Charlène Gadais

ZUSAMMENFASSUNG: G‐Protein‐gekoppelte Rezeptoren (GPCRs) sind an den meisten physiologischen Prozessen im Menschen beteiligt. Sie stellen eine der größten Proteinfamilien dar, über die eine Vielzahl zugelassener Medikamente wirken. Heterotrimere Gαβɣ‐Proteine binden über die C‐terminale α5‐Helix der Gα‐Untereinheit (GαCT) an die intrazelluläre Bindungsstelle des aktiven Rezeptors. Die Modulation der GPCR‐Aktivität über diese intrazelluläre GPCR‐Bindungsstelle stellt daher eine neue und vielversprechende Forschungsrichtung dar. In dieser Arbeit wurden hochaktive, von Gα _s CT abgeleitete Peptidomimetika entwickelt, die den β ₂ ‐Adrenorezeptor (β ₂ AR) in einer aktiven Konformation stabilisieren. Zu diesem Zweck wurde zunächst die helikale Konformation von GαsCT durch eine kovalent verankerte Brücke stabilisiert. Durch rationales Design, bei dem Informationen aus zwei Kristallstrukturen mit unterschiedlichen Bindungsmodi von Gα _s CT an β ₂ AR integriert wurden, konnte eine geeignete Position für die Verankerung der Brücke identifiziert werden. Um die Aktivität von GαsCT weiter zu verbessern, wurden die Schlüsselinteraktionen der beiden β ₂ AR‐Strukturen (β ₂ AR‐Gs(leer) und β ₂ AR‐T4L‐GsCT‐CC), nämlich Y391 und E392, durch nicht‐kanonische Aminosäuren ersetzt. Diese Optimierung führte schließlich zur Identifizierung eines potenten Peptidomimetikums, das in der Lage ist, die aktive Rezeptorkonformation zu stabilisieren und gleichzeitig die rezeptorvermittelte cAMP‐Bildung zu blockieren. Die mithilfe von Molekulardynamik‐Simulationen ermittelten Bindungsmuster des Peptidomimetikums an den Rezeptor deuten auf einen Zwischenzustand hin, der dem durch β ₂ AR‐Gs(leer) repräsentierten nukleotidfreien Zustand vorausgeht. Wir gehen davon aus, dass dieser Ansatz für die weitere strukturelle und funktionelle Erforschung anderer GPCRs oder als Werkzeug der Wirkstoffforschung von Nutzen sein wird.

G protein‐coupled receptors (GPCRs) are involved in most human physiological processes and one of the largest families of approved drug‐targeted proteins. Heterotrimeric Gαβɣ proteins bind to the intracellular cavity of the activated receptor mainly through the C‐terminal α5 helix of the Gα subunit (GαCT). Modulation of GPCR activity through intracellular GPCR binding sites is emerging. Here we develop highly active Gα _s CT‐derived peptidomimetics that stabilize the β ₂ adrenergic receptor (β ₂ AR) in an active‐like conformation when the helical conformation of Gα _s CT is preserved by a covalent tether (“staple”). By rational design, integrating the information of two crystal structures showing different binding modes of Gα _s CT with β ₂ AR, an appropriate staple position was identified. The key interactions observed in the two distinct β ₂ AR structures (β ₂ AR‐G _s (empty)) and β ₂ AR‐T4L‐G _s CT‐CC), namely Y391 and E392, were substituted with non‐canonical amino acids to improve activity. Optimization resulted in the identification of a potent peptidomimetic capable of stabilizing an active‐like receptor conformation, whilst blocking receptor‐mediated cAMP formation. Molecular dynamics simulations indicated a peptidomimetic binding mode that may represent another intermediate state preceding that of β ₂ AR‐G _s (empty). We envision this approach to be useful for further structural and functional exploration of other GPCRs or as a tool in drug discovery.

Les thiénopyrimidinones et les thiénofuranes sont considérés comme des structures prometteuses dans le design de molécules bioactives, notamment des composés anti-angiogéniques. L'angiogenèse pathologique est l'un des évènements majeurs dans le développement tumoral. Une des stratégies thérapeutiques étudiée repose donc sur la conception de petites molécules organiques susceptibles d'inhiber l'activité tyrosine kinase du récepteur VEGFR-2, un des acteurs clé du processus angiogénique. Dans ce contexte, nos travaux ont essentiellement porté sur la synthèse d'une nouvelle série de thiénopyrimidinones et thiénofuranes fonctionnalisés à potentiel inhibiteur du VEGFR-2. Pour chacune des nouvelles structures, nous avons étudié, par 3D-QSAR, l'influence sur le potentiel biologique de différentes modulations structurelles (fonctionnalisations et isomérie du noyau central, et structure d’une chaîne latérale). Puis, plusieurs stratégies de synthèse ont été élaborées et discutées afin de pouvoir préparer les molécules cibles. Pour accéder sélectivement à l'un des isomères du thiénofurane, nous avons mis au point une méthode de synthèse chimiosélective à partir de thiophènes 3-O-alkylés judicieusement fonctionnalisés. Ces composés peuvent ensuite être fonctionnalisés pour accéder aux molécules cibles. Parallèlement, la synthèse des thiéno[2,3-d]pyrimidinones a été abordée selon deux stratégies complémentaires : une voie de synthèse convergente permettant d'articuler autour du noyau central les modulations envisagées pour les cibles en série azotée (hétéroatome de jonction), et une voie de synthèse linéaire plus adéquate à la modulation des cibles en séries oxygénée et soufrée

Fungi pose a global threat to humankind due to the increasing emergence of multi-drug-resistant fungi. There is a rising incidence of invasive fungal infections. Due to the structural complexity of fungal cell membranes, only a few classes of antifungal agents are effective and have been approved by the U.S. FDA. Hence, researchers globally are focusing on developing novel strategies to cure fungal infections. One of the potential strategies is the "Trojan horse" approach, which uses the siderophore-mediated iron acquisition (SIA) system to scavenge iron to deliver potent antifungal agents for therapeutics and diagnostics. These siderophore conjugates chelate to iron and are taken up through siderophore-iron transporters, which are overexpressed exclusively on microbes such as bacteria or fungi, but not mammalian cells. Our comprehensive review delves into recent advancements in the design of siderophore-conjugated antifungal agents to gain fungal cell entry. Notably, our focus extends to unraveling the intricate relationship between the structure of natural siderophores or siderophore-like molecules and the resulting antifungal activity. By exploring these design strategies, we aim to contribute to the ongoing discourse on combating drug-resistant fungal infections and advancing the landscape of antifungal theranostics.

RAS proteins control various intracellular signaling networks. Mutations at specific locations were shown to stabilize their active guanosine triphosphate (GTP)‐bound state, which is associated with the development of multiple cancers. An attractive approach to modulate RAS signaling is through its regulatory guanine nucleotide exchange factor (GEF) son of sevenless 1 (SOS1). With the recent discovery of Nanobody14 (Nb14), which potently enhances SOS1‐catalyzed nucleotide exchange on RAS, we explored the feasibility of developing peptide mimetics by structurally mimicking the complementarity‐determining region 3 (CDR3). Guided by a biochemical GEF assay and X‐ray co‐crystal structures, successive rounds of optimization and gradual conformational rigidification led to CDR3 mimetics showing half of the maximal activation potential of Nb14 with an EC ₅₀ value of 29 μM. Altogether, this study demonstrated that peptides able to modulate a protein‐protein interaction can be obtained by structural mimicry of a Nb paratope.

RAS proteins control various intracellular signaling networks. Mutations at specific locations were shown to stabilize their active guanosine triphosphate (GTP)‐bound state, which is associated with the development of multiple cancers. An attractive approach to modulate RAS signaling is through its regulatory guanine nucleotide exchange factor (GEF) son of sevenless 1 (SOS1). With the recent discovery of Nanobody14 (Nb14), which potently enhances SOS1‐catalyzed nucleotide exchange on RAS, we explored the feasibility of developing peptide mimetics by structurally mimicking the complementarity‐determining region 3 (CDR3). Guided by a biochemical GEF assay and X‐ray co‐crystal structures, successive rounds of optimization and gradual conformational rigidification led to CDR3 mimetics showing half of the maximal activation potential of Nb14 with an EC ₅₀ value of 29 μM. Altogether, this study demonstrated that peptides able to modulate a protein‐protein interaction can be obtained by structural mimicry of a Nb paratope.

Terpene compounds probably represent the most diversified class of secondary metabolites. Some classes of terpenes, mainly diterpenes (C20) and sesterterpenes (C25) and to a lesser extent sesquiterpenes (C15), share a common bicyclo[3.6.0]undecane core which is characterized by the presence of a cyclooctane ring fused to a cyclopentane ring, i.e., a [5-8] bicyclic ring system. This review focuses on the different strategies elaborated to construct this [5-8] bicyclic ring system and their application in the total synthesis of terpenes over the last two decades. The overall approaches involve the construction of the 8-membered ring from an appropriate cyclopentane precursor. The proposed strategies include metathesis, Nozaki–Hiyama–Kishi (NHK) cyclization, Pd-mediated cyclization, radical cyclization, Pauson–Khand reaction, Lewis acid-promoted cyclization, rearrangement, cycloaddition and biocatalysis.

RAS proteins control various intracellular signaling networks. Mutations at specific locations were shown to stabilize their active GTP-bound state, which is associated with the development of multiple cancers. An attractive approach to modulate RAS signaling is through its regulatory guanine nucleotide exchange factor (GEF) son of sevenless 1 (SOS1). With the recent discovery of Nanobody14, which potently enhances SOS1-catalyzed nucleotide exchange on RAS, we explored the feasibility of developing peptide mimetics by structurally mimicking the complementarity-determining region 3 (CDR3). Guided by a biochemical GEF assay and X-ray co-crystal structures, successive rounds of optimization and gradual conformational rigidification led to CDR3 mimetics showing half of the maximal activation potential of the native nanobody. Altogether, this study provides the first proof-of-concept that peptides able to functionally modulate a protein-protein interaction can be obtained by structural mimicry of a nanobody paratope.

Opioid agonists are well-established analgesics, widely prescribed for acute but also chronic pain. However, their efficiency comes with the price of drastically impacting side effects that are inherently linked to their prolonged use. To answer these liabilities, designed multiple ligands (DMLs) offer a promising strategy by co-targeting opioid and non-opioid signaling pathways involved in nociception. Despite being intimately linked to the Substance P (SP)/neurokinin 1 (NK1) system, which is broadly examined for pain treatment, the neurokinin receptors NK2 and NK3 have so far been neglected in such DMLs. Herein, a series of newly designed opioid agonist-NK2 or -NK3 antagonists is reported. A selection of reported peptidic, pseudo-peptidic, and non-peptide neurokinin NK2 and NK3 ligands were covalently linked to the peptidic μ-opioid selective pharmacophore Dmt-DALDA (H-Dmt-d-Arg-Phe-Lys-NH₂) and the dual μ/δ opioid agonist H-Dmt-d-Arg-Aba-βAla-NH₂ (KGOP01). Opioid binding assays unequivocally demonstrated that only hybrids SBL-OPNK-5, SBL-OPNK-7 and SBL-OPNK-9, bearing the KGOP01 scaffold, conserved nanomolar range μ-opioid receptor (MOR) affinity, and slightly reduced affinity for the δ-opioid receptor (DOR). Moreover, NK binding experiments proved that compounds SBL-OPNK-5, SBL-OPNK-7, and SBL-OPNK-9 exhibited (sub)nanomolar binding affinity for NK2 and NK3, opening promising opportunities for the design of next-generation opioid hybrids.

The neurokinins are indisputably essential neurotransmitters in numerous pathoand physiological events. Being widely distributed in the Central Nervous System (CNS) and peripheral tissues, their discovery rapidly promoted them to drugs targets. As a necessity for molecular tools to understand the biological role of this class, endogenous peptides and their receptors prompted the scientific community to design ligands displaying either agonist and antagonist activity at the three main neurokinin receptors, called NK1, NK2 and NK3. Several strategies were implemented for this purpose. With a preference to small non-peptidic ligands, many research groups invested efforts in synthesizing and evaluating a wide range of scaffolds, but only the NK1 antagonist Aprepitant (EMENDT) and its prodrug Fosaprepitant (IVEMENDT) have been approved by the Food Drug Administration (FDA) for the treatment of Chemotherapy-Induced and Post-Operative Nausea and Vomiting (CINV and PONV, respectively). While non-peptidic drugs showed limitations, especially in side effect control, peptidic and pseudopeptidic compounds progressively regained attention. Various strategies were implemented to modulate affinity, selectivity and activity of the newly designed ligands. Replacement of canonical amino acids, incorporation of conformational constraints, and fusion with non-peptidic moieties gave rise to families of ligands displaying individual or dual NK1, NK2 and NK3 antagonism, that ultimately were combined with non-neurokinin ligands (such as opioids) to target enhanced biological impact.