Collaborations Help Drive Parkinson’s Disease Research

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Tamlyn Oliver: Hi everyone, welcome to Biocompare’s Tech Insights Podcast, where we speak to scientific experts about new tools and technology that can help advance your research. I’m Tamlyn Oliver, Managing Editor of Biocompare. Today’s guest is Julianna Tomlinson, Senior Research Associate at Ottawa Hospital Research Institute. She’s going to talk to us about Parkinson’s disease related research and novel reagents for studying α-Synuclein and Parkin biology. Thanks for joining us today, Julianna.

Julianna Tomlinson: Thank you so much and thank you for your interest in our work.

Tamlyn Oliver: You’re welcome. I have quite a few questions for you, so let’s get started. Despite considerable efforts, researchers have not yet developed effective measures to predict and stop the progression of Parkinson’s disease. What do you think is holding them back?

Julianna Tomlinson: I think that there are a few challenges, and I think some of those are that this disease has such an insidious slow onset and progression that by the time a person is diagnosed, the disease has progressed considerably, and critical nerve cells have been lost. Common late-onset sporadic-type Parkinson’s is also typically multifactorial in that it doesn’t have just one cause. So, both genetic and environmental factors play a role in its initiation and also not all Parkinson’s is the same. It’s a heterogenous syndrome. And so when you take all of that to be able to identify at risk persons in early stages of the disease, to stop its progression after diagnosis or to treat patients in advanced stages of illness, we really need to be able to understand where and how the disease starts, how it progresses and then ultimately how it leads to the death of vulnerable neurons. I’m hopeful that we’re getting closer though because the field is beginning to understand the underlying risk elements that conspire to generate typical Parkinson’s disease, and the underlying mechanisms that ultimately lead to neuronal death. Capturing these will inform our ability to predict onset and ultimately to modify the risk in still healthy individuals as is done for example, to delay heart failure, diabetes and stroke. Once we identify these pathological mechanisms, we’ll also be able to treat the diseases with cause directed therapies that are matched to specific individuals in a form of personalized medicine. Related to predictions, specially currently there is no one easy inexpensive blood test that captures all of the stages of the disease, such as who’s at risk, who has it, who progresses faster, et cetera. With new discoveries into the disease processes, supported by new tool development, I’m confident that we can get there.

Tamlyn Oliver: BioLegend’s novel immunoreagents for studying α-Synuclein and Parkin were developed by collaborating with leaders in the Parkinson’s disease field. Why is collaboration so important especially when it comes to Parkinson’s disease research?

Julianna Tomlinson: Our collaboration with BioLegend has been highly rewarding and I thank the individuals that have been working on this project both from our lab as well as the BioLegend team. The collaboration was made possible with support from the Michael J. Fox Foundation for which we’re very grateful and ultimately that support allowed our two teams to come together and each team had complimentary skills. The academics drove the hypothesis testing from the initial idea that there were different forms of Parkin in the human brain that the currently available antibodies on the market could not detect. BioLegend brought their expertise and resources in regard to tool development. Together, we screened and validated them in a multitude of assays, which for us lead to new discoveries in Parkin biology. It’s highly rewarding to see that two of the clones were ultimately brought to the market and made available to the research community.

Tamlyn Oliver: Alright, next question… BioLegend was first to market with antibodies able to specifically detect oxidized and aggregated Parkin. What was the rational behind developing these reagents and can you please explain how they’ve been used to advance Parkinson’s research?

Julianna Tomlinson: Yeah, so we’ve been interested in Parkin metabolism in the human brain for well over 10 years now in the lab, with a specific focus on its oxidation and what this could mean for its function in the aging human brain. Animal models of Parkin deficiency don’t recapitulate the human disease, and the current known function such as E3 ligase and its role in mitophagy don’t explain the selective vulnerability of cell loss observed in Parkinson disease as of yet. And so, we wanted to go back into the human brain, to study this protein with a renewed effort. Part of that effort was to be able to visualize the protein in the human brain where we suspected it was oxidized and to be able to purify these forms. And the challenge there has been that Parkin is notoriously hard to detect in human brain sections by microscopy. We hypothesize that this could be in part because it’s oxidized in such a way that the epitopes are masked. Our goal was to create antibodies that would detect these forms in the human brain and to be better able to study oxidized Parkin. Already, the antibodies proved to be valuable, in that they allowed us to see where Parkin is in human mid-brain sections and specifically, in the dopamine neurons. There we already found that Parkin was predominantly associated with neuromelanin and for us, this association was particularly exciting as we had been doing studies in parallel that showed that Parkin can participate in melanin formation in vitro, and it does this by donating its own cysteines to neutralize what would otherwise be toxic to dopamine metabolites. We’ve recently validated the specificity of these antibodies using human Parkin deficient mid-brain sections and now using these tools, we are in the process of using indirect immunofluorescence, and co-labeling to look at the localization of Parkin with other organelles. That being said, there’s still more work to be done in Parkin tool development. These antibodies appear to be specific for a form of oxidized or modified Parkin in human mid-brain sections as we don’t see predominant signal in dopaminergic neurons from younger individuals that we know express the protein. We also hope to be able to develop antibodies that specifically detect dopamine modified Parkin which proved to be difficult in our first attempts, and we’d like to be able to develop a sandwich ELISA for the screening and quantification of wild-type Parkin in blood and CSF.

Tamlyn Oliver: My final question for you…In addition to developing antibodies for studying α-Synuclein and Parkin biology, you’ve also developed novel α-Synuclein ELISA kit. Can you comment on some of the key features of this product and explain how it can benefit large studies, such as the Parkinson’s Progression Markers Initiative, or PPMI?

Julianna Tomlinson: Yeah, so the ELISA came out of work that was done in collaboration with Doctors Michael Schlossmacher, Brit Mollenhauer, Omar El-Agnaf and they worked together with Doctor Peggy Taylor, of BioLegend, who at the time it was Covance, and a subcontractor Epitomics, and again this project was supported by the Michael J. Fox Foundation. So Epitomics has pioneered a novel way to make monoclonal antibodies in animals other than mice, and the shared goal of the team was to create a human specific ELISA, that reliably worked on CSF plasma cells conditioned media and grain, thinking that this would be key to biomarker efforts, basic research and drug discovery. And I think that it’s fair to say that this project, that objective has been achieved. Together they worked out the frequent problem of matrix and analyte interactions in ELISA kits for amyloidogenic proteins and created a well-validated high throughput product that’s now available from BioLegend and is widely used in the field. I think that a standardized validated assay and then cross-validated by being used across multiple sites and by multiple teams is a strength in large studies such as the PPMI to reduce intra-study variability.

Tamlyn Oliver: Thanks Julianna, for providing an update on Parkinson’s disease research as well as providing some background and explaining the benefits of the some of the novel immunoreagents now available. For a more in-depth description of the novel reagents that Julianna has spoken about here, please read the related article “Novel Reagents for Studying α-Synuclein and Parkin Reveal New Insights into Parkinson’s Disease.” And thanks so much for listening. For more information on products, technologies and the latest scientific advancements, please visit Biocompare.com, and have a great day.