EWS-FLI1 targeted vectorized antisense oligonucleotides inhibit a model of Ewing sarcoma in mice

A. Maksimenko phD bc, G. Lambert phD a, J.R. Bertrand phD b, E. Fattal phD a, F. Subra phD b, H. Pinto-Alphandary phD a, C. Malvy phD b, C. Auclair phD b, P. Couvreur phD a

aLaboratoire de physico-chimie, pharmacotechnie et biopharmacie, UMR CNRS 8612, Faculté de Pharmacie, Chatenay Malabry, France b Laboratoire de physico-chimie et pharmacologie des macromolécules biologiques, UMR CNRS 8532, Institut G. Roussy, 94800 Villejuif, France cBioalliance Pharma SA, 59, Boulevard du Général Martial Valin, 75015, Paris

Aim : It is to block in a very specific way in mice the growth of tumors generated by EWS-Fli1. Vectorized antisense oligonucleotides (ODNs) complementary of the junction of both genes at the mRNA level allow this inhibition because the target is only present in the cancer cells. Methods : We have used two types of vectors for ODNs : poly-isobutylcyanoacrylate nanocapsules and transdrug® (Bioalliance Pharma). One million EWS-Fli1 transformed NIH 3T3 cells were injected subcuteanously to irradiated nude mice where a tumor appears after two weeks. Several injections of vectorized ODNs at non toxic doses were then performed in the tumor within a period of two weeks. The protection of ODNs by the vectors was measured using denaturing polyacrylamide gel electrophoresis after incubation of radioactively labeled ODNs in various serums. Results : We have used two antisense ODNs : Firstly a full phosphorothioate ODN vectorized by nanocapsules, secondly a structured chimeric phosphodiester phosphorothioate ODN vectorized by transdrug®. Both vectors improve the resistance of ODNs to degradation. Both antisense vectorized ODNs significantly inhibit the tumor growth in mice at a total dose ranging between 10 and 15 nanomoles whereas either the non vectorized antisense ODNs or the vectorized control ODNs have no effect. Conclusions : Thanks to the use of vectors associated with antisense ODNs we show that it is possible to inhibit the growth of a tumor in a very specific way. Tanaka et al (J.Clin Invest. 99, 239-247) have obtained a similar result with a 35 to 50 times higher dose of non vectorized phosphorothioate ODN targeted against the EWS part of EWS-FLI1, therefore much less specific than our junction targeted ODN. Our result shows that it should be possible to treat with specificicity tumors caused by chromosomic translocations without any major toxicity.

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Paper presented at the International Symposium on Predictive Oncology and Intervention Strategies; Paris, France; February 9 - 12, 2002; in the section on Novel Therapies, Part 1.