Project Summary Immunotherapy has tremendous promise for eradicating cancers. One approach to boost immune response to attack cancer is adoptive T cell transfer (ACT) using tumor-specific cytotoxic T cells. ACT cancer immunotherapy has demonstrated tremendous success against melanoma, leukemia, and lymphoma, but application to other solid tumors requires new developments. Despite decades of research, prognosis for patients with malignant brain tumors is extremely poor. Although these patients account for only ~2% of new cases, they are the fourth highest cause of cancer mortality in the USA. A critical step for the success of ACT immunotherapy in solid cancers is achieving trafficking and persistence of T cells at tumor sites, while avoiding toxicities due to T cell attack of off-target tissues and organs. As such, there is a real and present need to understand the location of adoptively transferred T cells. This Parent F31 – Diversity Fellowship proposal seeks to establish application of Magnetic Particle Imaging (MPI) to enable the non-invasive and quantitative monitoring of tumor accumulation and whole-body distribution of adoptively transferred T cells used to treat brain cancer. MPI is a new molecular imaging modality that enables non-invasive, unambiguous, and tomographic analysis of the whole-body distribution of magnetic tracers, typically consisting of biocompatible superparamagnetic iron oxide (SPIO) nanoparticles. The signal in MPI is proportional to tracer mass, does not suffer from tissue penetration depth limitations, and there is no body background signal, making quantification of tracer distribution straightforward. Furthermore, cell tracking sensitivity of MPI is already better than with other whole-body imaging modalities. This proposal seeks to optimize T cell labeling using magnetic nanoparticles for the non- invasive in vivo tracking of adoptively transferred T cells. Preliminary studies demonstrated that labeling T cells with the commercial MPI tracer ferucarbotran has no deleterious effects on T cell viability, effector phenotype, or anti-cancer cytotoxic activity. Furthermore, SPIO- labeled T cells accumulated in the brain, as detected non-invasively in vivo using MPI. However, improvements are needed to achieve the full potential of MPI for T cell tracking. The proposed research builds on this success through two specific aims. Research in AIM 1 will optimize conditions for fast and efficient labeling of T cells using commercially available SPIOs that outperform ferucarbotran in MPI sensitivity. Research in AIM 2 will demonstrate the non-invasive in vivo tracking of adoptive T cell cancer immunotherapy using MPI. By enabling non-invasive and quantitative evaluation of adoptive T cell biodistribution and accumulation in tumors, the proposed research has the potential to greatly accelerate the development of safer and more effective adoptive T cell immunotherapy treatments for brain cancer.