The Journey of Gene Therapy in Sickle Cell Disease: How Molecular Advances Meet Clinical Care

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

Overall this is a very substantial review article that is very thoroughly researched review on sickle cell disease gene therapy that spans the entire translational spectrum from basic science to clinical practice. 

There are no major problems. 

Minor issues:

1. In Section 3.2.1 and 3.2.2, it would be helpful to mention newer CRISPR/Cas9 based therapies such as base editing and prime editing.  An adenine base editor has already been in clinical trials for other conditions. 

2. In Section 3.2.5 the discussion of Biological and Safety Challenges of genome editing, it may be worthwhile to emphasize that the approved gene edited product, exa-cel (Casgevy) does not rely on HDR, but rather involves the introduction of indels.  This can still lead to genomic instability, but the absence of and HDR template most likely will lessen some forms of off-target insertion.

3. In Section 3.2.5, the base editing and prime editing technologies do not entail full DSBs and so may avoid some toxicities. 

4. In section 3.3.2, the authors are correct about the use of AAV for delivery of HDR templates, but in the third paragraph, when describing the long-term genomic safety, it may be worthwhile to cite the literature which indicates that when AAV vectors with ITRs flanking the homology arms may be inserted with the ITRs included rather than by homologous recombination within the homology arm sequences.  This would both introduce extraneous sequences at the intended site of correction, but could have unknown effects.   In addition, if AAV is also used to introduce the Cas9, the Cas9 may also be inserted instead of the HDR template.

Author Response

Overall this is a very substantial review article that is very thoroughly researched review on sickle cell disease gene therapy that spans the entire translational spectrum from basic science to clinical practice. 

There are no major problems. 

We thank the reviewer for the overall comment.

Minor issues:

1. In Section 3.2.1 and 3.2.2, it would be helpful to mention newer CRISPR/Cas9 based therapies such as base editing and prime editing.  An adenine base editor has already been in clinical trials for other conditions. 

We thank the reviewer for the suggestions. We have now mentioned these approaches in the two sections.

2. In Section 3.2.5 the discussion of Biological and Safety Challenges of genome editing, it may be worthwhile to emphasize that the approved gene edited product, exa-cel (Casgevy) does not rely on HDR, but rather involves the introduction of indels.  This can still lead to genomic instability, but the absence of and HDR template most likely will lessen some forms of off-target insertion.

 We have now clarified this point in the section 3.2.5.

 

3. In Section 3.2.5, the base editing and prime editing technologies do not entail full DSBs and so may avoid some toxicities. 

 We have now modified this point.

4. In section 3.3.2, the authors are correct about the use of AAV for delivery of HDR templates, but in the third paragraph, when describing the long-term genomic safety, it may be worthwhile to cite the literature which indicates that when AAV vectors with ITRs flanking the homology arms may be inserted with the ITRs included rather than by homologous recombination within the homology arm sequences.  This would both introduce extraneous sequences at the intended site of correction, but could have unknown effects.   In addition, if AAV is also used to introduce the Cas9, the Cas9 may also be inserted instead of the HDR template.

 We have now added this nuance and supporting literature.

 

 

Reviewer 2 Report

Comments and Suggestions for Authors

Tardif et al.

The journey of gene therapy in sickle cell disease: how molecular advances meet clinical care

This review of gene therapy for sickle cell disease could be viewed as one of many. However, this is a very comprehensive overview taking clinical aspects, patient perspectives and health economics into account, in addition to the usual discussion of approaches to gene therapy. It is commendable that the eligibility criteria and contraindications for autologous gene therapy in SCD are discussed (Table 1), and followed by a proposed clinical decision structure for curative therapy selection in SCD (Table 2). These aspects are often ignored.

Specific comments:

Figure 1: references are needed to support time line. To my knowledge, the first paper linking BCL11A to HbF was published in 2007 (ref 32 in the manuscript), not 1998 as the figure appears to suggest. Although HU treatment is not a gene therapy approach, it should be included as this is the standard for treatment of patients with SCD.

A table with clinical trials of gene therapy for patients with SCD is lacking. This would be a useful addition.

Author Response

This review of gene therapy for sickle cell disease could be viewed as one of many. However, this is a very comprehensive overview taking clinical aspects, patient perspectives and health economics into account, in addition to the usual discussion of approaches to gene therapy. It is commendable that the eligibility criteria and contraindications for autologous gene therapy in SCD are discussed (Table 1), and followed by a proposed clinical decision structure for curative therapy selection in SCD (Table 2). These aspects are often ignored.

 

We thank the reviewer for the overall comment.

 

Specific comments:

Figure 1: references are needed to support time line. To my knowledge, the first paper linking BCL11A to HbF was published in 2007 (ref 32 in the manuscript), not 1998 as the figure appears to suggest. Although HU treatment is not a gene therapy approach, it should be included as this is the standard for treatment of patients with SCD.

 

We thank the reviewer for having identified the typo and for the suggestion. We have now modified the figure 1 and provided references.

 

A table with clinical trials of gene therapy for patients with SCD is lacking. This would be a useful addition.

 

We have now added such table (Table 1).