Osteosarcoma (OS) is a primary malignant tumor of bone. Chemotherapy is one of the crucial approaches to prevent its metastasis and improve prognosis. Despite continuous improvements in the clinical treatment of OS, tumor resistance and metastasis remain dominant clinical challenges. Macropinocytosis, a form of non-selective nutrient endocytosis, has received increasing attention as a novel target for cancer therapy, yet its role in OS cells remains obscure. Benzethonium chloride (BZN) is an FDA-approved antiseptic and bactericide with broad-spectrum anticancer effects. Here, we described that BZN suppressed the proliferation, migration, and invasion of OS cells in vitro and in vivo, but simultaneously promoted the massive accumulation of cytoplasmic vacuoles as well. Mechanistically, BZN repressed the ERK1/2 signaling pathway, and the ERK1/2 activator partially neutralized the inhibitory effect of BZN on OS cells. Subsequently, we demonstrated that vacuoles originated from macropinocytosis and indicated that OS cells might employ macropinocytosis as a compensatory survival mechanism in response to BZN. Remarkably, macropinocytosis inhibitors enhanced the anti-OS effect of BZN in vitro and in vivo. In conclusion, our results suggest that BZN may inhibit OS cells by repressing the ERK1/2 signaling pathway and propose a potential strategy to enhance the BZN-induced inhibitory effect by suppressing macropinocytosis.

Bladder cancer (BC) is one of the most common malignant tumors in the urinary system. Due to the poor prognosis and high mortality rate of the disease, it is urgent to develop new drugs with high efficacy and low toxicity to treat BC. Echinatin (Ecn) is a bioactive natural flavonoid oflicorice that has attracted special attention for its promising anti-tumor potential. Herein, we explored the inhibitory effects of Echinatin on BC cells and probed the possible molecular mechanism. We found that Ecnin vitro inhibited the proliferation, migration, and invasion, arrested the cell cycle at the G2/M phase, and promoted apoptosis in BC cells. Besides, Ecn had no notable cytotoxicity towards human normal cells. We subsequently confirmed that Ecn restrained xenograft tumor growth and metastasis of BC cells in vivo. Mechanistically, Ecn activated the p38 signaling pathway but inactivated the Wnt/β-catenin signaling pathway, while over-expression of β-catenin and the p38 inhibitor both attenuated the inhibitory effects of Ecn on BC cells. Remarkably, Ecn combined with cisplatin (DDP) or gemcitabine (Gem) had synergistic inhibitory effects on BC cells. In summary, our results validate that Ecn inhibits the tumor growth of human BC cells via p38 and Wnt/β-catenin signaling pathways. More meaningfully, our results suggest a potential strategy to enhance DDP- or Gem-induced inhibitory effects on BC cells by combining with Ecn.