School of Pharmacy, Graduate School of Health, University of Technology, Sydney, NSW 2007, Australia; Vascular Immunology Unit, Discipline of Pathology, Sydney Medical School and Bosch Institute, The University of Sydney, NSW 2006, Australia.
Eur J Pharmacol. 2013 Dec 5;721(1-3):116-25. doi: 10.1016/j.ejphar.2013.09.044. Epub 2013 Oct 3.
Expanding on our previous findings demonstrating that microparticles (MPs) spread cancer multidrug resistance, we now show that MPs sequester drugs, reducing the free drug concentration available to cells. MPs were isolated from drug-sensitive and drug-resistant sub-clones of a human breast adenocarcinoma cell line and from human acute lymphoblastic leukemia cells. MPs were assessed for size, mitochondria, RNA and phospholipid content, P-glycoprotein (P-gp) expression and orientation and ATPase activity relative to drug sequestration capacity. Of the drug classes examined, MPs sequestered the anthracycline class to a significant degree. The degree of sequestration was likely due to the size of MPs and thus the amount of cargo they contain, to which the anthracyclines bind. Moreover, a proportion of the P-gp present on MPs was inside-out in orientation, enabling it to influx drugs rather than its typical efflux function. This was confirmed by surface immunofluorescence and by assessment of drug-stimulated ATPase activity following MP permeabilization. Thus we determined that breast cancer MPs carried a proportion of their P-gp oriented inside-out, providing active sequestration within the microvesicular compartment. These results demonstrate a capacity for MPs to sequester chemotherapeutic drugs, which has a predominantly active sequestration component for MPs derived from drug-resistant cells and a predominantly passive component for MPs derived from drug-sensitive cells. This reduction in available drug concentration has potential to contribute to a parallel pathway and complements that of the intercellular transfer of P-gp. These findings lend further support to the role of MPs in limiting the successful management of cancer.
基于我们之前的研究发现，微粒（MPs）会传播癌症多药耐药性，现在我们表明 MPs 会隔离药物，从而降低细胞中可用的游离药物浓度。我们从人乳腺癌腺癌细胞系的药物敏感和耐药亚克隆以及人急性淋巴细胞白血病细胞中分离出 MPs。我们评估了 MPs 的大小、线粒体、RNA 和磷脂含量、P-糖蛋白（P-gp）表达和定向以及相对于药物隔离能力的 ATP 酶活性。在所检查的药物类别中，MPs 显著隔离了蒽环类药物。这种隔离程度可能归因于 MPs 的大小以及它们所包含的货物数量，而蒽环类药物与货物结合。此外，MPs 上存在一部分 P-gp 呈外翻取向，使其能够摄取药物，而不是其典型的外排功能。这通过表面免疫荧光和评估 MPs 通透后药物刺激的 ATP 酶活性得到了证实。因此，我们确定乳腺癌 MPs 携带一部分 P-gp 呈外翻取向，在微囊泡隔室中提供主动隔离。这些结果表明 MPs 具有隔离化疗药物的能力，对于源自耐药细胞的 MPs，主要是主动隔离成分，对于源自敏感细胞的 MPs，主要是被动隔离成分。这种可用药物浓度的降低有可能促成平行途径，并补充 P-gp 的细胞间转移。这些发现进一步支持了 MPs 在限制癌症成功治疗中的作用。
