PONE-D-22-20618Gulf war illness inflammation reduction trial: a randomized controlled trial of low-dose prednisone chronotherapy, effects on health-related quality of lifePLOS ONE

Dear Dr. Bach,

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.

 Your manuscript has been assessed by three peer-reviewers and their reports are appended below.  The reviewers comment that aspects of the study require additional detail or clarification, including the methodology and the results section. One of the reviewers comments that the limitations section needs additional considerations. Furthermore, the reviewers raise concerns that the statistical method has not been reported adequately, and that essential details are missing from the manuscript.  Could you please carefully revise the manuscript to address all comments raised?

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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: Yes

Reviewer #3: Yes

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

Reviewer #1: Yes

Reviewer #2: Yes

Reviewer #3: No

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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: Yes

Reviewer #2: Yes

Reviewer #3: 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 study using a novel intervention of prednisone to treat neuroinflammation in symptomatic GW veterans provides hope that these veterans will finally get relief from their symptoms. However, I think that a few things need to be addressed prior to publishing and prior to going on to Phase 3 trial.

How do the researchers explain the placebo effect and is it potentially related to dosage of the drug or something else. Can this be explained? Is it possible to discuss why the PCS went back to baseline during the washout period? Should there be carry over and if not does this mean that chronic use of steroids (which can be harmful) would need to be used to improve veterans' health.

Although group data are meaningful, is it possible to determine which of the KS symptom clusters are most changed by this medication and if individual changes can be more meaningful descriptors.

Reviewer #2: My general impression of this article is that it is well-written, and addresses a topic of great importance for the field of Gulf War Illness (GWI) research. Gulf War veterans have waited over 30 years for treatments for their chronic and debilitating illness and this manuscript addresses an important potential treatment avenue for the illness. Suggested edits below should be considered minor revisions.

General Comments:

This manuscript addresses an important topic of identifying treatments for GWI, however it has several limitations and caveats that should be more fully stated. First, there are questions about rebound effects of steroid medications on inflammation that have not been addressed in the manuscript. If this low dose MR-prednisone does not cause this issue, it would be important to state this. In addition, both the prednisone and the placebo group showed an improvement over time on the VR36 primary outcome measure. This should be more clearly stated in the discussion and limitations.

1. Introduction. Please add a reference after the first sentence.

2. Introduction. On lines 45-46, it describes a biomarker study of a group ‘who met a GWI definition’ and one who did not. Please state what GWI definition was used in this prior study.

3. Introduction. In addition to the authors work, there are many other published articles in the GWI literature identifying inflammatory markers in blood and brain imaging of GW veterans that could be cited to strengthen this argument for using an anti-inflammatory treatment for the disorder.

4. Introduction. The introduction is quite brief while the discussion has much more background information than actual discussion of the treatment results and integration of those results within the GWI research field. It would be helpful to the reader to add some of the background information about why the low dose time-release formulation of the prednisone medication was chosen in the introduction.

5. Methods. Please state the composition of the placebo.

6. Methods. In the statistical methods section, please describe what analyses were used to compare demographics of the two groups.

7. Results. Please report how many participants reported comorbid PTSD, if known.

8. Discussion. The first two paragraphs of the discussion provide important information about potential pathophysiology but seem more appropriate for the introduction than the discussion. This manuscript reports results of a treatment trial not a translational study. I would suggest rewriting this first paragraph to state the results of this treatment trial and then the following paragraphs should describe how these results compare with other relevant work in the field of GWI treatment research as well as the RA study.

9. Discussion. There is much mentioning of the blood markers related to GWI but none are reported in this manuscript. It would make for a much stronger argument for a phase III trial if blood marker changes were also reported for this treatment trial. Please consider adding the results of the pre and post blood markers from this trial to the manuscript.

10. Discussion. On page 21, please add references for the statement that IL-6 is a blood biomarker for GWI. There are both publications confirming and those to the contrary for this marker.

11. Discussion. On page 21, lines 271-281 regarding glucocorticoids and the effects of time-released steroids on them is very important and could be moved to the introduction section of the manuscript rather than in the middle of the discussion.

12. Discussion. This discussion of glucocorticoids also brings up the question again of how many participants also had comorbid PTSD as this strongly affects glucocorticoid functioning and could result in different effects in those with and without comorbid PTSD. If known, controlling or stratifying for this could potentially account for the MCS differences reported.

13. Discussion. On page 21, line 296-298, it reads as though this treatment trial compared RA individuals with GWI subjects on HRQOL outcomes. Please reword for clarity. It should also be made clear that GWI and RA are two different disorders.

14. Discussion. The statement that ‘The two-state model of GWI, in which low PCS subjects respond to prednisone and high PCS subjects are unresponsive, is consistent with the hypothesis that GWI is a relapsing-remitting inflammatory illness’ does not seem logical. Some veterans may just have more severe illness and they are the ones that benefit from this therapy rather than the results indicating a relapsing-remitting inflammatory illness per se. I would suggest removing this line of reasoning or clarifying it.

15. Strengths. Please add ‘of’ after ‘component’ and before ‘GWI.’

16. Strengths. The sentence that starts, ‘Also, the translational nature of this work is noteworthy and at every stage had strict adherence to scientific method’ reads awkwardly and is redundant with the prior sentence. Please consider removing it.

17. Limitations: The fact that there was a significant change in the placebo as well as the treatment group should be mentioned here as well as it slightly weakens the study results.

18. Figures. The figures are difficult to read and look blurry. Please upload higher resolution figures.

Reviewer #3: A phase 2 randomized-controlled clinical trial was conducted which aimed to measure the effects of prednisone on health related quality of life (HRQOL) of veterans with Gulf War illness. Prednisone treatment showed a statistically significant improvement in physical HRQOL scores from baseline to follow up.

Major revision:

Provide a thorough statistical analysis section. List and describe the use of all statistical testing methods.

a. Line 201: State the statistical testing methods used to conclude, “There were no statistically

significant differences between the two arms of the study.”

b. Line 210: Include details of the linear correlation and the statistical method used to estimate it.

c. Provide details of the linear regression analysis.

Minor revisions:

1- Abstract: List the statistical method and provide the p-value to support the following statement. Also clarify the time points in which the HRQOL was compared. “Treatment with prednisone produced a statistically significant improvement in the physical HRQOL.”

2- Lines 162-166: Provide more complete details of the pre-study statistical power calculation. The power calculation should also include the statistical testing method. If a paired t-test was used, the standard deviation should also be stated.

3- Table 1: Provide measures of dispersion of continuous variables. If continuous variables are normally distributed, it is standard practice to summarize them with means and standard deviations. If the variables are non-normally distributed, use median, first and third quartiles.

4- All acronyms and abbreviations must be spelled out at first mention.

5- When presenting means in the text, also provide the corresponding standard deviations.

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Reviewer #1: No

Reviewer #2: No

Reviewer #3: No

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Gulf war illness inflammation reduction trial: a phase 2 randomized controlled trial of low-dose prednisone chronotherapy, effects on health-related quality of life

PONE-D-22-20618R1

Dear Dr. Bach,

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,

Kimberly Sullivan

Guest Editor

PLOS ONE

Additional Editor Comments (optional):

The authors have now addressed the concerns of the reviewers from the first submission. Although reviewer 3 has raised an issue regarding the statistical analyses, this issue was not raised on the first review and given that the first and second reviewers did not raise this issue, it appears that this may not be a pressing enough issue to require a further delay and re-review of this submission before publication.

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.

The authors have now addressed the concerns raised in the previous round of reviews and I feel that this manuscript is now acceptable for publication.

Reviewer #1: All comments have been addressed

Reviewer #3: (No Response)

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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: Partly

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

Reviewer #1: Yes

Reviewer #3: No

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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

**********

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: (No Response)

Reviewer #3: Major revision:

For the analysis of the primary aim (including subset analyses), comparing changes in PCS and MCS between the groups, tests of the interaction of group by time are statistically superior to repeatedly applying a paired t-test. A comprehensive re-analysis of all the paired t-test results is recommended.

Minor revision:

Line 199: State the estimate of variability and the statistical testing method which achieves 80% power.

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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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