The Immunosuppressive Drug Rapamycin Inhibits Several Molecular Mechanisms Involved in Tummor Development

PANELO L.C.; ALVARADO C V; RUIZ GRECCO M; RUBIO M F; FERNANDEZ LARROSA P N; AGUIRRE C; COSTAS M. A.

Berlin

Congreso; 24 TH International Congress of Transplantation Society; 2012

Transplantation Society

Immunosuppressive drugs are used to prevent organ rejection in transplanted patients, but this is associated with a high incidence of malignant tumors. However, the immunosuppresive drug Rapamycin (Rapa) is currently been investigated because their anti-tumor effect. It can inhibit tumor growth, angiogenesis and metastasis through inhibition of AKT / mTOR pathway.RAC3 is a member of the p160 coactivator family. It was first described as steroid receptor coactivator and its overexpression was associated to the development of hormone-dependent tumors. However, we then demonstrated that RAC3 is also a coactivator of the transcription factor NF-kB, which regulates the expression of genes involved in tumorigenesis. Then, RAC3 overexpression was found linked to the development of hormone-independent tumors. Additional evidences demonstrated latter that RAC3 is as an oncogene involved in tumorigenesis through several mechanisms that involves an anti-apoptotic role, the cell growth increase and metastasis. Although we have previously observed that renal tumors show high levels of RAC3, it is unclear how its expression is regulated. In this work we investigated the effect of Rapa on cell proliferation and senescence and the regulation of the RAC3 expression levels. The non tumoral human embrionary kidney HEK293 cell line with low levels of RAC3 and the human colon tumoral T84 cell line, that naturally overexpresses RAC3 were treated or not with Rapa 0.5mM for 24 and 48 h, and surviving was then analyzed by staining with crystal violet. The results show that Rapa can inhibit proliferation in both lines (p < 0.01). This is consistent with the increase of senescent cells: the non tumoral WI38 cells were treated or not with 50 nM Rapa for 3 days and then senescence was determined by the appearance of large heterochromatic nuclei or acid b-Gal activity. We found that Rapa induces premature senescence (p< 0.01). Because RAC3 exerts an anti-apoptotic role, we decided to investigate whether Rapa can reverse this effect. HEK cells with low or high levels RAC3 (transfected with an expression vector for the constitutive expression of this coactivator) were treated or not with H2O2 1.5mM for 24 h in the presence or absence of Rapa 100nM. The results showed that Rapa inhibits the antiapoptotic effect of RAC3. Finally, we investigated if Rapa is able to regulate the expression levels of RAC3. The HEK293 cells were stimulated or not with Rapa 100 nM. The RAC3 levels were determined by western blot and q-PCR. We found that Rapa inhibits RAC3 expression (p< 0.01). Similar results were obtained in WI38 cells treated or not with Rapa 50nM for 1, 3 or 6 days. In view of these results, Rapa is an immunosuppressive drug that inhibits all the actions that are stimulated by RAC3 overexpresion, like growth and apoptosis protection, as well as the coactivator expression. Because RAC3 is usually overexpressed in almost all the tummor types, these results support the use of Rapa as a good antitumor agent and immunosuppressive drug in transplanted patients.