PONE-D-19-27424

Lack of an association between clinical INSTI-related body weight gain and direct interference with MC4 receptor (MC4R), a key central regulator of body weight

PLOS ONE

Dear Dr. McMahon,

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ADDITIONAL ACADEMIC EDITOR COMMENTS: There were 4 expert reviewers that handled your manuscript. All reviewers found this study interesting and it contains merit. However, there were several suggestions that would benefit the authors to include in the paper. There are suggestions to correct the figure labels/legends and include graphs of the data; there is information that needs to be added to the abstract and introduction; and in the discussion, future studies using in vivo models, which includes examining how other signaling molecules involved in food intake and body weight homeostasis might interact with this system, should included along with how these findings could be translated in the clinic.

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PLOS ONE

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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

Reviewer #4: Yes

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

Reviewer #1: N/A

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 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

Reviewer #4: 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

Reviewer #4: 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: In this study, several INSTIs were interrogated for their capacity for antagonism or agonism of MC4R in an in vitro cell-based assays including at concentrations far exceeding plasma

concentrations reached at the recommended dosages. The results indicate that while

INSTIs do exhibit the capacity to antagonize MC4R, this occurs at concentrations well

above predicted clinical exposure and is thus an implausible explanation for INSTIassociated

weight gain.

This is an interesting topic.

However, the discussion is very short and simplified and only two figures are shown in the main text.

Furthermore, I think more detail with regards to other studies investigating the effects on drugs/chemicals interacting the MC4R is interesting to include and discuss within the context of obesity.

E.g. the GLP-1 receptor analogue treatment and the MC4R both in cells, mice (eg. PubMed Central PMCID: PMC5550563.) and humans (eg PubMed PMID: 29861388) and many more...

From this it seems that weight loss can be induced even though you have no MC4R with GLP-1 RA. Thus obesity and MC4R seems complicated and treatment of obesity is not only dependent on the MC4R but also on other brain areas/receptors etc.

Furthermore actual and potential MC4R agonist treatment could be discussed.

Reviewer #2: McMahon and colleagues examined the impact of several integrase strand transfer inhibitors (INSTIs) on melanocortin-4 receptor (MC4R) activity. Given the recent increase in prevalence of excess weight gain in patients with HIV, particularly after initiating treatment with INSTIs, several hypotheses have been suggested to explain this weight gain including a potential interaction of INSTIs with the MC4R, leading to its inhibition promoting hyperphagia and reduced energy expenditure. Using in vitro assays in CHO cells transfected with human MC4R, the authors showed that although every INSTI compound exhibit significant binding to the MC4R and evoked inhibition of MC4R-induced activation of cAMP, the doses required for significant inhibition were far too greater than anticipated clinical circulating levels of free INSTI. Thus, the authors concluded that potential inhibition of MC4R by INSTIs is likely not the main mechanism behind the excess weight gain observed in INSTI-treated HIV patients.

Comments:

The manuscript is very well written, concise and easy to follow.

1) As already acknowledged by the authors, MC4R located in the brain, particularly in the hypothalamus, are responsible for body weight regulation and plasma levels of free INSTI may not necessarily reflect the concentration of INSTIs seen by MC4R in the hypothalamus. Also, all experiments were performed using in vitro techniques which may also not reflect 100% of the potential interaction of INSTIs with MC4R in vivo. How difficult would it be for the authors to include a separate protocol where MC4R KO animals (rats or mice) would be treated peripherally and/or intracerebroventricularly with INSTIs while their appetite, metabolic function and body weight are followed for a couple of weeks? I am not requiring the authors to perform such experiments, but they would significantly enhance the study's impact in the field.

Reviewer #3: In this paper, authors perform in vitro assay for some antiviral drugs effect on MC4R binding to explain its potental role in a side effect of the therapy associated with weight gain. The study found no agonistic effects of compounds and the antagonistic affects at concentrations which are much higher than blood level of these drugs in patients. The authors conclude that interactions of these drugs with MC4R is an unlikely mechanisms underlying weight gain in patients.

Specific comments

1. Abstract, specify that dolutegravir is an INSTI

2. In the introduction, please explain the chemical nature of the drugs

3. Is effect on appetite of INSTI known?

4. Result, can you provide graphs illustrating binding of both test and control compounds

5. Figure 1 legend is missing

6. Table 1, Agonists and antagonists columns have been switched between control substances

Reviewer #4: This paper investigates the effect of INSTIs on weight gain through MC4R. Despite INSTIs' capacity to antagonize MC4R, the authors have reported that at theurapeutic concentrations, antagonism of MC4R might not be an explanation for weight gain in treated patients. They are aware of the limitations which were mentioned.However, they need to speculate on what needs to be done from the research point more spesifically for practical purposes.

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Reviewer #1: No

Reviewer #2: Yes: Alexandre Alves da Silva

Reviewer #3: Yes: Sergueï Fetissov

Reviewer #4: No

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Lack of an association between clinical INSTI-related body weight gain and direct interference with MC4 receptor (MC4R), a key central regulator of body weight

PONE-D-19-27424R1

Dear Dr. McMahon,

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

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With kind regards,

Frank T. Spradley

Academic Editor

PLOS ONE

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 #2: All comments have been addressed

Reviewer #3: All comments have been addressed

Reviewer #4: 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 #2: Yes

Reviewer #3: Yes

Reviewer #4: Yes

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

Reviewer #2: Yes

Reviewer #3: Yes

Reviewer #4: Yes

**********

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 #2: Yes

Reviewer #3: Yes

Reviewer #4: 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 #2: Yes

Reviewer #3: Yes

Reviewer #4: 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 #2: No additional comments except that inclusion of in vivo studies in MC4R KO model would strengthen the manuscript.

Reviewer #3: (No Response)

Reviewer #4: (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 #2: Yes: Alexandre A. da Silva

Reviewer #3: Yes: Serguei Fetissov

Reviewer #4: Yes: Oya Ercan