Targeted knockdown of IQGAP1 inhibits the progression of esophageal squamous cell carcinoma in vitro and in vivo
Authors: Kang Wang
Xiao-Xia Wang
Yan Zhao
Chong-Xiao Qu
Zhi-Rong Liu
Hui-Zhen Wang
Xiao-Zhong Li
Xu-Hong Wang
Li-Qin Zhai
Li-Wei Jing
Qi-Jun An
Journal: PLoS ONE (2014)
Discipline: Multidisciplinary
Keywords: IQGAP1; tumorigenesis; tumor progression; esophageal squamous cell carcinoma; overexpression
Affiliations: Department of Biochemistry and Molecular Biology, Shanxi Medical University, Taiyuan, China., Department of Emergency, Shanxi Provincial People's Hospital, Taiyuan, China., Department of Pathology, Shanxi Provincial People's Hospital, Taiyuan, China., Department of Statistics, Shanxi Medical University, Taiyuan, China., Department of Surgery, Shanxi Cancer Hospital, Taiyuan, China.

Abstract:  IQGAP1 is a scaffolding protein that can regulate several distinct signaling pathways. The accumulating evidence has demonstrated that IQGAP1 plays an important role in tumorigenesis and tumor progression. However, the function of IQGAP1 in esophageal squamous cell carcinoma (ESCC) has not been thoroughly investigated. In the present study, we showed that IQGAP1 was overexpressed in ESCC tumor tissues, and its overexpression was correlated with the invasion depth of ESCC. Importantly, by using RNA interference (RNAi) technology we successfully silenced IQGAP1 gene in two ESCC cell lines, EC9706 and KYSE150, and for the first time found that suppressing IQGAP1 expression not only obviously reduced the tumor cell growth, migration and invasion in vitro but also markedly inhibited the tumor growth, invasion, lymph node and lung metastasis in xenograft mice. Furthermore, Knockdown of IQGAP1 expression in ESCC cell lines led to a reversion of epithelial to mesenchymal transition (EMT) progress. These results suggest that IQGAP1 plays crucial roles in regulating ESCC occurrence and progression. IQGAP1 silencing may therefore develop into a promising novel anticancer therapy. 

Retracted: Yes

PubPeer comments:
 Figure 5A. * Red boxes: The KYSE control and the K2 panels appear to show a region of overlap. Reported to the journal in May 2014. ![file] , 
 In addition, some tumors shown in Figure 6A B appear to be larger than 2 cm in diameter (the maximum diameter in the guidelines I am familiar with), while there is advanced ulceration. Could the authors please comment on the ethical approval and implementation of these animal experiments? ![file] , 
 Dear Dr. Bik, Thank you very much for your pointing out this mistake. We apologize to the readers for this issue. Because of negligence, we made a mistake without awareness when editing the picture for Figure 5A, resulting in a wrong image of the 0 h KYSE150 control panel. Here we provide the corrected Figure 5A as below. This correction doesnu2019t change the result and conclusion. We apologize for any confusion it may have caused. Thanks again for your comment. ![file] , 
 Dear Dr. Bik, We are very sorry about this. In this article we wanted to investigate the effect of IQGAP1 knockdown on tumor cell growth in vivo, subcutaneous tumour transplantation was used that is an accepted experimental procedure for studying various cancers and cancer treatments. Further, we wanted to study the effect of IQGAP1 knockdown on tumor cell metastasis in vivo. There are mainly three models to study metastasis in nude mice. Experimental metastasis, as a model of metastasis refers to the injection of tumor cells directly to the systemic circulation. However, bolus injection of cells does not mimic natural dissemination. Moreover, experimental metastasis will increase numbers of animals in the experiment. Orthotopic transplantation refers to the delivery of cancer cells to the anatomic location or tissue from which a tumor was derived. The use of orthotopic injection transplantation has resulted in tumor models that may more closely resemble human cancers. However, this model requires delicate surgical procedures which not only increases numbers of animals used in experiment but also increases risk of infection and pain. Some studies reported that spontaneous metastases maybe develop from subcutaneous tumour of nude mice through a long period of observation. According to the above reports, so we applied subcutaneous tumour transplantation to observe the growth and spontaneous metastasis of tumor cell. Moreover, it will not increase numbers of animals and avoid pain and injury caused by surgery just as orthotopic injection transplantation. All animal experiments should incorporate, as far as possible, the 3Rs: replacement (of animals with alternative methods), reduction (in the numbers of animals used to achieve scientific objectives) and refinement (of methods to minimize animal suffering). However, we really neglect the size of the tumor limits because tumor size reported in some literature exceeds standard limits for mouse tumor studies. We are so sorry for the mistakes. We certainly abide by scientific and ethical requirements and fully consider the animalsu2019 welfare in future scientific research. Once again, we highly appreciate your comments. , 
 Retracted [20 April 2022] ------------------- Following the publication of this article [1], concerns were raised regarding the methods and results. Specifically, - In Fig 6, tumor volume appears to exceed 2000 mm3, and tumors appear ulcerated. The tumor data raise concerns about animal welfare and the adherence of the study to animal research ethics standards. - In the Methods section, the sequence of the control siRNA used in the siRNA knockdown experiment was not provided. - In Fig 5A, the KYSE150 control and K2 0 h panels appear to partially overlap. - From the raw western blot image data provided during discussions, it appears that the following cropped panels did not originate from the same location on the membrane or were from different membranes: - - Fig 2A: IQGAP1 and u03b2-actin panels. - - Fig 2C: IQGAP1 and u03b2-actin panels for KYSE150. - - Fig 4B: all panels. - - S1A Fig: IQGAP1 and u03b2-actin panels for EC9706 and KYSE150. The corresponding author acknowledged that tumor volume was calculated incorrectly in the published figure, and they provided updated graphs presenting tumor volume calculated using the following formula: Length*Width2 2. In the updated graphs, the control groups exceed 2000 mm3. The corresponding author provided details about welfare checks and stated that they were performed initially three times per week, and daily when tumors became larger. They stated that in the final two weeks of the experiment, some mice lost weight. The size and condition of tumors, and the fact that some mice lost weight, call into question whether the humane endpoint criteria or monitoring frequency were sufficient to mitigate animal welfare concerns in the reported experiments. The PLOS ONE Editors consulted with a member of the Editorial Board, who confirmed that tumor volumes reported in the updated graphs for Fig 6 exceed internationally-accepted standards. Concerns regarding tumor volume indicate that the article is not in compliance with the PLOS Animal Research policy which requires that, u201cstudies involving animals must be conducted according to internationally-accepted standards.u201d The corresponding author provided the following sequence for the control siRNA used in the siRNA knockdown experiment: 5u2019-TTCTCCGAACGTGTCACGT-3u2019. For Fig 5A, the corresponding author stated that the KYSE150 Control 0 h panel was included in error. They provided a replacement panel, along with the raw data for this figure. For western blot panels in Figs 2A and 4, the corresponding author confirmed that proteins were measured on different membranes. For Fig 4, they explained that this was due to difficulty probing for proteins of a similar size. They originally attempted to strip and re-probe the same membrane but were unsuccessful. For Figs 2C and S1A, the corresponding author stated that IQGAP1 and u03b2-actin were measured on the same membranes, but different lanes were presented for each protein to improve presentation. In light of the above concerns regarding animal welfare which indicate that the article is not in compliance with the PLOS Animal Research policy, the PLOS ONE Editors retract this article. The editors regret that the issues in this article were not identified prior to publication. XXW agreed with the retraction, stands by the articleu2019s findings, and apologizes for the issues with the published article. All other authors either did not respond directly or could not be reached. ,