Delivery of Kinase-Modified LRRK2 to Dopaminergic Neurons Using High-Capacity Viral Vectors

Promising Outcomes of Original Grant:
Mutations in LRRK2 are the most common known cause of PD, and LRRK2 demonstrates kinase-dependent neurotoxicity in vitro but these results have not been confirmed in vivo. We seek to determine whether LRRK2 over-expression causes dopaminergic toxicity in the intact mouse brain, and whether this is kinase dependent. In year one, we created a panel of high-capacity viral vector systems to deliver full-length wild type and kinase modified LRRK2 (G2019S or D1994A) into neurons in the intact adult mouse brain, based on helper-virus free herpes simplex virus (HSV) amplicons and adenovirus (Ad) vectors.

Objectives for Supplemental Investigation:
In year two, we will stereotactically deliver viral vectors encoding wild-type or kinase-modified LRRK2 to the mouse brain and measure corresponding dopaminergic toxicity. In addition to overt cell loss, measurements of cell morphology, density of tyrosine-hydroxylase positive axons in the striatum, and biochemical measurements of alpha-synuclein, ubiquitin and caspase reactivity will be taken at various time points. Finally, we will continue to refine LRRK2 viral vectors with the goal of generating neuronal-specific vectors with high potency and low immunoreactivity.

Importance of This Research for the Development of a New PD Therapy
This work will potentially validate LRRK2 kinase activity as an appropriate target for disease therapeutics, create a new and robust model for PD-relevant neurodegeneration, and enhance the understanding of the relationship between LRRK2 and cell death.