Yves Letourneux

Characterization of the Saccharomyces cerevisiae RTA1 gene involved in 7-aminocholesterol resistance.

Abstract 7-aminocholesterol has been described as being a strong inhibitor of yeast and of Gram+-bacteria proliferation. In order to determine the precise molecular target of the toxicity of this compound, we searched for yeast resistance linked to gene over-expression. We named the new yeast gene that was isolated RTA1 (EMBL X84736). This gene led to strong resistance to the inhibitor. Gene sequencing revealed that RTA1 is adjacent to the NAB1 gene which is orientated in an opposite direction and localized on chromosome VII. The RTA1 gene, which encodes a putative protein with seven potential membrane-spanning segments, was shown to be a non-essential gene in yeast.

Squalamine: A Polyvalent Drug of the Future?

The purpose of this mini-review is to summarize and highlight the different advances in our understanding of the antimicrobial and antiangiogenic activity of squalamine, a cationic steroid isolated in 1993 from the dogfish shark Squalus Acanthias. Indeed, squalamine has shown to be useful for the treatment of important diseases such as cancers (lung, ovarian, brain and others), age-related macular degeneration (AMD) and the control of body weight in man. All these results led to a question: could we consider squalamine as a polyvalent drug of the future?

A new conotoxin isolated from Conus consors venom acting selectively on axons and motor nerve terminals through a Na+‐dependent mechanism

A novel conotoxin was isolated and characterized from the venom of the fish‐hunting marine snail Conus consors. The peptide was identified by screening chromatography fractions of the crude venom that produced a marked contraction and extension of the caudal and dorsal fins in fish, and noticeable spontaneous contractions of isolated frog neuromuscular preparations. The peptide, named CcTX, had 30 amino acids and the following scaffold: X11CCX7CX2CXCX3C. At the frog neuromuscular junction, CcTx at nanomolar concentrations selectively increased nerve terminal excitability so that a single nerve stimulation triggered trains of repetitive or spontaneous synaptic potentials and action potentials. In contrast, CcTx had no noticeable effect on muscle excitability even at concentrations 100 × higher than those that affected motor nerve terminals, as revealed by direct muscle stimulation. In addition, CcTx increased miniature endplate potential (MEPP) frequency in a Ca2+‐free medium supplemented with ethylene glycol‐bis‐(β‐aminoethyl ether)‐N,N,N′,N′‐tetraacetic acid (EGTA). Blockade of voltage‐dependent sodium channels with tetrodotoxin (TTX) either prevented or suppressed the increase of MEPP frequency induced by the toxin. CcTx also produced a TTX‐sensitive depolarization of the nodal membrane in single myelinated axons giving rise, in some cases, to repetitive and/or spontaneous action potential discharges. In addition, CcTx increased the nodal volume of myelinated axons, as determined using confocal laser scanning microscopy. This increase was reversed by external hyperosmolar solutions and was prevented by pretreatment of axons with TTX. It is suggested that CcTx, by specifically activating neuronal voltage‐gated sodium channels at the resting membrane potential, produced Na+ entry into nerve terminals and axons without directly affecting skeletal muscle fibres. CcTx belongs to a novel family of conotoxins that targets neuronal voltage‐gated sodium channels.

Squalamine: An Appropriate Strategy against the Emergence of Multidrug Resistant Gram-Negative Bacteria?

We reported that squalamine is a membrane-active molecule that targets the membrane integrity as demonstrated by the ATP release and dye entry. In this context, its activity may depend on the membrane lipid composition. This molecule shows a preserved activity against bacterial pathogens presenting a noticeable multi-resistance phenotype against antibiotics such as polymyxin B. In this context and because of its structure, action and its relative insensitivity to efflux resistance mechanisms, we have demonstrated that squalamine appears as an alternate way to combat MDR pathogens and by pass the gap regarding the failure of new active antibacterial molecules.

Synthesis and in Vitro Study of a Diglyceride Prodrug of a Peptide

A diglyceride derivative of a pentapeptide renin inhibitor, the 1,3-dipalmitoyl-[Iva-Phe-Nle-Sta-Ala-Sta-acetyl]-glycerol was synthesized and tested in vitro as a potential prodrug for oral administration. The ability of the diglyceride analog to inhibit the renin activity was equivalent to that of the parent peptide after predigestion with pancreatic lipase. Furthermore, the presence of the palmitoyl groups was found to induce, in vitro, an efficient protection of the peptide from gastric and intestinal hydrolysis. During incubation with intestinal and gastric fluids, and with α-chymotrypsin and pancreatic lipase, the glycerolipidic derivative was more stable than the peptide alone. These results support the use of glycerolipidic prodrug for oral administration of peptides.

Cell binding, uptake and cytosolic partition of HIV anti-gag phosphodiester oligonucleotides 3'-linked to cholesterol derivatives in macrophages.

The purpose of this study is to evaluate the cell interactions of a new class of compounds composed of phosphodiester oligonucleotides linked to the cholesterol group at position 3, 7, or 22 of the steroid structure. The resulting conjugates were assessed for their capacity to bind, penetrate and partition in the cytoplasmic compartment of murine macrophages. The results showed that lipophilic conjugates bind to cells much faster (t(1/2) < or = 10 min) than do underivatized oligomers. Oligomers tethered to the cholesterol at positions 3 and 7 (PO-GEM-3-Chol and PO-GEM-7-Chol) interacted more efficiently with cell membranes and were better internalized than oligomers attached to the cholesterol moiety at position 22 (PO-GEM-22-Chol). The cytosolic fraction of internalized oligomers was studied by a digitonin-based membrane permeabilization method. The recovered fraction of oligomers that can freely diffuse from the cytosol was comparable for GEM-91, a phosphorothioate congener, and for PO-GEM-7-Chol (50-60% of the internalized oligomers), while that of PO-GEM-3-Chol was less (30% of the internalized oligomers) indicating a higher membrane affinity of the latter derivative as compared to the other investigated compounds. Membrane binding and cell internalization correlated well with the hydrophobicity of the conjugates as characterized by their partition coefficients in a water-octanol system. Due to their capacity of rapid binding and cytosolic partition in cells, cholesterol-derivatized oligonucleotides at position 3 or 7 of the steroid molecule appeared as good candidates for systemic delivery of anti-HIV antisense compounds.

Synthesis of 6(α,β)-aminocholestanols as ergosterol biosynthesis inhibitors

Abstract Δ 7 -5-Desaturase catalyses one of the last steps in ergosterol biosynthesis in fungi. Moreover Δ 5 -unsaturation is necessary for the sparking function. Synthesis of three pairs of C-6 epimeric cholestanol derivatives are described as potential growth inhibitors. Preliminary results suggest that 6β-aminocholestanol is a potent antifungal agent.

Study of Lymphotropic Targeting and Macrofilaricidal Activity of a Melphalan Prodrug on the Molinema dessetae Model

This study deals with the design of a new macrofilaricidal drug derived from melphalan and having a lymphotropism to avoid the hepatic first pass effect and enhance bioavailability after oral administration. Melphalan was linked to a ligand leading to a prodrug called 1,3-dp-melphalan which has structural analogy to triglycerides. The Molinema dessetae/Proechimys oris model was used for antiparasitic evaluation. Melphalan was macrofilaricidal in vitro against Molinema dessetae at 1mM, inactive in vivo after an oral single dose at 164 mumol/kg while the prodrug 1,3-dp-melphalan was active against adult worms after a single dose at 82 mumol/kg. After an oral administration of the prodrug to rats, the maximum concentration and the cumulated quantities of melphalan in lymph were about 45-fold higher than those observed with the free drug under the same conditions. Moreover, the plasma concentration of melphalan was 2-fold higher than those observed after the administration of the free drug. These results are in favor of lymphotropic targeting as a novel approach to develop new orally active macrofilaricides.

Antimicrobial Activities of 3-Amino- and Polyaminosterol Analogues of Squalamine and Trodusquemine

A series of 3-amino- and polyaminosterol analogues of squalamine and trodusquemine were synthesized and evaluated for their in vitro antimicrobial properties against human pathogens. The activity was highly dependent on the structure of the different compounds involved and the best results were obtained with aminosterol derivatives 4b, 4e, 8b, 8e and 8n exhibiting minimum inhibitory concentrations (MICs) against yeasts, Gram positive and Gram negative bacteria at average concentrations of 3.12–12.5 μM.

Efficient Synthesis of Various Secondary Amines Through a Titanium(IV) isopropoxide-Mediated Reductive Amination of Ketones

An efficient method for the synthesis of various secondary amines through a titanium(IV)isopropoxide-mediated reductive amination reaction of ketones is reported. Thus, a series of different ketones and amines were involved leading to the expected products in moderate to excellent yields up to 98% in numerous cases. The mechanistic rationale of this reaction has been demonstrated through 1 H and 13 C NMR experiments and occurs exclusively via the formation of a transient imine species.