Journal Article

Identification of autophagic target RAB13 with small‐molecule inhibitor in low‐grade glioma via integrated multi‐omics approaches coupled with virtual screening of traditional Chinese medicine databases

Chen, Yanmei, Liu, Bo, Jin, Wenke, Liao, Minru, Zhang, Yiwen, Zhu, Shiou, Tan, Huidan, Zhao, Rongyan, Su, Wei, He, Li

2021

Overview

Objectives Autophagy, a highly conserved lysosomal degradation process in eukaryotic cells, has been widely reported closely related to the progression of many types of human cancers, including LGG; however, the intricate relationship between autophagy and LGG remains to be clarified. Materials and methods Multi‐omics methods were used to integrate omics data to determine potential autophagy regulators in LGG. The expression of ZFP36L2 and RAB13 in SW1088 cells was experimentally manipulated using cDNAs and small interfering RNAs (siRNA). RT‐qPCR detects RNAi gene knockout and cDNA overexpression efficiency. The expression levels of proteins in SW1088 cells were evaluated using Western blot analysis and immunofluorescence analysis. Homology modelling and molecular docking were used to identify compounds from Multi‐Traditional Chinese Medicine (TCM) Databases. The apoptosis ratios were determined by flow cytometry analysis of Annexin‐V/PI double staining. We detect the number of autophagosomes by GFP‐MRFP‐LC3 plasmid transfection to verify the process of autophagy flow. Results We integrated various omics data from LGG, including EXP, MET and CNA data, with the SNF method and the LASSO algorithm, and identified ZFP36L2 and RAB13 as positive regulators of autophagy, which are closely related to the core autophagy regulators. Both transcription level and protein expression level of the four autophagy regulators, including ULK1, FIP200, ATG16L1 and ATG2B, and LC3 puncta were increased by ZFP36L2 and RAB13 overexpression. In addition, RAB13 participates in autophagy through ATG2B, FIP200, ULK1, ATG16L1 and Beclin‐1. Finally, we screened multi‐TCM databases and identified gallic acid as a novel potential RAB13 inhibitor, which was confirmed to negatively regulate autophagy as well as to induce cell death in SW1088 cells. Conclusion Our study identified the key autophagic regulators ZFP36L2 and Rab13 in LGG progression, and demonstrated that gallic acid is a small molecular inhibitor of RAB13, which negatively regulates autophagy and provides a possible small molecular medicine for the subsequent treatment of LGG. The two positive autophagy regulators ZFP36L2 and RAB13 in Low‐grade Glioma patients were discovered by SNF method and Lasso method, and it was demonstrated that gallic acid can negatively regulate autophagy by inhibiting RAB13 for future LGG therapeutics.

• Publisher: John Wiley & Sons, Inc,John Wiley and Sons Inc
• Subject: Algorithms / Apoptosis / Autophagy / Cancer therapies / cancer therapy / Cell death / Cell Line, Tumor / Chemotherapy / Cytotoxicity / Databases, Factual / DNA methylation / Drug Screening Assays, Antitumor / Flow cytometry / Gallic acid / Gene expression / Generalized linear models / Glioma / Glioma - drug therapy / Glioma - enzymology / Herbal medicine / Homology / Humans / Immunofluorescence / Inhibitors / Kinases / Low‐grade gliomas (LGG) / Lysosomes / Medical prognosis / Medicine, Chinese Traditional / Molecular docking / Molecular Docking Simulation / Original / Phagocytosis / Phagosomes / Proteins / rab GTP-Binding Proteins - antagonists & inhibitors / rab GTP-Binding Proteins - metabolism / RAB13 / Radiation therapy / RNA-mediated interference / siRNA / small‐molecule inhibitor / Traditional Chinese medicine / Traditional Chinese Medicine (TCM) / Transcription / Transfection / Tumors / ZFP36L2