Inhibitory Effect of the Multi-target TKI, Anlotinib, in 5-FU Resistant Colorectal Cancer HCT-8/15 Cells: Down Regulation of Drug Resistance-associated Protein Expression.

Purpose Colorectal cancer is the most prevalent gastrointestinal malignancy. Treatment usually includes 5-fluorouracil (5-FU), oxaliplatin, and irinotecan, with 5-FU usually being the first choice. 5-FU treatment failure occurs when cancer cells acquire resistance. Therefore, it is crucial to identify compounds effective against 5-FU-resistant tumors. Herein, we determined the efficacy and mechanism of anlotinib in 5-FU-resistant colon cancer cells. Materials and methods Human colon cancer cells (HCT-8/5-FU and HCT-15/5-FU) resistant to 5-FU were subjected to treatment with anlotinib, 5-FU, or both. Cell proliferation was assessed via MTS and clone formation assays. Cell cycle progression was studied using flow cytometry. Through immunoblotting, we evaluated changes in the protein levels of p-AKT and multidrug resistance 1. Results MTS assays indicated that HCT-8/5-FU and HCT-15/5-FU cells were sensitive to anlotinib and resistant to 5-FU. At 48 h, HCT-8/5-FU had an IC50 of 2246.5 ± 204.5 µM, while HCT-15/5-FU had an IC50 of 18.49 ± 3.23 mM for 5-FU. The IC50 of anlotinib for HCT-8/5-FU cells was 53.69 ± 8.10µM at 24 h and 17.39 ± 1.98µM at 48 h. The IC50 values for HCT-15/5-FU at 24 and 48 h were 55.03 ± 3.44µM and 8.83 ± 3.02µM, respectively. Anlotinib enhanced 5-FU sensitivity in resistant cells, with low concentrations (IC10) considerably enhancing the antiproliferative effects of 5-FU. Further, anlotinib significantly increased the number of cells in the G0-G1 phase dose-dependently, while the proportion of cells entering S phase decreased. MDR1 and AKT expression decreased with increasing anlotinib concentration. Conclusion Anlotinib suppressed the proliferation of 5-FU-resistant colon cancer cells by preventing entry into S phase, thus sensitizing cells to 5-FU. Moreover, anlotinib may reverse the effect of 5-FU on drug-resistant cells by down-regulating the expression of multidrug-resistant proteins, in which the AKT signaling pathway may play an important role.