Gene therapy for aromatic L-amino acid decarboxylase deficiency by MR-guided direct delivery of AAV2-AADC to midbrain dopaminergic neurons

Aromatic L-amino acid decarboxylase (AADC) deficiency is a rare genetic disorder characterized by deficient synthesis of dopamine and serotonin. It presents in early infancy, and causes severe developmental disability and lifelong motor, behavioral, and autonomic symptoms including oculogyric crises (OGC), sleep disorder, and mood disturbance. We investigated the safety and efficacy of delivery of a viral vector expressing AADC (AAV2-hAADC) to the midbrain in children with AADC deficiency (ClinicalTrials.gov Identifier NCT02852213). Seven (7) children, aged 4–9 years underwent convection-enhanced delivery (CED) of AAV2-hAADC to the bilateral substantia nigra (SN) and ventral tegmental area (VTA) (total infusion volume: 80 µL per hemisphere) in 2 dose cohorts: 1.3 × 1011 vg (n = 3), and 4.2 × 1011 vg (n = 4). Primary aims were to demonstrate the safety of the procedure and document biomarker evidence of restoration of brain AADC activity. Secondary aims were to assess clinical improvement in symptoms and motor function. Direct bilateral infusion of AAV2-hAADC was safe, well-tolerated and achieved target coverage of 98% and 70% of the SN and VTA, respectively. Dopamine metabolism was increased in all subjects and FDOPA uptake was enhanced within the midbrain and the striatum. OGC resolved completely in 6 of 7 subjects by Month 3 post-surgery. Twelve (12) months after surgery, 6/7 subjects gained normal head control and 4/7 could sit independently. At 18 months, 2 subjects could walk with 2-hand support. Both the primary and secondary endpoints of the study were met. Midbrain gene delivery in children with AADC deficiency is feasible and safe, and leads to clinical improvements in symptoms and motor function.


Introduction
This is a good review of the available studies and of the rationale for this study.

Results
A clear explanation for the study findings.  Do the authors have an explanation for the persistent elevation of 3-OMD after the gene therapy (i.e., was the gene therapy effective but not enough that it ameliorated the effect of the AADC deficiency and suggests that 100% gene function is not needed for improvement in this population, perhaps reflecting the redundancy of the systems that protect as one gets older)?
Line 275-276 For the reader who does not know, consider explaining briefly (perhaps in the Discussion) why was there a transient worsening in irritability and sleep and appearance of dyskinesia after the gene delivery.

Discussion
Line 313-315 A question -did the families comment on which improvement was thought to have the greatest impact, motor or OGC/behavioral/sleep? In many clinical situations, improvement in sleep/behavior are very much appreciated by caregivers. If they made any comments, is it worthwhile to inform the reader?
Supplementary Table S1 -It may be worthwhile to put a comment at the bottom to inform the reader why data on Subjects 1, 6, 7 were not acquired.
Reviewer #4 (Remarks to the Author): The authors present a well-designed and well-executed study of targeted AAV2-AADC gene replacement for AADC deficiency. The evidence is convincing that midbrain delivery is associated with improvement in motor function and prevention of oculogyric crises. I have no major comments.
In terms of the previous reviewer's concerns regarding small sample size, and uncontrolled trial design, I agree these are limitations. However, in an ultra-rare disease such as AADC, I believe this is the most appropriate study design, and it standard for similar disorders. The authors clearly reference the robust natual history data that exists for this disorder, and serves as an appropriate control.

Introduction
This is a good review of the available studies and of the rationale for this study.

Results
A clear explanation for the study findings.
Line 193-194 Do the authors have an explanation for the persistent elevation of 3-OMD after the gene therapy (i.e., was the gene therapy effective but not enough that it ameliorated the effect of the AADC deficiency and suggests that 100% gene function is not needed for improvement in this population, perhaps reflecting the redundancy of the systems that protect as one gets older)?

The persistent elevation of 3-OMD would be consistent with residual deficiency of AADC function, and we agree that this point deserves additional comment, so have added the following 2 sentences to the Discussion of p. 15: "We did observe a persistent elevation of 3-OMD after gene delivery in all subjects, which is consistent with some degree of residual AADC deficiency. The marked clinical improvements we observed in our subjects therefore did not require complete restoration of AADC function, but the localization of that function, not only the extent, may be a key determinant of clinical outcomes."
Line 275-276 For the reader who does not know, consider explaining briefly (perhaps in the Discussion) why was there a transient worsening in irritability and sleep and appearance of dyskinesia after the gene delivery.

Discussion
Line 313-315 A question -did the families comment on which improvement was thought to have the greatest impact, motor or OGC/behavioral/sleep? In many clinical situations, improvement in sleep/behavior are very much appreciated by caregivers. If they made any comments, is it worthwhile to inform the reader?

Caregivers did mention to the study team that subjects' improvement in sleep and irritability positively impacted the family's sleep and quality of life. We have added the following sentence to the Discussion on p. 13: "Caregivers reported that these changes, particularly the resolution of OGC and improvements in sleep, had a major positive impact on the child's and family's quality of life."
Supplementary Table S1 -It may be worthwhile to put a comment at the bottom to inform the reader why data on Subjects 1, 6, 7 were not acquired.

Reviewer #4 (Remarks to the Author):
The authors present a well-designed and well-executed study of targeted AAV2-AADC gene replacement for AADC deficiency. The evidence is convincing that midbrain delivery is associated with improvement in motor function and prevention of oculogyric crises. I have no major comments.
In terms of the previous reviewer's concerns regarding small sample size, and uncontrolled trial design, I agree these are limitations. However, in an ultra-rare disease such as AADC, I believe this is the most appropriate study design, and it standard for similar disorders. The authors clearly reference the robust natual history data that exists for this disorder, and serves as an appropriate control.