Vectorisation de complexes d'iridium (III) par des analogues de sidérophores : une stratégie de cheval de Troie utilisant la lumière contre les bactéries pathogènes

Therapeutic alternatives must be developed to cope with the increasing bacterial resistance to antibiotics. The use of light and photodynamic therapy is still in its infancy in the field of infections treatment, particularly using Ir(III)-based organometallic photosensitizers. In order to limit the toxicity of such molecules for host cells, the vectorization of these complexes by siderophores can increase antibacterial efficacy and cell selectivity. Siderophores are chelators synthesized by the bacteria to meet their iron needs and can serve as vectors in antibiotic Trojan horse strategies. We reported here the first conjugates between a siderophore, the desferrioxamine, and an Ir(III) complex, and showed that these compounds exhibited a potent antibacterial activity, thus validating our approach from a chemical point of view. The extension of this concept to catechol vectors mimicking the siderophore enterobactin has been attempted but the synthesis of the expected conjugates remains to be finalized so far. Finally, experimental serendipity has made it possible to discover a new reactivity and to carry out a methodological study on the mechanism of hydrolysis of the dipyridyl-amine acetamide (Dpaa) function and the use of this pattern as a protective group of the amine function.Face à l’augmentation des résistances bactériennes aux antibiotiques, des alternatives thérapeutiques doivent être développées. L’utilisation de la lumière au travers de la thérapie photodynamique n’en est qu’à ses balbutiements dans le domaine du traitement des infections, en particulier avec l’utilisation de photosensibilisateurs organométalliques à base d’Ir(III). Afin de pallier la toxicité de telles molécules pour les cellules de l’hôte, la vectorisation de ces complexes par des sidérophores peut permettre d’augmenter l’efficacité et la sélectivité cellulaire. Les sidérophores sont des chélateurs synthétisés par la bactérie pour subvenir à ses besoins en fer et qui peuvent servir de vecteurs dans le cadre d’une stratégie de type cheval de Troie. Nous avons décrit les premiers conjugués entre un sidérophore, la desferrioxamine, et un complexe d’Ir(III), et montré que ces composés présentaient une activité antibactérienne affirmée, validant ainsi notre approche du point de vue chimique. L’extension de ce concept à des vecteurs catécholés mimant le sidérophore entérobactine a été tentée et la synthèse des conjugués attendus reste à finaliser. Enfin, les aléas expérimentaux ont permis de mettre en évidence une nouvelle réactivité et de procéder à une étude méthodologique sur le mécanisme d’hydrolyse de la fonction dipyridyl-amine acétamide (Dpaa) et l’utilisation de ce motif comme groupe protecteur de la fonction amine

Vectorization of iridium(III) complexes using siderophores surrogates : a Trojan horse strategy using light against pathogenic bacteria

Face à l’augmentation des résistances bactériennes aux antibiotiques, des alternatives thérapeutiques doivent être développées. L’utilisation de la lumière au travers de la thérapie photodynamique n’en est qu’à ses balbutiements dans le domaine du traitement des infections, en particulier avec l’utilisation de photosensibilisateurs organométalliques à base d’Ir(III). Afin de pallier la toxicité de telles molécules pour les cellules de l’hôte, la vectorisation de ces complexes par des sidérophores peut permettre d’augmenter l’efficacité et la sélectivité cellulaire. Les sidérophores sont des chélateurs synthétisés par la bactérie pour subvenir à ses besoins en fer et qui peuvent servir de vecteurs dans le cadre d’une stratégie de type cheval de Troie. Nous avons décrit les premiers conjugués entre un sidérophore, la desferrioxamine, et un complexe d’Ir(III), et montré que ces composés présentaient une activité antibactérienne affirmée, validant ainsi notre approche du point de vue chimique. L’extension de ce concept à des vecteurs catécholés mimant le sidérophore entérobactine a été tentée et la synthèse des conjugués attendus reste à finaliser. Enfin, les aléas expérimentaux ont permis de mettre en évidence une nouvelle réactivité et de procéder à une étude méthodologique sur le mécanisme d’hydrolyse de la fonction dipyridyl-amine acétamide (Dpaa) et l’utilisation de ce motif comme groupe protecteur de la fonction amine.Therapeutic alternatives must be developed to cope with the increasing bacterial resistance to antibiotics. The use of light and photodynamic therapy is still in its infancy in the field of infections treatment, particularly using Ir(III)-based organometallic photosensitizers. In order to limit the toxicity of such molecules for host cells, the vectorization of these complexes by siderophores can increase antibacterial efficacy and cell selectivity. Siderophores are chelators synthesized by the bacteria to meet their iron needs and can serve as vectors in antibiotic Trojan horse strategies. We reported here the first conjugates between a siderophore, the desferrioxamine, and an Ir(III) complex, and showed that these compounds exhibited a potent antibacterial activity, thus validating our approach from a chemical point of view. The extension of this concept to catechol vectors mimicking the siderophore enterobactin has been attempted but the synthesis of the expected conjugates remains to be finalized so far. Finally, experimental serendipity has made it possible to discover a new reactivity and to carry out a methodological study on the mechanism of hydrolysis of the dipyridyl-amine acetamide (Dpaa) function and the use of this pattern as a protective group of the amine function

Dipyridylamine-acetamide (Dpaa): A primary amine protecting group orthogonally cleavable under acidic conditions in the presence of t-butyloxycarbonyl (Boc) and t-butylester

International audienceWe show that dipyridylamine-acetamide (Dpaa), can be cleaved under mild acidic conditions (30% formic acid in dichloromethane). The release of the amine function is orthogonal to other acid-labile protecting groups. Calculations suggest that the ease of Dpaa cleavage relies on activation of the carbonyl function by the protonated dipyridylamine nitrogen and activation of a water molecule by a hydrogen-bond network

2‐thiophenyl‐isoquinoline Ir(III) complex as red absorbing photosensitizer: a promising tool in antipseudomonal photodynamic therapy

International audienceInnovative therapeutic strategies are more than ever needed to counter the rise of antibiotic‐resistant bacterial pathogens worldwide. The use of light, and especially photodynamic therapy (PDT) appears as a promising alternative or complement to antibiotic treatments, fostered by the development of new photosensitizers. In this study, eight luminescent Ir(III) complexes were synthesized and evaluated for their photoactivation properties and capacity to generate radical species under blue (452 nm), green (525 nm), and red (631 nm) LED light, respectively. Their antibacterial properties were assessed on Pseudomonas aeruginosa, Acinetobacter baumannii, Escherichia coli, and Staphylococcus aureus with most of these complexes exhibiting potentially useful activities upon light irradiation, at concentrations below 10 mg/L. A complex of Ir(III) cyclometallated to thiophenyl‐isoquinoline (tiq) and bearing 2,2’‐bipyridine (bipy) as ancillary ligand was further investigated. This latter showed a concentration‐ and light intensity‐dependent bactericidal activity on P. aeruginosa when irradiated under blue to red lights, proving that such complexes would be suitable candidates for PDT. Importantly, this lead complex remained active against antibiotic resistant clinical strains and was unaffected by active efflux systems. These data open interesting perspectives for the development of new treatments to tackle antibiotic resistant Gram‐negative bacteria

Synthesis and antibacterial properties under blue LED light of conjugates between the siderophore desferrioxamine B (DFOB) and an Iridium(III) complex

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Skewed X-chromosome inactivation drives the proportion of DNAAF6-defective airway motile cilia and variable expressivity in primary ciliary dyskinesia

International audienceBackground. Primary ciliary dyskinesia (PCD) is a rare airway disorder caused by defective motile cilia. Only male patients have been reported with pathogenic mutations in X-linked DNAAF6, which result in the absence of ciliary dynein arms, whereas their heterozygous mothers are supposedly healthy. Our objective was to assess the possible clinical and ciliary consequences of X-chromosome inactivation (XCI) in these mothers. Methods. XCI patterns of 6 mothers of male patients with DNAAF6-related PCD were determined by DNA-methylation studies and compared to their clinical phenotype (6/6 mothers), as well as their ciliary phenotype (4/6 mothers), as assessed by immunofluorescence and high-speed videomicroscopy analyses. The mutated X chromosome was tracked to assess the percentage of cells with a normal inactivated DNAAF6 allele. Results. The mothers’ phenotypes ranged from absence of symptoms to mild/moderate or severe airway phenotypes, closely reflecting their XCI pattern. Analyses of the symptomatic mothers’ airway ciliated cells revealed the coexistence of normal cells and cells with immotile cilia lacking dynein arms, whose ratio closely mirrored their XCI pattern. Conclusion. This study highlights the importance of searching for heterozygous pathogenic DNAAF6 mutations in all female relatives of male PCD patients with a DNAAF6 defect, as well as in females consulting for mild chronic respiratory symptoms. Our results also demonstrate that about one third - ranging from 20-50% - normal ciliated airway cells sufficed to avoid severe PCD, a result paving the way for gene therapy

Skewed X-chromosome inactivation drives the proportion of DNAAF6 -defective airway motile cilia and variable expressivity in primary ciliary dyskinesia

International audienceBackground Primary ciliary dyskinesia (PCD) is a rare airway disorder caused by defective motile cilia. Only male patients have been reported with pathogenic mutations in X-linked DNAAF6 , which result in the absence of ciliary dynein arms, whereas their heterozygous mothers are supposedly healthy. Our objective was to assess the possible clinical and ciliary consequences of X-chromosome inactivation (XCI) in these mothers. Methods XCI patterns of six mothers of male patients with DNAAF6 -related PCD were determined by DNA-methylation studies and compared with their clinical phenotype (6/6 mothers), as well as their ciliary phenotype (4/6 mothers), as assessed by immunofluorescence and high-speed videomicroscopy analyses. The mutated X chromosome was tracked to assess the percentage of cells with a normal inactivated DNAAF6 allele. Results The mothers’ phenotypes ranged from absence of symptoms to mild/moderate or severe airway phenotypes, closely reflecting their XCI pattern. Analyses of the symptomatic mothers’ airway ciliated cells revealed the coexistence of normal cells and cells with immotile cilia lacking dynein arms, whose ratio closely mirrored their XCI pattern. Conclusion This study highlights the importance of searching for heterozygous pathogenic DNAAF6 mutations in all female relatives of male PCD patients with a DNAAF6 defect, as well as in females consulting for mild chronic respiratory symptoms. Our results also demonstrate that about one-third—ranging from 20% to 50%—normal ciliated airway cells sufficed to avoid severe PCD, a result paving the way for gene therapy

TTC12 Loss-of-Function Mutations Cause Primary Ciliary Dyskinesia and Unveil Distinct Dynein Assembly Mechanisms in Motile Cilia Versus Flagella

International audienceCilia and flagella are evolutionarily conserved organelles whose motility relies on the outer and inner dynein arm complexes (ODAs and IDAs). Defects in ODAs and IDAs result in primary ciliary dyskinesia (PCD), a disease characterized by recurrent airway infections and male infertility. PCD mutations in assembly factors have been shown to cause a combined ODA-IDA defect, affecting both cilia and flagella. We identified four loss-of-function mutations in TTC12, which encodes a cytoplasmic protein, in four independent families in which affected individuals displayed a peculiar PCD phenotype characterized by the absence of ODAs and IDAs in sperm flagella, contrasting with the absence of only IDAs in respiratory cilia. Analyses of both primary cells from individuals carrying TTC12 mutations and human differentiated airway cells invalidated for TTC12 by a CRISPR-Cas9 approach revealed an IDA defect restricted to a subset of single-headed IDAs that are different in flagella and cilia, whereas TTC12 depletion in the ciliate Paramecium tetraurelia recapitulated the sperm phenotype. Overall, our study, which identifies TTC12 as a gene involved in PCD, unveils distinct dynein assembly mechanisms in human motile cilia versus flagella