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Eline Willemse, et al. Neurogranin as Cerebrospinal Fluid Biomarker for Alzheimer Disease: An Assay Comparison Study. Clin Chem 2018;64:927-37.


Dr. Eline Willemse is a postdoctoral researcher at the VU University Medical Center in Amsterdam, The Netherlands.


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Bob Barrett:
This is a podcast from Clinical Chemistry, sponsored by the Department of Laboratory Medicine at Boston Children’s Hospital. I am Bob Barrett.

Dementia is a devastating disease with a huge global impact. Today, over 46 million people live with dementia worldwide. Alzheimer’s disease is the most common type of dementia. The disease process of Alzheimer’s in the brain starts approximately 20 years before the patient presents with clinical symptoms. Treatment of the disease in an early stage provides a huge opportunity to halt the disease before clinical symptoms are apparent.

It is possible that promising treatments for Alzheimer’s disease have failed thus far because treatment was started at a late stage of the disease process. So, early treatment could be beneficial, but how do you recognize a patient with Alzheimer’s disease if he or she does not present with clinical symptoms? This is a key question, and presents an enormous opportunity for laboratory test based on biological markers of ongoing brain pathology.

One example is measurement of neurogranin, a protein in cerebrospinal fluid that correlates with cognitive decline and is a potential novel biomarker for Alzheimer’s disease dementia. There are multiple antibody-based neurogranin assays, which all target different segments of the protein.

An original article in the June 2018 issue of Clinical Chemistry compared the analytical and the diagnostic performance of three commonly used neurogranin assays on the same cohort of patients.

For this podcast, we’re joined by Dr. Eline Willemse, the first author with this article. She is a postdoctoral researcher at the neurochemistry laboratory in the Department of Clinical Chemistry at the VU University Medical Center in Amsterdam, The Netherlands. Her work focuses on biomarkers for dementia. So, Dr. Willemse, let’s get basic. Why did you study neurogranin as a novel biomarker for Alzheimer’s disease?

Eline Willemse:
Well, for Alzheimer’s disease, now three biomarkers in the cerebrospinal fluid exist, which are able to detect Alzheimer’s disease in an early disease stage. But a few years ago, a novel biomarker candidate for Alzheimer’s disease has been discovered. This one is called neurogranin and it was found to have increased levels in the cerebrospinal fluid of Alzheimer’s disease patients compared to cognitively healthy persons.

The importance of neurogranin as a novel biomarker for Alzheimer’s disease is based on two reasons. First, neurogranin directly correlated with cognitive performance and that’s a new feature that the other biomarkers did not do. This is very promising as a response biomarker when treatments are tested in clinical trials.

Second, neurogranin seems to be higher in Alzheimer’s disease compared to other types of dementia, and the biomarker that can discriminate between different types of dementia is currently not available and patient management can highly differ between different types of dementia, therefore, certainty on the exact dementia diagnosis is very important.

In the past years, neurogranin in cerebrospinal fluid has been studied by several research groups which have proven that, first, neurogranin directly reflects cognitive worsening. And two, neurogranin concentrations are highest in Alzheimer’s disease compared to other types of dementia. And these studies were performed by different research groups that used different types of assays to measure neurogranin. They all used an immunoassay principle that they use different antibodies, reagents, and instruments.

As a result, the outcomes of these different studies are hard to compare among each other. Translation of results from one study to another is difficult as we don’t know whether they measure the exact same thing. For example, the exact same protein, as proteins can be truncated or clustered together, especially in pathological circumstances. We therefore asked all three research groups involved in this assay to participate in this comparison study for the novel biomarker neurogranin.

In this study, we focused on the details of which part of the neurogranin protein was measured by the three different assays and in what amounts. We also sent out the exact same set of cerebrospinal fluid samples from 108 individuals with dementia, so Alzheimer’s disease or other types of dementia, or controls, to the three different research groups to measure neurogranin with their assay to directly compare the results.

Bob Barrett:
And doctor, what do you feel was the most important finding of your study?

Eline Willemse:
Surprisingly, when taking a closer look on what part of the neurogranin protein was targeted by the three assays, we found that all three assays targeted a different part of the protein. One assay that was developed at Washington University in St. Louis, targeted the full protein. The second assay, developed at Gothenburg University in Sweden, targeted only the C-terminal parts of the neurogranin protein. And the third assay, that was developed by the assay development company, ADx NeuroSciences in Ghent in Belgium, targeted the C-terminal part of neurogranin with the truncation in a specific part of the protein. So, three assays targeting three different forms of neurogranin, so to say.

The surprising part is that all three assays measured the same pattern of neurogranin concentrations amongst the different dementia types and the controls. So, neurogranin concentrations were increased in Alzheimer’s disease patients in all assays and were lower in the controls and in the other types of dementia.

Bob Barrett:
Does this mean that neurogranin can be used to differentiate between Alzheimer’s disease patients and patients with other types of dementia?

Eline Willemse:
Well, not on individual basis, so not as diagnostic use. But on group level, we find differences and this was also confirmed in other studies.

Bob Barrett:
Why is it important to compare different assays that target the same protein? The results in the end were similar for all three assays, correct?

Eline Willemse:
That is correct. The pattern we observed from neurogranin concentrations in the clinical cohort is similar between types of dementia for all assays. Nevertheless, we made the important observation that different parts of neurogranin are targeted. In this cross-sectional study, we did not see an effect of this, although this could have an effect in other disease stages. For example, protein truncation of neurogranin could occur maybe even as part of the pathogenic process.

Truncation of protein is often observed in neurodegenerative diseases. So, it’s good that we know the exact part of the neurogranin proteins that the different assays measure as they might turn out to be different among types of dementia or they might change during the disease process. Longitudinal measurements will be necessary to detect these kinds of changes and direct assay comparison studies are an effective method to study these differences between assays.

Moreover, direct assay comparison studies address the problem of preanalytical variation, which is a hot topic in the field currently. So, pre-analytical variation means variation in biomarker concentration that is induced artificially during the so-called preanalytical phase.

Preanalytical phase, we mean everything that happens to a sample, in this case, the cerebrospinal fluid sample from withdrawal to measurement of the analyte. You could think of transport time to the laboratory, time delay to processing at the laboratory, the temperature at the laboratory, the storage time, and the temperature between processing and measurement of the analyte, pipetting errors, machine settings, small differences in composition of the assays, et cetera, et cetera.

So, as you can imagine, these circumstances are different in every laboratory and thus can introduce differences in results between research centers. To examine the influence of these effects on biomarker results, direct assay comparison studies are very effective.

Bob Barrett:
Well, finally, doctor, let’s look ahead. How did the results of this study move the field forward? Will we soon have neurogranin as a biomarker clinically?

Eline Willemse:
Well, the results we obtained help to translate results of the biomarker neurogranin between studies. Neurogranin is still in the research phases as a biomarker. It’s not yet ready for implementation in the clinic. More studies will be needed to define a reference range in a healthy population and in diseased populations.

We need to know all about this before we can take any steps to take the biomarker to the clinic. But our study showed that independent studies give similar results of neurogranin and that helps us to translate results from neurogranin studies that were performed in different research center and that are performed worldwide. And the outcome of our study is positive as the clinical outcomes of the measurements appeared very comparable amongst the three assays that we tested.

Bob Barrett:
Dr. Eline Willemse is a postdoctoral researcher in the neurochemistry laboratory in the Department of Clinical Chemistry at the VU University Medical Center in Amsterdam, The Netherlands. She has been our guest in this podcast from Clinical Chemistry. I’m Bob Barrett. Thanks for listening.