Dear Dr. Bailey,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

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ACADEMIC EDITOR: Please revise the reviewers' comments and modify the manuscript accordingly. In addition, I would like to highlight the need to include a sample size estimation showing how representative your sample is. Also, I suggest validating putative protein markers of oesophageal carcinoma. You can validate some markers by running a secondary analysis using external data, demonstrating their labeling by targeted methods such as western blot, or running an independent validation with additional samples.

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Please submit your revised manuscript by Dec 19 2024 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org . When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.

• A rebuttal letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'.
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We look forward to receiving your revised manuscript.

Kind regards,

Alexis G. Murillo Carrasco

Academic Editor

PLOS ONE

Journal Requirements:

When submitting your revision, we need you to address these additional requirements.

1. Please ensure that your manuscript meets PLOS ONE's style requirements, including those for file naming. The PLOS ONE style templates can be found at https://journals.plos.org/plosone/s/file?id=wjVg/PLOSOne_formatting_sample_main_body.pdf and https://journals.plos.org/plosone/s/file?id=ba62/PLOSOne_formatting_sample_title_authors_affiliations.pdf

2. We note that the grant information you provided in the ‘Funding Information’ and ‘Financial Disclosure’ sections do not match.

When you resubmit, please ensure that you provide the correct grant numbers for the awards you received for your study in the ‘Funding Information’ section.

3. Thank you for stating the following financial disclosure:

 [This study was supported by a Cancer Research UK Centres Network Accelerator Award Grant (C328/A21998). Instrumentation in the Centre for Proteomic Research is supported by the BBSRC (BM/M012387/1).]. 

Please state what role the funders took in the study.  If the funders had no role, please state: ""The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.""

If this statement is not correct you must amend it as needed.

Please include this amended Role of Funder statement in your cover letter; we will change the online submission form on your behalf.

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This information should be included in your cover letter; we will change the online submission form on your behalf.

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Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. Is the manuscript technically sound, and do the data support the conclusions?

Reviewer #1: Yes

Reviewer #2: Yes

Reviewer #3: Yes

Reviewer #4: Partly

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2. Has the statistical analysis been performed appropriately and rigorously? -->?>

Reviewer #1: I Don't Know

Reviewer #2: Yes

Reviewer #3: Yes

Reviewer #4: Yes

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3. Have the authors made all data underlying the findings in their manuscript fully available??>

The PLOS Data policy

Reviewer #1: Yes

Reviewer #2: Yes

Reviewer #3: Yes

Reviewer #4: Yes

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4. Is the manuscript presented in an intelligible fashion and written in standard English??>

Reviewer #1: Yes

Reviewer #2: No

Reviewer #3: Yes

Reviewer #4: Yes

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Reviewer #1: Your PICO isn't very clear in your abstract noe introduction, I would recommend clarifying it. I would recommend splitting your "Results" section to methods, analysis, and results instead of joining it all into one

Reviewer #2: Overall, this is an interesting observation study however the major limitations are first the sample size and second the lack of further analysis. The study would benefit from a more detailed focus in the discussion and avoid usign rather generic statements such as 'suggesting that tumourigenesis may impact cellular phenotype'.

There are also quite a number of typographical errors - a few examples below:

spelling- 'mass spectrometery proteomics'

missing word and punctuation - For enrichment of 'Reactome pathways [13] also found changes in'

capital letters- 'Bligh EG, Dyer WJ. A RAPID METHOD OF TOTAL LIPID EXTRACTION AND PURIFICATION.'

avoid discussing the results in the results section - for eg 'Anterior Gradient 2 (AGR2) , involved protein

folding and secretion was the most DEP for OAC. High AGR2 gene expression is a known

unfavourable prognostic marker in renal and liver cancer [8].'

avoid single sentence paragraphs

eg 'The principal value of the results presented here is the quantification of 3,500 proteins of which

nearly 500 were differently expressed as a resource to other OAC researchers.'

It is not clear if this refers to a previous study as the references do not correspond, or whether it was done during this study? 'Additionally, the finding that the G95R mutation decreases the stability of NOP58 suggests...'

Reviewer #3: In the manuscript entitled "Comparative analysis of protein expression between oesophageal adenocarcinoma and normal adjacent tissue," the authors present the proteomics analysis of differential protein expression data obtained from mass spectrometry from 7 cancer patients.

The differential protein expression highlights many previously known biomarkers along with a handful of OAC-specific potential candidate proteins across multiple functional classes. Although this kind of analysis will be even more powerful and informative with a larger sample size, I think this study still provides firsthand information about potential candidates for further scrutiny.

Minor queries

The PCA shows that group PC1 contributes significantly higher variation. What is the nature of this group in component analysis?

The relative heterogeneity between the 7 patents seems quite obvious even in the DEP analysis. Does this heterogeneity stem from the small sample size or the personalized nature of each patient?

Reviewer #4: The manuscript "Comparative analysis of protein expression between oesophageal adenocarcinoma and normal adjacent tissue" has been provided by the authors in a comprehensive manner. . The results complement their previous proteogenomics study to identify neoantigens oesophageal adenocarcinoma patients. In this study, differential expression of oesophageal adenocarcinoma and normal adjacent tissue has been carried out. Below are some concerns

1) How did you identify G95R mutation decreases the stability of NOP58?

2) In this study, Is NOP58 differentially expressed that it gives a defective ribosomal product (DRiP)?

3) Apart from G2/M checkpoint related protein, CDC5L others genes/proteins are also differentially expressed, Why only these genes are targeted?

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what does this mean? ). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy

Reviewer #1: No

Reviewer #2: No

Reviewer #3: Yes:  Hariharan Parameswaran

Reviewer #4: Yes:  Dicky John Davis G

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[NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files.]

While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com/ . PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email PLOS at figures@plos.org . Please note that Supporting Information files do not need this step.

Dear Dr. Bailey,

Please submit your revised manuscript by Feb 13 2025 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org . When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.

• A rebuttal letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'.
• A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'.
• An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'.

If applicable, we recommend that you deposit your laboratory protocols in protocols.io to enhance the reproducibility of your results. Protocols.io assigns your protocol its own identifier (DOI) so that it can be cited independently in the future. For instructions see: https://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols . Additionally, PLOS ONE offers an option for publishing peer-reviewed Lab Protocol articles, which describe protocols hosted on protocols.io. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols .

We look forward to receiving your revised manuscript.

Kind regards,

Alexis G. Murillo Carrasco

Academic Editor

PLOS ONE

Journal Requirements:

Please review your reference list to ensure that it is complete and correct. If you have cited papers that have been retracted, please include the rationale for doing so in the manuscript text, or remove these references and replace them with relevant current references. Any changes to the reference list should be mentioned in the rebuttal letter that accompanies your revised manuscript. If you need to cite a retracted article, indicate the article’s retracted status in the References list and also include a citation and full reference for the retraction notice.

[Note: HTML markup is below. Please do not edit.]

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

Reviewer #2: All comments have been addressed

Reviewer #3: All comments have been addressed

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2. Is the manuscript technically sound, and do the data support the conclusions??>

Reviewer #2: Yes

Reviewer #3: Yes

**********

3. Has the statistical analysis been performed appropriately and rigorously? -->?>

Reviewer #2: I Don't Know

Reviewer #3: Yes

**********

4. Have the authors made all data underlying the findings in their manuscript fully available??>

The PLOS Data policy

Reviewer #2: Yes

Reviewer #3: Yes

**********

5. Is the manuscript presented in an intelligible fashion and written in standard English??>

Reviewer #2: Yes

Reviewer #3: Yes

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Reviewer #2: There are still some minor corrections that need to be made, for example:

- Notable in NAT were high expression in NAT of

- be consistent when referring to companies and provide countries/headquarters for eg there is Waters, UK but no country etc for other companies.

- (Thermo Fisher Scientific,UK) but other times it is just Thermo Scientific

- use lowercase n for 'specifically Neutrophil degranulation'

I think there should be a bit more emphasis on studies that have previously identified the same proteins eg for AGR in Barretts https://pmc.ncbi.nlm.nih.gov/articles/PMC2575112/

Reviewer #3: The revised draft entitled, "Comparative analysis of protein expression between oesophageal adenocarcinoma and normal adjacent tissue.", addresses my queries raised in the initial submission.

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what does this mean? ). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy

Reviewer #2: No

Reviewer #3: Yes:  Hariharan Parameswaran

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[NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files.]

While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com/ . PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email PLOS at figures@plos.org

Comparative analysis of protein expression between oesophageal adenocarcinoma and normal adjacent tissue

PONE-D-24-45100R2

Dear Dr. Bailey,

We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements.

Within one week, you’ll receive an e-mail detailing the required amendments. When these have been addressed, you’ll receive a formal acceptance letter and your manuscript will be scheduled for publication.

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If your institution or institutions have a press office, please notify them about your upcoming paper to help maximize its impact. If they’ll be preparing press materials, please inform our press team as soon as possible -- no later than 48 hours after receiving the formal acceptance. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information, please contact onepress@plos.org.

Kind regards,

Alexis G. Murillo Carrasco

Academic Editor

PLOS ONE

Additional Editor Comments (optional):

Reviewers' comments: