Cancer metastasis, the spread of tumor cells from a primary site to secondary sites in the body, is the most common cause of cancer mortality. Prior to metastasis, primary tumor-derived factors are transported to secondary sites to suppress host immunity and to form an environment that supports metastatic tumor growth, called a pre-metastatic niche. Among these factors, extracellular vesicles (EVs) have been reported to play crucial roles. In the first part of this work, I investigate the routes of EV transport from primary tumors to secondary sites, and the consequences of modulating EV transport on premetastatic niche formation and metastasis. In the second part of this work, I analyzed EVs in another disease context, namely lipedema, with the goal of identifying disease-specific biomarkers.In Chapter 1, I introduce the fields of lymphatic biology and EVs. I describe the mechanisms of lymphatic drainage and the roles of lymphatic vessels in health and disease development. Then, I describe the production, properties and functions of EVs. In Chapter 2, I adapt methods to study EVs in vitro and in vivo. Such methods include producing, purifying and fluorescently labeling EVs. Additionally, I optimize the measurement of EV contents in various biological milieus. In Chapter 3, I investigate the roles of lymphatic vessels in the distribution of EVs from the healthy skin and melanoma tumors to draining lymph nodes and distant sites. For this, I perform EV biodistribution studies in wild type mice and in a transgenic mouse model which lacks dermal lymphatics. In Chapter 4, I investigate the consequences of tumor lymphangiogenesis on EV transport and on pre-metastatic niche formation. I analyze these mechanisms in two different mouse tumor models which were transduced to stably overexpress the main lymphangiogenic growth factor VEGFC. In Chapter 5, I characterize the fluid part of lipoaspirate samples as a source of adipose tissue-specific factors. In these samples, I analyze adipokines and EVs in lipedema patients compared with control patients. In Chapter 6, I discuss the consequences of this thesis in the field of cancer metastasis and biomarker development. Finally, I suggest future work that will address remaining questions following upon this thesis.