Ethanol extract of Vanilla planifolia stems reduces PAK6 expression and induces cell death in glioblastoma cells.

Glioblastoma multiforme (GBM) is a malignant tumour with a poor prognosis. Therefore, potential treatment strategies and novel therapeutic targets have gained increased attention. Our data showed that the ethanol extract of Vanilla planifolia stem (VAS) significantly decreased the viability and the colony formation of GBM cells. Moreover, VAS induced the cleavage of MAP1LC3, a marker of autophagy. Further RNA-seq and bioinformatic analysis revealed 4248 differentially expressed genes (DEGs) between VAS-treated GBM cells and the control cells. Protein-protein interactions between DEGs with fold changes less than -3 and more than 5 were further analysed, and we found that 16 and 9 hub DEGs, respectively, were correlated with other DEGs. Further qPCR experiments confirmed that 14 hub DEGs was significantly downregulated and 9 hub DEGs was significantly upregulated. In addition, another significantly downregulated DEG, p21-activated kinase 6 (PAK6), was correlated with the overall survival of GBM patients. Further validation experiments confirmed that VAS significantly reduced the mRNA and protein expression of PAK6, which led to the abolition of cell viability and colony formation. These findings demonstrated that VAS reduced cell viability, suppressed colony formation and induced autophagy and revealed PAK6 and other DEGs as potential therapeutic targets for GBM treatment.

A Selective Histone Deacetylase Inhibitor Induces Autophagy and Cell Death via SCNN1A Downregulation in Glioblastoma Cells.

Glioblastoma multiforme (GBM) is a grade IV, highly malignant brain tumor. Because of the heterogeneity of GBM, a multitarget drug is a rational strategy for GBM treatment. Histone deacetylase inhibitors (HDACis) regulate the expression of numerous genes involved in cell death, apoptosis, and tumorigenesis. We found that the HDAC4/HDAC5 inhibitor LMK235 at 0.5 µM significantly reduced the cell viability and colony formation of patient-derived, temozolomide-resistant GBM P#5 TMZ-R, U-87 MG, and T98G cells. Moreover, LMK235 also significantly increased TUBA acetylation, which is an indicator of HDAC inhibition. Interestingly, LMK235 induced MAP1LC3 robust readout and puncta accumulation but did not enhance PARP1 cleavage or the proportion of annexin V-positive cells, suggesting that LMK235-induced cell death occurred via autophagy activation. Further RNA-seq analysis after LMK235 treatment showed that 597 different expression genes compared to control. After bioinformatic analysis by KEGG and STRING, we focused on 34 genes and validated their mRNA expression by qPCR. Further validation showed that 2 µM LMK235 significantly reduced the mRNA and protein expression of SCNN1A. Cell viability of SCNN1A-silenced cells were reduced, but cells were rescued while treated with an autophagy inhibitor bafilomycin A1. Conclusively, SCNN1A plays a role in LMK235-induced autophagy and cell death in GBM cells.