Repetitive noninvasive monitoring of HSV1-tk-expressing T cells intravenously infused into nonhuman primates using positron emission tomography and computed tomography with 18F-FEAU

Abstract

Adoptive transfer of antigen-specific cytotoxic T lymphocytes (CTLs) has been successfully used to treat patients with different types of cancer. However, the long-term spatial-temporal dynamics of the distribution of systemically infused CTLs remains largely unknown. Noninvasive imaging of adoptively transferred CTLs using molecular-genetic reporter imaging with positron emission tomography and computed tomography (PET-CT) represents an innovative approach to understanding the long-term migratory patterns and therapeutic potential of adoptively transferred T cells. Here we report the application of repetitive PET-CT imaging with [18F]fluoro-5-ethyl-1-beta-D-arabinofuranosyluracil (18F-FEAU) in two nonhuman primates demonstrating that autologous polyclonal macaque T lymphocytes activated and transduced with a retroviral vector encoding for the sr39 mutant herpes simplex virus 1 thymidine kinase (sr39HSV1-tk) reporter gene can be detected after intravenous infusion in discrete lymphoid organs and in sites of inflammation. This study represents a proof of principle and supports the application of 18F-FEAU PET-CT imaging for monitoring the distribution of intravenously administered sr39HSV1-tk gene-transduced CTLs in humans.

Figure 1

Repetitive PET-CT images of ¹⁸F-FEAU distribution in the rhesus macaque 1 before and after injection of sr39HSV1-tk⁺ autologous T lymphocytes. A illustrates the percentage of LNGFR⁺ T lymphocytes, as assessed by FACS analysis using a monoclonal antibody detecting NGFR (Becton-Dickinson), 7 days after retroviral transduction and at the time of infusion after selection using LNGFR microbeads. B illustrates the uptake of [³H]-FEAU by sr39HSV1-tk⁺ T lymphocytes. Each assay was performed in triplicate. Sr39HSV1-tk⁺ T cells exhibited a cell to medium uptake ratio of 40.6, 24.5× higher than the control. Columns represent means; bars indicate ± standard deviation. C to N illustrate the whole-body PET scan 60 minutes after the tracer injection. PET images were reconstructed, analyzed, and normalized as previously described. C, D, and E represent maximum-intensity three-dimensional projections of whole-body PET images collected before T-cell infusion (C) and 90 minutes (D) and 7 days (E) after T-cell infusion. F, G, H, I, J, and K demonstrate the differences in ¹⁸F-FEAU accumulation in the parotid glands and cervical lymph nodes between baseline (F, I), 90 minutes (G, J), and 7 days (H, K) after T-cell administration. L, M, and N represent coronal PET or PET-CT images through the area of target lesions in the left thigh visible at 90 minutes after T-cell administration and 60 minutes after ¹⁸F-FEAU injection (M), which is absent in the baseline image (L) and 7 days later (N).

Figure 2

Repetitive PET-CT images of ¹⁸F-FEAU distribution in the rhesus macaque 2 before and after injection of sr39HSV1-tk⁺ autologous T lymphocytes. A illustrates the characteristics of the T-cell product as described in Figure 1. B illustrates the uptake of [³H]-FEAU by sr39HSV1-tk⁺ T lymphocytes. Sr39HSV1-tk⁺ T cells exhibited a cell to medium uptake ratio of 45.2, 32.2 × higher than the control. Columns represent means; bars indicate ± standard deviation. C to F represent maximum-intensity three-dimensional projections of whole-body PET images obtained at baseline (C) and 90 minutes (D), 1 day (E), and 7 days (F) after T-cell infusion. G to J represent transaxial PET-CT images through the area of posterior cervical lymph nodes at each time point.

Figure 3

Detection of sr39HSV1-tk⁺ T lymphocytes in peripheral blood and in a posterior neck lymph node biopsy. A illustrates the detection of circulating NGFR⁺ T cells as assessed by phenotypic analysis in blood samples collected before and after T-cell infusion. B illustrates the detection of sr39HSV-tk⁺ expression by polymerase chain reaction in peripheral blood mononuclear cells (PBMCs) preinfusion, 4 days postinfusion, and 7 days postinfusion and in a posterior neck lymph node biopsy sample obtained 7 days after T-cell infusion. Ex vivo expanded T nontransduced cells and ex vivo expanded sr39HSV1-tk⁺ T cells were also used as negative and positive control, respectively.