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The authors have made the requested changes.
[# PeerJ Staff Note - this decision was reviewed and approved by Paula Soares, a PeerJ Section Editor covering this Section #]
As commented by reviewer 1, text in several images are a little difficult to read. Please revise these images to assist the reader.
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1. While there have been improvements compared to previous drafts,
there are still a few areas where the text is small and difficult to read.
I suggest to enlarge the size of texts in several figures.
2. In Supplementary Table 1, there is a gene with an unfamiliar notation. This is probably due to an automatic correction by the spreadsheet software when it should be SEPTIN11. Please rewrite it to the correct gene name.
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The reviewers has indicated that a few minor changes still need to be made before the manuscript can be considered for publication.
I think it is good that this manuscript is more detailed than the first draft.
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In the added Fig. 3G, the data showed that the expression level of ANXA9 was lower in diffuse gastric cancer specimens than in intestinal specimens. Can you confirm the possibility that the expression of ANXA9 is elevated in the intestinal specimen but is less elevated in the diffuse specimen when normal and tumor tissues are paired?
The size of the dots in Figure 3 is different in several figures, and I request that they be unified.
Although improved, Figures 8 and 9 are still difficult to read. I suggest that the methylation levels of the ANXA9 promoter region and expression levels of that shown in Figure 8 be shown in a separate figure as a scatter plot.
In Fig. 9a and b, the gene names are illegible due to the small size of the diagrams. We suggest that a part of the network diagram be enlarged to create a diagram that clearly shows the gene names.
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Authors made notable changes, however, in line 222, it is better to mention that also lower expression of ANXA9 in UCEC is correlated with good prognosis and better OS.
As the reviewers indicated, this work is indeed interesting. The reviewers have identified a few areas where the work could be strengthened further. For example, one of the co-author's previous work related to CRC (Ref 20) as well as reference 19, indicate an opposite effect on survival in CRC compared to GC. Work on this, potentially using gastric cancer cell lines, and a detailed discussion would indeed strengthen the manuscript. Additionally, there are a handful of language/grammar errors. Since copy editing is not provided as a standard service, please ensure that the manuscript is thoroughly proofread to fix these errors. I look forward to receiving your revised submission.
[# PeerJ Staff Note: Please ensure that all review and editorial comments are addressed in a response letter and any edits or clarifications mentioned in the letter are also inserted into the revised manuscript where appropriate. It is a common mistake to address reviewer questions in the response letter but not in the revised manuscript. If a reviewer raised a question then your readers will probably have the same question so you should ensure that the manuscript can stand alone without the response letter. Directions on how to prepare a response letter can be found at: https://peerj.com/benefits/academic-rebuttal-letters/ #]
[# PeerJ Staff Note: The Academic Editor has identified that the English language must be improved. PeerJ can provide language editing services - please contact us at [email protected] for pricing (be sure to provide your manuscript number and title) #]
I think it is a very interesting finding that in gastric cancer, the prognosis is worse in the ANXA9 low-expression group, contrary to that in colon cancer. However, I feel that the impact of the other analysis results is weak, so I think the paper will be more accurate if the authors conduct experiments using knockdown and forced expression of ANXA9 in gastric cancer cell lines, and then add a discussion of this confliction between in gastric cancer and colon cancer.
Authors noted that the clinical specimens were analyzed using UALCAN website and that the classification by some clinical information did not show significant differences in ANXA9 expression. I propose to test whether significant differences in the expression of ANXA9 occur when using the gene expression-based classifications (GC, MSI, CIN, and EBV) or the Lauren classification (diffuse or intestinal) described in the TCGA report (Nature 513, 202–209 (2014). https://doi.org/10.1038/nature13480).
The authors analyzed most of the data sets except for Figure 1 and 2 using the TCGA data set; does the data set registered in GEO contain any survival data or clinical information? I think that adding the validation results of other datasets will make the results more reliable.
The five boxplots in Figure 1 are all different in size. It would be better to unify them.
The captions in Figure 1 and Figure 4 are difficult to read because the text is small and the font is not properly selected.
In Fig. 8, the width of the image is wider to fit the Legend, but I think this unfriendly due to the small size of the heatmap.
In Figure 7b, the red text is not only difficult to decipher because it is covered by dots, but also some of the numbers are obscured. I suggest that the correlation coefficient and p-value be listed elsewhere.
In Figure 9B, the individual gene names cannot be deciphered because the letters overlap. How about adding the gene names to the volcano plot in Figure 9A and omit Figure 9B?
There are some typos of gene names.
1-The English language should be improved.
2-In some parts authors mentioned AHNAK2 while the study was related to the ANXA9 gene, for example in abstract.
3-The figure quality could be improved.
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Did Author use the same tumour samples compared to their matched normal tissues for all bioinformatic analysis? If no, the use of the same samples could strengthen their results and data validity.
1- In abstract with following sentence :"Although the mRNA expression level of ANXA9 is associated with the poor prognosis of colorectal cancer, the function of ANXA9 in GC remains largely unknown", it was better to indicate that lower mRNA expression of ANXA9 is associated with the poor prognosis of colorectal cancer.
2- Line 302-303: "Kaplan-Meier survival analysis showed that the OS time were prolonged in gastric cancer patients with a high expression of BUB", it seems that BUB should be BUB1.
3- Lines 330-334 could be summarised and merged.
4- Did author observe correlation between ANXA9 expression and OS in other cancers with the TCGA data analysis. It could be performed using UALCAN.
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