Skip to main content

Advertisement

SpringerLink
Log in

Hepatic Intra-arterial Delivery of a “Trojan-horses” Gene Therapy: A Pilot Study on Rabbit VX2 Hepatic Tumor Model

  • Laboratory Investigation
  • Published:
CardioVascular and Interventional Radiology Aims and scope Submit manuscript

Abstract

Purpose

Gene-directed enzyme prodrug therapy (GDEPT) is a “Trojan-horses” suicide gene therapy that consists of tumor-targeted gene delivery (vectorized by mesenchymal stem cells MSCs) encoding an enzyme that converts a harmless prodrug into cytotoxic metabolites in situ. Then, cytotoxic metabolites passively diffuse in the neighboring tumor cells and kill them (bystander effect). The goal of our study was to assess the feasibility and efficacy of intra-arterial administration of MSCs transduced with an optimized gene (MSC-CYP2B6TM-RED) followed by intravenous administration of cyclophosphamide (CPA) into the VX2 rabbit liver tumor.

Materials and Methods

Nine rabbits with a VX2 liver tumor were randomly assigned into three groups: Control group A (one rabbit) free of any treatment; Control group B (two rabbits) receiving intravenous injection of cyclophosphamide at day 3 and CPA at day 14; and Group C (six rabbits) receiving the GDEPT treatment, consisting of successive intra-arterial injection of transduced-MSCs at days 0 (n = 6) and 11 (n = 3), followed by injection of CPA at days 3 (n = 6) and 14 (n = 3). The tumor response was assessed by ultrasound scan every 7 days and histopathological analysis at sacrifice (D25).

Results

There was a significant difference in the tumor volume between control groups (A + B) and group C at D7: 38/19 cm3 (p = 0.024); D11: 51/20 cm3 (p = 0.024), and D25: 121/37 cm3 (p = 0.048). Tumor necrosis was significantly greater and metastatic spread was lower for rabbits who received GDEPT (78% of total tumor surface) than for control animals (A + B) (22% of total tumor surface (p = 0.006).

Conclusion

Intra-arterial delivery of transduced-MSCs is feasible and, after CPA injection, resulted in 78% tumor necrosis (p = 0.006) and less metastasis in a VX2 liver tumor model.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6

Similar content being viewed by others

References

  1. Amara I, Pramil E, Senamaud-Beaufort C, et al. Engineered mesenchymal stem cells as vectors in a suicide gene therapy against preclinical murine models for solid tumors. J Controll Release. 2016;239:82–91.

    Article  Google Scholar 

  2. Amara I, Touati W, Beaune P, de Waziers I. Mesenchymal stem cells as cellular vehicles for prodrug gene therapy against tumors. Biochimie. 2014;105:4–11.

    Article  CAS  PubMed  Google Scholar 

  3. Atencio IA, Grace M, Bordens R, et al. Biological activities of a recombinant adenovirus p53 (SCH 58500) administered by hepatic arterial infusion in a Phase 1 colorectal cancer trial. Cancer Gene Ther. 2006;13(2):169–81.

    Article  CAS  PubMed  Google Scholar 

  4. Ayash LJ, Wright JE, Tretyakov O, et al. Cyclophosphamide pharmacokinetics: correlation with cardiac toxicity and tumor response. J Clin Oncol. 1992;10(6):995–1000.

    Article  CAS  PubMed  Google Scholar 

  5. Bruix J, Reig M, Sherman M. Evidence-based diagnosis, staging, and treatment of patients with hepatocellular carcinoma. Gastroenterology. 2016;150(4):835–53.

    Article  PubMed  Google Scholar 

  6. Cantore M, Fiorentini G, Zamagni D, Muttini MP, Mambrini A, Rabbi C. Molecular therapy: clinical applications. Intra-arterial adenoviruses administration. J Exp Clin Cancer Res. 2003;22(4 Suppl):47–9.

    CAS  PubMed  Google Scholar 

  7. Chen CS, Lin JT, Goss KA, He YA, Halpert JR, Waxman DJ. Activation of the anticancer prodrugs cyclophosphamide and ifosfamide: identification of cytochrome P450 2B enzymes and site-specific mutants with improved enzyme kinetics. Mol Pharmacol. 2004;65(5):1278–85.

    Article  CAS  PubMed  Google Scholar 

  8. Culver KW, Ram Z, Wallbridge S, Ishii H, Oldfield EH, Blaese RM. In vivo gene transfer with retroviral vector-producer cells for treatment of experimental brain tumors. Science. 1992;256(5063):1550–2.

    Article  CAS  PubMed  Google Scholar 

  9. de Baere T, Arai Y, Lencioni R, et al. Treatment of liver tumors with lipiodol TACE: technical recommendations from experts opinion. Cardiovasc Intervent Radiol. 2016;39(3):334–43.

    Article  PubMed  Google Scholar 

  10. De Waziers I, Touati W, Diry M, Flinois JP, Dansette P, Beaune P Mutant cytochrome p450 2b6 proteins and uses thereof. Google Patents. 2014.

  11. Fujii T, Le Du F, Xiao L, et al. Effectiveness of an adjuvant chemotherapy regimen for early-stage breast cancer: a systematic review and network meta-analysis. JAMA Oncol. 2015;1(9):1311–8.

    Article  PubMed  PubMed Central  Google Scholar 

  12. Greco O, Dachs GU. Gene directed enzyme/prodrug therapy of cancer: historical appraisal and future prospectives. J Cell Physiol. 2001;187(1):22–36.

    Article  CAS  PubMed  Google Scholar 

  13. Kim YI, Chung JW, Park JH, Han JK, Hong JW, Chung H. Intraarterial gene delivery in rabbit hepatic tumors: transfection with nonviral vector by using iodized oil emulsion. Radiology. 2006;240(3):771–7.

    Article  PubMed  Google Scholar 

  14. Lee KH, Liapi E, Buijs M, et al. Considerations for implantation site of VX2 carcinoma into rabbit liver. J Vasc Interv Radiol JVIR. 2009;20(1):113–7.

    Article  PubMed  Google Scholar 

  15. Nguyen TA, Tychopoulos M, Bichat F, et al. Improvement of cyclophosphamide activation by CYP2B6 mutants: from in silico to ex vivo. Mol Pharmacol. 2008;73(4):1122–33.

    Article  CAS  PubMed  Google Scholar 

  16. Pan JG, Zhou X, Luo R, Han RF. The adeno-associated virus-mediated HSV-TK/GCV suicide system: a potential strategy for the treatment of bladder carcinoma. Med Oncol. 2012;29(3):1938–47.

    Article  CAS  PubMed  Google Scholar 

  17. Pascale F, Ghegediban SH, Bonneau M, et al. Modified model of VX2 tumor overexpressing vascular endothelial growth factor. J Vasc Interv Radiol JVIR. 2012;23(6):809–17.

    Article  PubMed  Google Scholar 

  18. Reid T, Galanis E, Abbruzzese J, et al. Hepatic arterial infusion of a replication-selective oncolytic adenovirus (dl1520): phase II viral, immunologic, and clinical endpoints. Can Res. 2002;62(21):6070–9.

    CAS  Google Scholar 

  19. Reid T, Warren R, Kirn D. Intravascular adenoviral agents in cancer patients: lessons from clinical trials. Cancer Gene Ther. 2002;9(12):979–86.

    Article  CAS  PubMed  Google Scholar 

  20. Romieu-Mourez R, Francois M, Boivin MN, Stagg J, Galipeau J. Regulation of MHC class II expression and antigen processing in murine and human mesenchymal stromal cells by IFN-gamma, TGF-beta, and cell density. J Immunol. 2007;179(3):1549–58.

    Article  CAS  PubMed  Google Scholar 

  21. Suckow MA, Stevens KA, Wilson RP. The laboratory rabbit, guinea pig, hamster, and other rodents. Cambridge: Academic Press; 2012.

    Google Scholar 

  22. Sze DY, Freeman SM, Slonim SM, et al. Dr. Gary J. Becker Young Investigator Award: intraarterial adenovirus for metastatic gastrointestinal cancer: activity, radiographic response, and survival. J Vasc Interv Radiol JVIR. 2003;14(3):279–90.

    Article  PubMed  Google Scholar 

  23. Tan SL, Ahmad TS, Selvaratnam L, Kamarul T. Isolation, characterization and the multi-lineage differentiation potential of rabbit bone marrow-derived mesenchymal stem cells. J Anat. 2013;222(4):437–50.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Touati W, Tran T, Seguin J, et al. A suicide gene therapy combining the improvement of cyclophosphamide tumor cytotoxicity and the development of an anti-tumor immune response. Curr Gene Ther. 2014;14(3):236–46.

    Article  CAS  PubMed  Google Scholar 

  25. Wu H, Exner AA, Shi H, Bear J, Haaga JR. Dynamic evolutionary changes in blood flow measured by MDCT in a hepatic VX2 tumor implant over an extended 28-day growth period: time-density curve analysis. Acad Radiol. 2009;16(12):1483–92.

    Article  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

This work has been selected by the CIRSE 2017 Scientific Programme Committee and the CVIR Editorial Board, and cordially invited to send a complete manuscript for publication in the official journal of CIRSE, the CardioVascular and Interventional Radiology (CVIR). This work was supported by SATT Ile de France and the Ligue Nationale contre le Cancer, comités d’Ile de France. Ikrame Amara was a graduate student funded by the Cancéropole Ile-de-France. Baxter freely provided Cyclophosphamid. We acknowledge Julien Namur, Florentina Pascale from Archimmed SARL for their advice and help in the present work, and Dr. Jean Bénard PharmD, Ph.D, for his comments and editing.

Author information

Authors and Affiliations

  1. INSERM URM-S 970, Paris, France

    Olivier Pellerin & Marc Sapoval

  2. Université Paris Descartes, Sorbonne Paris Cité, Paris, France

    Olivier Pellerin, Ikram Amara, Marc Sapoval, Philippe Beaune & Isabelle de Waziers

  3. Department of Interventional Radiology, Hôpital Européen Georges Pompidou, Assistance Publique Hôpitaux de Paris, 20 rue Leblanc, 75015, Paris, France

    Olivier Pellerin, Marc Sapoval & Carole Déan

  4. INSERM, UMR-S 1147, Paris, France

    Ikram Amara, Philippe Beaune & Isabelle de Waziers

  5. Department of Pathology, Hôpital Européen Georges Pompidou, Assistance Publique Hôpitaux de Paris, 20 rue Leblanc, 75015, Paris, France

    Tchao Méachi

  6. Department of Biochimistry, Hôpital Européen Georges Pompidou, Assistance Publique Hôpitaux de Paris, 20 rue Leblanc, 75015, Paris, France

    Philippe Beaune

Authors
  1. Olivier Pellerin
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Ikram Amara
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Marc Sapoval
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Tchao Méachi
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Carole Déan
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Philippe Beaune
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Isabelle de Waziers
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Olivier Pellerin.

Ethics declarations

Conflict of interest

This work was supported by SATT Ile de France and the Ligue Nationale contre le Cancer, comités d’Ile de France. Baxter freely provided Cyclophosphamid. Second author was a graduate student funded by the Cancéropole Ile-de-France. Sixth and seventh authors are the inventor of patent PCT/EP2012/058219, US-2014-0127180A1.

Appendix

Appendix

CYP2B6-TM-RED

As described in previous works [8, 12], we built a CYP2B6TM-RED gene (a fusion of a triple mutant CYP2B6 with NADPH cytochrome P450 reductase) that catalyzed very efficiently the transformation of cyclophosphamide (CPA) into cytotoxic metabolites that causes intra- and inter-strand DNA cross-links and cell death.

Transduced-MSCs

Transduced-MSCs were obtained as previously described by Amara et al. [12]. Briefly, rabbit MSCs from bone-marrow of New Zealand White rabbits (Clinisciences, Nanterre, France) were transduced by lentiviral particles expressing luciferase (Luc2-MSCs). We checked that the pool of rabbit MSCs expressed luciferase at high level by the luminescence assay system (Promega, Madison, Wisconsin); this pool was further used to study MSCs distribution after intra-arterial injection (Fig. 3).

Murine MSCs from the bone-marrow of C57BL/6 mice were provided by Life technologies (GIBCO, Saint Aubin, France) and were tranduced by lentiviral particle LV-CYP2B6TM-RED. After two series of limiting dilutions, the most sensitive clone to CPA (clone 42) [13] was used in in vitro (Fig. 2) and in vivo experiments (Figs. 4, 5, 6).

VX2 Cell Liver Implantation

Tumor carriers were induced by intramuscular injection of 107 VX2 cells (CR2i, APHP-INRA, France) in each hind leg of adult New Zealand White male rabbits. Tumor was allowed to grow for 2 weeks. Then the carrier tumors were excised and minced. Aliquots of fresh tumor chunks (~ 1 mm3) were then implanted in the left hepatic lobe of recipient animals as previously described by Lee et al. [14]. Two weeks following implantation, a 15 cm3 single VX2 liver tumor was identified in the left liver lobe by US scans examination, and treatment was initiated.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Pellerin, O., Amara, I., Sapoval, M. et al. Hepatic Intra-arterial Delivery of a “Trojan-horses” Gene Therapy: A Pilot Study on Rabbit VX2 Hepatic Tumor Model. Cardiovasc Intervent Radiol 41, 153–162 (2018). https://doi.org/10.1007/s00270-017-1833-8

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00270-017-1833-8

Keywords

Search

Navigation