High-Throughput Screens To Identify Autophagy Inducers That Function by Disrupting Beclin 1/Bcl-2 Binding
| Autor: | Yongjie Wei, José Carlos Rodríguez Pérez, Jenna B. Yehl, Matthew J. Ranaghan, Beth Levine, Zhongju Zou, Bruce A. Posner, Yi Chun Kuo, Adam Skepner, Shuguang Wei, Joshua A. Bittker, Wei Chung Chiang, Anwu Zhou |
|---|---|
| Rok vydání: | 2018 |
| Předmět: |
0301 basic medicine
Allosteric modulator Cell Survival Drug Evaluation Preclinical Cellular homeostasis Plasma protein binding Biochemistry Article 03 medical and health sciences Autophagy medicine Humans Protein Interaction Maps Cytotoxicity Chemistry Kinase Neurodegeneration General Medicine medicine.disease In vitro High-Throughput Screening Assays Cell biology 030104 developmental biology Proto-Oncogene Proteins c-bcl-2 Molecular Medicine Beclin-1 HeLa Cells Protein Binding |
| Zdroj: | ACS Chemical Biology. 13:2247-2260 |
| ISSN: | 1554-8937 1554-8929 |
| DOI: | 10.1021/acschembio.8b00421 |
| Popis: | Autophagy, a lysosomal degradation pathway, plays a crucial role in cellular homeostasis, development, immunity, tumor suppression, metabolism, prevention of neurodegeneration, and lifespan extension. Thus, pharmacological stimulation of autophagy may be an effective approach for preventing or treating certain human diseases and/or aging. We sought to establish a method for developing new chemical compounds that specifically induce autophagy. To do this, we developed two assays to identify compounds that target a key regulatory node of autophagy induction-specifically, the binding of Bcl-2 (a negative regulator of autophagy) to Beclin 1 (an allosteric modulator of the Beclin 1/VPS34 lipid kinase complex that functions in autophagy initiation). These assays use either a split-luciferase assay to measure Beclin 1/Bcl-2 binding in cells or an AlphaLISA assay to directly measure direct Beclin 1/Bcl-2 binding in vitro. We screened two different chemical compound libraries, comprising ∼300 K compounds, to identify small molecules that disrupt Beclin 1/Bcl-2 binding and induce autophagy. Three novel compounds were identified that directly inhibit Beclin 1/Bcl-2 interaction with an IC50 in the micromolar range and increase autophagic flux. These compounds do not demonstrate significant cytotoxicity, and they exert selectivity for disruption of Bcl-2 binding to the BH3 domain of Beclin 1 compared with the BH3 domain of the pro-apoptotic Bcl-2 family members, Bax and Bim. Thus, we have identified candidate molecules that serve as lead templates for developing potent and selective Beclin 1/Bcl-2 inhibitors that may be clinically useful as autophagy-inducing agents. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|