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Inverse electron demand Diels–Alder click chemistry for pretargeted PET imaging and radioimmunotherapy

Nature Protocols volume 16, pages 3348–3381 (2021)Cite this article

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Abstract

Radiolabeled antibodies have shown promise as tools for both the nuclear imaging and endoradiotherapy of cancer, but the protracted circulation time of radioimmunoconjugates can lead to high radiation doses to healthy tissues. To circumvent this issue, we have developed an approach to positron emission tomography (PET) imaging and radioimmunotherapy (RIT) predicated on radiolabeling the antibody after it has reached its target within the body. This in vivo pretargeting strategy is based on the rapid and bio-orthogonal inverse electron demand Diels–Alder reaction between tetrazine (Tz) and trans-cyclooctene (TCO). Pretargeted PET imaging and RIT using TCO-modified antibodies in conjunction with Tz-bearing radioligands produce high activity concentrations in target tissues as well as reduced radiation doses to healthy organs compared to directly labeled radioimmunoconjugates. Herein, we describe how to prepare a TCO-modified antibody (humanized A33-TCO) as well as how to synthesize two Tz-bearing radioligands: one labeled with the positron-emitting radiometal copper-64 ([64Cu]Cu-SarAr-Tz) and one labeled with the β-emitting radiolanthanide lutetium-177 ([177Lu]Lu-DOTA-PEG7-Tz). We also provide a detailed description of pretargeted PET and pretargeted RIT experiments in a murine model of human colorectal carcinoma. Proper training in both radiation safety and the handling of laboratory mice is required for the successful execution of this protocol.

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Fig. 1: In vivo pretargeting based on the IEDDA reaction.
Fig. 2: Several ligation mechanisms have been leveraged to facilitate in vivo pretargeting.
Fig. 3
Fig. 4
Fig. 5
Fig. 6: In vivo pretargeted radioimmunotherapy data.
Fig. 7: Representative radio-iTLC chromatograms for radiosynthesis.
Fig. 8: Representative pretargeted PET images.

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Data availability

The data described in ‘Anticipated results’ were derived from refs. 21 and 26, which are available in the public domain from the National Library of Medicine Database at https://pubmed.ncbi.nlm.nih.gov. Source data are provided with this paper.

References

  1. Deri, M. A., Zeglis, B. M., Francesconi, L. C. & Lewis, J. S. PET imaging with 89Zr: from radiochemistry to the clinic. Nucl. Med. Biol. 40, 3–14 (2013).

    Article  CAS  PubMed  Google Scholar 

  2. Verel, I. et al. Long-lived positron emitters zirconium-89 and iodine-124 for scouting of therapeutic radioimmunoconjugates with PET. Cancer Biother. Radiopharm. 18, 655–661 (2003).

    Article  CAS  PubMed  Google Scholar 

  3. Stillebroer, A. B. et al. Phase 1 radioimmunotherapy study with lutetium 177-labeled anti-carbonic anhydrase IX monoclonal antibody girentuximab in patients with advanced renal cell carcinoma. Eur. Urol. 64, 478–485 (2013).

    Article  CAS  PubMed  Google Scholar 

  4. Kramer, K. et al. Phase I study of targeted radioimmunotherapy for leptomeningeal cancers using intra-ommaya 131-I-3F8. J. Clin. Oncol. 25, 5465–5470 (2007).

    Article  PubMed  Google Scholar 

  5. Zeglis, B. M., Houghton, J. L., Evans, M. J., Viola-Villegas, N. & Lewis, J. S. Underscoring the influence of inorganic chemistry on nuclear imaging with radiometals. Inorg. Chem. 53, 1880–1899 (2014).

    Article  CAS  PubMed  Google Scholar 

  6. Kramer, K. et al. A phase II study of radioimmunotherapy with intraventricular 131I-3F8 for medulloblastoma. Pediatr. Blood Cancer 65, 10.1002/pbc.26754 (2018).

    Article  CAS  Google Scholar 

  7. van Loon, J. et al. PET imaging of zirconium-89 labelled cetuximab: a phase I trial in patients with head and neck and lung cancer. Radiother. Oncol. 122, 267–273 (2017).

    Article  PubMed  CAS  Google Scholar 

  8. Pandit-Taskar, N. et al. A phase I/II study for analytic validation of 89Zr-J591 immunoPET as a molecular imaging agent for metastatic prostate cancer. Clin. Cancer Res. 21, 5277–5285 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Maloney, R., Buuh, Z. Y., Zhao, Y. & Wang, R. E. Site-specific antibody fragment conjugates for targeted imaging. Methods Enzymol. 638, 295–320 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Jain, M., Venkatraman, G. & Batra, S. K. Optimization of radioimmunotherapy of solid tumors: biological impediments and their modulation. Clin. Cancer Res. 13, 1374–1382 (2007).

    Article  CAS  PubMed  Google Scholar 

  11. Hnatowich, D. J., Virzi, F. & Rusckowski, M. Investigations of avidin and biotin for imaging applications. J. Nucl. Med. 28, 1294–1302 (1987).

    CAS  PubMed  Google Scholar 

  12. Leonidova, A. et al. In vivo demonstration of an active tumor pretargeting approach with peptide nucleic acid bioconjugates as complementary system. Chem. Sci. 6, 5601–5616 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Salaun, P. Y. et al. Phase II trial of anticarcinoembryonic antigen pretargeted radioimmunotherapy in progressive metastatic medullary thyroid carcinoma: biomarker response and survival improvement. J. Nucl. Med. 53, 1185–1192 (2012).

    Article  CAS  PubMed  Google Scholar 

  14. Rondon, A. & Degoul, F. Antibody pretargeting based on bioorthogonal click chemistry for cancer imaging and targeted radionuclide therapy. Bioconjug. Chem. 31, 159–173 (2020).

    Article  CAS  PubMed  Google Scholar 

  15. Reiner, T. & Zeglis, B. M. The inverse electron demand Diels–Alder click reaction in radiochemistry. J. Label. Comp. Radiopharm. 57, 285–290 (2014).

    Article  CAS  Google Scholar 

  16. Keinänen, O. et al. Dual radionuclide theranostic pretargeting. Mol. Pharm. 16, 4416–4421 (2019).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  17. Keinänen, O. et al. Harnessing 64Cu/67Cu for a theranostic approach to pretargeted radioimmunotherapy. Proc. Natl. Acad. Sci. USA 117, 28316–28327 (2020).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  18. Rossin, R., Lappchen, T., van den Bosch, S. M., Laforest, R. & Robillard, M. S. Diels–Alder reaction for tumor pretargeting: in vivo chemistry can boost tumor radiation dose compared with directly labeled antibody. J. Nucl. Med. 54, 1989–1995 (2013).

    Article  CAS  PubMed  Google Scholar 

  19. Rossin, R. et al. In vivo chemistry for pretargeted tumor imaging in live mice. Angew. Chem. Int. Ed. Engl. 49, 3375–3378 (2010).

    Article  CAS  PubMed  Google Scholar 

  20. Zeglis, B. M. et al. A pretargeted PET imaging strategy based on bioorthogonal Diels–Alder click chemistry. J. Nucl. Med. 54, 1389–1396 (2013).

    Article  CAS  PubMed  Google Scholar 

  21. Zeglis, B. M. et al. Optimization of a pretargeted strategy for the PET imaging of colorectal cancer via the modulation of radioligand pharmacokinetics. Mol. Pharm. 12, 3575–3587 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Meyer, J. P. et al. 18F-based pretargeted PET imaging based on bioorthogonal Diels–Alder click chemistry. Bioconjug. Chem. 27, 298–301 (2016).

    Article  CAS  PubMed  Google Scholar 

  23. Meyer, J. P. et al. Exploring structural parameters for pretargeting radioligand optimization. J. Med. Chem. 60, 8201–8217 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Houghton, J. L. et al. Establishment of the in vivo efficacy of pretargeted radioimmunotherapy utilizing inverse electron demand Diels–Alder click chemistry. Mol. Cancer Ther. 16, 124–133 (2017).

    Article  CAS  PubMed  Google Scholar 

  25. Adumeau, P. et al. A pretargeted approach for the multimodal PET/NIRF imaging of colorectal cancer. Theranostics 6, 2267–2277 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Membreno, R., Cook, B. E., Fung, K., Lewis, J. S. & Zeglis, B. M. Click-mediated pretargeted radioimmunotherapy of colorectal carcinoma. Mol. Pharm. 15, 1729–1734 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Poty, S. et al. Leveraging bioorthogonal click chemistry to improve 225Ac-radioimmunotherapy of pancreatic ductal adenocarcinoma. Clin. Cancer Res. 25, 868–880 (2019).

    Article  CAS  PubMed  Google Scholar 

  28. Membreno, R., Cook, B. E. & Zeglis, B. M. Pretargeted radioimmunotherapy based on the inverse electron demand Diels–Alder reaction. J. Vis. Exp. 2019, 10.3791/59041 (2019).

    Google Scholar 

  29. Cook, B. E., Membreno, R. & Zeglis, B. M. Dendrimer scaffold for the amplification of in vivo pretargeting ligations. Bioconjug. Chem. 29, 2734–2740 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Siegl, S. J., Galeta, J., Dzijak, R., Dracinsky, M. & Vrabel, M. Bioorthogonal fluorescence turn-on labeling based on bicyclononyne-tetrazine cycloaddition reactions that form pyridazine products. Chempluschem 84, 493–497 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Meyer, J. P. et al. Bioorthogonal masking of circulating antibody-TCO groups using tetrazine-functionalized dextran polymers. Bioconjug. Chem. 29, 538–545 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. van Duijnhoven, S. M. et al. Diabody pretargeting with click chemistry in vivo. J. Nucl. Med. 56, 1422–1428 (2015).

    Article  PubMed  CAS  Google Scholar 

  33. Yazdani, A. et al. A bone-seeking trans-cyclooctene for pretargeting and bioorthogonal chemistry: a proof of concept study using 99mTc- and 177Lu-labeled tetrazines. J. Med. Chem. 59, 9381–9389 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Algar, W. R. et al. The controlled display of biomolecules on nanoparticles: a challenge suited to bioorthogonal chemistry. Bioconjug. Chem. 22, 825–858 (2011).

    Article  CAS  PubMed  Google Scholar 

  35. Lesch, H. P., Kaikkonen, M. U., Pikkarainen, J. T. & Yla-Herttuala, S. Avidin-biotin technology in targeted therapy. Expert Opin. Drug Deliv. 7, 551–564 (2010).

    Article  CAS  PubMed  Google Scholar 

  36. Paganelli, G. et al. Antibody-guided three-step therapy for high grade glioma with yttrium-90 biotin. Eur. J. Nucl. Med. 26, 348–357 (1999).

    Article  CAS  PubMed  Google Scholar 

  37. Breitz, H. B. et al. Clinical optimization of pretargeted radioimmunotherapy with antibody-streptavidin conjugate and 90Y-DOTA-biotin. J. Nucl. Med. 41, 131–140 (2000).

    CAS  PubMed  Google Scholar 

  38. Schoffelen, R. et al. Pretargeted immuno-positron emission tomography imaging of carcinoembryonic antigen-expressing tumors with a bispecific antibody and a 68Ga- and 18F-labeled hapten peptide in mice with human tumor xenografts. Mol. Cancer Ther. 9, 1019–1027 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. Goldenberg, D. M., Chatal, J. F., Barbet, J., Boerman, O. & Sharkey, R. M. Cancer imaging and therapy with bispecific antibody pretargeting. Update Cancer Ther. 2, 19–31 (2007).

    Article  PubMed  PubMed Central  Google Scholar 

  40. Hall, H. et al. In vitro autoradiography of carcinoembryonic antigen in tissue from patients with colorectal cancer using multifunctional antibody TF2 and 67/68Ga-labeled haptens by pretargeting. Am. J. Nucl. Med. Mol. Imaging 2, 141–150 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  41. Bodet-Milin, C. et al. Immuno-PET using anticarcinoembryonic antigen bispecific antibody and 68Ga-labeled peptide in metastatic medullary thyroid carcinoma: clinical optimization of the pretargeting parameters in a first-in-human trial. J. Nucl. Med. 57, 1505–1511 (2016).

    Article  CAS  PubMed  Google Scholar 

  42. Bodet-Milin, C. et al. Pharmacokinetics and dosimetry studies for optimization of pretargeted radioimmunotherapy in CEA-expressing advanced lung cancer patients. Front. Med. 2, 84 (2015).

    Article  Google Scholar 

  43. Sharkey, R. M., Rossi, E. A., McBride, W. J., Chang, C. H. & Goldenberg, D. M. Recombinant bispecific monoclonal antibodies prepared by the dock-and-lock strategy for pretargeted radioimmunotherapy. Semin. Nucl. Med. 40, 190–203 (2010).

    Article  PubMed  PubMed Central  Google Scholar 

  44. Liu, G. et al. 90Y labeled phosphorodiamidate morpholino oligomer for pretargeting radiotherapy. Bioconjug. Chem. 22, 2539–2545 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Gupta, A., Mishra, A. & Puri, N. Peptide nucleic acids: advanced tools for biomedical applications. J. Biotechnol. 259, 148–159 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  46. Kim, K. L. et al. Supramolecular latching system based on ultrastable synthetic binding pairs as versatile tools for protein imaging. Nat. Commun. 9, 1712 (2018).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  47. Sundhoro, M., Jeon, S., Park, J., Ramstrom, O. & Yan, M. Perfluoroaryl azide staudinger reaction: a fast and bioorthogonal reaction. Angew. Chem. Int. Ed. Engl. 56, 12117–12121 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  48. Agard, N. J., Prescher, J. A. & Bertozzi, C. R. A strain-promoted [3 + 2] azide-alkyne cycloaddition for covalent modification of biomolecules in living systems. J. Am. Chem. Soc. 126, 15046–15047 (2004).

    Article  CAS  PubMed  Google Scholar 

  49. Carroll, L., Evans, H. L., Aboagye, E. O. & Spivey, A. C. Bioorthogonal chemistry for pre-targeted molecular imaging—progress and prospects. Org. Biomol. Chem. 11, 5772–5781 (2013).

    Article  CAS  PubMed  Google Scholar 

  50. Ackerman, M. E. et al. A33 antigen displays persistent surface expression. Cancer Immunol. Immunother. 57, 1017–1027 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  51. Keinänen, O. et al. Pretargeting of internalizing trastuzumab and cetuximab with a 18F-tetrazine tracer in xenograft models. EJNMMI Res. 7, 95 (2017).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  52. Houghton, J. L. et al. Pretargeted immuno-PET of pancreatic cancer: overcoming circulating antigen and internalized antibody to reduce radiation doses. J. Nucl. Med. 57, 453–459 (2016).

    Article  CAS  PubMed  Google Scholar 

  53. Rossin, R., van Duijnhoven, S. M., Lappchen, T., van den Bosch, S. M. & Robillard, M. S. Trans-cyclooctene tag with improved properties for tumor pretargeting with the Diels–Alder reaction. Mol. Pharm. 11, 3090–3096 (2014).

    Article  CAS  PubMed  Google Scholar 

  54. Royzen, M., Yap, G. P. & Fox, J. M. A photochemical synthesis of functionalized trans-cyclooctenes driven by metal complexation. J. Am. Chem. Soc. 130, 3760–3761 (2008).

    Article  CAS  PubMed  Google Scholar 

  55. Rondon, A. et al. Antibody PEGylation in bioorthogonal pretargeting with trans-cyclooctene/tetrazine cycloaddition: in vitro and in vivo evaluation in colorectal cancer models. Sci. Rep. 7, 14918 (2017).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  56. Maggi, A. et al. Development of a novel antibody-tetrazine conjugate for bioorthogonal pretargeting. Org. Biomol. Chem. 14, 7544–7551 (2016).

    Article  CAS  PubMed  Google Scholar 

  57. Billaud, E. M. F. et al. Micro-flow photosynthesis of new dienophiles for inverse-electron-demand Diels–Alder reactions. Potential applications for pretargeted in vivo PET imaging. Chem. Sci. 8, 1251–1258 (2017).

    Article  CAS  PubMed  Google Scholar 

  58. Billaud, E. M. F. et al. Pretargeted PET imaging using a bioorthogonal 18F-labeled trans-cyclooctene in an ovarian carcinoma model. Bioconjug. Chem. 28, 2915–2920 (2017).

    Article  CAS  PubMed  Google Scholar 

  59. Steen, E. J. L. et al. Improved radiosynthesis and preliminary in vivo evaluation of the 11C-labeled tetrazine [11C]AE-1 for pretargeted PET imaging. Bioorg. Med. Chem. Lett. 29, 986–990 (2019).

    Article  CAS  PubMed  Google Scholar 

  60. Edem, P. E. et al. Evaluation of a 68Ga-labeled DOTA-tetrazine as a PET alternative to 111In-SPECT pretargeted imaging. Molecules 25, 463 (2020).

    Article  CAS  PubMed Central  Google Scholar 

  61. Edem, P. E. et al. Evaluation of the inverse electron demand Diels–Alder reaction in rats using a scandium-44-labelled tetrazine for pretargeted PET imaging. EJNMMI Res. 9, 49 (2019).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  62. Keinänen, O. et al. A new highly reactive and low lipophilicity fluorine-18 labeled tetrazine derivative for pretargeted PET imaging. ACS Med. Chem. Lett. 7, 62–66 (2016).

    Article  PubMed  CAS  Google Scholar 

  63. Reiner, T., Lewis, J. S. & Zeglis, B. M. Harnessing the bioorthogonal inverse electron demand Diels–Alder cycloaddition for pretargeted PET imaging. J. Vis. Exp. 2015, e52335 (2015).

    Google Scholar 

  64. Altai, M. et al. Feasibility of affibody-based bioorthogonal chemistry-mediated radionuclide pretargeting. J. Nucl. Med. 57, 431–436 (2016).

    Article  CAS  PubMed  Google Scholar 

  65. Vito, A. et al. A 99mTc-labelled tetrazine for bioorthogonal chemistry. Synthesis and biodistribution studies with small molecule trans-cyclooctene derivatives. PloS One 11, e0167425 (2016).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  66. Zhou, Z., Devoogdt, N., Zalutsky, M. R. & Vaidyanathan, G. An efficient method for labeling single domain antibody fragments with 18F using tetrazine-trans-cyclooctene ligation and a renal brush border enzyme-cleavable linker. Bioconjug. Chem. 29, 4090–4103 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  67. Litau, S., Seibold, U., Wangler, B., Schirrmacher, R. & Wangler, C. iEDDA conjugation reaction in radiometal labeling of peptides with 68Ga and 64Cu: unexpected findings. ACS Omega 3, 14039–14053 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  68. Lindmo, T., Boven, E., Cuttitta, F., Fedorko, J. & Bunn, P. A. Jr. Determination of the immunoreactive fraction of radiolabeled monoclonal antibodies by linear extrapolation to binding at infinite antigen excess. J. Immunol. Methods 72, 77–89 (1984).

    Article  CAS  PubMed  Google Scholar 

  69. Sharma, S. K. et al. A rapid bead-based radioligand binding assay for the determination of target-binding fraction and quality control of radiopharmaceuticals. Nucl. Med. Biol. 71, 32–38 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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Acknowledgements

The authors thank the National Institutes of Health (B.M.Z.: R01CA240963, U01CA221046, R01CA204167 and R01244327), the Academy of Finland (OMK) and the Tow Foundation (GDLR) for their generous financial support.

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Author notes

  1. These authors contributed equally: Samantha M. Sarrett, Outi Keinänen.

Authors and Affiliations

  1. Department of Chemistry, Hunter College, City University of New York, New York, NY, USA

    Samantha M. Sarrett, Outi Keinänen, Eric J. Dayts, Guillaume Dewaele-Le Roi, Cindy Rodriguez & Brian M. Zeglis

  2. PhD Program in Biochemistry, Graduate Center of the City University of New York, New York, NY, USA

    Samantha M. Sarrett & Brian M. Zeglis

  3. Department of Radiology, Memorial Sloan Kettering Cancer Center, New York, NY, USA

    Outi Keinänen, Guillaume Dewaele-Le Roi & Brian M. Zeglis

  4. Department of Chemistry, Radiochemistry, University of Helsinki, Helsinki, Finland

    Outi Keinänen

  5. Ph.D. Program in Chemistry, Graduate Center of the City University of New York, New York, NY, USA

    Guillaume Dewaele-Le Roi, Cindy Rodriguez & Brian M. Zeglis

  6. Brain and Mind Research Institute & Appel Institute for Alzheimer’s Disease Research, Weill Cornell Medical College, New York, NY, USA

    Kathryn E. Carnazza

  7. Department of Radiology, Weill Cornell Medical College, New York, NY, USA

    Brian M. Zeglis

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Key references using this protocol

Membreno, R. et al. Mol. Pharm. 15, 1729–1734 (2018): https://pubs.acs.org/doi/10.1021/acs.molpharmaceut.8b00093

Zeglis, B. M. et al. Mol. Pharm. 12, 3575–3587 (2015): https://pubs.acs.org/doi/10.1021/acs.molpharmaceut.5b00294

Adumeau, P. et al. Theranostics 6, 2267–2277 (2016): https://www.thno.org/v06p2267.htm

Houghton, J. L. et al. Mol. Cancer Ther. 16, 124–133 (2017): https://mct.aacrjournals.org/content/16/1/124

Keinänen, O. et al. Mol. Pharm. 16, 4416–4421 (2019): https://pubs.acs.org/doi/10.1021/acs.molpharmaceut.9b00746

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Source Data Fig. 7

Organ activity concentration data for each mouse to go along with biodistribution data.

Source Data Fig. 8

Organ activity concentration data for each mouse to go along with biodistribution data.

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Sarrett, S.M., Keinänen, O., Dayts, E.J. et al. Inverse electron demand Diels–Alder click chemistry for pretargeted PET imaging and radioimmunotherapy. Nat Protoc 16, 3348–3381 (2021). https://doi.org/10.1038/s41596-021-00540-2

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