PONE-D-24-21041Variations in De Novo Donor-Specific Antibody Development Among HLA-DQ Mismatches in Kidney Transplant RecipientsPLOS 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: Partly

Reviewer #2: Partly

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: 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: 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 is a single-centre, retrospective study conducted on a cohort of kidney transplant and SPK first-graft recipients over a 14yrs period aimed to identify the specific HLA-DQB1 mismatches associated with de novo DSA occurrence and their impact on graft survival. Authors found that patients with HLA-DQ7 and HLA-DQ9 MMs had the highest risk of developing de novo DSAs, and that DSAs against DQB1 MM were associated with detrimental graft outcomes.

It is interesting to see that authors have interestingly managed to cast some light on the importance of matching against DQB1 antigens, raising concern about the higher risk of developing DSAs against DQ7 and DQ9. The association between DSAs against DQB1 antigens and rejection / allograft loss is not a new finding and has extensively been described in the literature (of note - the works from Aleksandar Senev and co-workers should be cited as remarkable studies highlighting the role of DSAs against DQs and poor transplant outcomes). Therefore, the authors should revisit the survival analysis and address the followings:

- Is the survival analysis the result of a crude analysis? Although well known from the literature, it is difficult to state that DSAs against DQB1 MMs are associated with detrimental outcomes in the in-study cohort if the survival analysis doesn't consist in a cox-regression taking into account

1) HLA-A / HLA-B MM (and possibly HLA-Cw where available)

2) co-existence of class I and class II DSAs

3) prevalence of cellular rejection and BK nephropathy

- was the incidence of AMR under-estimated? according to the methods, a biopsy was done only if DSA MFI > 5,000 or change in function - early AMR stage may have been missed

- did the authors look into the selective pathogenicity of DSAs against specific DQs?

Minor revision: page 7 line 112: the term 'repeated DQ MM' is inappropriately used in my opinion, as it should instead refer to MMs which recurs in recipients of >1 graft.

Reviewer #2: In their manuscript "Variations in De Novo Donor-Specific Antibody Development Among HLA-DQ Mismatches in Kidney Transplant Recipients" Skulratanasak and colleagues retrospectively investigated a cohort of 491 kidney transplant recipients with HLA-DQB1 mismatches. They analyzed individually DQ loci carried by the recipient, and the likelihood of developing donor specific antibodies (DSAs). They found that patients who developed DSAs to HLA-DQ had increased risk of rejection and graft loss, in line with previously published results. They found that some recipient antigen mismatches such as DQ7 and DQ9 are at highest risk of inducing DSAs. The methodology used is mostly appropriate. The manuscript is sometimes difficult to follow. While there is some originality to the main finding of the manuscript, its clinical relevance needs to be further clarified. Specifically, it remains to be demonstrated whether the stratification in risk of developing DSAs (figure 3) also correlates with stratified rejection-free survivals and graft outcomes. I would encourage the authors to consider the following additional issues should they consider resubmitting the manuscript to a different journal:

- Data on DR mismatches should be provided. In its current form, it is impossible to determine whether clinical outcomes were influenced by DR mismatches;

- Prior studies have demonstrated that the number of DQ mismatches influences the development of DSAs. Here, the authors have focused on specific allele mismatches and development of DSAs. What was the distribution of single vs double mismatches in for each allele? In other words, were there allele mismatches that were more likely to be found in isolation and allele mismatches that were more likely to be found in a double mismatch? This could also influence differences in development of DSAs.

- How was AMR defined?

- Was there a specific protocol for induction immunosuppression, or was this left to the discretion of the treating provider? Similarly, can you provide additional information on how maintenance immunosuppression was decided?

- The tables, their abbreviations/notes and the figure legends are embedded in the main text and this makes following the text more difficult. I would recommend placing the tables and figure legends at the end of the manuscript.

- In the methods you mention that "Simultaneous kidney-pancreas patients should be removed from the analysis, given the different potential for Ab development and rejection" - if SPK patients were indeed included, I would recommend removing them from the analysis

- In the introduction, it is stated that "compatibility of HLA between donors and recipients is crucial in achieving successful KT" - I would revise this sentence, as many recipients achieve excellent outcomes despite receiving fully mismatched kidneys

- Line 199: "The prevalence of dnDSA development per each HLA-DQB1 MM was 11.25% (64/569 MM)." - the number of mismatches should be 566, consistently with the rest of the manuscript

- Table 1: "Immunosuppression at time of dnDSA detection" by definition cannot be applied to the "no DSA" column. So what was the time point used to assess the immunosuppression reported?

Reviewer #3: The manuscript describes a single-center, retrospective, cohort study in 491 kidney transplant recipients where they sought to identify the HLA-DQ mismatches with the highest risk of developing de novo DSA. Patients included in this study have at least one HLA-DQ mismatch, have not received a previous kidney transplant, and did not show DSA >1000 MFI pre-transplant. The authors found that 12% developed de novo DSA to HLA-DQ mismatches and multivariate analysis showed that those with DQ7 and DQ9 mismatches carried an increased risk for developing de novo DSA, despite not being the most found DQ mismatches. Finally, they show that patients who developed de novo DQ DSA, compared to patients without de novo DQ DSA, had an increased incidence of late active or chronic AMR and lower 10-year death-censored graft survival. This is a well-written paper on a currently popular topic. This manuscript adds to the existing knowledge about HLA-DQ mismatches and de novo DSA formation and its clinical relevance in transplantation.

Comments:

1. Materials and Methods section, page 6, starting in line 96: Please describe more extensively how the assignment of HLA-DQ DSA was done

1.a. Was DSA assigned at the antigen or allele level (manuscripts state that HLA typing was performed by intermediate or high resolution, depending on whether it was before vs after 2014)

1.b. When you assigned DSA by antigen, how did you handle cases where multiple beads with the same antigen, but different alleles were above your cutoff?

1.c. DQA DSA independent from DQB DSA or DQA-DQB DSA as heterodimer

2. Results section,

2.a. Page 8, line 142: Please briefly describe how the virtual XM was done.

2.b. Page 12

2.b.i. Line 197: “the most common specific anti-DQ dnDSA was against HLA-DQ7 (24%)”, however, figure 2A shows 23.44% which rounded to the nearest whole number is 23%. Please clarify.

2.b.ii. Line 198: 18% for DQ9 but figure 2A shows 17.19% which rounded to the nearest whole number is 17%. Please clarify.

2.b.iii. Line 200: The prevalence of DQ dnDSA per DQ antigen says DQ9 is 16.92% but figure 2B shows 16.67%, DQ2 says 12.85% but the figure shows 14.08%. Please clarify.

2.c. Page 13

2.c.i. Line 215-220 is confusing. Why are DQ7 mismatches compared to DQ5 and DQ6, and DQ9 compared to DQ6?

2.c.ii. Line 226-228, like 4.a., why are DQ7 and DQ9 compared to DQ6?

2.c.iii. Line 234 and figure 4A shows that patients with dnDSA to DQ have lower rejection-free survival compared to patients with no dnDSA to DQ.

2.c.iii.1. Were there any differences between different DQ DSA (i.e., DQ7 or DQ9 DSA worse than other DQ DSA?

2.c.iii.2. How about DQ DSA versus non-DQ DSA? In other words, is it clinically relevant that certain DQ mismatches seem to be more “immunogenic”?

3. Discussion section

3.a. Page 16, line 295: When the authors refer to ratio of dnDSA per MM, shouldn’t this use the rank and percentages on figure 2B rather than 2A?

3.b. Page 16, line 303, the study cited is from Canada and not from the US

3.c. Page 7, line 313, the study is from Canada and not from the US

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Reviewer #1: No

Reviewer #2: No

Reviewer #3: Yes:  Hugo Kaneku

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Variations in De Novo Donor-Specific Antibody Development Among HLA-DQ Mismatches in Kidney Transplant Recipients

PONE-D-24-21041R1

Dear Dr. Skulratanasak

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.

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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: (No Response)

Reviewer #3: All comments have been addressed

Reviewer #4: All comments have been addressed

Reviewer #5: 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

Reviewer #5: Yes

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

Reviewer #2: Yes

Reviewer #3: Yes

Reviewer #4: N/A

Reviewer #5: 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 #2: Yes

Reviewer #3: Yes

Reviewer #4: Yes

Reviewer #5: 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

Reviewer #5: 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 #2: (No Response)

Reviewer #3: (No Response)

Reviewer #4: (No Response)

Reviewer #5: The article Variations in De Novo Donor-Specific Antibody Development Among HLA-DQ

Mismatches in Kidney Transplant Recipients is very well tailored after extensive revision. Methods and results are well organized with coherent explanation and conclusion. This study is a significant addition to the literature on KT and the article can now be published.

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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

Reviewer #5: No

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