RESUMEN

Inositol-requiring enzyme 1 alpha (IRE1α) is a transmembrane sensor protein with both kinase and ribonuclease activity, which plays a crucial role in the unfolded protein response (UPR). Protein misfolding in the endoplasmic reticulum (ER) lumen triggers dimerization and subsequent trans-autophosphorylation of IRE1α. This leads to the activation of its endoribonuclease (RNase) domain and splicing of the mRNA of the transcriptional activator XBP1, ultimately generating an active XBP1 (XBP1s) implicated in multiple myeloma survival. Previously, we have identified human IRE1α as a target for the development of kinase inhibitors that could modulate the UPR in human cells, which has particular relevance for multiple myeloma and other secretory malignancies. Here we describe the development and validation of a 384-well high-throughput screening assay using DELFIA technology that is specific for IRE1α autophosphorylation. Using this format, a focused library of 2312 potential kinase inhibitors was screened, and several novel IRE1α kinase inhibitor scaffolds were identified that could potentially be developed toward new therapies to treat multiple myeloma.

Asunto(s)

Endorribonucleasas/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Animales , Línea Celular , Endorribonucleasas/química , Endorribonucleasas/metabolismo , Ensayos Analíticos de Alto Rendimiento/métodos , Humanos , Insectos , Proteínas de la Membrana/antagonistas & inhibidores , Proteínas de la Membrana/metabolismo , Mieloma Múltiple/tratamiento farmacológico , Mieloma Múltiple/enzimología , Mieloma Múltiple/metabolismo , Fosforilación/efectos de los fármacos , Pliegue de Proteína/efectos de los fármacos , Proteínas Serina-Treonina Quinasas/química , Proteínas Serina-Treonina Quinasas/metabolismo , Respuesta de Proteína Desplegada/efectos de los fármacos