Ex Vivo Radiolabeling and In Vivo PET Imaging of T Cells Expressing Nuclear Reporter Genes

  • Maxim A. Moroz
  • Pat Zanzonico
  • Jason T. Lee
  • Vladimir PonomarevEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 1790)


Recent advances in T cell-based immunotherapies from bench to bedside have highlighted the need for improved diagnostic imaging of T cell trafficking in vivo and the means to noninvasively investigate failures in treatment response. T cells expressing tumor-associated T cell receptors (TCRs) or engineered with chimeric antigen receptors (CARs) face multiple challenges, including possible influence of genetic engineering on T cell efficacy, inhibitory effects of the tumor microenvironment, tumor checkpoint proteins and on-target, off-tissue toxicities (Kershaw et al., Nat Rev Cancer 13:525–541, 2013; Corrigan-Curay et al., Mol Ther 22:1564–1574, 2014; June et al., Sci Trans Med 7:280–287, 2015; Whiteside et al., Clin Cancer Res 22:1845–1855, 2016; Rosenberg and Restifo, Science 348:62–68, 2015). Positron emission tomography (PET) imaging with nuclear reporter genes is potentially one of the most sensitive and noninvasive methods to quantitatively track and monitor function of adoptively transferred cells in vivo. However, in vivo PET detection of T cells after administration into patients is limited by the degree of tracer accumulation per cell in situ and cell density in target tissues. We describe here a method for ex vivo radiolabeling of T cells, a reliable and robust technique for PET imaging of the kinetics of T cell biodistribution from the time of administration to subsequent localization in targeted tumors and other tissues of the body. This noninvasive technique can provide valuable information to monitor and identify the potential efficacy of adoptive cell therapies.

Key words

T cell Immunotherapy Adoptive cell therapy PET Human nuclear reporter gene Reporter probe Ex vivo radiolabeling Tumor microenvironment 



This work was supported by NIH P50 CA86438, R01 CA163980 and R01 CA161138 grants, Mr. William H. Goodwin and Mrs. Alice Goodwin and the Commonwealth Foundation for Cancer Research and The Experimental Therapeutics Center of Memorial Sloan Kettering Cancer Center, NIH Small-Animal Imaging Research Program (SAIRP), NIH Shared Instrumentation Grant No 1 S10 RR020892-01, NIH Shared Instrumentation Grant No 1 S10 RR028889-01, and NIH Center Grants P30 CA08748 and P30 CA08748.


  1. 1.
    Asperti-Boursin F, Real E, Bismuth G, Trautmann A, Donnadieu E (2007) CCR7 ligands control basal T cell motility within lymph node slices in a phosphoinositide 3-kinase-independent manner. J Exp Med 204:1167–1179CrossRefPubMedPubMedCentralGoogle Scholar
  2. 2.
    Salmon H, Rivas-Caicedo A, Asperti-Boursin F, Lebugle C, Bourdoncle P, Donnadieu E (2011) Ex vivo imaging of T cells in murine lymph node slices with widefield and confocal microscopes. J Vis Exp: JoVE (53):e3054Google Scholar
  3. 3.
    Ahrens ET, Bulte JW (2013) Tracking immune cells in vivo using magnetic resonance imaging. Nat Rev Immunol 13:755–763CrossRefPubMedGoogle Scholar
  4. 4.
    Koehne G, Doubrovin M, Doubrovina E et al (2003) Serial in vivo imaging of the targeted migration of human HSV-TK-transduced antigen-specific lymphocytes. Nat Biotechnol 21:405–413CrossRefPubMedGoogle Scholar
  5. 5.
    Adonai N, Nguyen KN, Walsh J et al (2002) Ex vivo cell labeling with 64Cu-pyruvaldehyde-bis(N4-methylthiosemicarbazone) for imaging cell trafficking in mice with positron-emission tomography. Proc Natl Acad Sci U S A 99:3030–3035CrossRefPubMedPubMedCentralGoogle Scholar
  6. 6.
    Bansal A, Pandey MK, Demirhan YE et al (2015) Novel (89)Zr cell labeling approach for PET-based cell trafficking studies. EJNMMI Res 5:19CrossRefPubMedPubMedCentralGoogle Scholar
  7. 7.
    Liu Z, Li Z (2014) Molecular imaging in tracking tumor-specific cytotoxic T lymphocytes (CTLs). Theranostics 4:990–1001CrossRefPubMedPubMedCentralGoogle Scholar
  8. 8.
    Yaghoubi SS, Campbell DO, Radu CG, Czernin J (2012) Positron emission tomography reporter genes and reporter probes: gene and cell therapy applications. Theranostics 2:374–391CrossRefPubMedPubMedCentralGoogle Scholar
  9. 9.
    Zanzonico P, Koehne G, Gallardo HF et al (2006) [131I]FIAU labeling of genetically transduced, tumor-reactive lymphocytes: cell-level dosimetry and dose-dependent toxicity. Eur J Nucl Med Mol Imaging 33:988–997CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  • Maxim A. Moroz
    • 1
  • Pat Zanzonico
    • 1
  • Jason T. Lee
    • 1
  • Vladimir Ponomarev
    • 1
    Email author
  1. 1.Memorial Sloan Kettering Cancer CenterNew YorkUSA

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