BIX-01294 sensitizes renal cancer Caki cells to TRAIL-induced apoptosis through downregulation of survivin expression and upregulation of DR5 expression

Abstract

BIX-01294 (BIX), a G9a histone methyltransferase inhibitor, has been reported for its anti-proliferative and anticancer activities against various cancer cell lines. In this study, we investigated whether BIX could sensitize TRAIL-mediated apoptosis in various cancer cells. Combined treatment with BIX and TRAIL markedly induced apoptosis in human renal carcinoma (Caki, ACHN, and A498), breast carcinoma (MCF-7), and lung carcinoma (A549) cells. In contrast, BIX and TRAIL co-treatment did not induce apoptosis in normal cells, specifically mouse kidney cell (TCMK-1) and human skin fibroblast (HSF). BIX downregulated protein expression levels of XIAP and survivin at the post-translational level. Overexpression of survivin markedly blocked combined BIX and TRAIL treatment-induced apoptosis, but XIAP had no effect. Furthermore, BIX induced upregulation of DR5 expression at the transcriptional levels, and knockdown of DR5 expression using small interfering RNAs (siRNAs) markedly attenuated BIX and TRAIL-induced apoptosis. Interestingly, siRNA-mediated G9a histone methyltransferase knockdown also enhanced TRAIL-induced apoptosis in Caki cells. However, knockdown of G9a did not change expression levels of XIAP, survivin, and DR5. Therefore, BIX-mediated TRAIL sensitization was independent of histone methyltransferase G9a activity. Taken together, these results suggest that BIX facilitates TRAIL-mediated apoptosis via downregulation of survivin and upregulation of DR5 expression in renal carcinoma Caki cells.▶ BIX facilitates TRAIL-mediated apoptosis in human renal carcinoma Caki cells.▶ Downregulation of survivin contributes to BIX plus TRAIL-induced apoptosis.▶ Upregulation of DR5 is involved in BIX plus TRAIL-mediated apoptosis.▶ BIX-mediated TRAIL sensitization is independent of ROS production.

Upregulation of DR5 and Downregulation of Survivin by IITZ-01, Lysosomotropic Autophagy Inhibitor, Potentiates TRAIL-Mediated Apoptosis in Renal Cancer Cells via Ubiquitin-Proteasome Pathway

“…To isolate the total RNA, we used TriZol reagent (Life Technologies, Gaithersburg, MD, USA), and obtained cDNA using M-MLV reverse transcriptase (Gibco-BRL, Gaithersburg, MD, USA). For PCR, we used Blend Taq DNA polymerase (Toyobo, Osaka, Japan) with primers targeting DR5, Cbl, survivin and actin as mentioned in our previous studies [ 46 , 47 ]. For qPCR we utilize SYBR Fast qPCR Mix (Takara Bio Inc., Shiga, Japan) and reactions were performed on Thermal Cycler Dice ® Real Time System III (Takara Bio Inc., Shiga, Japan).…”

WP1130 Enhances TRAIL-Induced Apoptosis through USP9X-Dependent miR-708-Mediated Downregulation of c-FLIP

“…To obtain cDNA, total RNA was prepared using the TriZol reagent (Life Technologies, Gaithersburg, MD, USA), and M-MLV reverse transcriptase (Gibco-BRL, Gaithersburg, MD, USA) was used. For PCR, we used DNA Taq polymerase with primers targeting DR5, c-FLIP and actin [45,46]. The amplified products were separated by electrophoresis on a 2% agarose gel and detected under UV light.…”

Honokiol Enhances TRAIL-Mediated Apoptosis through STAMBPL1-Induced Survivin and c-FLIP Degradation

“…For PCR, we used Blend Taq DNA polymerase (Toyobo, Osaka, Japan) with primers targeting DR5, c-FLIP, survivin, and actin. The used primers were referred to previous studies [38,39]. For qPCR, SYBR Fast qPCR Mix (Takara Bio Inc., Shiga, Japan) was used, and reactions were performed on Thermal Cycler Dice ® Real Time System III (Takara Bio Inc., Shiga, Japan).…”