PCOMPBIOL-D-24-02103

Anatomically aware simulation of patient-specific glioblastoma xenografts

PLOS Computational Biology

Dear Dr. Malik,

Thank you for submitting your manuscript to PLOS Computational Biology. After careful consideration, we feel that it has merit and are pleased to inform you that we are recommending a minor revision. While the manuscript does not fully meet PLOS Computational Biology's publication criteria as it currently stands, 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 within 30 days. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at ploscompbiol@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pcompbiol/ 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 editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'. This file does not need to include responses to formatting updates and technical items listed in the 'Journal Requirements' section below.

* 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, competing interests statement, or data availability statement, please make these updates within the submission form at the time of resubmission. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter.

We look forward to receiving your revised manuscript.

Kind regards,

Heber L. Rocha, Ph.D.

Guest Editor

PLOS Computational Biology

Christoph Kaleta

Section Editor

PLOS Computational Biology

Journal Requirements:

1) Please ensure that the CRediT author contributions listed for every co-author are completed accurately and in full.

At this stage, the following Authors/Authors require contributions: Adam Malik, Cecilia Krona, Soumi Kundu, Philip Gerlee, and Sven Nelander. Please ensure that the full contributions of each author are acknowledged in the "Add/Edit/Remove Authors" section of our submission form.

The list of CRediT author contributions may be found here: https://journals.plos.org/ploscompbiol/s/authorship#loc-author-contributions

2) We ask that a manuscript source file is provided at Revision. Please upload your manuscript file as a .doc, .docx, .rtf or .tex. If you are providing a .tex file, please upload it under the item type u2018LaTeX Source Fileu2019 and leave your .pdf version as the item type u2018Manuscriptu2019.

3) Please provide an Author Summary. This should appear in your manuscript between the Abstract (if applicable) and the Introduction, and should be 150-200 words long. The aim should be to make your findings accessible to a wide audience that includes both scientists and non-scientists. Sample summaries can be found on our website under Submission Guidelines:

https://journals.plos.org/ploscompbiol/s/submission-guidelines#loc-parts-of-a-submission

4) Your manuscript's sections are not in the correct order. Please amend to the following order: Abstract, Introduction, Results, Discussion, and Methods

5) Thank you for including an Ethics Statement for your study. Please include:

i) The full name(s) of the Institutional Review Board(s) or Ethics Committee(s).

6) Please upload all main figures as separate Figure files in .tif or .eps format. For more information about how to convert and format your figure files please see our guidelines:

https://journals.plos.org/ploscompbiol/s/figures

7) Please ensure that all Figure files have corresponding citations and legends within the manuscript. Currently, Figure 2 does not have in-text citations. Please include the in-text citation of the figure. Please also ensure that Figure 2 is labeled.

8) We notice that your supplementary Figures, and information are included in the manuscript file. Please remove them and upload them with the file type 'Supporting Information'. Please ensure that each Supporting Information file has a legend listed in the manuscript after the references list. Please cite and label the supplementary figure as "S1 Figure".

9) Some material included in your submission may be copyrighted. According to PLOSu2019s copyright policy, authors who use figures or other material (e.g., graphics, clipart, maps) from another author or copyright holder must demonstrate or obtain permission to publish this material under the Creative Commons Attribution 4.0 International (CC BY 4.0) License used by PLOS journals. Please closely review the details of PLOSu2019s copyright requirements here: PLOS Licenses and Copyright. If you need to request permissions from a copyright holder, you may use PLOS's Copyright Content Permission form.

Please respond directly to this email and provide any known details concerning your material's license terms and permissions required for reuse, even if you have not yet obtained copyright permissions or are unsure of your material's copyright compatibility. Once you have responded and addressed all other outstanding technical requirements, you may resubmit your manuscript within Editorial Manager.

Potential Copyright Issues:

i) Figure 1. Please confirm whether you drew the images / clip-art within the figure panels by hand. If you did not draw the images, please provide (a) a link to the source of the images or icons and their license / terms of use; or (b) written permission from the copyright holder to publish the images or icons under our CC BY 4.0 license. Alternatively, you may replace the images with open source alternatives. See these open source resources you may use to replace images / clip-art:

- https://commons.wikimedia.org

- https://openclipart.org/.

10) Please amend your detailed Financial Disclosure statement. This is published with the article. It must therefore be completed in full sentences and contain the exact wording you wish to be published.

1) State the initials, alongside each funding source, of each author to receive each grant. For example: "This work was supported by the National Institutes of Health (####### to AM; ###### to CJ) and the National Science Foundation (###### to AM)."

2) State what role the funders took in the study. If the funders had no role in your study, please state: "The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript."

Reviewers' comments:

Reviewer's Responses to Questions

Reviewer #1: General summary: In this article the authors present a method to estimate tumor growth model parameters from histological sections from patient-derived xenografts. Using four different PDX models with 2 replicates each, the authors collected tissue sections which were subsequently binarized for comparison against the output of an agent based model. The agent base model described tumor growth and invasion which was sensitive to white matter fiber tracts (DTI collected in a different animal) and tissue-cleared maps of vasculature (Also collected in different animals). The authors then applied approximate bayesian computation (ABC) requiring sampling parameter ranges to yield 5000 simulations per animal, the top 1% (50) were selected using a geometry based measure. In general, this is a very interesting paper that presents a novel contribution to Computational Biology/Mathematical Oncology demonstrating an approach for comparing agent based model results to experimental data. I do have several concerns that should be addressed before further consideration.

Comment 1: Comparison to existing literature and selection of method: In the introduction you provide an overview of existing PDE (proliferation-invasion) and ABM based models. What is the main motivation for using ABM methods over PDE methods for these type of problems? This is covered in the discussion, but some of this topic should be covered in the introduction to help support the modeling choices made in this manuscript. Some PDE-based methods to consider discussing that include vasculature-coupled growth and/or white matter coupled growth:

(1) Human application: Hawkins-Daarud A, Rockne RC, Anderson AR, Swanson KR. Modeling Tumor-Associated Edema in Gliomas during Anti-Angiogenic Therapy and Its Impact on Imageable Tumor. Front Oncol. 2013 Apr 4;3:66. doi: 10.3389/fonc.2013.00066. PMID: 23577324; PMCID: PMC3616256.

(2) Mice/rats: Hormuth, D.A., Jarrett, A.M., Feng, X. et al. Calibrating a Predictive Model of Tumor Growth and Angiogenesis with Quantitative MRI. Ann Biomed Eng 47, 1539–1551 (2019). https://doi.org/10.1007/s10439-019-02262-9

(3) Mice/rats: Rutter, E.M., Stepien, T.L., Anderies, B.J. et al. Mathematical Analysis of Glioma Growth in a Murine Model. Sci Rep 7, 2508 (2017). https://doi.org/10.1038/s41598-017-02462-0

Comment 2: Third paragraph introduction: I think there is a word or phrase missing here. “This paper seeks to align two distinct approaches to brain tumor invasion”. Is it “two distinct approaches to model/study brain tumor invasion”?

Comment 3: Missing figure label and or caption? At the top of page 6, there are two panels showing the 3D maps of white matter and the vasculature. These should be given a figure number and referenced in the text.

Comment 4: Figure 3 is missing a figure caption.

Comment 5: Section 2.2, third paragraph. On the first read it is not clear why 50 accepted parameters were chosen. I would provide more background in this section to say how many total simulations/samples were collected and what percent of those you saved. (You might add this in your discussion of ABC in section 2.1.

Comment 6: Section 2.3, Some of the comparisons between replicates in this section are vague or contradictory to the results shown in Table 1. For example,for U3230MG you state that the parameters agree well except for w_bv, but the median or mean value of p_m is nearly 2 times different between the replicates. Similar comments for U3062MG.

Comment 7: Section 2.3, I would consider defining delta here as the acceptance threshold to improve the clarity of this section.

Comment 8: For figure 4, I would consider adding the error or the value of the Jaccard score to provide reference quantifications for the best fits.

Comment 9: Section 2.5. I am not entirely convinced that this section is needed for this manuscript. Is the proposed in silico drug mimicking a clinical available option? If not, is there a reason why standard of care treatment (e.g., fractionated radiotherapy and/or temozolomide) options were not modeled on with this framework

Comment 10: Section 3.1, “data” should be changed to “Data”.

Comment 11: Section 3.2. Second paragraph. What are the units of the radius of 8? (8 microns, lattice/grid positions?). How closely does this radius of 8 correspond to the volume of injected cells. Does the volume/radius of injected cells in the initial condition influence the estimated model parameters? Have you considered using an ellipsoid to model the initial condition (assuming cells are distributed along the injection track rather than at a precise location?

Comment 12: Section 3.3, “We choose a carrying capacity of K = 3”. How much does the selection of K influence the estimated parameters? Would a higher K (K = 5 or 10) or a lower K (K = 1 or 2) skew the ratio of the probability to proliferate and probability to migrate?

Comment 13: Discussion, Is only one tissue section available from the experimental data? If so, is a 3D simulation truly needed to estimate these parameters, and if not could the number of cells in the 2D simulation be increased?

Reviewer #2: Reviewer Summary:

This manuscript presents a promising computational approach that explicitly incorporates both white matter and blood vasculature to predict glioblastoma xenograft growth. The overall structure and summary of the work are well articulated. However, the manuscript would benefit from clarification and elaboration in several key areas.

Major Comments:

1) In the Results section, there is ambiguity in the use of the terms metric, method, and algorithm within the ABC framework. The manuscript should clarify that the same ABC rejection algorithm is used throughout, with a fixed tolerance (e.g., 1% acceptance rate). It is unclear what the authors mean by “regression adjustment”—does this refer to the use of the Epanechnikov kernel, or to a change in the summary statistics used to compare geometric metrics? Please revise the following paragraph accordingly, and ensure consistent terminology throughout the manuscript:

“Given such ‘ground truth’ data, we evaluated three versions of the ABC algorithm. First, in the simplest version, we matched simulations to experiments using the Jaccard index (the fraction of tumor-containing pixels that overlapped between experiment and simulation). We further proposed a geometric similarity measure with regression adjustment (as explained in Section 3.3), and a ‘direct’ method without regression adjustment.”

In addition, the explanation of regression adjustment in Section 3.3 is not sufficiently clear. More detail, possibly in the Supplementary Information (SI), would be helpful.

2) In Section 2.4, it is interesting that the full model demonstrated greater plausibility than the white-matter-only model for tumors 4, 5, and 7. Since these are the largest tumors, could this be due to their increased dependence on vasculature? A discussion of the biological rationale behind this finding would strengthen the manuscript.

3) If the time scale is adjusted for each experiment, how can the model be used to make predictions in scenarios where no prior data are available? Since the model uses a real-time unit, one could potentially estimate the growth pattern from other data sources (e.g., RNA-seq). Additionally, is the spatial unit also adjusted for each experiment? Please clarify.

4) The manuscript lacks information regarding model uncertainty. Given that this is a stochastic model, how many replicates per parameter set are required to produce an accurate summary statistic of the model output?

Minor Comments:

1) In the last paragraph of Section 2.2, please describe the meaning of “formula 4 and 5” to improve readability and flow.

2) Figures 2 and 3 lack captions and are not clearly referenced in the main text.

3) In Section 2.5, the authors state:

“For tumors 1–4 and 6, it appears that the cell count is lowest when the tumor is treated with only anti-migration or only anti-proliferation drugs, with a maximum somewhere between the two extreme cases.”

Does this observation not apply to tumor 5? Please clarify.

4) In SI Section 5.2.4, the comparison steps need more detail:

Step 1: What criterion was used for decision-making? Was it based on error?

Step 2: What technique was used in the binarization step?

Step 4: Please describe the “closest point” method in one sentence to give readers a quick understanding.

Step 5: Provide more detail about how the transformation was applied.

**********

Have the authors made all data and (if applicable) computational code underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data and code 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 and code 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 or code —e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

Reviewer #2: Yes

**********

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.]

Figure resubmission:

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. If there are other versions of figure files still present in your submission file inventory at resubmission, please replace them with the PACE-processed versions.

Reproducibility:

To enhance the reproducibility of your results, we recommend that authors of applicable studies deposit laboratory protocols in protocols.io, where a protocol can be assigned its own identifier (DOI) such that it can be cited independently in the future. Additionally, PLOS ONE offers an option to publish peer-reviewed clinical study protocols. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols

PCOMPBIOL-D-24-02103R1

Anatomically aware simulation of patient-specific glioblastoma xenografts

PLOS Computational Biology

Dear Dr. Malik,

Thank you for submitting your manuscript to PLOS Computational Biology. After careful consideration, we feel that it has merit but does not fully meet PLOS Computational Biology'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 pay particular attention to the comments of reviewer 2, asking to integrate discussion points from the response letter into the manuscript.

Please submit your revised manuscript by Jan 05 2026 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 ploscompbiol@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pcompbiol/ 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 editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'. This file does not need to include responses to formatting updates and technical items listed in the 'Journal Requirements' section below.

* 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, competing interests statement, or data availability statement, please make these updates within the submission form at the time of resubmission. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter.

We look forward to receiving your revised manuscript.

Kind regards,

Christoph Kaleta

Section Editor

PLOS Computational Biology

Christoph Kaleta

Section Editor

PLOS Computational Biology

Journal Requirements:

If the reviewer comments include a recommendation to cite specific previously published works, please review and evaluate these publications to determine whether they are relevant and should be cited. There is no requirement to cite these works unless the editor has indicated otherwise.

1) Please amend your detailed Financial Disclosure statement. This is published with the article. It must therefore be completed in full sentences and contain the exact wording you wish to be published.

1) State the initials, alongside each funding source, of each author to receive each grant. For example: "This work was supported by the National Institutes of Health (####### to AM; ###### to CJ) and the National Science Foundation (###### to AM)."

2) State what role the funders took in the study. If the funders had no role in your study, please state: "The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript."

3) If any authors received a salary from any of your funders, please state which authors and which funders..

If you did not receive any funding for this study, please simply state: u201cThe authors received no specific funding for this work.u201d

2) Please ensure that the funders and grant numbers match between the Financial Disclosure field and the Funding Information tab in your submission form. Note that the funders must be provided in the same order in both places as well.

3) Please ensure that the files are uploaded in the online submission form in a correct numerical order

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Authors:

Please note here if the review is uploaded as an attachment.

Reviewer #1: I have no further comments, the authors have addressed my concerns in in their revised manuscript.

Reviewer #2: The following points were addressed satisfactorily in the response but should still be integrated into the revised manuscript:

Q2) The authors provide a satisfactory conceptual response, but this point should be explicitly mentioned in the Discussion section as a potential model extension and limitation, particularly regarding proliferation dependency on vascular proximity and nutrient/oxygen availability.

Q3) The response is satisfactory and well reasoned, but the discussion should be expanded in the manuscript to reflect these limitations and potential extensions (e.g., need for prior data, possible integration of RNA-seq).

Q4) The justification is sound, but the manuscript should explicitly mention that one replicate per parameter set was used because of observed low variance, and that parameter estimation accounted for this low stochastic variability.

Minor comment:

Q3) The authors’ response indicates the description has been rewritten, but I cannot find this change in the revised manuscript. The clarification therefore appears incomplete. Please update the manuscript to include the full corrected explanation and indicate where this change was made.

**********

Have the authors made all data and (if applicable) computational code underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data and code 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 and code 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 or code —e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

Reviewer #2: Yes

**********

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: Yes: David A. Hormuth, II

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.]

Figure resubmission:

While revising your submission, we strongly recommend that you use PLOS’s NAAS tool (https://ngplosjournals.pagemajik.ai/artanalysis) to test your figure files. NAAS can convert your figure files to the TIFF file type and meet basic requirements (such as print size, resolution), or provide you with a report on issues that do not meet our requirements and that NAAS cannot fix.

After uploading your figures to PLOS’s NAAS tool - https://ngplosjournals.pagemajik.ai/artanalysis, NAAS will process the files provided and display the results in the "Uploaded Files" section of the page as the processing is complete. If the uploaded figures meet our requirements (or NAAS is able to fix the files to meet our requirements), the figure will be marked as "fixed" above. If NAAS is unable to fix the files, a red "failed" label will appear above. When NAAS has confirmed that the figure files meet our requirements, please download the file via the download option, and include these NAAS processed figure files when submitting your revised manuscript.

Reproducibility:

To enhance the reproducibility of your results, we recommend that authors of applicable studies deposit laboratory protocols in protocols.io, where a protocol can be assigned its own identifier (DOI) such that it can be cited independently in the future. Additionally, PLOS ONE offers an option to publish peer-reviewed clinical study protocols. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols

Dear Dr. Malik,

We are pleased to inform you that your manuscript 'Anatomically aware simulation of patient-specific glioblastoma xenografts' has been provisionally accepted for publication in PLOS Computational Biology.

Before your manuscript can be formally accepted you will need to complete some formatting changes, which you will receive in a follow up email. A member of our team will be in touch with a set of requests.

Please note that your manuscript will not be scheduled for publication until you have made the required changes, so a swift response is appreciated.

IMPORTANT: The editorial review process is now complete. PLOS will only permit corrections to spelling, formatting or significant scientific errors from this point onwards. Requests for major changes, or any which affect the scientific understanding of your work, will cause delays to the publication date of your manuscript.

Should you, your institution's press office or the journal office choose to press release your paper, you will automatically be opted out of early publication. We ask that you notify us now if you or your institution is planning to press release the article. All press must be co-ordinated with PLOS.

Thank you again for supporting Open Access publishing; we are looking forward to publishing your work in PLOS Computational Biology.

Best regards,

Christoph Kaleta

Section Editor

PLOS Computational Biology

Christoph Kaleta

Section Editor

PLOS Computational Biology

***********************************************************

Reviewer's Responses to Questions

Comments to the Authors:

Please note here if the review is uploaded as an attachment.

Reviewer #2: Given that the authors have successfully incorporated the necessary changes into their revised manuscript, my concerns are fully resolved.

**********

Have the authors made all data and (if applicable) computational code underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data and code 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 and code 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 or code —e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #2: Yes

**********

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