Date of Award
Doctor of Philosophy
Anumantha G. Kanthasamy
Parkinson's disease (PD) is a chronic neurodegenerative disorder that affects about 1% of people over the age of 60. PD is characterized pathologically by the progressive loss of dopaminergic neurons and the presence of Lewy bodies in the substantia nigra, dramatic depletion of dopamine in striatum, and activation of glial cells. Although the etiology of PD is not fully understood, evidence points strongly to the involvement of oxidative stress and mitochondrial dysfunction resulting from impaired mitochondrial biogenesis. Therefore, identifying cell signaling mechanisms regulating mitochondrial biogenesis is critically important to the development of new treatment strategies for PD. We have recently shown that activation of protein kinase D1 (PKD1) is neuroprotective and that positive modulation of PKD1 protects against neuronal cell death in cell culture models of PD. The goals of my Ph.D. thesis work were to understand the molecular mechanisms of PKD1-mediated neuroprotection using cell culture and animal models of PD and to test the efficacy of positive modulators of PKD1 signaling as potential neuroprotective agents in preclinical models of PD. Herein, we demonstrate for the first time that PKD1 activation positively regulates PGC-1α transcriptional activity. Overexpression of constitutively active PKD1 increased PGC-1α promoter activity, mRNA and protein expression in MN9D dopaminergic neuronal cells. Moreover, treatment of MN9D cells with rationally designed PKD1 activator peptide (AKP4T) enhanced expression of PGC-1α and other markers of mitochondrial biogenesis. Interestingly, treatment of cells with a PKD1 inhibitor (kbNB-14270) strongly suppressed mRNA expression of PGC-1α and TFAM. Importantly, AKP4T treatment protected against dopaminergic neurotoxicity in human dopaminergic neurons. Next, we adopted a rationale-based pharmacological screening approach to identify natural compounds that activate PKD1 and found that quercetin effectively activated PKD1-mediated neuroprotective signaling and promoted mitochondrial biogenesis and bioenergetics capacity in dopaminergic cells. Furthermore, quercetin protected against dopaminergic neurodegeneration by reversing striatal dopamine depletion, TH neuronal cell loss and behavioral deficits in the MitoPark transgenic mouse model of PD. Finally, we synthesized Mito-Met, a mitochondria-targeted analog of the anti-diabetic drug metformin. We demonstrate that Mito-Met induces activation of pro-survival PKD1 kinase more potently than its parent compound metformin in dopaminergic neuronal cells. Interestingly, Mito-Met stimulated mitochondrial biogenesis and bioenergetics capacity and also protected against MPP+-induced neurotoxicity. More importantly, Mito-Met treatment significantly reduced motor deficits and striatal dopamine depletion in MitoPark mice. Collectively, the research described herein provides evidence that PKD1 activation promotes mitochondrial biogenesis in dopaminergic neurons. We also show that positive modulation of PKD1 signaling affords neuroprotection against dopaminergic neurodegeneration in preclinical models of PD. Our results suggest that PKD1 is a promising druggable target that merits further preclinical investigations for the treatment of PD.
Ay, Muhammet, "Translational drug discovery approaches targeting the PKD1 signaling in Parkinson's disease" (2016). Graduate Theses and Dissertations. 15113.
Available for download on Thursday, March 29, 2018