Ginsenoside 20(S)-Rg3 Inhibits the Warburg Effect Via Modulating DNMT3A/ MiR-532-3p/HK2 Pathway in Ovarian Cancer Cells

Abstract

Background/Aims: The Warburg effect is one of the main energy metabolism features supporting cancer cell growth. 20(S)-Rg3 exerts anti-tumor effect on ovarian cancer partly by inhibiting the Warburg effect. microRNAs are important regulators of the Warburg effect. However, the microRNA regulatory network mediating the anti-Warburg effect of 20(S)-Rg3 was largely unknown. Methods: microRNA deep sequencing was performed to identify the 20(S)-Rg3-influenced microRNAs in SKOV3 ovarian cancer cells. miR-532-3p was overexpressed by mimic532-3p transfection in SKOV3 and A2780 cells or inhibited by inhibitor532-3p transfection in 20(S)-Rg3-treated cells to examine the changes in HK2 and PKM2 expression, glucose consumption, lactate production and cell growth. Dual-luciferase reporter assay was conducted to verify the direct binding of miR-532-3p to HK2. The methylation status in the promoter region of pre-miR-532-3p gene was examined by methylation-specific PCR. Expression changes of key molecules controlling DNA methylation including DNMT1, DNMT3A, DNMT3B, and TET1-3 were examined in 20(S)-Rg3-treated cells. DNMT3A was overexpressed in 20(S)-Rg3-treated cells to examine its influence on miR-532-3p level, HK2 and PKM2 expression, glucose consumption and lactate production. Results: Deep sequencing results showed that 11 microRNAs were increased and 9 microRNAs were decreased by 20(S)-Rg3 in SKOV3 cells, which were verified by qPCR. More than 2-fold increase of miR-532-3p was found in 20(S)-Rg3-treated SKOV3 cells. Forced expression of miR-532-3p reduced HK2 and PKM2 expression, glucose consumption and lactate production in SKOV3 and A2780 ovarian cancer cells. Inhibition of miR-532-3p antagonized the suppressive effect of 20(S)-Rg3 on HK2 and PKM2 expression, glucose consumption and lactate production in ovarian cancer cells. Dual-luciferase reporter assay showed that miR-532-3p directly suppressed HK2 rather than PKM2. miR-532-3p level was controlled by the methylation in the promoter region of its host gene. 20(S)-Rg3 inhibited DNMT3A expression while exerted insignificant effect on DNMT1, DNMT3B and TET1-3. 20(S)-Rg3 reversed DNMT3A-mediated methylation in the promoter of the host gene of miR-532-3p, and thus elevated miR-532-3p level followed by suppression of HK2 and PKM2 expression, glucose consumption and lactate production. Conclusions: 20(S)-Rg3 modulated microRNAs to exert the anti-tumor effect in ovarian cancer. 20(S)-Rg3 lessened the DNMT3A-mediated methylation and promoted the suppression of miR-532-3p on HK2 to antagonize the Warburg effect of ovarian cancer cells.

Ginsenoside 20(S)-Rg3 Prevents PKM2-Targeting miR-324-5p from H19 Sponging to Antagonize the Warburg Effect in Ovarian Cancer Cells

“…Our previous studies have found that 20(S)-Rg3 antagonized the Warburg effect in ovarian cancer and downregulates the two key glycolytic enzymes HK2 and PKM2 [5,6]. Mechanistic studies have revealed that 20(S)-Rg3 downregulates HK2 via STAT3 [5] or DNMT3A/miR-532-3p [6] pathways. But the mechanism of PKM2 downregulation induced by 20(S)-Rg3 remains unknown.…”

“…20(S)-Rg3 exhibits anti-tumor activity in various cancers via inducing apoptosis or weakening proliferation, migration and invasion of cancer cells [18][19][20]. Our previous studies have found that 20(S)-Rg3 antagonized the Warburg effect in ovarian cancer and downregulates the two key glycolytic enzymes HK2 and PKM2 [5,6]. Mechanistic studies have revealed that 20(S)-Rg3 downregulates HK2 via STAT3 [5] or DNMT3A/miR-532-3p [6] pathways.…”

“…We proceeded to investigate the PKM2 downregulation pathway based on previously acquired microRNA sequencing data [6]. PKM2 was predicted and experimentally confirmed as the direct target of 20(S)-Rg3-upregulated miR-324-5p.…”

“…A series of study have shown that 20(S)-Rg3 significantly inhibits ovarian cancer cells growth and metastasis partly via antagonizing the Warburg effect [5,6]. The Warburg effect, also termed as aerobic glycolysis, is the main metabolic feature of cancers sustaining cancer cell growth under sufficient oxygen via rapidly generating ATP and biomaterials [7].…”

Panax ginseng C.A. Meyer (Rg3) Ameliorates Gastric Precancerous Lesions in Atp4a−/− Mice via Inhibition of Glycolysis through PI3K/AKT/miRNA‐21 Pathway

“…Previous studies have shown that Rg3 inhibits tumor cell growth, invasion, metastasis, and neovascularization [22][23][24]. Glycolytic regulation by Rg3, a potential anticancer mechanism, has been previously reported [25,26]. Pharmacological studies also showed that Rg3 suppressed gastric cancer cell growth [27][28][29].…”