Extracellular vesicles : mediators of immune modulation in the lung and as therapeutic vehicles

Extracellular vesicles (EVs) are released from all cell types, and carry a wide setup of proteins, nucleic acids, lipids and other cargo. The overall aim of this thesis is to explore EV-based immune therapy, but also to find clues on mechanisms of the inflammatory disease sarcoidosis, and of lung cancer. Exosomes from dendritic cells (DCs) pulsed with antigen can induce antigen-specific responses in vitro and in vivo.

Study I is an investigation comparing exosomes and microvesicles (MVs), which may complement exosomes therapeutically. We found surprisingly similar phenotypes of the two EV subtypes, including size distribution and immunestimulatory molecule expression. However, when tested in vivo, only exosomes induced a significant antigen-specific CD8+ T cell response. Antigenic restimulations ex vivo did, however, suggested that also MVs had such capacity, and both vesicle types induced antigen-specific IgG production. We further targeted the inflammatory disease pulmonary sarcoidosis in study II and study III with the aim to increase understanding of disease mechanisms, but also to search for disease biomarker candidates, and possible new treatment regimens. Broad proteomic characterizations of exosomes from patients revealed high abundance of proinflammatory molecules including leukotriene (LT)-forming enzymes. Large portions of the complement system were elevated, and we flagged vitamin Dbinding protein as a possible biomarker for sarcoidosis. Functional tests of patient exosomes further suggested that they can engage monocytes and favor release of pro-inflammatory cytokines. The effects were partly dependent on LTs, and we could reduce cytokine production using the commercially available LT receptor antagonist Montelukast. Study IV on exosomes in lung cancer is focused on how exosomes may contribute to tumor progression via LTs. Exosomes from pleural effusions of patients favored generation of tumorigenic LTD4, as well as tumor cell migration, which could be reduced using Montelukast.

In summary, this thesis highlights the importance of investigating all EV subtypes in both basic and applied research. Further, the ability of EVs to contribute to inflammatory processes in the lungs underscores the potential of EVs in understanding disease mechanisms and finding diagnostic and prognostic disease markers. Finally, all three lung studies IIIV point to the possibility of interfering with LTs in inflammatory conditions, with possible applications also in cancer therapy.

List of scientific papers

I. Wahlund CJE, Güclüler G, Hiltbrunner S, Veerman RE, Näslund TI, Gabrielsson S. Exosomes from antigen-pulsed dendritic cells induce stronger antigen-specific immune responses than microvesicles in vivo. Sci Rep. 2017 Dec 6;7(1):17095.
https://doi.org/10.1038/s41598-017-16609-6

II. Martinez-Bravo MJ, Wahlund CJE, Qazi KR, Moulder R, Lukic A, Rådmark O, Lahesmaa R, Grunewald J, Eklund A, Gabrielsson S. Pulmonary sarcoidosis is associated with exosomal Vitamin D-binding protein and inflammatory molecules. J Allergy Clin Immunol. 2017 Apr;139(4):1186-1194.
https://doi.org/10.1016/j.jaci.2016.05.051

III. Wahlund CJE, Güclüler G, Lepzien R, Smed-Sörensen A, Kullberg S, Eklund A, Grunewald J, Gabrielsson S. Sarcoidosis exosomes stimulate monocytes to produce proinflammatory cytokines and CCL2, which can be inhibited by Montelukast. [Manuscript]

IV. Lukic L, WahlundCJE, Gomez C, Brodin D, Samuelsson B, Wheelock CE, Gabrielsson S, Radmark O. Exosomes from lung cancer pleura exudates form LTD4, promoting cell migration and survival via CysLT1. [Manuscript]