Immunotherapy is a promising strategy for cancer patients, particularly for those nonresponsive to more conventional therapeutic regimens. Quantitative imaging of immune evasion biomarkers can help identify patients responsive to immunotherapy. Chemical exchange saturation transfer magnetic resonance imaging (CEST MRI) is a molecular imaging technology that is already being translated to humans. CEST MRI has the unique ability to monitor multiple agents simultaneously through selectively labeling their exchangeable protons with radiofrequency saturation, which can be encoded as “colors” similar to optical methods. We propose to develop (1) agents with unique colors and (2) analysis methods to quantify these agents and generate multicolor MR images. This award will support Dr. Aline Thomas in her transition to an independent research career. Candidate Training: Dr. Thomas has a background in biomedical engineering, investigating factors contributing to pathophysiology and regeneration failure in disease settings with an immune component using material, computational, genetic (therapeutic and reporter genes), and through collaboration, conventional MRI methods. With the guidance of her mentors, Dr. Thomas will learn how to develop clinically translatable CEST agents and quantify their targets using post-processing methods to probe multiple cellular processes in vivo in order to monitor cell behavior in disease and in response to therapy. She will also be trained in the practical aspects of a leading a research program—lab management, trainee mentorship, conference presentation, networking and fundraising—which are critical to her success as an independent investigator in the field of Molecular Imaging. The proposed work will lead to the preparation of a R01 grant proposal. Research Plan: A recognized mechanism of cancer to evade immunological attack is glutamine transport to deactivate (via PD- L1) and to deprive energy (independent of PD-L1) from infiltrating immune cells. The central hypothesis is that quantitative imaging of immune evasion mechanisms using CEST MRI can ultimately be used to help predict patient response to immunotherapy. Dr. Thomas proposes to develop CEST MRI agents and detection methods that can quantify glutamine transport and PD-L1 expression simultaneously. She will also develop post-processing methods to quantify these agents. To validate their potential as complementary imaging biomarkers of cancer immune evasion, she will evaluate their uptake in prostate cancer cell lines (in vitro; cancerous: PC3, LNCaP, DU-145; noncancerous: RWPE-1 and RWPE-2) and in a prostate tumor model (in vivo) and compare their uptake to conventional immune suppression (PD-L1 histology, flow cytometry, cytokines) and immune evasion (surface marker expression, tumor infiltration of leukocytes) metrics. Aim 1: Develop methods to visualize and quantify PD-L1 expression in prostate cancer. Aim 2: Develop methods to visualize and quantify glutamine t...