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Novel AAV Capsid, Truncated ChAT Promoter Address Challenges in Intracranial Gene Therapy

Key findings

  • The striatum plays a major role in movement and cognition, and transduction of striatal neurons is needed in gene therapies that target neurodegenerative diseases
  • This study addressed two problems with intra-striatal injection of adeno-associated virus (AAV) vectors: limited diffusion from the injection site and inefficient, nonspecific transgene expression in cholinergic neurons
  • Compared with the conventionally used AAV9, a novel engineered AAV capsid, AAV-S, showed greater transduction throughout the striatum after direct injection into mice
  • Combining AAV9 with a truncated human choline acetyltransferase (ChAT) promoter provided a seven-fold higher specificity of transduction of cholinergic neurons than the CAG promoter and a three-fold increase in the efficiency of transduction
  • The AAV-delivered ChAT promoter construct should be immediately useful to the neuroscience field, and the findings support further development of the AAV-S capsid for transduction of brain

Loss of striatal neuron circuits in the striatum is associated with several neurodegenerative diseases, including Parkinson's and Alzheimer's. Intracranial gene therapy is still investigational in the U.S., but the European Medicines Agency has approved Upstaza, a one-time gene therapy for severe aromatic L-amino acid decarboxylase deficiency that's injected directly into the striatum.

Adeno-associated virus (AAV) vectors are currently the most efficient option for intracranial gene therapies for neurodegenerative diseases. Massachusetts General Hospital researchers are working to increase AAV diffusion throughout the brain and increase transgene expression in cholinergic neurons, which regulate the activity of the most abundant striatal neurons and fine-tune a variety of brain functions.

Miguel C. Santoscoy, PhD, a research fellow in the Department of Neurology and Molecular Neurogenetics Unit at Mass General, Casey A. Maguire, PhD, a gene delivery expert and associate investigator in the Mass General Research Institute, and colleagues report two improvements to AAV-based intracranial gene therapy in Molecular Therapy: Methods & Clinical Development.

Comparison of AAV9 and AAV-S

In an earlier study, also published in Molecular Therapy: Methods & Clinical Development, Dr. Maguire and other colleagues engineered AAV-S, a derivative of AAV9 that showed much greater transduction of the brain than AAV9 after intra-cortical and intra-hippocampal injection. In this study, AAV-S transduced a greater area of striatum than AAV9 after direct intra-striatal injection in C57BL/6 mice.

Intra-striatal gene therapy in humans presently requires multiple burr holes and multiple injections to produce sufficient "coverage" of transduced cells in the striatum. A capsid that spreads a greater distance in the striatum would be highly desirable to reduce the number of injections and the potential for related complications.

AAV9 With ChAT Promoter

In an unpublished study, the researchers observed that after intracranial injection of AAV vectors using the broadly active CAG promoter, relatively few of the cells transduced were cholinergic neurons. To address that problem, the team engineered an AAV9 plasmid that contains a truncated human choline acetyltransferase (ChAT) promoter.

Intra-striatal injection into mice showed:

  • Specificity (number of transduced cholinergic neurons out of all transduced neurons)—64% for AAV9-ChAT vs. 8.9% for AAV-CAG (P=0.01)
  • Efficiency (number of transduced cholinergic neurons out of all cholinergic neurons)—42% vs. 14.2% (P=0.0551)

There were slight increases in the specificity and efficiency of cholinergic neuron transduction with AAV9-ChAT compared with AAV-S–ChAT, but the differences weren't statistically significant.

Immediate Utility

For applications in which broad transduction of striatal neurons is desired, the AAV-S capsid with the CAG promoter may be immediately useful. In applications in which high specificity and efficiency for striatal cholinergic neurons is desired, the AAV9 capsid and ChAT promoter may be the better choice.

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