PONE-D-20-13006

NHD2-15, a Novel Antagonist of Growth Factor Receptor-Bound Protein-2 (GRB2), Inhibits Leukemic Proliferation

PLOS ONE

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Reviewers' comments:

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Reviewer #1: Partly

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Reviewer #1: No

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Reviewer #1: No

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Reviewer #1: Yes

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5. Review Comments to the Author

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Reviewer #1: In this manuscript, Lewis and coworkers present a new small-molecule inhibitor of GRB2, a downstream effector of the important leukemia oncogene BCR-Abl1. The presented results are novel and interesting, the methodologies are scientifically sound. Overall I think the paper should be a good fit for PLOS ONE, but some of its conclusions/claims are weakly supported, therefore the authors should thoroughly revise it based on the following points.

1. The article seems to merge results from two independent studies: one for optimizing a novel synthetic reaction to yield a specific set of molecules, and one for testing some molecules against a specific oncotarget (GRB2). It is unclear how these studies were linked or what was the motivation to test the compounds against GRB2 in particular (or vice versa, to test these particular compounds against GRB2), even though the authors mention that NHD2-15 was "designed towards the SH2 domain of a specific protein". How was it designed? Did the authors establish a structural basis for its binding to the GRB2 SH2 domain (via ligand docking or similar computational methodologies)? Or were there similar compounds reported as SH2 inhibitors? Please elaborate on this in the manuscript.

2. Important details about statistical testing are left out. What statistical tests were applied to establish statistical significance of differences? Also, the authors claim that "the combination of NHD2-15 with imatinib significantly killed K562 cells better when compared to cells exposed to imatinib alone.", but on Figure 8, it is not reported whether these differences (1st striped column vs rest of striped columns) are significant or not.

3. Non-toxicity was established by testing the smallest effective concentration (15uM) in zebrafish. However, the Kd of NHD2-15 to GRB2 is significantly higher (119uM) and the reported data do not suggest that 15uM would be a sufficient therapeutic concentration in vivo. Combined with the fact that the other three (structurally related) compounds were toxic to zebrafish, I think that non-toxicity should be established at a higher concentration.

4. As per journal guidelines on data availability, the raw data should be published in a tabular format as supplementary data.

Minor:

- Figure 2 x axis title is confusing, as the native peptide is not an antagonist of GRB2, please revise.

- Figure 6 takes up space unnecessarily: it would be enough to state that none of the zebrafish were killed under the specific conditions.

- Results/Library synthesis section: "Finally, deprotonation of 2 with sodium hydride..." I believe it is the beta-diketone that is deprotonated, please double-check this sentence.

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Reviewer #1: No

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NHD2-15, a Novel Antagonist of Growth Factor Receptor-Bound Protein-2 (GRB2), Inhibits Leukemic Proliferation

PONE-D-20-13006R1

Dear Dr. Stachura,

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.

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Kind regards,

Laszlo Buday

Academic Editor

PLOS ONE

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