14 Dec 2018

Never mind the wrinkles and gray hair. Embattled microglia, waning neurogenesis, and withering synapses are also part and parcel of the aging process. What drives those seemingly inexorable changes inside the brain? At the Society for Neuroscience meeting, held November 3–7 in San Diego, researchers blamed factors from the outside. Specifically, they identified circulating immune cells as key purveyors of aging in the hippocampus, culminating in memory loss in mice. One such factor, eotaxin, rises in plasma as people age. Blocking its action restored memory in aging mice, and protected mouse models of Parkinson’s disease from motor deficits, sparking early stage clinical trials in humans.

The findings extend striking previous,evidence, conducted largely by the same scientists, that systemic factors fuel age-related decline in brain function. In 2014, a trio of papers reported that blood from old mice brought on markers of brain aging in young mice, while blood from young mice ablated aging markers in old mice (May 2014 news on Villeda et al., 2014). Young blood slowed synapse loss and neuroinflammation in mice modeling Alzheimer’s disease (Sep 2016 news). 

Immune Orchestra of Aging?
At SfN, researchers from the lab of Saul Villeda at the University of California, San Francisco, hypothesized that circulating immune cells were likely responsible for these parabiotic effects. To investigate whether these cells trigger aging in the brain, graduate student Luke Smith transplanted hematopoietic stem cells (HSCs) from 24-month-old mice into two-month-old mice. As a control, he transplanted other two-month-old mice with HSCs from same-age littermates. Four months later, Smith found that compared with controls, mice with old HSCs performed worse on tests of hippocampal-dependent memory, such as the water maze and fear conditioning. A look at their brains revealed less neurogenesis in the hippocampal dentate gyrus, more activated microglia, and fewer dendritic spines on hippocampal neurons than in controls.

The finding suggests that hematopoietic cells impart some of the blood’s aging influence on the brain, said Smith. Whatever the specific cell type, it likely exerts its sway by some soluble factors, he added, as nary a transplanted cell was found within the brains of recipients.

Could such factors act via microglia? After all, chronic activation of microglia is a hallmark of brain aging, and thought to modulate neurodegeneration. While exposure to old plasma can boost activation of microglia (May 2014 news), Jeremy Shea, a postdoc in Villeda’s lab, wanted to investigate whether microglia are also influenced by their local environment in the brain. Focusing on the hippocampus, Shea first simply asked how microglia there change with age. As expected, he found they became more activated, rounding up and expressing markers such as CD68. Surprisingly, this age-related activation depended on the ZIP code within the hippocampus. Microglia in the granule cell layer and hilus, but not the molecular layer, were activated.

To understand how the environment of the aging brain itself affects microglia, Shea next transplanted fledgling microglia extracted from the brains of newborn mice into the hippocampi of young or old mice. These naïve microglia assumed a more activated state after settling into old but not young hippocampi.

Returning to parabiosis, the researchers found that microglia in young mice ramped up an age-related gene expression signature after being conjoined to old mice, upregulating disease-related and immune genes. This signature was also present in hippocampal microglia from old mice, and the researchers found they could lessen it by joining old mice to youngsters. Shea said that the findings cast microglia, in keeping with their function, as exquisitely sensitive responders to both the local brain environment as well as systemic factors coming in from the blood.

Eo-taxin’ the Brain with Age
What, then, are those secret ingredients in blood that age the brain? According to Sakura Minami of the San Carlos-based biotech company Alkahest, one is eotaxin, a ligand for the C-C chemokine receptor type 3 (CCR3). Using proteomics approaches, Minami and colleagues saw that eotaxin shot up in the blood with age. Also known as CCL11, eotaxin is known to play a role in inflammation, for example in the recruitment of eosinophils upon CCR3 engagement. Given that these infection-fighting white blood cells can become damaging in allergic conditions such as asthma, CCR3 is an established drug target.

The researchers previously demonstrated that injecting eotaxin into young mice caused neurogenesis in the dentate gyrus to slow to a trickle, and that neutralizing eotaxin with an antibody blocked this effect (Aug 2011 news). At SfN, Minami reported preclinical findings from efforts to stifle the consequences of age-related eotaxin elevation. Rather than target eotaxin directly, Alkahest scientists blocked CCR3 with a small molecule antagonist acquired from Boehringer-Ingelheim. The drug, first called ALK4290, then AKST4290, was reportedly well-tolerated in previous clinical trials, though Boehringer-Ingelheim did not make results public.

First, the researchers established that rising eotaxin was detrimental to memory in mice. They put the mice through a Barnes maze test, placing them on a circular platform with holes surrounding its circumference, only one of which led to an escape box below. The mice had to use visual cues in the room to remember the escape hole. To make the test even more challenging, Minami modified the set-up to include rotating target holes. The researchers found that injecting young wild-type mice with recombinant eotaxin docked their performance on the test. Much like old mice, they were slow to locate target holes. For both sets of mice, treatment with the CCR3 antagonist corrected the deficit to the point where they performed as well as young, untreated animals, Minami reported. The researchers obtained similar results using the Y-maze, another spatial memory test.

Minami told the SfN audience that she noticed treated mice moved faster than controls through the Barnes maze, though the quickened pace did not explain their performance boost. Was there a motor benefit? Exploring this, Minami found that old mice injected with the CCR3 antagonist maintained their balance on a spinning rod nearly as long as young mice, while old control mice fell off quickly.

They next tested the drug in Line 61, Parkinson’s disease mice that overexpresses human, wild-type α-synuclein (Chesselet et al., 2012). After spiking the mice’s drinking water with the drug for six weeks, the researchers saw dramatic improvement in grip strength, and modest improvement in balance, compared with untreated Line 61 controls. Minami said plans to test AKST4290 in people with PD are in the works.

Researchers questioned Minami on AKST4290’s mechanism of action. Does it affect dopamine levels, which wane in PD? Minami said that while they are investigating whether the drug alters dopamine or its metabolites, she believes its benefit stems from anti-inflammatory properties. In support of this, Minami reported that AKST4290 markedly reduced microgliosis in the striatum of PD mice. In addition, the drug staved off the microglial activation and cognitive decline usually seen in response to chronic, low doses of lipopolysaccharide, a model of chronic inflammation.

Exactly how eotaxin in the blood manages to trigger neuroinflammation in the brain remains uncertain. Researchers in Tony Wyss-Coray’s lab at Stanford Medical School recently reported that the brain vasculature itself might play a role in transmitting such aging effects into the brain: expression of VCAM1, an adhesion molecule that tethers passing leukocytes, on brain endothelial cells was critical to allow aged blood to accelerate brain aging (Jul 2018 news). 

Alkahest recently completed two Phase 2 clinical trials testing AKST4290 in people with age-related macular degeneration, another condition in which inflammation plays a starring role. Results from those trials are not yet public.

In addition, the company is testing GRF6019, a proprietary plasma fraction, in people with AD (see clinicaltrials.gov and clinicaltrials.gov),  and has started a trial testing yet another fraction, GRF6021, in people with Parkinson’s disease with cognitive impairment or dementia (aka PDD).

At SfN, Alkahest scientists Eva Czirr and Viktoria Kheifets described how they identified the plasma fraction currently being tested in AD, GRF6019, as a booster of neuronal activity, synaptic function, and neurogenesis, and a douser of neuroinflammation, in mice and in neuronal culture models (Dec 2017 conference news and Jul 2018 conference news).—Jessica Shugart

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References

News Citations

1. In Revival of Parabiosis, Young Blood Rejuvenates Aging Microglia, Cognition 5 May 2014
2. Young Blood a Boon for APP Mice 14 Sep 2016
3. Paper Alert: Do Blood-Borne Factors Control Brain Aging? 31 Aug 2011
4. VCAM1: Gateway to the Aging Brain? 17 Jul 2018
5. Blood, the Secret Sauce? Focus on Plasma Promises AD Treatment 14 Dec 2017

Therapeutics Citations

1. GRF6019

Paper Citations

1. Villeda SA, Plambeck KE, Middeldorp J, Castellano JM, Mosher KI, Luo J, Smith LK, Bieri G, Lin K, Berdnik D, Wabl R, Udeochu J, Wheatley EG, Zou B, Simmons DA, Xie XS, Longo FM, Wyss-Coray T. Young blood reverses age-related impairments in cognitive function and synaptic plasticity in mice. Nat Med. 2014 Jun;20(6):659-63. Epub 2014 May 4 PubMed.
2. Chesselet MF, Richter F, Zhu C, Magen I, Watson MB, Subramaniam SR. A Progressive Mouse Model of Parkinson's Disease: The Thy1-aSyn ("Line 61") Mice. Neurotherapeutics. 2012 Apr;9(2):297-314. PubMed.

External Citations

1. clinicaltrials.gov
2. clinicaltrials.gov
3. GRF6021

Further Reading

News

1. VCAM1: Gateway to the Aging Brain? 17 Jul 2018