PONE-D-20-10368

Signatures of somatic mutations and gene expression from p16INK4A positive head and neck squamous cell carcinomas (HNSCC)

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PLOS ONE

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Reviewers' comments:

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Comments to the Author

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

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Reviewer #1: Yes

Reviewer #2: Partly

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

Reviewer #1: Yes

Reviewer #2: No

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Reviewer #1: Yes

Reviewer #2: No

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Reviewer #1: Yes

Reviewer #2: Yes

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5. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: This work presents the signatures of somatic mutation and gene expression from p16ink4a positive HNSCC. Thank you for your valuable research on genetic variants and gene expression signatures of significant HPV infection in head and neck cancer. However, while reviewing this manuscript, it was difficult to check all the details because all the resolutions of the figures were low. Attaching high resolution figures will be helpful in the next review.

In this study, p16 of head and neck cancer FFPE was classified by IHC staining. In case of ‘p16 unknown’, didn't you perform IHC? Or did you stain IHC and the staining result wasn't clear? I recommend you need to define p16? group. I am still not sure why you compared p16 separately in this study. Therefore, the reason for comparing the p16 unknown group with the p16 positive group in this study should be clearly described in the result section.

In addition, in the various experiments conducted in this study, some were targeted only to OPSCC, and some were confirmed in HNSCC, including cancers of a few other origins. Personally, I think it might have been clearer for this study to focus only on OPSCC. Or, in some experiments, please add the reason why only the OPSCC group was tested separately.

Reviewer #2: comments:

1. figures are fuzzy, hardly to read

2. Please explain what you found in details in figure 1. you could not see the results is summarized in figure..'

3.the manuscript is disorganized and need to be re-written. ie. figure legends were inserted in the Results part. Please be careful. Legends for fig 4 and 5 were separated. Hardly to read.

4. Figure 2 A. What is the percentage means? what is the figure 2B telling us?

5. Figure 4A, is confusing, looks mismatch what depicted in the manuscript. What is' p16?' in the figure? is it unknown HPV? confused

6. Are those aberrant genetic features correlating with any patient survival?

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Reviewer #1: No

Reviewer #2: No

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Signatures of somatic mutations and gene expression from p16INK4A positive head and neck squamous cell carcinomas (HNSCC)

PONE-D-20-10368R1

Dear Dr. Saba,

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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Kind regards,

Hyunseok Kang, MD, MPH

Academic Editor

PLOS ONE

Additional Editor Comments (optional):

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation.

Reviewer #1: All comments have been addressed

Reviewer #2: All comments have been addressed

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

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Yes

Reviewer #2: Partly

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

Reviewer #1: I Don't Know

Reviewer #2: I Don't Know

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

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

Reviewer #2: Yes

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

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: Yes

Reviewer #2: Yes

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6. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: All comments have been addressed. On discussion section you mentioned that 'Using RNA-seq data in parallel with targeted sequencing of a panel of 151 genes, we demonstrate that gene expression data from FFPE samples can identify gene signatures characteristic of p16 + versus p16- OPSCC which is a subsite of HNSCC.' but you used some of p16 non OPSCC samples (GHN-25). This paper analyzes data not only from OSCC but also from all HNSCC, and since data is mixed(p16+ all OSCC, p16- some OSCC and non-OSCC, and p16? all non-OSCC), accuracy of expression is considered to be important. Overall, if you care about this part, I think the rest of all is a well designed study.

Reviewer #2: The authors basically answered the question, although limitation for this research exist, it is qualified to be published.

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7. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

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Reviewer #1: No

Reviewer #2: No