Faculty of Health Sciences Department of Pharmacy The Metabolome and Lipidome of Ulcerative Colitis Joseph Diab A dissertation for the degree of Philosophiae Doctor - June 2020 A dissertation for the degree of Philosophiae Doctor The Metabolome and Lipidome of Ulcerative Colitis Joseph Diab Tromsø, June 2020 Natural Product and Medicinal Chemistry Research Group Department of Pharmacy Faculty of Heath Science UiT The Arctic University of Norway Norway ‘’The best workers, like the happiest livers, look upon their work as a kind of game: the harder they play the more enjoyable it becomes’’ Robert Baden-Powell “If you want to go fast, go alone. If you want to go far, go together” African Proverb Acknowledgments This work is funded by UiT – The Arctic University of Norway and Helse Nord. It was carried out at the Natural Products and Medicinal Chemistry Research Group, Department of Pharmacy, UiT The Arctic University of Norway. I would like to express my gratitude to everyone that has helped me along the way. First, I want to express my deep gratitude to my supervisors; To Guro Forsdahl for being the most understanding, patient, and motivating person ever. Thanks for guiding my PhD journey, and for providing all the scientific and the personal support. To Terkel Hansen for being there every time I needed a guidance, and for the numerous conversations on Mass spectrometry, data analysis, beers, and dogs. To Einar Jensen for establishing the collaboration with the Swedish Metabolomics centre (SMC). To Jon Florholmen for being the father of ASIB, for providing his extent knowledge, and for his amazing passion for science. For Rasmus Goll, for dedicating his time to share his valuable opinion and experience, for letting me experiencing his cooking skills, and for being the best ECCO traveling companion. I really hope to continue being part of your cool (poop) projects. I could never complete my PhD work without the help of many collaborators. Thanks to all personals at SMC, especially Thomas Moritz. Thanks a lot for hosting me in your lab. I feel privileged to be able to perform metabolomic analysis in SMC under the supervision of the best in the field! Thanks to Sandra Gouveia for her significant contribution in my project, and to Maria Ahnlund for providing me with hands-on training on data processing. Many thanks to Hans Stenlund for babysitting me, and for providing me with the toolbox that I needed to complete my PhD. I am honoured to get to know (and to have access) to such a great mind like yours. Thanks for my co-author, Rania al Mahdi. Thanks for Ines Heiland and Roland Sauter for helping me putting our finding into a wider biological context. I want to thank former and current colleagues at the Department of Pharmacy. I will never forget our legendary Christmas parties, ‘summer’ BBQs, wine tasting, seminars, and cakes. I want to thank all formers and current members of my research group, especially Martina Havelkova and Trude Andersen for being my trust circle, and for Terje Vasskog for sharing his wisdom and experience with me. I am still looking forward for our first fishing trip Over the last 4 years, I was blessed to have an amazing family of friends in Tromsø. Thanks for Beate for taking me on amazing adventures. Thanks for Julia and Theresa for their great friendship. Thanks for Fabrizio for his advices, his authentic ice cream and pizza. Thanks for Vidar and Joao for all the bromance. Thanks for Christina and Jan Håkon for making me be part of their family. Thanks for Merete for being there in every great moment of my PhD. Thanks for Dominik for helping me since the very first moment I started at UiT. I will miss our ritual of B&B. Thanks for my PhD colleagues, my brother and sister in arms, especially Nina, Margherita, Jennifer, Julie, Anup, Lorenz, and Chris. I wish you all the best in your future. Last, I am overwhelmed by the love and the unconditional support of my family. My amazing wife Katrine whom filled my life with happiness, and provided me with the purpose. My father and mother, Marwane and Rima, who made me the person I am. I will always try to make you proud of me. To my beloved brother Rami and his wife Marie, who stood by me through my worst episode. To my in laws, Mette and Kim, who took me as son and provided me with care and beers . Seven years ago, I left Syria for a journey that took me to Lebanon, Spain, Germany, and Norway. I was lucky to survive one of the deadliest war in the last 50 years, while many could not… I am grateful for Norway for hosting me, for allowing me to chase my dreams, and for giving me a safe home. However, I will always be proud of my mother country Syria and of all the sacrifices of its daughters and sons. Table of Contents List of Papers ............................................................................................................................................ I Summary ............................................................................................................................................... III Abbreviations ........................................................................................................................................ IV 1.Introduction .......................................................................................................................................... 1 1.1 Inflammatory Bowel Disease ............................................................................................................ 1 1.1.1 Epidemiology ................................................................................................................................. 1 1.1.2 Pathogenesis of Ulcerative Colitis ................................................................................................. 1 1.1.3 Management of Ulcerative Colitis ................................................................................................. 6 1.1.4 Biomarkers for UC ......................................................................................................................... 9 1.1.5 Clinical outcome........................................................................................................................... 10 1.1.6 ASIB study ................................................................................................................................... 12 1.2 Metabolomics and Lipidomics ........................................................................................................ 13 1.2.1 Definition ..................................................................................................................................... 13 1.2.2 Analytical approaches for metabolomics ..................................................................................... 15 1.2.3 Analytical platforms for metabolomics ........................................................................................ 15 1.2.4 Sample preparation ....................................................................................................................... 16 1.2.5 Metabolite separation ................................................................................................................... 18 1.2.6 Metabolites detection and quantification by MS .......................................................................... 19 1.2.7 Data processing ............................................................................................................................ 21 1.2.8 Data normalization and scaling .................................................................................................... 22 1.2.9 Data analysis ................................................................................................................................ 22 1.2.10 Biomedical interpretation ........................................................................................................... 23 1.2.11 Metabolomics in IBD ................................................................................................................. 23 2. Aims of the study .............................................................................................................................. 26 3. Methods ............................................................................................................................................. 27 3.1. Biopsies collection ......................................................................................................................... 27 3.2. Global metabolomics by CG-MS ................................................................................................... 27 3.3. Global metabolomics by LC-MS ................................................................................................... 27 3.3. Global lipidomics by LC-MS ......................................................................................................... 28 3.4. Targeted metabolomics by LC-MS ................................................................................................ 28 3.5. Data analysis .................................................................................................................................. 29 4. Summary of results ............................................................................................................................ 30 4.1 Paper I ............................................................................................................................................