5-Azacytidine (Ladakamycin) is a cytidine nucleoside analog, a DNA methylation inhibitor with specificity. 5-Azacytidine regulates gene expression by decreasing the level of DNA methylation. 5-Azacytidine induces autophagy and has antitumor activity.

方法：人胰腺癌细胞 PANC-1、Capan-2、PL45 和 SU.86-86 用 5-Azacytidine (2.5-30 μM) 处理 1-5 天，使用 MTT 检测细胞活力。
结果：5-Azacytidine 显示出明显的剂量-反应效应。Capan-2 和 PANC-1 细胞对 5-Azacytidine 的耐药性最强，暴露 48 h 后 IC50 分别为 71.3 和 45.6 μM。SU.86.86 细胞对 5-Azacytidine 更敏感，48 h 的 IC50 为 19.2 μM。[1]
方法：人多发性骨髓瘤细胞 MM.1S 用 5-Azacytidine (1.25-5 μM) 处理 16-72 h，使用 Flow Cytometry 方法检测细胞凋亡情况。
结果：5-Azacytidine 治疗以剂量依赖的方式增加了 24 h 后 Annexin V 阳性的细胞比例，在 72 h 达到峰值。5-Azacytidine 诱导细胞凋亡。[2]

方法：为检测体内抗肿瘤活性，将 5-Azacytidine (3 mg/kg) 腹腔注射给携带人胰腺癌肿瘤 PANC-1 的 athymic nude 小鼠，每周六次，持续四周。
结果：肿瘤在注射 5-Azacytidine 后明显变小，肿瘤进展受到显著抑制。[1]
方法：为研究在白血病中的作用，将 5-Azacytidine (5 mg/kg) 腹腔注射给用 C1498 细胞构建白血病模型的 C57BL/6 小鼠，每天一次，持续三天。
结果：5-Azacytidine 治疗在白血病小鼠中重新建立了免疫相关转录物的表达，抑制了白血病负荷并延长了生存期。5-Azacytidine 治疗的效果是短暂的，对免疫微环境的分析揭示了耐药性的可能机制，例如免疫检查点蛋白表达的同时增加。[3]

A crude cell-free extract is isolated from LI 210 cells in culture by suspension of the cells in a given volume of 0.05mol/LTris-HCl buffer, pH 7.4, and sonic extraction with a Biosonik at 70% maximal output for 30 sec. The supernatant is collected after centrifugation at 105,000 × g for 60 min (4°C) in a Model L Spinco ultracentrifuge. The final protein concentration of the cell-free extracts is approximately 3 mg/mL. The extracts are used as the source of enzymes. Ribonucleotide reductase activity is measured. A unit of enzyme is defined as the amount that catalyzed dCMP synthesis at a rate of 1 mμmole/hr. The assay systems for the measurement of pyrimidine nucleoside (CR) and deoxynucleoside (TdR, CdR) kinases are essentially those described by Chu and Fischer. However, reactions are terminated by heating for 2 min in a boiling water bath, and the phosphorylated derivatives are isolated according to the method of Bach. Fifty-jul aliquots are applied to 1-inch discs of diethylaminoethyl paper, which are then placed in counting vials and eluted with 0.5 mL of 0.5 mol/LPCA. After 1 hr, 12 mL of Diotol are added, and the radioactivity is determined.

5 mL of L1210 cells (5 × 103 cells/mL) are incubated with Azacitidine at 37 ℃ for 3 days. Cell number is determined twice a day for 3 days by means of a Model A Coulter counter.(Only for Reference)