Department of Pharmaceutical Technology and Biochemistry, Faculty of Chemistry, Gdańsk University of Technology, 11/12 Narutowicza Str., Gdańsk 80-233, Poland.
Department of Analytical Chemistry, Faculty of Chemistry, Gdańsk University of Technology, 11/12 Narutowicza Str., Gdańsk 80-233, Poland.
J Pharm Biomed Anal. 2025 Jan 1;252:116493. doi: 10.1016/j.jpba.2024.116493. Epub 2024 Sep 30.
Unsymmetrical bisacridines (UAs) represent a novel class of anticancer agents. Their high cytotoxicity towards multiple human cancer cell lines and inhibition of human tumor xenograft growth in nude mice signal their potential for cancer treatment. Therefore, the mechanism of their strong biological activity is broadly investigated. Here, we explore the efflux and metabolism of UAs, as both strongly contribute to the development of drug resistance in cancer cells. We tested two highly cytotoxic UAs, C-2028 and C-2045, as well as their glucuronic acid and glutathione conjugates in human cancer cell lines (HepG2 and LS174T). As a point of reference for cell-based systems, we examined the rate of UA metabolic conversion in cell-free systems. A multiple reaction monitoring (MRM)-mass spectrometry (MS) method was developed in the present study for analysis of UAs and their metabolic conversion in complex biological matrices. Individual analytes were identified by several features: their retention time, mass-to-charge ratio and unique fragmentation pattern. The rate of UA uptake and metabolic transformation was monitored for 24 h in cell extracts and cell culture medium. Both UAs were rapidly internalized by cells. However, C-2028 was gradually accumulated, while C-2045 was eventually released from cells during treatment. UAs demonstrated limited metabolic conversion in cells. The glucuronic acid conjugate was excreted, whereas the glutathione conjugate was deposited in cancer cells. Our results obtained from cell-free and cell-based systems, using a uniform MRM-MS method, will provide valuable insight into the mechanism of UA biological activity in diverse biological models.
不对称双吖啶（UAs）代表了一类新型的抗癌药物。它们对多种人癌细胞系的高细胞毒性和对裸鼠人肿瘤异种移植物生长的抑制作用表明了它们在癌症治疗中的潜力。因此，广泛研究了它们强生物活性的机制。在这里，我们探讨了 UAs 的外排和代谢，因为这两者都强烈促成了癌细胞耐药性的发展。我们测试了两种高细胞毒性的 UAs，C-2028 和 C-2045，以及它们在人癌细胞系（HepG2 和 LS174T）中的葡萄糖醛酸和谷胱甘肽缀合物。作为细胞为基础的系统的参考点，我们检查了无细胞系统中 UA 代谢转化的速度。本研究开发了一种多重反应监测（MRM）-质谱（MS）方法，用于分析复杂生物基质中的 UAs 及其代谢转化。通过几个特征来鉴定各个分析物：保留时间、质荷比和独特的碎片模式。在细胞提取物和细胞培养液中监测了 UA 摄取和代谢转化的速度 24 小时。两种 UAs 都被细胞迅速内化。然而，C-2028 逐渐积累，而 C-2045 在治疗过程中最终从细胞中释放出来。UAs 在细胞中的代谢转化有限。葡萄糖醛酸缀合物被排泄，而谷胱甘肽缀合物则沉积在癌细胞中。使用统一的 MRM-MS 方法从无细胞和细胞为基础的系统中获得的结果将为不同生物模型中 UA 生物学活性的机制提供有价值的见解。
