DAPK1 overcomes vincristine resistance in osteosarcoma by inhibiting ER-phagy through the ERK signaling pathway.

Background

The treatment of osteosarcoma still relies on surgery combined with chemotherapy. However, the development of chemotherapy resistance severely weakens treatment efficacy and leads to poor prognosis. Endoplasmic reticulum‑selective autophagy (ER-phagy) can maintain intracellular homeostasis by clearing ER fragments, and has been reported to contribute to chemotherapy resistance. However, the role of ER-phagy in chemotherapy resistance in osteosarcoma remains unclear. This study aims to investigate the potential role and molecular mechanisms of death-associated protein kinase 1 (DAPK1) in vincristine (VCR) resistance in osteosarcoma.

Methods

VCR-resistant osteosarcoma cell lines, U2OS/VCR and MG63/VCR, were constructed. Effect of DAPK1 on VCR resistance was evaluated using qRT-PCR, Western blot, CCK-8, colony formation, and immunofluorescence. Furthermore, pathway activation experiments explored the involvement of the ERK pathway in the molecular mechanism of VCR resistance. Finally, a xenograft mouse model was established to validate the role of DAPK1 in vivo.

Results

DAPK1 expression was significantly reduced in VCR-resistant osteosarcoma cell lines. DAPK1 overexpression markedly suppressed MG63/VCR proliferation and enhanced its sensitivity to VCR. Mechanistically, DAPK1 overexpression downregulated p-ERK levels, thereby inhibiting ER-phagy, activating ER stress, and promoting apoptosis. Further analysis revealed that the ERK pathway activation partially reversed the inhibitory effect of DAPK1 overexpression on ER-phagy and weakened cellular sensitivity to VCR. Additionally, DAPK1 overexpression rendered MG63/VCR xenografts more sensitive to VCR treatment in vivo.

Conclusion

We conclude that DAPK1 enhances osteosarcoma cell sensitivity to VCR by inhibiting ERK-mediated ER-phagy, which may represent a promising therapeutic strategy to overcome drug resistance.