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Comprehensive Study of the Fungal Endophyte Community of the Long-lived Amazonian Palm <i>Astrocaryum Sciophilum</i>: A Model for Deciphering Plant-microbe Interactions at Chemical and Bioactive Levels
No full textIl est désormais établi que les plantes ne sont pas des entités distinctes, mais qu'elles ont évolué en association avec des assemblages microbiens très complexes et diversifiés. Ces communautés de symbiotes microbiens des plantes (microbiote) et leur matériel génétique collectif constituent le microbiome végétal. Le microbiome semble jouer des rôles clés dans l'écologie et la physiologie des plantes, y compris leur croissance et survie. Notre intérêt se porte sur certains des habitants fongiques du microbiome: les champignons endophytes.
Dans cette thèse, réalisée dans le cadre du projet franco-suisse SECIL (Study of Endophytes Communities In a Leaf, SECIL_ANR), nous avons voulu étudier le crosstalk microbien dans les endophytes fongiques des feuilles de plantes au niveau chimique et bioactif, en recherchant des métabolites biologiquement pertinents. Le palmier typique de la forêt primaire de Guyane française, Astrocaryum sciophilum (Miq.) Pulle a été choisi pour une étude approfondie en tant que modèle d'hôte, en raison de son cycle de vie particulièrement long et de la diversité des communautés endophytes trouvées dans ses feuilles. Son long âge de maturation suggère que ce palmier pourrait maintenir une association stable avec les communautés microbiennes sur une période de temps substantielle. Cela implique que les endophytes potentiels sont capables de survivre et de résister à l'environnement de la plante, probablement en développant des anti- pathogènes et des métabolites cytotoxiques impliqués dans la longévité des pousses et des feuilles du palmier. De plus, l'écosystème hautement compétitif de la forêt tropicale rend la pression de sélection intense, incitant probablement les microbes associés à cet hôte tropical à produire des entités chimiques intéressantes. Nous avons également émis l'hypothèse que des interactions chimiques réciproques se produisent entre la plante et les champignons, via une chimie spécialisée potentiellement exprimée dans les tissus de l'hôte. Nous avons donc développé une stratégie pour étudier, à un moment donné, cette interaction plante-champignon d'un point de vue moléculaire, de la manière la plus complète possible, ainsi que le potentiel bioactif qui résulte de cette interaction.
Le workflow que nous avons développé dans cette thèse vise à prendre un apercu métabolomique de la feuille hôte et de sa communauté fongique endophyte et à étudier les interactions moléculaires et les interactions chimiques réciproques entre eux. Nous avons ensuite exploité, lorsque c'était possible, les métabolites spécialisés de la communauté fongique pour leur bioactivité antivirulentes ou anticancéreuses. Il a permis plus particulièrement de caractériser le métabolome de la communauté fongique et celui de la plante hôte. L’analyse poussée de ces données révèle des interactions jamais analysées auparavant de manière aussi détaillée et non ciblée au niveau d'un modèle hôte-feuille. Notre stratégie a également conduit à l'identification de 24 nouveaux métabolites secondaires d'une souche de Lasiodiplodia venezuelensis. Certains d'entre eux ont montré une activité intéressante d'inhibition de quorum sensing, avec un effet sur les gènes régulés par le quorum sensing et certains facteurs de virulence, et un composé mineur a montré une inhibition significative et spécifique de l'activité Wnt et de la prolifération de lignées cellulaires de cancer du sein triple négatif.It is now established that plants are not distinct entities, but have evolved in association with highly complex and diverse microbial assemblages. These plant microbial symbiont communities (microbiota) and their collective genetic material constitute the plant microbiome, thought to play key roles in plant ecology and physiology, including plant fitness (growth and survival). Our interest is turned towards some of the fungal inhabitants of the microbiome: endophytic fungi. Endophytes colonize the healthy internal tissues of plants in an asymptomatic way and have been extensively studied in the last decades for their ability to produce bioactive molecules as part of a defense response against pathogens for the host or for the fungi themselves.
In this thesis, carried out in the scope of the Franco-Swiss project SECIL (Study of Endophyte Communities In a Leaf, SECIL_ANR), we aimed to study microbial crosstalk in plant leaf fungal endophytes at the chemical and bioactive level, searching for biologically relevant metabolites.
The typical palm tree from the primary forest in French Guiana, Astrocaryum sciophilum (Miq.) Pulle was chosen for deep investigations as a host model, due to its remarkable long-life cycle and the diversity of endophytic communities found in its leaves. Its long maturation age suggests that this palm might maintain a stable association with microbial communities over a substantial period of time. This implies that potential endophytes are able to survive and resist the environment of the plant, likely by developing antipathogens and cytotoxic metabolites involved in the longevity of the shoots and leaves of the palm. Moreover, the highly competitive rainforest ecosystem makes the selection pressure intense, probably prompting the microbes associated with this tropical host to produce interesting chemical entities. We also hypothesized that reciprocal chemical interactions occur between plant and fungi, via specialised chemistry potentially expressed in host tissues. We thus developed a strategy to study at a given time, this plant-fungi interaction from a molecular point-of-view, in the most comprehensive way possible, along with the bioactive potential which results from the interaction.
The workflow we developed in this thesis aims to take a metabolomic snapshot of the host leaf and its fungal endophytic community and study molecular interplay and reciprocal chemical interactions between them. We further exploited when possible specialised metabolites of the fungal community for their bioactivity (antivirulence or anticancer).
It allowed more particularly to characterise the metabolome of the fungal community and that of the host plant. The in-depth analysis of these data revealed interactions never before analysed in such a detailed and untargeted manner at the level of a host-leaf model. Our workflow also led to the identification of 24 new secondary metabolites of a Lasiodiplodia venezuelensis strain. Some of which showed interesting quorum sensing inhibition activity, with effect on the quorum sensing- regulated genes and some virulence factors, a minor compound showed significant and specific inhibition of Wnt activity and of the proliferation of Triple Negative Breast Cancer cell lines.</p
Synthesis and Biological Evaluation of Electrophilic Fragments as Covalent Selective Blockers of TNFR1-DD
No full textDeregulation of the Tumor Necrosis Factor (TNF) pathway is responsible for the pathological onset of various autoimmune disorders and the perpetuation of chronic inflammation that adversely affects more than 50 million people worldwide. The current standard of care for a broad spectrum of chronic inflammatory conditions consists of using TNF blockers that directly prevent TNFα binding with both TNF receptor 1 (TNFR1) and TNF receptor 2 (TNFR2). However, total TNF signaling inhibition often leads to severe side effects, mainly caused by the impairment of TNFR2-mediated homeostatic signaling. Indeed, achieving the selective inhibition of TNFR1 over TNFR2 would address a challenging, unmet medical need in the field of autoimmunity by mitigating the risk associated with non-selective anti-TNF therapies.
My Ph.D project aims at exploring new pharmacological approaches that would exploit the intracellular structural difference between TNFR isoforms, namely the presence of a Death Domain (DD) in TNFR1, to mediate its selective inactivation and/or degradation. This new mechanism of action would prevent the pathological overstimulation of the NF-κB-mediated pathway leading to chronic inflammatory conditions while sparing TNFR2-mediated protective response to avoid undesired off-target effects. No therapeutic agent is currently reported as a selective antagonist of the TNFR1-DD and, considering the lack of information on the presence of potential binding hot spots within this particular region, TNFR1-DD targeting could not be addressed via rational inhibitor drug design. Indeed, we chose to employ electrophilic fragments as covalent probes for the biochemical investigation of TNFR1-DD activity, in the perspective of discovering new druggable sites within unexplored cryptic or shallow pockets.
By combining multiple binding affinity techniques for the screening of an in-house fragment library, with organic chemistry synthesis of structural analogs and their in-cell functional characterization (including proximity-based assays, protein NMR and ad-hoc investigational tools of TNFR1-pathway specific components), we discovered low molecular weight molecules (MW<400 Da) able to destabilize TNFR1-DD in a dose-dependent fashion. Further, mutagenesis studies included in this work shed light on the chemical space around the identified TNFR1-DD ligandable site to investigate the Structure-Activity Relationship (SAR) of our hits and their putative Mechanism of Action (MoA). Eventually, this would allow a better understanding of the complex TNFR1 cell-internalization mechanism and provide the fundamental basis for the design of more refined DD ligands following a hit-to-lead optimization workflow.</p
A System Approach for the Analysis of Natural Hazard Processes in Mountainous Areas
No full textMountainous regions cover about 24% of the global land area and are a living space for about 10 % of the world's population. They also play an important economic role (e.g. tourism). Mountainous regions are susceptible to natural hazard processes and are severely affected by climate change impacts. This poses major challenges for mountain areas when it comes to securing these areas as a sustainable living space. Detailed knowledge and assessment of natural hazard processes is therefore key when it comes to sustaining livelihoods in mountain regions, This PhD thesis addresses four research objectives and aims to overcome existing challenges in natural hazard research in complex mountain environments. With a comprehensive compilation of process chains from all over the world a basis for characterization and classification was established. A methodology based on fuzzy logic was developed to assess the climate sensitivity of natural hazard processes. The actual state and the further development of permafrost degradation is simulated, using a numerical model. Finally, the potential of system dynamics is being examined to get a better understanding of complex system behavior of natural hazard processes
Gestion de la constipation induite par les opioïdes : les anciens et les modernes
No full textThe use of opioids has increased over the past 20 years. Among their side effects, constipation is probably the most common. Managing opioid-induced constipation primarily involves laxatives from the onset and addressing any additional causes of constipation. The laxatives of choice for this condition are, firstly, macrogol, followed by stimulant laxatives. If these first-line measures fail, peripheral mu-opioid receptor agonists (PAMORA) can be considered ; however, their coverage by Swiss health insurances remains limited to date.L’utilisation des opioïdes a augmenté durant ces vingt dernières années. Parmi leurs effets secondaires, la constipation est probablement le plus courant. La gestion de la constipation induite par les opioïdes repose principalement sur l’administration de laxatifs dès leur introduction et le contrôle de causes surajoutées de constipation. Les laxatifs de choix dans cette indication sont en premier lieu le macrogol puis les laxatifs stimulants. En cas d’échec de ces mesures et traitements, des agonistes périphériques des récepteurs mu (PAMORA) peuvent être discutés, bien que leur prise en charge par les assurances sociales reste à ce jour limitée
Les atteintes ORL de l’infection congénitale à cytomégalovirus
No full textLe cytomégalovirus est la cause la plus fréquente d’infection congénitale dans le monde. 90 % des enfants infectés in utero naissent sans symptôme, mais 15 % d’entre eux vont développer des atteintes au cours des cinq premières années de vie. Les plus fréquentes touchent l’oreille interne, engendrant une surdité neurosensorielle et/ou une dysfonction vestibulaire (DV). La DV est souvent méconnue et confondue avec des atteintes du système nerveux central. Elle peut provoquer des retards du développement psychomoteur et prédisposer à un retard global du développement. Un diagnostic et une prise en charge précoces sont essentiels pour prévenir ou limiter ces séquelles. Un traitement antiviral en période pré et néonatale doit être considéré.Cytomegalovirus is the most common cause of congenital infection worldwide. 90 % of children infected in utero are born without symptoms, but 15 % of them will develop disorders within the first five years of life. The most common disorders affect the inner ear, resulting in sensorineural hearing loss and/or vestibular dysfunction (VD). VD is often unrecognized and confused with conditions -affecting the central nervous system. It can cause delays in psychomotor development and predispose to overall developmental delay. Early diagnosis and treatment are essential to prevent or limit these sequelae. Antiviral treatment during the pre- and neonatal periods should be considered
Exploring transition of care beyond hospital walls: patient perspectives, interprofessional medication management, and community pharmacists' clinical reasoning
No full textBackground: Transitioning care from hospital to outpatient settings is a critical and high-risk period for patients. It also presents substantial challenges for healthcare professionals, who frequently operate in fragmented and uncoordinated systems. Medication safety may be jeopardised, with issues such as medication reconciliation errors, drug-related problems and medication nonadherence leading to potential hospital readmissions. Interprofessional collaborative interventions across care settings have shown promising results by reducing readmissions and enhancing integrated care for patients transitioning from hospital to home. Still, these services are rarely implemented and sustained in standard care. Community pharmacists play a key role in the interprofessional team during hospital discharge; however, their practices and roles in outpatient care at hospital discharge remain insufficiently defined.
Objective: This thesis aimed to explore the perspectives, experiences, and needs of healthcare professionals and polypharmacy patients with long-term conditions during the post-hospitalisation care transition, particularly emphasising outpatient care, interprofessional collaboration, and medication management. Specifically, the ItPaDiab study explored the perspectives of discharged polypharmacy patients regarding their medications and their care journey from hospital to two months after discharge. The FGinterpro study focused on identifying and describing the current state and areas for improvement of interprofessional collaboration in medication management during the transition from hospital to outpatient care. The CPreasoning study aimed to evaluate community pharmacists’ practices and clinical reasoning regarding hospital discharge prescriptions.
Methods: This thesis employed a range of qualitative methodologies to explore care transition and interprofessional and professional practices. The ItPaDiab study focused on patients with type 2 diabetes and at least two long-term conditions discharged home, using four longitudinal semi-structured interviews per participant and self-reported questionnaires over the two-month period following discharge. The FGInterpro study was conducted through interprofessional serial focus groups with patients and healthcare professionals from both hospital and outpatient settings and analysed through the Canadian Interprofessional Health Collaborative (CIHC) Framework. The CPreasoning study used simulations to explore community pharmacists’ practices and retrospective think-aloud methodologies to better understand their clinical reasoning. The clinical reasoning was analysed using the Charlin et al.'s model and the encounter structure through the Calgary-Cambridge model. All studi</p
Rational Design of Thermosensitive Drug Delivery Systems for Biologics - a Roadmap Towards Industrialization
No full textThermosensitive polymers such poly(N-isopropylacrylamide) (PNIPAM) and poly(2-alkyl-2-oxazoline)s (PAOxs) have been under investigation as drug delivery systems for over three decades but did not reach clinical phases. The challenges impeding their regulatory approval include difficulties in translating experimental research into pharmaceutical development, vague regulatory guidelines, incomplete biocompatibility studies, and the absence of defined critical quality attributes (CQAs) outlined for parenteral administration. In our dissertation, we defined that as in situ forming depot PNIPAM- and PAOx-based drug delivery systems classify as implantable medical devices. When coupled with biotherapeutics, they can stabilize the active pharmaceutical ingredient and supply extended-release features. Generally, this combination needs a type 5 new drug application, and specific subset studies are needed depending on the physical and chemical nature of the polymer-protein loading. The scientific literature highlights thermosensitive PAOx as favored compared to PNIPAM for biomedical applications. This due to the antifouling properties, synthetic options, and the wide range of chemical designs which distinct PAOx from PNIPAM. Nonetheless, until differently proven, both express good biocompatibility and excretion by renal pathway for polymers below 50 kDa.
To evaluate the developability of thermosensitive injectables, it is essential to precisely determine their lower critical solution temperatures (LCSTs). We established a standardized threshold for LCST detection by harmonizing and cross-validating orthogonal methods, employing techniques such as nuclear magnetic resonance spectroscopy, light scattering, turbidimetry, background membrane imaging, and rheology. These techniques aligned and revealed unique features and potential biases in the characterization of thermosensitive polymers. Hyaluronic acid-poly(N-isopropylacrylamide) copolymers were synthesized using strain-promoted azide-alkyne cycloaddition (SPAAC) to clarify their critical material attributes and set the fundaments for a modulable drug product design. We assessed the structure-function relationships and behaviors of these copolymers under in vitro conditions simulating in vivo environments.
We later tested the universal applicability of the developed analytical workflow with an optimized subset of techniques. Also, the developed modular drug product approach was evaluated as a platform technology strategy. The aim was to guide the chemical design of thermosensitive, in situ forming injectables according to the intended target product profile. Self-assembly, mechanical properties, physical state, and thermal transition behavior were evaluated using nuclear magnetic resonance, oscillatory rheology, turbidimetry, and visual inspection. Both PNIPAM and PAOxs with LCSTs below body temperature were studied, both before and after grafting onto azido-substituted hyaluronic acid (HA) via SPAAC. We identified critical material attributes crucial for advancing the pharmaceutical development of in situ gelling thermosensitive polymers.
Our final investigation assessed the use of PNIPAM and PAOx derivatives, conjugated to azido-substituted hyaluronic acid copolymers (HA) through SPAAC, as thermosensitive, in situ form.</p
[Compte rendu de :] Les livres des Chroniques / Philippe Abadie. - Paris : Cerf, 2022. - (Mon ABC de la Bible)
No full textTraitements médicamenteux séquentiels de l’ostéoporose
No full textOsteoporosis is a chronic disease requiring lifelong treatment, both non-pharmacological and pharmacological. If life expectancy is long enough, drug treatment must be sequential and adapted to the risk of fracture and to comorbidities. With other treatments than bisphosphonates, sequences are mandatory. When denosumab is stopped, bisphosphonates reduce the rebound effect and limit the risk of vertebral fractures. Osteoanabolic treatments are used for 12 (romosozumab) to 24 months (teriparatide). Their anti-fracture benefits can be maintained with subsequent antiresorptive treatment (bisphosphonates, denosumab), which is an integral part of the management of patients pre-treated with an osteoanabolic agent. Regular monitoring is necessary to assess whether drug treatment should be interrupted, continued, or extended. The aim is to preserve bone health into old age and prevent fractures.L’ostéoporose nécessite une prise en charge tout au long de la vie. Si l’espérance de vie est longue, le traitement médicamenteux doit être séquentiel, adapté au risque fracturaire et aux comorbidités. Avec plusieurs traitements, des séquences sont obligatoires. À l’arrêt du dénosumab, les bisphosphonates atténuent l’effet rebond et limitent le risque de fractures vertébrales. Les traitements anaboliques osseux sont utilisés pour 12 (romosozumab) à 24 mois (tériparatide). Leur bénéfice antifracturaire peut être maintenu avec un traitement antirésorbeur consécutif qui fait partie intégrante de la prise en charge des patients prétraités par un anabolique osseux. Un suivi régulier est nécessaire pour adapter le traitement (interrompu, poursuivi ou élargi). Le but est de préserver la santé osseuse jusqu’à un âge avancé et prévenir les fractures