Low miR-224-5p in exosomes confers colorectal cancer 5-FU resistance by upregulating S100A4

This study aimed to identify molecular markers that mediate 5-fluorouracil (5-FU) resistance in colorectal cancer (CRC). Exosomes from 5-FU resistant CRC cells (HCT-15/FU) significantly enhanced the resistance to 5-FU and the malignant properties of HCT-15 cells. Double screening of miRNAs in CRC cell-exosomes and serum-exosomes from clinical CRC patients revealed that miR-224-5p was expressed at significantly lower levels in the 5-FU resistant type than in the 5-FU sensitive type. Moreover, the overall survival rates of 5-FU-resistant CRC patients were much lower than those of 5-FU-sensitive CRC patients. Furthermore, cellular miRNA sequencing (miR-Seq) and proteomic studies revealed that several miRNAs such as miR-224-5p, were significantly downregulated and that calcium-related proteins, including S100 calcium-binding protein A4 (S100A4), were upregulated in HCT-15/FU cells. An analysis of data from public databases revealed that patients with CRC with lower S100A4 expression had a better prognosis. In addition, miR-224-5p was shown to directly target S100A4. Functionally, in vitro and in vivo experiments verified that the downregulation of miR-224-5p promoted malignant properties and resistance to 5-FU in HCT-15 cells, whereas the upregulation of miR-224-5p in HCT-15/FU cells attenuated these effects. Notably, 5-FU combined with verapamil reversed 5-FU resistance in CRC by regulating the miR-224-5p/S100A4 pathway. Triptolide inhibited the malignant properties of HCT-15/FU cells by affecting the miR-224-5p/S100A4 axis. Overall, miR-224-5p is involved in CRC 5-FU resistance by regulating S100A4, and might serve as a molecular marker for the early prediction and intervention of 5-FU resistance in CRC patients in the clinic. Triptolide or 5-FU combined with a calcium antagonist could be used as a trial therapy for 5-FU resistant CRC patients.