Ravi Chavali

Introduction: Colorectal cancer is the second leading cause of death among cancers that affect both men and women¹. In most metastatic cancer patients, failure of chemotherapy is due to multidrug resistance (MDR) arising due to overexpression of ATP binding cassette (ABC) transporters that efflux out the therapeutic drugs from within the cells^2,3.  This reduces intracellular concentration of anti-cancer drugs too low to effectively interact with target molecules to have a therapeutic effect. Direct inhibition of these transporters, while successful in-vitro, has not been clinically successful due to undesired side effects and toxicity⁴. Recent efforts have identified some mechanisms – Human antigen R⁵, Epithelial to Mesenchymal Transition (EMT) ^6-9  – responsible for overproduction of ABC transporters in colorectal cancer^5-9 . This study aims to determine the role of Twist1, a transcription regulator of EMT, in MDR of vincristine resistant cancer cells⁶. Methods. Three types of colon cancer cells  – Immortalized: NCM460; Low Invasive: HT-29; High Invasive: HCT-8 – were used to determine the basal expression of ABC transporters and Twist1 in these cells. Vincristine resistant colon cancer cells, HCT-8/V, were used to demonstrate existence of EMT and MDR in these cells. HCT-8 cells were used to establish vincristine treatment induces EMT and MDR. Over expression of Twist1 in HCT-8 was used to determine if Twist1 caused EMT in these cells. siRNA was used for silencing Twist1 in HCT-8/V. HCT-8/V tumor-bearing mice were used to verify drug reversal effects of Twist1 silencing in-vivo. Results were considered statistically significant for P <0.05. Results. Basal expression of ABC transporters and Twist1 is highest in HCT-8 cells compared to NCM460 and HT-29 cells. Vincristine treatment of HCT-8 cells induced EMT and MDR. Overexpression of Twist1 in HCT-8 cells induced EMT and MDR. Silencing Twist1 in HCT-8/V cells reversed EMT and MDR as well as reduced expression levels of ABC transporters. Twist1 silencing treatment of HCT-8/V tumor bearing mice resulted in reduced tumor growth, lung metastasis and expression of ABC transporters. Conclusions. Twist1 plays an important role in EMT of colon cancer cells and their MDR. Its expression is upregulated in MDR colon cancer cells. Overexpression of ABC transporters is directly related to overexpression of Twist1. Silencing Twist1 in vincristine resistance cells has reversed EMT and increased their chemosensitivity providing a possible target for ABC transporter associated MDR in colon cancer treatment.

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