Molecular predictors of brain metastasis-related microRNAs in lung adenocarcinoma
Authors: Weimin Zhang
LuHua Wang
Zhiwu Wang
Xin Dong
Zitong Zhao
Yongmei Song
Jing Fan
Wei Zhou
Guogui Sun
WenJue Zhang
Xiao Ding
Nan Bi
Qimin Zhan
Lihong Wu
Ning Lv
Jingbo Wang
Journal: PLoS Genetics (2019)
Discipline: MOLECULAR BIOLOGY & GENETICS
Keywords: Brain metastasis; lung adenocarcinoma; miRNAs; tumorigenesis; molecular markers
Affiliations: Department of Radiation Oncology, National Cancer Center/Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China., State Key Laboratory of Molecular Oncology, National Cancer Center/Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China., Department of Radiation Oncology, North China University of Science and Technology Affiliated People's Hospital, Hebei, China., Department of Radiation Oncology, Shandong Provincial Hospital, Shandong, China., Department of Pathology, National Cancer Center/Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China., Laboratory of Molecular Oncology, Peking University Cancer Hospital and Institute, Beijing, China.

Abstract:  Brain metastasis (BM) is a major complication of lung adenocarcinoma (LAD). An investigation of the pathogenic mechanisms of BM, as well as the identification of appropriate molecular markers, is necessary. The aim of this study was to determine the expression patterns of microRNAs (miRNAs) in LAD with BM, and to investigate the biological role of these miRNAs during tumorigenesis. miRNA array profiles were used to identify BM-associated miRNAs. These miRNAs were independently validated in 155 LAD patients. Several in vivo and in vitro assays were performed to verify the effects of miRNAs on BM. We identified six miRNAs differentially expressed in patients with BM as compared to patients with BM. Of these, miR-4270 and miR-423-3p were further investigated. miR-4270 and miR-423-3p directly targeted MMP19 and P21, respectively, to influence cell viability, migration, and colony formation in vitro. miR-4270 downregulation and miR-423-3p upregulation was associated with an increased risk of BM in LAD patients. Thus, our results suggested that miR-4270 and miR-423-3p might play an important role in BM pathogenesis in LAD patients, and that these miRNAs might be useful prognostic and clinical treatment targets. 

Retracted: No

PubPeer comments:
 Fig.7F: Migration assay image of ct-mimic+ct-plasmid group reappeared in mimic+P21-plasmid. Could authors give an explanation? ![file] , 
 Fig.2C and Fig.3C of this paper shows two mice, IVIS images showed Inhibitor or Mimic of which carries very large tumors on its side. Line charts indicated that the tumors were smaller than 5mm3 in size - but they appear to be much larger on the IVIS photos. Could the authors clarify how big (mean and range) the tumors were obtained for this study? Were these in line with animal ethics guidelines? ![file] , 
 #1 Dear Cicindela Chrysippe, Thank you for your interest of our study and pointing this mistake out. It s strange that we have responded to your comments a few days ago, but for unknown reasons, the website only displayed the u201cAuthor responseu201d, but not reply content. To avoid any confusion, I use my colleague s account to reply again. Figure 7F showed migration assay results of H1299 cell line when received with different treatments. Regrettably, we put the same picture for control mimic + control plasmid group and miR-423-3p mimic + P21 plasmid group by mistake. We have checked the original images of these assays, and found that this picture was actually for miR-423-3p mimic + P21 plasmid group, in other words, we didnu2019t include the correct image for control mimic + control plasmid group. However, we should declare that this mistake was due to copy-and-paste error, the results and the conclusions of this assay were not affected. We have corrected the Figure 7F in the following figure and contacted the editor office of PLoS Genetics to make correction of this figure. Once again, many thanks for your comment. ![file] Fig 7. Rescue assay of miR-4270 and miR-423-3p. (A) The mRNA and protein expression levels of MMP-19. (B) The results of the migration assay after the co-transfection of cells with the miR-4270 mimic and the pEGFP-C1 plasmid containing an MMP-19 CDS sequence. (C) MMP-19 protein expression, as measured by immunohistochemical staining, in LAD samples. (D) Spearman correlation analysis showing the negative correlation between MMP-19 and miR-4270 expression. (E) The mRNA and protein expression levels of P21. (F) The results of the migration assay after the co-transfection of cells with the miR-423-3p mimic and pEGFP-C1 plasmid containing a P21 CDS sequence. (G) P21 protein expression, as measured by immunohistochemical staining, in LAD samples. (H) Spearman correlation analysis showing the negative correlation between P21 and miR-423-3p expression. Data are presented as the means u00b1 SD of three experiments. ** P 0.01. #2 In this study, we used luciferase expression to track growth of xenografts and bioluminescent signal were acquired by IVIS system. The photon flux from the tumor is proportional to the number of light emitting cells and the signal can be measured to represent the relative tumor size. However, the emission light from the tumor expressed luciferase will scatter through the tissue, showing a larger halo than the actual tumor size. The maximum diameter of any xenograft did not exceed 2.0 cm, which was consistent with the Institutional Animal Care and Use Committee (IACUC) Guideline of many universities, such as UCSF , University of Pennsylvania , Purdue University , et al. In this study, all animal care and procedures were in accordance with national and institutional policies for animal health and well-being. Mouse experimentations were approved by the Animal Care and Use Committee of the Institute. All mouse surgery was performed under anesthesia, and all efforts were made to minimize suffering of animals. Best Regards, Guogui Sun , 
 *[above]* Figs 3g h and 5c from  MiR-423-5p in brain metastasis: potential role in diagnostics and molecular biology ([Sun et al 2018] ) (*again*). *[middle]* Fig 5c d from  MicroRNA-1253 suppresses cell proliferation and invasion of non-small-cell lung carcinoma by targeting WNT5A ([Liu et al 2018] ). *[below]* **Fig 4c g**. ![] The descriptions of the duplicated panels do not seem to be consistent. , 
 *[left]* Fig 2H from  miR-769-5p suppressed cell proliferation, migration and invasion by targeting TGFBR1 in non-small cell lung carcinoma ([Yang et al 2017] ). *[right]* **Fig 5C**. ![file] , 
 For the sake of completeness, a few last overlaps between - Fig 3g h from  MiR-423-5p in brain metastasis: potential role in diagnostics and molecular biology ([Sun et al 2018] ). - Fig 5d from Sun et al. - **Fig 7F**. ![file] Panels in the first column of 5d and the first column of 7F also match, but they are described in both cases as a control condition with H157 cells, so not concerning. ,