Abstract 2594: Exosomes co-expressing aquaporin-5-targeting miRNAs and IL-4 receptor-binding peptide inhibit the migration of human breast cancer cells

Abstract

Abstract Water channel aquaporins (AQPs) play a critical role in transcellular water transport. Recent studies revealed that dysregulation of AQPs is highly correlated with the migration of cancer cells and metastasis of tumor in vitro and in vivo models. In particular, we have demonstrated that increased expression of aquaporin-5 (AQP5) is associated with breast cancer cell migration, metastasis, and poor prognosis in human patients. We aimed to identify novel AQP5-targeting miRNAs and to examine the strategy for the post-transcriptional regulation of AQP5 using exosome-mediated microRNA (miRNA) delivery. Multiple tools of bioinformatics were used to identify AQP5-targeting miRNAs and miRNA target enrichment analysis revealed the cellular signaling pathways associated with the identified miRNAs. To examine the exosome-mediated miRNA delivery systems, exosomes containing both AQP5-targeting miRNAs and a peptide (CRKRLDRNC: IL-4RPep1) targeting the interleukin-4 receptor (IL-4R), which is highly expressed in breast cancer cells, were applied to triple-negative breast cancer cell line MDA-MB-231. Putative AQP5-targeting miRNAs were identified using DIANA Tool. Thereafter, three additional bioinformatic tools (TargetScan, miRDB and MiRanda Tools) were applied to select the miRNAs that were commonly identified from at least 3 different programs. Three miRNAs, miR-1226-3p, miR-19a-3p and miR-19b-3p, were identified as putative regulators of AQP5. Protein expression of AQP5 was decreased when mimic of each miRNA was transfected into MDA-MB-231 cells. Quantitative real-time PCR demonstrated a decrease of AQP5 mRNA expression by miR-1226-3p mimic and luciferase reporter assay revealed a reduction of AQP5 translation by miR-19b mimic in MDA-MB-231 cells. Consistently, cell migration was inhibited by mimic of each miRNA. miRNA target enrichment analysis showed that target genes of three miRNAs were predominantly enriched in the gap junction regulatory pathway. This suggested that these miRNAs are associated with breast cancer cell migration and, also, they might be involved in cell-cell adhesion. For the delivery of AQP5-targeting miRNAs to MDA-MB-231 cells, exosomes containing both AQP5-targeting miRNAs and a peptide (IL-4RPep1) were constructed in HEK293T cells. Both AQP5 protein expression and cell migration were significantly decreased in MDA-MB-231 cells when they were incubated with the exosome-containing culture media obtained from the culture of HEK293T cells expressing both AQP5-targeting miRNAs and IL-4RPep1. In conclusion, AQP5-regulating miRNAs are identified and their target genes are found to be mainly involved in the gap junction regulatory pathway. The exosome-mediated delivery of AQP5-targeting miRNAs could be exploited for the inhibition of breast cancer cell migration associated with AQP5. Citation Format: Eui-Jung Park, Hyun Jun Jung, Hyo-Jung Choi, Hyo-Ju Jang, Hye-Jeong Park, Tae-Hwan Kwon. Exosomes co-expressing aquaporin-5-targeting miRNAs and IL-4 receptor-binding peptide inhibit the migration of human breast cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2594.