PONE-D-21-03224

Targeting neovascularization and lactate-fueled respiration in model tumors: assessment of anti-cancer treatment efficacy through chick embryo chorioallantoic membrane assays

PLOS ONE

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PLOS ONE

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"TAG: Krebsliga Zürich and Marie-Louise von Muralt-Stiftung für Kleintiere

HA: Swisslife

MG: Stiftung für wissenschaftliche Forschung an der Universität Zürich

The funders had no role in study design, data collection and analysis, decision to

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Reviewers' comments:

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Comments to the Author

1. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Yes

Reviewer #2: Yes

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2. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

Reviewer #2: Yes

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Reviewer #1: Yes

Reviewer #2: Yes

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Reviewer #1: No

Reviewer #2: Yes

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5. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: In this paper Dr. Ademi et al. show their ability to evaluate the ovo chorioallantoic membrane (CAM) assay/tumor assay by simultaneously targeting tumor angiogenesis and hypoxic/glycolytic tumor cells. With this approach they assess tumor growth, measure blood flow, stain tissue hypoxia and quantify dissemination of human and canine cancer cells, demonstrating the suitability of the CAM/tumor approach as a pre-clinical alternative for low cost, time saving and animal-free experimental setup in cancer research.

Data are concordant with recent literature papers as regards the angiogenesis-related assays and animal-free experimental setup in an anti-cancer approach using canine cell lines.

Major Remarks:

• Since the paper is based on the use of CAM authors should explain better this assay as an extraembryonic membrane commonly used in vivo to study both new vessel formation and its inhibition in response to tissues, cells, or soluble factors (i.e. Ribatti D, Gualandris A, et al. J Vasc Res. 1997; Ribatti D, Vacca A et al. Curr Pharm Biotechnol. 2000; Ribatti D, Nico B, et al. Anat Rec. 2001).

Minor Remarks:

• In the Figure 5b, the specific primers used are lacking. In the same Figure, in the plots d and e the figure legends are missing. The authors should indicate this to make the figure understandable.

• In the Figure 6, the figure legends are missing. The authors should add them.

• In the Materials and Methods section the sources and relatives affiliations of several reagents are missing. The authors should complete the informations.

• Please remove the abbreviations that are not necessary and introduce the missing abbreviations.

• All figures layout issues should be resolved in order to submit the paper for publication.

Reviewer #2: In the study” Targeting neovascularization and lactate-fueled respiration in model tumors: assessment of anti-cancer treatment efficacy through chick embryo chorioallantoic membrane assays” the authors report the interesting application of antiangiogenic molecules, demonstrating the results with different types of dynamic experiments, which do not involve animal models. Although the study brings new insights into angiogenesis and its inhibition, it nevertheless needs some modifications.

Minor revision

The text should be revised according to the following points:

- Reduce the "Introduction" paragraph, focusing on the main aspects and the purpose of the study

- Discussion: the part between 304-311 lines should be eliminated or merged with the introduction

- Update the "References" section with more recent citations, if possible

- Insert an "Abbreviations" section

- Figure 3: make the stairs uniform to the same unit

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Reviewer #1: No

Reviewer #2: Yes: Dr Schiano Concetta

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Targeting neovascularization and respiration of tumor grafts grown on chick embryo chorioallantoic membranes

PONE-D-21-03224R1

Dear Dr. Gorr,

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,

Filomena de Nigris, M.D., Ph.D.

Academic Editor

PLOS ONE

Additional Editor Comments (optional):

The revision has addressed the issues previously raised by the reviewers and editor.

Reviewers' comments: