Center for Cancer Research Nanobiology Program, National Cancer Institute at Frederick, National Institutes of Health, Frederick, MD 21702-1201, USA.
Exp Mol Pathol. 2010 Apr;88(2):238-49. doi: 10.1016/j.yexmp.2010.01.006. Epub 2010 Feb 1.
The CD22 antigen is a viable target for therapeutic intervention for B-cell lymphomas. Several therapeutic anti-CD22 antibodies as well as an anti-CD22-based immunotoxin (HA22) are currently under investigation in clinical settings. Coupling of anti-CD22 reagents with a nano-drug delivery vehicle is projected to significantly improve treatment efficacies. Therefore, we generated a mutant of the targeting segment of HA22 (a CD22 scFv) to increase its soluble expression (mut-HA22), and conjugated it to the surface of sonicated liposomes to generate immunoliposomes (mut-HA22-liposomes). We examined liposome binding and uptake by CD22(+) B-lymphocytes (BJAB) by using calcein and/or rhodamine PE-labeled liposomes. We also tested the effect of targeting on cellular toxicity with doxorubicin-loaded liposomes. We report that: (i) Binding of mut-HA22-liposomes to BJAB cells was significantly greater than liposomes not conjugated with mut-HA22 (control liposomes), and mut-HA22-liposomes bind to and are taken in by BJAB cells in a dose and temperature-dependent manner, respectively; (ii) This binding occurred via the interaction with the cellular CD22 as pre-incubation of the cells with mut-HA22 blocked subsequent liposome binding; (iii) Intracellular localization of mut-HA22-liposomes at 37 degrees C but not at 4 degrees C indicated that our targeted liposomes were taken up through an energy dependent process via receptor-mediated endocytosis; and (iv) Mut-HA22-liposomes loaded with doxorubicin exhibited at least 2-3 fold more accumulation of doxorubicin in BJAB cells as compared to control liposomes. Moreover, these liposomes showed at least a 2-4 fold enhanced killing of BJAB or Raji cells (CD22(+)), but not SUP-T1 cells (CD22(-)). Taken together these data suggest that these 2nd-generation liposomes may serve as promising carriers for targeted drug delivery to treat patients suffering from B-cell lymphoma.
CD22 抗原是治疗 B 细胞淋巴瘤的一个可行的治疗靶点。目前，几种治疗性抗 CD22 抗体以及一种基于抗 CD22 的免疫毒素（HA22）正在临床研究中。将抗 CD22 试剂与纳米药物递送载体偶联预计将显著提高治疗效果。因此，我们生成了 HA22（CD22 scFv）的靶向片段的突变体，以增加其可溶性表达（mut-HA22），并将其偶联到超声处理的脂质体表面，以生成免疫脂质体（mut-HA22-脂质体）。我们使用 calcein 和/或 rhodamine PE 标记的脂质体来检查 CD22(+)B 淋巴细胞（BJAB）的脂质体结合和摄取。我们还测试了用载有阿霉素的脂质体进行靶向治疗对细胞毒性的影响。我们报告：（i）mut-HA22-脂质体与 BJAB 细胞的结合明显大于未与 mut-HA22 偶联的脂质体（对照脂质体），mut-HA22-脂质体以剂量和温度依赖性方式与 BJAB 细胞结合并被其摄取；（ii）这种结合是通过与细胞 CD22 的相互作用发生的，因为细胞预先与 mut-HA22 孵育会阻止随后的脂质体结合；（iii）在 37°C 而非 4°C 下 mut-HA22-脂质体的细胞内定位表明，我们的靶向脂质体通过受体介导的内吞作用通过能量依赖的过程被摄取；（iv）与对照脂质体相比，mut-HA22 负载的脂质体在 BJAB 细胞中至少积聚了 2-3 倍的阿霉素。此外，这些脂质体对 BJAB 或 Raji 细胞（CD22(+))的杀伤作用至少增强了 2-4 倍，但对 SUP-T1 细胞（CD22(-))没有杀伤作用。总的来说，这些数据表明，这些第二代脂质体可以作为有前途的载体，用于靶向药物递送，以治疗患有 B 细胞淋巴瘤的患者。
