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MR Reporter Gene Imaging of Endostatin Expression and Therapy

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Abstract

Purpose

The aim of this study is to monitor endostatin gene expression and therapy using transferrin receptor (TfR) as reporter gene and transferrin conjugate of ultrasmall supramagnetic iron oxide nanoparticle (Tf–USPIO) as magnetic resonance (MR) reporter probe.

Procedure

A retroviral plasmid (pLP-LNCX) encoding mouse endostatin and TfR was constructed, and packaged with a titer of 4 × 107colony-forming units per millimeter. MDA-MB-231 breast tumors were established in BALB/c mice by subcutaneous injection of 2 × 106 MDA-MB-231 cells. Mice were intratumorally injected with recombinant retrovirus and imaged with MR using Tf–USPIO. Western blot, Prussian blue, and immunohistochemical staining were performed to validate the magnetic resonance imaging results. The antitumor effect of retro-endostatin (ES)-TfR was also evaluated by intratumoral injection of the viral vector.

Results

The expression of both endostatin and TfR genes in MDA-MB-231 cells after retroviral transfection was confirmed by Western blot and flow cytometry. Tf–USPIO conjugate binds specifically to cells stably transfected with retro-ES-TfR. After intravenous injection of the Tf–USPIO conjugate, there was a more pronounced decrease in T2 relaxation time in tumors treated with retro-ES-TfR than in tumors treated with empty retrovirus retro-LNCX. The expression of ES gene significantly delayed the growth of MDA-MB-231 tumor and reduction of microvessel density and VEGF level as compared to those without viral transfection or transfected with empty retro-LNCX vector.

Conclusions

Endostatin therapeutic gene expression was visualized successfully using TfR reporter gene and Tf–USPIO MR reporter probe, which indicates that MR reporter gene imaging may be valuable in gene therapy to evaluate therapeutic gene expression and treatment efficacy.

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References

  1. Carmeliet P, Jain RK (2000) Angiogenesis in cancer and other diseases. Nature 407:249-257

    Article  CAS  PubMed  Google Scholar 

  2. Shojaei F, Ferrara N (2008) Role of the microenvironment in tumor growth and in refractoriness/resistance to anti-angiogenic therapies. Drug Resist Updat 11:219-230

    Article  CAS  PubMed  Google Scholar 

  3. Tozer GM, Kanthou C, Lewis G, Prise VE, Vojnovic B, Hill SA (2008) Tumour vascular disrupting agents: combating treatment resistance. Br J Radiol 81(Spec No 1):S12-S20

    Article  CAS  PubMed  Google Scholar 

  4. Wickstrom SA, Alitalo K, Keski-Oja J (2005) Endostatin signaling and regulation of endothelial cell-matrix interactions. Adv Cancer Res 94:197-229

    Article  CAS  PubMed  Google Scholar 

  5. Li J, Dong X, Xu Z, Jiang X, Jiang H, Krissansen GW et al (2008) Endostatin gene therapy enhances the efficacy of paclitaxel to suppress breast cancers and metastases in mice. J Biomed Sci 15:99-109

    Article  PubMed  Google Scholar 

  6. Xu HM, Zhang GY, Ji XD, Cao L, Shu L, Hua ZC (2005) Expression of soluble, biologically active recombinant human endostatin in Escherichia coli. Protein Expr Purif 41:252-258

    Article  CAS  PubMed  Google Scholar 

  7. Dhanabal M, Ramchandran R, Volk R, Stillman IE, Lombardo M, Iruela-Arispe ML et al (1999) Endostatin: yeast production, mutants, and antitumor effect in renal cell carcinoma. Cancer Res 59:189-197

    CAS  PubMed  Google Scholar 

  8. Huang X, Wong MK, Zhao Q, Zhu Z, Wang KZ, Huang N et al (2001) Soluble recombinant endostatin purified from Escherichia coli: antiangiogenic activity and antitumor effect. Cancer Res 61:478-481

    CAS  PubMed  Google Scholar 

  9. Hansma AH, Broxterman HJ, van der Horst I, Yuana Y, Boven E, Giaccone G et al (2005) Recombinant human endostatin administered as a 28-day continuous intravenous infusion, followed by daily subcutaneous injections: a phase I and pharmacokinetic study in patients with advanced cancer. Ann Oncol 16:1695-1701

    Article  CAS  PubMed  Google Scholar 

  10. Ning T, Yan X, Lu ZJ, Wang GP, Zhang N, Yang J, et al. Gene therapy in orthotopic lung cancer murine model with angiogenesis inhibitor, endostatin. Hum Gene Ther. 2009. (in press)

  11. Coutinho EL, Andrade LN, Chammas R, Morganti L, Schor N, Bellini MH (2007) Anti-tumor effect of endostatin mediated by retroviral gene transfer in mice bearing renal cell carcinoma. FASEB J 3153–3161

  12. Liu F, Tan G, Li J, Dong X, Krissansen GW, Sun X (2007) Gene transfer of endostatin enhances the efficacy of doxorubicin to suppress human hepatocellular carcinomas in mice. Cancer Sci 98:1381-1387

    Article  CAS  PubMed  Google Scholar 

  13. Waerzeggers Y, Monfared P, Viel T, Winkeler A, Voges J, Jacobs AH (2009) Methods to monitor gene therapy with molecular imaging Methods. 2009. (in press)

  14. Wei X, Lin L, Hou Y, Fu X, Zhang J, Mao Z et al (2008) Construction of recombinant adenovirus co-expression vector carrying the human transforming growth factor-β1 and vascular endothelial growth factor genes and its effect on anterior cruciate ligament fibroblasts. Chin Med J 121:1426-1432

    CAS  PubMed  Google Scholar 

  15. Jin X, Bookstein R, Wills K, Avanzini J, Tsai V, LaFace D et al (2001) Evaluation of endostatin antiangiogenesis gene therapy in vitro and in vivo. Cancer Gene Ther 982–989

  16. Huszthy PC, Brekken C, Pedersen TB, Thorsen F, Sakariassen PO, Skaftnesmo KO et al (2006) Antitumor efficacy improved by local delivery of species-specific endostatin. J Neurosurg 104:118-128

    Article  CAS  PubMed  Google Scholar 

  17. Jiang Z, Teng GJ, Chen BA, Xu ZF, Hu J, Shao ZY (2008) Local effects of retrovirally transduced endostatin-expressing human umbilical cord blood CD34+ cells on transplanted malignancy in a mouse model of hepatic cancer. Cell Transplant 17:969-975

    Article  CAS  PubMed  Google Scholar 

  18. Pawliuk R, Bachelot T, Zurkiya O, Eriksson A, Cao Y, Leboulch P (2002) Continuous intravascular secretion of endostatin in mice from transduced hematopoietic stem cells. Mol Ther 5:345-351

    Article  CAS  PubMed  Google Scholar 

  19. Hu F, Wei L, Zhou Z, Ran Y, Li Z, Gao M (2006) Preparation of Biocompatible Magnetite Nanocrystals for In Vivo Magnetic Resonance Detection of Cancer. Adv Mater Deerfield 18:2553-2556

    Article  CAS  Google Scholar 

  20. Zhang C, Jugold M, Woenne EC, Lammers T, Morgenstern B, Mueller MM et al (2007) Specific targeting of tumor angiogenesis by RGD-conjugated ultrasmall superparamagnetic iron oxide particles using a clinical 1.5-T magnetic resonance scanner. Cancer Res 67:1555-1562

    Article  CAS  PubMed  Google Scholar 

  21. Xie J, Chen K, Lee HY, Xu C, Hsu AR, Peng S et al (2008) Ultrasmall c(RGDyK)-coated Fe3O4 nanoparticles and their specific targeting to integrin alpha(v)beta3-rich tumor cells. J Am Chem Soc 130:7542-7543

    Article  CAS  PubMed  Google Scholar 

  22. Mowat P, Franconi F, Chapon C, Lemaire L, Dorat J, Hindré F et al (2007) Evaluating SPIO-labelled cell MR efficiency by three-dimensional quantitative T2* MRI. NMR Biomed 20:21-27

    Article  CAS  PubMed  Google Scholar 

  23. Yamanaka R, Zullo SA, Ramsey J, Onodera M, Tanaka R, Blaese M et al (2001) Induction of therapeutic antitumor antiangiogenesis by intratumoral injection of genetically engineered endostatin producing Semliki Forest virus. Cancer Gene Ther 8:796-802

    Article  CAS  PubMed  Google Scholar 

  24. Wang X, Liu FK, Li X, Li JS, Xu GX (2002) Retrovirus-mediated gene transfer of human endostatin inhibits growth of human liver carcinoma cells SMMC7721 in nude mice. World J Gastroenterol 1045–1049.

  25. Hogemann D, Basilion JP (2002) “Seeing inside the body”: MR imaging of gene expression. Eur J Nucl Med Mol Imaging 29:400-408

    Article  PubMed  Google Scholar 

  26. Brack SS, Dinkelborg LM, Neri D (2004) Molecular targeting of angiogenesis for imaging and therapy. Eur J Nucl Med Mol Imaging 31:1327-1341

    Article  PubMed  Google Scholar 

  27. Schäfer R, Kehlbach R, Wiskirchen J, Bantleon R, Pintaske J, Brehm BR et al (2007) Transferrin receptor upregulation: in vitro labeling of rat mesenchymal stem cells with superparamagnetic iron oxide. Radiology 244:514-523

    Article  PubMed  Google Scholar 

  28. Ichikawa T, Högemann-Savellano D, Saeki Y, Tyminsk E, Terada K, Weissleder R et al. MRI of transgene expression: correlation to therapeutic gene expression Neoplasia. 2002;4:523–530.

  29. Sauter BV, Martinet O, Zhang WJ, Mandeli J, Woo SL (2000) Adenovirus-mediated gene transfer of endostatin in vivo results in high level of transgene expression and inhibition of tumor growth and metastases. Proc Natl Acad Sci USA 97:4802-4807

    Article  CAS  PubMed  Google Scholar 

  30. Moore A, Josephson L, Bhorade RM, Basilion JP, Weissleder R (2001) Human transferrin receptor gene as a marker gene for MR imaging. Radiology 221:244-250

    Article  CAS  PubMed  Google Scholar 

  31. Hogemann D, Josephson L, Weissleder R, Basilion JP (2000) Improvement of MRI Probes To Allow Efficient Detection of Gene Expression. Bioconjug Chem 11:941-946

    Article  CAS  PubMed  Google Scholar 

  32. Moore A, Basilion JP, Chiocca EA, Weissleder R (1998) Measuring transferrin receptor gene expression by NMR imaging. Biochim Biophys Acta 1402:239-249

    Article  CAS  PubMed  Google Scholar 

  33. Thorek DL, Tsourkas A (2008) Size, charge and concentration dependent uptake of iron oxide particles by non-phagocytic cells. Biomaterials 29:3583-3590

    Article  CAS  PubMed  Google Scholar 

  34. Nakase I, Gallis B, Takatani-Nakase T, Oh S, Lacoste E, Singh NP et al (2009) Transferrin receptor-dependent cytotoxicity of artemisinin-transferrin conjugates on prostate cancer cells and induction of apoptosis. Cancer Lett 274:290-298

    Article  CAS  PubMed  Google Scholar 

  35. Inoue T, Cavanaugh PG, Steck PA, Brunner N, Nicolson GL (1993) Differences in transferrin response and numbers of transferrin receptors in rat and human mammary carcinoma lines of different metastatic potentials. J Cell Physiol 156:212-217

    Article  CAS  PubMed  Google Scholar 

  36. Kealy B LA, McMahon JM, Ritter T, O'Doherty A, Hoare M, Greiser U, Vaughan EE, Maenz M, O'Shea C, Barry F, O'Brien T. Comparison of viral and nonviral vectors for gene transfer to human endothelial progenitor cells. Tissue Eng Part C Methods. 2009. (in press)

  37. Evans V, Foster H, Graham I, Foster K, Athanasopoulos T, Simons J et al (2008) Human apolipoprotein E expression from mouse skeletal muscle by electrotransfer of nonviral DNA (plasmid) and pseudotyped recombinant adeno-associated virus (AAV2/7). Hum Gene Ther 19:569-578

    Article  CAS  PubMed  Google Scholar 

  38. Morizono K, Xie Y, Ringpis G, Johnson M, Nassanian H, Lee B et al (2005) Lentiviral vector retargeting to P-glycoprotein on metastatic melanoma through intravenous injection. Nat Med 11:346-352

    Article  CAS  PubMed  Google Scholar 

  39. Blasberg R (2003) In vivo molecular-genetic imaging: multi-modality nuclear and optical combinations Nucl Med Biol 30:879-888

    Article  CAS  PubMed  Google Scholar 

  40. Bhaumik S, Gambhir S (2002) Optical imaging of Renilla luciferase reporter gene expression in living mice. Proc Natl Acad Sci USA 99:377-382

    Article  CAS  PubMed  Google Scholar 

  41. MacLaren D, Toyokuni T, Cherry S, Barrio J, Phelps M, Herschman H et al (2000) PET imaging of transgene expression. Biol Psychiatry 1:337-348

    Article  Google Scholar 

Download references

Acknowledgments

This work was supported, in part, by the Foundation of Heilongjiang Educational Committee (10551123), Heilongjiang Technical Plan Project (WC30307), the Key Program of Heilongjiang Scientific Problem Tackling Project (GB04C30201), and the Ministry of Personnel Foundation for Returnees (2007275).

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Authors and Affiliations

  1. Department of Medical Imaging and Nuclear Medicine, the 4th Affiliated Hospital, Harbin Medical University, Harbin, 150001, China

    Kai Wang, Kezheng Wang, Baozhong Shen, Tao Huang, Xilin Sun, Weihua Li, Gang Jin, Lin Li, Lihong Bu, Renfei Li & Dan Wang

  2. Department of Radiology, Bio-X & Biophysics, Stanford University School of Medicine, 1201 Welch Rd P095, Stanford, CA, 94305-5484, USA

    Xiaoyuan Chen

  3. Department of Medical Imaging and Nuclear Medicine, Harbin Medical University, Yiyuan Street 135, Nan gang District, Harbin, 150001, China

    Baozhong Shen

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  1. Kai Wang
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  2. Kezheng Wang
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  3. Baozhong Shen
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  4. Tao Huang
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  10. Renfei Li
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  11. Dan Wang
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  12. Xiaoyuan Chen
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Corresponding authors

Correspondence to Baozhong Shen or Xiaoyuan Chen.

Additional information

Kai Wang and Kezheng Wang contributed equally to the work.

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Wang, K., Wang, K., Shen, B. et al. MR Reporter Gene Imaging of Endostatin Expression and Therapy. Mol Imaging Biol 12, 520–529 (2010). https://doi.org/10.1007/s11307-009-0286-0

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  • DOI: https://doi.org/10.1007/s11307-009-0286-0

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