P(HPMA-APMA)-ATRA的合成及其对HL-60细胞诱导分化能力的评估

Preparation of P(HPMA)-APMA-ATRA and Assessment of Its Induction of HL-60 Cell Differentiation

以N-（2-羟丙基）甲基丙烯酰胺（HPMA），N-（3-氨基丙基）甲基丙烯酰胺（APMA）和金反式维甲酸（ATRA）为原料，采用自由基溶液聚合法设计合成P（HPMA-APMA）．ATRA，并用核磁共振氢谱对该化合物进行结构表征．相比于单体ATRA，聚合物的水溶性显著增加，同时可通过胞吞作用进入细胞．3-（4，5-二甲基噻唑-2）-2，5-二苯基四氮唑溴盐（MTT）法评估聚合物和单体ATRA对人早幼粒白血病细胞HL-60生长的抑制作用，流式细胞术检测两者对HL-60细胞周期分布及细胞表面抗原CDllb表达的影响，进一步结合氯化硝基四氮唑蓝（NBT）还原法评估聚合物诱导HL-60细胞分化的能力．结果显示，聚合物比单体ATRA具有更强的细胞生长抑制活性，其IC_50值分别为1．03和4．09μmol／L；聚合物还具有更高的G_0／G_1期细胞阻滞效应，1．2μmol／L时，聚合物比单体ATRA的G_0／G_1期细胞率高出17．7％；同样，0．4μmol／L聚合物与2．4μmaol／L单体ATRA诱导HL-60的NBT还原能力相当，0．8μmaol／L聚合物与2．4μmol／L单体ATRA诱导HL-60细胞表面抗原CDllb表达相当，表明聚合物比单体ATRA具有更强的诱导HL-60细胞向粒细胞分化的能力，其药效增强3～4倍．

P（HPMA-APMA）-ATRA was synthesized by free-radical solution polymerization with N-（2-hydroxypropyl methacrylamide） （HPMA）, N-（3-aminopropyl） methacrylamide （APMA） and all-trans retinoic acid （ATRA） as raw materials. The structure of the polymer was confirmed by 1H NMR. Compared with the monomeric ATRA, the water-solubility of the polymer increased significantly and could enter the cells through endocytosis. The inhibition of polymerized and monomeric ATRA on HL-60 cell growth was assessed by MTT（3-（4,5-dimethylthiazol- 2-yl）-2,5-diphenyl-tetrazolium bromide）. Then flow cytometry was used to detect the effects of the two compounds on cell cycle distribution and expression of cell surface antigen CDllb in HL-60 cell. Finally, a further experiment with NBT （nitroblue tetrazolium） was conducted to analyze the cell differentiation of HL-60 induced by the two compounds. The P-ATRA showed a stronger suppressive ability on cell growth than monomeric ATRA, the IC50 was 1.03 and 4.09 μmol L-1, respectively. The P-ATRA had a higher G0/G1 phase cell block effect. At the concentration of 1.2 μmol L-1, G0/G1 phase cell rate induced by P-ATRA was 17.7% higher than that of monomeric ATRA. The NBT reduction ability of HL-60 induced by 0.4 μmol L-1 P-ATRA was similar with that of 2.4 μmol L-： ATRA, the expression of cell surface antigen CD1 lb in HL-60 cell induced by 0.8 μmol L-1 P-ATRA was similar with that of 2.4 μmol L-1 ATRA, and the results indicated that the differentiation of HL-60 cell induced by P-ATRA increased 3-4 folds compared to that of ATRA.