Structural Characterization and Molecular Docking Screening of Most Potent 1,2,4-Triazine Sulfonamide Derivatives as Anti-Cancer Agents

Round 1

Reviewer 1 Report

The authors have chosen several 1,2,4-triazine sulfonamide derivatives and performed in silico evaluation of their anticancer properties. Density functional theory calculations, Molecular Docking, ADME calculations were carried out. Such purely theoretical evaluation of the chosen molecules might be interesting for some researchers as an initial information for development of novel molecules of the same class. However, the authors should improve the manuscript before it could be accepted for publication. Here are some suggestions:

 

-        The manuscript is lacking the discussion part. The authors present results, in some of the parts there is some discussion included, but, for example, ADME results are not discussed at all. The authors should analyse the results and include information about related work of other researchers.

-        The authors should clearly indicate the authorship of the compounds chosen for this evaluation. My suggestion would be to cite the suitable references in Figure 1 next to the names of the compounds. In these names N should be in italics as symbols of chemical elements in the names of organic compounds are written in italics.

-        The numbering of the compounds chosen for evaluation is very strange. The standard numbering used in organic chemistry is by using Arabic numbers. Here authors have chosen lower case letters. There is no need at all to enclose those letters into quotation marks. The same would apply to the numbers. In line 126 the authors use numbers while referring to the compounds.

-        Figure 10 contains ten molecules, however in several places the authors refer to twelve compounds evaluated, for example, line 35.

-        1,2,4-Triazine is written without spaces after commas, the name of the chemical compound should start from the lower case letter in the middle of the sentence.

-        The authors' contributions and references are not written according to the journal requirements and should be corrected.

-        The authors should check the English language of the manuscript for grammar and style mistakes as well as to avoid such phrases as „is an essential part of how living things function“ (line 54).

Author Response

Reviewer 1

The authors have chosen several 1,2,4-triazine sulfonamide derivatives and performed in silico evaluation of their anticancer properties. Density functional theory calculations, Molecular Docking, ADME calculations were carried out. Such purely theoretical evaluation of the chosen molecules might be interesting for some researchers as an initial information for development of novel molecules of the same class. However, the authors should improve the manuscript before it could be accepted for publication. Here are some suggestions:

 -        The manuscript is lacking the discussion part. The authors present results, in some of the parts there is some discussion included, but, for example, ADME results are not discussed at all. The authors should analyse the results and include information about related work of other researchers.

Response: Respected reviewer, we have added details about ADME results and compared it with the standard values.

All compounds were evaluated for their drug-likeness using the Lipinski's rule of five, which takes into account their molecular weights (MW), lipophilicity (log P), number of hydrogen bond acceptors (HBA), and number of hydrogen bond donors (HBD). Ten compounds showed no breaches of Lipinski's rule, while the remaining. It is also said that substances are more likely to be ingested if they meet Jorgensen's rule of three (QPlogS > -5.7, QPPCaco > 22 nm/s, and # Primary Metabolites 7) with fewer infractions. Jorgensen's rule of three was found to be followed by the all compounds. The solubility and permeability of the medication, as well as interactions of the drug with transporters and metabolic enzymes in the gut wall, all have a role in absorption. The majority of the substances demonstrated high values for projected qualitative human oral absorption and 100% for expected human oral absorption, it was also demonstrated.

It is noteworthy that drug binding to plasma significantly reduces the amount of the drug in the blood circulation; hence, the less bound a medication is, the better its ability to disperse or cross cell membranes is. Almost all compounds are discovered in the required QPlogKhsa range of 1.5 to 1.5. (prediction of binding to human serum albumin). This indicates that the majority of the chemicals are probably free to circulate inside the bloodstream and hence reach the target region. The results of ADME demonstrated that practically all medications contain characteristics of other drugs.

-        The authors should clearly indicate the authorship of the compounds chosen for this evaluation. My suggestion would be to cite the suitable references in Figure 1 next to the names of the compounds. In these names N should be in italics as symbols of chemical elements in the names of organic compounds are written in italics.

Response: Response: Respected reviewer, the synthesis and kinase inhibitory activity of these ten compound were reported by Mariusz et. al. in European Journal of Medicinal Chemistry but they did not explore the structural characterization, and molecular docking screening of these compound against cancer proteins. So in this study we explored their computational studies and found them excellent to good candidates for cancer treatment. This study will open new directions for the researchers in this field. Respected reviewer, we have updated figure 1 in the manuscript. As the names and structures of the compounds have presented in the form of figure, so references numbers would be difficult to manage with EndNote in figure. Therefore we have presented all references in the caption of the figure 1.

-        The numbering of the compounds chosen for evaluation is very strange. The standard numbering used in organic chemistry is by using Arabic numbers. Here authors have chosen lower case letters. There is no need at all to enclose those letters into quotation marks. The same would apply to the numbers. In line 126 the authors use numbers while referring to the compounds.

Response: Respected reviewer, we have changed numbering the compounds from lower case letters to Arabic style in all figures and tables.

-        Figure 1 contains ten molecules, however in several places the authors refer to twelve compounds evaluated, for example, line 35.

Response: Dear reviewer, it was a typo mistake we have corrected it.

-        1,2,4-Triazine is written without spaces after commas, the name of the chemical compound should start from the lower case letter in the middle of the sentence.

Response: Respected reviewer, we have corrected this mistake in manuscript.

-        The authors' contributions and references are not written according to the journal requirements and should be corrected.

Response: Respected reviewer, we have updated authors' contributions and references list in the manuscript.

Author Contributions: Conceptualization, S.M. and M.W.; methodology, M.A.K.; software, A.M.N., M.A.A.O., and A.A.A. validation, B. H., A.S.A.; formal analysis, A.S.A., M.S.R.; investigation, S.M.; resources, S.M. and A.M.N.; writing—original draft preparation, S.M. and M.A.K.; writing—review and editing, A.M.N., visualization, M.S.R.; supervision, S.M. and M.A.K.; project administration, S.M. and M.A.K.;. All authors have read and agreed to the published version of the manuscript.

-        The authors should check the English language of the manuscript for grammar and style mistakes as well as to avoid such phrases as „is an essential part of how living things function“ (line 54).

Response: Respected reviewer, we have revised the whole manuscript carefully and tried to avoid any grammar or syntax errors. In addition, we have asked several colleagues who are skilled authors and reviewers of English language papers/journals to check the English of our manuscript. We believe that the language is now acceptable for publication.

Reviewer 2 Report

See attachment

Comments for author File: Comments.pdf

Author Response

Reviewer 2

This manuscript reports the structural characterization and molecular docking study of ten 1.2.4-triazine sulfonamides. The information presented in the manuscript is interesting. The docking study demonstrated that several compounds have higher binding affinity to the corresponding receptors than the reference drugs, however the authors did not provide/perform any real (wet) experiment to validate the results. Some data are missing in Figure 5-8. The writing of the manuscript needs improvement. There are many grammar errors and typo in the manuscript including the title. In addition, the current study seems beyond the scope of the journal Crystals, therefore, it is recommended for publication in another specialized journal after major revision. Some specific comments and suggestions are as follows:

• The abstract needs to be written concisely. In addition, it is almost identical to the Conclusion.

Response: Respected reviewer, we have updated abstract and conclusion the manuscript.

Abstract: One of the biggest problems facing contemporary medicine is cancer. New approaches to therapy are required due to the difficult and prolonged treatment, the numerous adverse properties of the medications employed, and the developing confrontation of neoplastic cells to treatment. Ten 1, 2, 4-triazine sulfonamide derivatives (1–10) were chosen for the first time in the current work, and their chemical structures were examined by DFT studies. The in silico flexible docking analysis to the chosen receptors involved in cancer development and metastasis (3RHK, 5GTY, 6PL2 and 7JXH) revealed that the selected compounds are the most promising. The binding affinity of compounds 10, 2, 6, and 4 is much better than the standard drugs, Erlotinib, whereas compounds 9, 3, 1, and 7 showed better affinities as compared to standard drugs Neratinib, and Tepotinib in case of 3RHK receptor. The binding affinity against 5GTY receptor of compounds 10, 5 and 3 is much better than the standard drug Tepotinib, and compounds 7, 6, 2, 4, 1, 8 and 9 showed better than Erlonitib and Neratinib. The binding affinity against 6PL2 receptor of compounds 8, 3, 5, 4, 9 and 1 are much better than the standard drugs Tepotinib. Compounds 10, 6, 7 and 2 were better than Erlotinib and Neratinib. All selected drugs showed better binding affinities than the standard anticancer drug Neratinib in case 7JXH receptor, whereas compounds 2, 10, 5, 9 and 8 are better than Erlotinib. In silico ADME experiments supported the identified compounds' drug similarity. According to the MEP calculations, chemicals 3 through 10 can interact non-covalently. The interactions might take the form of σ- and π-hole interactions. Softest compound 4 has the smallest energy gap, with an E-gap value of 3.25 Ev. Compound 4 has the largest energy gap, at 3.41 eV. Compound 5 superior electron donor has the highest HOMO energy (6.5470 eV for HOMO). Compound 2 has the lowest LUMO energy, which suggests that it would be the best electron acceptor (E_LUMO = 5.766364 eV).

4. CONCLUSION

1, 2, 4-triazine sulfonamide derivatives (1–10) were selected for their virtual screening and DFT calculations to evaluate cancer inhibition potential and results were compared against standard (Erlotinib, Neratinib, and Tepotinib) anticancer drugs. The in silico flexible docking analysis to the chosen receptors involved in cancer development and metastasis (3RHK, 5GTY, 6PL2 and 7JXH) revealed that the selected compounds are the most promising. The binding affinity of compounds 10, 2, 6, and 4 is much better than the standard drugs, Erlotinib, whereas compounds 9, 3, 1, and 7 showed better affinities as compared to standard drugs Neratinib, and Tepotinib in case of 3RHK receptor. The binding affinity against 5GTY receptor of compounds 10, 5 and 3 is much better than the standard drug Tepotinib, and compounds 7, 6, 2, 4, 1, 8 and 9 showed better than Erlonitib and Neratinib. The binding affinity against 6PL2 receptor of compounds 8, 3, 5, 4, 9 and 1 are much better than the standard drugs Tepotinib. Compounds 10, 6, 7 and 2 were better than Erlotinib and Neratinib. All selected drugs showed better binding affinities than the standard anticancer drug Neratinib in case 7JXH receptor, whereas compounds 2, 10, 5, 9 and 8 are better than Erlotinib. In silico ADME experiments supported the identified compounds' drug similarity. Chemicals 1 through 10 are capable of non-covalent interaction, according to the MEP calculations. Potential interactions include σ- and π-hole interactions. With an E-gap value of 3.25 Ev, the softest compound 6, has the smallest energy gap. The biggest energy gap, 3.41 eV, is found in compound 1. HOMO energy is higher in compound 4, a superior electron donor (-6.00 eV for HOMO). Since compound 5 has the lowest LUMO energy (ELUMO = -2.50 eV), it could be the best electron acceptor. The results suggest that the selected triazine sulfonamide motifs have excellent to good potential to be adopted as futures anticancer drugs.

 

• Page 2, line 45, The sentence “Several prescription drugs used in medicine include a heterocyclic core, indicating that this field of chemistry is still developing and offers a wide range of opportunities for the creation of novel drugs” does not read good. Many drugs containing a heterocyclic core have already approved for clinic use. Do the authors refer to recently approved drugs or drugs currently in clinical trial? Otherwise, why state “is still developing”?

Response: Respected reviewer, we have modified this line to make it clear for readers.

Several prescription drugs used in medicine include a heterocyclic core, indicating the importance of this field of chemistry and offer a wide range of opportunities for the creation of novel drugs.

• It is not clear why these ten compounds were selected for study. Do they have potent anticancer activity? The papers need to be cited if the result has been published. On page 2, line 98, the cited paper (Gornowicz, A., 2021) does not include those ten compounds.

Response: Respected reviewer, the synthesis and kinase inhibitory activity of these ten compound were reported by Mariusz et. al. in European Journal of Medicinal Chemistry but they did not explore the structural characterization, and molecular docking screening of these compound against cancer proteins. So in this study we explored their computational studies and found them excellent to good candidates for cancer treatment. This study will open new directions for the researchers in this field.

• Page 3, line 103, give full name of ADME.

Response: Respected reviewer, we have added full name of ADME in the manuscript.

• Page 3, the name of compound b in Figure 1 is duplicated.

Response: Respected reviewer, we have updated the compound numbering to Arabic style in the manuscript.

• In addition to the role of the four selected receptors in cancer development and metastasis, what is the other reason (e.g., drug target of the three reference drugs?)? Please clarify.

Response: Respected reviewer, Compounds 1 through 10 were docked after the docking process was validated, and their binding affinity was determined. Their performance was compared to the binding affinities of Erlotinib, Neratinib, and Tepotinib, which are recognized inhibitors of the cMet, EGFR, HER2, and hTrkA receptors.

• Please give name of the four receptors in addition to the PDB code which is easier for readers to understand the proteins for study.

Response: Respected reviewer, We have provided names of four receptors alongwith PDB IDs. cMet, EGFR, HER2 and hTrkA (PDB codes: 3RHK, 5GTY, 7JXH and 6PL2 respectively)

• In Figure 5-8, pictures of compound g. h, i, are missing. The figure legends also need correction/clarification.

Response: Respected reviewer, in Figure 5-8, pictures of only most potent compound have been presented.

• Page 5, line 168, change “Table 5” to “Table 1”.

Response: Respected reviewer, we have corrected this typo mistake.

• Page 5, change “atoms” to “compounds” in table title.

Response: Respected reviewer, Respected reviewer, we have corrected this typo mistake.

• Page 11, line 286 and 287, it is not clear in sentence “For Rule of Three, these parameters must have a numerical value of 0, and 3, for Rule of Five, they must have a value of 4, respectively (Demir 2020).” as well as the value of the two rules in Table 3. Does the value refer to the maximum violation allowed?

Response: Respected reviewer, Number of violations of Lipinski’s rule of five. The rules are: MW < 500, logP < 5, DHB ≤5, AHB ≤ 10, Positive PSA value. VRT: Number of violations of Jorgensen’s rule of three. The three rules are: logS > -5.7, PCaco > 22 nm/s, PM < 7

• One piece of important data to support the conclusion is missing. The author need provide proof (cite the biological results if published) or perform the real experiment to validate the docking results, at least in vitro cell proliferation inhibition assay is needed.

Response: Respected reviewer, synthesis and kinase inhibitory activity of these sulfonamide derivatives of pyrazolo[4,3-e][1,2,4]triazines has already been published.

Mojzych, M.; Šubertová, V.; Bielawska, A.; Bielawski, K.; Bazgier, V.; Berka, K.; Gucký, T.; Fornal, E.; Kryštof, V. Synthesis and kinase inhibitory activity of new sulfonamide derivatives of pyrazolo [4, 3-e][1, 2, 4] triazines. European Journal of Medicinal Chemistry 2014, 78, 217-224.

Round 2

Reviewer 1 Report

The authors have revised the manuscript. However, the compound name 1,2,4-triazine still needs to be corrected at there should be no spaces in between commas and numbers.

The English language needs revision. It is not correct to refer to the synthesized compounds as chemicals.

Author Response

Dear and respected,

Ms. Ploy Assavajamroon

Assistant Editor

E-Mail: [email protected]

Manuscript Ref. No.: MDPI: crystals-2314903

Article Type: Full-length article

Article Title: Structural Characterization, and Molecular Docking Screening of Most Potent 1,2,4-triazine Sulfonamide Derivatives as Anticancer Agents

Thank you very much for letting us revise our manuscript. No doubt the reviewer’s comments are valuable and very helpful for improving our research paper. We have studied the comments carefully and have tried our best to revise the manuscript, which we hope to meet with acceptance requirements. The reviewer’s comments have been yellow highlighted in the manuscript.

REVIEWER'S COMMENTS AND RESPONSES

Reviewer 1

 

Comments and Suggestions for Authors

1. The authors have revised the manuscript. However, the compound name 1,2,4-triazine still needs to be corrected at there should be no spaces in between commas and numbers.

Response: Respected reviewer, we have corrected this mistake in manuscript according the following published articles.

2022. Strzelecka, M., Glomb, T., Drąg-Zalesińska, M., Kulbacka, J., Szewczyk, A., Saczko, J., Kasperkiewicz-Wasilewska, P., Rembiałkowska, N., Wojtkowiak, K., Jezierska, A. and Świątek, P., 2022. Synthesis, Anticancer Activity and Molecular Docking Studies of Novel N-Mannich Bases of 1,3,4-Oxadiazole Based on 4, 6-Dimethylpyridine Scaffold. International Journal of Molecular Sciences, 23(19), p.11173.
2023. Cascioferro, S., Parrino, B., Spano, V., Carbone, A., Montalbano, A., Barraja, P., Diana, P. and Cirrincione, G., 2017. An overview on the recent developments of 1,2,4-triazine derivatives as anticancer compounds. European Journal of Medicinal Chemistry, 142, pp.328-375.
2024. El-Agrody, A.M., Abd El-Latif, M.S., El-Hady, N.A., Fakery, A.H. and Bedair, A.H., 2001. Heteroaromatization with 4-Hydroxycoumarin Part II: Synthesis of Some New Pyrano [2,3-d] pyrimidines,[1,2,4] triazolo [1, 5-c] pyrimidines and Pyrimido [1,6-b]-[1,2,4] triazine Derivatives. Molecules, 6(6), pp.519-527.

 

2. The English language needs revision. It is not correct to refer to the synthesized compounds as chemicals.

Response: Respected reviewer, the corresponding authors of this manuscript belong to the country where the official language is English and they have published dozens of research articles in international journals. Also, they have training and collaborations in countries where the first language is English. Besides this, we have revised the whole manuscript carefully and tried to avoid any grammar or syntax errors. In addition, we have asked several colleagues who are skilled authors and reviewers of English language papers/journals to check the English of our manuscript. We believe that the language is now acceptable for publication.

Author Response File: Author Response.docx

Reviewer 2 Report

See attachment

Comments for author File: Comments.pdf

Author Response

Dear and respected,

Ms. Ploy Assavajamroon

Assistant Editor

E-Mail: [email protected]

Manuscript Ref. No.: MDPI: crystals-2314903

Article Type: Full-length article

Article Title: Structural Characterization, and Molecular Docking Screeing of Most Potent 1,2,4-triazine Sulfonamide Derivatives as Anticancer Agents

Thank you very much for letting us revise our manuscript. No doubt the reviewer’s comments are valuable and very helpful for improving our research paper. We have studied the comments carefully and have tried our best to revise the manuscript, which we hope to meet with acceptance requirements. The reviewer’s comments have been yellow highlighted in the manuscript.

REVIEWER'S COMMENTS AND RESPONSES

Reviewer 2

Comments
The authors have revised the manuscript according to reviewer’s comments. However, the following comments need to be addressed before accepting for publication.

• Page 11, line 287, change “the Rules-of-Three and Rules-of-Five” to “the rule of three and rule of five”.

Response: Respected reviewer, we have corrected this mistake in manuscript.

Among all the estimated parameters, the rule of three and rule of five are the two most crucial parameters.

• Page 11, line 288-289, “For Rule of Three, these parameters must have a numerical value of 0, and 3, for Rule of Five, they must have a value of 4, respectively [34]”. This sentence reads awkward and is confusing. Please rewrite it. If you want to refer the value range, then the numerical number is 0-3 in rule of three and 0-4 in rule of five. If you intended to say the maximum value (parameters), the number should be 3 and 4 for rule of three and rule of five, respectively.

Response: Respected reviewer, we have corrected this mistake in manuscript.

For rule of three, these parameters must have a numerical values ranging from 0-3, and 0-4 in rule of five

• Page 11, line 295, add “<” between “Primary Metabolites” and “7”.

Response: Respected reviewer, we have corrected this mistake in manuscript.

(QPlogS > -5.7, QPPCaco > 22 nm/s, and # Primary Metabolites < 7)

• Table 3 needs improvement, e.g., change “title” to “Compd” on the first column. What is “QPlog Po/w”? What is the recommended value? Is it related to LogP? This reviewer suggests changing “Donor HB” and “accpt” to “HBD” and “HBA”, respectively because their full name have been mentioned in the text.

Response: Respected reviewer, we have corrected these mistakes in manuscript. QPlogPo/w = Predicted octanol/water partition coefficient: Recommended Values from –2.0–6.5.Yes it is related to LogP. We have also updated table 3.

• Table 3, how come the HBA value is a decimal number? In addition, some of numbers are greater than 10 which is a violation of Lipinski’s rule of five. However, the value on the right column of rule of five is 0. Lipinski’s rule of five suggests HBD<5 and HBA<10, why the authors use recommended values as “Donor HB (0.0-6.0), Accpt HB (2.0-20.0). Please explain/discuss this in the main text.

Response: Respected reviewer, we have taken these ranges from QikProp User Manual of Schrodinger Software. (http://gohom.win/ManualHom/Schrodinger/Schrodinger_2015-2_docs/qikprop/qikprop_user_manual.pdf )

donorHB/HBD  (0.0 – 6.0)

Estimated number of hydrogen bonds that would be donated by the solute to water molecules in an aqueous solution. Values are averages taken over a number of configurations, so they can be non-integer.

accptHB/HBA (2.0 – 20.0)

Estimated number of hydrogen bonds that would be accepted by the solute from water molecules in an aqueous solution. Values are averages taken over a number of configurations, so they can be non-integer.

• Table 3, apparently, the numbers on the columns of rule of three and rule of five refer to the violation of the rules. Please add a footnote below the table. This reviewer suggests deleting “Rule of Five (maximum is 4), Rule of Three (maximum is 3”. It will confuse the readers.

Response: Respected reviewer, we have modified footnote below table 3 as follows.

 Rule of five (0-4), Rule of three (0-3).

• Since the biological evaluation of the ten compounds has been published, is the finding of this manuscript in agreement with the published results? Please discuss this in the manuscript.

Response: Respected reviewer, we have chosen the compounds whose in vivo studies are reported against anticancer agents. According to the reported data in vivo studies compounds 3 and 5 are highly potent inhibitor and our in silico results also aligned with the reported data, among 3 and 5, compound 3 is potent against all four selected cancer proteins, whereas compound 5 shows potency against 5GTY, 6PL2 and 7JXH.

• The figure ligands of Figure 5-8 in current version were not clearly written. For example, which compounds correspond to (a)-(f)? What difference between the picture on left side and right side?

Response: Respected reviewer, we have modified figure ligands of Figure 5-8 as follows.

Figure 5: 3D molecular docking images (a, c, e) and 2D ligand interaction images (b, d, f) of 1,2,4-triazine sulfonamides derivatives (10 (a, b), 2 (c, d), 3 (e, f), and 3RHK).

Figure 6: 3D molecular docking images (a, c, e) and 2D ligand interaction images (b, d, f) of 1,2,4-triazine sulfonamides derivatives (10 (a, b), 5 (c, d), 3 (e, f), and 5GTY).

Figure 7: 3D molecular docking images (a, c, e) and 2D ligand interaction images (b, d, f) of 1,2,4-triazine sulfonamides derivatives (8 (a, b), 3 (c, d), 5 (e, f), and 6PL2).

Figure 8: 3D molecular docking images (a, c, e) and 2D ligand interaction images (b, d, f) of 1,2,4-triazine sulfonamides derivatives (2 (a, b), 10 (c, d), 5 (e, f), and 7JXH).

Author Response File: Author Response.docx