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Inhibición de pequeñas moléculas independiente de la quinasa de la señalización JAK-STAT

Danny Hung-Chieh Chou¹, Amedeo Vetere, Amit Choudhary²

¹‡Department of Chemistry and Chemical Biology, Harvard University, 12 Oxford Street, Cambridge, Massachusetts 02138, United States.

Journal of the American Chemical Society

|June 5, 2015

Resumen

La pequeña molécula BRD0476 protege las células pancreáticas inhibiendo la vía JAK-STAT a través de USP9X, no suprimiendo la actividad de la JAK quinasa. Esto revela USP9X como un nuevo objetivo para regular la inflamación.

Área de la Ciencia:

Biología celular Biología celular.
Biología Molecular Biología Molecular
Farmacología Farmacología.

Sus antecedentes:

El cribado fenotípico es vital para el descubrimiento de fármacos, pero la identificación de objetivos moleculares y mecanismos de acción (MoA) para los éxitos sigue siendo un desafío.
La apoptosis de las células beta pancreáticas es un factor clave en la diabetes, y comprender su regulación es crucial para el desarrollo terapéutico.

Objetivo del estudio:

Para dilucidar el objetivo intracelular y el MOA de la pequeña molécula BRD0476, un nuevo supresor de la apoptosis de las células beta pancreáticas.
Para investigar el efecto del compuesto sobre la Janus quinasa 2 (JAK2) y el transductor de señal y la activación de la vía de señalización de la transcripción 1 (STAT1).

Principales métodos:

Análisis proteómico cuantitativo en células beta de rata para identificar el objetivo intracelular de BRD0476.6.
La interferencia de ARN (RNAi) y la edición de genes CRISPR/Cas9 para validar el objetivo.
Genética química inversa utilizando un inhibidor conocido del objetivo identificado.
Mutagénesis dirigida al sitio para investigar la interacción entre la fosforilación y la ubiquitinación.

Principales resultados:

BRD0476 inhibe la señalización JAK2 / STAT1 inducida por interferón gamma (IFN-γ), promoviendo la supervivencia de las células beta sin suprimir la actividad de la quinasa JAK.
La proteómica cuantitativa identificó la peptidasa 9X (USP9X) específica de la ubiquitina como el objetivo intracelular.
La eliminación de USP9X y la inhibición de USP9X imitaron los efectos de BRD0476.
La mutagenesis sugirió que el MOA de BRD0476 implica competencia entre la fosforilación de JAK2 y la ubiquitinación.

Conclusiones:

El cribado fenotípico junto con estudios de MOA puede producir nuevos conocimientos sobre las vías de señalización establecidas.
USP9X se identifica como un objetivo terapéutico potencial para modular la actividad de JAK2 en la inflamación celular y promover la supervivencia de las células beta.

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