New evidence on the mechanisms of chaga mushroom against oral cancer

In a recent study published in the journal Scientific Reports, researchers examined the mechanisms of antitumor activity of chaga mushroom extracts on human oral cancer HSC-4 cells.

Oral cancer is a global health problem with limited treatment options due to its side effects and sequelae. The main treatments are surgery, radiation therapy and chemotherapy, although they can harm healthy tissue, affect speech and reduce quality of life.

Understanding and targeting metabolic pathways in tumor cells provides a possible avenue for the development of new therapeutic agents. Chaga mushroom has anticancer properties against several types of cancer; however, the mechanism is unclear.

In this study, the researchers tested whether chaga mushroom affects the development and metabolism of oral cancer.

After treatment with the mushroom extract, the researchers studied cell survival, proliferative capacity, glycolytic pathways, apoptosis, and mitochondrial respiration mechanisms.

They treated HSC-4 cells with the mushroom extract at doses of 0 μg/ml, 160 μg/ml, 200 μg/ml, 400 μg/ml and 800.0 μg/ml for a day to evaluate its effects on oral cancer cell behavior, including cell cycle, proliferation, viability, mitochondrial respiration, apoptosis and glycolysis.

The team analyzed the treated cells in terms of their cell cycle using cell counting kit-8 (CCK-8) assays to determine cell viability.

To investigate whether the suppressive effects of chaga mushroom on tumor proliferation and survival in treated cells involved signal transducer and activator of transcription 3 (STAT3), they measured STAT3 activation after treatment with a 200.0 μg/mL dose of the extract.

In addition, they performed flow cytometry to analyze cell distribution and Western blotting to extract total cellular proteins.

The researchers used liquid chromatography followed by tandem mass spectrometry (LC-MS) to identify the components responsible for the anti-cancer properties of chaga mushroom extract.

Concentrations of candidate compounds were determined using high-performance liquid chromatography with photodiode detector (HPLC-DAD).

They investigated the regulation of glycolysis by extracts among treated cells using extracellular acidification rate (ECAR) assay. They recorded real-time ECAR measurements in treated cells after administration of glucose, oligomycin, and 2-deoxy-D-glucose (2-DG).

The team examined the activation of an energy sensor called adenosine monophosphate-activated protein kinase (AMPK) and the cellular oxygen consumption rate (OCR).

They also assessed the effect of chronic energy deficit on autophagy associated with apoptotic cell death in treated cells.

They examined whether a concentration of 200.0 μg/mL chaga extract affected p38 mitogen-activated protein kinases (MAPKs) and nuclear factor kappa B (NF-κB)-stimulated apoptosis in treated cells.

The extract slowed the growth of HSC-4 cells by inhibiting cell cycle and proliferation, reducing the energy consumption of cancer cells, and enhancing cell death through autophagy and apoptosis.

The extract significantly increased the growth phases of oral cancer cells (G0/G1) while decreasing the synthesis phase (S). Western blot analysis revealed that the extract significantly decreased the expression of phospho-STAT3 after 15 minutes and maintained it for 120 minutes.

LC-MS identified three possible anticancer compounds: 2-hydroxy-3,4-dimethoxybenzoic acid, syringic acid and protocatechuic acid. The extract inhibited glycolysis, glycolytic capacity and glycolytic reserves in treated cells.

It also activated AMPK, promoting autophagy and inhibiting glycolytic pathways in treated cells. Induction of autophagy by the extract showed a dose-dependent increase in basal mitochondrial respiratory rates and adenosine triphosphate (ATP) turnover.

However, no significant changes in maximal mitochondrial respiratory rates were observed except at the highest extract concentration. In addition, the researchers observed a dose-dependent significant decrease in mitochondrial respiratory reserve capacity.

The results showed that chaga mushrooms reduced mitochondrial membrane potentials in treated cells through persistent autophagy mediated by inhibition of glycolysis, implying that mitochondrial dysfunction induces apoptosis.

Activation of NF-κB and p38 MAPK by the extract increased apoptosis. The extract increased early apoptosis of treated cells in a dose-dependent manner.

However, no significant differences in late apoptosis were observed at extract concentrations ranging from 0 to 400 μg/mL. High doses of chaga extract may affect other cell physiologies and reduce maximal mitochondrial respiratory capacity.

The researchers found that chaga extract suppressed mitochondrial membrane potentials and glycolytic activity in the HSC-4 cell line, resulting in reduced ATP levels and autophagy.

AMPK activation resulted in effects by inducing autophagy. STAT3 dephosphorylation inhibits the cell cycle by stimulating apoptotic pathways through activation of NF-κB and p38 MAPK.

Various cell signaling mechanisms mediated the inhibitory effects of the extract. The extract contained three anticancer compounds: 2-hydroxy-3,4-dimethoxybenzoic acid, syringic acid, and protocatechuic acid.

Although more preclinical studies are needed to determine whether the extract suppresses tumor growth, the study results suggest that the mushroom extract may be a potential adjunctive therapeutic agent for the treatment of patients with oral cancer.