Discovery of novel benzophenone integrated derivatives as anti-Alzheimer’s agents targeting presenilin-1 and presenilin-2 inhibition: A computational approach

Reshma Mary Martiz; Shashank M. Patil; Ramith Ramu; Jayanthi M. K.; Ashwini P.; Lakshmi V. Ranganatha; Shaukath Ara Khanum; Ekaterina Silina; Victor Stupin; Raghu Ram Achar

doi:10.1371/journal.pone.0265022

Peer Review History

Original SubmissionNovember 1, 2021
13 Dec 2021 Decision Letter - Jie Zheng, Editor PONE-D-21-34824Discovery of novel benzophenone integrated derivatives as anti-Alzheimer’s agents targeting presenilin-1 and presenilin-2 inhibition: a computational approachPLOS ONE Dear Dr. Ramu, 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 Jan 27 2022 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, Jie Zheng, Ph.D 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. Thank you for stating the following in the Funding Section of your manuscript: The author(s) reported there is no funding associated with the work featured in this article. We note that you have provided funding information that is not currently declared in your Funding Statement. However, funding information should not appear in the Acknowledgments section or other areas of your manuscript. We will only publish funding information present in the Funding Statement section of the online submission form. Please remove any funding-related text from the manuscript and let us know how you would like to update your Funding Statement. Currently, your Funding Statement reads as follows: The author(s) reported there is no funding associated with the work featured in this article Please include your amended statements within your cover letter; we will change the online submission form on your behalf. 3. PLOS requires an ORCID iD for the corresponding author in Editorial Manager on papers submitted after December 6th, 2016. Please ensure that you have an ORCID iD and that it is validated in Editorial Manager. To do this, go to ‘Update my Information’ (in the upper left-hand corner of the main menu), and click on the Fetch/Validate link next to the ORCID field. This will take you to the ORCID site and allow you to create a new iD or authenticate a pre-existing iD in Editorial Manager. Please see the following video for instructions on linking an ORCID iD to your Editorial Manager account: https://www.youtube.com/watch?v=_xcclfuvtxQ 4. Please amend your authorship list in your manuscript file to include author Reshma Mary Martiz and Shashank M Patil. 5. We note that Figures 1 and 5 in your submission contain copyrighted images. All PLOS content is published under the Creative Commons Attribution License (CC BY 4.0), which means that the manuscript, images, and Supporting Information files will be freely available online, and any third party is permitted to access, download, copy, distribute, and use these materials in any way, even commercially, with proper attribution. For more information, see our copyright guidelines: http://journals.plos.org/plosone/s/licenses-and-copyright. We require you to either (1) present written permission from the copyright holder to publish these figures specifically under the CC BY 4.0 license, or (2) remove the figures from your submission: a. You may seek permission from the original copyright holder of Figures 1 and 5 to publish the content specifically under the CC BY 4.0 license. We recommend that you contact the original copyright holder with the Content Permission Form (http://journals.plos.org/plosone/s/file?id=7c09/content-permission-form.pdf) and the following text: “I request permission for the open-access journal PLOS ONE to publish XXX under the Creative Commons Attribution License (CCAL) CC BY 4.0 (http://creativecommons.org/licenses/by/4.0/). Please be aware that this license allows unrestricted use and distribution, even commercially, by third parties. Please reply and provide explicit written permission to publish XXX under a CC BY license and complete the attached form.” Please upload the completed Content Permission Form or other proof of granted permissions as an ""Other"" file with your submission. In the figure caption of the copyrighted figure, please include the following text: “Reprinted from [ref] under a CC BY license, with permission from [name of publisher], original copyright [original copyright year].” b. If you are unable to obtain permission from the original copyright holder to publish these figures under the CC BY 4.0 license or if the copyright holder’s requirements are incompatible with the CC BY 4.0 license, please either i) remove the figure or ii) supply a replacement figure that complies with the CC BY 4.0 license. Please check copyright information on all replacement figures and update the figure caption with source information. If applicable, please specify in the figure caption text when a figure is similar but not identical to the original image and is therefore for illustrative purposes only. [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: Partly ******** 2. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: No Reviewer #2: Yes ****** 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: The study analyzed the binding affinity between benzophenones and PSEN-1/2 for demonstrating the pharmacotherapeutic potential by inhibiting PSEN-1/2. However, the study only focused on analyzing the affinities and interactions, without further investigation of the preferential binding pocket for the inhibition effect. Herein, it is hard to conclude the inhibition efficiency of screened benzophenone from these preliminary data, and many issues need to be addressed as followings: 1. Figure 3-4, the binding pocket and binding sequence labeled in different colors should be clearly explained in the figure caption. 2. Table 3-4, the in parallel comparison of binding affinity and non-bonding interactions of various BIDs are encouraged to present in column figures for better illustration. Also, please highlight the molecule structural differences of these BIDs in different color. 3. Figure 5-6, it is hard to distinguish between the Pi-alkyl and alkyl interactions. 4. The binding interactions between BIDs and PSEN-1/2 were analyzed in silico, but for the inhibition effect, how this binding contribute to the inhibition? Whether by stabilize the PSEN-1/2 conformation or block the active sites? All these need to be further introduced to demonstrate the inhibition benefits. 5. In addition to the binding affinity, the binding pocket should also be analyzed for screening the best pose of BID-16 with PSEN-1/2. Reviewer #2: In the manuscript PONE-D-21-34824, the study evaluates the interaction of BIDs through molecular docking simulations, molecular dynamics simulations, and binding free energy calculations. Combinations of these approaches are acceptable. Although the manuscript is informative, there are still some questions that need to be addressed (see comments listed below). 1. The false-positive rate of molecular docking is very high in the absence of experimental support. The literature listed by the author only mentions the inhibition of AChE/BACE-1 by Benzophenones, not PSEN-1/PSEN-2. Therefore, there is little significance in performing molecular docking study in such cases. 2. The authors used CHARMM27 force field to get the parameter for protein (Line 126 to 129 in manuscript). However, their studies were based on an obsolete force field, which need a revision with the latest force fields. This can impact the results, weakening some analysis. One should use CHARMM36 instead of 27 (please see doi.org/10.1021/ct300400x). 3. Typically, the two compounds with the best activity and the worst activity are usually selected for comparison in MD study. The authors should add a compound as the negative control. 4. Table 3 and Table 4 contain the same dataset and should be merged into one table. 5. One gets the impression that the work was assembled very hastily and without careful control. For instance, there are miscellaneous lines on the Fig.8B, which may be caused by the incorrect modification of the residue number. Also, Fig. 7B and 8B are missing ordinate labels. The units in the pictures should be unified (e.g. Fig7A, 7C, 7D, 7E, 8A, 8C, 8D, 8E). As a suggestion, the authors should use professional drawing software to improve the quality of these pictures. ****** 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. https://doi.org/10.1371/journal.pone.0265022.r001
Revision 1
18 Jan 2022 Author Response Reviewer 1 comments The study analysed the binding affinity between benzophenones and PSEN-1/2 for demonstrating the pharmacotherapeutic potential by inhibiting PSEN-1/2. However, the study only focused on analysing the affinities and interactions, without further investigation of the preferential binding pocket for the inhibition effect. Herein, it is hard to conclude the inhibition efficiency of screened benzophenone from these preliminary data, and many issues need to be addressed as followings: 1. Figure 3-4, the binding pocket and binding sequence labelled in different colours should be clearly explained in the figure caption. Response: As per the reviewer’s suggestion, binding pocket and binding sequence labelled in different colours (for both PSEN-1 and PSEN-2) have been explained in figure captions of Figure 3 (line 201-204) and Figure 4 (line 206-209). 2. Table 3-4, the in parallel comparison of binding affinity and non-bonding interactions of various BIDs are encouraged to present in column figures for better illustration. Also, please highlight the molecule structural differences of these BIDs in different colour. Response: As per the reviewer’s suggestion, Table 3 and Table 4 have been merged. The merged table (Table 3, line 225) now contains the structural differences of the BIDs highlighted in different colour. Also, binding affinity, total number of non-bonding interactions, total number of hydrogen bonds formed by BIDs during the docking simulation have also been given in adjacent columns. For the better illustration of these interaction parameters, Figure 5 and Figure 6 have been added, which represent the different docking parameters of PSEN-1 and PSEN-2, respectively. 3. Figure 5-6, it is hard to distinguish between the Pi-alkyl and alkyl interactions. Response: As per the reviewer’s suggestion, colour of the Pi-alkyl interactions have been changed to dark pink colour (now Figure 7 and Figure 8). Also, row headings have been added in Table 4, which clearly describes the types of interactions to avoid confusion. 4. The binding interactions between BIDs and PSEN-1/2 were analysed in silico, but for the inhibition effect, how this binding contributes to the inhibition? Whether by stabilize the PSEN-1/2 conformation or block the active sites? All these need to be further introduced to demonstrate the inhibition benefits. Response: As per the reviewer’s suggestion, in silico interaction of BIDs have now been supported with suitable information in “Discussion” section (lines 439-462) and supplementary data. In order to do this, authors have carefully performed the binding pocket analysis of both PSEN proteins, followed by the interaction analysis of BIDs with specific amino acid residues. We have identified a few key amino acid residues from the literature analysis, binding to which induces the inhibition activity. Binding interactions of BIDs also reveal that the key amino acids have been bound with the BIDs. Therefore, the interaction is oriented towards the inhibition of the protein activity by blocking the active binding residues. From previous studies conducted on γ-secretase, which consists of PSEN-1 and PSEN-2 proteins as functional subunits, we have provided the information of in silico inhibition of γ-secretase. The same inhibitor compound (LY450139) used in the study was docked with our homology model built PSEN-1 and PSEN-2 proteins. This has produced similar binding interactions like BIDs, which depicts that the binding interactions of BIDs have been oriented towards the inhibition, not the activation of the target proteins. 5. In addition to the binding affinity, the binding pocket should also be analysed for screening the best pose of BID-16 with PSEN-1/2. Response: As per the reviewer’s suggestion, a thorough analysis of binding pockets of both PSEN-1 and PSEN-2 was conducted using the literature given in the UniProt database website. The literature survey resulted in the identification of few key amino residues, binding to which causes the inhibition of the protein activity. Results from our molecular docking studies reveal that BID-16 binds to the same amino acid residues, which shows that BID-16 as a potential inhibitor of both the PSEN-1 and PSEN-2 proteins. Difference in MD simulation plots were also given to highlight the impact of interaction (lines 476-481). During the virtual screening of the different binding poses, the first binding pose with zero RMSD and most negative binding affinity was selected. The same binding pose was found to interact with the key residues, in comparison with the other 9 binding poses. Reviewer 2 comments In the manuscript PONE-D-21-34824, the study evaluates the interaction of BIDs through molecular docking simulations, molecular dynamics simulations, and binding free energy calculations. Combinations of these approaches are acceptable. Although the manuscript is informative, there are still some questions that need to be addressed (see comments listed below). 1. The false-positive rate of molecular docking is very high in the absence of experimental support. The literature listed by the author only mentions the inhibition of AChE/BACE-1 by Benzophenones, not PSEN-1/PSEN-2. Therefore, there is little significance in performing molecular docking study in such cases. Response: As per the reviewer’s suggestion, we conducted an extensive literature survey on the involvement of BIDs in the induction of anti-Alzheimer’s activity. However, there have been no studies performed in vitro for the inhibition of PSEN-1 and PSEN-2 by BIDs. The only available study on the anti-Alzheimer’s activity of BIDs is the inhibition of AChE/BACE-1 by Belluti et al. (2014) (reference no. 5). Therefore, we aimed to investigate the anti-Alzheimer’s activity through PSEN inhibition. This study provides an insight to the molecular mechanism of inactivating PSEN proteins through blocking their active sites. As this study is an in silico investigation, in vitro and in vivo studies in future could be performed using BIDs aiming for PSEN inhibition. 2. The authors used CHARMM27 force field to get the parameter for protein (Line 126 to 129 in manuscript). However, their studies were based on an obsolete force field, which need a revision with the latest force fields. This can impact the results, weakening some analysis. One should use CHARMM36 instead of 27 (please see doi.org/10.1021/ct300400x). Response: As per the reviewer’s suggestion, the force field used for molecular dynamics simulation was changed from CHARMM27 to CHARMM36. According to the reviewer 2 comments the negative control was need to be added in case of both docking and dynamics simulation processes. Therefore, MD simulations of all the BIDs for both PSEN-1 and PSEN-2 were performed again using CHARMM36 force field. 3. Typically, the two compounds with the best activity and the worst activity are usually selected for comparison in MD study. The authors should add a compound as the negative control. Response: As per the reviewer’s suggestion, negative controls with worst activity were selected for the comparative study. In case of PSEN-1, BID-19 was selected as the negative control, whereas in case of PSEN-2, BID-20 was selected as the same. The visualization/representation of docking studies, dynamics simulation studies, binding free energy calculations, druglikeliness, and pharmacokinetic analysis were also completed by comparing the BID-16 with negative controls. 4. Table 3 and Table 4 contain the same dataset and should be merged into one table. Response: As per the reviewer’s suggestion, both Table 3 and Table 4 were merged into one table (now Table 3). The table now contains data of structural differentiation of BIDs as well as their interaction details. 5. One gets the impression that the work was assembled very hastily and without careful control. For instance, there are miscellaneous lines on the Fig.8B, which may be caused by the incorrect modification of the residue number. Also, Fig. 7B and 8B are missing ordinate labels. The units in the pictures should be unified (e.g. Fig7A, 7C, 7D, 7E, 8A, 8C, 8D, 8E). As a suggestion, the authors should use professional drawing software to improve the quality of these pictures. Response: As per the reviewer’s suggestion, negative controls were added for the MD simulation study. Using CHARMM36 forcefield, all the simulations were re-run for 100 ns. All the trajectories of MD simulations were plotted using XMGRACE. The plots were unified and put in one single picture using InkScape software. Attachments Attachment Submitted filename: Author Response (1).docx https://doi.org/10.1371/journal.pone.0265022.r002
21 Feb 2022 Decision Letter - Jie Zheng, Editor Discovery of novel benzophenone integrated derivatives as anti-Alzheimer’s agents targeting presenilin-1 and presenilin-2 inhibition: a computational approach PONE-D-21-34824R1 Dear Dr. Ramu, 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, Jie Zheng, Ph.D Academic Editor PLOS ONE Additional Editor Comments (optional): Reviewers' comments: https://doi.org/10.1371/journal.pone.0265022.r003
Formally Accepted
22 Mar 2022 Acceptance Letter - Jie Zheng, Editor PONE-D-21-34824R1 Discovery of novel benzophenone integrated derivatives as anti-Alzheimer’s agents targeting presenilin-1 and presenilin-2 inhibition: a computational approach Dear Dr. Ramu: I'm pleased to inform you that your manuscript has been deemed suitable for publication in PLOS ONE. Congratulations! Your manuscript is now with our production department. If your institution or institutions have a press office, please let them know about your upcoming paper now to help maximize its impact. If they'll be preparing press materials, please inform our press team within the next 48 hours. 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. If we can help with anything else, please email us at plosone@plos.org. Thank you for submitting your work to PLOS ONE and supporting open access. Kind regards, PLOS ONE Editorial Office Staff on behalf of Dr. Jie Zheng Academic Editor PLOS ONE https://doi.org/10.1371/journal.pone.0265022.r004

Open letter on the publication of peer review reports

PLOS recognizes the benefits of transparency in the peer review process. Therefore, we enable the publication of all of the content of peer review and author responses alongside final, published articles. Reviewers remain anonymous, unless they choose to reveal their names.

We encourage other journals to join us in this initiative. We hope that our action inspires the community, including researchers, research funders, and research institutions, to recognize the benefits of published peer review reports for all parts of the research system.

Learn more at ASAPbio .