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Genetic Engineering of Therapeutic Cells with the Sodium Iodide Symporter (NIS) to Enable Noninvasive In Vivo Therapy Tracking

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Positron Emission Tomography

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2729))

Abstract

Noninvasive long-term imaging of therapeutic cells in preclinical models can be achieved through introducing a reporter gene into the cells of interest. Despite important recent developments such as gene editing, cell engineering based on lentiviruses remains a mainstream tool for gene transfer applicable to a variety of different cell types.

In this chapter, we describe how to use lentivirus-based genetic engineering to render different candidate cell therapies in vivo traceable by radionuclide imaging. We illustrate this reporter gene technology using the sodium iodide symporter (NIS), which is compatible with both positron emission tomography (PET) and single-photon emission computed tomography (SPECT). For preclinical experimentation, we fused NIS with a suitable fluorescent protein such as monomeric GFP or RFP to streamline cell line generation and downstream analyses of ex vivo tissue samples. We present protocols for reporter gene engineering of human cardiac progenitor cells, regulatory T cells, and effector T cells as well as for the characterization experiments required to validate NIS-fluorescent protein reporter function in these candidate therapeutic cells.

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References

  1. Caplan AI (2017) Mesenchymal stem cells: time to change the name! Stem Cells Transl Med 6(6):1445–1451. https://doi.org/10.1002/sctm.17-0051

    Article  Google Scholar 

  2. Heathman TR, Nienow AW, McCall MJ, Coopman K, Kara B, Hewitt CJ (2015) The translation of cell-based therapies: clinical landscape and manufacturing challenges. Regen Med 10(1):49–64. https://doi.org/10.2217/rme.14.73

    Article  CAS  Google Scholar 

  3. Bioinformant (2019) Bioinformant cell therapy industry database. BioInformant. https://bioinformant.com/product/cell-therapy-industry-database/. Accessed 3 Oct 2019

  4. Ashmore-Harris C, Iafrate M, Saleem A, Fruhwirth GO (2020) Non-invasive reporter gene imaging of cell therapies, including T cells and stem cells. Mol Ther. https://doi.org/10.1016/j.ymthe.2020.03.016

  5. Krekorian M, Fruhwirth GO, Srinivas M, Figdor CG, Heskamp S, Witney TH, Aarntzen E (2019) Imaging of T-cells and their responses during anti-cancer immunotherapy. Theranostics 9(25):7924–7947. https://doi.org/10.7150/thno.37924

    Article  CAS  Google Scholar 

  6. Iafrate M, Fruhwirth GO (2020) How non-invasive in vivo cell tracking supports the development and translation of cancer immunotherapies. Front Physiol 11:154. https://doi.org/10.3389/fphys.2020.00154

    Article  Google Scholar 

  7. Jacob J, Volpe A, Peng Q, Lechler RI, Smyth LA, Lombardi G, Fruhwirth GO (2023) Radiolabelling of Polyclonally Expanded Human Regulatory T Cells (Treg) with 89Zr-oxine for Medium-Term In Vivo Cell Tracking Molecules 28:1482. https://doi.org/10.3390/molecules28031482

  8. Keu KV, Witney TH, Yaghoubi S, Rosenberg J, Kurien A, Magnusson R, Williams J, Habte F, Wagner JR, Forman S, Brown C, Allen-Auerbach M, Czernin J, Tang W, Jensen MC, Badie B, Gambhir SS (2017) Reporter gene imaging of targeted T cell immunotherapy in recurrent glioma. Sci Transl Med 9(373). https://doi.org/10.1126/scitranslmed.aag2196

  9. Fruhwirth GO, Diocou S, Blower PJ, Ng T, Mullen GE (2014) A whole-body dual-modality radionuclide optical strategy for preclinical imaging of metastasis and heterogeneous treatment response in different microenvironments. J Nucl Med 55(4):686–694. https://doi.org/10.2967/jnumed.113.127480

    Article  CAS  Google Scholar 

  10. Volpe A, Man F, Lim L, Khoshnevisan A, Blower J, Blower PJ, Fruhwirth GO (2018) Radionuclide-fluorescence reporter gene imaging to track tumor progression in rodent tumor models. J Vis Exp 133(133):e57088. https://doi.org/10.3791/57088

    Article  CAS  Google Scholar 

  11. Ashmore-Harris C, Blackford SJ, Grimsdell B, Kurtys E, Glatz MC, Rashid TS, Fruhwirth GO (2019) Reporter gene-engineering of human induced pluripotent stem cells during differentiation renders in vivo traceable hepatocyte-like cells accessible. Stem Cell Res 41:101599. https://doi.org/10.1016/j.scr.2019.101599

    Article  CAS  Google Scholar 

  12. Kang JH, Lee DS, Paeng JC, Lee JS, Kim YH, Lee YJ, Hwang DW, Jeong JM, Lim SM, Chung JK, Lee MC (2005) Development of a sodium/iodide symporter (NIS)-transgenic mouse for imaging of cardiomyocyte-specific reporter gene expression. J Nucl Med 46(3):479–483

    CAS  Google Scholar 

  13. Jacob J, Nadkarni S, Volpe A, Peng Q, Tung SL, Hannen RF, Mohseni YR, Scotta C, Marelli-Berg FM, Lechler RI, Smyth LA, Fruhwirth GO, Lombardi G (2020) Spatiotemporal in vivo tracking of polyclonal human regulatory T cells (Treg) reveals a role for innate immune cells in Treg transplant recruitment. Mol Ther – Methods Clin Dev 20:324–336. https://doi.org/10.1016/j.omtm.2020.12.003

  14. Volpe A, Lang C, Lim L, Man F, Kurtys E, Ashmore-Harris C, Johnson P, Skourti E, de Rosales RTM, Fruhwirth GO (2020) Spatiotemporal PET imaging reveals differences in CAR-T tumor retention in triple-negative breast cancer models. Mol Ther 28(10):2271–2285. https://doi.org/10.1016/j.ymthe.2020.06.028

    Article  CAS  Google Scholar 

  15. O’Doherty J, Jauregui-Osoro M, Brothwood T, Szyszko T, Marsden PK, O’Doherty MJ, Cook GJR, Blower PJ, Lewington V (2017) (18)F-Tetrafluoroborate, a PET probe for imaging sodium/iodide symporter expression: whole-body biodistribution, safety, and radiation dosimetry in thyroid cancer patients. J Nucl Med 58(10):1666–1671. https://doi.org/10.2967/jnumed.117.192252

    Article  CAS  Google Scholar 

  16. Meller J, Becker W (2002) The continuing importance of thyroid scintigraphy in the era of high-resolution ultrasound. Eur J Nucl Med Mol Imaging 29(Suppl 2):S425–S438. https://doi.org/10.1007/s00259-002-0811-8

    Article  Google Scholar 

  17. Goswami R, Subramanian G, Silayeva L, Newkirk I, Doctor D, Chawla K, Chattopadhyay S, Chandra D, Chilukuri N, Betapudi V (2019) Gene therapy leaves a vicious cycle. Front Oncol 9:297. https://doi.org/10.3389/fonc.2019.00297

    Article  Google Scholar 

  18. Ghani K, Boivin-Welch M, Roy S, Dakiw-Piaceski A, Barbier M, Pope E, Germain L, Caruso M (2019) Generation of high-titer self-inactivated gamma-retroviral vector producer cells. Mol Ther Methods Clin Dev 14:90–99. https://doi.org/10.1016/j.omtm.2019.05.013

    Article  CAS  Google Scholar 

  19. Ronald JA, Cusso L, Chuang HY, Yan X, Dragulescu-Andrasi A, Gambhir SS (2013) Development and validation of non-integrative, self-limited, and replicating minicircles for safe reporter gene imaging of cell-based therapies. PLoS One 8(8):e73138. https://doi.org/10.1371/journal.pone.0073138

    Article  CAS  Google Scholar 

  20. Lufino MM, Edser PA, Wade-Martins R (2008) Advances in high-capacity extrachromosomal vector technology: episomal maintenance, vector delivery, and transgene expression. Mol Ther 16(9):1525–1538. https://doi.org/10.1038/mt.2008.156

    Article  CAS  Google Scholar 

  21. Maggio I, Goncalves MA (2015) Genome editing at the crossroads of delivery, specificity, and fidelity. Trends Biotechnol 33(5):280–291. https://doi.org/10.1016/j.tibtech.2015.02.011

    Article  CAS  Google Scholar 

  22. Bressan RB, Dewari PS, Kalantzaki M, Gangoso E, Matjusaitis M, Garcia-Diaz C, Blin C, Grant V, Bulstrode H, Gogolok S, Skarnes WC, Pollard SM (2017) Efficient CRISPR/Cas9-assisted gene targeting enables rapid and precise genetic manipulation of mammalian neural stem cells. Development 144(4):635–648. https://doi.org/10.1242/dev.140855

    Article  CAS  Google Scholar 

  23. Ashmore-Harris C, Fruhwirth GO (2020) The clinical potential of gene editing as a tool to engineer cell-based therapeutics. Clin Transl Med 9(1):15. https://doi.org/10.1186/s40169-020-0268-z

    Article  Google Scholar 

  24. US Food & Drug Administration (2017) FDA approval brings first gene therapy to the United States. https://www.fda.gov/newsevents/newsroom/pressannouncements/ucm574058.htm. Accessed 25 Feb 2018

  25. Abou-El-Enein M, Bauer G, Reinke P, Renner M, Schneider CK (2014) A roadmap toward clinical translation of genetically-modified stem cells for treatment of HIV. Trends Mol Med 20(11):632–642. https://doi.org/10.1016/j.molmed.2014.08.004

    Article  CAS  Google Scholar 

  26. Diocou S, Volpe A, Jauregui-Osoro M, Boudjemeline M, Chuamsaamarkkee K, Man F, Blower PJ, Ng T, Mullen GED, Fruhwirth GO (2017) [(18)F]tetrafluoroborate-PET/CT enables sensitive tumor and metastasis in vivo imaging in a sodium iodide symporter-expressing tumor model. Sci Rep 7(1):946. https://doi.org/10.1038/s41598-017-01044-4

    Article  CAS  Google Scholar 

  27. Zufferey R, Nagy D, Mandel RJ, Naldini L, Trono D (1997) Multiply attenuated lentiviral vector achieves efficient gene delivery in vivo. Nat Biotechnol 15(9):871–875. https://doi.org/10.1038/nbt0997-871

    Article  CAS  Google Scholar 

  28. Lewis-McDougall FC, Ruchaya PJ, Domenjo-Vila E, Shin Teoh T, Prata L, Cottle BJ, Clark JE, Punjabi PP, Awad W, Torella D, Tchkonia T, Kirkland JL, Ellison-Hughes GM (2019) Aged-senescent cells contribute to impaired heart regeneration. Aging Cell 18(3):e12931. https://doi.org/10.1111/acel.12931

    Article  CAS  Google Scholar 

  29. Scalise M, Torella M, Marino F, Ravo M, Giurato G, Vicinanza C, Cianflone E, Mancuso T, Aquila I, Salerno L, Nassa G, Agosti V, De Angelis A, Urbanek K, Berrino L, Veltri P, Paolino D, Mastroroberto P, De Feo M, Viglietto G, Weisz A, Nadal-Ginard B, Ellison-Hughes GM, Torella D (2020) Atrial myxomas arise from multipotent cardiac stem cells. Eur Heart J 41(45):4332–4345. https://doi.org/10.1093/eurheartj/ehaa156

    Article  CAS  Google Scholar 

  30. Boardman DA, Philippeos C, Fruhwirth GO, Ibrahim MA, Hannen RF, Cooper D, Marelli-Berg FM, Watt FM, Lechler RI, Maher J, Smyth LA, Lombardi G (2017) Expression of a chimeric antigen receptor specific for donor HLA class I enhances the potency of human regulatory T cells in preventing human skin transplant rejection. Am J Transplant 17(4):931–943. https://doi.org/10.1111/ajt.14185

    Article  CAS  Google Scholar 

  31. Fraser H, Safinia N, Grageda N, Thirkell S, Lowe K, Fry LJ, Scotta C, Hope A, Fisher C, Hilton R, Game D, Harden P, Bushell A, Wood K, Lechler RI, Lombardi G (2018) A rapamycin-based GMP-compatible process for the isolation and expansion of regulatory T cells for clinical trials. Mol Ther Methods Clin Dev 8:198–209. https://doi.org/10.1016/j.omtm.2018.01.006

    Article  CAS  Google Scholar 

  32. Safinia N, Vaikunthanathan T, Fraser H, Thirkell S, Lowe K, Blackmore L, Whitehouse G, Martinez-Llordella M, Jassem W, Sanchez-Fueyo A, Lechler RI, Lombardi G (2016) Successful expansion of functional and stable regulatory T cells for immunotherapy in liver transplantation. Oncotarget 7(7):7563–7577. https://doi.org/10.18632/oncotarget.6927

    Article  Google Scholar 

  33. Kutner RH, Zhang XY, Reiser J (2009) Production, concentration and titration of pseudotyped HIV-1-based lentiviral vectors. Nat Protoc 4(4):495–505. https://doi.org/10.1038/nprot.2009.22

    Article  CAS  Google Scholar 

  34. Harlow E, Lane D (2006) Attaching suspension cells to slides using a cytocentrifuge. CSH Protoc 2006(3). https://doi.org/10.1101/pdb.prot4291

  35. Cronin J, Zhang XY, Reiser J (2005) Altering the tropism of lentiviral vectors through pseudotyping. Curr Gene Ther 5(4):387–398. https://doi.org/10.2174/1566523054546224

    Article  CAS  Google Scholar 

  36. Zacharias DA, Violin JD, Newton AC, Tsien RY (2002) Partitioning of lipid-modified monomeric GFPs into membrane microdomains of live cells. Science 296(5569):913–916. https://doi.org/10.1126/science.1068539

    Article  CAS  Google Scholar 

  37. Snapp EL, Hegde RS, Francolini M, Lombardo F, Colombo S, Pedrazzini E, Borgese N, Lippincott-Schwartz J (2003) Formation of stacked ER cisternae by low affinity protein interactions. J Cell Biol 163(2):257–269. https://doi.org/10.1083/jcb.200306020

    Article  CAS  Google Scholar 

  38. Merzlyak EM, Goedhart J, Shcherbo D, Bulina ME, Shcheglov AS, Fradkov AF, Gaintzeva A, Lukyanov KA, Lukyanov S, Gadella TW, Chudakov DM (2007) Bright monomeric red fluorescent protein with an extended fluorescence lifetime. Nat Methods 4(7):555–557. https://doi.org/10.1038/nmeth1062

    Article  CAS  Google Scholar 

  39. Jiang W, Hua R, Wei M, Li C, Qiu Z, Yang X, Zhang C (2015) An optimized method for high-titer lentivirus preparations without ultracentrifugation. Sci Rep 5:13875. https://doi.org/10.1038/srep13875

    Article  Google Scholar 

  40. Hasan AN, Selvakumar A, Shabrova E, Liu XR, Afridi F, Heller G, Riviere I, Sadelain M, Dupont B, O’Reilly RJ (2016) Soluble and membrane-bound interleukin (IL)-15 Ralpha/IL-15 complexes mediate proliferation of high-avidity central memory CD8(+) T cells for adoptive immunotherapy of cancer and infections. Clin Exp Immunol 186(2):249–265. https://doi.org/10.1111/cei.12816

    Article  CAS  Google Scholar 

  41. Zhou J, Jin L, Wang F, Zhang Y, Liu B, Zhao T (2019) Chimeric antigen receptor T (CAR-T) cells expanded with IL-7/IL-15 mediate superior antitumor effects. Protein Cell 10(10):764–769. https://doi.org/10.1007/s13238-019-0643-y

    Article  CAS  Google Scholar 

  42. Alizadeh D, Wong RA, Yang X, Wang D, Pecoraro JR, Kuo CF, Aguilar B, Qi Y, Ann DK, Starr R, Urak R, Wang X, Forman SJ, Brown CE (2019) IL15 enhances CAR-T cell antitumor activity by reducing mTORC1 activity and preserving their stem cell memory phenotype. Cancer Immunol Res 7(5):759–772. https://doi.org/10.1158/2326-6066.CIR-18-0466

    Article  CAS  Google Scholar 

  43. Wilkie S, Burbridge SE, Chiapero-Stanke L, Pereira AC, Cleary S, van der Stegen SJ, Spicer JF, Davies DM, Maher J (2010) Selective expansion of chimeric antigen receptor-targeted T-cells with potent effector function using interleukin-4. J Biol Chem 285(33):25538–25544. https://doi.org/10.1074/jbc.M110.127951

    Article  CAS  Google Scholar 

  44. Portulano C, Paroder-Belenitsky M, Carrasco N (2014) The Na+/I- symporter (NIS): mechanism and medical impact. Endocr Rev 35(1):106–149. https://doi.org/10.1210/er.2012-1036

    Article  CAS  Google Scholar 

  45. Khoshnevisan A, Chuamsaamarkkee K, Boudjemeline M, Jackson A, Smith GE, Gee AD, Fruhwirth GO, Blower PJ (2017) 18F-Fluorosulfate for PET imaging of the sodium-iodide symporter: synthesis and biologic evaluation in vitro and in vivo. J Nucl Med 58(1):156–161. https://doi.org/10.2967/jnumed.116.177519

    Article  CAS  Google Scholar 

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Acknowledgments

BG and AS were supported by PhD studentships from the MRC-funded Doctoral Training Programme in Health Sciences at King’s College London. AV is supported by The Center for Experimental Immuno-Oncology Fellowship Award (FP00001443), the Tow Foundation Fellowship Award (FP000004141) and the Fiona and Stanley Druckenmiller Center for Lung Cancer Research Fellowship Award (FP00005072) at Memorial Sloan Kettering Cancer Center. Further support was received by an NIH/NCI Cancer Center Support Grant to MSKCC (P30 CA008748). This work was further supported by a Cancer Research UK grant [C48390/A21153] to GOF. This work was further supported by the Cancer Research UK Centre of London, the National Institute for Health Research (NIHR) Biomedical Research Centre based at Guy’s and St Thomas’ NHS Foundation Trust and King’s College London, the MRC Centre for Transplantation at King’s College London [MR/J006742/1], and the Wellcome/EPSRC Centre for Medical Engineering at King’s College London [WT 203148/Z/16/Z]. The views expressed are those of the authors and not necessarily those of the NIHR, the National Health Service, or the Department of Health.

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Author notes

  1. Adeel Saleem and Alessia Volpe contributed equally with all other contributors.

Authors and Affiliations

  1. Imaging Therapies and Cancer Group, Comprehensive Cancer Centre, School of Cancer and Pharmaceutical Sciences, King’s College London, London, UK

    Ben Grimsdell, Adeel Saleem & Gilbert O. Fruhwirth

  2. Molecular Imaging Group, Department of Radiology, Memorial Sloan Kettering Cancer Center, New York, NY, USA

    Alessia Volpe

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  1. Ben Grimsdell
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  2. Adeel Saleem
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  3. Alessia Volpe
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  4. Gilbert O. Fruhwirth
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Corresponding author

Correspondence to Gilbert O. Fruhwirth .

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Editors and Affiliations

  1. School of Biomedical Engineering and Imaging Sciences, King’s College London, London, UK

    Timothy H. Witney

  2. Department of Chemistry and Biomolecular Sciences, University of Ottawa, Ottawa, ON, Canada

    Adam J. Shuhendler

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Grimsdell, B., Saleem, A., Volpe, A., Fruhwirth, G.O. (2024). Genetic Engineering of Therapeutic Cells with the Sodium Iodide Symporter (NIS) to Enable Noninvasive In Vivo Therapy Tracking. In: Witney, T.H., Shuhendler, A.J. (eds) Positron Emission Tomography. Methods in Molecular Biology, vol 2729. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-3499-8_18

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  • DOI: https://doi.org/10.1007/978-1-0716-3499-8_18

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-3498-1

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