Investigating the Epigenetic Therapeutic Potential of Natural Compounds in Cancer

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

1. The review effectively summarizes the epigenetic effects of natural compounds; however, it would benefit from a clearer mechanistic comparison among alkaloids, terpenoids, and polyphenols to identify shared molecular targets or signaling pathways.
2. The manuscript should include a brief section discussing bioavailability and pharmacokinetic limitations of natural compounds that may affect their clinical applicability.
3. Several subsections describe in vitro results in great detail; integrating these with in vivo and clinical trial data would strengthen the translational relevance.
4. The role of combination therapy (natural compound + epigenetic drug) is mentioned but not deeply analyzed—please expand with examples where synergistic or antagonistic effects were observed.
5. The figures summarizing epigenetic mechanisms (e.g., Figures 1–3) could incorporate unified pathway diagrams linking DNA methylation, histone modification, and miRNA regulation.
6. Some compounds, such as apigenin and oleanolic acid, are discussed for multiple cancers; a comparative table summarizing their specific molecular effects across cancer types would enhance clarity.
7. The manuscript focuses on canonical epigenetic marks (DNA methylation, histone acetylation); emerging marks like histone lactylation and RNA methylation (m6A) could also be briefly discussed for completeness.
8. There is minimal discussion of the dose-dependent or concentration-specific dual roles (protective vs. toxic) of these compounds; this is crucial for therapeutic relevance.
9. The conclusion should better integrate the challenges of translating preclinical epigenetic modulation to human therapies, including compound stability, specificity, and delivery strategies.
   
   
   

   Technical Comments:

   1. There are multiple instances of placeholder errors (“Error! Reference source not found.”) throughout the text—these must be corrected before submission.
   2. Ensure consistency in compound nomenclature (e.g., “3,3’-Diindolylmethane (DIM)” vs. “3,3’-diindolylmethane”) and follow IUPAC or accepted pharmacological naming conventions.
   3. Some figure legends (e.g., Figure 1–3) lack sufficient detail on abbreviations and experimental context; add concise, self-contained explanations.
   4. Tables 1 and 2 contain numerous upward/downward arrows (↑/↓); include a clear legend and maintain uniform formatting for readability.
         5. The manuscript would benefit from language refinement for conciseness and       scientific tone—consider reducing repetitive phrases like “has been shown” and “studies have demonstrated.”

Author Response

Comment 1. The review effectively summarizes the epigenetic effects of natural compounds; however, it would benefit from a clearer mechanistic comparison among alkaloids, terpenoids, and polyphenols to identify shared molecular targets or signaling pathways.

Response 1: We appreciate the reviewer’s valuable suggestion. To address this comment, we have added a new summary figure (Figure 3 ) that provides a comparative overview of the epigenetic mechanisms through which alkaloids, terpenoids, and polyphenols regulate cancer-related cellular processes. The figure highlights the shared molecular targets and signaling pathways, as well as the key biological outcomes, including apoptosis, cell cycle regulation, and modulation of tumor suppressor genes, which collectively influence cancer progression (proliferation, migration, invasion, and angiogenesis). This addition aims to enhance the clarity and translational relevance of the mechanistic comparison among the major classes of natural compounds discussed in the review.

Comment 2: The manuscript should include a brief section discussing bioavailability and pharmacokinetic limitations of natural compounds that may affect their clinical applicability.

Response 2: In response, we have added a new Section 3, which discusses the bioavailability, metabolic stability, and pharmacokinetic limitations of natural compounds that may influence their clinical applicability. This section also outlines potential strategies to overcome these challenges, such as the use of nanoformulations, structural modification, and targeted delivery systems.

 Comment 3: Several subsections describe in vitro results in great detail; integrating these with in vivo and clinical trial data would strengthen the translational relevance.

Response 3: We thank the reviewer for this valuable suggestion. In response, we have expanded Section 4, which now provides a more detailed discussion of clinical trial results to better integrate preclinical and clinical findings. Additionally, we have updated Table 5 by adding a new column summarizing the corresponding clinical outcomes, thereby enhancing the translational relevance and practical context of the reviewed data.

 

 Comment 4. The role of combination therapy (natural compound + epigenetic drug) is mentioned but not deeply analyzed—please expand with examples where synergistic or antagonistic effects were observed.

Response 4. We thank the reviewer for this constructive comment. In response, we have expanded the relevant section to provide a more detailed analysis of combination therapies involving natural compounds and epigenetic drugs. The revised text now includes specific examples illustrating both synergistic and antagonistic effects, clearly highlighting how such interactions influence epigenetic regulation, tumor suppression, and therapeutic outcomes. These additions aim to clarify the translational potential of combinatorial strategies in cancer therapy.

 Comment 5: The figures summarizing epigenetic mechanisms (e.g., Figures 1–3) could incorporate unified pathway diagrams linking DNA methylation, histone modification, and miRNA regulation.

Response 5. In response, due to the extensive amount of information contained in these figures, Figures 1 and 2 were divided into two parts each to enhance clarity and readability. Figures 4–6, which originally contained detailed information on polyphenols, were converted into a comprehensive summary table illustrating the effects of various polyphenol subclasses on epigenetic regulation. This approach allows for a clearer and more structured presentation of the complex interactions between different epigenetic mechanisms while maintaining figure readability and avoiding excessive visual complexity.

 Comment 6. Some compounds, such as apigenin and oleanolic acid, are discussed for multiple cancers; a comparative table summarizing their specific molecular effects across cancer types would enhance clarity.

Response 6: We thank the Reviewer for this insightful comment. In response to the suggestion, due to the extensive amount of information regarding polyphenols, including apigenin and oleanolic acid, we have summarized their effects in a comparative table. This table presents the specific molecular mechanisms and anticancer effects of these compounds across different cancer types, providing a clearer and more concise overview of their multifaceted epigenetic actions.

 Comment 7. The manuscript focuses on canonical epigenetic marks (DNA methylation, histone acetylation); 8. emerging marks like histone lactylation and RNA methylation (m6A) could also be briefly discussed for completeness.

Response 7. We appreciate the Reviewers’ suggestion to include emerging epigenetic marks. In the revised manuscript, we have now incorporated histone lactylation and crotonylation. Additionally, we have briefly discussed RNA methylation (m6A) in the context of chrysin and fisetin. These additions provide a more comprehensive overview of both canonical and emerging epigenetic mechanisms , enhancing the completeness of the manuscript.

 Comment 8. There is minimal discussion of the dose-dependent or concentration-specific dual roles (protective vs. toxic) of these compounds; this is crucial for therapeutic relevance.

Response 8. We thank the Reviewer for highlighting the importance of dose-dependent and concentration-specific effects. In the revised manuscript, information regarding the dual roles (protective versus toxic) of the polyphenolic compounds has been incorporated into Section 3. This section now addresses the concentration-specific effects, providing a clearer context for their therapeutic relevance.

Comment 9. The conclusion should better integrate the challenges of translating preclinical epigenetic modulation to human therapies, including compound stability, specificity, and delivery strategies.

Response 9. We appreciate the Reviewer’s suggestion. The Conclusion section has been revised to better integrate the challenges of translating preclinical findings on epigenetic modulation into human therapies. Specifically, we now discuss issues related to compound stability, target specificity, and delivery strategies, providing a more comprehensive perspective on the therapeutic potential of natural compounds.

Technical Comments:

Comment 1. There are multiple instances of placeholder errors (“Error! Reference source not found.”) throughout the text—these must be corrected before submission.

Response 1. All instances of placeholder errors (“Error! Reference source not found.”) have been corrected throughout the manuscript in accordance with the Reviewer’s recommendation.

Comment 2. Ensure consistency in compound nomenclature (e.g., “3,3’-Diindolylmethane (DIM)” vs. “3,3’-diindolylmethane”) and follow IUPAC or accepted pharmacological naming conventions.

Response 2. The nomenclature of all compounds has been standardized throughout the manuscript in accordance with IUPAC and accepted pharmacological conventions.

 Comment 3. Some figure legends (e.g., Figure 1–3) lack sufficient detail on abbreviations and experimental context; add concise, self-contained explanations.

Response 3. All figures now include detailed legends, providing concise, self-contained explanations of abbreviations and experimental context. This ensures that each figure and table can be understood independently of the main text.

Comment 4. Tables 1 and 2 contain numerous upward/downward arrows (↑/↓); include a clear legend and maintain uniform formatting for readability.

Response 4. We have implemented the Reviewer’s suggestion. All tables now include a clear legend explaining the meaning of upward (↑) and downward (↓) arrows, and formatting has been standardized to ensure uniformity and improved readability.

Comment 5. The manuscript would benefit from language refinement for conciseness and       scientific tone—consider reducing repetitive phrases like “has been shown” and “studies have demonstrated.”

Response 5. We appreciate the Reviewer’s suggestion. The manuscript has been carefully revised to improve conciseness and scientific tone, with repetitive phrases such as “has been shown” and “studies have demonstrated” reduced throughout the text.

 

Reviewer 2 Report

Comments and Suggestions for Authors

The manuscript is a comprehensive review of the potential of natural compounds to serve as anticancer agents.

The writing style is clear and engaging, and the authors addressed the influence of numerous molecules on epigenetic mechanisms involved in cancer development.

I have several comments for the authors:

• the main problem in the manuscript is the errors related to the references in the text (their absence)
• Figure 1 should be split into 2 parts, which should be enlarged
• Figure 2 should also be enlarged
• In Tables 2 and 3, the first line of the table repeats in the middle of the table
• Subtitle 2.2.2. must be corrected
• Rephrase the section between lines 852-853
• I recommend adding an additional column to Table 4 to briefly present the main conclusions of all completed clinical trials. While the authors discuss some trials results, including all would enhance the table's comprehensiveness.

Author Response

Comments 1. The main problem in the manuscript is the errors related to the references in the text (their absence)

Response 1. All instances of placeholder errors (“Error! Reference source not found.”) have been corrected throughout the manuscript in accordance with the Reviewer’s recommendation.

Comment 2. Figure 1 should be split into 2 parts, which should be enlarged, Figure 2 should also be enlarged

Response 2. In response, due to the extensive amount of information contained in these figures, Figures 1 and 2 were divided into two parts each to enhance clarity and readability. Figures 4–6, which originally contained detailed information on polyphenols, were converted into a comprehensive summary Table 4 illustrating the effects of various polyphenol subclasses on epigenetic regulation. This approach allows for a clearer and more structured presentation of the complex interactions between different epigenetic mechanisms while maintaining figure readability and avoiding excessive visual complexity.

Comment 3. In Tables 2 and 3, the first line of the table repeats in the middle of the table

Response 3.We thank the Reviewer for pointing this out. The repeated first lines in Tables 2 and 3 have been removed, and the tables have been carefully revised to ensure correct formatting and readability.

Comment 4. Subtitle 2.2.2. must be corrected

Response 4. The subtitle “2.2.2.” has been corrected as suggested to ensure proper formatting and consistency throughout the manuscript.

Comment 5. Rephrase the section between lines 852-853

Response 5. The section between lines 852–853 has been rephrased as suggested to improve clarity and readability.

Comment 6. I recommend adding an additional column to Table 4 to briefly present the main conclusions of all completed clinical trials. While the authors discuss some trials results, including all would enhance the table's comprehensiveness.

Response 6. In response to the Reviewer’s suggestion, an additional column has been added to Table 5 summarizing the main results of completed clinical trials. This enhancement provides a more comprehensive overview of the clinical evidence.

Reviewer 3 Report

Comments and Suggestions for Authors

It is a very interesting and comprehensive review, certainly worth to be published. However, there is one serious issue that needs to be clearified. Thus, paragraph 2.2.1 describes valeric acid, which it is not a terpenoid (despite the fact that it contains five carbon atoms). From the attached literature (two references) it seems that it is indeed valeric acid (in original paper it is declared as bought from some producer). However, is it possible that the data relate to more structuraly complex anticancer valerenic acid, a major component of Valeriana officinalis, please refer to : Valeriana officinalis extract and its main component, valerenic acid, Exp Gerontol 2013 ;48(11):1369-77. doi: 10.1016/j.exger.2013.09.002. There are more papers on anticancer activity of this terpenic compound.

Other small comments are as follows:

1./ in my copy references are not shown in the text - system says: "error......"

2./ line 83 should be demethylases;

3./ line 86: (PIN) should precede "lesions".

Author Response

Comments 1. It is a very interesting and comprehensive review, certainly worth to be published. However, there is one serious issue that needs to be clearified. Thus, paragraph 2.2.1 describes valeric acid, which it is not a terpenoid (despite the fact that it contains five carbon atoms). From the attached literature (two references) it seems that it is indeed valeric acid (in original paper it is declared as bought from some producer). However, is it possible that the data relate to more structuraly complex anticancer valerenic acid, a major component of Valeriana officinalis, please refer to : Valeriana officinalis extract and its main component, valerenic acid, Exp Gerontol 2013 ;48(11):1369-77. doi: 10.1016/j.exger.2013.09.002. There are more papers on anticancer activity of this terpenic compound.

Response 1. We thank the Reviewer for this insightful comment. The compound in paragraph 2.2.1 has been carefully reviewed, and the reference to valeric acid has been removed, as it is not a terpenoid. We appreciate the suggestion regarding valerenic acid, a terpenoid with anticancer activity, and have revised the text accordingly to avoid any confusion.

Comment 2. Other small comments are as follows:

1./ in my copy references are not shown in the text - system says: "error......"

2./ line 83 should be demethylases;

3./ line 86: (PIN) should precede "lesions".

 

Comment 3. We thank the Reviewer for these detailed observations. The following corrections have been made:

Response 3:

     1.All reference errors in the text (e.g., “Error…”) have been corrected, and references are now properly displayed.

2. The term on line 83 has been corrected to “demethylases.”
3. On line 86, “(PIN)” has been moved to precede “lesions” as suggested.

Round 2

Reviewer 1 Report

Comments and Suggestions for Authors

The manuscript has been thoroughly revised in accordance with all suggested comments. All suggested changes and clarifications have been carefully incorporated into the text, figures, and supplementary materials. I believe that the revised version satisfactorily addresses all concerns and significantly improves the quality and clarity of the paper.

Accordingly, I consider the manuscript now suitable for acceptance in its current form.

Reviewer 3 Report

Comments and Suggestions for Authors

Thank You for the corrections done - paper is now suitable for publication