Dual Magnetic Particle Imaging and Akaluc Bioluminescence Imaging for Tracking Cancer Cell Metastasis

Round 1

Reviewer 1 Report

In the manuscript entitled “Dual Magnetic Particle Imaging and Akaluc Bioluminescence,” the authors report an MPI and Akaluc BLI dual imaging platform for tracking metastatic cancer cells. The paper is interesting, and I recommend its publication after the authors address the following questions:

Comment 1: In figure 4 a colocalization between BLI and MPI signal has been shown, after 14 days signal using both techniques was observed in other parts of the body, suggesting metastasis. Could the authors explain why there is no overlap in the signal from sites other than the primary tumor?  

Comment 2: Figure 4: In the MPI channel, the area of the region from which the signal is coming has significantly increased from day 8 to day 14, is it because of an increase in tumor size? A similar increase is not noticed in the BLI channel.

Comment 3: Figure 1G showing total BLI signal for unlabeled (Black), and Synomag-D labeled (Red) cells, up to 1 week post-loading. For day 1, the changes are labeled as ns but it seems there are significant changes.

Comment 4: How do the authors suggest selective targeting of metastatic cells within the primary tumor using the developed platform can be achieved? It is important to image the metastatic potential of tumor cells before they get transported to other parts of the body.

Author Response

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Reviewer 2 Report

The authors presented an important study for cell tracking using magnetic nanoparticles. 

 

The authors showed the comparison of two SPIO candidates: Synomag-D and Vivotrax, but only showed the iron loading using Synomag-D (Fig. 2). It will be more convincing to compare the iron loading for Synomag-D and Vivotrax because the final signals of MPI not only depend on the SPIO magnetic property but also the amount of iron loaded into the cells. What is the zeta potential of those two SPIO? The charge of nanoparticles could be related to the iron loading efficiency. Additionally, the figure legend for the Fig. 2 needs to be corrected.

 

For the in vivo testing, the authors took images for MPI and BLI on different days (MPI: Day 1, 8, 14 but BLI: Day 0, 6, 13). Why not compare the signals on the same day? Since there were 4 mice per group, it is necessary to show all the images from all the mice so we could see the variabilities from different subjects. 

 

It is very interesting to investigate the secondary signals from the lymph nodes where the metastases occurred. It seems to me that the MPI signals in Fig. 5 were saturated. However, the MPI signals in Fig. 4 decreased over the time of the tumor growth. I would suggest that the authors could explain the reason. 

 

I noticed the authors used nude mice instead of BALB/c mice but the 4T1 cells can grow well in immunocompetent mice. Did the authors try to establish the tumor model in BALB/c using 4T1Br5 cells?

 

I would suggest the authors revising the manuscript to improve and cover a more complete story. 

Author Response

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Reviewer 3 Report

The article by Wiliams et al. is devoted to an important and relevant topic in bioimaging: a comparison of different imaging modalities in vivo, which is useful for both basic and applied research. In his manuscript, the authors describe bioluminescence and magnetic imaging of dual-labeled breast cancer cell growth and metastasis in nude mice. 

 

The manuscript is well-written, relevant, and attractive. The authors preemptively addressed the major concerns of the manuscript, but I have some minor concerns preventing me from endorsing its acceptance at the present stage. I would like to address issues with some panels, figure legends, and references.

 

The figures in the article are clear and concise but would likely benefit from certain improvements.

Authors should add descriptions of panels C-F to the legend for Figure 2 and use the same way of addressing panels as in other figure legends.

It would be better to increase the font size of the figure legend (mice numbering) on panels С and D of Figure 3.

 

Issues with references relate to their relevance. More than 50% of all references are to articles published more than 5 years ago – 30/57, and the third of all references are more than 10 years old (n = 20). Since bioimaging is a rapidly developing and changing area of research, I suggest updating or removing references to the old literature whenever possible. Below is a non-exhaustive list of references that need to be replaced or removed.

Ref. 2 is too old and irrelevant considering the present-day data.

Ref. 5-6 are too old to prove the emergent nature of the technology.

The same applies to references 22-24 and 30.

 

The statement in the Materials and Methods section 2.1 about the preferential metastasis site of the 4T1Br5 cells requires a reference.

The technique of killing cultured cells by heating in the Materials and Methods section 2.5 requires a reference to the protocol.

The comparison of Red-Fluc and Akaluc in the Discussion section would be better if their respective emission maxima were cited and/or the papers characterizing their spectral properties with their main substrates referenced

 

As a side note, it would be interesting to discuss why the bioluminescence signal increases with time in Fig. 1G.

Author Response

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Round 2

Reviewer 2 Report

The authors have reasonable explanations and responses to improve the manuscript. I would recommend to accept the manuscript for publication.