College of Biology, Hunan Province Key Laboratory of Plant Functional Genomics and Developmental Regulation, Hunan University, Changsha, 410082, China.
Precision Pharmacy & Drug Development Center, Department of Pharmacy, Second Affiliated Hospital, Air Force Medical University, Xi'an, 710038, China.
Analyst. 2024 Oct 7;149(20):5073-5080. doi: 10.1039/d4an01059k.
Cisplatin (DDP) is a potent chemotherapeutic drug, which can regulate tumor cell apoptosis by up-regulating caspase-3 activity. Thus, monitoring caspase-3 activity in breast cancer cells can directly illustrate the efficiency of DDP treatment. In this study, by using reduced graphene oxide (rGO) as a quencher of a fluorescence labeled peptide, we developed an "off to on" method to monitor the effect of DDP on caspase-3 in breast cancer cells. In this method, the rGO quenched fluorescence with an ultra-high level of efficiency. Caspase-3 hydrolyzed the polypeptide probe, generating two segments of different lengths. The release of a short segment marked with fluorophores led to the recovery of the fluorescence signal (Ex/Em = 450/521 nm). Under the optimal conditions, the linear range of caspase-3 was 0.4-7 U mL and the limit of detection was 0.33 U mL. The upregulating effect of DDP on intracellular caspase-3 activity was visualized with the "off to on" method and flow cytometry assay showed that caspase-3 activity increased along with the apoptosis rate of tumor cells. The above results show the practical application of the method for evaluating the efficacy of drugs against cancer cells.
顺铂（DDP）是一种有效的化疗药物，可通过上调半胱天冬酶-3 活性来调节肿瘤细胞凋亡。因此，监测乳腺癌细胞中半胱天冬酶-3 的活性可以直接说明 DDP 治疗的效果。在本研究中，我们使用还原氧化石墨烯（rGO）作为荧光标记肽的猝灭剂，开发了一种“关-开”方法来监测 DDP 对乳腺癌细胞中半胱天冬酶-3 的作用。在该方法中，rGO 以超高效率猝灭荧光。半胱天冬酶-3 水解多肽探针，生成两个长度不同的片段。带有荧光标记的短片段的释放导致荧光信号的恢复（Ex/Em = 450/521nm）。在最佳条件下，半胱天冬酶-3 的线性范围为 0.4-7 U mL，检测限为 0.33 U mL。用“关-开”法可视化 DDP 对内源性半胱天冬酶-3 活性的上调作用，流式细胞术检测表明，随着肿瘤细胞凋亡率的增加，半胱天冬酶-3 的活性也随之增加。上述结果表明，该方法可用于评估药物对癌细胞的疗效。
