Suppressive effect of an analog of the antimicrobial peptide of LL-37 on colon cancer cells via exosome-encapsulated miRNAs

Antimicrobial peptides (AMPs) are multifunctional factors with an important role in the innate immune system. Our previous studies revealed that the human cathelicidin LL-37 and its analog, FF/CAP18, limit the proliferation of colon cancer cell lines. In the present study, the exosomes released by HCT116 cells treated with FF/CAP18 were analyzed. After the treatment, exosomes were isolated from the culture supernatant by ultrafiltration and using the miRCURY™ Exosome Isolation Kit. Membrane vesicles 40-100-nm expressing CD63 and CD81 were identified before and after FF/CAP18 treatment. Exosome concentration in the culture supernatant was increased after treatment with FF/CAP18. Exosomes formed in HCT116 cells treated with FF/CAP18 induced growth suppression of the cells in a dose-dependent manner. By contrast, the exosomes formed in non-treated HCT116 cells did not affect cell viability. Microarray analysis of miRNA expression indicated that FF/CAP18 treatment induced increases in the expression of three miRNAs (miR-584-5p, miR-1202 and miR-3162-5p) in both HCT116 cells and exosomes. These results suggest that FF/CAP18 treatment increases exosome formation, and that exosome-encapsulated miRNAs suppress HCT116 cell proliferation. Exosomal miRNAs are considered to be involved in the dissemination of cell signals to control local cellular microenvironments. The present findings suggest that FF/CAP18 regulates cancer growth by modulating cell-to-cell communication. AMPs localize in the cytoplasm of cancer cells and enhance the expression of growth-suppressing miRNAs. These miRNAs are also transported to other cancer cells via exosomes. Therefore, transportation of these miRNAs has the potential to suppress cancer growth. AMPs exert their effects directly by targeting cancer cells and indirectly via exosomes.