Scicinski Jan, Oronsky Bryan, Cooper Vance, Taylor Michael, Alexander Mike, Hadar Rebecca, Cosford Rebecca, Fleischmann Thomas, Fitch William L
RadioRx Inc., 800 W El Camino Real, Suite 180, Mountain View, CA 94040, USA.
Bioanalysis. 2014 Apr;6(7):947-56. doi: 10.4155/bio.13.331.
Bioanalytical methods were required to study the novel anticancer drug, RRx-001 preclinically and for clinical pharmacokinetic analysis; however, RRx-001 quickly and completely disappeared on intravenous administration in preclinical species.
Quantification of RRx-001 directly or by derivatization was unsuccessful. On exposure to whole blood, RRx-001 formed the glutathione (GSH) adduct very rapidly, suggesting this metabolite as the bioanalyte. However, rapid enzymatic degradation in the blood matrix of RRx-001-GSH posed significant technical problems. Herein, we describe a novel and broadly applicable solution to stabilize GSH conjugates in blood samples by inhibiting the degrading enzyme. Liquid chromatography-tandem mass spectrometry methods for analysis of RRx-001-GSH in rat, dog and human plasma were developed and successfully validated to good laboratory practice standards.
Extensive breakdown of RRx-001-GSH was effectively stopped by addition of the enzyme inhibitor, acivicin. The developed liquid chromatography-tandem mass spectrometry assay for RRx-001-GSH was validated for use in preclinical toxicology studies and the Phase I first-in-human clinical trial.
在临床前研究新型抗癌药物RRx-001以及进行临床药代动力学分析时，需要生物分析方法；然而，在临床前物种中静脉给药后，RRx-001迅速且完全消失。
直接或通过衍生化对RRx-001进行定量均未成功。暴露于全血后，RRx-001非常迅速地形成了谷胱甘肽（GSH）加合物，表明该代谢物为生物分析物。然而，RRx-001-GSH在血液基质中的快速酶促降解带来了重大技术问题。在此，我们描述了一种通过抑制降解酶来稳定血样中GSH缀合物的新颖且广泛适用的解决方案。开发了用于分析大鼠、犬和人血浆中RRx-001-GSH的液相色谱-串联质谱方法，并成功按照良好实验室规范标准进行了验证。
通过添加酶抑制剂阿西维辛有效地阻止了RRx-001-GSH的大量分解。所开发的用于RRx-001-GSH的液相色谱-串联质谱测定法已验证可用于临床前毒理学研究和I期首次人体临床试验。
