A Graphene Oxide-Angiogenin Theranostic Nanoplatform for the Therapeutic Targeting of Angiogenic Processes: The Effect of Copper-Supplemented Medium

Round 1

Reviewer 1 Report

In this paper, authors report a new GO@ANG nanohybrid prepared by assembling GO with a fluorescent derivative of ANG (ANGF488) obtained by conjugating the primary amines of the protein with the reactive dye TFP. 

First of all, the physicochemical characterization carried out by UV-visible and fluorescence confirm the effective immobilization of the protein by physisorption on bwGO and GO120s nanosheets. 

Then the theranostic capacity of the GO@ANG nanohybrid was tested on prostate cancer cells (PC-3 line). The GO@ANG nanotoxicity was assessed, both in the absence and in the presence of copper ions, by in vitro cellular experiments on PC-3 cell line with the MTT colorimetric assay. Laser confocal microscopy (LSM) cell imaging evidenced an enhanced internationalization of GO@ANG than bare GO nanosheets, as well as significant changes compared to free ANG upon analysis of cell cytoskeleton organization and mitochondrial staining.

This is a careful work with very encouraging results for the treatment of prostate cancer. I strongly recommend this paper for publication.

Author Response

We thank the reviewer for the comments.

Reviewer 2 Report

I have reviewed the paper by Riela et al. The paper relates to the utilization of graphene oxide as anti-angiogenic effects a therapeutic agents. The work is well done and the case is well presented and convincing. I find the work done by the authors is significant and highly relevant to the readers of "Inorganics". Barring minor grammatical errors, I recommend publication of the manuscript.

Author Response

We thank the reviewer for their comment. The text has been carefully scanned and corrected for grammatical errors.

Reviewer 3 Report

The manuscript by Riela et al. investigates the bioconjugation of graphene oxide with angiogenin protein aiming to modulate angiogenic processes in cancer therapies. The manuscript is clearly and soundly written, and I could not find any major problems. However, I would like to make the following two comments:

1. The prepared bioconjugates were prepared by labeling with the fluorophore Alexa fluor 488, which makes the complex detectable by LSM. I think it is worth mentioning that there are localization strategies that do not require labeling, using label-free confocal Raman imaging. See, e.g., https://doi.org/10.1039/C8AN00225H.

2. Various strategies have been adopted to bioconjugate graphene oxide. Therefore, I believe that the review section of the introduction can be enriched by including the following paper - https://doi.org/10.3390/cancers11060753 - where a different strategy for bioconjugation of graphene oxide is presented.

The overall impression is that the work is suitable for publication after a minor revision.

Author Response

We thank the reviewer for their comment. The proposed references have been added to the revised manuscript