PONE-D-23-30500Identification of potential anti-tumor targets and mechanisms of HuaChanSu injection using network pharmacology and cytological experiments in Breast cancerPLOS ONE

Dear Dr. Yang,

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PLOS ONE

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This work is supported by the Youth Program Research Project of the Affiliated Traditional Chinese Medicine Hospital of Guangzhou Medical University (No. 20210131), Guangzhou Chinese Medicine and Integrative Medicine Science and Technology Project (No. 20232A011005), and Scientific Research Project of Guangdong Provincial Administration of Traditional Chinese Medicine (No. 20221324 and No. 20210078). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

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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?

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

Reviewer #3: Partly

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

Reviewer #1: Yes

Reviewer #2: Yes

Reviewer #3: Yes

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3. 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

Reviewer #3: Yes

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4. 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: No

Reviewer #2: Yes

Reviewer #3: No

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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: 

The study gave some preliminary insights about the application and underlying mechanism of Traditional Chinese Medicine HuaChanSu (HCS) injection in controlling human Breast cancer.

My major concern about this manuscript would be the writing as it is too colloquial and few errors in grammar. However, the authors have addressed this concern.

Minor comments

Please explain why MCF-7 cells are used for migration studies as they are not metastatic.

Please explain why MCF-7 cells are not used in Western Blotting experiment

Please emphasize the importance of p13-AKT and mTOR pathways in developing the cancer and also the active component HuaChanSu that aids in regulating the pathway.

Also if possible mention how HuaCHanSu can be further evaluated in clinical trails.

In Abstract

Line 19 Rewrite the sentence

"Subsequently" is repeated many times

In Introduction

Line 47-48 both the sentences are conveying same meaning.Please Merge sentences and rewrite

Line 67 please correct "no studies have not"

In Line 75 replace "we used integrated several techniques" with "we have used several integrated techniques"

Line 240 Please replace BRCA cells with MCF-7 and MDA-MB-231 cell lines

Line 274 Authors have mentioned "reviewed previously published literature to screen for active components......". Please give the reference for this.

Line 294 replace Finally with and

Line 375 replace influences with appropriate word like affects or alters

Line 385 replace the word cyberpharmacology with Network Pharmacology

Reviewer #2: 

1- My observations are overlapped with the reviewer 1 an 2.

2- Also still fonts and numbers need some clarification specially in the figures and photos.

3- There are few writing mistakes and need correction.

Reviewer #3: 

In this paper, authors used network pharmacology and cytological experiments to explore the potential anti-tumor targets and the mechanisms of HuaChanSu injection. However, there are still some questions that should be better explained in the revised manuscript. I have the following concerns:

Major comments:

1. In the introduction section, the clinical efficacy of HuaChanSu or its patent medicine in the treatment of cancer should be clearly described.

2. In the screening for active components, authors only consider literatures. It is suggested that authors also consider pharmacokinetics such as oral bioavailability.

3. In the GO and KEGG enrichment analysis, authors need to describe the significance criterion of enrichment analysis results.

4. In the molecular docking verification section, authors should describe how they used the same docking method to calculate the energy score of the positive drug binding to key targets.

5. In the result section, authors wrote: HCS downregulates the PI3K-Akt/mTOR signaling pathway to mediate the anti-tumor effect in BRCA. Recently, the network pharmacology evaluation method guidance is issued. It is suggested that authors compare their work with this guidance and provide more solid experimental evidence to support their analysis in the revised manuscript. Please see: Network pharmacology evaluation method guidance‐Draft. World J Tradit Chin Med, 2021; 7(1):146-154.

6. In the discussion section, the advantages and limitations of integrating molecular docking and cytological experiments into the network pharmacology approach used in this study should be discussed.

7. In the introduction and discussion section, it is suggested that authors provide more background and applications on network pharmacology integrating docking and cell experimental validation such as PMID: 30618762 and 30762338.

Minor comments:

1. What is the screening criteria for targets in the 3B?

2. What is the meaning of different node and edge color in the Figure 3C?

3. It is suggested that authors mark enrichment significance in the Figure 3

4. There still exist some typos, spelling and grammar mistakes. The manuscript should be thoroughly checked, and the language also needs to be further improved.

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Reviewer #1: No

Reviewer #2: No

Reviewer #3: No

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Identification of potential anti-tumor targets and mechanisms of HuaChanSu injection using network pharmacology and cytological experiments in Breast cancer

PONE-D-23-30500R1

Dear Dr. Yang,

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,

Mohammad Sadegh Taghizadeh, Ph.D.

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 #3: 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 #3: Yes

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

Reviewer #1: Yes

Reviewer #3: Yes

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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 #3: 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 #3: 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: The manuscript is technically sound and well written. The purpose of the study and experimentation is clear. The results presented are statistically significant and supports the conclusion in the study. The public data repository mentioned in the manuscript is accessible. However, I request Authors to carefully revise the paper as few typographical errors are noticed.

The authors have well addressed all the comments made by reviewers and also considerably improved the quality of the paper for publication. Therefore, the manuscript can be considered for publication.

Reviewer #3: (No Response)

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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.

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 #3: No

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