HSP90AA1-mediated Autophagy Promotes Drug Resistance in Osteosarcoma

Background: Osteosarcoma is the most common primary bone tumor in children and adolescents. Unfortunately, osteosarcoma treatments often fail due to the development of chemoresistance, of which the underlying molecular mechanisms still remain unclear. In this study, we demonstrated that HSP90AA1 gene is responsible for drug resistance in osteosarcoma through an autophagy-related mechanism.

Methods: shRNAs were transfected into osteosarcoma cells for knockdown of HSP90AA1 gene. Stable HSP90AA1 overexpressing osteosarcoma cell lines were obtained by lentivirus infection. mRNA and protein expressions of HSP90AA1 in osteosarcoma cells were tested by quantitative real-time PCR and western blot, respectively. Autophagy of osteosarcoma cells was detected by western blot of LC3, transmission electron microscopy and fluorescence microscope. mRFP-GFP-LC3 lentiviral transfection was also performed to detect autophagic flux. NOD/SCID mices were inoculated with MG-63 tumor cells transfected with HSP90AA1 specific shRNA. TUNEL and LC3 staining were performed to detect apoptosis and autophagy of resected tumor tissues.

Results: Doxorubicin, cisplatin, and methotrexate, which are commonly used in chemotherapy, each induced HSP90AA1 upregulation in human osteosarcoma cells. Suppression of HSP90AA1 restored the sensitivity of osteosarcoma cells to chemotherapy both in vivo and in vitro. Mechanism study indicated that autophagy is responsible for the chemoresistance in osteosarcoma cells. HSP90AA1 increased drug resistance by inducing autophagy and inhibiting apoptosis. Suppression of HSP90AA1 diminished autophagic protection in response to chemotherapy in osteosarcoma cells. Moreover, HSP90AA1 promotes autophagy through PI3K/Akt/mTOR pathway and inhibits apoptosis through JNK/P38 pathway.

Conclusion: We showed that chemotherapy agents can induce HSP90AA1 expression in osteosarcoma cells. And HSP90AA1, acting as an important regulator of autophagy, is a critical factor in the development of osteosarcoma chemoresistance both in vitro and in vivo. HSP90AA1 provides a novel therapeutic target for improving osteosarcoma treatment.