10 Dec 2018

Dieticians recommend consuming mostly unsaturated fats for cardiovascular health and a slim waistline. Now, in a surprising twist, researchers claim one such fat, oleic acid, may worsen α-synuclein pathology. In the December 4 Molecular Cell, researchers led by Dennis Selkoe, Ulf Dettmer, and Saranna Fanning at Brigham and Women’s Hospital, Boston, report that oleic acid, a monounsaturated fat that happens to be the major constituent of olive oil, promoted toxicity in a handful of cellular and animal models of Parkinson’s disease. Oleic acid caused aggregated and phosphorylated α-synuclein to build up in the cytoplasm, and the cells were more likely to die. But don’t change your dietary habits just yet—this oleic acid was made on the spot in cells, and may have little to do with consumption. In fact, in a vicious pathological circle, its production seems to be promoted by high levels of α-synuclein. Both yeast and neurons that accumulated α-synuclein overproduced the fatty acid.

The enzyme that converts stearic acid to oleic, stearoyl-CoA-desaturase (SCD), might be a promising new drug target, the authors believe. Inhibiting SCD reduced oleic acid levels, lessened α-synuclein aggregates, and protected neurons from degeneration. The researchers have obtained a grant from the Michael J. Fox Foundation to test SCD inhibitors in mouse models.

A paper in the December 4 Cell Reports supports this approach. Researchers at Yumanity Therapeutics in Cambridge, Massachusetts, independently identified SCD inhibitors as protective against α-synuclein toxicity in an unbiased screen. Inhibitors preserved neuronal health and smoothed out vesicle trafficking, which slows in Parkinson’s disease. Yumanity plans to test an SCD inhibitor in clinical studies next year. “We think this is an intriguing new approach, and we’re excited to test this mechanism in patients,” CEO Kenneth Rhodes told Alzforum.

Lipid Inhibitor for Parkinson’s? Neuroblastoma cells (top) expressing mutant α-synuclein (green) made fewer α-synuclein inclusions (bright dots) when given an SCD inhibitor (bottom). [Courtesy of Molecular Cell, Fanning et al.]

Other scientists agreed the approach has potential, but urged that the research first gather detailed pharmacologic and toxicity data in mice. They note a potential for side effects since oleic acid is abundant in cell membranes throughout the body. “While this new finding is tremendously exciting and highlights novel areas of research for the Parkinson’s community, some caution may be required moving forward toward the clinic,” Simon Stott at The Cure Parkinson’s Trust, a charity based in London, wrote to Alzforum (full comment below). Subhojit Roy at the University of Wisconsin, Madison, said that α-synuclein has been known for some time to interact with lipids, but few therapeutic strategies have exploited this relationship.

Lipidomics Screen: Out Pops Oleic Acid

It’s known that α-synuclein associates with acidic phospholipids in membranes, allowing the protein to hop on and off vesicles (Davidson et al., 1998; Perrin et al., 2000). Selkoe and others reported that α-synuclein directly binds fatty acids, including oleic (Sharon et al., 2001; Kubo et al., 2005). These interactions seemed to nudge α-synuclein to assume a helical shape and promoted its oligomerization (Perrin et al., 2001; Feb 2003 news).

Fanning wondered how α-synuclein might affect the lipid composition of cells. Working initially in the lab of the late Susan Lindquist at MIT, Fanning analyzed the lipidomic profile of yeast that expressed human α-synuclein driven by an estrogen promoter. Within six hours of α-synuclein induction, these cells massively overproduced oleic acid, up to 60-fold excess. Fatty acids are incorporated into diglycerides, and in keeping with this, Fanning saw a threefold excess of lipid droplets that contain these fats in the yeast as well.

Oleic acid and diglycerides both seemed to harm cells. With the buildup of diglycerides came sluggish vesicle trafficking and cytotoxicity. Adding exogenous oleic acid to yeast cultures that expressed α-synuclein aggravated cell death. Oleic acid did not harm wild-type yeast. Other fatty acids had no effect on cell health, however. Turning off the yeast ortholog of SCD, Ole1, which converts stearic acid to oleic acid, rescued cells.

Would the link between α-synuclein and fat production hold true in neurons? Results from several model systems suggest as much, the scientists report. In a roundworm model of PD, suppressing oleic acid production cut dopaminergic neuron death in half. In rat primary cortical neurons, overexpression of human α-synuclein led to a twofold excess of oleic acid and sped cell death. As in yeast, knocking down SCD1 preserved cells. Mice that express human mutant E46K α-synuclein accumulated excess diglycerides whose levels correlated with motor deficits. Human neurons derived from iPS cells followed the same pattern, whereby overexpressing wild-type α-synuclein in these cells doubled oleic acid levels. Neurons derived from patients with an α-synuclein triplication, and neurons carrying the familial E46K mutation, also made more diglycerides than control lines.

To get an idea of how oleic acid could end up being toxic, the authors used a human neuroblastoma cell line that expresses α-synuclein with E46K mutations in several KTKEGV motifs that repeat in the protein. E46K α-synuclein adopts monomeric form more readily than wild-type, and this causes neuronal inclusions, as seen in a mouse E46K repeat model developed in the Selkoe lab (Oct 2018 news). Knocking down or inhibiting SCD1 suppressed inclusions. Inhibiting SCD also lessened other signs of α-synuclein toxicity in these neuroblastoma cells. The tetramer-to-monomer ratio rose, levels of phosphorylated α-synuclein sank, and more of the protein was soluble rather than membrane-bound. Selkoe and colleagues argue that tetramers represent a physiological, beneficial form of α-synuclein (Aug 2011 news; Feb 2012 news; Apr 2015 news). Exposing these cells to oleic, but not other, fatty acids dose-dependently increased the number of α-synuclein inclusions.

The exact mechanism of how a cell’s lipid composition affects these α-synuclein properties remains unclear. When monounsaturated fatty acids, such as oleic, slide into membranes, the lipid bilayers become more fluid. Fanning thinks this might matter. “The way α-synuclein interacts with those membranes apparently causes problems in some way and increases toxicity to the cell,” she told Alzforum. Likewise, it is a mystery how α-synuclein boosts oleic acid production. Dettmer emphasized the circular relationship. “I found it fascinating that α-synuclein seems to upregulate the exact factor that makes it more toxic,” Dettmer said.

Drug Screen: Out Pops SCD Inhibitor

Yumanity researchers arrived at SCD from a different direction. Joint first authors Daniel Tardiff and Benjamin Vincent screened for compounds that rescued growth in yeast overexpressing α-synuclein. A clutch of compounds containing an oxadiazole core did the best. Their benefit was specific for α-synuclein; it did not reduce other toxicities, such as Aβ. The most potent one, YTX-465, boosted growth more than fourfold, to 40 percent of that in wild-type cells, at a concentration of 50 nM. It also restored vesicle trafficking to normal and reduced the number of α-synuclein foci.

Further experiments revealed the SCD ortholog Ole1 as a target of YTX-465. Ole1 produces both oleic and palmitoleic acid. Adding either to yeast cultures abrogated YTX-465’s effects, supporting these fatty acids as key downstream effectors for the compound. At 500 nM, YTX-465 inhibited the growth of wild-type yeast, indicating that suppressing oleic acid by too much can be toxic.

The authors generated human cortical neurons from iPS cells, transfected them with human mutant A53T α-synuclein, and treated them with an YTX-465 analogue that was active against SCD1. Levels of monounsaturated fatty acids rose, and neuron viability improved. This analogue will not advance to the clinic, however; Rhodes noted that Yumanity has selected a molecule with a different chemical scaffold and distinct properties for human studies.

Overall, despite differences in the model systems, both groups’ findings dovetail closely. “It’s encouraging that the two papers are in agreement on so many levels,” Fanning said. Rhodes agreed, noting, “This lends credence to the importance of this target in α-synuclein biology.”

One discrepancy was that the Yumanity researchers saw no change in fatty acid composition in yeast expressing α-synuclein. Fanning thinks that new fatty acids might have been missed because they were incorporated into triglycerides. The Yumanity researchers did see an increase in triglyceride content. Selkoe noted that Yumanity used a galactose-inducible promoter to express α-synuclein in yeast, rather than an estradiol one. The addition of galactose to yeast cultures by itself changes the fatty acid composition, and could have masked the effects of α-synuclein, Selkoe said. Rhodes agreed this could be the case.

Implications for Health?

Is SCD inhibition promising as a therapeutic approach? David Standaert at the University of Alabama, Birmingham, noted that many cellular factors have been found to modulate α-synuclein aggregation and toxicity. This includes molecules involved in trafficking, lysosomal degradation, and protein folding. “Oleic acid, however, may be more amenable to therapeutic intervention than some of these other pathways,” he wrote to Alzforum (full comment below).

Nonetheless, others wondered about safety. “If oleic acid is important for the toxicity of α-synuclein, it might also be important for its normal function,” said Robert Edwards at the University of California, San Francisco. Stott pointed to preclinical studies where inhibiting SCD exacerbates inflammation in models of colitis, and is detrimental in at least one diabetes model (Chen et al., 2008; Flowers et al., 2007). “There could be complications in peripheral tissues,” Stott suggested.

Dettmer agrees. Ideally, SCD inhibition should be directed to the brain, he said. One way to do this might be to target SCD5, which is confined to the central nervous system, rather than SCD1. Selkoe plans to investigate the pros and cons of various existing SCD inhibitors in mouse models of Parkinson’s. He believes the target range for suppressing oleic acid production will be low, perhaps inhibition of less than 30 percent.

So what about dietary fats? Oleic acid is found in many foods considered healthy, such as olive oil, avocadoes, and nuts. Rhodes noted that fatty acids in the bloodstream barely enter the brain. “It looks like most of the lipids we are studying are generated within the CNS,” he said. Still, Selkoe believes the issue is worth studying. He suggested putting a PD mouse model on a diet low in monounsaturated fats and examining the effects, if any, on α-synuclein pathology. In the meantime, it’s probably still safe to enjoy those foods. Adherence to a Mediterranean diet, which is rich in monounsaturated fats, has been linked to a lower risk of Parkinson’s in small epidemiological studies (Alcalay et al., 2012; Maraki et al., 2018; Agarwal et al., 2018).—Madolyn Bowman Rogers