stained with Alexa 488-phalloidin. Scale bar in panel F = 20 μm (applicable to panels A, B, E). Pooled measurements of spine number (<b>C, G</b>) and area (<b>D, H</b>) were collected from 20–24 cells from 4 independent experiments. N.S., not significantly different by Student’s t-test.</p