(4gj) Deciphering Immune Cell Signaling Pathways and Transcriptome Responses to Colorectal Cancer-Derived Extracellular Vesicles

Excreted lipid-based vesicles—known as extracellular vesicles (EVs), by aggressive cancer cells have been shown to aid in metastatic dissemination and immune suppression. Once thought to be filled with cellular waste, EVs (comprising both exosomes, 50-500 nm, and microvesicles, up to 1-2 microns) can serve as endogenous delivery vehicles for amplified oncogenic cargo (mRNA, miRNA, ncRNA, DNA, etc.), pro-survival proteins, and enzymes originating from the parental organ. The broad class of colloidally stable nanoparticle-like vesicles, called exosomes, can promote tumor expansion and metastatic spread, immune suppression, chemosurvivability, and even an increase in vascular permeability. The relationship between the different size populations of EVs and their signaling influences on immune cells, however, is still not well understood. In this work, I will report differences between the responses of peripheral blood lymphocyte and solid tumor infiltrating lymphocyte models exposed to EVs, and how EVs inhibit CD8⁺ T cells from attacking tumor cells. First, comparative transcriptomic analyses from bulk mRNA of cells exposed to either microvesicles or exosomes will highlight genomic differences between these two EV subsets and how the differences affect peripheral blood lymphocytes (T cells, B cells, and Natural Killer (NK) Cells). Next, a dose-dependent analysis of various compositions of EVs incubated with peripheral blood mononuclear cells (comprising lymphocytes, monocytes, and dendritic cells) will be considered using a single-cell mRNA sequencing platform (Seq-Well). These data will reveal transcriptomic responses to the EVs among individual subsets of cells as well as cell-cell EV based communication pathways and their effects on lymphocyte suppression. Finally, 3D adenocarcinoma organoid cell culture systems, which recapitulate morphological and genetic features of the original tumor, to mimic tumor immune microenvironments will be evaluated. Such systems will provide insight on how vesicles send signals through tumor tissues and model degree of invasion of lymphocytes after encountering EVs. The data together will reveal new insights to signaling pathways mediated by EVs and suggest new hypotheses for which immunotherapies or engineered nanomedicines can be utilized to circumvent tumor growth and inhibit metastatic spread.