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Non-invasive monitoring of tumor-vessel infarction by retargeted truncated tissue factor tTF–NGR using multi-modal imaging

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Abstract

The fusion protein tTF–NGR consists of the extracellular domain of the thrombogenic human tissue factor (truncated tissue factor, tTF) and the peptide GNGRAHA (NGR), a ligand of the surface protein CD13 (aminopeptidase N), upregulated on endothelial cells of tumor vessels. tTF–NGR preferentially activates blood coagulation within tumor vasculature, resulting in tumor vessel infarction and subsequent tumor growth retardation/regression. The anti-vascular mechanism of the tTF–NGR therapy approach was verified by quantifying the reduced tumor blood-perfusion with contrast-enhanced ultrasound, the reduced relative tumor blood volume by ultrasmall superparamagnetic iron oxide-enhanced magnetic resonance imaging, and by in vivo-evaluation of hemorrhagic bleeding with fluorescent biomarkers (AngioSense680) in fluorescence reflectance imaging. The accumulation of tTF–NGR within the tumor was proven by visualizing the distribution of the iodine-123-labelled protein by single-photon emission computed tomography. Use of these multi-modal vascular and molecular imaging tools helped to assess the therapeutic effect even at real time and to detect non-responding tumors directly after the first tTF–NGR treatment. This emphasizes the importance of imaging within clinical studies with tTF–NGR. The imaging techniques as used here have applicability within a wider scope of therapeutic regimes interfering with tumor vasculature. Some even are useful to obtain predictive biosignals in personalized cancer treatment.

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References

  1. Ferrara N, Kerbel RS (2005) Angiogenesis as a therapeutic target. Nature 438(7070):967–974

    Article  CAS  PubMed  Google Scholar 

  2. Kessler T, Bayer M, Schwöppe C, Liersch R, Mesters RM, Berdel WE (2010) Compounds in clinical Phase III and beyond. Recent Results Cancer Res 180:137–163

    Article  CAS  PubMed  Google Scholar 

  3. Carmeliet P, Jain RK (2011) Molecular mechanisms and clinical applications of angiogenesis. Nature 473(7347):298–307

    Article  CAS  PubMed  Google Scholar 

  4. Blankenberg FG, Levashova Z, Goris MG, Hamby CV, Backer MV, Backer JM (2011) Targeted systemic radiotherapy with scVEGF/177Lu leads to sustained disruption of the tumor vasculature and intratumoral apoptosis. J Nucl Med 52(10):1630–1637

    Article  CAS  PubMed  Google Scholar 

  5. Mohamedali KA, Niu G, Luster TA, Thorpe PE, Gao H, Chen X, Rosenblum MG (2012) Pharmacodynamics, tissue distribution, toxicity studies and antitumor efficacy of the vascular targeting fusion toxin VEGF121/rGel. Biochem Pharmacol 84(11):1534–1540

    Article  CAS  PubMed  Google Scholar 

  6. Brooks PC, Clark RA, Cheresh DA (1994) Requirement of vascular integrin alpha V beta 3 for angiogenesis. Science 264:569–571

    Article  CAS  PubMed  Google Scholar 

  7. Burg MA, Pasqualini R, Arap W, Ruoslahti E, Stallcup WB (1999) NG2 proteoglycan-binding peptides target tumor neovasculature. Cancer Res 59:2869–2874

    CAS  PubMed  Google Scholar 

  8. Carnemolla B, Balza E, Siri A, Zardi L, Nicotra MR, Bigotti A, Natali PG (1989) A tumor-associated fibronectin isoform generated by alternative splicing of messenger RNA precursors. J Cell Biol 108(3):1139–1148

    Article  CAS  PubMed  Google Scholar 

  9. Curnis F, Arrigoni G, Sacchi A, Fischetti L, Arap W, Pasqualini R, Corti A (2002) Differential binding of drugs containing the NGR motif to CD13 isoforms in tumor vessels, epithelia, and myeloid cells. Cancer Res 62:867–874

    CAS  PubMed  Google Scholar 

  10. Dvorak HF, Brown LF, Detmar M, Dvorak AM (1995) Vascular permeability factor/vascular endothelial growth factor, microvascular hyperpermeability, and angiogenesis. Am J Pathol 146:1029–1039

    CAS  PubMed  Google Scholar 

  11. Felding-Habermann B, Ruggeri ZM, Cheresh DA (1992) Distinct biological consequences of integrin alpha v beta 3-mediated melanoma cell adhesion to fibrinogen and its plasmic fragments. J Biol Chem 267(8):5070–5077

    CAS  PubMed  Google Scholar 

  12. Kessler TA, Pfeifer A, Silletti S, Mesters RM, Berdel WE, Verma I, Cheresh D (2002) Matrix metalloproteinase/integrin interactions as target for anti-angiogenic treatment strategies. Ann Hematol 8(Suppl. 2):S69–S70

    Google Scholar 

  13. Kessler T, Fehrmann F, Bieker R, Berdel WE, Mesters RM (2007) Vascular endothelial growth factor and its receptor as drug targets in hematological malignancies. Curr Drug Targets 8:257–268

    Article  CAS  PubMed  Google Scholar 

  14. Pasqualini R, Koivunen E, Kain R, Lahdenranta J, Sakamoto M, Stryhn A, Ashmun RA, Shapiro LH, Arap W, Ruoshlahti E (2000) Aminopeptidase N is a receptor for tumor-homing peptides and a target for inhibiting angiogenesis. Cancer Res 60:722–727

    CAS  PubMed  Google Scholar 

  15. Pfeifer A, Kessler T, Silletti S, Cheresh DA, Verma IM (2000) Suppression of angiogenesis by lentiviral delivery of PEX, a noncatalytic fragment of matrix metalloproteinase 2. Proc Natl Acad Sci 97(22):12227–12232

    Article  CAS  PubMed  Google Scholar 

  16. Rettig WJ, Garin-Chesa P, Healey JH, Su SL, Jaffe EA, Old LJ (1992) Identification of endosialin, a cell surface glycoprotein of vascular endothelial cells in human cancer. Proc Natl Acad Sci 89:10832–10836

    Article  CAS  PubMed  Google Scholar 

  17. Arap W, Pasqualini R, Ruoslahti E (1998) Cancer treatment by targeted drug delivery to tumor vasculature in a mouse model. Science 279:377–380

    Article  CAS  PubMed  Google Scholar 

  18. Ellerby HM, Arap W, Ellerby LM, Kain R, Andrusiak R, Rio GD, Krajewski S, Lombardo CR, Rao R, Ruoslahti E, Bredesen DE, Pasqualini R (1999) Anti-cancer activity of targeted pro-apoptotic peptides. Nat Med 5:1032–1038

    Article  CAS  PubMed  Google Scholar 

  19. Hood JD, Bednarski M, Frausto R, Guccione S, Reisfeld RA, Xiang R, Cheresh DA (2002) Tumor regression by targeted gene delivery to the neovasculature. Science 296:2404–2407

    Article  CAS  PubMed  Google Scholar 

  20. Ruoslahti E (2000) Targeting tumor vasculature with homing peptides from phage display. Semin Cancer Biol 10:435–442

    Article  CAS  PubMed  Google Scholar 

  21. Curnis F, Sacchi A, Borgna L, Magni F, Gasparri A, Corti A (2000) Nat Biotechnol 18:1185–1190

    Article  CAS  PubMed  Google Scholar 

  22. Curnis F, Arrigoni G, Sacchi A, Fischetti L, Arap W, Pasqualini R, Corti A (2002) Differential binding of drugs containing the NGR motif to CD13 isoforms in tumor vessels, epithelia, and myeloid cells. Cancer Res 62:867–874

    CAS  PubMed  Google Scholar 

  23. Pastorino F, Brignole C, Marimpietri D, Cilli M, Gambini C, Ribatti D, Longhi R, Allen TM, Corti A, Ponzoni M (2003) Cancer Res 63(21):7400–7409

    CAS  PubMed  Google Scholar 

  24. Sacchi A, Gasparri A, Curnis F, Bellone M, Corti A (2004) Crucial role for interferon gamma in the synergism between tumor vasculature-targeted tumor necrosis factor alpha (NGR-TNF) and doxorubicin. Cancer Res 64(19):7150–7155

    Article  CAS  PubMed  Google Scholar 

  25. Sacchi A, Gasparri A, Gallo-Stampino C, Toma S, Curnis F, Corti A (2006) Synergistic antitumor activity of cisplatin, paclitaxel, and gemcitabine with tumor vasculature-targeted tumor necrosis factor-alpha. Clin Cancer Res 12(1):175–182

    Article  CAS  PubMed  Google Scholar 

  26. van Laarhoven HW, Gambarota G, Heerschap A, Lok J, Verhagen I, Corti A, Toma S, Gallo Stampino C, van der Kogel A, Punt CJ (2006) Effects of the tumor vasculature targeting agent NGR-TNF on the tumor microenvironment in murine lymphomas. Invest New Drugs 24(1):27–36

    Article  CAS  PubMed  Google Scholar 

  27. Di Matteo P, Curnis F, Longhi R, Colombo G, Sacchi A, Crippa L, Protti MP, Ponzoni M, Toma S, Corti A (2006) Immunogenic and structural properties of the Asn-Gly-Arg (NGR) tumor neovasculature-homing motif. Mol Immunol 43(10):1509–1518

    Article  PubMed  Google Scholar 

  28. Morrissey JH, Macik BG, Neuenschwander PF, Comp PC (1993) Quantitation of activated factor VII levels in plasma using a tissue factor mutant selectively deficient in promoting factor VII activation. Blood 81:734–744

    CAS  PubMed  Google Scholar 

  29. Kessler T, Bieker R, Padró T, Schwöppe C, Persigehl T, Bremer C, Kreuter M, Berdel WE, Mesters RM (2005) Inhibition of tumor growth by RGD peptide-directed delivery of truncated tissue factor to the tumor vasculature. Clin Cancer Res 11:6317–6324

    Article  CAS  PubMed  Google Scholar 

  30. Kessler T, Schwöppe C, Liersch R, Schliemann C, Hintelmann H, Bieker R, Berdel WE, Mesters RM (2008) Generation of fusion proteins for selective occlusion of tumor vessels. Curr Drug Discov Technol 5:1–8

    Article  CAS  PubMed  Google Scholar 

  31. Bieker R, Kessler T, Schwöppe C, Padró T, Persigehl T, Bremer C, Dreischalück J, Kolkmeyer A, Heindel W, Mesters RM, Berdel WE (2009) Infarction of tumor vessels by NGR-peptide directed targeting of tissue factor. Experimental results and first-in-man experience. Blood 113:5019–5027

    Article  CAS  PubMed  Google Scholar 

  32. Schwöppe C, Kessler T, Persigehl T, Liersch R, Hintelmann H, Dreischalück J, Ring J, Bremer C, Heindel W, Mesters RM, Berdel WE (2010) Tissue-factor fusion proteins induce occlusion of tumor vessels. Thromb Res 125(Suppl. 2):S143–S150

    Article  PubMed  Google Scholar 

  33. Nilsson F, Kosmehl H, Zardi L, Neri D (2001) Targeted delivery of tissue factor to the ED-B domain of fibronectin, a marker of angiogenesis, mediates the infarction of solid tumors in mice. Cancer Res 61:711–716

    CAS  PubMed  Google Scholar 

  34. Liu C, Huang H, Donate F, Dickinson C, Santucci R, El-Sheikh A, Vessella R, Edgington TS (2002) Prostate-specific membrane antigen directed selective thrombotic infarction of tumors. Cancer Res 62:5470–5475

    CAS  PubMed  Google Scholar 

  35. Ran S, Gao B, Duffy S, Watkins L, Rote N, Thorpe PE (1998) Infarction of solid Hodgkin’s tumors in mice by antibody-directed targeting of tissue factor to tumor vasculature. Cancer Res 58:4646–4653

    CAS  PubMed  Google Scholar 

  36. Huang X, Molema G, King S, Watkins L, Edgington TS, Thorpe PE (1997) Tumor infarction in mice by antibody-directed targeting of tissue factor to tumor vasculature. Science 275:547–550

    Article  CAS  PubMed  Google Scholar 

  37. Persigehl T, Wall A, Kellert J, Ring J, Remmele S, Heindel W, Dahnke H, Bremer C (2010) Tumor blood volume determination by using susceptibility-corrected ∆R2* multiecho MR. Radiology 255(3):781–789

    Article  PubMed  Google Scholar 

  38. Dreischalück J, Schwöppe C, Spieker T, Kessler T, Tiemann K, Liersch R, Schliemann C, Kreuter M, Kolkmeyer A, Hintelmann H, Mesters RM, Berdel WE (2010) Vascular infarction by subcutaneous application of tissue factor targeted to tumor vessels with NGR-peptides: activity and toxicity profile. Int J Oncol 37:1389–1397

    PubMed  Google Scholar 

  39. Von Maltzahn G, Park J-H, Lin KY, Singh N, Schwöppe C, Mesters R, Berdel WE, Ruoslahti E, Sailor MJ, Bhatia SN (2011) Nanoparticles that communicate in vivo to amplify tumour targeting. Nat Mater 10:545–552

    Article  Google Scholar 

  40. Schwöppe C, Zerbst C, Fröhlich M, Schliemann C, Kessler T, Liersch R, Overkamp L, Holtmeier R, Stypmann J, Dreiling A, König S, Höltke C, Lücke M, Müller-Tidow C, Mesters RM, Berdel WE (2013) Anticancer therapy by tumor vessel infarction with polyethylene glycol conjugated retargeted tissue factor. J Med Chem 56(6):2337–2347

    Article  PubMed  Google Scholar 

  41. Bailey GS (1994) Labeling of peptides and proteins by radioiodination. Methods Mol Biol 32:441–448

    CAS  PubMed  Google Scholar 

  42. Dennie J, Mandeville JB, Boxerman JL, Packard SD, Rosen BR, Weisskoff RM (1998) NMR imaging of changes in vascular morphology due to tumor angiogenesis. Magn Reson Med 40(6):793–799

    Article  CAS  PubMed  Google Scholar 

  43. Allkemper T, Bremer C, Matuszewski L, Ebert W, Reimer P (2002) Contrast-enhanced blood-pool MR angiography with optimized iron oxides: effect of size and dose on vascular contrast enhancement in rabbits. Radiology 223(2):432–438

    Article  PubMed  Google Scholar 

  44. Zhu H, Melder RJ, Baxter LT, Jain RK (1996) Physiologically based kinetic model of effector cell biodistribution in mammals: implications for adoptive immunotherapy. Cancer Res 56(16):3771–3781

    CAS  PubMed  Google Scholar 

  45. Lohmaier S, Ghanem A, Veltmann C, Sommer T, Bruce M, Tiemann K (2004) In vitro and in vivo studies on continuous echo-contrast application strategies using SonoVue in a newly developed rotating pump setup. Ultrasound Med Biol 30:1145–1151

    Article  PubMed  Google Scholar 

  46. Persigehl T, Bieker R, Matuszewski L, Wall A, Kessler T, Kooijmann H, Meier N, Ebert W, Berdel WE, Heindel W, Mesters RM, Bremer C (2007) Antiangiogenic tumor treatment: early non-invasive monitoring with USPIO-enhanced MR Imaging in mice. Radiology 244(2):449–456

    Article  PubMed  Google Scholar 

  47. Von Wallbrunn A, Waldeck J, Höltke C, Zühlsdorf M, Mesters RM, Heindel W, Schäfers M, Bremer C (2008) In vivo optical imaging of CD13/APN-expression in tumor xenografts. J Biomed Opt 13(1):011007

    Article  Google Scholar 

  48. Salmon BA, Salmon HW, Siemann DW (2007) Monitoring the treatment efficacy of the vascular disrupting agent CA4P. Eur J Cancer 43(10):1622–1629

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  49. Nielsen T, Bentzen L, Pedersen M et al (2012) Combretastatin A-4 phosphate affects tumor vessel volume and size distribution as assessed using MRI-based vessel size imaging. Clin Cancer Res 18(23):6469–6477

    Article  CAS  PubMed  Google Scholar 

  50. Kim KW, Lee JM, Jeon YS, et al. (2013) Vascular disrupting effect of CKD-516: preclinical study using DCE-MRI. Invest New Drugs. doi:10.1007/s10637-012-9915-6

  51. Shenoi MM, Iltis I, Choi J, et al. (2013) Nanoparticle Delivered Vascular Disrupting Agents (VDAs): Use of TNF-alpha conjugated Gold Nanoparticles for Multimodal Cancer Therapy. Mol Pharm. doi:10.1021/mp300505w

  52. Wang H, Sun X, Chen F et al (2009) Treatment of rodent liver tumor with combretastatin a4 phosphate: noninvasive therapeutic evaluation using multiparametric magnetic resonance imaging in correlation with microangiography and histology. Invest Radiol 44(1):44–53

    Article  CAS  PubMed  Google Scholar 

  53. Bohndiek SE, Kettunen MI, Hu DE et al (2010) Detection of tumor response to a vascular disrupting agent by hyperpolarized 13C magnetic resonance spectroscopy. Mol Cancer Ther 9(12):3278–3288

    Article  CAS  PubMed Central  PubMed  Google Scholar 

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Acknowledgments

We would like to thank Ina Winkler, Kathrin Höke, Klaudia Niepagenkemper, Justina Mbah, Rebecca Roß, Heike Hintelmann and Dirk Reinhardt for technical assistance. J.R. and C.Z. contributed experiments in partial fulfillment of the requirements to obtain a PhD title. This work was supported by grants of the Deutsche Krebshilfe e.V. (109245 to W.E. Berdel), the Deutsche Forschungsgemeinschaft [SFB656, projects C08, C03, C06, and Z05, EXC 1,003 Cells in Motion-Cluster of Excellence), the Sybille-Hahne-Stiftung, and the Interdisziplinäres Zentrum für Klinische Forschung (IZKF, Core Unit PIX (SmAP, SAMRI, ECHO, OPTI)].

Conflict of interest

R.M.M. and W.E.B. share a patent on vascular targeting with TF constructs. The other authors declare that they have no conflict of interest.

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  1. Thorsten Persigehl

    Present address: Department of Clinical Radiology, University Hospital Cologne, Cologne, Germany

Authors and Affiliations

  1. Department of Clinical Radiology, University of Muenster, Albert-Schweitzer-Campus 1, 48149, Münster, Germany

    Thorsten Persigehl, Janine Ring, Christoph Bremer & Walter Heindel

  2. Interdisciplinary Center of Clinical Research (IZKF Muenster), University of Muenster, Albert-Schweitzer-Campus 1, 48149, Münster, Germany

    Richard Holtmeier & Jörg Stypmann

  3. Division of Cardiology, Department of Cardiovascular Medicine, University of Muenster, Albert-Schweitzer-Campus 1, 48149, Münster, Germany

    Jörg Stypmann

  4. Department of Nuclear Medicine, University of Muenster, Albert-Schweitzer-Campus 1, 48149, Münster, Germany

    Michael Claesener & Michael Schäfers

  5. European Institute for Molecular Imaging (EIMI), Mendelstr. 11, University of Muenster, Albert-Schweitzer-Campus 1, 48149, Münster, Germany

    Sven Hermann & Michael Schäfers

  6. Department of Medicine A (Hematology, Hemostaseology, Oncology and Pneumology), University of Muenster, Albert-Schweitzer-Campus 1, 48149, Münster, Germany

    Caroline Zerbst, Christoph Schliemann, Rolf M. Mesters, Wolfgang E. Berdel & Christian Schwöppe

  7. Cluster of Excellence EXC 1003-Cells in Motion, University of Muenster, Albert-Schweitzer-Campus 1, 48149, Münster, Germany

    Wolfgang E. Berdel

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  1. Thorsten Persigehl
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Correspondence to Christian Schwöppe.

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Persigehl, T., Ring, J., Bremer, C. et al. Non-invasive monitoring of tumor-vessel infarction by retargeted truncated tissue factor tTF–NGR using multi-modal imaging. Angiogenesis 17, 235–246 (2014). https://doi.org/10.1007/s10456-013-9391-4

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