目的:探究抗AGGF1中和抗体(RDD-Ab),RDD-Ab + 抗CTLA-4抗体联合治疗对小鼠黑色素瘤的治疗作用。方法:合成AGGF1的GTFQRDDAPASVHSE肽并制备多克隆中和抗体(RDD-Ab)。为了评估RDD-Ab对血管生成活性和黑色素瘤生长的影响,我们实施了小管生...目的:探究抗AGGF1中和抗体(RDD-Ab),RDD-Ab + 抗CTLA-4抗体联合治疗对小鼠黑色素瘤的治疗作用。方法:合成AGGF1的GTFQRDDAPASVHSE肽并制备多克隆中和抗体(RDD-Ab)。为了评估RDD-Ab对血管生成活性和黑色素瘤生长的影响,我们实施了小管生成实验、迁移实验、细胞增殖实验、黑色素瘤细胞皮下移植性模型实验和免疫组化等实验。结果:实验结果显示,制备的RDD-Ab可以识别细胞中天然的AGGF1蛋白和过表达的AGGF1蛋白。RDD-Ab可显著抑制血管内皮细胞小管形成、迁移和增殖。与IgG对照组相比,RDD-Ab治疗显著减缓黑色素瘤生长,RDD-Ab + CTLA-4抗体联合治疗时肿瘤生长速度最慢。免疫组化实验也表明,RDD-Ab显著减少瘤内微血管生成与肿瘤细胞增殖,同时,联合治疗可显著增加瘤内CD4+和CD8+淋巴细胞的浸润。结论:RDD-Ab可在体外抑制血管内皮细胞的血管新生功能,具有高效的黑色素瘤生长抑制作用,RDD-Ab + CTLA-4联合治疗黑色素瘤效果更佳(显著增加瘤内CD4+和CD8+淋巴细胞的浸润),这为未来黑色素瘤的临床干预提供了一种新的潜在治疗方案。Objective: To investigate the therapeutic effect of neutralizing antibody (against AGGF1, RDD-Ab), RDD-Ab + CTLA-4 therapy on mouse melanoma. Methods: Synthesize GTFQRDAPASVHSE peptide of AGGF1 and prepare polyclonal neutralizing antibody (RDD-Ab). In order to evaluate the effects of RDD-Ab on angiogenesis and melanoma growth, we conducted tube formation, migration, cell proliferation, subcutaneous melanoma cell transplantation model and immunohistochemistry (IHC). Results: RDD-Ab can recognize both natural AGGF1 protein and overexpressed AGGF1 protein in cells. The experimental results showed that RDD-Ab significantly inhibits the formation, migration, and proliferation of endothelial cell tubules. Compared with the IgG control group, RDD-Ab significantly slowed down the growth of melanoma. IHC experiments showed that RDD-Ab significantly suppressed tumor angiogenesis and proliferation. The combination therapy of RDD-Ab + CTLA-4 antibody has the slowest tumor growth rate, and the combination therapy increases the infiltration of CD4+ and CD8+ lymphocytes robustly in solid tumors. Conclusion: RDD-Ab can inhibit the angiogenesis function of endothelial cells in vitro and has a highly effective inhibitory effect on melanoma growth has an efficient inhibitory effect on melanoma growth, and the combined treatment with CTLA-4 antibody is more effective. This provides a new potential treatment option for clinical intervention of melanoma in the future.展开更多
背景与目的本研究旨在探索细胞毒性T淋巴细胞相关抗原(cytotoxic T lymphocyte associated antigen-4,CTLA-4)、程序坏死因子(programmed death 1,PD-1)和程序坏死因子配体(programmed death ligand 1,PD-L1)在小细胞肺癌(small cell lu...背景与目的本研究旨在探索细胞毒性T淋巴细胞相关抗原(cytotoxic T lymphocyte associated antigen-4,CTLA-4)、程序坏死因子(programmed death 1,PD-1)和程序坏死因子配体(programmed death ligand 1,PD-L1)在小细胞肺癌(small cell lung cancer,SCLC)患者外周血中的分布情况,探索其免疫作用机制并评估其作为生物标志物的临床价值。方法招募290例SCLC患者及60例健康志愿者,收集治疗前和治疗2^(nd)周期末SCLC患者EDTA抗凝血2 mL。应用流式细胞仪检测CTLA-4、PD-1和PD-L1在外周血CD3、CD4、CD8及CD25的分布,分析其与临床病理特征的相关性;采用细胞免疫化学法和流式细胞法检测PD-L1在SCLC细胞系H446中的表达。结果 SCLC患者外周血中CTLA-4^+细胞和PD-1^+细胞水平分别为(1.56±1.24)%和(8.07±3.97)%、CTLA-4在CD3细胞和CD4细胞中的表达水平无明显差异,分别为(4.87±5.18)%和(3.85±2.60)%,均低于PD-1在CD3^+或CD4^+细胞中的表达(26.63±9.04)%和(20.79±9.41)%,与健康对照组相比,SCLC中CD4^+CD25^+CTLA-4^+细胞水平明显升高(1.91±1.27)%vs(7.09±5.09)%,P<0.001;PD-1^+(CD8^+)细胞表达水平明显降低,分别为(22.56±4.21)%vs(11.47±5.85)%,P<0.001。CD4^+CD25^+CTLA-4^+细胞或CD8^+PD-1^+细胞水平与患者的年龄、性别、吸烟状况、临床分期以及是否转移等因素无关(P>0.05)。化疗两周期末CD4^+CD25^+CTLA-4^+和CD8^+PD-1^+细胞的水平对比化疗前明显下降,分别为(5.11±2.60)%vs(6.94±4.91)%;(8.74±3.39)%vs(11.48±5.91)%,P值均<0.000,1,但与无疾病进展生存和总生存无显著相关性。PD-L1高表达于SCLC细胞系H446中并定位在细胞膜和细胞浆,但在外周血中未见表达。结论本研究首次证实SCLC外周血中CTLA4高表达于调节性T细胞中,而PD-1低表达于效应性T细胞,该结果为揭示SCLC免疫监测点免疫逃逸机制提供了理论依据,可能作为一种新的无创性且可实时监测的生物标志物。展开更多
文摘目的:探究抗AGGF1中和抗体(RDD-Ab),RDD-Ab + 抗CTLA-4抗体联合治疗对小鼠黑色素瘤的治疗作用。方法:合成AGGF1的GTFQRDDAPASVHSE肽并制备多克隆中和抗体(RDD-Ab)。为了评估RDD-Ab对血管生成活性和黑色素瘤生长的影响,我们实施了小管生成实验、迁移实验、细胞增殖实验、黑色素瘤细胞皮下移植性模型实验和免疫组化等实验。结果:实验结果显示,制备的RDD-Ab可以识别细胞中天然的AGGF1蛋白和过表达的AGGF1蛋白。RDD-Ab可显著抑制血管内皮细胞小管形成、迁移和增殖。与IgG对照组相比,RDD-Ab治疗显著减缓黑色素瘤生长,RDD-Ab + CTLA-4抗体联合治疗时肿瘤生长速度最慢。免疫组化实验也表明,RDD-Ab显著减少瘤内微血管生成与肿瘤细胞增殖,同时,联合治疗可显著增加瘤内CD4+和CD8+淋巴细胞的浸润。结论:RDD-Ab可在体外抑制血管内皮细胞的血管新生功能,具有高效的黑色素瘤生长抑制作用,RDD-Ab + CTLA-4联合治疗黑色素瘤效果更佳(显著增加瘤内CD4+和CD8+淋巴细胞的浸润),这为未来黑色素瘤的临床干预提供了一种新的潜在治疗方案。Objective: To investigate the therapeutic effect of neutralizing antibody (against AGGF1, RDD-Ab), RDD-Ab + CTLA-4 therapy on mouse melanoma. Methods: Synthesize GTFQRDAPASVHSE peptide of AGGF1 and prepare polyclonal neutralizing antibody (RDD-Ab). In order to evaluate the effects of RDD-Ab on angiogenesis and melanoma growth, we conducted tube formation, migration, cell proliferation, subcutaneous melanoma cell transplantation model and immunohistochemistry (IHC). Results: RDD-Ab can recognize both natural AGGF1 protein and overexpressed AGGF1 protein in cells. The experimental results showed that RDD-Ab significantly inhibits the formation, migration, and proliferation of endothelial cell tubules. Compared with the IgG control group, RDD-Ab significantly slowed down the growth of melanoma. IHC experiments showed that RDD-Ab significantly suppressed tumor angiogenesis and proliferation. The combination therapy of RDD-Ab + CTLA-4 antibody has the slowest tumor growth rate, and the combination therapy increases the infiltration of CD4+ and CD8+ lymphocytes robustly in solid tumors. Conclusion: RDD-Ab can inhibit the angiogenesis function of endothelial cells in vitro and has a highly effective inhibitory effect on melanoma growth has an efficient inhibitory effect on melanoma growth, and the combined treatment with CTLA-4 antibody is more effective. This provides a new potential treatment option for clinical intervention of melanoma in the future.
文摘背景与目的本研究旨在探索细胞毒性T淋巴细胞相关抗原(cytotoxic T lymphocyte associated antigen-4,CTLA-4)、程序坏死因子(programmed death 1,PD-1)和程序坏死因子配体(programmed death ligand 1,PD-L1)在小细胞肺癌(small cell lung cancer,SCLC)患者外周血中的分布情况,探索其免疫作用机制并评估其作为生物标志物的临床价值。方法招募290例SCLC患者及60例健康志愿者,收集治疗前和治疗2^(nd)周期末SCLC患者EDTA抗凝血2 mL。应用流式细胞仪检测CTLA-4、PD-1和PD-L1在外周血CD3、CD4、CD8及CD25的分布,分析其与临床病理特征的相关性;采用细胞免疫化学法和流式细胞法检测PD-L1在SCLC细胞系H446中的表达。结果 SCLC患者外周血中CTLA-4^+细胞和PD-1^+细胞水平分别为(1.56±1.24)%和(8.07±3.97)%、CTLA-4在CD3细胞和CD4细胞中的表达水平无明显差异,分别为(4.87±5.18)%和(3.85±2.60)%,均低于PD-1在CD3^+或CD4^+细胞中的表达(26.63±9.04)%和(20.79±9.41)%,与健康对照组相比,SCLC中CD4^+CD25^+CTLA-4^+细胞水平明显升高(1.91±1.27)%vs(7.09±5.09)%,P<0.001;PD-1^+(CD8^+)细胞表达水平明显降低,分别为(22.56±4.21)%vs(11.47±5.85)%,P<0.001。CD4^+CD25^+CTLA-4^+细胞或CD8^+PD-1^+细胞水平与患者的年龄、性别、吸烟状况、临床分期以及是否转移等因素无关(P>0.05)。化疗两周期末CD4^+CD25^+CTLA-4^+和CD8^+PD-1^+细胞的水平对比化疗前明显下降,分别为(5.11±2.60)%vs(6.94±4.91)%;(8.74±3.39)%vs(11.48±5.91)%,P值均<0.000,1,但与无疾病进展生存和总生存无显著相关性。PD-L1高表达于SCLC细胞系H446中并定位在细胞膜和细胞浆,但在外周血中未见表达。结论本研究首次证实SCLC外周血中CTLA4高表达于调节性T细胞中,而PD-1低表达于效应性T细胞,该结果为揭示SCLC免疫监测点免疫逃逸机制提供了理论依据,可能作为一种新的无创性且可实时监测的生物标志物。
