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1.
Acta Crystallogr D Struct Biol ; 79(Pt 1): 1-9, 2023 Jan 01.
Artículo en Inglés | MEDLINE | ID: mdl-36601802

RESUMEN

Formation of the Aurora-A-MYCN complex increases levels of the oncogenic transcription factor MYCN in neuroblastoma cells by abrogating its degradation through the ubiquitin proteasome system. While some small-molecule inhibitors of Aurora-A were shown to destabilize MYCN, clinical trials have not been satisfactory to date. MYCN itself is considered to be `undruggable' due to its large intrinsically disordered regions. Targeting the Aurora-A-MYCN complex rather than Aurora-A or MYCN alone will open new possibilities for drug development and screening campaigns. To overcome the challenges that a ternary system composed of Aurora-A, MYCN and a small molecule entails, a covalently cross-linked construct of the Aurora-A-MYCN complex was designed, expressed and characterized, thus enabling screening and design campaigns to identify selective binders.


Asunto(s)
Neuroblastoma , Humanos , Proteína Proto-Oncogénica N-Myc/metabolismo , Proteína Proto-Oncogénica N-Myc/uso terapéutico , Neuroblastoma/tratamiento farmacológico , Neuroblastoma/metabolismo , Línea Celular Tumoral
2.
J Med Chem ; 64(15): 10682-10710, 2021 08 12.
Artículo en Inglés | MEDLINE | ID: mdl-33980013

RESUMEN

Histone H3K4 methylation serves as a post-translational hallmark of actively transcribed genes and is introduced by histone methyltransferase (HMT) and its regulatory scaffolding proteins. One of these is the WD-repeat-containing protein 5 (WDR5) that has also been associated with controlling long noncoding RNAs and transcription factors including MYC. The wide influence of dysfunctional HMT complexes and the typically upregulated MYC levels in diverse tumor types suggested WDR5 as an attractive drug target. Indeed, protein-protein interface inhibitors for two protein interaction interfaces on WDR5 have been developed. While such compounds only inhibit a subset of WDR5 interactions, chemically induced proteasomal degradation of WDR5 might represent an elegant way to target all oncogenic functions. This study presents the design, synthesis, and evaluation of two diverse WDR5 degrader series based on two WIN site binding scaffolds and shows that linker nature and length strongly influence degradation efficacy.


Asunto(s)
Antineoplásicos/farmacología , Compuestos de Bifenilo/farmacología , Dihidropiridinas/farmacología , Diseño de Fármacos , Péptidos y Proteínas de Señalización Intracelular/antagonistas & inhibidores , Antineoplásicos/síntesis química , Antineoplásicos/química , Compuestos de Bifenilo/síntesis química , Compuestos de Bifenilo/química , Células Cultivadas , Dihidropiridinas/síntesis química , Dihidropiridinas/química , Relación Dosis-Respuesta a Droga , Femenino , Humanos , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Ligandos , Masculino , Estructura Molecular , Relación Estructura-Actividad
3.
Nat Chem Biol ; 16(11): 1179-1188, 2020 11.
Artículo en Inglés | MEDLINE | ID: mdl-32989298

RESUMEN

The mitotic kinase AURORA-A is essential for cell cycle progression and is considered a priority cancer target. Although the catalytic activity of AURORA-A is essential for its mitotic function, recent reports indicate an additional non-catalytic function, which is difficult to target by conventional small molecules. We therefore developed a series of chemical degraders (PROTACs) by connecting a clinical kinase inhibitor of AURORA-A to E3 ligase-binding molecules (for example, thalidomide). One degrader induced rapid, durable and highly specific degradation of AURORA-A. In addition, we found that the degrader complex was stabilized by cooperative binding between AURORA-A and CEREBLON. Degrader-mediated AURORA-A depletion caused an S-phase defect, which is not the cell cycle effect observed upon kinase inhibition, supporting an important non-catalytic function of AURORA-A during DNA replication. AURORA-A degradation induced rampant apoptosis in cancer cell lines and thus represents a versatile starting point for developing new therapeutics to counter AURORA-A function in cancer.


Asunto(s)
Antineoplásicos/química , Aurora Quinasa A/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/química , Proteolisis/efectos de los fármacos , Talidomida/química , Ubiquitina-Proteína Ligasas/metabolismo , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Apoptosis/efectos de los fármacos , Aurora Quinasa A/genética , Benzazepinas/química , Dominio Catalítico , Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Replicación del ADN/efectos de los fármacos , Diseño de Fármacos , Femenino , Humanos , Masculino , Terapia Molecular Dirigida , Polietilenglicoles/química , Unión Proteica , Conformación Proteica
4.
Structure ; 27(8): 1195-1210.e7, 2019 08 06.
Artículo en Inglés | MEDLINE | ID: mdl-31230944

RESUMEN

Ubiquitin-conjugating enzymes (E2s) govern key aspects of ubiquitin signaling. Emerging evidence suggests that the activities of E2s are modulated by posttranslational modifications; the structural underpinnings, however, are largely unclear. Here, we unravel the structural basis and mechanistic consequences of a conserved autoubiquitination event near the catalytic center of E2s, using the human anaphase-promoting complex/cyclosome-associated UBE2S as a model system. Crystal structures we determined of the catalytic ubiquitin carrier protein domain combined with MD simulations reveal that the active-site region is malleable, which permits an adjacent ubiquitin acceptor site, Lys+5, to be ubiquitinated intramolecularly. We demonstrate by NMR that the Lys+5-linked ubiquitin inhibits UBE2S by obstructing its reloading with ubiquitin. By immunoprecipitation, quantitative mass spectrometry, and siRNA-and-rescue experiments we show that Lys+5 ubiquitination of UBE2S decreases during mitotic exit but does not influence proteasomal turnover of this E2. These findings suggest that UBE2S activity underlies inherent regulation during the cell cycle.


Asunto(s)
Lisina/metabolismo , Enzimas Ubiquitina-Conjugadoras/química , Enzimas Ubiquitina-Conjugadoras/metabolismo , Ubiquitina/metabolismo , Dominio Catalítico , Línea Celular , Cristalografía por Rayos X , Cisteína/metabolismo , Regulación de la Expresión Génica , Células HeLa , Homeostasis , Humanos , Mitosis , Simulación de Dinámica Molecular , Ubiquitinación
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