Investigators at the Stanford University School of Medicine have pinpointed a molecular defect that seems almost universal among patients with Parkinson’s disease and those at a high risk of acquiring it.
The discovery could provide a way of detecting the neurodegenerative disorder in its earliest stages, before symptoms start to manifest.
And it points to the possibility of halting the disease’s progression. The defect appears to be exclusive to individuals with Parkinson’s disease.
“We’ve identified a molecular marker that could allow doctors to diagnose Parkinson’s accurately, early and in a clinically practical way,” said Xinnan Wang, MD, Ph.D., associate professor of neurosurgery.
“This marker could be used to assess drug candidates’ capacity to counter the defect and stall the disease’s progression.”
The scientists also identified a compound that appears to reverse the defect in cells taken from Parkinson’s patients. In animal models of the disease, the compound prevented the death of the neurons whose loss underlies the disease.
These steps are described in a study to be published online Sept. 26 in Cell Metabolism. Wang is the study’s senior author. Postdoctoral scholars Chung-Han Hsieh, Ph.D., and Li Li, MD, Ph.D., share lead authorship.
Common neurodegenerative disease
Parkinson’s, the second most common neurodegenerative disease, affects 35 million people worldwide.
Whereas 5%-10% of cases are familial – the inherited result of known genetic mutations – the vast majority are sporadic, involving complex interactions of multiple unknown genes and environmental factors.
So it’s encouraging, Wang said, that both the diagnostic marker and the treatment worked in cells from Parkinson’s patients with either familial or sporadic versions of the condition.
An age-related progressive movement disorder, the disease stems from the mysterious die-off of a set of nerve cells, or neurons, in the brain that fine-tunes bodily movement.
These neurons, which originate in a midbrain structure, the substantia nigra, are referred to as dopaminergic because they secrete a substance, dopamine, to transmit motion-modulating signals to other neurons.
By the time a person starts manifesting symptoms of the disease, an estimated 50% of the substantia nigra’s dopaminergic neurons have already died.
What makes these particular neurons die is unknown.
A leading theory holds that the special intensity with which they perform their duties frazzles their mitochondria.
These bacteria-sized cellular components generate energy for cells in exchange for a steady supply of raw materials: oxygen and carbon-rich carbohydrates or fats.
This process, known as respiration, has a downside: It inevitably generates toxic byproducts called free radicals, which not only can cause cellular damage but are extremely harmful to the mitochondria themselves.
Parkinson’s is known to involve a defect in mitochondrial function.
The harder a cell has to work, the more energy its mitochondria have to churn out—and the more likely they’ll burn out.
Dopaminergic neurons in the substantia nigra are among the body’s hardest-working cells.
Mitochondria spend much of their time attached to a grid of protein “roads” that crisscross cells.
Like old cars that can no longer pass a smog test because they can’t stop spewing noxious exhaust fumes, defective mitochondria have to be taken off the road.
Our cells have a technique for clearing mitochondrial clunkers: a series of proteins that shuffle them off to the cell’s recycling centers.
But first, those proteins have to remove an adaptor molecule called Miro that attaches mitochondria, damaged or healthy, to the grid.
Wang’s group previously identified a mitochondrial-clearance defect in Parkinson’s patients’ cells: Their inability to remove Miro from damaged mitochondria.
In the new study, Wang’s team obtained skin samples from 83 Parkinson’s patients, five asymptomatic close relatives considered to be at heightened risk, 22 patients diagnosed with other movement disorders and 52 healthy control subjects.
They extracted fibroblasts – cells that are common in skin tissue – from the samples, cultured them in petri dishes and subjected them to a stressful process that messes up mitochondria.
This should result in their clearance, necessarily preceded by removal of Miro molecules tethering them to the grid.
Yet the researchers found the Miro-removal defect in 78 of the 83 Parkinson’s fibroblasts (94%) and in all 5 of the “high-risk” samples, but not in fibroblasts from the control group or other or from patients with other movement-disorders.
Screening small molecules
Next, the investigators screened 6,835,320 small molecules, whose structures reside in a commercially available database, in collaboration with Atomwise Inc.
The biotechnology company’s software predicted that 11 of these molecules would bind to Miro in a way that would facilitate its separation from mitochondria and would, in addition, be nontoxic, orally available and able to cross the blood-brain barrier, the study reports.
After feeding these compounds to fruit flies for seven days, the researchers determined that four of them significantly reduced the flies’ Miro levels without toxicity.
They tested one compound, which appeared to target Miro most exclusively, on fibroblasts from a patient with sporadic Parkinson’s disease. It substantially improved Miro clearance in these cells after their exposure to mitochondria-damaging stress.
The scientists also fed the compound to three different fruit-fly strains bioengineered to develop Parkinson’s-like climbing difficulty. Administering the compound to those flies throughout their 90-day life spans produced no evident toxicity and prevented dopaminergic neurons’ death in all three strains and, in two, preserved their climbing ability.
Wang said she believes clinical trials of the compound or a close analog are no more than a few years off.
“Our hope,” she said, “is that if this compound or a similar one proves nontoxic and efficacious and we can give it, like a statin drug, to people who’ve tested positive for the Miro-removal defect but don’t yet have Parkinson’s symptoms, they’ll never get it.”
Stanford’s Office of Licensing Technology has filed a provisional patent for the use of the study’s lead compound in Parkinson’s disease and other neurodegenerative disorders. Wang has formed a company, CuraX, with the goal of speeding its development.
Journal information: Cell Metabolism
Provided by Stanford University Medical Center
Alzheimer’s disease (AD) is the leading cause of dementia worldwide. Mitochondrial abnormalities have been identified in many cell types in AD, with deficits preceding the development of the classical pathological aggregations. Ursodeoxycholic acid (UDCA), a treatment for primary biliary cirrhosis, improves mitochondrial function in fibroblasts derived from Parkinson’s disease patients as well as several animal models of AD and Parkinson’s disease.
In this paper, we investigated both mitochondrial function and morphology in fibroblasts from patients with both sporadic and familial AD. We show that both sporadic AD (sAD) and PSEN1 fibroblasts share the same impairment of mitochondrial membrane potential and alterations in mitochondrial morphology.
Mitochondrial respiration, however, was decreased in sAD fibroblasts and increased in PSEN1 fibroblasts.
Morphological changes seen in AD fibroblasts include reduced mitochondrial number and increased mitochondrial clustering around the cell nucleus as well as an increased number of long mitochondria.
We show here for the first time in AD patient tissue that treatment with UDCA increases mitochondrial membrane potential and respiration as well as reducing the amount of long mitochondria in AD fibroblasts.
In addition, we show reductions in dynamin-related protein 1 (Drp1) level, particularly the amount localized to mitochondria in both sAD and familial patient fibroblasts. Drp1 protein amount and localization were increased after UDCA treatment.
The restorative effects of UDCA are abolished when Drp1 is knocked down. This paper highlights the potential use of UDCA as a treatment for neurodegenerative disease.
Abbreviations: Drp1, dynamin-related protein; PSEN1, presenilin 1; Mfn1, mitofusin 1; Mfn2, mitofusin 2; Opa1, optic atrophy 1
Alzheimer’s disease (AD) is the leading cause of dementia worldwide and is characterized by the build-up of amyloid plaques and neurofibrillary tangles with a loss of neurons later in the disease course [1].
Mounting evidence indicates that amyloid plaques and neurofibrillary tangles do not correlate well with disease severity [2].
Mitochondrial dysfunction is a well-established mechanism in familial and sporadic forms of AD (sAD), with evidence from both post-mortem and peripheral patient tissue as well as animal models.
Fluorodeoxyglucose positron emission tomography imaging in living patients has identified hypometabolism in parietal and temporal brain regions, even in early disease. Alterations in glucose metabolism and cellular respiration have also been found in AD patient fibroblasts [3], [4], [5], [6], [7].
Post-mortem data from AD patients show reduced activity of tricarboxylic acid enzymes and reduced complex IV activity, with complex IV activity decreasing during disease progression [8], [9].
Mitochondrial enzymatic failure, reduced glucose metabolism and increased reactive oxygen species production have all been shown to occur before amyloid pathology [10].
Expression of mitochondrial subunits from all respiratory chain complexes is reduced in the entorhinal cortex (which is an area of early pathological change in AD) of AD patients at post-mortem [11].
In addition, similar changes in expression of mitochondrial genes have been shown early in disease progression in whole blood samples of AD patients [12].
It is not only mitochondrial function that is altered in AD; of particular importance in neurons is mitochondrial dynamics. Mitochondria are in a constant state of flux undergoing fission and fusion events allowing them to adapt and meet local energy requirements.
Evidence from both neurons and patient fibroblasts shows that mitochondria are more elongated and have altered distribution throughout the cell [6].
In particular mitochondria are localized around the perinuclear region in sAD fibroblasts suggesting a collapse of the mitochondrial network [13].
Mitochondrial dysfunction is a shared mechanism between sAD and familial forms of AD.
Transgenic models of familial AD that incorporate amyloid precursor protein (APP) show impaired mitochondrial function and changes in mitochondrial morphology, specifically reduced mitochondrial membrane potential and tricarboxylic acid enzyme enzymes as well as reduced ATP levels [14], [15].
In addition, genetic risk factors for AD alter mitochondrial function. Possession of the APOE4 allele is the largest genetic risk factor for sAD, and possession of this allele is associated with reduced expression of respiratory chain complex proteins and activity of complex IV [16], [17].
Much work has been done trying to elucidate the mechanisms which cause AD with a view to finding therapeutic targets to slow or stop the progression of AD.
To date, however, these interventions have not succeeded in modifying clinical outcome. The search for therapeutic targets has focused mostly around the amyloid cascade.
Mitochondrial abnormalities are also found in fibroblasts of patients with other neurodegenerative diseases. We and others have extensively characterized these changes in Parkinson’s disease (PD) genetic subtypes [18], [19], [20], [21], [22], [23] and MND sporadic and genetic subtypes [24], [25].
We were the first to use the mitochondrial functional deficits as a primary screen in a drug screening cascade for PD [20].
We identified ursodeoxycholic acid (UDCA) in a drug screen of fibroblasts from parkin mutant PD patients, which we have subsequently validated in other forms of PD and other model systems [21].
UDCA is a promising compound as it is already in clinical use for the treatment of primary biliary cirrhosis.
We therefore hypothesized that mitochondrial abnormalities are detectable in fibroblasts from sAD and familial presenilin 1 (PSEN1) patients, and that these abnormalities could be improved with UDCA treatment.
Here we describe our findings of mitochondrial membrane potential, mitochondrial morphology and localization, metabolic activity and mitochondrial fission/fusion machinery expression in sAD and PSEN1 fibroblasts.
In addition, we describe a new mode of action of UDCA on mitochondrial respiration which is abolished when dynamin-related protein 1 (Drp1) is knocked down, indicating that Drp1 is involved in the recovery mechanism in AD.
