Decreased expression levels of RAC3 coactivator sensitizes colorectal cancer cells to the effect of chemotherapeutic drugs

M F RUBIO; F.D. ROSA; M.C. LIRA; L.C. PANELO; A.D. SAMBRESQUI; M.C. SALAZAR GÜEMES; M.A. COSTAS

Florencia

Conferencia; 2nd Special Conference EACR AACR SIC, The Challenges of Optimising Immuno and Targeted Therapies From Cancer Biology to the Clinic; 2017

EACR, AACR, SIC

Colorectal cancer (CRC) is one of the most commonly diagnosed cancers in the world. Despite recent advances in chemotherapies that have improved survival rates, patients with late-stage disease or elderly still have a poor prognosis.We investigate the role of RAC3 coactivator in tumor development and we have previously demostrated that RAC3 over expression contributes to inhibition of apoptosis and autophagy, being these two mechanisms involved in resistance to treatment with chemotherapeutic agents.The aim of this study was to evaluate the RAC3 expression levels in three cell lines of CRC and determine their sensitivity to chemotherapeutic drugs.Analyzing GEO data bank (GSE28702), we observed that patients who do not respond to Folic acid-5-Fluorouracil and Oxaliplatin (FOLFOX) treatment have higher RAC3 expression levels than thosepatients who respond to treatment, being this difference significantly greater in metastatic lesions.To explore the potencial role of RAC3 in CRC sensitivity to drugs, we determined the RAC3 expression levels in three human CRC cell lines (HT29, HCT116 and LoVo) by qPCR and Western blot and we showed RAC3 expression was higher in HT29> HCT116> LoVo (44> 9.6> 1-fold respect LoVo by qPCR). Then, we studied sensitivity to 5-fluorouracil (FUra 0-150 μM) and oxaliplatin (Oxa 0-50 μM).Cell viability was determined by crystal violet staining and the IC50 was calculated for each cell type (Oxa: IC50 HT29 0.8±0.2 μM, HCT116 0.6±0.3 μM and LoVo 0.05±0.02 μM; FUra: IC50 HCT116 4.5± 0.6 μM, LoVo 0.6±0.2 μM, HT29 did not respond to treatment with FUra in the doses used). We observed that LoVo were more sensitive to treatment with these drugs.To study whether sensitivity observed in the different CRC lines was due to the expression levels of RAC3, the HCT116 cell line was transfected with an shRNA for RAC3 (shRAC3) and Western blot and qPCR were performed to validate the knockdown efficiency (shRAC3 0.08-fold respect HCT116 control by qPCR).Compared to the control, the shRAC3-transfected group displayed significantly decreased viability (FUra: control 4.5±0.6 μM vs shRAC3 2.4±0.2 μM and Oxa: control 0.6±0.3 μM vs shRAC3 0.17±0.1 μM).In conclusion, our results show that the expression levels of RAC3 influence sensitivity to chemotherapeutic drugs. Therefore, the knowledge of the RAC3 expression levels in tumoral samples could beimportant in order to design new improved therapeutic strategies.