Pharmacokinetic Study of NADPH Oxidase Inhibitor Ewha-18278, a Pyrazole Derivative
, Kyung Min Kim 2, Kee-In Lee 3, Yun Soo Bae 4 and Hwa Jeong Lee 1,2,*Round 1
Reviewer 1 Report
Review comments to authors
This manuscript described the pharmacokinetics of Ewha-18278 after i.v. or oral administration of several formulation of Ewha-18278 to rats. I have major comments as follow:
The research question of the current study is ambiguous. Why did the authors compare the PK profiles between two formulations? I considered that the obtained results could not give the novelty of the study.
After i.v. administration of Ewha-18278, the plasma concentrations were gradually decreased because the compound were distributed to peripheral tissue and eliminated; however, that mean levels at 10 h was higher than at 6 h. Why were the levels increased?
The mean plasma concentrations data after i.v. administration in Figure 2 and 3 seems the same.
In figure 2, the error bar at 0.25 h (i.v. data) is too high. Why?
Blood sampling volume is unknown. Please describe it.
The analytical methods using LC-UV should be described in more detail.
I hope these comments would be valuable for your manuscript.
Author Response
Thank you for your comments and suggestions for our manuscript. We have taken your critiques into account for revising the manuscript. It has been led to strengthen the manuscript overall. Below we provided the point-to-point responses to your comments. In revised manuscript, our responses are marked as yellow highlight.
Point 1: The research question of the current study is ambiguous. Why did the authors compare the PK profiles between two formulations? I considered that the obtained results could not give the novelty of the study.
Response 1: The purposes of this study were to obtain pharmacokinetic (PK) parameters of Ewha-18278 in animals, as part of the preclinical process of drug development and to find optimal formulation of the compound applicable to humans. We obtained PK parameters of Ewha-18278 in DMSO-based formulation suitable for animals and in diazepam injection-based formulation applicable to humans. Then, we compared PK parameters between two formulations and found no significant differences in PK parameters between two formulations. Thus, we could conclude that the diazepam injection-based formulation of Ewha-18278 is a suitable candidate for dosage development for future toxicity test and clinical trial.
In some cases, different drug formulation provided significant changes in PK parameters because surfactants and solvents in the formulation can affect permeability of the compound in body organs. In our study, we confirmed that there were no significant differences in PK parameters of Ewha-18278 between both formulations and absolute BA was > 40%.
Point 2: After i.v. administration of Ewha-18278, the plasma concentrations were gradually decreased because the compound were distributed to peripheral tissue and eliminated; however, that mean levels at 10 h was higher than at 6 h. Why were the levels increased?
Response 2: The mean plasma level at 10 h was lower than that at 6 h when the number of animals increased to 5~6 in the PK studies. Please find the revised Table 1, Figures 2 and 3.
Point 3: The mean plasma concentrations data after i.v. administration in Figure 2 and 3 seems the same.
Response 3: No, the mean plasma concentration-time profile following IV injection were different between two formulations. Please find the revised Figures 2 and 3.
Point 4: In figure 2, the error bar at 0.25 h (i.v. data) is too high. Why?
Response 4: When the number of animals increased to 6 in the PK studies, the error bar at 0.5 h was dramatically decreased in Figure 2. Please find the revised Figure 2.
Point 5: Blood sampling volume is unknown. Please describe it.
Response 5: As followed by reviewer’s comments, we described the blood sampling volume (0.2 mL) at line 121.
Point 6: The analytical methods using LC-UV should be described in more detail.
Response 6: As followed by reviewer’s comments, we added more information on HPLC-UV analytical method in the method section.
Thank you for your consideration of this work.
Sincerely,
Hwa Jeong Lee, Ph.D.
Professor
Reviewer 2 Report
The manuscript reports a pharmacokinetics study of a NADPH oxidase inhibitor - Ewha 18278 designed for treating Osteoporosis. In this study, authors present in-vivo pharmacokinetics study parameters by administering the drug in two different formulations in rat model. Samples were analysed using a validated HPLC UV method. Author solved the issue of chemical instability with the lead compound and oral formulation of the optimized compound has > 40% bioavailability showing potential for further studies.
Author Response
Thank you for your comments and suggestions for our manuscript. We have taken your critiques into account for revising the manuscript. It has been led to strengthen the manuscript overall. Below we provided the point-to-point responses to your comments.
Point 1: The manuscript reports a pharmacokinetics study of a NADPH oxidase inhibitor - Ewha 18278 designed for treating Osteoporosis. In this study, authors present in-vivo pharmacokinetics study parameters by administering the drug in two different formulations in rat model. Samples were analyzed using a validated HPLC UV method. Author solved the issue of chemical instability with the lead compound and oral formulation of the optimized compound has > 40% bioavailability showing potential for further studies.
Response 1: We appreciate that you describe our research purpose exactly in your comments.
Thank you for your consideration of this work.
Sincerely,
Hwa Jeong Lee, Ph.D.
Professor
Reviewer 3 Report
The manuscript is interesting.
I have only minor comments.
1.The manuscript would benefit from inclusion of introducing/bridging sentences between the individual parts of the "Results" that explain the logical order and rationale for the experiments
Was the formulation toxic? Please indicate the DE504.The number of animals used per group should be at least n = 6
Author Response
Thank you for your comments on our manuscript. We have taken your critiques into account for revising the manuscript. It has been led to strengthen the manuscript overall. Below we provided the point-to-point responses to your comments. In the revised manuscript, our responses are marked as sky blue highlights.
Point 1: The manuscript would benefit from inclusion of introducing/bridging sentences between the individual parts of the "Results" that explain the logical order and rationale for the experiments.
Response 1: As followed by reviewer’s comments, we added or modified the introducing/bridging sentences between each result part to provide better understanding on our study. Please find the highlighted lines (150-151, 171-172, 178-181 and 184-185).
Point 2: Was the formulation toxic?
Response 2: No, both formulations were not toxic. We dissolved several lead compounds in DMSO-based formulation for animal PK studies. Diazepam dissolved in diazepam injection-based formulation is clinically applied. We found that both formulations themselves were not toxic when they applied to animals without compound.
Point 3: Please indicate the ED50.
Response 3: From the previous report (Sci. Rep. 2016, 6, 22389), the Ki doses of Ewha-18278 specific to NOX1, NOX2 and NOX4 on ROS production were introduced as 1.08, 0.57 and 0.63 µM, respectively (Fig. 1). Moreover, the working doses of the NOX inhibitor were 10-20 µM for in vitro study and 10-20 mg/kg for in vivo study. We mentioned the effective dose of NOX inhibitor used in vivo efficacy study at the second paragraph of ‘Introduction’, at the lines 48-51.
Point 4: The number of animals used per group should be at least n = 6.
Response 4: As followed by reviewer’s comments, the number of animals increased to 5-6 (Table 1, Figures 2 and 3). The number of animals above 5 could be acceptable for the study.
Thank you for your consideration of this work.
Sincerely,
Hwa Jeong Lee, Ph.D.
Professor
Round 2
Reviewer 1 Report
I considered that the revised version has been much improved.
