雷公藤红素通过TRAIL途径增强γδ T细胞免疫治疗骨肉瘤的机制研究

Our previous study had demonstrated that γδ T cells can be used as osteosarcoma (OS) immune effector cells, exhibiting potent lytic activity against OS cells. However, loss or downregulation of cell surface molecules expression which γδ T cells recognize was seen in OS, rendering OS resistant to γδ T cell mediated cytotoxicity. Celastrol is an active ingredient of the traditional chinese medicinal plant Tripterygium Wilfordii, which exhibits significant antitumor activity in different cancer. More importantly, celastrol up-regulates death receptor 4 (DR4) and death receptor 5(DR5) expression, sensitizing tumour cells to TRAIL/Apo-2L-induced apoptosis. However, up to now, we do not know whether celastrol could display antitumor activity in OS and enhance the susceptibility of OS cell lines to the cytotoxicity of γδ T cells through the TRAIL pathway. To explore the treatment of osteosarcoma with celastrol and γδ T cells and explore whether celastrol enhance osteosarcoma susceptibility to γδ T cell-mediated killing through the TRAIL pathway, we intend to use molecular biological methods and the establishment of animal models of OS.The anti-OS effect and the mechanism of the combination of celastrol and γδ T cells, which could be clarified, would provide a scientific basis for the immune adjuvant treatment of patients with OS.

我们前期研究证实γδT细胞可以作为骨肉瘤的免疫效应细胞，对骨肉瘤具有杀伤作用；但是发现骨肉瘤细胞表面表达的为 γδT细胞所识别的分子常常丢失，出现免疫逃逸，限制了γδT细胞治疗骨肉瘤的应用。因此，如何提高γδ T细胞对骨肉瘤细胞的识别成为目前亟需解决的问题。近来，发现来源于雷公藤根皮的雷公藤红素不仅具有广泛的抗癌作用；它还可上调肿瘤细胞表面死亡受体，致敏TRAIL途径介导的肿瘤细胞凋亡。但迄今为止，雷公藤红素是否具有抗骨肉瘤作用，及其能否通过TRAIL途径促进γδT细胞对骨肉瘤的识别和杀灭未见报道。我们拟采用分子生物学方法并建立动物模型，进行雷公藤红素联合γδ T细胞治疗骨肉瘤研究，探索雷公藤红素通过TRAIL途径增强γδ T细胞抗骨肉瘤效应的问题。雷公藤红素联合γδT细胞抗骨肉瘤细胞的效应及机制的阐明，将为骨肉瘤患者的免疫辅助治疗提供科学依据。

我们前期研究证实γδT细胞可以作为骨肉瘤的免疫效应细胞，对骨肉瘤具有杀伤作用；但是发现骨肉瘤细胞表面表达的为 γδT细胞所识别的分子常常丢失，出现免疫逃逸，限制了γδT细胞治疗骨肉瘤的应用。因此，如何提高γδ T细胞对骨肉瘤细胞的识别成为目前亟需解决的问题。近来，发现来源于雷公藤根皮的雷公藤红素不仅具有广泛的抗癌作用；它还可上调肿瘤细胞表面死亡受体，致敏TRAIL途径介导的肿瘤细胞凋亡。但迄今为止，雷公藤红素是否具有抗骨肉瘤作用，及其能否通过TRAIL途径促进γδT细胞对骨肉瘤的识别和杀灭未见报道。我们采用分子生物学方法，包括ELISA、细胞毒性试验、流式细胞技术、PCR、蛋白质印记技术、凋亡及细胞增殖试验，进行雷公藤红素联合γδ T细胞治疗骨肉瘤研究，探索雷公藤红素通过TRAIL途径增强γδ T细胞抗骨肉瘤效应的问题。结果显示雷公藤红素提高骨肉瘤细胞（HOS，U2OS）表面死亡受体DR4和DR5，通过TRAIL途径促进异体或自体γδT细胞对骨肉瘤的识别和杀灭。重要的是，小剂量雷公藤红素对γδT细胞活性没有影响。总之，我们数据表明雷公藤红素能上调骨肉瘤细胞表面死亡受体DR4和DR5，通过TRAIL/APO-2L 途径促进γδ T细胞对骨肉瘤细胞杀伤，提示雷公藤红素在骨肉瘤的临床应用价值，尤其是与γδ T细胞联合免疫治疗骨肉瘤。