Do Elevated YKL-40 Levels Drive the Immunosuppressive Tumor Microenvironment in Colorectal Cancer? Assessment of the Association of the Expression of YKL-40, MMP-8, IL17A, and PD-L1 with Coexisting Type 2 Diabetes, Obesity, and Active Smoking

Round 1

Reviewer 1 Report

The link between YKL-40 and molecules involved in cancer metastasis and immunosuppression in the tumor environment of colorectal cancer is not yet clear. In this manuscript, the authors used database information to examine the expression and correlation of YKL-40 and tumor-associated molecules. To determine these correlations, they also analyzed the expression of YKL-40 and tumor-associated molecules in samples from colorectal cancer patients.

However, the reviewer thinks this manuscript seems to present only preliminary findings. The data are primarily correlative, and the cause-or-effect relationship is unclear. Further, convincing data describing the novel aspects of this work are missing.

Overall, this manuscript is too preliminary to be considered for publication.

 

・ref6: It is inappropriate that this is the only reference to chronic inflammation.

・ref7: reference no.7 is inappropriate to insert in this position because it references e-cigarettes.

・ref11: The Citation in reference no.11 is incorrect.

・ref14: reference no.14 does not include this textbook in its reference list, although it contains many figures very similar to those in the book "Immunobiology".

・ref27: The information in the article in reference no.27 is not suitable as a reference.

・ref30: The authors should cite other papers in reference no.30 as it is a pre-peer-reviewed paper.

 

・The reviewer could not find Figure 4 & 5.

 

・line221: Figure7 -> Figure 4

 

・In the ELISA assay, the authors normalize the expression of the target protein by the total protein level. Still, the assay is usually performed using a recombinant target protein whose concentration is known in advance as a standard.

First, the reliability of evaluating the expression of target proteins by ELISA assay alone is questionable.

Why don't the authors use Western blotting to evaluate the expression of target proteins? They should.

Author Response

Dear reviewers

 

Thank you very much for your reviews.

We are thankful to one reviewer for pointing out the lack of two very important figures from the "results" section, which are very important for the reception of the whole work. The figures must have been lost during the manuscript conversion due to their incorrect format. Now the missing figures are available.

 

We have made corrections to the manuscript and corrected citations according to your recommendations:

1. We have expanded the introduction with additional information regarding the development of PD-L1 therapies in cancer.
2. We elaborated on the potential clinical significance of the obtained results and the proposals, as well as the importance of further studies on the interaction of CHI3L1 with PD-L1;
3. We corrected the citations from the previous version of the manuscript, the scope of the corrections was as recommended: citation no. 6, no. 7, no. 11, no. 14, no. 30.
4. Citation No. 27 (in the previous version of the manuscript, regarding CAMOIP, we left it unchanged - because CAMOIP on its website indicates this publication as the right one to be cited when using this tool);
5. We made some minor changes to the abstract; however, due to the 200-character limit; we didn't make big changes; it is hoped that the changes made have improved the abstract sufficiently for a cursory review of the entire manuscript;
6. Fixed minor mistakes as indicated in the reviews.

 

 

Answering the questions:

1. After conferring with the research team, we have concluded that we cannot organize cell culture research on that topic in the coming months. However, we believe that the analysis of expression in GEO genesets will constitute a sufficient validation of the ELISA results.
2. We did not evaluate the expression of the tested proteins using the Western blot method, because the ELISA kits used in the study were dedicated to tissue homogenates and used reference concentrations of the target protein. This is the reason why we did not further validate the ELISA results with Western blot.

 

An important limitation of the study, as indicated by one of the reviewers, is the lack of thorough examination of the still unclear direct impact of CHI3L1 expression on the target proteins under study. However, our study, together with high-dimensional analyses such as the GSEA, seem to fill important gaps in the current literature on CHI3L1 immunological relationships in colorectal cancer.

This may become more important shortly, due to the significant development of small molecules targeting CHI3L1, the importance of PD-L1 in photodynamic therapy, and chitosan Nanoparticles.

 

 

Thank you for your reviews that have helped us improve our work.

We look forward to hearing from you.

The Authors

Reviewer 2 Report

In this research, the authors elevated the role of different concentrations of YKL-40 in driving immunosuppressive tumor micro-environment in colorectal cancer. Generally, it’s a meaningful and interesting research but still needs to be improved before the possible acception. In my opinion, the current version of this manuscript fits the scope of Curr. Issues Mol. Biol. and could be accepted after major revision.

My specific comments are in detail listed below:

1.     The abbreviations should be explained at the place it first mentioned, such as the YKL-40 (CHI3L1).

2.     The abstract was poorly written. In my opinion, a better and brief summary and summarize could be more clearly added.

3.     In this introduction (Line 74-87), the current development of PD-1/PD-L1 axis should be more clearly discussed. Some references should be added including 10.1016/j.molcel.2019.04.005, 10.1002/adma.202206121, 10.1038/S41467-021-25416-7, doi.org/10.1016/j.apsb.2022.07.023, and 10.1016/j.molcel.2018.07.030. 

4.     All the figures are of low quality. If possible, the authors may could improve it to obtain a better reading experience.

5.     If possible, some in vitro cell experiments an in vivo anti-tumor experiments should be added to evaluate the discovered by the authors.

6.     In the conclusion and future perspective part, the clinical transformation prospect should be more clearly discussed and revealed. Some references may be helpful to the authors, including 10.1126/scitranslmed.aav7431, 10.1016/j.cej.2022.140164, 10.1016/j.jconrel.2022.11.004, and 10.1073/pnas.2114851119.

7.     Some minor mistakes existed in this paper. The authors should carefully check it.

Author Response

Dear reviewers

 

Thank you very much for your reviews.

We are thankful to one reviewer for pointing out the lack of two very important figures from the "results" section, which are very important for the reception of the whole work. The figures must have been lost during the manuscript conversion due to their incorrect format. Now the missing figures are available.

 

We have made corrections to the manuscript and corrected citations according to your recommendations:

1. We have expanded the introduction with additional information regarding the development of PD-L1 therapies in cancer.
2. We elaborated on the potential clinical significance of the obtained results and the proposals, as well as the importance of further studies on the interaction of CHI3L1 with PD-L1;
3. We corrected the citations from the previous version of the manuscript, the scope of the corrections was as recommended: citation no. 6, no. 7, no. 11, no. 14, no. 30.
4. Citation No. 27 (in the previous version of the manuscript, regarding CAMOIP, we left it unchanged - because CAMOIP on its website indicates this publication as the right one to be cited when using this tool);
5. We made some minor changes to the abstract; however, due to the 200-character limit; we didn't make big changes; it is hoped that the changes made have improved the abstract sufficiently for a cursory review of the entire manuscript;
6. Fixed minor mistakes as indicated in the reviews.

 

 

Answering the questions:

1. After conferring with the research team, we have concluded that we cannot organize cell culture research on that topic in the coming months. However, we believe that the analysis of expression in GEO genesets will constitute a sufficient validation of the ELISA results.
2. We did not evaluate the expression of the tested proteins using the Western blot method, because the ELISA kits used in the study were dedicated to tissue homogenates and used reference concentrations of the target protein. This is the reason why we did not further validate the ELISA results with Western blot.

 

An important limitation of the study, as indicated by one of the reviewers, is the lack of thorough examination of the still unclear direct impact of CHI3L1 expression on the target proteins under study. However, our study, together with high-dimensional analyses such as the GSEA, seem to fill important gaps in the current literature on CHI3L1 immunological relationships in colorectal cancer.

This may become more important shortly, due to the significant development of small molecules targeting CHI3L1, the importance of PD-L1 in photodynamic therapy, and chitosan Nanoparticles.

 

 

Thank you for your reviews that have helped us improve our work.

We look forward to hearing from you.

The Authors

Round 2

Reviewer 1 Report

Overall Recommendation: Reject (unless the author responds in good faith to the following comments)
  Comments and Suggestions for Authors, 1. The addition of two new authors to a revised manuscript should be explained. The addition of authors without proper justification has recently become a problem.   2. lane172(section 2.5): CHI3L1 (YKL17240) concentration was measured using MICROVUE YKL-40 ELISA Kit (Quidel, San Diego, 173 USA) We used MICROVUE YKL-40 ELISA Kit (Quidel, San Diego, USA) to measure CHI3L1(YKL-40) concentration. .....   The same sentences are repeated.   3.   3.3. Exploration of Immunogenicty and Immune Infiltration Landscape of YKL-40 based on TCGACOAD data.
The immune scores of Intratumor Heterogeneity, Macrophage regulation, lymphocyte infiltration signature score, IFN-gamma response, Th1 cells, and TGF-beta response were significantly up-regulated in the YKL-40 high expression group (Figure 7, A-B) unlike proliferation and Th2 cells score which were decreased in this group…….
  -> Isn't Figure 7 in lane 235 (section 3.3) a mistake for Figure 4?

Further, Figure 4 is very unclear, and it is impossible to determine if this data is adequate. It is not reader-friendly.
Since the reviewer cannot determine whether Figure 4 is appropriate for the results obtained and the text in Section 3.3, the reviewer cannot accept the revised manuscript.

Author Response

Dear Reviewer

 

Thank you very much for all your new comments and suggestions.

 

As you wrote in your review, we have corrected the duplication of the sentence in section 2.5, the wrong figure reference in section 3.3, and significantly improved the quality and readability of figure 4.

 

Regarding the change of the authorship of the manuscript, a few days after the submission of the manuscript, we became aware of our mistake. New authors were not added as a result of manuscript revision after the first reviews, but we agreed to add two co-authors with the editor before receiving the first-round review.

The authorship of the paper, together with the two added co-authors, is the same as during the previous submission of our manuscript to the Metabolites Special Issue, from where we were redirected to CIMB.

 

We thank the Reviewer for all comments and for the time devoted to our manuscript.

We look forward to hearing further information from you regarding our manuscript.

 

Sincerely

The authors

 

Reviewer 2 Report

The current version of this manuscript could be accepted.

Author Response

Dear Reviewer 

Thank you for your review and cooperation. 

 

Sincerely

The authors 

Round 3

Reviewer 1 Report

The manuscript has been improved. There are only minor points. Please refer to the attached file.

Comments for author File: Comments.pdf

Author Response

Dear Reviewer

Thank you very much for your reply.
We have corrected the errors you pointed out in the manuscript.
We look forward to hearing from you

Sincerely,
The authors