RESUMEN
Redirecting E3 ligases to neo-substrates, leading to their proteasomal disassembly, known as targeted protein degradation (TPD), has emerged as a promising alternative to traditional, occupancy-driven pharmacology. Although the field has expanded tremendously over the past years, the choice of E3 ligases remains limited, with an almost exclusive focus on CRBN and VHL. Here, we report the discovery of novel ligands to the PRY-SPRY domain of TRIM58, a RING ligase that is specifically expressed in erythroid precursor cells. A DSF screen, followed by validation using additional biophysical methods, led to the identification of TRIM58 ligand TRIM-473. A basic SAR around the chemotype was established by utilizing a competitive binding assay employing a short FP peptide probe derived from an endogenous TRIM58 substrate. The X-ray co-crystal structure of TRIM58 in complex with TRIM-473 gave insights into the binding mode and potential exit vectors for bifunctional degrader design.
RESUMEN
Acute kidney injury (AKI) is a life-threatening disease with no known curative or preventive therapies. Data from multiple animal models and human studies have linked dysregulation of bone morphogenetic protein (BMP) signaling to AKI. Small molecules that potentiate endogenous BMP signaling should have a beneficial effect in AKI. We performed a high-throughput phenotypic screen and identified a series of FK506 analogs that act as potent BMP potentiators by sequestering FKBP12 from BMP type I receptors. We further showed that calcineurin inhibition was not required for this activity. We identified a calcineurin-sparing FK506 analog oxtFK through late-stage functionalization and structure-guided design. OxtFK demonstrated an improved safety profile in vivo relative to FK506. OxtFK stimulated BMP signaling in vitro and in vivo and protected the kidneys in an AKI mouse model, making it a promising candidate for future development as a first-in-class therapeutic for diseases with dysregulated BMP signaling.
Asunto(s)
Lesión Renal Aguda/tratamiento farmacológico , Proteínas Morfogenéticas Óseas/metabolismo , Tacrolimus/farmacología , Animales , Células Cultivadas , Modelos Animales de Enfermedad , Ensayos Analíticos de Alto Rendimiento , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Estructura Molecular , Fenotipo , Tacrolimus/análogos & derivados , Tacrolimus/químicaRESUMEN
An amendment to this paper has been published and can be accessed via a link at the top of the paper.
RESUMEN
Biosynthesis of glycosylphosphatidylinositol (GPI) is required for anchoring proteins to the plasma membrane, and is essential for the integrity of the fungal cell wall. Here, we use a reporter gene-based screen in Saccharomyces cerevisiae for the discovery of antifungal inhibitors of GPI-anchoring of proteins, and identify the oligocyclopropyl-containing natural product jawsamycin (FR-900848) as a potent hit. The compound targets the catalytic subunit Spt14 (also referred to as Gpi3) of the fungal UDP-glycosyltransferase, the first step in GPI biosynthesis, with good selectivity over the human functional homolog PIG-A. Jawsamycin displays antifungal activity in vitro against several pathogenic fungi including Mucorales, and in vivo in a mouse model of invasive pulmonary mucormycosis due to Rhyzopus delemar infection. Our results provide a starting point for the development of Spt14 inhibitors for treatment of invasive fungal infections.
Asunto(s)
Antifúngicos/farmacología , Glicosiltransferasas/antagonistas & inhibidores , Policétidos/farmacología , Proteínas de Saccharomyces cerevisiae/antagonistas & inhibidores , Animales , Proliferación Celular , Modelos Animales de Enfermedad , Fermentación , Genes Reporteros , Glicosilfosfatidilinositoles/biosíntesis , Células HCT116 , Células Hep G2 , Humanos , Concentración de Iones de Hidrógeno , Concentración 50 Inhibidora , Células K562 , Pulmón/microbiología , Masculino , Ratones , Ratones Endogámicos ICR , Mucorales , Familia de Multigenes , Rhizopus , Saccharomyces cerevisiaeRESUMEN
Malaria continues to be one of the most devastating human diseases despite many efforts to limit its spread by prevention of infection or by pharmaceutical treatment of patients. We have conducted a screen for antiplasmodial compounds by using a natural product library. Here we report on cyclomarinâ A as a potent growth inhibitor of Plasmodium falciparum and the identification of its molecular target, diadenosine triphosphate hydrolase (PfAp3Aase), by chemical proteomics. Using a biochemical assay, we could show that cyclomarinâ A is a specific inhibitor of the plasmodial enzyme but not of the closest human homologue hFHIT. Co-crystallisation experiments demonstrate a unique binding mode of the inhibitor. One molecule of cyclomarinâ A binds a dimeric PfAp3Aase and prevents the formation of the enzymeâ substrate complex. These results validate PfAp3Aase as a new drug target for the treatment of malaria. We have previously elucidated the structurally unrelated regulatory subunit ClpC1 of the ClpP protease as the molecular target of cyclomarinâ A in Mycobacterium tuberculosis. Thus, cyclomarinâ A is a rare example of a natural product with two distinct and specific modes of action.