Blockade of STAT3 Signaling Contributes to Anticancer Effect of 5-Acetyloxy-6,7,8,4′-Tetra-Methoxyflavone, a Tangeretin Derivative, on Human Glioblastoma Multiforme Cells

Abstract

Glioblastoma multiforme (GBM) is the most aggressive primary brain tumor with poor prognosis, largely due to resistance to current radiotherapy and Temozolomide chemotherapy. The constitutive activation of Signal Transducer and Activator of Transcription 3 (STAT3) is evidenced as a pivotal driver of GBM pathogenesis and therapy resistance, and hence, is a promising GBM drug target. 5-acetyloxy-6,7,8,4′-tetramethoxyflavone (5-AcTMF) is an acetylated derivative of Tangeretin which is known to exert anticancer effects on breast, colon, lung, and multiple myeloma; however, its effect on GBM remains elusive. Herein, we reported that 5-AcTMF suppressed the viability and clonogenicity along with inducing apoptosis in multiple human GBM cell lines. Mechanistic analyses further revealed that 5-AcTMF lowered the levels of Tyrosine 705-phosphorylated STAT3 (p-STAT3), a canonical marker of STAT3 activation, but also dampened p-STAT3 upregulation elicited by Interleukin-6. Notably, ectopic expression of dominant-active STAT3 impeded 5-AcTMF-induced suppression of viability and clonogenicity plus apoptosis induction in GBM cells, confirming the prerequisite of STAT3 blockage for the inhibitory action of 5-AcTMF on GBM cell survival and growth. Additionally, 5-AcTMF impaired the activation of STAT3 upstream kinase JAK2 but also downregulated antiapoptotic BCL-2 and BCL-xL in a STAT3-dependent manner. Moreover, the overexpression of either BCL-2 or BCL-xL abrogated 5-AcTMF-mediated viability reduction and apoptosis induction in GBM cells. Collectively, we, for the first time, revealed the anticancer effect of 5-AcTMF on GBM cells, which was executed via thwarting the JAK2-STAT3-BCL-2/BCL-xL signaling axis. Our findings further implicate the therapeutic potential of 5-AcTMF for GBM treatment.

Inhibitory Effects of Tangeretin, a Citrus Peel-Derived Flavonoid, on Breast Cancer Stem Cell Formation through Suppression of Stat3 Signaling

“…The Stat3 protein is essential for the maintenance of BCSCs [4]. The acetylated derivative of tangeretin (5-AcTMF) had anticancer effects on human glioblastoma multiforme cells through blockade of Stat3 signaling [33]. Extracellular IL-8 protein is a factor for BCSCs formation [13].…”

Obatoclax, a Pan-BCL-2 Inhibitor, Downregulates Survivin to Induce Apoptosis in Human Colorectal Carcinoma Cells Via Suppressing WNT/β-catenin Signaling

“…The cytotoxic effect of Obatoclax on CRC cell lines was evaluated by measuring the viability of cells after 24 and 48 h treatment with Obatoclax using CellTiter 96 ® AQueous One Solution Cell Proliferation Assay (MTS) assay (Promega; Madison, WI, USA), as well as evaluating colony-formation capacity (clonogenicity) of Obatoclax-treated cells. Both cell viability and clonogenicity assays were performed in accordance to our established protocol [45,46].…”

“…All PCR products were first TA-cloned into the pGEM-Teasy vector (Promega; Madison, WI, USA) to verify sequence, followed by directional subcloning to the pBabe-HA vector, a pBabe.puro vector engineered to encode an in-frame N-terminal hemagglutinin (HA) epitope. These pBabe-based expression plasmids were then subjected to production of retroviral particles and subsequent infection to target cells according the established protocols in our laboratory [46].…”

“…CRC cells (4 × 10 5 ) were treated with Obatoclax (0, 100, 200 µM) for 24 h, or treated with Obatoclax for 22 h and then combined with MG132 (20 µM) for an additional 2 h. After drug treatment, cells were lysed, subjected to SDS-PAGE, blot transfer, incubation with primary and secondary antibodies, exposure to enhanced chemiluminescence (ECL) reagent, and final visualization of the antigen-of-interest using the LAS3000 system (Fujifilm; Tokyo, JPN) as stated previously [45,46]. Densitometry analysis of immunoblot signals was accomplished by ImageJ software (National Institute of Health, Bethesda, MD, USA).…”

Integrative Bioinformatics Study Reveals Tangeretin Targets and Molecular Mechanisms Against Metastatic Breast Cancer

“…Tangeretin, a citrus avonoid (Fig. 1A), may be developed as a speci c molecular-targeted anticancer agent because of its antimetastatic effects (5) on cancer cells (6)(7)(8). In speci c, this compound inhibits metastases of skin, breast, and gastric cancer cells.…”