PONE-D-22-28204Calcineurin-Inhibitor Free Immunosuppression After Lung Transplantation – a Single Center Case-control Study in 51 Patients converted to Mammalian Target of Rapamycin (mTOR) InhibitorsPLOS ONE

Dear Dr. Gottlieb,

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.

Please submit your revised manuscript by Mar 17 2023 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.

Please include the following items when submitting your revised manuscript:

• 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 you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter.

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,

H. Hakan Aydin, MD, FAACC

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. Please provide additional details regarding participant consent. In the ethics statement in the Methods and online submission information, please ensure that you have specified what type you obtained (for instance, written or verbal, and if verbal, how it was documented and witnessed). If your study included minors, state whether you obtained consent from parents or guardians. If the need for consent was waived by the ethics committee, please include this information

3. We note that you have indicated that data from this study are available upon request. PLOS only allows data to be available upon request if there are legal or ethical restrictions on sharing data publicly. For more information on unacceptable data access restrictions, please see http://journals.plos.org/plosone/s/data-availability#loc-unacceptable-data-access-restrictions. 

In your revised cover letter, please address the following prompts:

a) If there are ethical or legal restrictions on sharing a de-identified data set, please explain them in detail (e.g., data contain potentially sensitive information, data are owned by a third-party organization, etc.) and who has imposed them (e.g., an ethics committee). Please also provide contact information for a data access committee, ethics committee, or other institutional body to which data requests may be sent.

b) If there are no restrictions, please upload the minimal anonymized data set necessary to replicate your study findings as either Supporting Information files or to a stable, public repository and provide us with the relevant URLs, DOIs, or accession numbers. For a list of acceptable repositories, please see http://journals.plos.org/plosone/s/data-availability#loc-recommended-repositories.

We will update your Data Availability statement on your behalf to reflect the information you provide

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

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: Partly

Reviewer #2: Yes

**********

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

Reviewer #1: Yes

Reviewer #2: I Don't Know

**********

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

**********

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

**********

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: There are several problems with this manuscript:

- This is a retrospective analysis rather than prospective study and the mTOR inhibitor was started at varying intervals for the treatment group

- The group of patients with malignancies is very small (51, compared to 2049)

- Multiple types of cancer are being compared together

- The CNI free immunosuppression was initiated after the diagnosis of malignancy was made, thus not allowing for an analysis of the "preventive" potential of mTOR inhibitors used as CNI free immunosuppression

mTOR inhibitors have been used in Lung transplant programs for many years, although this reviewer acknowledges that they are seldom used in an elective or protocolized fashion. Effectiveness of immunosuppression is well established, so there is nothing novel in reporting that they are well tolerated. Unfortunately, this study does not help to answer the malignancy question as the immunosuppression was changed "after the fact". A better approach would be to develop a protocolized conversion timeline, with regular sequential follow up.

Reviewer #2: I was delighted to have a chance to peer review the submission to PLOS ONE entitled “Calcineurin-Inhibition Free Immunosuppression After Lung Transplantation – a Single Center Case-Control Study in 51 Patients converted to Mammalian Target of Rapamycin (mTOR)”, submitted by the Hannover group. In it, you report on a retrospective analysis of 51 patients over almost 30 years at a single center where patients had their CNI stopped and an alterative regimen was used. Your aim was to “investigate indications, safety, and outcomes of CNI-free immunosuppression based on mTOR inhibitors in the cohort” (last sentence of the introduction). About half of them were patients who had been changed because of active malignancies. Some later had CNI’s re-introduced.

You describe better than expected outcomes (in my estimation), with no acute rejections reported (but limited numbers having had biopsies), high mortality for the oncology patients (1-year survival of 36%), low mortality for the non-oncology patients (1-year survival 100%), improvement in neurologic side effects for those who had been changed for that reason, and a modest improvement in GFR for the cohort. The median duration of CNI-free immunosuppression was 338 days, with the period beginning a median of 6.1 years after transplantation.

1. Why were there not more biopsies done after the switch?

2. Other than “CLAD or not”, were there any assessments of graft health after the switch? What happened to the spirometry?

3.

You state in the results section that only 47% of the cohort of 51 patients had been biopsied at any time before the CNI free conversion. This is surprising to me and may need clarification. Are not all of your center’s patients getting transbronchial biopsies in their first year after transplant? If not, you may want to mention somewhere that this is your routine.

You state that 10 patients tested positive for donor specific antibodies before conversion. Please mention how you handled this information in general (did it influence decisions about converting to CNI-free?). Were patients routinely (or occasionally ad hoc) testing for DSAs after the conversion?

Your figures should each have a text box for the document with a description of what is in the figure.

It would be interesting and visually pleasing to see survival curves (ie Kaplan-Meier)

Under “2.2 Safety”, you describe incidence of hospitalization and infections in the group that was CNI free. How did this compare to patients who were on more standard therapy? You might consider a broader comparison looking at all patients (not just oncology patients) and outcomes with and without the conversion to CNI-free.

Minor:

1. In the first paragraph of the introduction, the word “haltering” was used. This is an irregular term and I might suggest using a different word such as “slowing”.

2. In the last sentence of the first paragraph of the introduction, change “CNI-free immunosuppression after lung transplantation is limited” to “CNI-free immunosuppression after lung transplantation are limited”.

3. Throughout the document, mTOR is referred to as “mammalian target of Rapamycin”. A more updated and common terminology is now “mechanistic target of Rapamycin”. Consider a change if you are inclined to agree.

4. The word “intermitted” just before section “2.1 outcomes” is an irregular word, please change.

5. In the “2.1 Outcomes” section, consider changing the description of “corona virus disease 2019” to “SARS CoV-2 infection”.

6. In Discussion, “Ina conference abstract Eiting” should be, “In a conference abstract Eiting”

7. In Discussion, “In our cohort, Minor improvement” should be, “In our cohort, minor improvement”

**********

6. 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 #2: No

**********

[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.

PONE-D-22-28204R1Calcineurin-Inhibitor Free Immunosuppression After Lung Transplantation – a Single Center Case-control Study in 51 Patients converted to Mechanistic Target of Rapamycin (mTOR) InhibitorsPLOS ONE

Dear Dr. Gottlieb,

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.

Please submit your revised manuscript by Apr 29 2023 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.

Please include the following items when submitting your revised manuscript:

• 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 you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter.

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,

H. Hakan Aydin, MD, FAACC, EuSpLM

Academic Editor

PLOS ONE

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

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 #2: All comments have been addressed

Reviewer #3: (No Response)

Reviewer #4: (No Response)

**********

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: Partly

Reviewer #4: Partly

**********

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

Reviewer #2: I Don't Know

Reviewer #3: Yes

Reviewer #4: No

**********

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: No

**********

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: No

**********

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: Thank you for your work in amending your submission. It is improved with your address of reviewers' comments.

Reviewer #3: I reveiwed a revised version of the manuscript entitled "Calcineurin-Inhibitor Free Immunosuppression After Lung Transplantation – a Single Center Case-control Study in 51 Patients converted to Mechanistic Target of Rapamycin (mTOR) Inhibitors" by Gttlieb et al.

First of all I have to state that I did NOT review the previous version.

Due to the nature of the study (retrospective single-center study) it is difficult to draw any conclusion.

For those patients who were converted from CNI-based IS to mTOR inhibitor-based IS the trough levels of sirolimus and everolimus are not mentioned.

On average after conversion from CNI- to mTOR-I-based IS how many times the patients were seen in the outpatient clinic?

mTOR-Is-associated side effects are poorly reported: when it occurred did it result in reverse conversion to CNI-based IS?

The paper is more suited for a pulmonology journal. Otherwise it could be considered as a Letter to the Editor.

Reviewer #4: Gottlieb and coll described their experience of CNI-free IS in lung transplant recipients.

In my opinion, the manuscript should be reoriented toward the main result of the study, ie the safety regarding regarding rejection, in lung transplant recipients with an advanced cancer.

In a methodological point of vue, I think the comparison with patients under CNI that presented different cancer types, and different cancer stages prevents analysis of patient survival grouped by IS

Other minor remarks:

* introduction: the statement that CNI free IS is a reality in the kidney and heart or pancreas transplant setting should be tempered. In fact all of these CNIfree protocols were stopped and antimetabolite are switched to everolimus in association with CNI.

A larger presentation of the burden of cancer after lung transplantation, and specificities of cancer in the lung transplantation could improve the intro

* Patients and Method

The authors stated that they used only nonparametric tests for quantitative comparisons, so with different different distribution in a small cohort. This suggest that cox-regression model to assess predictive factors is not really useful in this case-series. I suggest to remove this part, or at least to develop the statistical section to explain the number of variables included in the model, and the number of events per variables

* results: treatments used for cancer in association with mTOR should be developed to better understand the risk for rejection in this cohort

**********

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: No

Reviewer #3: No

Reviewer #4: No

**********

[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.

Calcineurin-Inhibitor Free Immunosuppression After Lung Transplantation – a Single Center Case-control Study in 51 Patients converted to Mechanistic Target of Rapamycin (mTOR) Inhibitors

PONE-D-22-28204R2

Dear Dr. Gottlieb,

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.

An invoice for payment will follow shortly after the formal acceptance. To ensure an efficient process, please log into Editorial Manager at http://www.editorialmanager.com/pone/, click the 'Update My Information' link at the top of the page, and double check that your user information is up-to-date. If you have any billing related questions, please contact our Author Billing department directly at authorbilling@plos.org.

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,

H. Hakan Aydin, MD, FAACC

Academic Editor

PLOS ONE