ZFP36L2 promotes cancer cell aggressiveness and is regulated by antitumor microRNA‐375 in pancreatic ductal adenocarcinoma

Abstract

Due to its aggressive nature, pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal and hard‐to‐treat malignancies. Recently developed targeted molecular strategies have contributed to remarkable improvements in the treatment of several cancers. However, such therapies have not been applied to PDAC. Therefore, new treatment options are needed for PDAC based on current genomic approaches. Expression of microRNA‐375 (miR‐375) was significantly reduced in miRNA expression signatures of several types of cancers, including PDAC. The aim of the present study was to investigate the functional roles of miR‐375 in PDAC cells and to identify miR‐375‐regulated molecular networks involved in PDAC aggressiveness. The expression levels of miR‐375 were markedly downregulated in PDAC clinical specimens and cell lines (PANC‐1 and SW1990). Ectopic expression of miR‐375 significantly suppressed cancer cell proliferation, migration and invasion. Our in silico and gene expression analyses and luciferase reporter assay showed that zinc finger protein 36 ring finger protein‐like 2 (ZFP36L2) was a direct target of miR‐375 in PDAC cells. Silencing ZFP36L2 inhibited cancer cell aggressiveness in PDAC cell lines, and overexpression of ZFP36L2 was confirmed in PDAC clinical specimens. Interestingly, Kaplan–Meier survival curves showed that high expression of ZFP36L2 predicted shorter survival in patients with PDAC. Moreover, we investigated the downstream molecular networks of the miR‐375/ZFP36L2 axis in PDAC cells. Elucidation of tumor‐suppressive miR‐375‐mediated PDAC molecular networks may provide new insights into the potential mechanisms of PDAC pathogenesis.

Molecular pathogenesis of pancreatic ductal adenocarcinoma: Impact of passenger strand of pre‐miR‐148a on gene regulation

“…Quantitative real‐time PCR (qRT‐PCR) of clinical samples was carried out by extracted total RNA from frozen specimens, immunohistochemistry was performed by PDAC tissues from formalin‐fixed paraffin‐embedded. Two human PDAC cell lines (PANC‐1 and SW1990) were used in this study as described previously …”

“…Protocols for total RNA extraction from clinical specimens and cell lines were described in previous studies. The procedure for quantification of miRNAs and mRNAs was described earlier . TaqMan qRT‐PCR probes and primers were obtained from Thermo Fisher Scientific (Waltham, MA, USA) as follows: miR‐148a‐5p (product ID: 002134); miR‐148a‐3p (product ID: 000470); PHLDA2 (product ID: Hs00169368_m1); LPCAT2 (product ID: Hs01044164_m1); AP1S3 (product ID: Hs00950999_m1); SMS (product ID: Hs01924834_u1); ENDOD1 (product ID: Hs00826684_m1); UHMK1 (product ID: Hs00332674_m1).…”

“…Protocols for functional assays (cell proliferation, migration and invasion) were described previously …”

“…Antibodies against PHLDA2 (product ID: ab99209; Abcam, Tokyo, Japan), poly (ADP‐ribose) polymerase (PARP) antibody (product ID: 9542; Cell Signaling Technology, Danvers, MA, USA) and GAPDH (product ID: SAF6698; Wako) were used in this study. The procedures for western blotting and tissue immunohistochemistry were as described previously …”

“…To identify genes targeted by miR‐148a‐5p and miR‐148a‐3p , we used a combination of in silico analyses and genome‐wide gene expression analyses as described previously . Expression data by microarray analyses were deposited into the Gene Expression Omnibus (GEO) database (accession numbers , ).…”