Department of Clinical Chemistry/Preclinical Pharmacology, The Netherlands Cancer Institute (Antoni van Leeuwenhoek Hospital), Amsterdam, The Netherlands.
J Chromatogr B Analyt Technol Biomed Life Sci. 2011 Dec 15;879(32):3823-31. doi: 10.1016/j.jchromb.2011.10.028. Epub 2011 Oct 29.
Palomid 529 (8-(1-Hydroxy-ethyl)-2-methoxy-3-(4-methoxy-benzyloxy)-benzo[c]chromen-6-one), is a novel non-steroidal small-molecule drug, which inhibits both mTORC1 and mTORC2 assembly, and elicits both anti-angiogenic and direct anti-tumor effects in vivo. We have developed and validated a sensitive and selective method for the quantification of Palomid 529 in human and mouse plasma and in a range of mouse tissue samples. Sample pretreatment involved liquid-liquid extraction with tert-butyl methyl ether yielding a recovery of >75%. Palomid 529 and the internal standard Palomid 545 were separated using a GraceSmart RP18 column (2.1 mm × 150 mm) packed with 5 μm C-18 material and a mobile phase comprised of 50% (v/v) acetonitrile and 50% (v/v) water delivered at a flow rate of 0.2 ml/min, and were detected by UV absorbance at a wavelength of 315 nm. Within the linear range of the calibration curve (10-10,000 ng/ml), acceptable accuracy and precision was achieved for all tested matrices. The validation results show that the method was selective and reproducible. Palomid 529 was stable in plasma upon 3 repeated freeze-thaw cycles and during storage for up to 24h at ambient temperature. However, pre-treated samples waiting for HPLC analyses need to be kept under dimmed light conditions at ambient temperature since a significant degradation of both Palomid 529 and Palomid 545 was observed when exposed to light. A pilot pharmacokinetic study in mice demonstrated the applicability of this method for pharmacokinetic purposes. Even at a low dose of 5.4 mg/kg this assay was still sensitive enough to determine the drug concentration in plasma samples obtained up to 24h after administration.
帕洛米德 529（8-（1-羟乙基）-2-甲氧基-3-（4-甲氧基苄氧基）-苯并[c]色烯-6-酮）是一种新型的非甾体小分子药物，可抑制 mTORC1 和 mTORC2 组装，并在体内产生抗血管生成和直接抗肿瘤作用。我们开发并验证了一种灵敏且选择性的方法，用于定量检测人血浆和小鼠血浆以及一系列小鼠组织样本中的帕洛米德 529。样品预处理涉及使用叔丁基甲基醚进行液-液萃取，回收率>75%。帕洛米德 529 和内标帕洛米德 545 采用 GraceSmart RP18 柱（2.1mm×150mm）分离，柱内填充 5μm C-18 材料，流动相由 50%（v/v）乙腈和 50%（v/v）水组成，流速为 0.2ml/min，通过 315nm 处的紫外吸光度进行检测。在校准曲线的线性范围内（10-10000ng/ml），所有测试基质均具有可接受的准确性和精密度。验证结果表明该方法具有选择性和重现性。帕洛米德 529 在经过 3 次反复冻融循环和在室温下储存长达 24 小时后，在血浆中稳定。然而，待进行 HPLC 分析的预处理样品需要在室温下处于避光条件下保存，因为当暴露于光线下时，帕洛米德 529 和帕洛米德 545 均会发生显著降解。在小鼠中进行的初步药代动力学研究表明，该方法适用于药代动力学研究。即使在 5.4mg/kg 的低剂量下，该测定法仍足够灵敏，可在给药后 24 小时内测定血浆样品中的药物浓度。
