BioTechniques
02/17/2016 Tori Tanenbaum

Despite spending millions of dollars on dementia drug research, the answer to reversing Alzheimer’s disease may lie in a non-invasive, non-pharmacological treatment after all.


With society’s elderly population growing, the prevalence of Alzheimer’s disease (AD) with its associated cognitive deterioration will affect an increasing number of lives. Symptoms of memory loss, confusion, and even personality change arise largely because of two types of brain lesions—amyloid-β plaques and Tau protein tangles—which lead to neuron death [1]. Despite significant efforts to combat this devastating disease, there is currently no known cure or preventive treatment.

In fact, the field has experienced some significant setbacks recently. In October 2015, Pfizer ended a Phase II trial of an AD drug that is supposed to preserve cognitive ability [2] because early results indicated that it would fail the endpoint efficacy criteria established at the beginning of the study. Axovant Sciences, a biotechnology corporation also developing pharmaceuticals for dementia, saw a steep drop in share prices following Pfizer’s announcement since Axovant’s lead investigational drug works through a similar mechanism to Pfizer’s terminated drug [3].
While multiple drugs have failed, hope remains. Now, two minds out of Queensland, Australia propose a groundbreaking solution for reversing AD-related dementia, one that suggests pharmaceutical drugs might not be the answer after all. Reporting in the journal Science Translational Medicine, researchers have harnessed scanning ultrasound technology to repeatedly open the blood-brain barrier (BBB), clearing amyloid-β plaques and restoring memory function [4].

"This scanning ultrasound approach shows that you don’t need antibodies or drugs, but simply opening the BBB with repeated short ultrasound treatments can evoke a similar clearance response and improve cognitive measures,” said Gregory Cole, associate director of UCLA’s Alzheimer’s Research Center, who was not involved with the study. “This is a very creative and cool approach.”

Brain Matters

As the first PhD candidate at the Queensland Brain Institute’s Clem Jones Center for Aging Dementia Research, Gerhard Leinenga is excited about the opportunity to work on a disease that affects others so profoundly. “I found looking at what happens to the brain when things go wrong—in this case, my current research into Alzheimer’s disease—to be very interesting work and a unique complimentary approach to studies already finding out what is happening in a normal brain,” he said.

Leinenga’s first daunting challenge was to find a way to access the amyloid-β plaques and Tau protein tangles contained in the brains of AD patients. One significant problem is the BBB, which filters blood transported to the brain and spinal cord in an effort to block harmful substances, including drugs, from entering. With only 5% of the 7000 available small-molecule drugs able to transport across the barrier, researchers have numerous obstacles to overcome in order to treat brain lesions [5].

Leinenga’s team aims to find ways to prevent and treat dementia, especially as it pertains to AD. In this time-sensitive battle, they will consider any and all possibilities for treatment.

“Scanning ultrasound technology is fairly new as it’s been developed by a couple of groups in North America, “ said Leinenga. “I was very intrigued by their papers and my supervisor was inspired by a presentation given by Elisa Kono***ou at an overseas conference, so we looked at all the literature and decided we wanted to apply [the technology] in our own lab.”

Breaching the Barrier

Kono***ou and her colleagues at Columbia University used a focused ultrasound technique to penetrate the seemingly impermeable BBB [5]. The ultrasound technique requires coupling with microbubbles, which are used as ultrasound contrast agents. These microbubbles—composed of lipid shells and gaseous centers—are inserted intravenously. Administered ultrasound pulses produce focal vibrations that cause the microbubbles to expand and contract, which essentially opens tight junctions in vasculature and enables diffusion across the BBB [4]. By demonstrating that the BBB can, in fact, be disrupted, Kono***ou’s research raised the possibility of treating many neurological disorders non-invasively.

Leinenga and his supervisor, Jürgen Götz, embarked on this project without any prior experience using ultrasound technology. The duo’s goal was clear; they intended to use a scanning ultrasound technique to open the BBB in AD mouse models and then remove amyloid-β lesions to reverse cognitive effects.

Leinenga and Götz designed three memory tests to gauge the efficacy of scanning ultrasound in reversing amyloid-β plaques and restoring memory function. They first placed the mice in a rotating arena with a grid. When mice enter one zone in the round arena, they get a small electric shock. To avoid this area in a rotating arena, they have to rely on cues outside of the maze by establishing where the shock zone lies in relation to the room.

“Another test involved the Y-maze, which is a commonly used test of spatial memory,” explained Leinenga. “The third test was the novel object test which is another frequently used test of memory where the mouse first explores two identical objects, and then after an interval —a retention period—one of the objects is replaced by an object the mouse has never seen before. Because mice are curious creatures, they should explore the novel object more if they remember that they have already seen the familiar object.”

Leinenga and Götz tested their mice prior to ultrasound treatment and then treated them 5 separate times over 6 weeks before subjecting the mice to the memory tests again. The researchers saw restored spatial and recognition memory results in the scanning-ultrasound-treated mice. The control, “sham-treated” mice did not perform as well in the second set of memory tests in comparison to the treated mice, with their performance being similar to wild-type mice without the AD-causing gene [4].

“Seeing that there were beneficial effects of the treatment on various aspects of the pathology that we were testing, that was very encouraging,” said Leinenga. “As we looked at different aspects such as behavior, pathology, and the neuronal cells, it was interesting to find that there were effects on all of these,”

To investigate the mechanism causing amyloid-β plaque reduction, the duo conducted additional testing and found that microglial activation induces the clearance by enabling microglial lysosomes to internalize amyloid-β [4].

“There was a risk involved in the sense that I might not be able to get it to work, or that other people working in the field a lot longer had been able to do things that I might not be able to do in a small amount of time,” Leinenga said. “But in the end it paid off; the technique is very versatile and can be applied quite broadly.”

Cole cautions that there are some caveats, however. “While the authors find no evidence of overt toxicity, the BBB is there for a reason and BBB damage can cause major damage. So it is essential to limit any opening of the BBB, and we don’t know what the margins for error will be in people,” he said. “It will be important to get a better idea of the mechanism. For example, it would be nice to rule out the possibility that this treatment allows peripheral macrophages and monocytes to enter the brain, as they are known to clear amyloid.”

Making Waves

In addition to investigating the mechanism of amyloid-β clearing by scanning ultrasound treatment, Leinenga is interested in exploring the new approach for administering pharmaceuticals. “Normally the BBB only allows very tiny amounts of drugs, like antibodies, to get into the brain, and in situations where we know that these drugs can be effective therapeutics, using this technique allows you to deliver larger amounts,” he speculated.

Leinenga is also trying to apply the technique to sheep since their skulls are similar in thickness to human skulls. The larger sheep brain is also closer in size and shape to human brains as well. Having a similar model is important for developing ultrasound technology, because the physics can differ in a mouse versus a human due to the difficulty of transmitting through a thicker human skull.

“We’re also looking at other types of brain pathologies to see whether ultrasound can cause an improvement in some of these other pathologies where there are protein aggregates in the brain,” Leinenga said.

References

1. “Alzheimer’s Disease.” Focused Ultrasound Foundation: Accelerating the Development and Adoption of Focused Ultrasound. Focused Ultrasound Foundation. Web. 7 February 2016.

2. “Study Evaluating The Safety And Efficacy Of PF-05212377 Or Placebo In Subjects With Alzheimer's Disease With Existing Neuropsychiatric Symptoms On Donepezil.” ClinicalTrials.gov. U.S. National Institutes of Health, 29 January 2016. Web. 7 February 2016.

3. “Stock information.” AXOVANT Dementia Solutions. Axovant Sciences Ltd., 3 February 2016. Web. 7 February 2016.

4. Leinenga G., Götz J. “Scanning ultrasound removes amyloid-β and restores memory in an Alzheimer’s disease mouse model.” Science Translational Medicine. 11 March 2015. DOI: 10.1126/scitranslmed.aaa2512

5. Kono***ou E.E., Tung Y-S., Choi J., Deffieux T., Baseri B., Vlachos F. “Ultrasound-Induced Blood-Brain Barrier Opening.” Current Pharmaceutical Biotechnology. June 2012. DOI: 10.2174/138920112800624364#sthash.loBiPxNW.dpuf