Evaluate the role of extracellular vesicles in tumor microenvironment modulation

PROJECT SUMMARY In the United States, it is estimated that >100,000 people will die from non-small cell lung cancer (NSCLC) in 2024. Emerging targeted therapies, such as checkpoint immunotherapies have resulted in significant success; still, 50% of patients treated with anti-PD1 or anti-CTLA4 have poor clinical efficacy. The effectiveness of immunotherapies has been strongly correlated with the abundance of immune T cells in the tumor stroma. Indeed, increased T cell infiltration is a strong prognostic indicator across 17 solid cancer types, including NSCLC. Nonetheless, immunotherapies have largely remained ineffective in tumors that have reduced T cell infiltration, also called immune-cold tumors. Therefore, understanding the mechanism(s) that lead to reduced tumor infiltrating T cells is of significant interest in the treatment of immunosuppressive NSCLC. Despite an extensive understanding of T cell suppression mechanisms in immune-cold NSCLC, current therapies have shown modest clinical benefit, indicating that additional mechanisms contribute to this process. One such emerging mechanism is tumor-derived extracellular vesicles (EVs), however, identification of specific EV subpopulations responsible for T cell suppression has remained a challenge. Overcoming this challenge, we have identified a specific subpopulation of NSCLC-derived EVs, called TGN46+ EVs, that strongly inhibit the proliferation of T cells. In an immunosuppressive NSCLC model (H358), total H358 EVs inhibit T cell proliferation; however, specifically depleting TGN46+ EVs from total H358 EVs significantly reduces their ability to suppress T cells. Our findings are clinically relevant, and we have further verified them in two independent NSCLC models as well as in prostate cancer, which are well-known to form immune-cold tumors. Given these findings, we hypothesize that TGN46+ EVs play a key role in modulating T cell tumor infiltration in immune-cold tumors. In the proposed work, we plan to evaluate how this single EV subpopulation, i.e. TGN46+ EVs, influences T cell infiltration and overall tumor microenvironment. To evaluate this, we will use (i) a humanized mouse model that supports the development of the human immune system and allows the study of human-specific tumor-immune interactions; (ii) a human NSCLC model (HCC827) that exhibits high T cell infiltration in humanized mice, and (iii) EVs from an immunosuppressive NSCLC model (H358). We will evaluate how T cell infiltration and abundance of other immune cell types in HCC827 tumors change upon introduction of (i) total H358 EVs (which includes TGN46+ EVs), or (ii) H358 EVs depleted of TGN46+ EV subpopulation, in the tumor stroma. Following the successful completion of this study, we anticipate that we will have evaluated EV-specific modulation of the tumor microenvironment and its physiological implications. These accomplishments would have an enormous positive impact on our understanding of intercellular communication in immune-cold NSCLC. In the long term, these insights have the potential to inform the development of effective strategies to overcome immunotherapy resistance, thereby benefiting a considerable number of lung cancer patients.