MicroRNA‑296 targets AKT2 in pancreatic cancer and functions as a potential tumor suppressor
Authors: Baolin Shi
Jilin Li
Feng Chen
Hailing Li
Journal: Molecular Medicine Reports (2017)
Discipline: CLINICAL MEDICINE
Keywords: microRNA-296; pancreatic cancer; expression level; biological role; mechanism of action
Affiliations: Department of Endocrinology, Weifang People's Hospital, Weifang, Shandong 261000, P.R. China., Department of Medical Laboratory Medicine, Family Planning Guidance Center of Weifang, Weifang, Shandong 261061, P.R. China., Department of Neurology, Weifang People's Hospital, Weifang, Shandong 261000, P.R. China., Department of Endocrine, Weifang Municipal Official Hospital, Weifang, Shandong 261041, P.R. China.

Abstract:  Although microRNA-296 (miR-296) has been studied in various types of human cancer, its expression, biological role and mechanism of action in pancreatic cancer remains to be elucidated. The aim of the current study was to investigate the expression level, possible roles and underlying molecular mechanisms of miR‑296 in pancreatic cancer. The present study revealed that miR‑296 is significantly downregulated in tissue from patients with pancreatic cancer and in human pancreatic carcinoma cell lines, when compared with matched healthy tissue and normal human pancreatic cell lines, respectively. In addition, restoration of miR‑296 expression was revealed to inhibit the proliferation, migration and invasive activity of pancreatic cancer cells. Furthermore, bioinformatics analysis and a luciferase reporter assay validated the AKT2 gene as a direct target of miR‑296 in pancreatic cancer. Reverse transcription‑quantitative polymerase chain reaction and western blot analysis revealed that miR‑296 was able to decrease AKT2 expression at the post‑transcriptional level. Notably, the effects of AKT2 knockdown were similar to miR‑296 overexpression in pancreatic cancer. In conclusion, the present findings indicate a role for miR‑296 as a tumor suppressor in pancreatic cancer through directly targeting AKT2, thus suggesting that miR‑296 may serve as a potential therapeutic target for the treatment of pancreatic cancer. 

Retracted: No

PubPeer comments:
 This publication seems to be weakened by what may be a misunderstanding of microRNA biology and measurement. Each miRNA precursor is processed into one or two  arms, or mature miRNAs, designated with -5p and -3p suffixes. For some miRNA precursors, one arm predominates. For miR-296, -5p and -3p arms can be at similar levels. Interestingly, this manuscript never explicitly identifies the arm that is under study. However, we can gather this from Figure 4, where the sequence for miR-296-5p is shown. Having identified the mature sequence that is being studied, we recognize that his is one of several papers from different author teams that report using the same forward and reverse primers for quantitative PCR of miR-296-5p or miR-296-3p. qPCR is said to be done with what are described as specific primers  (Forward, 5u02b9-TGCCTAATTCAGAGGGTTGG-3u02b9; Reverse, 5u02b9-CTCCACTCCTGGCACACAG-3u02b9) . Unfortunately, no reliable results for miR-296-5p would have been obtained with these primers. Here is the precursor sequence for miR-296, with the miR-296-5p arm (the one that is supposed to be amplified in this paper) in bold: AGGACCCUUCCAG**AGGGCCCCCCCUCAAUCCUGU**UGUGCCUAAUUCAGAGGGUUGGGUGGAGGCUCUCCUGAAGGGCUCU And here in bold is where the published  forward primer sequence would bind: AGGACCCUUCCAGAGGGCCCCCCCUCAAUCCUGUUG**UGCCUAAUUCAGAGGGUUGG**GUGGAGGCUCUCCUGAAGGGCUCU Obviously, this primer does not capture miR-296-5p at all. Also, the reverse primer recognizes a sequence adjacent to the miR-296 precursor in the genome, but this is not part of the precursor RNA. These forward and reverse primers might amplify genomic DNA, or maybe a primary transcript upstream of both the precursor and mature miRNA, but they would not tell us anything about the mature miR-296-5p that is supposed to be under investigation here. ,