Date of Award

Fall 2010

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Graduate Group

Pharmacology

First Advisor

Dr. Benoit I. Giasson

Abstract

Parkinson's disease (PD) is a debilitating and progressive neurodegenerative disorder that affects over 6 million people worldwide. Despite being the most common movement disorder in the U.S., there is still no effective treatment for halting the progression of disease. While generally considered a sporadic and idiopathic disorder, a number of mutations in genetic loci causal for PD have provided valuable insight into the etiology of disease. Mutations in the gene for leucine-rich repeat kinase 2 (LRRK2) are the single most common cause of both familial and sporadic forms of PD. LRRK2 is a large 2527‐amino acid protein with several distinct domains: leucine-rich repeats, Ras-like GTPase domain, C-terminal of ROC (COR) domain, serine/threonine kinase domain, and WD40 repeats; however the understanding of LRRK2 function or how its aberration may lead to disease is still rudimentary. In 2006, we identified and characterized 3 patients with the G2019S LRRK2 mutation; however this search was limited to a few sequenced exons. An expanded screen identified 2 new patients with LRRK2 mutation, and the clinical and neuropathological findings for all these patients are provided herein. A novel system to express and purify the full-length protein with active in-vitro kinase activity revealed that the most common disease causing alteration (G2019S) markedly increases kinase activity. This highlighted overactive kinase activity as a possible intervention point for its aberrant effects. Screening for molecular inhibitors of kinase activity identified several compounds (Gö6976, K252a, and staurosporine) that share a basic indolocarbazole structure, which act as potent inhibitors of LRRK2 at low nanomolar concentrations. Increased kinase activity in the absence of outside factors is unlikely to account for the pathogenicity of the G2019S mutation. A more careful analysis of LRRK2 kinase activity revealed that this mutation, relative to the wildtype and other pathogenic mutations, may act in a novel pathway leading to disease by disrupting LRRK2 sensitivity to manganese kinase inhibition. Furthermore, based on kinetic data, we propose a novel hypothesis that LRRK2 may act as a cellular sensor of manganese levels, and disruption of this function may contribute to disease.

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