A new modified gene therapy approach avoids a toxicity seen in some nonhuman primate studies using adeno-associated viral (AAV) vectors to treat neurological disorders. Penn Medicine researchers developed the technique, which uses a modified transgene with a microRNA target designed to reduce the level of transgene expression in the dorsal root ganglia (DRG). They report that the alteration lowered transgene expression in the DRG by more than 80 percent and reduced toxicity in primates, which is considered a major hurdle to gene therapy for central nervous system conditions. The Penn group’s findings were reported online this week in Science Translational Medicine.
“We believe it is a safe, straightforward way to ameliorate the safety of AAV therapy for the central nervous system,” said first author Juliette Hordeaux, DVM, PhD, senior director of Translational Research in Penn’s Gene Therapy Program. “This approach could be used to design other gene therapy vectors to repress transgene expression in the cell types that are affected by the toxicity and not others, which is critical, because you need the expression everywhere else to effectively treat the disorder.”
Transgene overexpression in the DRG is thought to cause axonal degeneration in spinal cord tracts and peripheral nerves. Although this side effect has only been seen in primates so far, it represents a major technical challenge to the field. There are already several gene therapy trials ongoing in CNS conditions such as spinal muscular atrophy and Parkinson’s, and many more potential CNS-related targets for gene therapy exist.
This side effect was first seen in nonhuman primate studies using AVV vectors to deliver corrected genes via the spinal cord fluid and intravenously. Those studies reported problems of axonal degeneration in some tracts of the spinal cord and peripheral nerves. The cause was traced back to the DRG, a cluster of neural cells on the outside of the spinal cord responsible for transmission of sensory messages.
For their studies, the Penn researchers first documented DRG toxicity in nonhuman primates, then they devised a way to overcome it. Though its asymptomatic in primates, the damage is clear under close study of CNS histopathology. It’s already established that damage to the DRG in humans can lead to the breakdown of axons responsible for delivering impulses from nerves to the brain. Numbness and weakness in limbs, among other side effects, follow suit. This observed toxicity prompted the U.S. Food and Drug Administration to recently place a partial hold on human trials administering a gene therapy vector into the spinal cord to treat spinal muscular atrophy, a genetic disease that severely weakens muscles and causes problems with movement.
To overcome this toxicity, the Penn researchers injected vectors with and without a microRNA target, first in mice and then primates. microRNA regulates gene expression and makes for an ideal target in the cells. They chose microRNA-183 because it is largely restricted to the neurons in the DRG.
They found that the unmodified AAV vectors resulted in robust delivery of the new gene into target tissue and toxicity in DRG neurons. Vectors with the miRNA target, on the other hand, reduced transgene expression significantly, as well as the toxicity of DRG neurons, without affecting transduction elsewhere in the primate’s brain, based on histological analyses of the specimens up to 90 days later. The researchers also examined whether the DRG toxicity was caused by an immune response. They carried out experiments that showed immunosuppressants and steroids were unsuccessful at alleviating the toxicity.
According to the authors, toxicity of DRGs is likely to occur in any gene therapy that relies on high doses of a vector or direct delivery of a vector into the spinal cord fluid.
“We were concerned about the DRG pathology that was observed in most of our [non human primate] NHP studies,” Wilson said. “This modified vector shows great promise to reduce DRG toxicity and should facilitate the development of safer AAV-based gene therapies for many CNS diseases.”