抗CD47抗体联合阿糖胞苷靶向治疗NOD/SCID小鼠单核细胞白血病的研究

Effectiveness of Anti-CD47 antibody and Ara-C combination targeting therapy NOD/SCID mouse of myeloid leukemia

摘要:

目的：探讨CD47在急性髓细胞白血病NOD/SCID小鼠模型中的预后意义及靶向治疗的最佳策略。方法：将分选的CD34+CD38-白血病干细胞（leukemia stem cells, LSCs）移植入NOD/SCID小鼠体内，建立小鼠急性单核细胞白血病模型；抗人CD47单克隆抗体单独或联合阿糖胞苷治疗白血病小鼠7~14 d，并进行疗效分析。将LSCs与小鼠巨噬细胞在含抗CD47单克隆抗体的培养液中共培养，观察CD47对巨噬细胞吞噬LSCs能力的影响。结果：THP-1细胞中存在CD34+CD38- LSCs，比例约为0.12%±0.06%，将分选后的CD34+CD38- LSCs（比例高达97.0%±1.7%）移植入NOD/SCID小鼠后成功建立白血病模型。体内实验表明，阿糖胞苷（7 d）联合抗CD47单克隆抗体（14 d）治疗后，白血病小鼠外周血和骨髓中CD33+CD45+白血病细胞下降最明显（P<0.01），生存时间明显长于其它各组。体外共培养2 h后，抗CD47单克隆抗体组吞噬指数（76.9%±12.2%）明显高于抗CD45单克隆抗体组（7.60%±2.4%，P<0.01）。结论：CD47高表达是急性髓细胞白血病的预后不良因素。阿糖胞苷联合抗CD47单克隆抗体可有效杀灭普通白血病细胞和白血病干细胞，对彻底治愈急性髓细胞白血病具有重要临床意义。

Abstract:

OBJECTIVE: To study the prognostic significance of CD47 in a NOD/SCID mouse model of acute myeloid leukemia (AML) and the best strategy for targeted therapy for this disease. METHODS: CD34+CD38- leukemia stem cells (LSCs) were separated and transplanted into NOD/SCID mice to establish a mouse model of acute monocytic leukemia (AMoL). Anti-human CD47 antibody, alone or combined with cytosine arabinoside (Ara-C), was used to treat the mice with AMoL for 7-14 days, and therapeutic efficacy was assessed. LSCs were cultured together with mouse macrophages in culture medium containing anti-CD47 or anti-CD45 monoclonal antibody for 2 hours, to observe the phagocytic ability of macrophages to LSCs. RESULTS: CD34+CD38- LSCs existed among THP-1 cells, with a content of about (0.12±0.06)%, and a mouse model of AML was successfully established after the purified CD34+CD38- LSCs (97.0%±1.7%) were transplanted into NOD/SCID mice. The in vivo experiment showed that mice with AMoL had the most significant decrease in CD33+CD45+ leukemia cells in peripheral blood and bone marrow and survived the longest after being treated with Ara-C (7 days) plus anti-CD47 monoclonal antibody (14 days) (P<0.01). After 2 hours of in vitro culture, the phagocytic index in the culture medium containing anti-CD47 monoclonal antibody was significantly higher than in the culture medium containing anti-CD45 monoclonal antibody (76.9%±12.2% vs 7.60%±2.4%; P<0.05). CONCLUSIONS: High expression of CD47 is an adverse prognostic factor in AML. Combination therapy with anti-CD47 monoclonal antibody and Ara-C can effectively eliminate leukemia cells and LSCs, demonstrating great clinical significance in curing AML.