Chengyi J. Shu

Molecular imaging of lymphoid organs and immune activation by positron emission tomography with a new [18F]-labeled 2'-deoxycytidine analog.

Monitoring immune function with molecular imaging could have a considerable impact on the diagnosis and treatment evaluation of immunological disorders and therapeutic immune responses. Positron emission tomography (PET) is a molecular imaging modality with applications in cancer and other diseases. PET studies of immune function have been limited by a lack of specialized probes. We identified [18F]FAC (1-(2′-deoxy-2′-[18F]fluoroarabinofuranosyl) cytosine) by differential screening as a new PET probe for the deoxyribonucleotide salvage pathway. [18F]FAC enabled visualization of lymphoid organs and was sensitive to localized immune activation in a mouse model of antitumor immunity. [18F]FAC microPET also detected early changes in lymphoid mass in systemic autoimmunity and allowed evaluation of immunosuppressive therapy. These data support the use of [18F]FAC PET for immune monitoring and suggest a wide range of clinical applications in immune disorders and in certain types of cancer.

Positron emission tomography with computed tomography imaging of neuroinflammation in experimental autoimmune encephalomyelitis

2-[18F]Fluoro-2-deoxy-d-glucose positron emission tomography ([18F]FDG PET) detection of the up-regulated glycolysis associated with malignant transformation is a noninvasive imaging technique used extensively in cancer diagnosis. Although striking similarities exist in glucose transport and metabolism between tumor cells and activated immune cells, the potential use of [18F]FDG PET for the diagnosis and evaluation of autoimmune disorders has not been systematically investigated. Here we ask whether [18F]FDG PET in conjunction with computed tomography (CT) could be used to monitor a complex autoimmune disorder such as murine experimental autoimmune encephalomyelitis (EAE) and whether this approach is sensitive enough to evaluate therapeutic interventions. We found that (i) coregistration of metabolic (i.e., microPET) and high-resolution anatomical (i.e., CT) images allows serial quantification of glycolysis with [18F]FDG in various spinal column segments; (ii) [18F]FDG PET/CT can detect the increased glycolysis associated with paralysis-causing inflammatory infiltrates in the spinal cord; and (iii) the [18F]FDG measure of glycolysis in the spinal cord is sensitive to systemic immunosuppressive therapy. These results highlight the potential use of serial [18F]FDG PET/CT imaging to monitor neuroinflammation in EAE and suggest that similar approaches could be applied to the diagnosis and evaluation of other autoimmune and inflammatory disorders in animal models and in humans.

Anti-tumor activity and trafficking of self, tumor-specific T cells against tumors located in the brain.

It is commonly believed that T cells have difficulty reaching tumors located in the brain due to the presumed “immune privilege” of the central nervous system (CNS). Therefore, we studied the biodistribution and anti-tumor activity of adoptively transferred T cells specific for an endogenous tumor-associated antigen (TAA), gp100, expressed by tumors implanted in the brain. Mice with pre-established intracranial (i.c.) tumors underwent total body irradiation (TBI) to induce transient lymphopenia, followed by the adoptive transfer of gp10025–33-specific CD8+ T cells (Pmel-1). Pmel-1 cells were transduced to express the bioluminescent imaging (BLI) gene luciferase. Following adoptive transfer, recipient mice were vaccinated with hgp10025–33 peptide-pulsed dendritic cells (hgp10025–33/DC) and systemic interleukin 2 (IL-2). This treatment regimen resulted in significant reduction in tumor size and extended survival. Imaging of T cell trafficking demonstrated early accumulation of transduced T cells in lymph nodes draining the hgp10025–33/DC vaccination sites, the spleen and the cervical lymph nodes draining the CNS tumor. Subsequently, transduced T cells accumulated in the bone marrow and brain tumor. BLI could also detect significant differences in the expansion of gp100-specific CD8+ T cells in the treatment group compared with mice that did not receive either DC vaccination or IL-2. These differences in BLI correlated with the differences seen both in survival and tumor infiltrating lymphocytes (TIL). These studies demonstrate that peripheral tolerance to endogenous TAA can be overcome to treat tumors in the brain and suggest a novel trafficking paradigm for the homing of tumor-specific T cells that target CNS tumors.

Novel PET Probes Specific for Deoxycytidine Kinase

Deoxycytidine kinase (dCK) is a rate-limiting enzyme in the deoxyribonucleoside salvage pathway and a critical determinant of therapeutic activity for several nucleoside analog prodrugs. We have previously reported the development of 1-(2′-deoxy-2′-18F-fluoro-β-d-arabinofuranosyl)cytosine (18F-FAC), a new probe for PET of dCK activity in immune disorders and certain cancers. The objective of the current study was to develop PET probes with improved metabolic stability and specificity for dCK. Toward this goal, several candidate PET probes were synthesized and evaluated in vitro and in vivo. Methods: High-pressure liquid chromatography was used to analyze the metabolic stability of 18F-FAC and several newly synthesized analogs with the natural d-enantiomeric sugar configuration or the corresponding unnatural l-configuration. In vitro kinase and uptake assays were used to determine the affinity of the 18F-FAC l-nucleoside analogs for dCK. The biodistribution of selected l-analogs in mice was determined by small-animal PET/CT. Results: Candidate PET probes were selected using the following criteria: low susceptibility to deamination, high affinity for purified recombinant dCK, high uptake in dCK-expressing cell lines, and biodistribution in mice reflective of the tissue-expression pattern of dCK. Among the 10 newly developed candidate probes, 1-(2′-deoxy-2′-18F-fluoro-β-l-arabinofuranosyl)cytosine (l-18F-FAC) and 1-(2′-deoxy-2′-18F-fluoro-β-l-arabinofuranosyl)-5-methylcytosine (l-18F-FMAC) most closely matched the selection criteria. The selection of l-18F-FAC and l-18F-FMAC was validated by showing that these two PET probes could be used to image animal models of leukemia and autoimmunity. Conclusion: Promising in vitro and in vivo data warrant biodistribution and dosimetry studies of l-18F-FAC and l-18F-FMAC in humans.

Quantitative PET reporter gene imaging of CD8+ T cells specific for a melanoma-expressed self-antigen

Adoptive transfer (AT) T-cell therapy provides significant clinical benefits in patients with advanced melanoma. However, approaches to non-invasively visualize the persistence of transferred T cells are lacking. We examined whether positron emission tomography (PET) can monitor the distribution of self-antigen-specific T cells engineered to express an herpes simplex virus 1 thymidine kinase (sr39tk) PET reporter gene. Micro-PET imaging using the sr39tk-specific substrate 9-[4-[18F]fluoro-3-(hydroxymethyl)-butyl]guanine ([18F]FHBG) enabled the detection of transplanted T cells in secondary lymphoid organs of recipient mice over a 3-week period. Tumor responses could be predicted as early as 3 days following AT when a >25-fold increase of micro-PET signal in the spleen and 2-fold increase in lymph nodes (LNs) were observed in mice receiving combined immunotherapy versus control mice. The lower limit of detection was ∼7 × 105 T cells in the spleen and 1 × 104 T cells in LNs. Quantification of transplanted T cells in the tumor was hampered by the sr39tk-independent trapping of [18F]FHBG within the tumor architecture. These data support the feasibility of using PET to visualize the expansion, homing and persistence of transferred T cells. PET may have significant clinical utility by providing the means to quantify anti-tumor T cells throughout the body and provide early correlates for treatment efficacy.

Non‐invasive imaging of adaptive immunity using positron emission tomography

Summary: Non‐invasive monitoring of adaptive immunity in infection, cancer, and autoimmunity remains a major challenge. Current techniques to monitor lymphocytes involve numeric and functional determinations of immune cells isolated from the peripheral blood (most often) and tissue (rarely). Invasive measurements are prone to sampling errors and are poorly reflective of the dynamic changes in the location, number, and movement of lymphoid cells. These limitations indicate the need for non‐invasive whole‐body imaging methodologies that allow longitudinal, quantitative, and functional analyses of the immune system in vivo. Positron emission tomography (PET), a clinically based whole‐body imaging modality, has the potential to revolutionize diagnostics and therapeutic monitoring in both clinical and pre‐clinical settings. This review discusses studies using PET to image adaptive immune responses in small animal models. We address the challenges inherent in assessing whole‐body immunity with PET and recent developments that can improve its performance. Finally, we discuss work to translate PET immune imaging into clinical practice.

Design and characterization of a biomedical device capable of pico-CI level beta detection for the study of cell metabolism

A new system is being developed to allow imaging of charged particles in microfluidic chip using position sensitive avalanche photodiodes (PSAPD). The new imaging system was created by placing the PSAPD detector with close proximity to the PDMS microchip. The new system therefore is capable of quantifying tiny amounts of these radiolabeled probes over time. The PSAPD can be used for the direct detection of positrons and other charged particles. In our studies, the performance of the system has been carried out. A gradient of solution containing radioactive positron emitting fluorodeoxyglucose (FDG) was imaged from a microchip. Results have shown that sensitivities as low as 56 plusmn 0.1 pCi in a 4 mm2 region of interest (ROI) (14 pCi/mm2) and line pair spatial resolution of 0.5 mm can be achieved. Moreover, the application to cell biology was successfully performed in which currently radioactive signal from less than one hundred cells can be detected and resolved clearly in the system.