Date of Graduation
Statler College of Engineering and Mineral Resources
Chemical and Biomedical Engineering
David J Klinke, II
Christopher F Cuff
Robin S Hissam
It is still unknown how tumor exosomes influence malignant cell survival and alter cell-to-cell communication to modulate the immune system by, in part, manipulating the activity of cytotoxic T lymphocytes. This study focuses on two critical parts regarding cancer exosomes. The first part is to tailor existing bionanotechnology methods to account for the nanoscale aspects of exosome biology. SEM, TEM, Bioanalyzer and flow cytometry were used to characterize exosome morphologies, identify specific protein biomarkers HER1 and HER2, as well as the quality of RNAs enclosed in exosomes. Competing methods related to exosome isolation, production, preservation, stability and analysis were evaluated. Based on these studies, we recommend improved experimental methods that aim to ensure a consistent framework to identify the roles that exosomes play. With these improved methods, the second part is to characterize the immunosuppressive role that melanoma exosomes play, especially from the perspective of delivering a payload of mRNAs to immune cells. Toward this second aim, melanoma exosomes were purified and cytokine receptor IL12Rbeta2 and specific mRNA enrichment were identified. Microarray and pathway analysis suggested that mRNAs derived from melanoma impact a variety of immune signaling pathways. Induction effects of PTPN11 and DNMT3A from the exosomal mRNAs were characterized in T lymphocytes. Specifically, we showed that PTPN11 upregulation impeded CTLL-2 cytotoxic T cell proliferation in response to IL2 stimulation, and DNMT3A upregulation hindered IFN-gamma production in 2D6 TH1 cells. These findings provide insights regarding the specific immunosuppression effects that tumor-infiltrating lymphocytes (TILs) may encounter in tumor microenvironment. Understanding those immunosuppression effects is important to engineer anti-tumor immunity for innovative and improved treatments against cancer.
Wu, Yueting, "Cancer exosomes are unique and complex mechanisms that suppress effector T lymphocyte functions" (2015). Graduate Theses, Dissertations, and Problem Reports. 6982.