Objective/Rationale:
Many genetic disorders are caused by the loss of a functional protein which protects cells from disease. These disorders are difficult to treat with drugs because most drugs block the function of a particular protein. This project seeks to rescue the activity of a mutant protein important in Parkinson’s disease. We have made a surprising discovery that the enzyme PINK1 in its Parkinson’s mutant form can be made active if we provide it with a designed non-natural substrate drug.
Project Description:
We must now confirm our initial results by producing the protein PINK1 in mammalian cells and understand how our molecule is able to rescue the activity of PINK1 mutants. Once we complete these studies with purified proteins we will study PINK1 mutants in neurons and test whether we can achieve in the cell what we have shown outside of the cell. This may require additional chemical changes to the rescue drug and may require understanding how the molecule is recognized by cellular enzymes. We will study the process of neurons which die because they express the Parkinson’s associated PINK1 enzyme—and show that by adding our rescue drug to the neurons we can protect them from death.
Relevance to Diagnosis/Treatment of Parkinson’s Disease:
This project is very ambitious and risky, since there are no real precedents for treating this kind of genetic disorder with a drug. However if we are successful, I think it will open the door to a new therapy for PARK6 gene containing Parkinson’s patients. A chemical which can rescue an inactive enzyme required for neuronal survival would be a very exciting medical advance. The ability to genotype patients and provide them a custom drug regimen would be particularly helpful because it would allow only the patients with a strong likelihood of benefiting from the drug to be given the drug. I want to stress again that this is a very risky approach but we are excited about the opportunity to explore it vigorously.
Anticipated Outcome:
We hope to show that it is possible to activate the disease gene PARK6 (PINK1) to phosphorylate its cellular targets by addition of a small molecule drug. If we can achieve this rescue of the disease-causing gene, then we will be in a position to try and develop a drug based on this innovative strategy. the work is with both purified proteins and cells, but if these experiments work as we hope, we can begin to investigate animal studies in the next phase.
Final Outcome
We have been able to confirm our original finding that our small molecule can activate the function of a catalytically compromised Parkinson’s disease mutatation in PINK1. We did this by developing for the first time a robust expression system for PINK1. We were able to express PINK1 only by co-expressing its molecular
chaperone, TRAP1, and the correct truncation of the mitochondrial localization sequence. Using this purified protein, we performed biochemical analyses that confirmed our original observation that PD mutated PINK1 activity is activated when using our designed non-natural substrate drug. With this confirmation, we now feel
confident enough to move into attempting drug rescue of the anti-apoptotic activity of PINK1 in dopaminergic neurons. If these experiments go well, we will also attempt to move into studies utilizing pre-clinical models to attempt the unprecedented rescue of a catalytically compromised disease gene.