Tracer molecules can distinguish between very similar brain diseases

Two diseases that affect the brain, Parkinson’s disease and multiple system atrophy, show the same characteristics: proteins form toxic aggregates. Scientists have now shown that tracer molecules developed at LiU can distinguish between these diseases. They hope that the discovery will lead to earlier molecular diagnosis.

Aggregat av alfa-synuklein i celler i mänsklig hjärnvävnad som färgats in med spårarmolekyl (grön) och antikropp mot alfa-synuklein (röd). Den turkosa eller blå färgen kommer av ett annat ämne (lipofuscin) som ansamlas i åldrande hjärnceller, men som inte har någon betydelse för studiernas resultat.Aggregates of alpha-synuclein in cells from human brain tissue stained with tracer molecules (green) and antibodies against alpha-synuclein (red). The turquoise or blue colour is from another substance (lipofuscin) that increases as the brain ages. It is not, however, significant for the study results. Photo credit Therese Klingstedt

A doctor faced with a person who has problems with tremors, difficulty moving and difficulty speaking finds it very difficult to make a diagnosis. These symptoms can be caused both by Parkinson’s disease and another, more rare, disease – multiple system atrophy (MSA). Both diseases are the result of gradual degeneration of nerves in increasing volumes of the brain. And it is the same protein in both diseases, alpha-synuclein, that is wrongly folded and forms aggregates. MSA can initially be mistaken for Parkinson’s disease, but its progression is more rapid and follows a different pathway. The two diseases may also require different treatments. There is currently only one way to distinguish the diseases with certainty: to wait and see how the symptoms develop.

Scientists have now developed a method to distinguish between Parkinson’s disease and MSA at an early stage. Tracer molecules developed by researchers at LiU play an important role. In collaboration with Claudio Soto’s research group at the University of Texas Health Science Center at Houston (UTHealth), Peter Nilsson and his research group have investigated the aggregates of wrongly folded alpha-synuclein that characterise both diseases.

Molecular differences

In the study, which has recently been published in Nature, the American scientists took samples from the cerebrospinal fluid of patients who had either Parkinson’s disease or MSA. Since the concentration of wrongly folded alpha-synuclein is very low in cerebrospinal fluid, the researchers used a method that they had developed to amplify the signal, like a molecular Xerox machine. They mix a small amount of protein from a patient sample with the normal form of alpha-synuclein. The wrongly folded protein from the patient then converts the normal alpha-synuclein such that it assumes the same, wrongly folded, structure.

The researchers then used special tracer molecules, developed by Peter Nilsson and his colleagues, to investigate whether it is possible to track the aggregates of damaged protein. The tracer molecules fluoresce with different colours, depending on the exact shape of the protein they are bound to. The researchers saw that the colour emitted by the tracer molecules differed between the protein aggregates in samples from people with Parkinson’s disease and those from people with MSA.Peter Nilsson, professor i organisk kemi.Peter Nilsson Photo credit Karin Söderlund Leifler

“This is the first time that we have been involved and shown that our tracer molecules work with samples from the cerebrospinal fluid from patients. Our American colleagues have also shown in the study that even though both types of protein aggregate contain alpha-synuclein, they have different structures and differ between the two diseases”, says Peter Nilsson, professor in the Department of Physics, Chemistry and Biology.

The American researchers then used cryo-electron microscopy to examine the aggregates of alpha-synuclein. They showed that these differ in structure between the two diseases. They also differed in the amount of damage they caused in cells cultured in the laboratory, where wrongly folded alpha-synuclein from MSA seems to be more harmful.

“It appears to be possible to use the tracer molecules to analyse cerebrospinal fluid, so we hope that this can be developed to become a molecular diagnosis method in healthcare, possibly in combination with a method to multiply the amount of protein present”, says Peter Nilsson.

Earlier diagnosis

In another study, published recently in the journal Acta Neuropathologica Communications, the researchers have examined aggregates of alpha-synuclein in the brains of people who have died either with Parkinson’s or with MSA. The study is a collaboration with British researchers and was carried out when Therése Klingstedt was a postdoc in Michel Goedert’s lab at the MRC Laboratory of Molecular Biology in Cambridge, Great Britain. This study also used tracer molecules to track wrongly folded alpha-synuclein.Therese Klingstedt.

“Since the colour emitted by our tracer molecules changes depending on the protein structure, we looked at the colour they emitted when bound to aggregates of alpha-synuclein from people with MSA or Parkinson’s disease. We saw different colours from the two groups, which suggests that the structures of the protein aggregates differ”, says Therése Klingstedt, principal research engineer in the Department of Physics, Chemistry and Biology.

Thus, both studies – one analysing brain tissue and the other cerebrospinal fluid – support the theory that it is possible to see differences in the structure of the wrongly folded proteins, and in this way distinguish between the two diseases.

Previous research has suggested that the aggregates of alpha-synuclein start to form several years or decades before the person affected displays clinical symptoms. Peter Nilsson finds the possibility of detecting the wrongly folded proteins in cerebrospinal fluid to be particularly interesting.

“It opens the possibility of making the right diagnosis while the disease is active. Early diagnosis is an important aspect for determining medication and other types of treatment. It should also be able to use the method to evaluate the effects of new potential drugs against the diseases.”

The American scientists have patented the protein amplification method and formed a company to commercialise it.

The articles:
"Discriminating α-synuclein strains in Parkinson’s disease and multiple system atrophy", Mohammad Shahnawaz, Abhisek Mukherjee, Sandra Pritzkow, Nicolas Mendez, Prakruti Rabadia, Xiangan Liu, Bo Hu, Ann Schmeichel, Wolfgang Singer, Gang Wu, Ah-Lim Tsai, Hamid Shirani, K. Peter R. Nilsson, Phillip A. Low and Claudio Soto, (2020), Nature, published online 5 February 2020, doi: 10.1038/s41586-020-1984-7

"Luminescent conjugated oligothiophenes distinguish between α-synuclein assemblies of Parkinson’s disease and multiple system atrophy", Therése Klingstedt, Bernardino Ghetti, Janice L. Holton, Helen Ling, K. Peter R. Nilsson and Michel Goedert, (2019), Acta Neuropathologica Communications, published online 3 December 2019, doi: 10.1186/s40478-019-0840-1

Translated by George Farrants

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