Department of Orthopedics, The Third Affiliated Hospital (Eastern Hepatobiliary Surgery Hospital), Naval Medical University (Second Military Medical University), Shanghai 201805, China.
Department of Biliary Tract Surgery, The Third Affiliated Hospital (Eastern Hepatobiliary Surgery Hospital), Naval Medical University (Second Military Medical University), Shanghai 201805, China.
Radiat Res. 2024 Nov 1;202(5):752-764. doi: 10.1667/RADE-24-00046.1.
The prognosis of osteosarcoma has not been improved for decades. As radioresistance is one of the major reasons, effective radiotherapy sensitization drugs need to be discovered. HOS and K7M2 osteosarcoma cell lines were treated with disulfiram (DSF) and radiation to assess cell viability, proliferation, migration ability, apoptosis level, ROS and Ca2+ level, and cell cycle in vitro. A HOS-derived subcutaneous tumor mouse model was constructed to evaluate tumor growth after DSF combined with radiation, and the Tunel assay and immunohistochemistry of Ki67 were conducted. Western blot was used to evaluate the protein expression level. The IC50 and working concentration of DSF in osteosarcoma cell lines were ascertained. When combined with radiation, DSF effectively suppressed cell viability, proliferation, and migration, while enhancing apoptosis in osteosarcoma cells. The cell cycle postirradiation exhibited a downward shift in the G1 phase, but the addition of DSF counteracted this trend. The combination of DSF and radiation exhibited inhibitory effects on tumor growth in vivo, which was corroborated by Ki67 staining and Tunel assay. Western blot analysis revealed that DSF upregulated the expression of P53, P21, CDKN2C, BAX, and cleaved Caspase-3 while downregulating BCL2, CDK4/6, and CyclinD1 after irradiation. Our results document that DSF exerts its radiosensitization effects in vivo and in vitro, and is a valuable radiosensitizing drug option for osteosarcoma. The radiosensitization effect is mainly achieved by activating the apoptotic pathway and promoting cell cycle arrest induced by P53/P21 and CDKN2C after irradiation.
几十年来，骨肉瘤的预后并未得到改善。由于放射抗性是主要原因之一，因此需要发现有效的放射增敏药物。用二硫化硒（DSF）和辐射处理 HOS 和 K7M2 骨肉瘤细胞系，以评估体外细胞活力、增殖、迁移能力、凋亡水平、ROS 和 Ca2+水平以及细胞周期。构建了 HOS 衍生的皮下肿瘤小鼠模型，以评估 DSF 联合辐射后肿瘤的生长情况，并进行 Tunel 测定和 Ki67 的免疫组化。采用 Western blot 评估蛋白表达水平。确定 DSF 在骨肉瘤细胞系中的 IC50 和工作浓度。当与辐射联合使用时，DSF 可有效抑制骨肉瘤细胞的活力、增殖和迁移，同时增强细胞凋亡。辐射后的细胞周期在 G1 期呈下降趋势，但添加 DSF 则抵消了这种趋势。DSF 与辐射的联合对体内肿瘤生长具有抑制作用，Ki67 染色和 Tunel 测定也证实了这一点。Western blot 分析显示，DSF 上调了 P53、P21、CDKN2C、BAX 和 cleaved Caspase-3 的表达，同时下调了 BCL2、CDK4/6 和 CyclinD1 的表达。我们的研究结果表明，DSF 在体内和体外均发挥放射增敏作用，是骨肉瘤有价值的放射增敏药物选择。放射增敏作用主要通过激活凋亡途径以及促进照射后 P53/P21 和 CDKN2C 诱导的细胞周期阻滞来实现。
