Nicklee T, Crump M, Hedley D W
Department of Oncologic Pathology, Ontario Cancer Institute/Princess Margaret Hospital, Toronto, Canada.
Cytometry. 1996 Nov 1;25(3):205-10. doi: 10.1002/(SICI)1097-0320(19961101)25:3<205::AID-CYTO1>3.0.CO;2-E.
A fluorescence image cytometry technique was developed to measure the effects of topotecan, a topoisomerase I inhibitor, on the nuclear expression of topoisomerase II alpha in a series of patients with refractory or relapsed acute myeloid leukemia (AML). We used a commercially available affinity-purified rabbit polyclonal antibody and a fluorescein-conjugated secondary antibody. By using DAPI as a DNA counterstain and dual wavelength excitation, it was possible to measure enzyme expression in the cell nucleus, and to examine its cell cycle phase distribution. In human acute leukemia cell lines, topoisomerase II alpha expression was greatest in late S and G2 phases, but in leukemia patient samples the enzyme expression appeared to be much less cell cycle dependent. There was considerable interpatient variation in the effects of topotecan on topoisomerase II alpha expression in the leukemia patients, with a threefold increase in the median value after 48 h followed by a decline to pretreatment levels after 5 days of treatment with the topoisomerase I inhibitor. Although these findings should be treated with caution because of the small number of cases studied, they support the prediction that topoisomerase I inhibitors might be capable of increasing sensitivity to topoisomerase II active drugs such as anthracyclines and epipodophyllotoxins by upregulating topoisomerase II expression. They also illustrate the potential value of fluorescence image cytometry for making sequential measurements of the effects of drug resistance modulating agents in cancer patients.
开发了一种荧光图像细胞术技术，用于测量拓扑异构酶I抑制剂拓扑替康对一系列难治性或复发性急性髓系白血病（AML）患者拓扑异构酶IIα核表达的影响。我们使用了市售的亲和纯化兔多克隆抗体和荧光素偶联二抗。通过使用DAPI作为DNA复染剂和双波长激发，能够测量细胞核中的酶表达，并检查其细胞周期阶段分布。在人急性白血病细胞系中，拓扑异构酶IIα表达在S期晚期和G2期最高，但在白血病患者样本中，酶表达似乎对细胞周期的依赖性小得多。拓扑替康对白血病患者拓扑异构酶IIα表达的影响在患者之间存在相当大的差异，48小时后中位数增加了三倍，随后在用拓扑异构酶I抑制剂治疗5天后降至预处理水平。尽管由于研究的病例数量较少，这些发现应谨慎对待，但它们支持这样的预测，即拓扑异构酶I抑制剂可能能够通过上调拓扑异构酶II表达来增加对拓扑异构酶II活性药物（如蒽环类药物和表鬼臼毒素）的敏感性。它们还说明了荧光图像细胞术在对癌症患者耐药性调节剂的效果进行连续测量方面的潜在价值。
