Abstract 4774: LC-MS/MS detection of the dansyl-modified biologically active CDK inhibitor VMY-1-103 using an in vivo model system

Abstract

Abstract The development of new small molecule kinase inhibitors as cancer therapeutic drugs has placed increased emphasis on determining the mechanism of action of the new compounds preclinically, such as in genetically engineered mice or using human tissue xenografts. The ability to accurately measure the pharmacokinetics and pharmacodynamics of new compounds is often hampered by the lack of suitable platforms to measure serum and tissue distribution with high sensitivity and specificity. We have recently developed a novel dansylated cyclin dependent kinase inhibitor called VMY-1-103 (VMY) that has purvalanol B as its base compound. We established that VMY was significantly more effective in blocking cell cycle progression and inducing apoptosis than purvalanol B. Since dansylation is frequently used to enhance mass spectrometry-based quantitative analyses, we hypothesized that this modality could similarly be used to evaluate the tissue distribution of VMY in mouse models. In the current study, we developed a rapid liquid phase-extraction of VMY and purvalanol B from mouse tissue and validated a sensitive liquid chromatography-tandem mass spectrometry based assay to quantify tissue delivery of these compounds in vivo. Comparisons of the standard curves for VMY versus Purvalanol B revealed a greater than 7.7-fold increase in sensitivity for VMY. Using both the ND-2 SmoA1, a genetically engineered mouse model of medulloblastoma, and normal mice, we quantified tissue delivery by mass spectrometry as well as Area Under the Curve (AUC) analysis by integrating the peak intensity normalized by an internal standard. Our analyses revealed that VMY delivered intraperitoneally at 20mg/kg was found at its highest concentration in serum 4 hours after injection. We further established that VMY was distributed in multiple tissues, including pancreas and prostate at varying levels and, importantly, we establish that VMY was able to traverse an intact blood-brain barrier as it was detected in the brain and cerebellum of normal mice. Purvalanol B delivered in the same manner as VMY was found at significantly lower levels in all tissues examined and unlike VMY, we failed to detect purvalanol B in brain or and cerebellar tissue. The significance of this study illustrates that a biologically active compound can be modified by dansylation and exhibit both increased sensitivity to mass spectrometry and enhanced bioavailability and tissue delivery, including the brain, in vivo. We also define a simple mass spectrometry-based tissue and serum assay. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4774. doi:1538-7445.AM2012-4774