微环DNA表达抗EpCAM/抗CD3双特异BiTE分子的抗卵巢癌细胞体外实验研究

Experimental study on anti-ovarian carcinoma effect of minicircle DNA expressing anti-EpCAM/anti-CD3 bispecific BiTE in vitro

目的探讨抗EpCAM/抗CD3 BsAb分子蛋白对体外卵巢癌细胞生长的抑制效果。方法采用非病毒载体微环DNA表达抗EpCAM/抗CD3 BsAb分子蛋白。观察不同浓度(1×10^-3、1×10^-4、1×10^-5、1×10^-6、1×10^-7、1×10^-8μg/mL)抗EpCAM/抗CD3 BsAb分子蛋白加至卵巢癌细胞及T细胞中,经6 h和12 h培育后,利用倒置荧光显微镜观察活细胞与死亡细胞数量及形态变化。结果倒置显微镜下可见加抗体后卵巢癌细胞与T细胞明显聚集成块,周围见大量死亡细胞。相同时间下,随着浓度的增加,T细胞介导的卵巢癌肿瘤细胞的杀伤效果并不会一直增强。当浓度达到1×10^-4μg/mL时,培育6 h和12 h时的杀伤均达到最大,差异均有统计学意义(P<0.05)。结论新型的微环DNA表达系统所表达的抗EpCAM/抗CD3 BsAb对体外卵巢癌细胞生长具有抑制作用,未来可作为一种有效的抗卵巢癌肿瘤的靶向治疗药物。

Objective To study the effect of anti-EpCAM/anti-CD3 BsAb protein inhibition on ovarian cancer cells in vitro.Methods Non-viral vector microcircle DNA was used to obtain the anti-EpCAM/anti-CD3 BsAb protein.Different concentrations(1×10^-3,1×10^-4,1×10^-5,1×10^-6,1×10^-7,1×10^-8μg/mL)of anti-EpCAM/anti-CD3 BsAb protein was added to ovarian cancer cells and T cells.After 6 h and 12 h,the number of live cells and dead cells were counted and morphological changes were observed by inverted fluorescence microscopy.Results Under the inverted microscope,ovarian cancer cells and T cells were obviously aggregated after the addition of anti-EpCAM/anti-CD3 BsAb,and a large number of dead cells were found around them.At the same time,the killing effect of ovarian cancer cells mediated by T cells did not always increased with the increase of concentration.When the concentration reached 1×10^-4μg/mL,the killing rate reached the maximum in both 6 h and 12 h time periods,and the difference were both statistically significant(P<0.05).Conclusion The anti-EpCAM/anti-CD3 BsAb expressed by the novel microcircle DNA expression system can inhibit the growth of ovarian cancer cells in vitro,and it may be used as an effective targeted anti-ovarian cancer tumor in the future.