The proposed Analytical Core will perform analyses of a wide variety of bioactive lipids in various mouse tissues and biological fluids for investigators in the Program Project Grant. The four projects in this PPG proposal will evaluate the role of COX-2, and associated metabolites such as PGE2 that are believed to be involved in the initiation and progression of colon, breast, and endometrial cancers. A strong advantage of the current PPG is that we will now also explore the important roles of a series of bioactive lipids including endocannabinoids and sphingolipids and their interactions with COX-2 pathways. This will require the Analytical Core to not only quantify a wide variety of prostaglandins such as PGE2 and PGE-M, but also to determine and quantify pharmacologically relevant endocannabinoids and sphingolipids in various animal tissues. These three categories of bioactive lipids will be measured predominantly by methods that involve high performance liquid chromatography/electrospray ionization tandem mass spectrometry (LC/ESIMS/ MS) using a stable isotope dilution technique developed by the Core Director and which has been extensively used in her laboratory. Use of eicosanoid assays by investigators involved in this PPG grant has already contributed to a number of publications that demonstrate the potential impact of these lipids on cancer development. The proposed research will enhance our understanding of the essential roles of bioactive lipids in the pathophysiological processes related to malignancy. In addition, the Core will also develop a novel LC/MS/MS method to simultaneously quantify all three classes of bioactive lipids. This will allow us to assess the interaction of bioactive lipids of interest and advance our knowledge of how the three categories of bioactive lipids are interrelated. Lastly, the Core will quantify blood and tissue levels of intervention compounds, such as the selective COX-2 inhibitor celecoxib, as well as prostaglandins, such as PGI2 and TXA2 to facilitate monitoring the pharmacologic effect of celecoxib proposed in all four projects. Thus, the overall objective of the Analytical Core is to provide sophisticated technical and analytical support that involves integration of sensitive isolation and detection procedures with the latest bioanalytical methods
The proposed research work involves identification and quantification of bioactive lipids, including eicosanoids, endocannanbinoids, and sphingolipids using state-of-the-art mass spectrometry instrumentation and associated lipid extraction and separation techniques. Bioactive lipids of interest will be isolated from animal tissues derived from mouse colon, breast and endometrial cancers. Sophisticated analytical methods as well as isolation and separation techniques to be used for determination of complex bioactive lipids are critical for the success of this PPG.
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