DIFFERENTIATION OF J-SERIES PROSTAGLANDINS AS PRO-APOPTOTIC PRODUCTS OF CANCER CELL METABOLISM
Kobet, Robert A.
Cyclopentenone J-series prostaglandins and prostaglandin-ethanolamide conjugates have been shown to selectively induce endoplasmic reticulum stress-apoptosis in different cancer cell types. These lipids have been identified as the downstream products of COX-2 metabolism of arachidonic acid and arachidonoyl ethanolamide, respectively. While techniques such as ELISA and UV-Vis spectroscopy can detect these molecules based on their class, they fail to offer the specificity necessary to differentiate those prostaglandins that are isomeric in structure. The J-series prostaglandins PGJ2 and [Delta]12PGJ2 are structural isomers that differ in the position of a single double bond. The differentiation of these prostaglandins is an important step in the elucidation of the metabolic pathway mediated by COX-2 and the mechanism of action of the downstream products. Mass spectrometry has been shown to be a powerful method in the study of biological lipids due to the sensitivity and specificity it offers. Tandem mass spectrometry (MS/MS) offers the ability to isolate molecules of identical masses and obtain information regarding their individual fragmentation patterns as well as energetics and stability characteristics. The goal of this project is to differentiate the prostaglandin isomers PGJ2 and [Delta]12PGJ2 using a variety of MS/MS methods. MS/MS experiments were performed on prostaglandin standard solutions using a linear quadrupole-time of flight (Q-ToF) mass spectrometer and a quadrupole ion trap (QIT). An initial set of experiments aimed to identify product ions specific to either isomer using MS/MS and pseudo-MS3 on the Q-ToF and MSn analysis using the QIT. Our results indicate that neither method yielded product ions with significant intensity and reproducibility to be designated as specific to either isomer. Our focus then shifted to attempting to exploit differences in the energies of onset of some of the common product ions (energy-resolved mass spectrometry). Under controlled sample preparation and ESI source parameters, this method was also shown to be unsuccessful in providing information specific to either PGJ2 or [Delta]12PGJ2. These results are predicted to be the result of in-source isomerization of PGJ2 to the conjugated [Delta]12 isomer. In order to confirm unique fragmentation of the double bond isomers using the established LC-ESI-MS/MS method initially used to identify these metabolic products, offline derivatization methods will likely be necessary to fix the double bond position in order to prevent the in-source isomerization that is believed to have hindered our ability to differentiate these molecules.
Kobet, Robert A.. (December 2016). DIFFERENTIATION OF J-SERIES PROSTAGLANDINS AS PRO-APOPTOTIC PRODUCTS OF CANCER CELL METABOLISM (Master's Thesis, East Carolina University). Retrieved from the Scholarship. (http://hdl.handle.net/10342/6003.)
Kobet, Robert A.. DIFFERENTIATION OF J-SERIES PROSTAGLANDINS AS PRO-APOPTOTIC PRODUCTS OF CANCER CELL METABOLISM. Master's Thesis. East Carolina University, December 2016. The Scholarship. http://hdl.handle.net/10342/6003. January 23, 2020.
Kobet, Robert A., “DIFFERENTIATION OF J-SERIES PROSTAGLANDINS AS PRO-APOPTOTIC PRODUCTS OF CANCER CELL METABOLISM” (Master's Thesis., East Carolina University, December 2016).
Kobet, Robert A.. DIFFERENTIATION OF J-SERIES PROSTAGLANDINS AS PRO-APOPTOTIC PRODUCTS OF CANCER CELL METABOLISM [Master's Thesis]. Greenville, NC: East Carolina University; December 2016.
East Carolina University