Gene therapy

Solid tumor treatment by an efficient suicide-gene

 

Project Leader: Dr Isabelle de Waziers (INSERM Researcher, CR1) and Pr Philippe Beaune (PU-PH, Univ. Paris Descartes/ hEGP)

Scientists Involved: Ikrame AMARA (PhD), Catherine Senamaud (Technician, TCN), Audrey Devillers (Research Engineer, IR).

 

Summary

Our goal is to develop a new method of anti-solid tumors therapy, consisting in sensitization of tumor cells to an anti-cancer drug by transfer of a suicide-gene. Thus, this approach aims at introducing, specifically into tumors, a gene transforming, inside the tumor, a drug (cyclophosphamide, CPA) into toxic metabolites.

We have built up a suicide-gene able to transform very efficiently CPA into toxic metabolites. The sequence of this optimized gene (CYP2B6TM, a mutated cytochrome P450) fused to NADPH-cytochrome P450 reductase (CYP2B6TM-RED) was patented (2012 PCT/EP2012/058219, and USA 2014 US-2014-012718A1).

We first showed in vitro that pulmonary tumor cells were made sensitive to CPA by introduction of our suicide gene. Then, in immuno-competent mouse models, we have shown that, when expressing our suicide-gene in tumors, CPA eradicated completely the tumor without any recurrence for 24 months after stopping the treatment (see figure). Finally we have shown that the immune system contributed to the efficacy of our approach. This implication of the immune system is synergic with the toxicity of CPA metabolites, since it protects against a recurrence of the primary tumor and could also block the development of secondary metastases

We thus have clearly shown that the efficacy of our approach was due to a strong toxicity towards tumor cells, a by-stander effect (diffusion of the toxic metabolites) and an immune response; these effects were not demonstrated in other similar gene-suicide treatments which failed.

To introduce our suicide-gene into the tumor we have tested, as vectorsmesenchymal  stem cells (MSCs) which migrate into the tumors and home there. They can be easily isolated from bone marrow or adipose tissue, without  any particular ethic problem, they can multiply rapidly in culture and the suicide gene can be easily introduced. We used commercially available human cell lines allowing, after transduction with a lentivirus and cloning, to dispose of a stable product. MSCs expressing the suicide-gene (CYP2B6TM-RED) transform, inside the tumor, CPA into toxic metabolites which diffuse into neighboring tumor cells. Thus these toxic products destroy both tumor cells and MSCs avoiding any uncontrolled proliferation and/or dissemination.

We have demonstrated, in mice, that MSCs cells injected into tumors stay there for at least 6 days without any diffusion in other organs and then disappeared at day 14.  Then we showed that intra-tumor administration of MSCs expressing CYP2B6TM-RED followed by CPA induced the complete eradication  of the tumors in  30% of mice without any recurrences more than 6 months later without any other treatment. Moreover the same immune reaction was observed in  cured mice.

These very encouraging results could nevertheless be improved by controlling the implantation of MSCs closeby the tumor. In collaboration with interventional radiologists (Pr. M. Sapoval and Dr O. Pellerin  HEGP Paris), we tested the efficacy of  our approach on a rabbit model with VX2 hepato-carcinoma; transduced MSCs were injected, intra-arterially close to the tumor. After two  injections of MSCs-CYP2B6TM-RED and CPA, tumor regressed significantly  and metastasis was absent in  2/3 rabbits.

These MSCs could be also used to screen the sensitivity of different human cell lines to CPA after CYP2B6TM-RED activation; we have already shown the sensitivity of several types of human tumor cell lines (colon, breast, head and neck). This will help to extend our approach to other types of cancer than hepato-carcinoma which will be our first target.

The demonstration of the efficacy of our approach in vivo in two animal models and in vitro in  several cell lines, is an important asset for the future clinical development  of this approach.

 

Image Isabelle English