Exosomal transfer of miR-15b-3p enhances tumorigenesis and malignant transformation through the DYNLT1/Caspase-3/Caspase-9 signaling pathway in gastric cancer

Abstract

Abstract Background Exosomes are essential for tumor growth, metastasis, and are used as novel signaling molecules in targeted therapies. Therefore, exosomal miRNAs can be used in new diagnostic and therapeutic approaches due to their involvement in the development of cancers. However, the detailed biological function, potential molecular mechanism and clinical application of exo-miR-15b-3p in gastric cancer (GC) remains unclear. Methods miR-15b-3p mRNA levels in tissues, serum, cells and exosomes were analyzed using qRT-PCR assays. qRT-PCR, immunohistochemical and western blotting analyses were utilized for the determination of DYNLT1 expression. The interrelationship connecting miR-15b-3p with DYNLT1 was verified using Dual-luciferase report, western blotting and qRT-PCR assays. Fluorescent PKH-26 or GFP-Lv-CD63 labeled exosomes, as well as Cy3-miR-15b-3p, were utilized to determine the efficacy of the transfer of exo-miR-15b-3p between BGC-823 and recipient cells. Several in vitro assays and xenograft tumor models were conducted to determine exo-miR-15b-3p impact on GC progression. Results This is the first study to confirm high miR-15b-3p expression in GC cell lines, tissues and serum. Exosomes obtained from 108 GC patient serum samples and GC cell-conditioned medium were found to show upregulation of exo-miR-15b-3p, with the area under the ROC curve (AUC) being 0.820 [0.763–0.876], which is superior to the AUC of tissues and serum miR-15b-3p (0.674 [0.600–0.748] and 0.642 [0.499–0.786], respectively). In addition, high exo-miR-15b-3p expression in serum was found to accurately predict worse overall survival. SGC-7901 and GES-1 cells are capable of internalizing BGC-823 cell-derived exosomes, allowing the transfer of miR-15b-3p. Migration, invasion, proliferation and inhibition of apoptosis in vitro and in vivo were enhanced by exo-miR-15b-3p, by restraining DYNLT1, Cleaved Caspase-9 and Caspase-3 expression. Conclusions This study identified a previously unknown regulatory pathway, exo-miR-15b-3p/DYNLT1/Caspase-3/Caspase-9, which promotes GC development and GES-1 cell malignant transformation. Therefore, serum exo-miR-15b-3p may be a potential GC diagnosis and prognosis biomarker, which can be used in precise targeted GC therapy.

Keywords

• Exosomes
• miR-15b-3p
• Gastric cancer
• DYNLT1
• Apoptosis
• Cleaved Caspase-3