A pair of congenic mice for imaging of transplants by positron emission tomography using anti-transferrin receptor nanobodies

Reviewer #1 (Public review):

Summary:

The topic of nanobody-based PET imaging is important and holds great potential for real-world applications since nanobodies have many advantages over full sized immunoglobulins and small molecules.

Strengths:

The submitted manuscript contains quite a bit of interesting data from a collaborative team of well-respected researchers. The authors are to be congratulated for presenting results that may not have turned out the way they had hoped, and doing so in a transparent fashion.

Weaknesses:

However, the manuscript could be considered to be a collection of exploratory findings rather than a complete and mature scientific exposition. Most of the sample sizes were 3 per group, which is fine for exploratory work, but insufficient to draw strong statistically robust conclusions for definitive results.

Reviewer #2 (Public review):

Summary:

This is a strong and well-described study showing for the first time the use and publicly available resources to use a specific PET tracer to track proliferating transplanted cells in vivo, in a full murine immunecompetent environment.

In this study the authors described a previously developed set of VHH-based PET tracers to track transplants (cancer cells, embryo's) in a murine immune-competent environment.

Strengths:

Unique set of PET tracer and mouse strain to track transplanted cells in vivo without genetic modification of the transplanted cells. This is a unique asset, and a first-in-kind.

Weaknesses:

-some methodological aspects and controls are missing

-no clinical relevance?

Author response:

Reviewer #1 (Public review):

Summary:

The topic of nanobody-based PET imaging is important and holds great potential for real-world applications since nanobodies have many advantages over full sized immunoglobulins and small molecules.

Strengths:

The submitted manuscript contains quite a bit of interesting data from a collaborative team of well-respected researchers. The authors are to be congratulated for presenting results that may not have turned out the way they had hoped, and doing so in a transparent fashion.

Weaknesses:

However, the manuscript could be considered to be a collection of exploratory findings rather than a complete and mature scientific exposition. Most of the sample sizes were 3 per group, which is fine for exploratory work, but insufficient to draw strong statistically robust conclusions for definitive results.

We thank reviewer #1 for the review of our work. We appreciate reviewer’s #1 comment on our intent to publish our results in the most transparent fashion, which is the case. We would point out that due to the technical challenges and cost of generating all the different nanobody-radiometal tracer conjugates, we included 3 repeats per group, which is the minimum required to perform statistical comparisons. We plan to add additional controls to the manuscript that were not initially included to limit the length of the manuscript. These additional controls will lend more weight to our conclusions.

Reviewer #2 (Public review):

Summary:

This is a strong and well-described study showing for the first time the use and publicly available resources to use a specific PET tracer to track proliferating transplanted cells in vivo, in a full murine immunecompetent environment.

In this study the authors described a previously developed set of VHH-based PET tracers to track transplants (cancer cells, embryo's) in a murine immune-competent environment.

Strengths:

Unique set of PET tracer and mouse strain to track transplanted cells in vivo without genetic modification of the transplanted cells. This is a unique asset, and a first-in-kind.

Weaknesses:

- Some methodological aspects and controls are missing

- No clinical relevance?

We thank reviewer #2 for their review of our work. We support reviewer’s 2 view on the strength of being able to track transplanted cells in vivo without the need of any sort of manipulation of the transferred cells. We plan to add additional controls to the manuscript that were not initially included to limit the length of the manuscript. These additional controls will lend more weight to our conclusions. We emphasize that although no clear clinical applications immediately derive from our studies, this work still offers better-suited tools for pre-clinical studies that require the ability to track transplanted cells in in vivo . We will resubmit a revised version shortly.