Urine aldehydeomics by CAX mass tag mass spectrometry

Abstract

The central theme of this dissertation is to develop a sensitive method for the nontargeted analysis of aldehydes as a biomarker in human urine. We are particularly interested in exploring the urine aldehydeome of ovarian cancer. This method is based on ESI (electrospray ionization) and MALDI (matrix assisted laser desorption ionization) mass spectrometric analyses.Recent findings in the literature suggest that aldehyde-levels could be elevated in cancer due to disruptions in metabolic processes. These aldehydes may in part be excreted into the urine, which can be collected non-invasively and analyzed. The challenge associated with evaluating urine is that aldehydes may be present in trace quantities and lack ionization efficiency. In order to boost the sensitivity and specificity of detection of aldehydes, our lab has developed a mass tag known as CAX-A, or cationic xylyl aniline.Aldehydes tagged with CAX-A can be readily detected in a mass spectrometer (MS) due to the nature of the CAX group and the higher order MS behavior of CAX-aldehydes. The overall research plan is to apply CAX-MS technology to 1) devise a sensitive tagging approach for the detection of aldehyde standards in buffer; and 2) apply this tagging methodology to detect aldehyde biomarkers in urine. For validation of the labeling methodology in buffer, we applied CAX-labeling to 6 standard aldehydes. All six standard CAX-aldehyde derivatives were detected as precursor ions after dilution to 830 fmol/injection by ESI-MS (signal-to-noise [S/N] ratio 587-1573) and to 1.1 fmol/spot via MALDI-MS (S/N 36-1945). CAX-MS was also used to detect aldehydes in 10 urine samples (5 ovarian cancer cases and 5 controls), yielding 5,419 features, of which 238 were putative aldehydes. This expanded the human urine putative aldehydeome about me to this day. I also owe a big thank you to Rafay Abu of the Barnette Institute for his kind help with the LTQ-Orbitrap instrument mechanics. A special thank you to Dr. Sumantha Garai, a former scientist at Dr. Thakur's Lab, for helping me gain a better understanding of the organic chemistry involved in this work. I would also like to extend my gratitude to our collaborator, Dr. Marsha Moses, at the Boston Children's Hospital for providing us with precious samples and making this research more meaningful. A special thank you to Rosalee Robinson, Sarom Lay, and Ainsley Watkins of the Pharmaceutical Sciences Department for helping me streamline my path towards graduation. I also extend my thanks to Drs. Joelle Carlo, Jane Saczynski, and Lara Milane, for their guidance during my 4.5 years as a teaching assistant.Finally, I offer my deepest gratitude to my family and friends for their unconditional love and support. My mom, Purnima, a trailblazer of her times, educated herself and made countless sacrifices to come to this country and to give us a better life. She is my idol. My dad, Sam, with his great sense of humor (dad jokes) has been a steady and gentle presence throughout my life. My wonderful sister, Syndhia, has always bee...