The presence of CD8 T cell (TCD8) in tumors is a positive indicator of patient survival. It has also become clear that patients who respond clinically to new generation immunotherapies are those in which an immunological tumor infiltrate is already evident prior to treatment. On the other hand, the fraction of adoptively transferred tumor-specific effectors that infiltrate tumors is surprisingly small, necessitating in vitro expansion of large numbers of T cells in conjunction with lymphodepletion to support further expansion in vivo. Thus, limited infiltration of T cells is a major barrier to tumor contro over and above the hurdle of immunosuppression. This is an application to understand the basis for this limited infiltration and seek ways to overcome it. Homing receptors (HR) on T cells and their corresponding vascular ligands are essential for TCD8 effectors to infiltrate peripheral tissues. However, we lack a full understanding of which HR control migration into tumors, how this is influenced by expression of the corresponding ligands on tumor vasculature, and the extent to which these same HR also enable infiltration into normal tissues, potentially in competition with tumor infiltration. In addition, the retention of TCD8 effectors after extravasatin may be determined by their ability to recognize Ag on tumor cells or intratumoral Ag presenting cells, or by the action of retention integrins. Most studies of intratumoral T cells have relied on flow cytometry, histology, and blockade or knockout approaches to investigate some of these issues at single time points. Here, we propose to use a new imaging technique, Fluorescence Molecular Tomography (FMT), to longitudinally monitor TCD8 in tumor bearing animals in real time. FMT has been used to visualize TCD8 in vivo in only a single preliminary study. Thus, some aspects of this application are designed to optimize the use of FMT, with the expectation that it will become widely used. The more fundamental aspects of the application will provide greater understanding of the trafficking, distribution and dynamics of T cell entry and exit into tumors and other peripheral tissues;the importance of homing receptors (HR) on T cells, their corresponding vascular ligands, and antigen in promoting tumor entry and retention;and the role of the tumor microenvironment in controlling these processes. This application is a collaboration between two investigators with complementary intellectual expertise in tumor immunology and tumor vascular biology and technical expertise in cellular immunology / single cell analysis by multi-color flow cytometry and advanced whole animal imaging techniques.
The specific aims of this application are: 1) To establish a system for long-term measurements of T cell distribution and trafficking using FMT;2) To investigate the distribution and dynamics of TCD8 effectors in tumor bearing mice;and 3) To establish the role of inflammation, endogenous TCD8 effectors, and Treg in controlling TCD8 infiltration into tumors and the properties of the tumor- associated vasculature. We expect the results from the work proposed in this application will support a future multi-investigator R01 or P01 project application in which such comprehensive studies will be undertaken.
CD8 T cell (TCD8) infiltration into tumors is a positive prognostic indicator of patient survival, and recent successes in controlling tumors by adoptive transfer of activated tumor-specific T cells validate its importance. Indeed, it has become clear that the patients who respond clinically to new generation immunotherapies are those in which an immunological infiltrate is already evident prior to treatment. Thus, limited infiltration of immune cells represents a major barrier to tumor control above the hurdle created by immunosuppression. The work proposed in this application will provide basic mechanistic understanding of the factors that limit T cell infiltration into tumors, and point towards strategis that may be used to enhance this process for improved therapy in cancer patients.