Compensatory mechanisms of neuroprotection by PKD signaling against oxidative damage in experimental models of Parkinson\u27s disease (PD): Relevance to PD drug discovery strategies

Oxidative stress is a key pathophysiological mechanism contributing to the selective degeneration of dopaminergic neurons in Parkinson\u27s disease. Unraveling the molecular mechanisms underlying various stages of oxidative neuronal damage is critical to better understanding the diseases and developing new treatment modalities. In this study, we identified that protein kinase D1 (PKD1) functions as a key anti-apoptotic kinase to protect neuronal cells against early stages of oxidative stress. Blockade of PKCδ cleavage, PKCδ knockdown or overexpression of a cleavage-resistant PKCδ mutant effectively attenuated PKD1 activation, indicating that PKCδ proteolytic activation regulates PKD1 phosphorylation. We also identified that phosphorylation of S916 at the C-terminal is a preceding event required for PKD1 activation loop phosphorylation. Further PKD1 signal transduction was characterized in a pre-clinical model of Parkinson\u27s disease. Exposure of dopaminergic neuronal cells or primary mesencephalic neurons to MPP+ induced PKD1 activation. PKD1 was not activated in the nigral dopaminergic neurons of PKCδ knock-out (PKCδ -/-) mice exposed to acute MPTP treatment. Earlier we reported that Fyn kinase regulates PKCδ in dopaminergic cells. PKD1 was not activated in nigral dopaminergic neurons of Fyn knockout (Fyn -/-) mice exposed to acute MPTP treatment. Further, dopaminergic neurons co-treated with the PKD1 inhibitor kbNB 142-70 and exposed to MPP+ exacerbated neuronal death, confirming the survival role of PKD1. Having confirmed that positive modulation of PKD1 can be a novel neuroprotective strategy, we took a translational approach by developing PKD1 activator and characterizing the protective function in pre-clinical models of Parkinson\u27s disease. Peptides were rationally designed and screened for their ability to activate PKD1 using various screening methods. Peptide AK-P4 was identified to activate PKD1 specifically and protect against MPP+ and 6-OHDA in both N27 cells and primary mesencephalic neurons. Further, AK-P4 tagged with the TAT sequence (AK-P4T) delivered using intravenous injections activated PKD1 in mice. Co-treatment with AK-P4T restored the neurotransmitter levels and the behavioral and locomotor activities of the MPTP treated mice significantly. Overall, positive modulation of PKD1 suggests its promise as a potential therapeutic strategy in PD