Madrid: ICAD Conference Draws to a Close

21 Jul 2006

The 10th International Conference on Alzheimer’s Disease and Related Disorders, ICAD for short, ended yesterday just outside the palatial capital of Spain. The conference attracted not only a record number of attendees—just above 5,000 from 50 countries—but also a visit by Queen Sofia, whose philanthropy supports AD care and research in her country. (The first ICAD conference, held 1988 in Las Vegas, hosted around 300 attendees.)

In several dozen informal interviews in hallways, ballrooms, on escalators and shuttle buses, attendees applauded that many scientists presented unpublished data in their talks and posters. Presentations included major new developments, such as the discovery that the progranulin gene causes a form of frontotemporal dementia (see ARF related news story) as well as the discovery of a physiological function for the enzyme BACE (see ARF related news story) Smaller nuggets of news solidified emerging trends or opened up new research directions. Some scientists grumbled about feeling crammed into a crowded poster area while the neighboring company exhibits enjoyed all the space and air that’s necessary for animated conversation. Others viewed the commercial presence, together with the large number of presentations on a variety of experimental therapies, as a positive sign of the field’s needed movement toward translational science as patient numbers grow inexorably.

Even a casual flick through the conference program makes clear that a large fraction of the presentations focused on a panoply of different early detection and diagnostic efforts. Research in this area has exploded compared even to as recently as 5 years ago. It included attempts to detect telltale fingerprints of preclinical AD in body fluids, such as innovative work on leukocyte gene expression profiles or more advanced attempts to validate known biomarkers in the cerebrospinal fluid. Efforts ranged from imaging methods and more refined neuropsychological tests to proxy markers derived from epidemiological research on risk factors in the cardiovascular and metabolic fields. On the imaging front, news included updates on the amyloid imaging agent PIB and its use in presymptomatic carriers of familial AD mutations, as well as talks on promising new approaches such as diffusion tensor imaging and perfusion MRI to measure the degradation of the brain’s white matter that is thought to precede overt AD.

This early detection work reflects an emerging consensus among scientists that Alzheimer disease develops for a decade, perhaps even longer, before clinical signs become apparent. In fact, researchers increasingly compare the phase of diagnosed AD as we know it to terminal, metastatic cancer—the end stage of a disease that ideally should be treated years before the patient shows up at the neurologist’s door. The need for early detection is pressing, and many scientists wonder whether some drugs fail partly because they are tested too late in the disease.

The lack of validated, consensus biomarkers notwithstanding, this year’s ICAD program indicates that therapeutic approaches appear to grow exponentially at the present stage of AD research. The majority of them dig into some aspect of the amyloid hypothesis, but not all. Outliers that buck this overwhelming trend include intranasal insulin, growth factor gene therapy, gonadotropal hormones, tau immunotherapy, and dietary supplements. Immunotherapy approaches appear to have mushroomed, but many scientists remain as wary of their potential side effects as they are hopeful about their ability to remove forms of Aβ or amyloid. Finally, if γ-secretase inhibitors could speak, they might borrow a quote from Mark Twain and declare that the reports of their death have been greatly exaggerated. β-secretase inhibitors, still largely guarded behind the doors of pharmaceutical companies, are beginning to show their face, as well.

What works? To date, some trials remain ongoing and blinded, other approaches look promising in pilot trials only to fail in larger ones, and yet others appear to be limping along with rather small effect sizes. Piecing together the shards of the shattered AN1792 trial, scientists appear to be finding hints that the patients might yet have benefited a whit from the prototype vaccine after all. That said, the croupier is still taking bets on which of the current approaches will survive the minefield of cost, trial design headaches, patient recruitment and dropout woes, side effects, bad press, and struggle with the FDA that together characterize today’s drug development environment.

Some issues were notable for their absence. For example, there were fewer discussions than at prior conferences about whether Aβ oligomers, fibrils, or plaques were the toxic species in AD. Researchers appeared to have adopted the stance that all of these are damaging when present in excess over a long period of time (though more are also coming around to think that, at the proper levels, Aβ may indeed have a physiological role to do with synaptic activity). Broadly speaking, oligomers are suspected of triggering acute cognitive deficits, whereas plaques are thought to damage the structure and transmission capability of the brain and to fuel chronic inflammatory states. Likewise, investigators no longer debated whether Aβ or tau are detrimental factors in AD. Most view both proteins as critical, and Aβ as upstream of tau. At the Alzforum symposium, Bart de Strooper referred to the “Amyloid Tau Hypothesis” as a framework for AD research. What’s more, some scientists suggested that tau pathology appears to occur downstream of many amyloids, not just amyloid-β but also prion amyloid and synuclein deposits, for example. Yet another shift appears to occur around the notion that, increasingly, basic scientists no longer focus as relentlessly on Aβ or tau as they tended to do before, but view them as but two players in a broader drama of oxidative, synaptic, and proteasomal stress, confounded by systemic changes such as high blood pressure, insulin resistance, and other components of what is collectively called metabolic syndrome. Consequently, future treatments will likely have to include combinations of drugs directed against Aβ, against tau, as well as neuroprotective, cardiovascular, and other agents.

Details from roughly 340 talks and 400+ posters in six days can make the most capacious brain overflow. More than 300 conventioneers took advantage of a break offered by the Alzforum and joined our 10th anniversary symposium. Called “Mapping the Next Decade of Alzheimer Research,” it took a crack at integrating major ideas and trends in AD research. Admission was possible only for those who passed a spatial learning paradigm called the Madrid Paper Maze. Probants had to navigate a trail of signs to find the hidden platform—that is, a location changed at the last minute to a different quadrant of the convention center. We will post coverage of this event, as well as a stream of news stories from the ICAD conference, over the next two weeks. As always, contributions from our readers are warmly invited.—Gabrielle Strobel.

—Gabrielle Strobel

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Madrid: BACE Found to Have Big Job in Wrapping Motoneurons

21 Jul 2006

By the wholly unscientific survey of a roving reporter, the single most convincing and surprising molecular biology story that stuck in the minds of scientists at the 10th ICAD meeting was Christian Haass’s demonstration of a physiological function for the β-secretase BACE-1. This aspartyl protease has for years been considered an ideal drug target that appeared to do little else of much consequence other than cleave APP to yield the Aβ peptide BACE-1. Now it turns out that BACE-1 signals Schwann cells to ensheath motoneurons in a developmental period of intense myelination after birth. Here’s the gist of the story, hot off the tail-end of the conference, which concluded yesterday here in Madrid, Spain. (Minji Kim, Alice Lu, and Rudy Tanzi first mentioned this work in their report of this spring's Keystone meeting. Here is a more detailed story.)

Haass’s group at Ludwig Maximilian University got onto the trail of myelin when they established a developmental profile of BACE-1 expression in the mouse. They saw no BACE expression in adult mice, but sky-high expression in the two weeks after birth. “It was a difference like night and day and night,” recalled Haass. “That gave us the idea that something must happen functionally after birth with BACE.”

In parallel, the scientists knocked down BACE in zebra fish and noticed a movement phenotype, where the fish no longer swam away as normal ones do in response to a shock. This pointed to a problem with the peripheral nervous system. Because myelination begins in earnest right after birth, it came up as a suspect at this point. Turning back to mice, the researchers made sections of the sciatic nerve of the BACE-1 knockout mice generated by Bart de Strooper’s lab in Belgium and saw that it was much less extensively myelinated than in wild-type mice. Some axons in the BACE-less sciatic nerve had no myelin around them, while others had an abnormally thin sheath. The axons in their sciatic nerve were also bundled aberrantly.

This hypomyelination persisted into adulthood. It is not a quirk of this particular mouse strain, either. Martin Citron at Amgen in Thousand Oaks, California, checked in adult mice of his groups’ BACE-1 single and BACE-1/2 double knockouts, and reported at the ICAD conference that their myelin, too, was thinner by about a third than that of wild-type. BACE-2 knockout mice do not show this phenotype, however, so it is specific to BACE-1. The mice show no overt motor deficit, but Citron noted that they have yet to be stressed or subjected to detailed strength tests.

The hypomyelination phenotype is well known in the literature. It resembles the phenotype of a heterozygous neuregulin-1 knockout (homozygous neuregulin deletion is lethal), as well as that of knockouts of ErbB, a neuregulin-1 ligand. This hinted that BACE function could lie in neuregulin-1 signaling.

Next, the German scientist, which included Linna Rabe and Michael Willem, looked for neuregulin-1 in mouse brain lysates. They reasoned that if it were a substrate for BACE, then in knockout mice the full length neuregulin-1 protein would accumulate. They indeed found unprocessed neuregulin-1 in the lysate and with further mechanistic work found evidence suggesting that BACE cuts neuregulin-1 twice, once in front of and once right behind neuregulin-1’s EGF domain. This releases the EGF domain from the axon, possibly to signal to the Schwann cell, Haass speculated.

As always with first data from basic science, their implications for drug development are hard to predict. Citron, Haass, and other scientists noted that they do not see it as a show stopper for BACE as a target, because the effect occurs mostly in development. Importantly, wholesale genetic knockouts represent a more extreme phenotype than a calibrated pharmacological reduction of BACE levels. Even so, after brain injury, BACE is known to become upregulated and conceivably aids in remyelination of damaged fiber tracks. AD patients who get a stroke or, more immediately relevant, a peripheral nerve injury, while on a BACE inhibitor, might in theory recover less well than they would without the drug. This is far from clear at this point, as the knockout mice have a peripheral phenotype, not a central one. Oligodendrocytes, which myelinate central axons in the brain, may be very different with regard to BACE and neuregulin. But the specter of a myelination side effect has put a caveat that bears watching on the radar screen of the many drug companies engaged in developing BACE inhibitors.

Haass believes that signaling the myelination of peripheral neurons is a major physiological function of BACE-1. An evolutionary tidbit supporting this view lies in the fact that invertebrates have no BACE-1. Myelination evolved with vertebrates, and BACE evolved in parallel.

The relevance of this finding for AD pathogenesis needs more research. White matter degeneration shows up as a frequent, if underappreciated, feature of early AD in various imaging modalities. Indeed, 46 presentations dealt with white matter changes in cognitive impairment and AD at the ICAD conference alone. Readers may want to consider George Bartzokis’s provocative hypothesis that AD is a disease of myelin breakdown that proceeds in a reverse ontogenetic order of myelination; that is, areas that are myelinated last in teenage development lose myelin first during AD pathogenesis, and early-myelinating areas in early childhood degenerate last in the course of AD (Bartzokis et al., 2003; Bartzokis, 2004). This is food for thought in a broader context, but the difference between peripheral and CNS BACE action precludes a direct analogy at this point.

The Haass group’s finding is equally relevant for trauma and schizophrenia. In particular, neuregulin-1 is one of the genetic risk factor genes for schizophrenia. By logic, BACE may then well be involved in schizophrenia, as well. As a next step, Haass is beginning to look for BACE upregulation and increases in neuregulin signaling in fresh postmortem schizophrenia brain tissue. This discovery, then, has brought a new lab to the growing field of schizophrenia research. The closely intertwined goings-on between molecular players in AD and schizophrenia represent an intriguing aspect in this research: BACE-1 cleaves neuregulin, and the protease that takes the baton from BACE-1 during AD pathogenesis, γ-secretase, is known to cleave a neuregulin receptor on Schwann cells, that is, ErbB4.

In summary, this work brings to light a broader parallel between APP and BACE function: Both have an important physiological role in development (neuronal migration in the case of APP, see related news story, myelination in the case of BACE), whereas in adult brains they appear to play useful repair roles in response to stressors such as ischemia or injury. Then where, exactly, do things first go awry?—Gabrielle Strobel.

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Bartzokis G, Cummings JL, Sultzer D, Henderson VW, Nuechterlein KH, Mintz J. White matter structural integrity in healthy aging adults and patients with Alzheimer disease: a magnetic resonance imaging study. Arch Neurol. 2003 Mar;60(3):393-8. PubMed.
Bartzokis G. Age-related myelin breakdown: a developmental model of cognitive decline and Alzheimer's disease. Neurobiol Aging. 2004 Jan;25(1):5-18; author reply 49-62. PubMed.

Madrid: Why Do People Drop Out of Trials?

26 Jul 2006

Designing AD trials well, drumming up the funding, and coordinating sites across the country and the world—all that would seem to be challenge enough for clinicians in the Alzheimer disease field. But even once that’s done, they don’t enjoy smooth sailing. A particularly vexing problem down the line is that even the best trials are hemorrhaging participants, and doing so in surprisingly large numbers. What’s more, participants drop out unevenly from study arms that were well balanced at trial outset, draining the remaining sample of precious power and leaving it biased.

Why are people quitting? Jennifer Emond, a statistician at the University of California, San Diego, tackled this question. She presented her results at the 10th International Conference on Alzheimer’s Disease and Related Disorders, held July 15 to 20 in Madrid. Emond chose as her example a secondary prevention trial that was praised for its careful design and seen as a forerunner for future trials of the earliest stage of AD. Led by Ronald Peterson, Leon Thal, and other members of the Alzheimer Disease Cooperative Study (ADCS), this publicly funded trial randomized 769 people with amnestic MCI into a vitamin E arm, a donepezil arm, or a placebo arm. It aimed to treat them for 3 years. The results have received detailed discussion on Alzforum (Petersen et al., 2005). For her part, Emond took the trial database to address the issue of how many people dropped out and why. If those reasons were better understood, future trials could focus specific training and retention efforts on people at risk for early withdrawal.

Emond found that, overall, 30 percent of enrollees did not finish the trial. The donepezil group lost the most subjects at 36 percent. That was no surprise, as side effects of the study medication often cause a disproportionate dropout from the treatment group. But there is more going on than side effects: The placebo group also lost 25 percent, and Emond noticed that the incidence of side effects from donepezil was actually the same among people who stayed and those who dropped out. The biggest other factors influencing early dropout that emerged from her multivariate analysis included race/ethnicity (non-white and Hispanic participants dropped out at twice the rate than white participants) and what type of center conducted the study. Of the 69 participating sites, non-academic non-ADRC (Alzheimer Disease Research Center) sites were about twice as likely to lose study participants than were academic ADRCs. By contrast, how severely a subject’s cognition was impaired at baseline had nothing to do with dropout.

Moreover, Emond discovered that, frequently, it was the caregiver more than the study subject who was unwilling or unable to continue the study. The influence of the caregiver seems to have been great, Emond said. At the univariate level, more unmarried people than married people left prematurely, as did more women than men, raising the question of whether these two factors might be related to the caregiver’s motivation, as well. People who discontinued the study also cited as reasons their doubt about the effectiveness of the study medication, and some noted that they or their caregiver had chosen an alternate treatment. Finally, the burden of having to travel to the center for assessments every six months played a role, as well.

This initial study cannot offer a definitive answer to the question, and more large trial samples should be analyzed for dropout reasons. All the same, Emond said one safe recommendation even now might be that the staff at non-academic sites could use more training in actively supporting patients, and particularly caregivers, through the course of the trial.—Gabrielle Strobel.

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Madrid: Stressed Brain—SNP Carriers Can Have Cake and Eat It

27 Jul 2006

Famous as they are for their powerful effects throughout the body, circulating stress hormones such as the glucocorticoid cortisol still pose somewhat of a riddle when it comes to the brain. It’s clear that chronic high exposure to them causes detrimental effects in the brain of some people, and they have been implicated in AD repeatedly, but exactly how they might play out in age-related neurodegeneration has been harder to pin down. Now, an endocrinologist/internist has come across a lucky gene that sheds new light on the question. Elisabeth van Rossum, at Erasmus Medical Center in Rotterdam, the Netherlands, introduced it at the 10th International Conference on Alzheimer’s Disease and Related Disorders, held July 15 to 20 in Madrid. The gene is a variant of the glucocorticoid receptor that not only turns its carriers into wholesome babes and studmuffins, respectively, but also gives their brains an edge when it comes to aging. The ER22/23EK polymorphism, it turns out, blunts the gene expression consequences of cortisol. Its aging carriers are not only quicker on certain tests of psychomotor speed, they also enjoy a sizable protection against dementia and white matter lesions of the brain. About that free lunch? One price, known so far, for this boon appears to be a high risk for depression.

“When it comes to stress hormones, we all think of circulating glucocorticoid levels, high or low. This work shows differences between people are also a matter of one’s receptors. The receptor can determine the actual consequences a person sustains in their tissues from a given level of cortisol,” Van Rossum said.

First, a little background. Both biorhythms and stress drive the production of glucocorticoids. Those who remember their Biology 101 will recall the hypothalamic-pituitary-adrenal axis, which hosts the cycle leading from CRH via ACTH to the release of cortisol. This hormone exerts a wide range of effects on target tissues in the body and the brain, and then throttles its own production in a negative feedback loop. Brain consequences of cortisol are less well established than peripheral ones, but they include effects on dendritic branching, synapse formation, norepinephrine uptake, and glucose utilization. What’s important is that acute cortisol exposure enhances memory performance, but chronically elevated levels have been blamed for cognitive impairment, shrunken hippocampuses, and an increased risk of cerebrovascular problems (for a review, see Belanoff et al., 2001).

Glucocorticoids act through their receptors, and humans have two basic kinds. The type1 or mineraloreceptor (MR) regulates basal and circadian cortisol release, and the type2 (or GR) receptor regulates stress-dependent functions. Both occur in the hippocampus and other brain regions, though their distribution is not identical. When the receptor, which belongs to the nuclear receptor family, binds cortisol, it comes off its chaperone, dimerizes, enters the nucleus, and turns transcription of certain target genes on, of other genes off.

While studying this receptor, Van Rossum came across a variant in its second exon. This ER22/23EK polymorphism resides in the GR receptor, and 7 percent of the population carry at least one allele of it. Otherwise, the glucocorticoid receptor gene is largely terra incognita in AD research (see Alzgene). Carriers of this new polymorphism seem to enjoy benefits across the cardiovascular/metabolic range. They have more muscle and less fat than the rest of the population (in fact, the sole homozygous carrier Van Rossum knows is an avid triathlete), lower blood pressure, lower total and LDL cholesterol levels, lower C-reactive protein levels (this inflammatory marker is implicated in both heart disease and cognitive dysfunction), and increased insulin sensitivity.

“Since carriers are protected against harmful glucocorticoid effects in the body, we asked whether that held true in the brain,” said Van Rossum. To answer this question, Van Rossum studied two elderly populations. By screening 6,034 community elderly from the long-standing Rotterdam study, she found that the prevalence of dementia in this group was a whopping 86 percent lower among ER22/23EK carriers than non-carriers. When she excluded all subjects who were impaired at baseline and looked forward in time at the incidence of further cases of dementia over the course of the study, she again found carriers were less likely to develop dementia. Among a separate 1,011 people in the Rotterdam Scan Study, ER22/23EK carriers were less likely to have cerebral white matter lesions and brain infarctions than non-carriers. Those carriers who did had progressed less in a second scan 3 years later than non-carriers. Atrophy of the medial temporal lobe, or hippocampus, showed no association with glucocorticoid receptor variant, however. Finally, in a cognitive test battery, carriers had no better memory or overall cognitive function than non-carriers. However, they aced tests of coordination and psychomotor speed—the kind of reaction skills that fast-acting videogames train and that may reflect the integrity of white matter tracts.

Van Rossum does not know whether the effect of the gene variant is direct—that is, by dampening a direct toxic effect of glucocorticoids in neurons—or represents an indirect benefit of the carriers’ favorable metabolic and cardiovascular profile. Certainly, she said, small vessel disease can induce white matter lesions. What’s clear is that carriers express more of a longer form of the receptor that is relatively indolent when it comes to transactivating target genes. That means carriers sense the cortisol and react to it, but some of the downstream tissue consequences are softened. In the brain, this change occurs not so much in the hippocampus, but mostly in white matter. In essence, people carrying the variant may experience as many stressful episodes in their lives as anyone, but their bodies suffer the physical consequences less intensely, Van Rossum said.—Gabrielle Strobel.

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Madrid: BACE News Roundup, Part 1

27 Jul 2006

This is part 1 of our 3-part series. Also see part 2 and part 3, or download PDF.

Advances in understanding BACE1, the β-secretase enzyme relevant to Alzheimer disease, stood out as a notable trend at the 10th International Conference on Alzheimer’s Disease and Related Disorders (ICAD), held from July 15 to 20 in Madrid. At ICAD, bits and pieces of news were rustling up a fresh breeze in the air that came as a welcome change after a doldrums of sorts. Hope that this enzyme would serve as a new drug target first rose when researchers led by Martin Citron cloned it in 1999. At first blush, BACE1 looked like a safer target than its big brother γ-secretase, because knockout mice generated by Robert Vassar and several other independent groups all appeared largely normal. When Jordan Tang’s group solved the BACE1 crystal structure a year later—why, it seemed that all that was left to do was for clever drug designers to get busy and, presto, serve up a suitable small molecule drug. But the going got tough when BACE1 proved to be a recalcitrant drug target. What’s more, basic scientists began to whisper that BACE1 might not be as straightforward a target as initially thought. In Madrid, researchers for the first time presented a potent BACE1 inhibitor, fledgling immunotherapy approaches, and new data on its biology and potential as a biomarker. Read on for summaries of a plenary lecture and some of the 48 other presentations on BACE1. As always, Alzforum encourages presenters and attendees to amend our selected notes with their own.

In the plenary reviewing current knowledge on BACE, Citron, of Amgen in Thousand Oaks, California, first recapped that BACE1 and 2 are single transmembrane aspartyl proteases. They are related to the HIV retropepsin, which is a thoroughly studied drug target. One reason why BACE1 is less well understood, besides having been known for only six years, is that it undergoes numerous post-translational modifications that influence its activity in still-mysterious ways, Citron noted. Some things are known, however. BACE2 appears to play little, if any, role in AD pathogenesis. Cell biologists have pieced together that BACE1 traffics through the secretory pathway, moving from the trans-Golgi network to the plasma membrane, where it becomes pinched off into to endosomes and from there is retrieved again for further transport. BACE1 is thought to cleave APP most readily in endosomes and the trans-Golgi network, said Citron. It forms homodimers, and appears to do its work in lipid rafts.

One of the hottest questions in BACE research these days is whether BACE1 is upregulated in AD, and whether this upregulation comes as an epiphenomenon in late-stage AD or plays an early role and contributes to pathogenesis. Numerous reports have found that BACE1 activity increases with age and even more so in AD. Yet no familial AD loci containing BACE1 polymorphisms, much less AD-causing mutations in the BACE1 gene, have been found. This raises the underlying question of what regulates BACE1 expression. Many interactions of BACE1 and other proteins are on the map, including with reticulons, GGA proteins, and sorLa, but which ones participate in AD pathogenesis remains a puzzle. Other research has implicated BACE1 in an inflammatory feed-forward loop, and energy depletion as occurs in an atherosclerotic, underperfused brain is also thought to trigger BACE1.

Tang’s BACE1 crystal structure, and Amgen’s, too, showed that the active site comprises eight subsites, and that it would be difficult for a single small molecule drug to touch them all. Studying which of these sites a drug needs to hit has taken up much of the intervening time since 2000, Citron said. Only clinical trials will show whether BACE1 can be inhibited safely. In the interim, basic research has put potential concerns to watch for on the drug developers’ radar screen. Potential risks include that interfering with APP metabolism could narrow the therapeutic wiggle room if indeed Aβ turns out to perform an essential biological function, for example, in synaptic activity. Moreover, BACE1 has proven to cleave other substrates more readily than APP, and any physiological consequences of inhibiting these reactions remain unclear at present. The list of published substrates includes ST6Gal I, Psgl-1, LRP, and neuregulin-1 (see ARF related Madrid story).

BACE1 knockout mice are fertile, viable, and appear to age normally, but little is known about how they fare when stressed while aging. Some studies have identified subtle memory deficits, though this issue remains controversial, and some BACE1/2 double knockout mice tend to die early. In Madrid, Alex Harper and colleagues from GlaxoSmithKline in Harlow, Great Britain, reported that BACE1 knockouts had trouble gaining weight with age. Removing BACE1 protected the mice against the weight gain usually seen on a high-fat diet. Lack of BACE1 also appeared to increase the mice's insulin sensitivity in the face of a glucose challenge test, pointing to some still-mysterious metabolic role for BACE1. The BACE1 knockout mice also tended to die earlier than did wild-type controls. On the plus side, however, a different safety concern that has been raised about inhibiting APP cleavage by either BACE or its downstream successor γ-secretase appears less worrisome upon further inspection. It concerns a loss of physiological gene expression signaled by the intracellular tail of APP, aka AICD. A few genes, including neprilysin, KAI1, APP itself, or GSK3β, had been implicated as AICD target genes. Yet subsequent studies in different labs have struggled to reproduce these findings, and in Madrid, Sebastian Hébert in Bart de Strooper’s group in Leuven, Belgium, reported that in their hands, too, reducing AICD through secretase inhibition had no major effect on any of those genes (see also Hébert et al., 2006).—Gabrielle Strobel.

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Madrid: BACE News Roundup, Part 2

28 Jul 2006

This is part 2 of our 3-part series. Also see part 1 and part 3 or download PDF.

Anti-BACE Drugs Appear on Horizon
On BACE inhibition, Martin Citron of Amgen in Thousands Oaks, California, noted that more than 100 patent applications have been filed, and a growing number of non-peptide inhibitors are being published now that researchers have gained more experience with BACE1. High-throughput screening against BACE was largely unsuccessful as nothing of use stuck to BACE1, so companies switched over to rational design based on the crystal structure. In the patent literature, hydroxyethylamines are a central chemical theme of BACE1 inhibition. In Madrid, scientists for the first time began introducing compounds that appear to work in vivo.

James McCarthy, of Eli Lilly and Co. in Indianapolis, presented the first data on a BACE1 inhibitor that seems to have turned the corner in BACE1 drug development after years of frustration. McCarthy noted that the Eli Lilly team had long worked with a group of sulfone and sulfonamide compounds that are highly potent, but whose physicochemical properties stubbornly kept them outside the blood-brain barrier. The scientists then decided to scrap this class of compounds and instead go after others that started out with a slightly lesser affinity to BACE1 but with more attractive physicochemical properties, such as a lower molecular weight, lower polar surface area, and other parameters.

After describing stereochemical modifications to their initial lead compounds, McCarthy presented an experimental BACE1 inhibitor, LY2434074. This is the first publicly shown BACE1 inhibitor that enters the brain of PDAPP mice and reduces sAPPβ, the product of BACE1 cleavage, in cortex and hippocampus in a dose-dependent manner, McCarthy noted. The product of the alternative α cleavage that processes APP in the absence of BACE1, that is, sAPPα, went up in the brains of the injected mice. Aβ levels decreased in CSF and in plasma, as detailed in a subsequent poster presented by Patrick May of the same group.

Other scientists confirmed that this approach for the first time has demonstrated proof of principle for BACE1 inhibition in brain by a chemical given systemically. They also pointed out that the compound McCarthy presented likely is not the one the company is pursuing for clinical development. It had to be injected in rather large doses, implying problems with its oral availability or possibly its metabolism. Indeed, McCarthy replied in response to a question that Eli Lilly has more suitable compounds in hand. Colleagues from other drug development companies applauded Eli Lilly’s decision to present a potent structure. They added that other firms also have overcome some of the structural challenges posed by BACE1’s unwieldy active site. Indeed, Sethu Sankaranarayanan and colleagues from Merck’s team in West Point, Pennsylvania, presented evidence that intravenous injection of their own inhibitor lowers Aβ in the brain of Bruce Lamb’s human wild-type APP-transgenic mice.

Toward a BACE Vaccine
If anti-Aβ antibodies hold promise, why not hit BACE1 in the same way? Two groups reported progress toward this goal in Madrid. Wan-Pin Chang, in Jordan Tang’s group at the Oklahoma Medical Research Foundation in Oklahoma City, followed in the footsteps of Aβ immunotherapists and injected Tg2576 mice with BACE1. Chang’s prior experiments had detected reduced Aβ production in cultured cells treated with polyclonal anti-BACE1 antibodies, and he had also noticed that a fraction of injected anti-BACE1 antibodies entered the brain of mice. The underlying rationale for his approach, Chang said, would be that antibodies stick to BACE1 on neuronal cell surfaces and prevent its internalization into endosomes, where BACE1 cleavage of APP finds a conducive pH of 4.5.

In Madrid, Chang described a two-pronged study of active immunization with recombinant BACE1. A prevention arm began injecting BACE1 into Tg2576 mice repeatedly at 1 month of age, and a treatment arm began injecting BACE1 at 10 months, when plaques form. The scientists tracked the mice’s behavior and measured Aβ levels at 15 months or 23 months, respectively. In both study arms, but more so in the preventive one, the scientists measured rising antibody titers and waning Aβ40 and Aβ42 levels in serum and brain, as well as a reduced plaque load in brain as the immunization protocol progressed. Immunized mice outdid the untreated mice in negotiating the Morris water maze, Chang added. T cells, microglia, and astrocytes showed no sign of activation.

Michal Arbel, who works with Beka Solomon at Tel-Aviv University in Israel, took a different tack. Because BACE1 cleaves not only APP, Arbel works on devising an immunotherapy that interferes specifically with the BACE1-APP interaction rather than inhibiting or eliminating BACE1 altogether. Arbel develops antibodies directed against the BACE1 cleavage site on APP, which bind to human wild-type APP and human APP carrying the Swedish FAD mutation, but not to Aβ itself (see ARF related news story). (On a broader note: Scientists are realizing, to their surprise, that immunotherapy in mice works quite well across the board. Ajodeji Azuni, working with Einar Sigurdsson at New York University School of Medicine reported initial data of a tau vaccine. A P301L tauopathy mouse model responded to active vaccination with a phospho-tau peptide by mounting a tau-specific antibody response, showing less tau pathology, and performing better on some sensorimotor tasks.)—Gabrielle Strobel.

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Madrid: BACE News Roundup, Part 3

28 Jul 2006

This is part 3 of our 3-part series. Also see part 1 and part 2, or download PDF.

BACE Biology: Who Are Its Handlers?
Investigators are probing intensely the question of which other proteins interact with BACE. Perhaps the interacting proteins control BACE activity, and in this way influence Aβ production? Among the list of proteins thought to bind BACE, the nogo family is emerging as an intriguing group. Long studied for its ability to repulse outgrowing neurites in the brain and spinal cord, nogo and its receptor are forming a point of convergence between the formerly separate problems of axonal regeneration and AD pathology.

In Madrid, Rabinder Prinjha of the GlaxoSmithKline research group in Harlow, Great Britain, asked how nogo modulates APP processing via its link to BACE. Prinjha first noted that nogo-A and the nogo receptor are members of a large family of proteins, called reticulons, the physiological function of which is barely known. Four human reticulon genes are known to date with reticulon-4A being better known as nogo-A.

Nogo-A (reticulon4-A) and BACE occur together in the endoplasmic reticulum of cultured cells, Prinjha said. To test if they might interact in vivo, his team looked in APP- and APP/PS1-transgenic mouse lines and saw nogo-A and BACE both being upregulated in cortical areas surrounding plaques. Then the researchers tested the effect of nogo-A and its relatives in the reticulon family on APP processing, and noticed that they all affected Aβ production in distinct ways. In short, both overexpression and RNAi knockdown studies were used to confirm that reticulon3-A1 functions to decrease Aβ production, whereas reticulon4-A (nogo-A) increases it. The precise location of the proteins within the cell drove the effect. Prinjha did not address the mechanism of the interaction, but said that he suspects it to change the endocytosis or subcellular localization of BACE. Notably, besides the large pool in the ER, a second, smaller pool of nogo-A occurs on the cell surface. Similarly, Wataru Araki, of Japan’s National Institute of Neuroscience in Tokyo, reported that reticulon3 and reticulon4-B and -C bind BACE1. (Reticulons4-B/C are smaller isoforms of reticulon4). He suggested that these reticulons appear to inhibit BACE1’s ability to cleave APP by some interaction with BACE1 outside of the enzyme’s active site. For more on this up-and-coming topic, see Park et al., 2006; Gil et al., 2006; Yan et al., 2006).

Different sorting proteins also appear able to control APP processing by directing BACE¹s journey through intracellular compartments, primarily between endosomes and the trans-Golgi network. They include GGA1 (see ARF 2006 Eibsee conference report; He et al., 2005) and sortilin. On the latter, Gina Finan, working with Tae-Wan Kim at Columbia University in New York, reported that postmortem brain samples of AD patients contain less sortilin than controls. Sortilin forms a complex with BACE1 and appears to reduce Aβ secretion through its role in trafficking BACE1, the researchers suggest.

BACE in the Aging Brain: What Goes Wrong?
A number of groups have established that BACE activity tends to go up with age, and more steeply in AD. What could cause this? One hypothesis came to the fore when Robert Vassar’s group at Northwestern University in Chicago picked up findings from brain imaging, which has shown mild hypometabolism in the aging brain and even more in the AD brain. Wondering if the BACE increase might follow diminished perfusion—that is, insufficient oxygen and glucose supplies to the brain—Rodney Velliquette began to model energy starvation. Last November, the scientists reported that a single injection of chemical inhibitors of ATP generation had a long-lasting effect on the brain in that BACE levels (and Aβ production) shot up, and stayed up for a week (Velliquette et al., 2005). Reflecting Citron’s comment about the importance of post-translational modifications, Vassar noted that this increase occurred at the level of BACE protein, not gene expression.

But one injection does not model a slow disease such as AD, and in Madrid, Vassar followed up with a second, chronic study. It mildly starved ATP production in Tg2576 mice for a 3-month period, beginning prior to amyloid deposition at 9 months of age until 12 months of age when plaques are forming. BACE, Aβ levels, and plaque load all went up in the treated mice, Vassar reported. This suggests that, perhaps, sporadic AD could have an upstream, stress-related beginning that would drive BACE. “It is established that cerebral blood flow decreases in aging and particularly in AD brain. We do not know if this is just a correlation or a pre-existing driving force. It is something to look into because aging is the major risk factor in AD. As we age, cardiovascular disease increases and could put the brain under chronic energy stress,” Vassar speculated.

To sort out the time course of these events, Vassar’s group needed to make a better monoclonal antibody. All antibodies they could get their hands on were “dirty,” showing non-specific binding on Western blots and in brain tissue. With Skip Binder of Northwestern, who is noted for his skill in generating antibodies, the Chicago scientists used BACE knockout mice as immunization hosts, because they have never seen this protein. Out came a cleaner anti-BACE monoclonal antibody that recognizes but a single band on blots, Vassar said. This antibody helped the scientists characterize the BACE increase in various materials, including the Tg2576 and the group’s aggressive 5x-transgenic strain (see ARF SfN meeting story). The BACE antibody co-stains with neuronal markers but not astrocytic ones, particularly in dystrophic neurons. What’s more, BACE staining correlates with plaque development, and visualizes BACE around plaque cores. The BACE increase appears to be associated with Aβ42 deposition. This raises a chicken-and-egg question about which comes first in the course of AD pathogenesis: Does BACE first go up and induce Aβ42 deposition, or do Aβ42 deposits induce a secondary BACE increase? Vassar suspects a stress-induced feedback loop at play here.

The new antibody, Vassar hopes, will help with the analysis of what upstream factors can trigger BACE. Vassar particularly wonders whether any of those upstream factors would make for a good drug target so that, ultimately, a drug would become available that prevents only the age- or AD-related BACE increase, not all BACE altogether. “We may need BACE around for other functions,” Vassar said.

BACE: The Newest Biomarker?
Last but not least, one ICAD presentation moved research on BACE into the bustling realm of biomarker research. Yong Shen at Sun Health Research Institute in Sun City, Arizona, reported results of a collaborative study that, tantalizingly, suggested BACE1 might make for a decent biomarker. Shen’s lab was among the first to notice the BACE1 increase in AD brain, (see Li et al., 2004), a finding others have since confirmed. In Madrid, Shen used the same ELISA assay used for that study to assess BACE1 levels in the CSF of 80 sporadic AD cases, 59 MCI cases, and 69 age-matched controls. His data suggest that BACE levels in these MCI cases were twice as high as in the healthy controls but—surprisingly—returned to control levels once a person has progressed to overt AD. If replicated, this data would suggest that BACE1 might eventually serve to predict AD. Much remains to be sorted out about this prospect. In Shen’s hands, a BACE activity assay tracked with the ELISA for BACE detection and with total Aβ levels in that all three measures correlated in the CSF. These are early days for BACE1 research in CSF, but one study published to date would tend to confirm that enzymatically active BACE1 can be detected in human CSF (Verheijen et al., 2006).

All told, AD researchers still consider BACE the ideal target to test the amyloid hypothesis of Alzheimer disease. To be sure, BACE1 gets more complex the deeper the field digs into its function and regulation. Still, many scientists agree that inhibiting BACE1 may be cleaner than inhibiting γ-secretase, because that enzyme complex has a startling array of functions other than snipping APP once BACE is through with it. If a BACE inhibitor were to get into the brain and remove Aβ as expected, yet failed to improve dementia, then the amyloid hypothesis would be in serious jeopardy. With such inhibitors now coming online, this day of reckoning may be drawing nearer.—Gabrielle Strobel.

References

News Citations

Paper Citations

Park JH, Gimbel DA, GrandPre T, Lee JK, Kim JE, Li W, Lee DH, Strittmatter SM. Alzheimer precursor protein interaction with the Nogo-66 receptor reduces amyloid-beta plaque deposition. J Neurosci. 2006 Feb 1;26(5):1386-95. PubMed.
Gil V, Nicolas O, Mingorance A, Ureña JM, Tang BL, Hirata T, Sáez-Valero J, Ferrer I, Soriano E, Del Río JA. Nogo-A expression in the human hippocampus in normal aging and in Alzheimer disease. J Neuropathol Exp Neurol. 2006 May;65(5):433-44. PubMed.
Yan R, Shi Q, Hu X, Zhou X. Reticulon proteins: emerging players in neurodegenerative diseases. Cell Mol Life Sci. 2006 Apr;63(7-8):877-89. PubMed.
He X, Li F, Chang WP, Tang J. GGA proteins mediate the recycling pathway of memapsin 2 (BACE). J Biol Chem. 2005 Mar 25;280(12):11696-703. PubMed.
Velliquette RA, O'Connor T, Vassar R. Energy inhibition elevates beta-secretase levels and activity and is potentially amyloidogenic in APP transgenic mice: possible early events in Alzheimer's disease pathogenesis. J Neurosci. 2005 Nov 23;25(47):10874-83. PubMed.
Li R, Lindholm K, Yang LB, Yue X, Citron M, Yan R, Beach T, Sue L, Sabbagh M, Cai H, Wong P, Price D, Shen Y. Amyloid beta peptide load is correlated with increased beta-secretase activity in sporadic Alzheimer's disease patients. Proc Natl Acad Sci U S A. 2004 Mar 9;101(10):3632-7. PubMed.
Verheijen JH, Huisman LG, van Lent N, Neumann U, Paganetti P, Hack CE, Bouwman F, Lindeman J, Bollen EL, Hanemaaijer R. Detection of a soluble form of BACE-1 in human cerebrospinal fluid by a sensitive activity assay. Clin Chem. 2006 Jun;52(6):1168-74. PubMed.

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Madrid: News from the Vaccine Front—Phase 2 Postmortem, Part 1

31 Jul 2006

This is part 1 of our 3-part series. Also see part 2 and part 3 or download PDF.

Immunotherapy may not be the most eclectic choice of subject for those news aficionados who thirst for something completely different. Yet as the bearer of much hope and expectation throughout the field, the topic clearly warrants an update from every conference that offers news. And news there was at the 10th International Conference on Alzheimer’s Disease and Related Disorders, held July 15 to 20 in Madrid. Here is a quick summary of some of the presentations this reporter managed to attend. Whip out your notes, and send in additions or corrections! (And for those who still want something completely different, check out our closing story, in part 3 of this series, of Beka Solomon’s phage-only nose spray.)

Remains of AN1792—did it do some good, after all?
Followers of the field have heard proponents repeat the refrain that the trial of Elan/Wyeth’s active immunotherapy prototype was valuable despite the aseptic meningoencephalitis that halted it prematurely. Skeptics might say: “Yes, yes, we’ve heard that the trial proved immunotherapy can remove Aβ from human brain. And yes, we know that the responders of the Zurich cohort appear to be doing a tad better than the non-responders even if that’s but a handful of people.” But what most may not know yet is that an ongoing follow-up study of the entire trial population is beginning to show inklings that the same may be true throughout.

Mike Grundman, who formerly worked at the University of California, San Diego site of the Alzheimer Disease Cooperative Study (ADCS) and now is at Elan Pharmaceuticals in South San Francisco, heads the study. UCSD’s Leon Thal, who is chief investigator of the ADCS, encouraged Elan/Wyeth to conduct the study and presented the data available to date. At a packed symposium sponsored by Elan, Thal first reminded the audience that the trial participants, who received one or two injections of antigen, showed a small but significant improvement in three memory and three executive function tests (Gilman et al., 2005). But how well they fare beyond this assessment is unknown. Indeed, for a while scientists worried that there would not be any formal follow-up study of the trial participants. This study now under way asks these questions:

How are the participants doing clinically in the long term?
Do their antibody titers persist?
Do their puzzling MRI findings persist?
Did they develop anymore side effects after the study ended?

The study began this past January, 4.5 years after the trial began. It is being conducted blinded. Participants who are still able to visit the clinics are asked to come in, while others receive home visits or phone calls from investigators. Many of the study participants have progressed in their AD so that neuropsychological testing is no longer possible. For this reason, the investigators are applying a dependence scale and interviews with caregivers instead, though neuropsychology tests and MRI scans are still taken where possible. To date, about 40 percent of the original study—372 participants—have been contacted, and 85 of them agreed to participate. “We expect to have twice the data eventually that we have now,” Thal said.

What does that initial data suggest? This long after the trial, the mean MMSE for the treated group is 13, and for those on placebo it is 10. Both groups started out at a mean of 20. Seven of the eight responders (defined as people who produced antibodies in response to the vaccine) whose blood has been tested so far have retained “respectable” titers, Thal said. Ninety-five percent of placebo recipients now require total care, whereas 65 percent of the responders do. On activities of daily living, all groups declined comparably, and the clinical dementia rating (CDR) changed for the worse in all groups, as well. The ADAS-Cog data are too premature to make a statement, Thal noted. Follow-up MRI scans are slowly trickling in; the two available to date from the placebo group and three from the responder group no longer show the differential brain shrinkage that had startled the field after an initial post-dose scan (see Fox et al., 2005). There, too, the data is too premature to make a claim, Thal cautioned.

A curious finding popped up around the basic measure of age. At baseline, both treatment and placebo groups had a mean age of just over 71. At the 2006 follow-up, the treatment group had aged, as one would expect, to a mean age of 76; however, the placebo recipients who have been contacted so far still come in at a mean age of 71. Time has not stood still for them; rather, it is possible that the older AD patients among the placebo group might have died at a higher rate than those in the treatment group.

Finally, no further cases of encephalitis beyond the 18 reported ones have cropped up since then, nor did other drug-related serious side effects, Thal noted. He emphasized that even though the data show a trend favoring patients who received AN1792 and responded to it, this data is highly preliminary and not yet fit for conclusions.

For his part, Roger Nitsch of University of Zurich offered further tidbits of data on the AN1792 trial Zurich cohort that his group is following separately. On the meningoencephalitis, Nitsch noted that of the three Zurich patients who developed it, two have antibodies and their Alzheimer disease remains stable to date, whereas one did not have antibodies and died three years after the immunization.

Nitsch then described a fourth autopsy case in addition to three published ones, from Southampton (Nicoll et al., 2003), Barcelona (Ferrer et al., 2004), and Arizona (Masliah et al., 2005). A 79-year-old man with a 7-year history of dementia from the Zurich group stopped speaking after a final MMSE of 12, then died four years after having received two shots of vaccine. He did not suffer the encephalitis and had low antibody titers in his blood and CSF. His Aβ levels in frontal and temporal cortex were low, as was amyloid deposition, Nitsch reported. Amyloid plaques had microglia around them, which stained with the 6E10 Aβ antibody, indicating the cells were ingesting the amyloid. This patient showed severe neuronal loss, gliosis, but no cerebral amyloid angiopathy. Alzforum has followed conference updates on this trial closely; for recent news, see ARF Eibsee report; ARF Sorrento story; and ARF St. Moritz story).—Gabrielle Strobel.

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References

News Citations

Paper Citations

Gilman S, Koller M, Black RS, Jenkins L, Griffith SG, Fox NC, Eisner L, Kirby L, Rovira MB, Forette F, Orgogozo JM, . Clinical effects of Abeta immunization (AN1792) in patients with AD in an interrupted trial. Neurology. 2005 May 10;64(9):1553-62. PubMed.
Fox NC, Black RS, Gilman S, Rossor MN, Griffith SG, Jenkins L, Koller M. Effects of Abeta immunization (AN1792) on MRI measures of cerebral volume in Alzheimer disease. Neurology. 2005 May 10;64(9):1563-72. PubMed.
Nicoll JA, Wilkinson D, Holmes C, Steart P, Markham H, Weller RO. Neuropathology of human Alzheimer disease after immunization with amyloid-beta peptide: a case report. Nat Med. 2003 Apr;9(4):448-52. PubMed.
Ferrer I, Boada Rovira M, Sánchez Guerra ML, Rey MJ, Costa-Jussá F. Neuropathology and pathogenesis of encephalitis following amyloid-beta immunization in Alzheimer's disease. Brain Pathol. 2004 Jan;14(1):11-20. PubMed.
Masliah E, Hansen L, Adame A, Crews L, Bard F, Lee C, Seubert P, Games D, Kirby L, Schenk D. Abeta vaccination effects on plaque pathology in the absence of encephalitis in Alzheimer disease. Neurology. 2005 Jan 11;64(1):129-31. PubMed.

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Madrid: News from the Vaccine Front—Phase 1 Hopefuls

01 Aug 2006

This is part 2 of our 3-part series. Also see part 1 and part 3 or download PDF.

Whatever the fallout of the AN1792 debacle, it has not stopped commercial or academic interest in immunotherapy. Elan Pharmaceuticals and its partner Wyeth have moved a passive vaccine based on a version of the plaque-binding 3D6 antibody into a multicenter phase 2 trial. Other companies, too, are beginning to report first clinical experiences with their own vaccines. In Madrid, two such vaccines that have entered human studies were made public in some detail, one by Novartis and its partner Cytos Biotechnology in Zurich, Switzerland, and another by Eli Lilly and Co. in Indianapolis.

Matthias Staufenbiel, of Novartis Institutes for BioMedical Research in Basel, disclosed preclinical data for a new active vaccine, dubbed CAD106, that the company is currently testing in humans in Sweden. Staufenbiel described the vaccine as having grown out of efforts to avoid the T cell activation widely blamed for the inflammatory side effect that stymied Elan/Wyeth’s AN1792 trial. The Swiss scientists fashioned a therapeutic vaccine out of the first six N-terminal amino acids of Aβ—a snippet able to stimulate human B cells but devoid of epitopes that arouse human T cells. The trick lies in hitching this stub of Aβ to a virus-like particle that generates the sort of T cell reaction needed to mount a full-fledged antibody response and to break the body’s self-tolerance against Aβ (see also Li et al., 2004). The virus-like particle comes with the added benefit that it is sufficiently potent at marshaling the immune system’s troops as to render additional adjuvants unnecessary.

All mice injected with this vaccine, young and old, mounted an antibody response with high titers, Staufenbiel said. Rabbits did, too. The researchers used two different strains of APP-transgenic mice—one depositing mostly diffuse amyloid but none around blood vessels, and one depositing plaques on blood vessel walls as well as between neurons (see part 3 for more on vessel amyloid). Both mice strains had reductions in Aβ levels and in their predicted amyloid pathology. Vaccination of young versus old mice indicated that the vaccine is more potent at reducing the accumulation of new plaques than removing existing plaques, Staufenbiel said. CAD106 removes parenchymal and vascular amyloid but, importantly, it does not cause the microhemorrhages that some other vaccines are reported to have caused in mice, Staufenbiel noted.

Tests with spleen cells isolated from immunized mice showed that CAD106 does not appear to stimulate T cells. Studies of CAD106 in rhesus monkeys confirmed the mouse data in that the primate antibodies stain plaques in APP-transgenic mouse brain and AD postmortem brain, do not cross-react with APP, and block Aβ-induced toxicity in cell-based assays. Results from the phase 1 trial are expected in 2007, according to a press release issued by Cytos Biotechnology.

Lilly’s Eric Siemers described an initial single-dose trial of a so-called capture antibody. The rationale of this trial is based on the peripheral sink hypothesis. The approach grew out of a widely noted study showing that a single injection of the m266 antibody, which sticks with high affinity to soluble Aβ, improved cognition in mice overnight (Dodart et al., 2002). Together with a paper describing how peripheral injection of the m266 antibody bound plasma Aβ (DeMattos et al., 2001), this line of investigation raised hope that certain antibodies might be able to “draw” Aβ out of the brain by way of shifting a series of presumably connected transport equilibria across the brain, CSF, and blood toward the side of the blood. The vision of a peripheral therapy for AD, or perhaps a diagnostic test similar to an insulin challenge shot for diabetes, took shape.

After reviewing preclinical research in PDAPP mice and rats, Siemers described the first human study in Lilly’s clinical program using LY206430. This humanized version of the m266 monoclonal antibody binds to Aβ16-23, the peptide’s midsection. Investigators infused one of four doses into the veins of four AD patients per dose, plus three placebo controls. Their mean age was 69, mean MMSE 20. The investigators took CSF samples and ran MRI scans at baseline and 21 days later. They tested the participants’ performance in the ADAS-Cog battery at baseline, three days later to check for immediate effects as seen in the animal studies, and again at 21 days. The patients were then followed for one year.

This is what the Lilly scientists found: The antibody appeared safe in terms of standard laboratory values such as liver enzymes, and it also produced no evidence of the side effects that make AD vaccinologists jittery these days, that is, inflammation or microhemorrhage. The antibody produced a typical infusion reaction in five patients, which soon resolved on its own, Siemers said.

Plasma Aβ40 levels shot up between roughly 150- to 600-fold, depending on how much antibody was infused. CSF Aβ40 levels nudged up roughly 1.2- to 1.8-fold; Siemers estimated that 0.1 percent of this antibody enters the CSF and binds Aβ there. He added that the ADAS-Cog values signaled a hint of improvement but that this single-dose study was not designed to support a statement on efficacy.

Other scientists noted that more work lies ahead for this approach. One question they debate concerns the origin and precise nature of the Aβ that binds to the antibody in plasma. How much of it comes from the brain as compared to coming from other large organs known to secrete Aβ, such as muscle? The body-wide economy of Aβ will become clearer as this approach progresses. In that regard, a poster by Yona Levites, in Todd Golde’s group at the Mayo Clinic in Jacksonville, showed that peripheral injection of an anti-Aβ1-16 monoclonal antibody caused a steep rise in plasma Aβ but did not change brain Aβ in wild-type and young APP-transgenic mice. This raises questions about stabilization of peripheral Aβ by the antibody, and appears to complicate the notion of a diagnostic challenge test based on injected antibody. Peter Seubert, of Elan Pharmaceuticals in South San Francisco, reported that in his group’s hands, even long-term treatment of PDAPP mice with the m266 capture antibody did not lower the mice’s cerebral amyloid burden or improve neuritic dystrophy, but it did reduce soluble brain Aβ levels significantly and produced some improvement in tests of acute function and of synaptic health. Seubert's and Golde’s findings would suggest that a capture antibody might prolong the peripheral clearance rate of Aβ by stabilizing Aβ in the blood.

On a more general note, the plasma half-life of all antibodies that are being developed as passive AD vaccines is roughly around a month. This raises questions about how often it would have to be infused and what price for such a therapy health care systems would be able, or willing, to sustain.—Gabrielle Strobel.

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References

News Citations

Paper Citations

Li Q, Cao C, Chackerian B, Schiller J, Gordon M, Ugen KE, Morgan D. Overcoming antigen masking of anti-amyloidbeta antibodies reveals breaking of B cell tolerance by virus-like particles in amyloidbeta immunized amyloid precursor protein transgenic mice. BMC Neurosci. 2004 Jun 8;5:21. PubMed.
Dodart JC, Bales KR, Gannon KS, Greene SJ, DeMattos RB, Mathis C, DeLong CA, Wu S, Wu X, Holtzman DM, Paul SM. Immunization reverses memory deficits without reducing brain Abeta burden in Alzheimer's disease model. Nat Neurosci. 2002 May;5(5):452-7. PubMed.
Demattos RB, Bales KR, Cummins DJ, Paul SM, Holtzman DM. Brain to plasma amyloid-beta efflux: a measure of brain amyloid burden in a mouse model of Alzheimer's disease. Science. 2002 Mar 22;295(5563):2264-7. PubMed.

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Madrid: News from the Vaccine Front—Bloody Complicated?

01 Aug 2006

This concludes our 3-part series. Also see part 1 and part 2 or download PDF.

Scientists continue to debate the relative merit of using N-terminal versus mid-section or C-terminal antibodies, and of using antibodies against soluble versus against fibrillar Aβ. The discussion turns, in part, on the issue of microhemorrhages. Concern about small bleeds inside the brain arose when several groups began reporting them in the brains of immunized mice, especially mice who carry a heavy load of amyloid deposits in the blood vessel walls of their brains, not just in the parenchymal spaces. Called cerebral amyloid angiopathy (CAA), this pathology exists in the majority of AD patients. CAA is estimated to occur in up to 30 percent of elderly people, though that number varies. CAA can cause hemorrhagic strokes and cognitive impairment, and by itself is thought to cause numerous small bleeds that often go unnoticed.

The condition is attracting increasing attention from researchers. At the 10th ICAD meeting, held July 15 to 20 in Madrid, 23 presentations dealt with the topic of CAA and microbleeds. Presentations ranged from one showing that CAA pathology intensifies along with AD pathology, albeit in somewhat different brain regions, to another suggesting that the amyloid imaging agent PIB-PET can detect the region-specific CAA pattern in non-demented, living people. At the epidemiological level, scientists led by Meike Vernooij and colleagues at Erasmus University Medical School in Rotterdam used MRI to measure the frequency of microbleeds in 491 community-based participants of the Rotterdam Scan study. They found that 17 percent had such bleeds in their brains. Older people were at higher risk than younger people, and high blood pressure and use of thrombolytic agents upped the risk some more.

The worry with regard to CAA and AD immunotherapy is that clearance of blood vessel amyloid by anti-Aβ antibodies might lead to ruptures of the already dysfunctional vessel wall (Pfeifer et al., 2002; Burbach et al., 2006). A related worry is that overly fast removal of parenchymal amyloid would overwhelm the clearance capacity at nearby blood vessels and lead to renewed deposition of the amyloid there (Wilcock et al., 2004). Intriguingly, the sugar groups decorating a given antibody appear to influence this phenomenon (Wilcock et al., 2006). RN1219, a C-terminal Aβ antibody being developed by the San Francisco biotechnology company Rinat Neuroscience, a Genentech spinoff, is said to have an edge on that score.

This issue is in flux, and in Madrid, Sally Schroeter of Elan Pharmaceuticals addressed it with data from a 6-month study comparing the effects of several Aβ antibodies on CAA in the PDAPP mouse model. In short, Schroeter reported that, to her surprise, the 3D6 antibody, which recognizes fibrillar Aβ, not only did not worsen CAA, but instead cleared it in a dose-dependent fashion. The m266 capture antibody predictably had no effect on CAA. A multiphoton imaging study by Claudia Prada and colleagues at Massachusetts General Hospital in Charlestown paralleled this data by observing in live mice that a different antibody given to Tg2576 mice caused CAA to regress.

Furthermore, in Schroeter’s study, microhemorrhages did occur in conjunction with the CAA clearance, she acknowledged, but could be limited by reducing the antibody dose, essentially tuning down amyloid clearance to a manageable rate. David Morgan, of University of Southern Florida, who has collaborated with Rinat Neuroscience but has no financial interest in the company, pointed out that at the age at which Schroeter and colleagues began treating the PDAPP mice—12 months—the mice’s brains do not yet have a full load of parenchymal amyloid and also have less CAA than many AD patients. Repeating the study in older mice might more closely mimic the amyloid and CAA burden of AD patients and would test the findings raised by Wilcock et al. and Pfeifer et al. under more comparable conditions. For a prior comparison of 3D6 and m266 in older PDAPP mice, led by Ron DeMattos at Lilly, see Racke et al., 2005.

Yet another factor that young PDAPP mice represent poorly is the inflammation in blood vessels laden with CAA. In Madrid, Manuel Buttini and colleagues of Elan Pharmaceuticals in South San Francisco characterized inflammatory infiltrates in brain blood vessels of elderly people with AD and young normal controls, and found that more than twice the percentage of CAA vessels than CAA-free vessels had monocytes and T cells near them. “Age still is the most important risk factor in AD,” Morgan summed up.

But Morgan also emphasized that it’s all but clear how important the microbleeds will prove to be clinically in people. Clot busters such as the stroke treatment tissue plasminogen activator are known to cause microbleeds and, at least in acute situations, the risk is tolerated. Morgan’s antibody-treated mice, for all they are worth, retained the behavioral improvement despite having more of these bleeds. “Minute hemorrhages are not the big worry. Large ones are,” Morgan said. In summary, numerous scientists asked about this issue tended to agree that they expected some form of immunotherapy to slow progression of AD. But they also worry that some patients who receive these therapies for extended periods of time might develop larger hemorrhages, forcing the FDA’s hand against the approach.

Little phage to the rescue?

If reading all this leaves you scratching your head about how the pitfalls of AD immunotherapy can possibly be avoided, you are not alone. Some scientists are also looking for alternatives. One of them, Beka Solomon of Tel Aviv University, who has worked on AD immunotherapy for a decade (Solomon et al., 1997) happily announced a fortuitous observation that led her to begin exploring such an alternative. Solomon has long studied the potential of bacteriophages for ferrying either Aβ antigens or antibodies into the brain. It was during one of those studies that a control—naked phage—surprised her when it performed just as well as the study drug—phage studded with single-chain Aβ antibody—on her measures of reduction in amyloid burden. It seemed to do so more safely, too. Perhaps this humble life form could make for a treatment?

Bacteriophages come in two basic varieties. The better-known ones that lyse bacteria have been used as an alternative to antibiotics in the former Soviet Union, but Solomon uses the non-lytic phages. At almost a micrometer in length but only nine nanometers in width, they look like microscopic filaments of DNA packaged into a narrow protein sheath. Their physicochemical properties enable them to slip through membranes easily. In prior years, Solomon has explored the phages’ potential to carry foreign peptides into the brain of animals (e.g., Lavie et al., 2004), to serve as immunogens (Frenkel et al., 2000), and to exert effects in the brain when sprayed into the nose (Frenkel and Solomon, 2002). (Incidentally, the scent of intranasal delivery wafted through the ICAD conference, with some presentations extolling the benefits of this route and others describing its use in the delivery of specific experimental drugs. Examples include a talk by Suzanne Craft at the University of Washington, Seattle, on intranasal insulin and verbal memory in early AD, and another by Illana Gozes of Tel Aviv University on the neuroprotective peptide NAP, which acts to stabilize microtubules and is in early clinical trials with an intranasal formulation.)

In her studies with the phages, Solomon discovered that they alone, even without sporting Aβ or an antibody at their tips, had useful effects in AD models. Apparently, their size and structure allow them not only to penetrate the brain when given through the nose but also to intercalate into the β-sheet structure of amyloid and disrupt it. Electron microscopy images showed immuno-gold labeled Aβ fibrils alone, and amorphous Aβ aggregates in the presence of filamentous phage. The phages’ threadlike shape did the trick, because when hooked into spheres, the phages no longer busted amyloid fibrils.

Solomon then showed in-vitro data on the phage’s disaggregating properties, and on their ability to stain amyloid plaques. Injection of filamentous phage alone into the brains of Tg2576 mice reduced the mice’s amyloid load over the course of three days. A subsequent one-year study of biweekly, then monthly, administration of phage sprayed up the noses of PDAPP mice improved the mice’s memory performance in an object recognition test and a smell test. No water maze data were given. The phage also reduced the amyloid burden in the mice’s brain and increased their synaptophysin levels, Solomon said. Notably, unlike the untreated transgenic mice, the phage-treated mice had no astrocytosis in their hippocampuses. Microgliosis showed no difference between the groups. Microhemorrhages were undetectable in the phage-treated mice, Solomon stressed. Peripheral organs also suffered no ill effects, in keeping with the established safety profile of these organisms. The phages leave the body within 3 weeks, Solomon said. They exit the brain with the help of microglia and are concomitantly eliminated from the body by urine and feces.

Solomon did not show data about the detailed in vitro-in vivo correlations, pharmacokinetic data, and dose-effect curves that would become necessary once drug developers became interested in this approach. Likewise, other scientists were curious to see short-term in-vivo studies that track how Aβ levels change in brain and CSF soon after phage delivery. Presumably, levels of Aβ would initially go up as it gets freed from fibrils, before eventually being cleared. Solomon said this was her initial presentation on the approach and much remains to be done. But already, it is clear that the phages are safe and ubiquitous in the environment, she said. “You and I can pick them up by swallowing a bit of water while swimming outdoors, and we won’t even know it,” Solomon added. They are dirt cheap, too.—Gabrielle Strobel.

References

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Wilcock DM, Rojiani A, Rosenthal A, Subbarao S, Freeman MJ, Gordon MN, Morgan D. Passive immunotherapy against Abeta in aged APP-transgenic mice reverses cognitive deficits and depletes parenchymal amyloid deposits in spite of increased vascular amyloid and microhemorrhage. J Neuroinflammation. 2004 Dec 8;1(1):24. PubMed.
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Madrid: γ-secretase Dimers, A New Model, A Drug in Clinic

02 Aug 2006

The 10th International Conference on Alzheimer’s Disease and Related Disorders, held earlier this month in Madrid, offered its share of data on the intricate ways by which the γ-secretase enzyme complex spews out a slew of different forms of Aβ peptide, some of which are more inclined to damage neurons than are others. Alzforum has summarized some new thinking on γ-secretase this past May (see ARF Eibsee meeting report), therefore, our present ICAD coverage will pick out a few selected presentations that add to our recent coverage. Read on for a notable mechanistic insight and news on efforts to inhibit this enzyme, including some human data. As always, write in about what this observer has missed.

In their hunt for compounds that divert γ-secretase away from generating Aβ42 without affecting its ability to cleave Notch, several groups independently are scrutinizing a small area of presenilin-substrate interaction that might be amenable to allosteric modulation. By studying APP dimerization, Gerd Multhaup of the Free University, Berlin, arrived at this very spot, as well. With graduate student Lisa Munter, Multhaup discovered a potentially critical dimerization motif on APP that influences the sequential cleavages of the γ-secretase substrate. By doing so, it helps determine what forms of Aβ come out of this process.

APP dimerization itself, by two different points of contact between monomers, had been published before (see Beher et al., 1996; Scheuermann et al., 2001). In her talk, Munter first noted that she and her colleagues had discovered a third, new site of contact between two APP molecules that mediates dimerization. This site includes three consecutive GxxxG motifs extending into the transmembrane sequence. Such motifs are known to promote interaction of protein helices, and a recent Alzforum Discussion raised the question of whether they might mediate Aβ toxicity. On APP, glycines 29 and 33 within the Aβ sequence represent particularly important residues of this GxxxG series, Munter and Multhaup observed. The scientists found that when they mutated the glycine 33 of APP, the mutant form not only failed to dimerize but also shifted Aβ generation away from Aβ42 and toward more of the smaller peptides Aβ37 and 38. Intriguingly, some NSAIDs have exactly the same effect on Aβ peptide distribution. This raises the question of whether they might do so by weakening the dimerization of the γ-secretase’s APP substrate C99, Munter and Multhaup speculated.

Here is how the German scientists explain their finding. The γ-secretase complex has been shown to process its C99 substrate by a series of sequential ε-, ζ-, and γ-cleavages. This is well-documented but has not yet been widely incorporated into people’s thinking about how this unusual protein machine works. Proceeding from the carboxyl terminus toward the N-terminus, the cleavages generate successively shorter Aβ peptides, from 49/48 to 46/45, to 43/40 and 42 (Qi-Takahara et al., 2005; Zhao et al., 2005; Chandu and Kopan, 2006). Quite possibly, ε and ζ cleavage proceed regardless of whether the substrate is a monomer or a dimer, yet after that, the dimer poses a steric obstacle that prevents the substrate from moving farther through the catalytic site. In essence, the idea is that dimers would stall successive cleavages at the Aβ42 site. Successive degradation of C99 would represent a physiological function of γ-secretase, and mutations that stabilize dimerization would increase production of an intermediate product of this normal degradation, namely Aβ42. By contrast, G29/G33 mutations—or perhaps some future therapeutic compounds—preventing dimerization would allow the substrate to be further processed into shorter peptides before it can leave the membrane.

γ-secretase inhibitors—alive and well?
When early classes of γ-secretase inhibitors began showing toxic side effects, especially severe ones in the intestinal tract and the maturation of lymphocytes, it looked like the approach might be dead in the water. Yet a more apt analogy might have been to not throw the baby out with the bathwater. It is true that companies are focusing on finding compounds that modulate γ-secretase rather than inhibit it outright, but they have quietly continued studying inhibitors, as well. “We really still would like to use these drugs. Some of them have great properties, if only we can get rid of their mechanism-based side effects,” said Christian Czech of Hoffmann-La Roche’s CNS Research group in Basel, Switzerland. With this comment, he echoed the sentiment of colleagues in other firms, and some in academia, as well.

One ray of hope in this area was implicit from one of a series of posters presented by Elan Pharmaceuticals in South San Francisco. In one, Guriqbal Basi and colleagues described an analysis of the structural determinants of different inhibitor classes. The upshot of this characterization of exactly which inhibitor binds to which amino acid residues on a given presenilin was that certain kinds of inhibitor clearly distinguish between presenilin-1 and presenilin-2. In this case, sulfonamides, as represented by a compound called BMS299897, acted more potently on the former than the latter, whereas DAPT and some other inhibitors acted comparably on both.

One can suspect that none of the inhibitors shown on the poster will be tomorrow’s drug, but the data is nonetheless important because it means that drugs can be found that will selectively block only certain kinds of γ-secretase complex. This might point a way toward finding a drug that acts in the brain but not in the gut, for example. Several labs have shown that different tissues of the body assemble the presenilin and Aph-1 isoforms in the human genome into γ-secretase complexes of distinct composition, and perhaps also distinct function.

Elan scientists showed more data on some new γ-secretase inhibitor series, and they were not alone. Nearly a dozen ICAD presentations dealt with γ-secretase inhibitors, some from academic labs, others from company labs such as Wyeth and Bristol-Myers Squibb. For his part, Mark Shearman, of Merck Research Laboratories, Boston, Massachusetts, described how scientists at his company used rational drug discovery to make new γ-secretase inhibitors. These inhibitors showed their ability to lower brain Aβ40 and 42 consistently when tested in Tg2576 mice, mice expressing full-length human APP off a yeast artificial chromosome, in guinea pigs, and then in a primate model.

To bridge the step from small animal to human, and to help predict an effective dose range in humans, the Merck scientists developed a rhesus monkey model that allowed them to monitor how a given inhibitor regimen affected Aβ levels in several fluid compartments over the course of days, weeks, even months. The partial nature of prior data sets—plasma time courses in this study, a spinal tap in that study—had made it difficult to compare between studies and especially to track how a given drug changed the separate pools of Aβ in the body over time (see ARF related news story on biomarker validation). Merck’s primate model overcomes this limitation. It is simple: a surgeon implants a catheter into the cisterna magna, a reservoir of cerebrospinal fluid at the lower back of the head. The catheter connects to a sterile port that allows the scientists to drain a few drops of CSF for analysis non-invasively whenever needed without having to sedate the animal. A cohort of about 30 monkeys has been living with this catheter for more than a year without ill effects, Merck scientists said. These animals have enabled the scientists to compare the effects of their γ-secretase inhibitors to those of γ-secretase modulators on Aβ40 and 42 levels in plasma and CSF. On a poster, Maria Michener, working with Lynn Cook and colleagues, showed data for 6-day studies, but internally the scientists have followed the effects of their compounds for longer.

Moreover, in an initial human safety and pharmacokinetics/ pharmacodynamics study presented by Laura Rosen, a lumbar catheter was implanted into 27 human volunteers at two clinical study sites in order to test a single oral dose of the MK-0752 γ-secretase inhibitor. CSF was then collected for up to 30 hours afterwards, as well as blood samples for up to 4 days. Besides being well-tolerated in this small study, MK-0752 made it into the CSF, where it replicated a dose-dependent Aβ40 reduction as seen in the primate model, Rosen reported. “We believe the basic biology of APP metabolism, and its monitoring in plasma and CSF in response to our inhibitors, is transferable from mouse through guinea pig through primates to the human,” Shearman said in an evening lecture detailing some of the company’s efforts on γ-secretase inhibitors, modulators, and BACE inhibitors. The MK-0752 γ-secretase inhibitor is currently in phase 1 trials as a candidate disease-modifying agent, and other compounds have recently been published (see Jelley et al., 2006; Shaw et al., 2006; Best et al., 2006).—Gabrielle Strobel.

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Beher D, Hesse L, Masters CL, Multhaup G. Regulation of amyloid protein precursor (APP) binding to collagen and mapping of the binding sites on APP and collagen type I. J Biol Chem. 1996 Jan 19;271(3):1613-20. PubMed.
Scheuermann S, Hambsch B, Hesse L, Stumm J, Schmidt C, Beher D, Bayer TA, Beyreuther K, Multhaup G. Homodimerization of amyloid precursor protein and its implication in the amyloidogenic pathway of Alzheimer's disease. J Biol Chem. 2001 Sep 7;276(36):33923-9. PubMed.
Qi-Takahara Y, Morishima-Kawashima M, Tanimura Y, Dolios G, Hirotani N, Horikoshi Y, Kametani F, Maeda M, Saido TC, Wang R, Ihara Y. Longer forms of amyloid beta protein: implications for the mechanism of intramembrane cleavage by gamma-secretase. J Neurosci. 2005 Jan 12;25(2):436-45. PubMed.
Zhao G, Cui MZ, Mao G, Dong Y, Tan J, Sun L, Xu X. gamma-Cleavage is dependent on zeta-cleavage during the proteolytic processing of amyloid precursor protein within its transmembrane domain. J Biol Chem. 2005 Nov 11;280(45):37689-97. PubMed.
Jelley RA, Elliott J, Gibson KR, Harrison T, Beher D, Clarke EE, Lewis HD, Shearman M, Wrigley JD. 3-Substituted gem-cyclohexane sulfone based gamma-secretase inhibitors for Alzheimer's disease: conformational analysis and biological activity. Bioorg Med Chem Lett. 2006 Jul 15;16(14):3839-42. PubMed.
Shaw D, Best J, Dinnell K, Nadin A, Shearman M, Pattison C, Peachey J, Reilly M, Williams B, Wrigley J, Harrison T. 3,4-Fused cyclohexyl sulfones as gamma-secretase inhibitors. Bioorg Med Chem Lett. 2006 Jun 1;16(11):3073-7. PubMed.
Best JD, Jay MT, Otu F, Churcher I, Reilly M, Morentin-Gutierrez P, Pattison C, Harrison T, Shearman MS, Atack JR. In vivo characterization of Abeta(40) changes in brain and cerebrospinal fluid using the novel gamma-secretase inhibitor N-[cis-4-[(4-chlorophenyl)sulfonyl]-4-(2,5-difluorophenyl)cyclohexyl]-1,1,1-trifluoromethanesulfonamide (MRK-560) in the rat. J Pharmacol Exp Ther. 2006 May;317(2):786-90. PubMed.

Madrid: Clinical Trials Update—Where Do Things Stand?

12 Sep 2006

Nerve growth factor gene therapy, diabetes drugs, a pooled antibody stew, hormones, and radical scavengers—it’s not like all researchers are focusing squarely on immunotherapy or secretase inhibitors in their search for better Alzheimer drugs. Last July at the 10th International Conference on Alzheimer’s Disease and Related Disorders in Madrid, several dozen groups presented data on human tests—ranging from tiny pilots to large controlled cohorts—of a panoply of approaches. None produced spectacular “light-bulb” effects, but some bear watching. The news moment of ICAD has passed as researchers are already beginning to look toward the Society for Neuroscience meeting next month in Atlanta, Georgia. But since trial data appear in the formal literature with long delays, if at all, here is one more ICAD roundup of selected trial presentations. As always, we invite additions and corrections. If this writer missed a presentation on “your” trial, we’ll gladly add your comments.

Nerve Growth Factor Gene Therapy
Mark Tuszynski, at the University of California, San Diego, explores the potential of delivering nerve growth factor specifically to cholinergic neurons of the forebrain’s nucleus basalis to protect neurons against degeneration in Alzheimer disease. Tuszynski has for years persisted against skeptics who maintain that his method of choice—gene therapy in the brain—is not ready for prime time, too dangerous, too expensive, and too high-tech. All of that might be so, except the approach looks like it might just work. That, at least, is where things stand currently.

Tuszynski noted that prior work on models ranging from aging to injury, excitotoxicity, and amyloid overexpression all substantiate the rationale of using nerve growth factor in an effort to stem neuronal death and support repair. The leap to success in humans, Tuszynski believes, hinges on delivering this potent agent to its designated area but nowhere else. Last year, Alzforum covered formal publication of phase 1 results, to 22 months of follow-up. In Madrid, Tuszynski added data for up to five years after the beginning of this trial.

This far out, the procedure appears safe when administered under anesthesia, causing no pain or weight loss. As is typical during neurosurgical procedures, the patients initially were awake during the surgery; however, having AD, some forgot to stay still and moved, causing one person’s death 5 weeks later from deep vein thrombosis following a brain hemorrhage. The person’s autopsy results suggested robust NGF expression and cholinergic axons sprouting into the graft area. “There was a classic trophic response to NGF, so AD neurons can respond to this growth factor in the aged brain,” Tuszynski said. Subsequent surgeries were done under anesthesia, without complication.

The long-term data showed that the patients who received bilateral injections of NGF-releasing cells maintained stable cognitive performance in MMSE and ADAS-Cog, as well as their ability to live independently. Patients who received injections into only one side of their brain declined on these scores. While Tuszynski considers these data encouraging, he noted that the absence of controls and the small number of patients preclude stronger conclusions. At this stage, Ceregene, a California biotech company he co-founded, has filed an investigational new drug (IND) application with the FDA for a phase 1/2 trial. Pending approval, it will test a new gene delivery method using adeno-associated virus (AAV), not the older method of using a patient’s own, individually engineered fibroblasts. The AAV method hopefully will prove to be simpler and ensure sustained gene expression over time, Tuszynski noted. In addition, six patients have been enrolled in a phase 1 trial overseen by David Bennett at Rush University Medical Center in Chicago, and Ceregene is planning a 50-patient phase 2 trial with sham surgery controls, Tuszynski said.—Gabrielle Strobel.

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Madrid: Pooled Antibody Cocktail, New Metal Quencher

13 Sep 2006

Norman Relkin, at Weill Medical College of Cornell University in New York, presented the latest data on his efforts to test intravenous immunoglobulin (IVIg), a somewhat mysterious mixture of purified human polyclonal antibodies originating from the plasma of blood donors. It has been used safely, with the FDA’s blessing, for immune deficiency and inflammatory conditions for 25 years. Researchers do not know precisely which component of this preparation does what with regard to Alzheimer disease pathogenesis, but what they do know indicates that some of the antibodies in the mix dissociate Aβ fibrils and enhance Aβ clearance (see Istrin et al., 2006). IVIg contains so-called natural antibodies that always course through people’s bodies rather than being made specifically in response to a given infection. The numbers of natural antibodies appear to decline in people with AD.

Relkin first mentioned a 6-month phase 1a study in which eight AD patients received monthly infusions of IVIg. They either stayed stable or improved by an average of 2.75 points on the MMSE. After 6 months, their CSF Aβ had decreased. When the researchers washed the drug out, the patients declined rapidly, reverting to their previous cognitive state after 3 months. In a second, 9-month, phase 1b trial in the same people, the investigators re-infused the preparation every other week, and tested the patients’ cognitive status at weeks 9, 12, 15, and 18. During the second treatment phase, the patients’ symptoms remained stable and some trended upward, Relkin reported.

The trial was an open-label, add-on study of IVIg. This means its inclusion criteria require that the patients be on cholinesterase inhibitors and/or memantine prior to IVIg and declined despite these conventional treatments. All patients took at least one of these drugs when they began IVIg treatment and kept doing so throughout the entire 18 months.

The trial included five spinal taps on each patient, three in the first 6 months, one after 9 months, and one after 18 months. Plasma Aβ increased after IVIg infusions, whereas CSF Aβ42 and 40 declined from baseline (compare to other AD immunotherapies). After several months of IVIg infusion, plasma Aβ levels, too, began to decline in half the patients. Relkin noted that he believes a peripheral sink effect is at work in his patients in that the presence of the IVIg antibodies slowly drives Aβ towards peripheral clearance. This is not mutually exclusive with central mechanisms, as the investigators also noted a dose-dependent increase of CSF immunoglobulin levels. Relkin cautioned that more work is needed to sort out the complicated relationship between the different forms of Aβ in plasma and CSF. To address this issue, Relkin collaborated with Gerald Boehm, who founded ACGT ProGenomics, a German biotech company that claims to have developed the first test to detect Aβ oligomers in physiological fluids. This test indicated a decrease in the level of detectable oligomers after IVIg treatment, suggesting the preparation might act against oligomers, Relkin said.

At present, a 24-subject, double-blind, placebo-controlled phase 2 trial with additional outcome measures is in progress, with results expected early next year. A phase 3 trial through the Alzheimer Disease Cooperative Study is being planned for 2007, Relkin noted. The product, by Baxter Bioscience, is expensive and in limited supply. This might raise thorny questions about who will get access to IVIg should larger trials confirm its promise.

A Better Clioquinol to Enter Phase 2
Clioquinol is a former antibiotic with a storied history. It has served to prove the point that a drug that dampens the metal-mediated redox activity of Aβ, and also revs up its clearance, has potential as an Alzheimer drug (White et al., 2006; Ritchie et al., 2004). Clioquinol fell short as a candidate for a new AD medicine due to manufacturing difficulties (see ARF related news story), but Prana Biotechnology, the Australian company behind it, has a new horse running.

A principal academic proponent of this approach has been Ashley Bush of the Mental Health Research Institute, Victoria, Australia. In Madrid, Bush reported that PBT2, Prana’s second-generation compound, is an 8-hydoxyquinoline that outdoes clioquinol on a range of preclinical parameters in vitro and in transgenic mouse models for AD. Importantly, PBT2 enters the brain at a more rapid clip and shows better pharmacokinetics, such that it can probably be given once a day. Two phase 1 trials of PBT2, a single-dose trial in healthy men and a multi-dose series in healthy, elderly men and women, showed no significant side effects. Prana is currently gearing up for a multi-site phase 2a trial of PBT2 in people with early AD, to be conducted in Sweden starting late this year. Also at ICAD in Madrid, Prana’s co-founder Colin Masters, University of Melbourne, Australia, received a Lifetime Achievement Award in Alzheimer Disease Research.—Gabrielle Strobel.

References

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Istrin G, Bosis E, Solomon B. Intravenous immunoglobulin enhances the clearance of fibrillar amyloid-beta peptide. J Neurosci Res. 2006 Aug 1;84(2):434-43. PubMed.
White AR, Du T, Laughton KM, Volitakis I, Sharples RA, Xilinas ME, Hoke DE, Holsinger RM, Evin G, Cherny RA, Hill AF, Barnham KJ, Li QX, Bush AI, Masters CL. Degradation of the Alzheimer disease amyloid beta-peptide by metal-dependent up-regulation of metalloprotease activity. J Biol Chem. 2006 Jun 30;281(26):17670-80. PubMed.
Ritchie CW, Bush AI, Mackinnon A, Macfarlane S, Mastwyk M, MacGregor L, Kiers L, Cherny R, Li QX, Tammer A, Carrington D, Mavros C, Volitakis I, Xilinas M, Ames D, Davis S, Beyreuther K, Tanzi RE, Masters CL. Metal-protein attenuation with iodochlorhydroxyquin (clioquinol) targeting Abeta amyloid deposition and toxicity in Alzheimer disease: a pilot phase 2 clinical trial. Arch Neurol. 2003 Dec;60(12):1685-91. PubMed.

Madrid: Highs and Lows of The Insulin Connection

14 Sep 2006

Frequently in Alzheimer research, new trends take form when epidemiologic studies suggest an association between the risk of developing Alzheimer disease and some second factor. One such area that is growing in strength is the overlap between type 2 diabetes and Alzheimer disease. More broadly, all components of what is loosely called metabolic syndrome—hypertension, high blood lipids, high blood sugar, insulin resistance, obesity—are linked with increased risk for age-related dementia. While mechanistic studies are ongoing and the epidemiologic connection is still growing in strength, some groups are already beginning to report results of some initial clinical trials (see below). Unfortunately, also frequently in AD research, tantalizing hints of a therapeutic effect show up in small pilot trials, only to fall flat when tested subsequently in larger, better-controlled studies. One problem is that, sometimes, trials are designed without sufficient input from basic scientists before underlying biologic processes of the new association, and specific biologic markers for it, are worked out for clinical trials to measure. The insulin/diabetes connection so far is no different.

One pilot trial reported at ICAD tested insulin itself. The underlying rationale is that plasma hyperinsulinemia and insulin resistance in the periphery paradoxically lead to a deficiency of insulin in the brain, probably because the peripheral condition changes the receptor-mediated transport of insulin at cells of the blood-brain barrier. Addressing this issue, Suzanne Craft and colleagues at the University of Washington, Seattle, attempted to deliver insulin directly to the brain by way of the nose. This route might be able to avoid the low blood sugar that would result from systemic insulin treatment (see also Born et al., 2002). The scientists used electronic atomizers to spray insulin into the noses of people with early AD or amnestic MCI for 3 weeks. Of 25 patients, 13 were randomized to receive 20 international units of insulin daily and 12 to placebo. Plasma glucose and insulin levels happily did not change in insulin sniffers, nor did they suffer other side effects, Craft reported. The placebo and insulin group performed equally on verbal recall tests at baseline, but at the end of the trial the insulin group outperformed the placebo group. Older patients responded less well than younger patients. Intriguingly, intranasal insulin appeared to change plasma Aβ and cortisol levels.

Instead of using insulin, perhaps current type 2 diabetes drugs might work for AD? After all, AD is sometimes called “type 3 diabetes,” and some widely used drugs effectively increase the body’s sensitivity to insulin and lower blood glucose levels. GlaxoSmithKline’s rosiglitazone and Takeda Pharmaceutical/Lilly’s pioglitazone both are thiazolidinedione compounds that act as agonists of the PPARγ nuclear receptors. First, consider rosiglitazone. A small trial by Craft and colleagues had suggested a cognitive benefit in AD patients (Watson et al., 2005), and in Madrid, scientists from GlaxoSmithKline presented data for a large follow-up study. In a 6-month double-blind, placebo-controlled, dose-ranging trial of an extended-release form of rosiglitazone in 518 non-diabetic AD patients, the drug showed a similar safety profile as was previously established for diabetic patients. Edema and cardiac complications occurred as anticipated (see also ARF related news story), but no additional side effects cropped up in this AD population. The trial at first looked good: the patients were newly diagnosed, did not also take cholinesterase inhibitors or memantine, and 85 percent completed the trial. Unfortunately, the drug did not significantly improve their ADAS-Cog or CIBIC scores, reported Marina Zvartau-Hind of GlaxoSmithKline in Greenford, United Kingdom. There was no significant difference between the rosiglitazone and the placebo group. (The placebo group barely declined, as sometimes happens in 6-month trials of this slow-moving disease.)

This disappointing result could have ended the effort. Yet when the investigators analyzed, as planned, the ApoE4-positive and negative trial participants separately, they found a ray of hope. Patients without an E4 allele had, in fact, improved on the highest dose given, whereas people with one ApoE4 allele showed no benefit, and people with two ApoE4 alleles declined (Risner et al., 2006). Subgroup analysis is weaker than the result on the primary endpoint. Zvartau-Hind noted that this exploratory finding can’t help a doctor decide whether to prescribe rosiglitazone to a given AD patient. It also is not sufficient to encourage patients to find out their ApoE status. But the finding has swayed GlaxoSmithKline to continue testing rosiglitazone for AD, and larger trials powered to study its effect both in ApoE4 carriers and non-carriers are planned.

Rosiglitazone’s competitor pioglitazone also was put to the test, though a smaller one. David Geldmacher of the University of Virginia, Charlottesville, with colleagues at University Hospitals and Case Western Reserve University in Cleveland, Ohio, reported results of their 18-month trial of this drug in 29 non-diabetic AD patients. They were randomized to take either the drug or placebo but unlike in the GlaxoSmithKline trial, also took cholinesterase inhibitors and/or memantine. More than a quarter of the people in the treatment group developed edema; otherwise, they tolerated the drug well. Cognition, function, and behavior did not improve significantly, but there was a positive trend that the investigators interpret to warrant a larger trial on this drug, as well.

If those drugs are no home run, how about going after the signal transduction cascade downstream of insulin, to boost the hormone’s downstream effects? The literature is ripe with evidence implicating reduced levels of insulin-like growth factor-1 (IGF-1) in aging, cognitive decline, AD, and amyloid degradation (e.g., Rivera et al., 2005; for a recent review, see Messier and Teutenberg, 2005). Led by J. Michael Ryan, scientists at Merck Research Laboratories in North Wales, Pennsylvania, took a cue from that body of work and tested MK-0677, a compound that induces secretion of IGF-1. They randomized 563 AD patients with baseline MMSE scores between 14 and 26 to take either MK-0677 or placebo daily for a year. In this double-blind trial, MK-0677 did increase IGF-1 serum levels by 60 percent. Sadly, this failed to move any of the clinical treatment endpoints. Both CIBIC-plus and ADAS-Cog scales showed little change; neither did secondary endpoints.

What gives? Does the failure of large trials mean the epidemiological data are wrong? No, scientists across the field generally agree. Epidemiologists cautioned that one possible reason why trials have shown little effect is that epidemiology data are converging to show a link between components of the diabetic syndrome in mid-life and elevated risk for AD a decade or two later. As happened with anti-inflammatory drugs and estrogen, the trials tested drugs that are based on a mid-life risk factor in the hope that the drug will still be able to help a much older brain that has since degenerated considerably. To design better intervention—or even prevention—trials in younger people, more mechanistic insight in the underlying processes of metabolic syndrome components in AD is needed. This is particularly urgent because most patients have mixed forms of AD and vascular dementia, said Monique Breteler of Erasmus University in Rotterdam, The Netherlands. Echoing a similar story for estrogen, Kristin Yaffe of University of California, San Francisco, noted that after the disheartening failure of conjugated horse estrogen in the Women’s Health study, researchers have tried to focus on a critical period of dementia initiation in late mid-life, when endogenous sex hormone levels decline. They are beginning to test designer estrogens such as raloxifene for their ability to protect against MCI, not dementia (Yaffe et al., 2005).

The association between a history of diabetes and risk for AD is undisputed, but the mechanisms are nebulous, agreed Richard Mayeux of Columbia University, New York. Leads for possible mechanisms include insulin’s role in Aβ clearance by competition for the enzyme IDE, its downregulation of the tau kinase GSK3β, and its effect on the neuroprotective Akt signaling pathway. Does insulin resistance change the outcome of these pathways toward AD? Insulin-resistant adults have lower CSF Aβ42 levels, which other work has suggested foreshadows future AD. Research should focus on how increased CSF insulin might damage the brain’s microvasculature and blood-brain barrier and, in turn, lower insulin signaling inside the brain. More broadly, mechanisms accounting for microvascular damage could explain some of the established overlap between vascular dementia and AD. The focus in this area is slowly shifting away from ischemia and toward small hemorrhages and vascular amyloid, noted Breteler.

Research also should focus on a clear delineation between the effect of central insulin on Aβ and peripheral insulin on Aβ. If blood insulin levels increase peripheral Aβ, especially large amounts produced in muscle, then the direction of transport could shift toward Aβ import into the brain. Insulin is one of several factors that affect APP metabolism, Mayeux added, all of which deserve a clear description of the mechanistic pathway. Examples include dietary factors and stress. Hormones released by fat in that dangerous potbelly, as well as elevated glucocorticoids, cause insulin resistance and can lead to the same functional hypoglycemia in the brain that is seen in diabetes (see also Green et al., 2006; see ARF Madrid story). Understanding these mechanisms could pay off not only in better trial design but also in early detection and, eventually, prevention. The scientists agreed that epidemiology and basic science need to move in concert toward this goal.

A final note on the anti-inflammatory treatment front, which has suffered from similar problems. A trial of triflusal, an antithrombotic drug that also appears to inhibit NF-κB in the brain, showed a hint of promise toward reducing progression from MCI to AD in 257 people. However, slow recruitment forced a premature end to this trial, led by Teresa Gomez-Isla of the Hospital Santa Creu i Sant Pau in Barcelona, Spain, and conducted there and in Lisbon, Spain. An Italian trial of ibuprofen, conducted by researchers in Brescia, Pavia, Turin, and Rome, failed to slow cognitive decline in patients with mild-to-moderate AD in 132 patients. A longer-term follow-up of R-flurbiprofen confirmed and extended a moderate positive effect on cognition in patients with mild AD reported earlier (see ARF related conference story).—Gabrielle Strobel.

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Watson GS, Cholerton BA, Reger MA, Baker LD, Plymate SR, Asthana S, Fishel MA, Kulstad JJ, Green PS, Cook DG, Kahn SE, Keeling ML, Craft S. Preserved cognition in patients with early Alzheimer disease and amnestic mild cognitive impairment during treatment with rosiglitazone: a preliminary study. Am J Geriatr Psychiatry. 2005 Nov;13(11):950-8. PubMed.
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Madrid: Beyond Aβ—Learning, Age, DNA Damage, and Pai-1

15 Sep 2006

This story closes our coverage of the 10th International Conference on Alzheimer’s Disease and Related Disorders, held last July in Madrid. See PDF of this story.

15 September 2006. Bid adios to this news-rich conference by reading about attempts to branch out beyond the Aβ peptide itself. Two studies summarized here work toward understanding how Aβ functions in a broader context of aging and synaptic plasticity. The final part highlights a novel drug development approach to promote its clearance rather than slow its production.

In a presentation on the continuing analysis of the triple transgenic mouse model of AD, Frank LaFerla of University of California, Irvine, focused on the effects of age and learning. Mental activity is thought to protect against AD. In a separate talk, Serge Gauthier of McGill University in Montreal, Canada, discussed the promise of cognitive training as a non-pharmaceutical measure that could be used to treat amnestic MCI even while practitioners have to make do without good mechanism-based drugs. But much confusion remains about exactly how this protection works at the cellular level. Synaptic activity appears to drive Aβ production (Cirrito et al., 2005), and too much synaptic activity, that is, excitotoxicity, certainly damages neurons. In his talk, LaFerla addressed the questions of how much good cognitive training can do and what pathology has to do with it. He used his lab’s triple transgenic mice to model these questions.

This work is a current focus in the ongoing behavioral characterization of the triple transgenic mouse model beyond the published data (e.g., Oddo et al., 2003; Billings et al., 2005). In brief, starting at 6 months of age, the triple transgenic mice diverge from controls in that they learn but do not remember well, the homozygotes more so than the heterozygotes. Both show a training effect in that repeated learners tend to perform better than transgenics of the same age who must plunge into the Morris water maze the first time. With increasing age, the mice’s cognitive performance goes down as intraneuronal Aβ levels go up, a finding made by Claudio Cuello’s lab, as well.

In Madrid, LaFerla described a longitudinal study from 2 to 18 months of age, in which his coworkers evaluated triple transgenic mice at multiple time points, at every step comparing them with naïve transgenics. As expected, the trained transgenics performed better at every step. Unexpectedly, however, the training had an effect on the pathology. The repeated learners had less amyloid deposition and soluble Aβ, similar to what Carl Cotman’s (Adlard et al., 2005) and Sam Sisodia’s groups have found (Lazarov et al., 2005). Unlike the latter study, however, LaFerla’s group found no evidence for an enrichment-induced upregulation in enzymes that degrade Aβ monomer. Instead, preliminary data suggest that the repeated learners might have less of certain kinds of Aβ oligomer, particularly the Aβ56* species (Lesne et al., 2006). The mice also had reduced levels of tau phosphorylation and of active GSK3β tau kinase.

Continued training carried the transgenics only so far. By 15 months of age, learners lost their advantage over naïve transgenics, presumably because the induced pathology increases with time and overwhelms whatever activity-induced protective mechanisms are at play. It could also be that aging itself induces gene expression changes that affect learning and pathology, or loss of neurogenesis, LaFerla noted.

The Cdk5 system of tau phosphorylation was unaffected in LaFerla’s learning series, but it clearly plays a role in learning- and memory-related synaptic activity. Li-Huei Tsai, of the Picower Institute for Learning and Memory at MIT, presented new data for the mechanisms by which this kinase participates in neurodegeneration in a mouse model. Cdk5 has been implicated in AD, Parkinson disease, stroke, and even neurodevelopmental conditions (e.g., Jacobs et al., 2006; Smith et al., 2006; Liu et al., 2004; Venturin et al., 2006). Prior work by Tsai and others has established Cdk5 as a kinase that is normally tightly controlled by its cofactor p35 but becomes hyperactive when the protease calpain cleaves p35 to release p25 (Cruz and Tsai, 2004). An inducible mouse model of p25 shows that a transient p25 sharpens synaptic plasticity and learning, but prolonged p25 exposure leads to a crash and massive neurodegeneration, suggesting that temporary p25 production helps accommodate increased demand for plasticity and learning but that neurons cannot sustain this state (Fischer et al., 2005).

In Madrid, Tsai laid out a temporal sequence of events inside neurons from p25 induction to neuronal death. Like AD patients, the p25 inducible mice suffer severe neuronal loss, astrogliosis, and brain shrinkage. Like the triple transgenics, the p25 inducible mice show increases first and predominantly in intraneuronal Aβ. Data for this include intraneuronal staining with oligomer-specific antibodies and immunogold electron microscopy labeling of filamentous Aβ near the nuclei of forebrain neurons and in distended axons. This led Tsai to suggest that p25 can somehow upregulate Aβ production. The mechanism is unclear, but options include effects on BACE1, on APP C-terminal phosphorylation, and on axonal transport. The mice also show tau hyperphosphorylation (Cruz et al., J. Neuroscience, in press).

Further analysis of the phenotype pointed to DNA damage and cell cycle induction as underlying mechanisms of the neurodegeneration. Tsai’s group performed a microarray analysis of brain mRNA preparations to compare gene expression 2 weeks after induction (prior to neuronal loss and gliosis) and 8 weeks after induction (well into degeneration). Of 236 genes, the expression of which was altered at the 2-week point, 74 were related to the cell cycle and DNA damage response. This pattern reflects the induction of neurodegeneration more than its progression, because these genes were much more highly upregulated at the 2-week than the 8-week point, Tsai noted.

Subsequent analyses showed evidence of DNA double-strand breaks in p25 transgenic mice. It also indicated that this damage is still reversible 2 weeks after p25 induction, suggesting that ongoing DNA repair mechanisms can handle damage for a while but that continuous stress will exhaust them. In cultured cortical neurons, p25 induced DNA double strand breaks. It did so 4 hours after p25 expression, but the neurons did not die until many hours later. This echoes the time course in the mice, where DNA damage precedes cell death. (It also fits with longitudinal multiphoton imaging on another severe neurodegeneration mouse model, performed by Tara Spires in Brad Hyman’s lab, where doomed neurons that express mutant human tau, bear tangles, and even have activated caspases still manage to survive for weeks.) In Tsai’s work, double-staining experiments indicate that those same neurons that accrue DNA damage at the same time also attempt to re-enter the cell cycle. In this sense, the p25 mice confirm earlier data by Karl Herrup, Inez Vincent, and Peter Davies, who suggest that cell cycle re-entry is an early sign of AD. Currently, the Tsai lab is testing human brain samples for evidence of DNA damage and double-strand breaks in AD.

Taken together, Tsai’s data would suggest, then, that chronic p25 generation leads to DNA damage, cell cycle reactivation, Aβ induction, and tau phosphorylation, and that these processes feed into each other before the neuron eventually dies. How p25 fits into synaptic activity in aging humans, however, remains a question.

Many scientists across the field share the goal of broadening their vista, and die-hard believers of the amyloid hypothesis practice their own version of this virtue. An example on display in Madrid was Steve Jacobsen’s presentation on his company’s attempt to reduce Aβ levels by a new mechanism. Wyeth Research, in Princeton, New Jersey, has started a drug development program around the notion that finding a small-molecule inhibitor of a target upstream of an Aβ-degrading enzyme might reduce rising Aβ levels in a way that is different from current attempts, such as secretase inhibition or immunotherapy. Researchers frequently have called for efforts to understand the broader regulation of Aβ metabolism so that targets several steps removed can be tackled for drug development. This strategy has been successful in heart disease, where statin drugs don’t attack cholesterol itself but inhibit an enzyme that acts upstream in its synthesis cascade.

In this case, the Wyeth researchers picked up earlier academic work by Steve Estus, Sidney Strickland, and others who had put the plasmin system on the map as one of a handful of Aβ-degrading enzyme systems now known (Tucker et al., 2000; Melchor et al., 2003; Melchor and Strickland, 2005). Plasmin looked attractive because, unlike IDE, ECE, and neprilysin, it cleaves oligomeric Aβ as well as the monomeric form. A growing number of scientists suspect Aβ oligomers of causing acute synaptic dysfunction, and plaques of rousing glial cells and inflammation and impeding impulse transmission in other ways. In AD, Jacobsen said, the plasmin system itself is probably intact but an upstream protein that downregulates it is abnormally increased. That protein is called plasminogen activator inhibitor-1 (Pai-1). Aging and inflammation both somehow elevate Pai-1 in the brain, Jacobsen said. Pai-1 feeds into the biochemical plasmin cascade by inhibiting the tissue plasminogen activator (TPA) complex, which turns into plasminogen and then into active plasmin. Consequently, small-molecule inhibitors of Pai-1 should spur Aβ degradation by plasmin and tip the balance between Aβ production and clearance toward the latter. Such a compound would be the first in a new class, Jacobsen said.

The company scientists have found Pai-1 inhibitors that enter the brain. Some lower plasma and brain Aβ levels in Tg2576 mice, and improve their performance in contextual fear conditioning. A safety concern with this approach is that TPA serves as a treatment for ischemic stroke, so fueling this cascade could cause bleeding. Jacobsen said that TPA is an endogenous molecule that would be induced at much lower concentrations than are given in acute ischemia. Pai-1 knockout mice are normal. Pai-1 loss-of-function mutations known to occur among the Old Order Amish cause no bleeding, though whether these people enjoy protection from AD is unknown, Jacobsen added. A separate genetic tie to AD exists in the form of association studies implicating a tissue plasminogen activator gene in AD; see Alzgene overview. Hasta luego, Madrid.—Gabrielle Strobel.

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References

Paper Citations

Cirrito JR, Yamada KA, Finn MB, Sloviter RS, Bales KR, May PC, Schoepp DD, Paul SM, Mennerick S, Holtzman DM. Synaptic activity regulates interstitial fluid amyloid-beta levels in vivo. Neuron. 2005 Dec 22;48(6):913-22. PubMed.
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Adlard PA, Perreau VM, Pop V, Cotman CW. Voluntary exercise decreases amyloid load in a transgenic model of Alzheimer's disease. J Neurosci. 2005 Apr 27;25(17):4217-21. PubMed.
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Cruz JC, Tsai LH. Cdk5 deregulation in the pathogenesis of Alzheimer's disease. Trends Mol Med. 2004 Sep;10(9):452-8. PubMed.
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Tucker HM, Kihiko M, Caldwell JN, Wright S, Kawarabayashi T, Price D, Walker D, Scheff S, McGillis JP, Rydel RE, Estus S. The plasmin system is induced by and degrades amyloid-beta aggregates. J Neurosci. 2000 Jun 1;20(11):3937-46. PubMed.
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Melchor JP, Strickland S. Tissue plasminogen activator in central nervous system physiology and pathology. Thromb Haemost. 2005 Apr;93(4):655-60. PubMed.

Madrid: ICAD Conference Draws to a Close

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