ADVANCES IN CLINICAL LIQUID BIOPSY: BIOFLUID EXTRACELLULAR VESICLES AS PROMISING SOURCES FOR DRUG METABOLISM AND EFFICACY SURVEILLANCE

Cell-derived extracellular vesicles (EVs) are increasingly being recognized as an important mediator of intercellular communication, immune response and waste disposal. Due to the prevalent dissemination in most biofluids, EVs provide a promising alternative for non-invasive liquid biopsy in clinics. With internal cargoes well-preserved by EV membranes, EV-containing DNA, RNA, proteins, lipids and metabolites are widely investigated for biomarker discovery and disease progression and therapeutics monitoring. Among them, proteins and their post-translational modifications (PTMs), are the direct phenotypic indicators of pathophysiological states. Besides, development of liquid chromatography-tandem mass spectrometry (LC-MS/MS) has allowed high-throughput and discovery-driven bioanalysis on proteomics, including EV proteomics. Nevertheless, the translational research and clinical applications on EVs are severely hampered due to the lack of suitable and efficient isolation methods as well as robust and sensitive proteomic approaches. This dissertation proposes EV proteomic strategies for therapeutic surveillance by using a novel EV isolation technique. Chapter One introduces overall status of proteomics and phosphoproteomics in translational and clinical analysis for to-date medical sciences. Chapter Two highlights a novel EV isolation method termed EV total recovery and purification (EVtrap). Chapter Three delves into the development of a pipeline proteomic strategy that utilizes EV P450 enzymes to monitor drug metabolic behaviors. Finally, Chapter Four explores the possibility of leveraging protein PTMs in EVs, especially tyrosine phosphorylation, to indicate drug therapeutic efficacy in a clinical setting. This dissertation primarily proposes EV-based liquid biopsy for revolutionary clinical analysis, especially on longitudinal monitoring for in vivo treatment response.