Chicago: Studies Probe Diminishing Placebo Decline, Part 1

28 Jul 2008

This is a two-part story. See also Part 2.

It is an observation noted so often that it’s taken as gospel: in Alzheimer disease clinical trials, placebo groups seem to be worsening at a more leisurely rate than they once did. If so, AD drug makers face serious problems. The newest generation of AD therapeutics tend to slow disease progression rather than improve symptoms, so if placebo groups do not deteriorate at predictable rates, perfectly effective drugs could slip under the statistical radar.

Much of the discussion on waning placebo decline has centered around the ADAS-Cog, an 11-item portion of the Alzheimer’s Disease Assessment Scale that measures cognition and serves as a primary outcome measure in many AD drug trials. “What was bandied around back in the early 1990s is that decline on the ADAS-Cog would be six to nine points per year,” said Rachelle Doody of Baylor College of Medicine in Houston, Texas, noting an earlier analysis (Stern et al., 1994) that produced an equation for predicting progressive ADAS-Cog decline from baseline scores. “We never see this much decline now. Four points at one year is typical.”

Several studies presented this week at International Conference on Alzheimer's Disease (ICAD) use different approaches to address this conundrum. Tomorrow afternoon, Lon Schneider of the University of Southern California Keck School of Medicine in Los Angeles, will present results from a meta-analysis that essentially debunks the notion that placebo participants have declined more slowly in recent AD trials. Roy Jones, director of the Research Institute for the Care of Older People (RICE) in Bath, U.K., and colleagues presented a poster yesterday showing that placebo patients in AD trials of donepezil initiated between 1995 and 1999 have declined more slowly than placebo groups of earlier (1990-1994) donepezil trials. And in a poster presentation Wednesday (see ARF companion story), Michael Irizarry and other GlaxoSmithKline colleagues in Research Triangle Park, North Carolina, and several U.K. sites will share their analysis identifying baseline participant characteristics that predict subsequent ADAS-Cog decline in placebo groups of AD trials. Despite differing design, data sets, and conclusions, these studies underscore the field’s intensifying need to gauge placebo decline more effectively in future AD trials.

A Test of Urban Legend
After “sitting in meetings and listening to experts say the placebo groups are not responding as they’re supposed to,” Schneider decided to apply some rigorous number-crunching toward a test of this claim. He and colleagues reasoned that an analysis of placebo data pooled from many trials could most effectively evaluate this widespread perception, based largely on unfavorable results from individual trials. The researchers combed the literature and hit up company officials for raw data from randomized, double-blind, placebo-controlled AD trials lasting at least six months. They found 103 such trials between 1991 and 2005 and were able to extract the following information from 87: trial size, countries, number of sites, treatment allocation ratios, enrollment dates, age, gender, ADAS-Cog, and Mini-Mental State Examination (MMSE) scores. More than a third of the studies in his meta-analysis were unpublished, Schneider told ARF.

Consolidating the placebo data from these 87 studies, Schneider and colleagues found no changes in baseline characteristics or cognitive change scores across the 15-year period of the trials. Collectively, the placebo subjects showed mean MMSE declines of -0.73 (SD = 0.94) and -2.77 (SD = 0.94) at six and 12 months, respectively. ADAS-Cog changes at those time points were 1.44 (SD = 1.58) and 4.13 (SD = 2.39).

"When you look at this, it becomes a numbers thing,” Schneider told Alzforum. “Sampling and play of chance alone may lead to not showing the decline.” Consider a six-month, European Phase 3 trial of phenserine for probable AD, whose lack of efficacy in cognition, global function, or activities of daily living was chalked up to an unusually slow rate of placebo decline. Those unpublished data—presented by Bengt Winblad (Karolinska Institutet, Stockholm) at the 2005 International Conference on Alzheimer’s Disease and Parkinson’s Disease in Sorrento, Italy—marked a turning point for the field, causing many to cast their first suspicious glare on placebo decline patterns. And rightly so—at 12 weeks, the placebo group had improved more than a point on the ADAS-Cog and returned to baseline by six months.

Although the phenserine study was relatively large (375 patients), the placebo group was small (76 patients). Therein lies a problem, Schneider said. In his analysis to be presented tomorrow at ICAD, the size of the placebo arm was a critical factor in subsequent cognitive decline. Smaller studies show greater variability in mean placebo progression—some improve, some worsen, Schneider said. In a recent phone conversation, he drew an analogy to a coin toss experiment in which laws of probability predict 50 percent of the flips to turn up heads. However, in smaller samples—say, 10 flips—the number of heads could swing widely, maybe from one to nine. If the coin were tossed 1,000 times, the outcomes would fall within a range much closer to the expected 50 percent.

Similarly, Schneider’s meta-analysis of AD trials showed that at six months, placebo sample sizes less than 100 had a 37 percent chance of not showing significant change, whereas placebo groups of more than 200 all showed significant worsening. In 12-month studies, 95 percent of placebo groups larger than 100 declined significantly. “This is just measurement and sampling theory that I am demonstrating is a major problem in AD study design,” Schneider said.

In his meta-analysis, placebo groups from studies that allowed use of cholinesterase inhibitors showed about the same ADAS-Cog decline as did placebo subjects in studies from an earlier time when cholinesterase inhibitors were not allowed. This finding would seem to discredit the idea that use of cholinesterase inhibitors has led to slower placebo decline in AD trials of newer, disease-modifying treatments. “What this practically means is that you don't have to exclude patients on cholinesterase inhibitors from clinical trials, all other things being equal,” Schneider noted. Less deterioration was also linked with trials having non-English speaking sites, perhaps reflecting the challenge of using clinical instruments in demographics for which they were not originally designed. In addition, the researchers found less variance associated with more frequent assessments. While this point suggests that boosting the number of evaluations in clinical trials may improve reliability, it should be considered in context with a recent study (Gold, 2007 and ARF companion story) that found frequent assessments associated with practice effects.

Sid Gilman, a neurologist at the University of Michigan, Ann Arbor, who has broad experience in AD drug trials, studied Schneider’s data when the two met informally during a meeting several weeks ago. In a phone interview with ARF, Gilman said he was struck by the similarity between the mean ADAS-Cog changes for the pooled placebo patients in Schneider’s analysis and for the placebo group in an 18-month trial of folate and vitamins B12 and B6 to lower homocysteine in AD. “A single trial comes very close to what Lon was showing overall,” Gilman said of the homocysteine lowering study, with which he was not involved.

In that trial, the mean ADAS-Cog decline in 169 placebo patients was 1.12 points at six months and 4.46 at 12 months, compared with 1.44 and 4.13 points, respectively, in Schneider’s analysis. Data from the homocysteine lowering study were first presented last year at the Alzheimer’s Association International Conference on Prevention in Washington, DC, by lead investigator Paul Aisen, then at Georgetown University and now at the University of California, San Diego. Aisen is director of the Alzheimer’s Disease Cooperative Study (ADCS).

Further support for Schneider’s ADAS-Cog figures comes from placebo data of several other large, 18-month AD trials—of simvastatin and tarenflurbil (aka Flurizan)—presented at ICAD this week. In the statin and Flurizan trials, involving placebo groups of around 200 and 800, respectively, placebo decline on the ADAS-Cog hovered around 1.5 points at six months and four points at 12 months—mirroring the figures from Schneider’s meta-analysis. Disappointing results from the highly anticipated tarenflurbil trial, in which Schneider was a co-lead investigator, were reported in brief last month by Myriad Genetics, Salt Lake City, Utah, which has decided to discontinue the drug (see ARF related news story). None of these three 18-month trials were included in Schneider’s meta-analysis of placebo data.

Doody told ARF in a phone interview that she found “no surprises” in Schneider’s findings. But “you can’t take this and apply it as a critique of specific studies,” she said. As lead investigator in a recently published Russian trial of Dimebon, Doody noted that though the small placebo group of 77 mild to moderate AD patients worsened significantly (more than five points on the ADAS-Cog at six months), the drug effect did not depend on that decline. In that trial, patients on Dimebon actually had better ADAS-Cog scores at six and 12 months than at baseline (see ARF related news story).

Placebo Behavior in a Decade of Donepezil Studies
Jones, director of the Research Institute for the Care of Older People (RICE) in Bath, U.K., and colleagues took a closer look at the placebo decline dilemma using a different approach: a meta-analysis of placebo data from AD trials of the acetylcholinesterase inhibitor, donepezil, the world’s most prescribed Alzheimer’s medication. The researchers collected data on 3,403 patients who participated in 13 randomized, double-blind, placebo-controlled AD trials of donepezil initiated between 1990 and 1999. All pooled data came from mild to moderate AD patients (MMSE scores 10-26) who took part in AD trials that used the MMSE and/or ADAS-Cog to measure post-baseline cognition and were within a donepezil clinical development program run by Eisai Inc. and Pfizer Inc., the drug’s developers, wrote Jones in an e-mail to ARF. He said that data from trials run after 1999 were not included because double-blind studies of donepezil after this date focused on MCI, severe AD, and vascular dementia.

Jones and colleagues found that patients in more recent (1995-1999) trials had significantly lower mean MMSE decline (-0.56 points) from baseline to week 24, compared with those enrolling in earlier (1990-1994) trials (-1.28 points). Over the same time frame, placebo subjects of more recent donepezil trials also worsened less on the ADAS-Cog (1.03 points versus 1.82 points in placebo participants of earlier trials), but this difference was not significant. Jones noted that their data included more information on MMSE changes and less on ADAS-Cog changes, reflecting availability of information from the studies used in their analysis.

As disclosed in the ICAD meeting abstract, this study resulted from discussions and analyses by an expert working group initiated and funded by Eisai and Pfizer. In the U.S., donepezil is co-promoted by these two companies and distributed by Eisai. Jones received honoraria for his participation in the analysis, which was presented in an ICAD poster yesterday.

Jeff Cummings, director of the Alzheimer’s Disease Research Center at the University of California, Los Angeles, said in a recent ARF interview that he does not find Jones’s and Schneider’s findings incompatible. “Lon’s point is that larger, longer trials are more likely to be associated with placebo decline. That probably is also true of Roy’s data,” said Cummings, who has consulted and served on speakers’ bureaus for Eisai and Pfizer. “But Roy’s data also alert us to the fact that at least within some sample sets, there seems to be a trend toward less placebo decline in more recent trials.”

As they were done by the same companies, the trials in Jones’s study had relatively similar design. In addition, the researchers observed a comparable donepezil-placebo treatment difference in the earlier and later trial sets, suggesting a consistent treatment effect across the decade of studies. Such similarities presumably made their trials more homogeneous than those in Schneider’s analysis and allowed more in-depth analysis of design details, Jones noted. He did mention that the donepezil studies post-1995 tended to have somewhat smaller sample sizes than did the pre-1995 trials, lending support to Schneider’s suggestion that sample size is important for demonstrating placebo decline.

Strangely enough, the apparent “smarting up” of placebo subjects in more recent trials did not correlate with patient characteristics that would predict this effect. Jones and colleagues found that individuals enrolling in later (post-1995) trials had, in fact, lower baseline MMSE scores. Low screening MMSE was identified in the study by Irizarry and other GlaxoSmithKline colleagues as a significant independent predictor of 24-week ADAS-Cog decline in AD trials. However, that same study found that patients whose ADAS-Cog performance deteriorated markedly between screening and baseline were less likely to show significant 24-week cognitive decline. This would seem consistent with the finding by Jones and colleagues that patients with lower baseline MMSE tended to decline less at 24 weeks.

Also somewhat unexpected was the finding by Jones’s team that placebo subjects in later (post-1995) donepezil studies—who seemed to decline at slower rates—were older, had other medical conditions, and used more medications than did patients in the earlier group of trials. Jones speculated that the increased comorbidity and medication use could have stemmed from less restrictive entry criteria in later studies, presumably because more experience had been gained about the use of donepezil. That patients in the later studies appeared to deteriorate less could also result from advances in care and social support for AD patients, improvements in the management of AD risk factors, and general advances in medical care for older people, Jones noted.—Esther Landhuis.

This is a two-part story. See also Part 2.

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Chicago: Studies Probe Diminishing Placebo Decline, Part 2

29 Jul 2008

This is a two-part story. See also Part 1.

Identifying Predictors of Placebo Decline in AD Trials
Are oddly behaving placebo groups scuppering clinical trials, or are potential treatments simply not living up to expectation? As discussed in Part 1 of this two-part story, some Alzheimer disease clinicians worry that cognitive function among placebo groups is not declining as quickly as it used to. If so, treatment effects could be harder to spot. For example, the phenserine study data reported at the 2005 AD/PD conference in Sorrento, Italy (see ARF companion news story), and several other large AD trials—of galantamine (Brodaty et al., 2005) and rosiglitazone (Risner et al., 2006)—have failed to show a difference between treatment and placebo groups in at least one primary endpoint. “When you looked at the data and at why you didn't pick up a treatment effect, what became a recurrent theme was that you didn't see a decline in the placebo groups,” said Michael Gold, vice president of neurology at GlaxoSmithKline, Research Triangle Park, North Carolina. “That prompted the question of whether something had changed in the patients, or in the way we conducted our trials.” These issues were addressed in a recent meta-analysis by Gold (Gold, 2007), and in an ICAD poster presented today by Michael Irizarry and other GlaxoSmithKline colleagues.

For his study, Gold collected published data from prospective, placebo-controlled, double-blind, parallel group studies that included subjects with mild to moderately severe probable AD and that used the ADAS-Cog as an endpoint. Emerging from his analysis of 69 AD trials, which met these criteria and were published between 1992 and 2006, were several broad observations: patients enrolling in trials are progressively older, and trials are becoming longer. In fact, trial length came up as the most robust predictor of decline in placebo groups. Gold acknowledged that this finding was no surprise, as greater deterioration would be expected in longer studies of a progressive disease. However, this could become a sticking point for more recent AD clinical studies that have bucked the overall trend toward longer trials—a move Gold attributes to the intensifying need to cut costs and gain approval by institutional review boards (IRBs) that are increasingly wary of side effect risks in prolonged studies.

His analysis also identified baseline dementia severity as a predictor of decline—another no-brainer, Gold conceded, given that milder AD patients tend to progress more slowly (Morris et al., 1993). However, his finding becomes relevant in light of the field’s greater emphasis on early intervention, which requires testing experimental drugs on patients with fewer cognitive symptoms. “If patients are coming in milder and you have IRBs that are reluctant to do longer studies, then you’ll have a problem detecting significant decline,” he said.

Throughout the 1990s, pivotal AD drug trials tended to be six-month studies. Toward the late 1990s, some trials went out to 12 months with the aim of showing longer-lasting drug benefit. As disease-modifying drugs entered the clinical pipeline in the early 2000s, companies began launching 18-month trials in hopes of being able to demonstrate disease modification.

But longer is not necessarily better. In Gold’s analysis, lengthier trials were associated with greater cognitive decline, but they also tend to involve more evaluations—a factor Gold found to be linked, unexpectedly, with less placebo decline, possibly due to practice effects that kick in with increased exposure to psychometric testing. In a separate discussion, Lon Schneider of the University of Southern California, Los Angeles—who presented a study deflating the idea that placebo groups have declined less in more recent AD trials (see ARF companion news story)—noted that longer studies can also pose problems because their data sets are often riddled with features that make them harder to analyze—for example, more dropouts and broader standard deviations due to greater range and trajectory in cognitive decline of individual patients.

A bottom-line problem, Gold told ARF, is that “clinical trials have become prohibitively expensive, so everyone is trying to cut costs.” Add to this scenario the skyrocketing demand for research subjects that comes from testing a growing number of experimental AD drugs, and companies have started going into areas of the world where trials are cheaper to conduct. However, increasing the number of trial sites often means smaller sample sizes per location, more raters, and greater diagnostic and treatment variability—factors that can decrease a study’s statistical power. It is not just “a matter of more noise in the ADAS-Cog instrument; there's also more noise in the populations,” said Gold, whose analysis found increased numbers of investigational sites to be associated with less decline among placebo subjects. If the higher “noise” within patient populations from multicenter trials arises from increased frequency of misdiagnosis—a scenario Gold finds plausible, especially for global trials—cognitive decline would be blunted in trials with more sites. Perhaps related to Gold’s finding that increased numbers of sites predict less decline is Schneider’s data, which drew an association between less worsening and trials with non-English-speaking sites (see ARF companion news story).

Gold’s meta-analysis of placebo populations was extended in an analysis to be presented at ICAD tomorrow in a poster by GlaxoSmithKline colleagues Michael Irizarry and others at several of the company’s U.K. sites. Using pooled data from 773 placebo participants in six AD trials conducted in 1996-1997, Irizarry’s team performed multivariable linear regression analysis to identify patient characteristics that predict cognitive decline. When the number crunching was finished, baseline cognitive status and screening-to-baseline change in the ADAS-Cog turned up as the strongest predictors of 24-week ADAS-Cog change. In other words, lower MMSE scores at baseline were associated with greater ADAS-Cog decline at 24 weeks, as was marked worsening on the ADAS-Cog during the four-week stretch between screening and baseline. If investigators detect a large cognitive drop in a patient at the end of those interim weeks—a period during which medications get tweaked and caregiver education takes place—“that’s a red flag,” Gold said.

Future AD Trials: What Should Change?
These findings should help guide the design of AD drug studies, which often struggle to demonstrate significant efficacy at Phase 2. “There’s a tendency to do Phase 2 studies with too few patients and, if the results are negative, to kill it and blame it on cholinesterase inhibitors,” Schneider said.

Eric Siemers, medical director for the Alzheimer disease research team at Eli Lilly and Company in Indianapolis, agreed that lack of statistical power hinders many Phase 2 studies. Phase 2 trials typically involve between 30 and 300 patients. “So even with 300 people, if it’s a three-arm study (1 placebo, 2 treatment), you’d be at just 100 per arm,” he said—too small for reliable cognitive decline by Schneider’s standards. “If you use cognitive measures as your primary outcome, you’re always going to be left with this statistical variability. There are some potentially good drugs that you’re going to kill because you don’t see the effect when in fact it was there,” Siemers said. With cognitive tests, “it takes longer and more people to see decline.”

This difficulty with cognitive measures is what led Eli Lilly to design Phase 2 trials that rely less on cognitive decline and more on biomarkers, which generally require fewer subjects to show statistical significance. Last year, Siemers presented data from a Phase 2 study of Eli Lilly’s γ-secretase inhibitor LY450139 that used plasma and cerebrospinal fluid Aβ levels as its endpoint (see ARF Washington news story). However, the risk of using biomarkers as Phase 2 readouts is that they might not translate into the cognitive responses needed in Phase 3 trials. Ideally, Phase 2 studies would use surrogate markers—biomarkers with a demonstrated ability to substitute for clinically meaningful outcomes (for more on biomarkers and surrogates, see ARF related news story). Very few biomarkers are able to meet validated surrogate criteria outlined previously (Temple, 1999), but “from a drug development standpoint, it's not so important that something is a validated surrogate marker,” Siemers told ARF. “What you’re really looking for is evidence that your drug has the right mechanism, the right immediate biochemical effect.”

Time will tell whether biomarkers will increasingly replace cognitive measures as endpoints in Phase 2 trials of AD therapeutics. Meanwhile, the studies of placebo decline described in this two-part story highlight several concerns the field will continue to confront. As Rachelle Doody, Baylor College of Medicine, Houston, Texas, sees it, “the issues to be grappled with are—either develop compounds that are expected to improve people over baseline as well as slow their decline, or deal with the fact that in a one-year trial, if all you’re expecting is for your drug to hold people flat, you won’t have a lot of differentiation from placebo.”

If AD trials using cognitive decline as a primary readout are further constrained by recruitment of milder patients, the challenges intensify. “Either we do longer studies, or the standards for calling something efficacious has to change,” Gold said. In March, a workshop involving AD activist group leaders, U.S. Food and Drug Administration representatives, clinicians, and industry leaders explored the latter possibility (see ARF related news story).

Further insight into the problem of unpredictable placebo decline could come from a large-scale effort—led by Marilyn Albert of Johns Hopkins University in Baltimore, Maryland, and Ron Petersen of the Mayo Alzheimer's Disease Research Center in Rochester, Minnesota—to analyze placebo data from AD and MCI clinical trials. Plans for collecting and analyzing such data were put on the table at a workshop last year (see ARF workshop report) and discussed further at a 21 July meeting in Washington, DC.—Esther Landhuis.

This is a two-part story. See also Part 1.

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Brodaty H, Corey-Bloom J, Potocnik FC, Truyen L, Gold M, Damaraju CR. Galantamine prolonged-release formulation in the treatment of mild to moderate Alzheimer's disease. Dement Geriatr Cogn Disord. 2005;20(2-3):120-32. PubMed.
Risner ME, Saunders AM, Altman JF, Ormandy GC, Craft S, Foley IM, Zvartau-Hind ME, Hosford DA, Roses AD, . Efficacy of rosiglitazone in a genetically defined population with mild-to-moderate Alzheimer's disease. Pharmacogenomics J. 2006 Jul-Aug;6(4):246-54. PubMed.
Gold M. Study design factors and patient demographics and their effect on the decline of placebo-treated subjects in randomized clinical trials in Alzheimer's disease. J Clin Psychiatry. 2007 Mar;68(3):430-8. PubMed.
Morris JC, Edland S, Clark C, Galasko D, Koss E, Mohs R, van Belle G, Fillenbaum G, Heyman A. The consortium to establish a registry for Alzheimer's disease (CERAD). Part IV. Rates of cognitive change in the longitudinal assessment of probable Alzheimer's disease. Neurology. 1993 Dec;43(12):2457-65. PubMed.
Temple R. Are surrogate markers adequate to assess cardiovascular disease drugs?. JAMA. 1999 Aug 25;282(8):790-5. PubMed.

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Chicago: ICAD Offered Mix Bag of Setbacks and Hopeful News

04 Aug 2008

On 31 July, the 11th International Conference on Alzheimer’s Disease, run by the Alzheimer’s Association in its hometown of Chicago, drew to a close amid a varied buzz of some good and some disheartening news. Held from 26 July on, the conference posted record attendance at more than 5,400 registrants and over 2,000 abstracts. This resulted in a conference where morning and afternoon slide talks typically required four to six parallel sessions and conference attendees increasingly had to settle on a few areas to follow.

One major take-home message from this meeting that leading scientists portrayed is that the state of the AD research field should be judged by the diversity of therapeutic approaches that are developing in preclinical science as well as wending their way through the clinical pipeline. Human trials, for example, center not only on the amyloid hypothesis, but are also beginning to test tau targets, various neurotransmitter-based targets beyond acetyl cholinesterase, neuroprotective therapies, NMDA antagonists, antioxidants, nutritional approaches, and more, as well as drugs with as yet poorly understood mixed effects, especially on mitochondria. “A marker of the field is how many trials with many different mechanisms of action are entering Phase 2 now, some entering Phase 3. The news is that there is both enough science, and will on the part of funding institutions, to push therapy development of Alzheimer disease,” Steve DeKosky of University of Pittsburgh told reporters at the meeting. Paul Aisen of University of California, San Diego agreed that the field must test a diversity of approaches for AD. “There have been many papers here on disease-modifying strategies, many around the amyloid hypothesis, but we must be open-minded to all.”

DeKosky at a news briefing urged the media and colleagues in the field not to stake too much news value on the success or failure of a single trial, especially in Phase 2. Particularly on that point, however, there was a palpable disconnect between how drug development proceeds and how it is being rated by observers in the press and investment community. For example, when Elan/Wyeth on July 30 summarized data of their Phase 2 trial of bapineuzumab in a 12-minute presentation, financial analysts in the room were piping results through instantaneously to their firms, and in after-hour trading Wall Street sliced a third off Elan’s stock price and a tenth off Wyeth’s the same evening. Many scientists at the conference who interpreted the results more positively were taken aback at this swift punishment. They argued that observers should not expect too much from a single Phase 2 trial, but instead take a longer view of immunotherapy. Along the same vein, the definitive failure of Flurizan in Phase 3 to some meant a shot across the bow of the amyloid hypothesis, while others maintained the trial had not truly tested the amyloid hypothesis because the drug was too weak.

Often in medical news coverage, when the perceived frontrunner is judged to be down, another drug steals the show as a binary media looks to fill the airwaves with thumbs up-thumbs down stories. At this conference, a U.K.-Singaporean biotech company largely unknown to the field filled the void with a completed Phase 2 trial of a compound targeting tau. With its catchy name, Rember^™ enjoyed a day in the sun, streaming from television networks to newspapers and online sources (see upcoming ICAD story for details.)

The Woes of Trial Design
More broadly, methodological problems of trial design, dry as that sounds, are an obstacle at this stage. Pharmaceutical and biotech companies are pushing intensely to move from symptomatic drugs (i.e., the ones on the market) to the coveted label of disease modification. Amid intense technical discussion on how to achieve this, some scientists recommended that companies not get hung up on the label and focus instead on finding a drug that shows a reasonably robust effect, period, no matter whether it is called symptomatic or disease-modifying. “Getting the dm phrase in your label for a marketed drug is not critically important,” said Aisen, and Rachelle Doody of Baylor College of Medicine in Houston agreed. Both these scientists run numerous clinical trials in AD. For their part, patients won’t care about the distinction and doctors will use a new drug so long as it has an obvious effect. Problems of trial design invariably came up in interviews with scientists no matter which particular trial was at issue; therefore, the Alzforum news coverage will summarize this topic first (it’s not that dry, really) in an effort to provide context for our subsequent summaries of individual trials.

The meeting illustrated that the field has made progress in the basic research that paves the way for drug development. A large proportion of the presentations explored the neurobiology and pathophysiology of AD, and there is by now a wealth of targets to try to attack therapeutically.

Consensus was palpable around several themes. Above all, scientists from all corners of AD research felt that trials of experimental drugs, and eventually treatment of people with AD, need to start earlier in the disease process than the mild to moderate stage defined by NINDS-ADRDA criteria that remain the standard today. “There is a focus on moving the detection threshold earlier,” Ron Petersen of the Mayo Clinic in Rochester, Minnesota, told reporters. (Petersen is the incoming chairman of the Alzheimer’s Association’s Medical and Scientific Advisory Council; he will replace Sam Gandy who currently holds this position.) Reflecting this shift toward early detection and drug testing, presentations on various aspects of fluid markers and brain imaging numbered a proud 775. Overall, data from different labs are converging on the conclusion that both CSF Aβ42/tau as well as some imaging modalities, possibly even certain cognitive tests but not the current ADAS-Cog, will be able to move detection back in the disease process and in this way improve the odds that experimental therapies will do some good.

Big Science Initiatives in AD?
Another topic that cropped up frequently at ICAD was the need to find ways to share data, standardize measurements, and conduct multisite biomarker research in such a way that results can be compared side by side. In other words, research consortia. Sticking to standard operating procedures is an anathema to the creative innovator, but without that, advances at an individual research institution tend to stay just that, and cannot pull the whole field forward. Two notable big-science initiatives are laying the groundwork in this direction. One is the Alzheimer’s Disease Cooperative Study (ADCS)), which has done much to help clinical sites of large multicenter trials standardize appropriate methods and procedures so any effect a drug might exert does not get lost amid the sites’ different ways of doing things. The other is the Alzheimer’s Disease Neuroimaging Initiative (ADNI) (see upcoming ICAD story), a 58-site, five-year natural history study run by a public-private consortium. ADNI has carved out what is called “pre-competitive space,” i.e., areas of common ground to all drug developers in which cooperation and data sharing benefit all participants. In this case, ADNI aims to enable both companies and publicly funded drug developers to define useful antecedent biomarkers and methods that will help them as they each subsequently test their proprietary compounds individually. This study is ongoing, but preliminary data suggest that both CSF and neuroimaging markers look promising (see upcoming story.)

ICAD Every Year
One final note on the conference itself. In Chicago, the Alzheimer’s Association president, Harry Johns, announced that ICAD will from now on be held annually, in lieu of the Prevention Conference the Association previously hosted every June in Washington, DC. The next ICAD conference will take place next July in Vienna, Austria. Johns said the decision was taken in consultation with scientists, yet an utterly unscientific poll of some 40 scientists randomly collared in the hallways suggested that the field at large may still have to come around. At first blush, none welcomed the news. Many scientists in the field enjoy the European AD/PD meeting organized by Abraham Fisher of the Israel Institute for Biological Research in Ness-Ziona. AD/PD is held in the spring of what used to be ICAD off years and in 2009 will convene in Prague, Czech Republic. Many scientists also attend the Society for Neuroscience conference each fall. In addition, they travel to small, topical conferences that have no parallel sessions and facilitate the exchange of ideas in one’s own area of research, such as Keystone, Gordon, or Cold Spring Harbor conferences. “Besides creating unhelpful competition for speakers, adding yet another major conference to our travel schedule keeps us from working productively on our research,” said one scientist who did not want to offend the Association, a granting institution in the field.—Gabrielle Strobel.

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Chicago: Trial Design Bedevils Search for New AD Drugs, Part 1

05 Aug 2008

This is Part 1 of a two-part story. See also Part 2.

On the last day of the International Conference on Alzheimer’s Disease (ICAD), held 26-31 July in Chicago, Paul Aisen of University of California, San Diego, used his plenary lecture before a still-respectable audience to put recent disappointing clinical trial results in Alzheimer disease into a broader perspective and to paint a way forward. Aisen heads the Alzheimer’s Disease Study Group, an NIA-funded cooperative involving some 50 U.S. and Canadian centers that was originally set up by the late Leon Thal and colleagues. He consults extensively for companies developing AD drugs.

In the past year, Neurochem’s Alzhemed and Myriad’s Flurizan failed Phase 3 trials, and Elan’s Phase 2 results of its passive immunotherapy bapineuzumab were clearly positive only in a subgroup of patients. These results have revived debate about the validity of the amyloid hypothesis, with some scientists arguing the hypothesis is wrong and others insisting it has still not been truly tested in the clinic. A recent paper about how the Phase 1 participants of the original AN1792 vaccine fared years later has fueled the dispute further (see Holmes et al., 2008). Suddenly, the terms “Baptists” and “Tauists,” which the field had retired years ago to most everyone’s relief, were back in currency again. Rather than enter into the heat of this controversy, Aisen took a step back and explained broadly how clinical trials in AD have evolved over the past 15 years, why current experimental treatments are facing a particular technical hurdle in Phase 2, and what could be done to overcome it. This story is largely a summary of Aisen’s plenary lecture.

On a timeline of drug development, the 1980s were the decade of acetyl cholinesterase development, the 1990s the decade of their clinical trials, 1997 to 2001 saw the approval of the three such drugs that are in wide use (donepezil, rivastigmine, galantamine), then memantine came along in 2003. All those are symptomatic. Currently, the field focuses intensely on generating and testing disease-modifying drugs. By Aisen’s estimate, the years 2010 to 2015 are when the first ones might reasonably be expected to come on line; however, to make their regulatory approval possible, methods of trial design and drug testing have to change.

In the 1990s, the FDA and leaders in the field jointly developed approval guidelines for the first wave of drugs (see also ACT-AD workshop). Trials had to have twin primary outcomes—a memory/cognitive test plus a global or functional measure. For the former, researchers largely settled on using the ADAS-Cog; it worked well for the mild to moderate population that enrolled in the trials, Aisen said. The CIBIC-plus scale worked as a global measure, as did the CDR-SB, the ADCS-ADL, and the DAD. For these symptomatic drugs, these measures enabled a separation between placebo and treatment curves on both cognition and clinical outcome. This was true even in short trials of 12 weeks, in which placebo groups generally do not decline. The effects were small—about two points on the ADAS-Cog. But they were statistically significant, and the FDA never mandated a minimum effect size.

“This was a straightforward pathway for symptomatic AD,” Aisen said. It led to approval of five drugs (including tacrine, which sees little use because it can damage the liver), and the FDA’s guidelines are still the same. But many scientists feel that the guidelines are not adequate for current science anymore. To this day, Aisen said, there has been neither a successful MCI trial nor prevention trial. Perhaps most urgently, no disease-modifying drug has garnered regulatory approval. One part of the problem is that the ADAS-Cog is insensitive to changes in very mild AD, a population that by a strong consensus of AD researchers stands a better chance of responding to drugs than people who have sustained damage to their brain for several years longer.

What can be done? For one, the ADAS-Cog can be made more sensitive for earlier stages of AD, i.e., MCI or CDR 0.5, for example, by adding executive function and delayed recall tests. Moreover, there is room to improve its analysis as well as its standardization, Aisen said. Alternative measures such as the NTB deserve attention, and computerized batteries are being developed, though in Aisen's view they are not quite ready for widespread use in ADCS trials. All these areas were topics of presentations at ICAD in Chicago. These are nuts-and-bolts kinds of tweaks that are entirely doable in the near future, Aisen said. In addition, it is hugely important to make an effort to recruit and especially to train more clinical sites, that is, principal investigators, study coordinators, raters, trial monitors. Equally important are efforts to recruit patients and design the practical experience of trials such that more patients and their caregivers stay in the trial to the end. All those efforts are necessary as a greater number and range of experimental therapies are leaving preclinical research and getting ready for human tests.

The goal of obtaining a disease modification label from the FDA poses a trickier challenge. From the regulatory point of view, the FDA at present demands a complex trial design of randomized start and withdrawal, whereas the European counterpart EMEA is open to simpler designs. Both agencies are open to the field’s increasing use of biomarker data, though not yet in lieu of cognitive clinical outcomes. The randomized start design means that one half of patients takes drug from the beginning of the trial, whereas a second set is first randomized to placebo and at 18 months switches over to active drug. This is meant to distinguish between symptomatic and disease-modifying drugs in this way: with a symptomatic drug, the second group of patients should catch up cognitively with the first, because a symptomatic drug merely produces an initial hump of benefit but then patients decline at the same rate, or parallel slope, as placebo-treated patients. With a disease-modifying drug, the second group of patients would never catch up to the first, because the earlier group’s rate of decline has slowed already in the previous treatment period and the curves of the two sets of patients diverge.

While this is nice in theory, it’s fair to say researchers and drug companies do not love this design. The second randomization doubles the length of a typical Phase 3 trial to 36 months; given present dropout rates of 15 to 35 percent, that is simply impractical, Aisen said. Aisen showed two recent examples of trial designs. The Alzhemed Phase 3 design of parallel 18-month placebo/treatment arms followed by a joint open-label extension is a fine design that should work. With an effective drug, that is. In this case, the failure was due to the drug itself, as well as to variations between the clinical sites. In the Dimebon Phase 2 trial, the data at 12 months looked like the combined curves of a disease-modifying and a symptomatic effect. That is, treated patients rapidly improved above baseline in the first 12 weeks and then declined more slowly than the placebo group (Doody et al., 2008). In this case, the slopes behaved in this mixed way in a standard trial design that also did not employ randomized start.

The message to take away at this point, Aisen said, is that getting the disease modification label is really not critical. It’s not the label companies should worry about. The bigger and more treacherous methodological challenge is how to optimize trial design in Phase 2 for drugs that are believed to work by a disease-modifying mechanism. This is the Phase 2 problem.

Where exactly is the dilemma? It is that with a disease-modifying drug, scientists cannot expect to see proof of efficacy in a Phase 2 trial. With the symptomatic drugs, they could. Why is that? Aisen explained that the underlying biology suggests that disease-modifying drugs will produce no short-term benefit but rather lessen the slope of a person’s decline curve over time. They still get worse, but less so. Over the long run, that would serve the patient well, but in the relatively short time frame accessible to Phase 2 trials, it’s a disadvantage over a quick symptomatic boost. What’s more, people with very mild AD or with MCI/CDR 0.5 do not decline at all in six months and only minimally by 12 months on the existing standard measures. To see the slope of decline curves diverge, a trial needs to enroll many hundreds or thousands of people and follow them over 18 months—but that is basically the recipe for a Phase 3, not a Phase 2. So companies are faced with the choice of running hugely expensive Phase 2 trials or moving into Phase 3 without proof of efficacy. “There currently is no rational progression from a small Phase 2 to a large Phase 3,” Aisen said. This means, in essence, that Phase 3s can be as risky as Phase 2s. In the absence of Phase 2 efficacy data, Phase 3 trials of disease-modifying drugs can be expected to fail eight out of 10 times, Aisen said. A Phase 3 program costs $200 million. The annual cost of AD to the nation currently is estimated at $100 billion. “The potential gains justify even this high risk,” Aisen said. To see how companies deal with the Phase 2 problem and how it can be circumnavigated, see Part 2 of this story, to be posted tomorrow.—Gabrielle Strobel.

This is Part 1 of a two-part story. See also Part 2.

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References

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Paper Citations

Holmes C, Boche D, Wilkinson D, Yadegarfar G, Hopkins V, Bayer A, Jones RW, Bullock R, Love S, Neal JW, Zotova E, Nicoll JA. Long-term effects of Abeta42 immunisation in Alzheimer's disease: follow-up of a randomised, placebo-controlled phase I trial. Lancet. 2008 Jul 19;372(9634):216-23. PubMed.
Doody RS, Gavrilova SI, Sano M, Thomas RG, Aisen PS, Bachurin SO, Seely L, Hung D, . Effect of dimebon on cognition, activities of daily living, behaviour, and global function in patients with mild-to-moderate Alzheimer's disease: a randomised, double-blind, placebo-controlled study. Lancet. 2008 Jul 19;372(9634):207-15. PubMed.

Chicago: The Phase 2 Problem and the Way to a Surrogate, Part 2

06 Aug 2008

This concludes a two-part story. See also Part 1.

Companies have dealt with the uncertainty created by the Phase 2 problem in clinical trial design (see accompanying story) in several ways. They either looked for hints of clinical efficacy in Phase 2 data, or used biomarkers, or combined both. In his plenary lecture on the last morning of the ICAD conference last week in Chicago, Paul Aisen of UCSD cited two recent examples, Alzhemed and Flurizan, to illustrate how well this has worked. Both drugs sank in Phase 3. Neurochem ran a short, small Phase 2 and used CSF Aβ and tau as a biomarker to give them some confidence; Myriad ran a longer, larger Phase 2 without biomarkers. The Alzhemed trial showed a change in the CSF biomarker; the Flurizan trial was negative but later during the follow-on period showed a separation, though there was no placebo comparison at that point anymore.

Two current high-profile immunotherapies, i.e., the passive vaccines by Elan/Wyeth and Eli Lilly and Co., similarly tried to glean meaning from Phase 2 trials. Elan reported efficacy across cognitive measures in a subgroup of patients, as well as a signal on CSF biomarkers. This company took a look-see at this data during the still blinded Phase 2 trial to decide whether to go ahead with Phase 3. Lilly took a perhaps even riskier leap, deciding to move into Phase 3 based on an extensive biomarker analysis but without any hint of a clinical benefit in Phase 2 (see subsequent ICAD stories on these experimental therapies).

In the context of the recent failures of Alzhemed and Flurizan, Aisen cited a recent Newsweek column that came up frequently in conversations with scientists at ICAD. Aisen took exception to the charge that a narrow-minded bias in favor of the amyloid hypothesis blinkers the field. Presentations at ICAD in Chicago included results of clinical trials of two tau-based drugs, of lithium, of antioxidants, of curcumin, of a nicotinic agonist, of hormones, of gingko biloba, insulin, omega-3 fatty acids and various nutraceutical formulations. No home runs yet (though see details on trial results in subsequent story). Clinical trials of anti-inflammatory compounds to date have failed, and trials of statins and other compounds are ongoing. It’s not for lack of interest in alternatives, Aisen insisted. The two reasons behind recent high-profile disappointments were that the compounds were weak and that deep methodological problems need to be solved.

Toward this goal, Aisen reiterated a growing tide of consensus at the ICAD meeting, namely that future trials must enroll patients who are at an earlier stage in the neurobiological progression of AD. This requires changes in early detection and in the regulatory pathway to approval (see also ARF Live Discussion on early detection).

In practice, early detection means biomarkers. Several hundred presentations at ICAD dealt with various aspects of this growing area. The main candidate markers are plasma Aβ, CSF Aβ, tau, oxidative or inflammatory markers, and various modalities of brain imaging. Of these, CSF Aβ/tau, as well as structural, amyloid, and FDG-PET imaging appear to be the most robust at present for early detection, Aisen said. This impression is based not only on individual studies, but also on preliminary data from the Alzheimer’s Disease Neuroimaging Study (ADNI), which compares these markers in the same set of people. ADNI allows scientists to calculate how large hypothetical clinical trials must be to detect a treatment effect when using these markers to measure outcome. In a separate presentation at ICAD, Michael Donohue of University of California, San Diego, presented some of these power calculations. The upshot is that if these measures were incorporated into trials of people with MCI as baseline covariates, studies could be at least a third smaller. This would enable the community to conduct more trials on a greater variety of compounds and would place less pressure on the success of any given trial.

Using these markers to detect people prior to dementia and to power trials accordingly, will become doable within the next two years, Aisen said. Also possible will be efforts to exclude people with other forms of dementia from trials. Thinking bigger, however, all this is but a step toward the larger goal of developing a surrogate marker, Aisen said. Biology dictates the search for such a higher-level marker because the pathology begins years to decades before people become demented. This means primary prevention trials would take too long to detect a cognitive or clinical effect. Trials cannot follow people for 15 or 20 years.

“We need to develop biomarkers as surrogate outcome measures. This is the future of the development of disease-modifying drugs in AD,” said Aisen.

How can the field anoint a marker to this vaunted status? The steps are clear, Aisen said. Clinical research must establish a close relationship between the biomarker and the disease neurobiology and progression. This work is well underway. Most importantly, the field has to demonstrate that the effect of a drug on the biomarker correlates with cognitive and clinical benefit. This must be done in more than one trial, with more than one agent, i.e., with a secretase inhibitor and an immunotherapy. “We need to work harder to strengthen the link between biomarker and outcome to elevate them to the status of surrogate marker. Unless we do that, the case for a surrogate marker is unacceptable to the FDA,” Aisen said.

On this goal, too, the leading candidates at present include the CSF Aβ42 and tau/phospho-tau markers, brain atrophy and amyloid imaging, even certain neuropsychology tests, Aisen said. (It is important to remember that one biomarker may not serve all purposes—markers for asymptomatic prediction, progression, a drug hitting its target must all be individually defined and may each turn out to be different ones, scientists said.) As a next step, prediction and progression markers must be established in asymptomatic or mildly symptomatic patients.

Aisen left the field with a vision for how to test drugs in a brain that still has a fighting chance to stem disease, before it has suffered extensive damage. That is to screen people in mid- and late life for abnormalities in this future surrogate marker. This would detect preclinical AD neurobiology. Then treat to reduce the person’s risk for AD. In essence it’s to treat the surrogate marker in a person who could be said to have “surrogate AD,” or, to quote a term by Jeffrey Cummings from UCLA, treat “AD without dementia.”

This is in the future, Aisen said, but not a distant one. It is clearly within reach. ADNI, as well as some individual institutional research programs, such as that at Washington University, St. Louis, are encouraging in that their data are beginning to link both neuroimaging and CSF biomarkers to a risk of decline in a subset of people with normal cognition at baseline. This is the research front at this point, Aisen said. The field must continue to build the case linking a potential surrogate to AD diagnosis and progression in mild AD, show that a drug effect on the potential surrogate is linked with clinical effect, then establish a link between the potential surrogate in asymptomatic people and later disease—and all along the way, gradually persuade regulators that this data warrants updating the agency’s guidelines.

This goal requires sustained cooperation between otherwise competing groups in academia and industry. Several efforts are underway, or just gearing up, to bring people together around shared goals in so-called pre-competitive settings. Aside from ADNI, these include the Dominantly Inherited Alzheimer’s Disease Network (aka DIAN, see subsequent story), as well as the Alzheimer’s Study Group (ASG)), the Alzheimer’s Association Research Roundtables. Advocacy groups such as ACT-AD help, as well. Finally, Aisen said, a European initiative has helped to push AD diagnosis, backed with amyloid biomarkers, toward a very mild symptomatic stage (Dubois et al., 2007; Gauthier et al., 2008). This is an improvement over the standard NINDS-ADRDA criteria of possible and probable AD (McKhann et al., 1984), which are viewed as outdated yet remain in wide use. AD starts insidiously with a presymptomatic phase, and that is where detection ultimately must go to find it.—Gabrielle Strobel.

This concludes a two-part story. See also Part 1.

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References

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Paper Citations

Dubois B, Feldman HH, Jacova C, Dekosky ST, Barberger-Gateau P, Cummings J, Delacourte A, Galasko D, Gauthier S, Jicha G, Meguro K, O'brien J, Pasquier F, Robert P, Rossor M, Salloway S, Stern Y, Visser PJ, Scheltens P. Research criteria for the diagnosis of Alzheimer's disease: revising the NINCDS-ADRDA criteria. Lancet Neurol. 2007 Aug;6(8):734-46. PubMed.
Gauthier S, Dubois B, Feldman H, Scheltens P. Revised research diagnostic criteria for Alzheimer's disease. Lancet Neurol. 2008 Aug;7(8):668-70. PubMed.
McKhann G, Drachman D, Folstein M, Katzman R, Price D, Stadlan EM. Clinical diagnosis of Alzheimer's disease: report of the NINCDS-ADRDA Work Group under the auspices of Department of Health and Human Services Task Force on Alzheimer's Disease. Neurology. 1984 Jul;34(7):939-44. PubMed.

External Citations

Chicago: Out of the Blue—A Tau-based Treatment for AD?

07 Aug 2008

Of all clinical trials reported at the International Conference on Alzheimer’s Disease (ICAD) held 26-31 July in Chicago, one targeting the protein tau was the therapy du jour, both in terms of buzz among scientists and media coverage. Never mind that the scientific buzz ranged from puzzled to intensely skeptical, whereas the media coverage was mostly breathless (see Chicago Sun Times headline Breakthrough Drug Fights Alzheimer’s; Alzheimer’s ”wonder drug”). This first tau-based Phase 2 trial reported at an AD conference fell on fertile ground in part because the research community is so urgently awaiting the development of tau-based therapies. None as yet directly address tau pathology, the second hallmark of AD, and consensus is growing that truly effective, disease-modifying therapy will have to attack both amyloid and tau. Some tau-related targets, such as its phosphorylating kinases or the GSK3β enzyme, have been extensively studied in academia and industry, and pharmaceutical anti-tau programs in general are said to be revitalized; however, no drugs based on these programs have yet posted any success in Phase 2. Moreover, the definitive failure of Flurizan in Phase 3 and the complex, mixed results of Bapineuzumab in Phase 2, made it easy for this newcomer to steal the show.

And a newcomer it is. PubMed lists no publications on the new drug, or on preclinical research to prepare its human testing. From the perspective of AD researchers, the sponsors of the new drug burst on the scene out of left field, with eight abstracts at ICAD and a press briefing that garnered multiple television interviews (see, e.g., CNN clip).

The name behind the potential drug is Claude Wischik. Younger readers may not remember that Wischik made his mark in the AD field 20 years ago, when he published two groundbreaking papers on tau with Nobel Laureate Aaron Klug, Sir Martin Roth, and tau experts Michel Goedert and R. Anthony Crowther (Wischik et al., 1988; Goedert et al., 1988). After that, Wischik continued publishing in the tau field, though he has not reported new AD research findings in the public literature since 2001. Besides holding an appointment at the University of Aberdeen, Scotland, Wischik now is co-founder and chairman of TauRx Therapeutics, a privately held Scottish-Singaporean biotech company that officially launched its website 30 July this year during the ICAD conference.

Here is a summary of the drug and the data. Its trade name is Rember^TM, its active compound is methyl thioninium chloride (MTC), a reducing agent better known as methylene blue. This is a deep blue dye used in analytical chemistry, as a tissue stain in biology, and in various industrial products such as ink, for example. Wischik told reporters and scientists that MTC interferes with tau aggregation by acting on self-aggregating truncated tau fragments.

The company conducted a Phase 2 study randomizing 321 people with mild or moderate AD to treatment with either placebo or one of three oral doses of MTC: 30 mg, 60 mg, or 100 mg three times a day. People taking AD drugs, i.e., acetylcholinesterase inhibitors or memantine, were excluded. The trial’s primary objective was to compare the effect of MTC to placebo on cognitive abilities measured by the ADAS-Cog battery at 24 weeks. In this phase of the study, patients were assessed every six weeks. Wischik did not show results of the entire cohort at 24 weeks; he showed results stratified by mild and moderate AD. For patients with mild AD, there was no difference between the groups at six months. For patients with moderate AD, Wischik reported a roughly 5.5-point decline on ADAS-Cog for the placebo patients versus a 1.5-point decline in the treated groups, resulting in an approximately four-point treatment effect. (If this holds up, it would be larger than currently approved drugs typically achieve.)

Secondary objectives included assessments at 50 and 84 weeks, brain imaging at 25 weeks, and safety and tolerability. For the 50-week time point, Wischik presented pooled results of mild to moderate patients. By 50 weeks, people on placebo had declined seven points on ADAS-Cog and those on 60 mg MTC by one point, yielding a treatment effect size of about six points. This translated to an 81 percent reduction in the rate of decline on the higher dose, Wischik said. (The low dose slope showed a 3.5-point decline.)

At 84 weeks/21 months, 30 mg data were not shown, but people on 60 mg appeared to have stabilized. If this finding stands the test of time, it would be a major advance in AD therapy. However, leading scientists cautioned that this was a large claim to stake on a single Phase 2 study, not to mention the first human study publicly reported on the compound. There were no placebo data by 84 weeks anymore, as the placebo period ended at 50 weeks; presumptive comparisons would have to be made against historical controls.

This clinical trial featured an imaging marker component, whose data looked supportive at first glance. Some patients (138 as per ICAD abstract, 125 as per presented poster) received a SPECT scan at baseline and again between 18 and 28 weeks into the study. Some patients (18 as per online abstract, 19 as per presented poster) received an FDG-PET scan at baseline and at around 26 weeks. The images were taken at participating sites in England and Singapore, and analyzed at the University of Aberdeen. With SPECT, patients on placebo showed reduced regional blood flow in AD-relevant areas, whereas patients on MTC did not. With FDG-PET, patients on MTC showed increased glucose use, whereas patients on placebo did not. (In both imaging modalities, the data on the posters indicated that the placebo groups were two to five years older than the MTC groups.)

On safety and tolerability, Wischik showed no data to the press but said in the subsequent scientific session that the major side effects were diarrhea, urinary urgency, and painful urination; there were also some dizziness and falls. Wischik said the side effect profile overall is similar to the three acetylcholinesterase inhibitors that are in wide use, but that diarrhea was more common.

On that point, it is worth noting that the talks excluded certain data that have become standard for pharmaceutical company presentations. For example, it is unclear how many people completed each arm of the trial, and what were the reasons for discontinuation. Other scientists in the field later wondered about how intent-to-treat analysis was handled, and how dropouts may have affected the power of the final data.

The 100 mg dose was ineffective. Wischik said that was due to interactions between the study drug and gelatine in the capsule wall. According to Wischik, this delayed absorption of the drug from the stomach to the intestine. However, rather than analyzing this group for what it was, warts and all, or leaving this group of patients out of the analysis, the investigators instead decided to combine the 100 mg group with the placebo group and compared this pooled set to the 30 mg and 60 groups. This is unusual. It means that all side effects people sustained from 300 mg MTC per day were tallied on the placebo side, making the 30 and 60 mg doses look better by comparison. (For sake of argument, it’s fair to assume that if Elan/Wyeth had pooled the placebo group with the ApoE4 carrier group, in whom the immunotherapy did not work so well, and then compared safety in this combined group with the ApoE4 non-carriers, that practice would have drawn protest.)

On MTC’s efficacy, as well, it is possible that including the highest dose with placebo affected the progression of the true placebo group, inflating the effect size. “If the investigators could have gotten statistical significance or a statistic with a p value Wischik noted that the study was double-blind for the full 84 weeks. However, methylene blue colors the urine green, raising the question of how a study with this substance can stay blinded at all. MTC also has been reported to sometimes tint eyes blue. (Sorry folks, not the iris. It goes into the white part.)

During the press briefing and again after his talk, Wischik was asked whether the company had included CSF tau/phospho-tau measurements in the trial. (Biomarker data are increasingly built into clinical trials in AD to establish evidence that the drug at hand hits its intended target. Research shows that CSF tau and phospho-tau go up before AD diagnosis and during its course.) Wischik replied that TauRx did not, because prior research made it impossible to predict whether CSF tau should go up or down in response to treatment with MTC, and that he would have had to pre-specify this issue prior to the trial. In fact, biomarker pre-specification is not necessary for all Phase 2 trials. For example, a Phase 2 trial of Prana’s PBT2 compound, conducted in Europe, measured CSF Aβ42 precisely to find out which way it would change in response to the drug, as well as to obtain some measure of how well the drug crosses the blood-brain barrier in humans (see subsequent ICAD story). It’s unclear at this point how well methylene blue enters the human brain.

Scientists from TauRx did, however, present posters on tau-transgenic mouse lines developed by scientists at University of Aberdeen who are also employees at WisTa Laboratories, a company also headed by Wischik. According to the ICAD abstracts, MTC given intravenously to these mice at 5 mg/kg for 17 days ameliorated tau pathology and cognitive deficits in the Morris water maze. Interested AD researchers can attempt to reproduce and expand these findings in a variety of tau-transgenic mouse lines known to the AD research community. For comparative listings, see tau-transgenic mice, double-crosses, others; strains available through The Jackson Laboratory.

In terms of the compound’s potency, Wischik’s presentation stated that MTC dissolves tangle filaments isolated from brain with an effective concentration (EC50) of 0.15 μMolar and stops tau aggregation in cells with an EC50 of 0.56 μMolar in cells. For background, a compound’s half-maximal inhibitory concentration (IC50) is used for in-vitro assays where there is no absorption, distribution, metabolism, excretion (ADME) component. That is, the assay simply contains a target and a drug, and the investigator can assume that 100 percent of the drug finds the target. The EC50 is typically used in cell culture and is lower than the IC50 because the compound has to cross the cell membrane and distribute through the cells. The key parameter for humans is the effective dose (ED50). That is because the ADME component is large as the drug spreads in the body, and only a small fraction finds the desired target, especially if it is in the brain and intracellular. Between the EC50 and the ED50, there can be two to three orders of magnitude difference. Because of these issues, companies typically prefer compounds that start out with an IC50 in the low nanomolar range.

Other researchers at ICAD noted that TauRx plans to exploit MTC interactions with α-synuclein, as well, to develop Rember^TM for Parkinson disease. The company’s website states this as well. These scientists questioned how specific the underlying molecular mechanism of a single pan-AD/tauopathy/synucleinopathy medication will prove to be.

MTC is available in the U.S. in tablet form. Called Urolene Blue, it is a grandfathered drug that predates the existence of the FDA, Wischik said. Urolene Blue is being used to treat the blood disorder methemoglobinemia. According to Wischik, it is also a mild antibiotic and in its earlier days used to be prescribed to treat urinary tract infections. According to Wikipedia, methylene blue was used against malaria a century ago.

TauRx has a use patent on a particular formulation of methylene blue, which Wischik said is purer, and hence possibly safer, than Urolene Blue. Both Rember^TM and TauRx have new Wikipedia pages, though the former was on the site’s watch list for deletion on August 3.

In summary, Wischik said: “These Phase 2 data show for the first time that it is possible to halt the progression of AD with an anti-tangle treatment.” Most commentators felt that if there is a real treatment effect in this data, even just a kernel, this would be exciting as a general signal that targeting tau aggregation can in fact make people better. Researchers tend to express healthy skepticism when data are not available in peer-reviewed publications. Here, too, they reserved judgment of whether, or how much of, this signal is real. Many will attempt to reproduce the reported effects in tau models with available forms of methylene blue. Scientists voiced enthusiasm about the general approach of inhibiting tau aggregation, but tempered it with some doubt about whether this particular dye formulation will prove to be enough like a drug to be approvable for chronic consumption. The ICAD presentation has raised awareness of this approach; both regulators and investors will scrutinize the quality of the data in detail before deciding on next steps.—Gabrielle Strobel.

References

Paper Citations

Wischik CM, Novak M, Thøgersen HC, Edwards PC, Runswick MJ, Jakes R, Walker JE, Milstein C, Roth M, Klug A. Isolation of a fragment of tau derived from the core of the paired helical filament of Alzheimer disease. Proc Natl Acad Sci U S A. 1988 Jun;85(12):4506-10. PubMed.
Goedert M, Wischik CM, Crowther RA, Walker JE, Klug A. Cloning and sequencing of the cDNA encoding a core protein of the paired helical filament of Alzheimer disease: identification as the microtubule-associated protein tau. Proc Natl Acad Sci U S A. 1988 Jun;85(11):4051-5. PubMed.

Other Citations

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Chicago: Dimebon Safe for 18 Months

08 Aug 2008

An oral antihistamine first marketed 25 years ago in Russia has emerged as a contender in the race to become the first Alzheimer disease drug with stronger, more lasting benefit than any of the drugs currently available. Dimebon drew notice for its ability to improve AD patients for at least 12 months on all of five cognitive measures in a recently published Russian trial of the compound (see ARF related news story). It’s odd to consider how a drug that fights sniffles could also boost cognition, and fresh data for how that might be turned more than a few heads at the International Conference on Alzheimer’s Disease (ICAD), held 26-31 July in Chicago. New data from the Russian trial extension also came in.

The new work extends earlier evidence that Dimebon stabilizes mitochondria. This target is distinct from those of available AD drugs and has traditionally attracted scant attention alongside big guns amyloid and tau. Interest in mitochondrial mechanisms in AD is growing, however, and the lecture hall was packed when Andrew Protter presented details on Dimebon’s proposed mitochondrial mechanism of action. Protter works for Medivation, Inc., the San Francisco-based company that is developing Dimebon as a treatment for Alzheimer and Huntington diseases.

The current batch of AD drugs work by inhibiting acetylcholinesterases (enzymes that break down a key learning/memory neurotransmitter) or NMDA receptors (proteins whose hyperactivation may underlie the degeneration of cholinergic cells). Dimebon, too, acts on these molecules, as shown in earlier work by Russian scientists (Bachurin et al., 2001; Bachurin et al., 2003). It does so more weakly, however, suggesting that other mechanisms mediate the drug’s cognitive effects if indeed those benefits hold up in larger trials. Because potency is a critical factor in drug development, researchers at Medivation have confirmed the earlier acetylcholinesterase findings in two sets of experiments—one using recombinant enzyme and another using fresh blood samples from three volunteers. In these follow-up studies presented at ICAD, Dimebon blocked acetylcholinesterase activity with a half-maximal inhibitory concentration (IC50) in the 31-72 μMolar range—several thousand-fold lower than that of the widely prescribed anti-cholinesterase drug donepezil. In NMDA receptor binding assays, Dimebon’s inhibition constant (Ki—a related measure of potency, defined as the concentration of test compound required to bind 50 percent of receptors in the absence of agonist) was 97 μMolar, compared with 0.5 μMolar for the AD drug memantine, the Medivation scientists claimed.

Another hint that Dimebon functions differently from current AD drugs came from its side effect profile in human studies: less than 3 percent of clinical trial participants who took Dimebon have reported gastrointestinal problems. In contrast, Protter said, more than 20 percent of AD patients taking donepezil routinely experience such symptoms. “Just looking at the low side effect profiles says (Dimebon) is not working like the other guys,” he told ARF.

Then how might it work? The same earlier studies provided initial clues to mitochondrial effects, and these have been confirmed and extended in unpublished experiments Protter presented at ICAD. A search for Dimebon’s mechanism of action that ranged from enzyme inhibition and cell-based activity assays to kinase profiling to commercial compound screening services identified mitochondrial enhancement under cellular stress conditions as the most potent activity. Mitochondrial dysfunction has been linked to aging and neurodegenerative diseases, including Alzheimer’s, in various ways. As the primary power plants of the cell, mitochondria churn out toxic compounds that can accelerate Aβ production in AD mouse models (see ARF related news story). A more recent study (see ARF related news story) highlights the importance of mitochondrial waste management: in transgenic mice with a mitochondrial enzyme defect leading to buildup of harmful aldehydes, age-related neurodegeneration kicked in and led to early death. (For more on how mitochondrial dysfunction might tie in with sporadic AD, see ARF Live Discussion).

To get a look at Dimebon’s mitochondrial activities, the researchers treated SK-N-SH human neuroblastoma cells with ionomycin, which reduces the electrochemical gradient across mitochondrial membranes, triggering apoptosis. They assessed mitochondrial function using JC-1, a dye that fluoresces red when it accumulates in healthy mitochondria and green when forced to remain in the cytoplasm due to collapsed mitochondrial membrane potentials. This allowed the researchers to gauge mitochondrial health as a percentage of cells fluorescing red versus green. The red/green ratio for cells hit with ionomycin dropped to 70 percent of that seen for untreated cells, Protter said. Picomolar concentrations of Dimebon brought the ratio up to 85-90 percent of control cells in a concentration-dependent fashion. As measured in MTT (3-[4,5-dimethyl-2-thiazolyl]-2,5-diphenyl-2H-tetrazolium bromide) assays, ionomycin treatment reduced cell viability to 40 percent of control cells, and picomolar Dimebon restored survival to 75-85 percent. Separate experiments examined mitochondrial function in a related neuroblastoma line (SH-SY5Y) using a different probe (tetramethyl rhodamine methyl ester, or TMRM). With this, too, the researchers reported that pre-treatment with one nanoM Dimebon protected cells from ionomycin-induced stress. Together, these findings suggest that Dimebon’s mitochondria-stabilizing effects are potent and correlate with improved cell viability, Protter said.

Because dendritic mitochondria are important for synaptic plasticity, a process critical for learning and memory (see ARF related news story), the company researchers examined Dimebon’s effects on the growth of neurites. In primary rat cortical neurons cultured for three days with either brain-derived neurotrophic factor (BDNF) or Dimebon, picomolar concentrations of Dimebon increased neurite outgrowth as well as did BDNF applied at its maximally effective concentrations, Protter reported.

“Dimebon has shown quite encouraging results in the clinic,” Protter said in a post-ICAD phone conversation. “Combined with the pharmacology studies we’ve done thus far, we can say that Dimebon is working with a novel mechanism.” So far, signs point to the mitochondria, though researchers have yet to clearly demonstrate that Dimebon binds these organelles. Protter said Medivation has not been keen thus far on doing experiments with purified mitochondria. For starters, he said that it is hard to isolate the organelles cleanly. In addition, the literature is increasingly ascribing functional roles to proteins that are bound to mitochondria, yet these interactions could be disrupted in the purified preparations. Protter claimed the company has preliminary data showing that Dimebon “is affecting proteins that are bound to mitochondria” but did not show that data at ICAD. No independent confirmation of these data by other laboratories is available to date.

Also at ICAD, Jeffrey Cummings of the University of California, Los Angeles, presented clinical results of the six-month open-label extension of Dimebon’s Phase 2 trial in Russia, which brought exposure to the drug out to 18 months in these patients with mild to moderate AD. In this extension, patients who had been on drug during the blinded phase continued on Dimebon. Patients who had been randomized to placebo crossed over to Dimebon. This created, at the 18-month time point, a mixture of people who had been on drug for 18 months and had actually still enjoyed a slight improvement over baseline at 12 months, and of other people who been declining in the placebo group for 12 months. People who had been on drug all along declined somewhat after 12 months, but ended the 18-month time point with only an ADAS-Cog change of about minus 2 from the start of the trial. They appeared to hold their own, i.e., around baseline, on a psychiatric rating scale, as well. People who had previously been on placebo at 18 months remained stable at the lower level of cognitive function at which they had ended their 12 months on placebo.

Perhaps the most straightforward value of open-label extensions lies in safety and tolerability, because they give an indication of long-term use. On that score, “no safety or tolerability issues came up that would concern us,” Cummings said. The one side effect reported more frequently in the treatment group was depression. This depression was self-reported, not measured by depression scales, raising the question of whether people might become more depressed as their cognition improves slightly and they regain insight into their still-difficult condition. Cummings noted that a Phase 3 trial of Dimebon monotherapy held at 30 sites in the U.S., as well as in Europe and South America, is presently enrolling. Cummings added that the FDA had accepted the Phase 2 trial, which was conducted in Russia with the guidance of established clinical leaders including Rachelle Smith Doody, of Baylor College of Medicine in Houston, as a first pivotal trial in the process, and that the Phase 3 would be the second pivotal trial to support a New Drug Application. If all goes well, that is.—Esther Landhuis and Gabrielle Strobel.

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References

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Paper Citations

Bachurin S, Bukatina E, Lermontova N, Tkachenko S, Afanasiev A, Grigoriev V, Grigorieva I, Ivanov Y, Sablin S, Zefirov N. Antihistamine agent Dimebon as a novel neuroprotector and a cognition enhancer. Ann N Y Acad Sci. 2001 Jun;939:425-35. PubMed.
Bachurin SO, Shevtsova EP, Kireeva EG, Oxenkrug GF, Sablin SO. Mitochondria as a target for neurotoxins and neuroprotective agents. Ann N Y Acad Sci. 2003 May;993:334-44; discussion 345-9. PubMed.

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Chicago: Bapineuzumab’s Phase 2—Was the Data Better Than the Spin?

11 Aug 2008

It is tempting to say that the story of how Elan/Wyeth’s antibody therapy fared at the 11th International Conference on Alzheimer’s Disease, held last month in Chicago, is one of the stranger tales in present-day AD drug development. (Then, again, weren’t all the recent ones a little strange? Take Alzhemed’s inglorious demise and subsequent resurrection as a nutraceutical, or Flurizan’s sudden death just weeks after its sponsor had announced a European commercialization agreement.)

Bapineuzumab’s story is different, of course. This humanized monoclonal anti-Aβ antibody is very much alive as an experimental treatment. Four separate Phase 3 trials in the U.S., Canada, and Europe are presently enrolling up to 4,100 Alzheimer disease patients. Elan/Wyeth are running an open-label extension of the present Phase 2 trial and are gearing up to test subcutaneous bapineuzumab delivery in a Phase 2 trial. But like the previous two anti-amyloid drugs, bapineuzumab rode to ICAD on high expectations, and its luster has dimmed somewhat after the company’s presentation there. This is not only because investors promptly dumped Elan stock but also because scientists felt that the presentation could have been more straightforward.

What happened? The company presented an overall negative result balanced against a positive effect on a subgroup of patients who don’t carry the AD risk allele ApoE4. Many listeners did not buy it but interpreted the presentation as having massaged the data. In conversations with scientists, most of whom spoke privately, reactions ranged from “they stratified until they got the result they wanted” to “the effect on the ApoE4 negatives makes sense; it’s reassuring,” to “told you so, anti-amyloid drugs lead nowhere” to “I am cautiously optimistic.” Several agreed, however, that the way in which Elan/Wyeth sought to portray the Phase 2 data in the best possible light backfired. In essence, emphasizing a post-hoc pharmacogenomic interpretation shortchanged the trial’s overall decent results, they said. Here’s the lowdown:

At ICAD, Sid Gilman of the University of Michigan, Ann Arbor, presented a more detailed version of the previous top-line data Elan had released on June 16 (see ARF related news story). In this talk, Gilman said that the trial had randomized 234 patients to receive six infusions, one every 13 weeks, of one of four different treatment doses or placebo. He reported that 26 percent of patients dropped out of the treatment groups versus 21 percent in the placebo group.

As a Phase 2, this was first and foremost a safety trial. The safety data Gilman showed received less attention than the efficacy data summarized below, but several scientists noted afterward that some of the reported side effects might bear watching. The most widely discussed side effect is vasogenic edema. Gilman showed that 12 patients developed this, all in the treatment group. Ten occurred in ApoE4 carriers and two in non-carriers, though the trial had twice as many people with ApoE4 than without, meaning that this pharmacogenomic distinction works out to roughly 10/4 rather than 10/2 in this trial. The edemas showed up on MRI and all resolved. Most caused few clinical symptoms, but one patient needed steroid treatment. Six people dropped out of the study; the other six resumed treatment. It is unclear at present how troublesome these edemas really are, and whether they may eventually be deemed an acceptable side effect for treating a deadly disease such as Alzheimer’s. This question generated animated discussion at ICAD and is the subject of ongoing research. Some scientists felt that vasogenic edemas may reflect the underlying mechanism of the drug, as Aβ peptide gets released from plaques, protein concentrations in the interstitial space and in capillary beds rise, osmotic pressure rises along with that, and clearance mechanisms, perhaps some involving ApoE, gradually dispose of the surplus protein. Others doubted whether such a therapy would get the FDA’s blessing.

Concern over vasogenic edema overshadowed the fact that Gilman’s presentation included additional side effects that were more common in the treated patients. The list of unwanted effects that occurred twice as often as in placebo, and in more than 5 percent of patients on active drug, included back pain, anxiety, vomiting, hypertension, weight loss, paranoia, skin lacerations, gait disturbance, and muscle spasms. Syncope and cataracts, among some other side effects, occurred in fewer than 5 percent of patients on active drug. Adverse events were generally mild or moderate and transient, Gilman said. Three men in treatment groups died. According to Gilman, one died of an aortic aneurysm, one succumbed to a fall and Alzheimer’s, and one to infection and Alzheimer’s. None of these deaths was attributed to study drug.

Moving on to efficacy signals, Gilman then briefly noted that the companies had decided to change the originally planned intent-to-treat analysis away from a slope analysis, which shows how the patients do at every time point, to a different analysis he called “MITT repeated measures model without assumption of linearity.” This mouthful essentially compares baseline to the final assessment at 78 weeks/19.5 months and incorporates the time points in between in ways that are not directly shown. That left many in the audience puzzled. To some, the decision to show the data this way reflects a biological argument that a plaque-clearing antibody, unlike a conventional drug, cannot be expected to show a linear effect over time as it might make a patient temporarily worse before it makes him/her better in the long run. To others, this change in the statistical model to a complex one they don’t fully understand implied that Elan/Wyeth might not want to show outcome data over all seven time points because those data looked bad. These scientists would have welcomed trend lines, i.e., graphs plotting the slope of outcome measures along the time axis, because that is how they have come to expect clinical trial data to be shown.

Gilman then showed efficacy data according to the modified analysis. In the total population, the trial posted a 2.3-point improvement on the ADAS-Cog, with a p value of 0.078. On the NTB battery, the improvement was 0.13 with a p value of 0.068. Thus, the trial failed to achieve statistical significance on pre-specified endpoints. Gilman stated this clearly and then moved right on to present a pharmacogenomic stratification of the data. But wait, not so fast! Commentators noted that this overall finding is important. “This is a respectable efficacy signal for a Phase 2 trial. It is a consistent effect, the trial is powered to detect it, and you cannot expect a larger effect from a trial of 230 patients,” Lon Schneider of the University of Southern California told this reporter. Other scientists agreed with this assessment.

In post-hoc analysis, Elan/Wyeth split the patients into those who received all six injections, i.e., the “completers,” versus those who missed one or more doses. Analyzed in this way, the patients who completed the trial (N = 78, ApoE4 carriers and non-carriers combined), showed a 4.3-point improvement over placebo on the ADAS-Cog with a p value of 0.003. “My gut feeling is that [bapineuzumab] is probably modestly effective. But my other gut feeling is that they are chasing their tail by emphasizing efficacy in the E4 non-carriers,” said Schneider.

What, then, is this pharmacogenomic result? Prior research suggests biologic differences between ApoE4 carriers and non-carriers, which might be expected to affect the course of an amyloid-removing therapeutic approach. ApoE4 carriers tend to get the disease earlier than people with ApoE3 or the rare allele ApoE2, and they develop more amyloid in their brain (Rowe et al., 2007). Numerous studies have implicated ApoE in amyloid deposition and removal, and a recent one showed that ApoE4 is less effective at clearing Aβ than the other two alleles (Jiang et al., 2008). Hence, Elan/Wyeth split trial participants into ApoE4 carriers and non-carriers, and then analyzed these groups separately. This created some imbalance between the groups because two-thirds of the patients in the trial happened to have an ApoE4 allele. At 78 weeks, the ApoE4 carriers showed a respectable 4.5-point improvement in ADAS-Cog on the lowest dose but a worsening at the highest dose. Results were inconsistent across the other outcome measures (DAD, NTB, CDR-SB). Some 70 ApoE carriers were in the four treatment groups combined. Of these ApoE4 carriers, a total of 42 across all treatment groups completed the study, i.e., about 10 per group on average. They showed a larger and more consistent benefit. ApoE non-carriers (N = 47) showed a benefit primarily with the second-lowest dose. The completers among this group numbered only 36 patients, i.e., on average less than 10 per dose comparison. But they had larger effects, some reaching a whopping 20 points on ADAS-Cog. (This was relative to an 11-point decline in the placebo group.)

Looking at the best result of all those different groupings, it seemed that a small subset of patients, i.e., the ApoE non-carriers who received the second-lowest of the four doses six times, responded truly well by 78 weeks. The presentation did not state how many people were in this group; it could have been 30, it could have been one. At the same time, the overall appearance of the various bar graphs of these subgroups was one of considerable variation across doses and across the different cognitive/clinical assessments. Subgroup analysis gets statistically weaker the smaller the group sizes become, opening it to criticism. “It could be noise,” Schneider cautioned.

Gilman did show slope curves for one subgroup, i.e., the ApoE4 non-carriers. They suggest that the curves diverged for ADAS-Cog after 36 weeks, on the NTB and CDR-SB right after baseline, but not until 63 weeks on the DAD.

Furthermore, the trial featured three kinds of biomarker, two imaging, and one fluid. Scanned at 71 weeks, neither brain volume nor ventricular volume (both measured by the boundary shift integral) changed in the total population, but again, stratification by ApoE appeared to yield a signal. ApoE carriers on bapineuzumab showed an increase in ventricular brain volume over placebo. Whether this is good or bad is unclear, Gilman said. It could reflect ongoing protein clearance or something else. ApoE4 non-carriers on bapineuzumab suffered less brain atrophy than those on placebo, and here, too, a slope diagram of diverging curves was shown. The CSF biomarker measurements, gleaned from spinal taps at baseline and 52 weeks in 20 antibody- and 15 placebo-treated patients, showed a trend toward lower phospho-tau in treated versus placebo with a p value of 0.056. CSF Aβ and total tau were the same between treated and placebo groups.

“This was a relatively small Phase 2 trial, and it was not realistic that a trial of this size with multiple doses would yield a definitive proof of efficacy. The trial was clearly a success, however, in demonstrating the feasibility of completing multiple IV infusions with intensive safety monitoring in AD patients,” Reisa Sperling of Brigham and Women’s Hospital in Boston, commented by e-mail. “There are some potential safety issues that will need to be followed closely in the larger trials. There may be some encouraging signals that we are on the right track with immunotherapy, but it is important not to over-interpret post-hoc analyses with small sample sizes. We will just have to wait and see."

It is notoriously difficult to extract a solid efficacy signal from Phase 2 trials for AD drugs (see ARF ICAD story). Interestingly, Elan/Wyeth’s immunotherapy competitor Eli Lilly and Company dealt with this dilemma in the opposite way. Rather than slice the data to find efficacy in Phase 2, Lilly placed its bets on biomarker characterization alone and decided to move to Phase 3 just based on that and safety (see upcoming ICAD story). It’s fitting, perhaps, to end with a philosophical remark from a leading basic researcher in the field. “We are at a turning point with regard to developing therapeutics. Everybody is very nervous about the outcome of current anti-amyloid trials because so much is at stake,” said Bart de Strooper at K.U. Leuven and the Flanders Institute for Biotechnology (VIB) in Belgium. “The basis for this test of the amyloid hypothesis is strong, but clearly amyloid is not everything that is going wrong in AD. We should interpret any negative results in this context and try to learn from it. We also need to think about prevention. We need to develop safer medicines that we can test for prevention.”—Gabrielle Strobel.

References

News Citations

Paper Citations

Rowe CC, Ng S, Ackermann U, Gong SJ, Pike K, Savage G, Cowie TF, Dickinson KL, Maruff P, Darby D, Smith C, Woodward M, Merory J, Tochon-Danguy H, O'Keefe G, Klunk WE, Mathis CA, Price JC, Masters CL, Villemagne VL. Imaging beta-amyloid burden in aging and dementia. Neurology. 2007 May 15;68(20):1718-25. PubMed.
Jiang Q, Lee CY, Mandrekar S, Wilkinson B, Cramer P, Zelcer N, Mann K, Lamb B, Willson TM, Collins JL, Richardson JC, Smith JD, Comery TA, Riddell D, Holtzman DM, Tontonoz P, Landreth GE. ApoE promotes the proteolytic degradation of Abeta. Neuron. 2008 Jun 12;58(5):681-93. PubMed.

Chicago: Lilly’s Antibody Appears to Do No Harm, But Will It Help?

13 Aug 2008

As new experimental drugs are getting ready for clinical testing against Alzheimer disease, it’s instructive to learn how pharmaceutical companies are navigating the transition from Phase 2 to 3. At this juncture, they sometimes base the decision on whether to start a long and costly Phase 3 program on less than rock-solid efficacy data (see ARF ICAD story). At the International Conference on Alzheimer’s Disease, held 26-31 July in Chicago, the pharmaceutical company Eli Lilly and Company took the opposite approach to Elan/Wyeth. Both competitors are developing a humanized therapeutic antibody, which would be the first biologic treatment for AD. Elan/Wyeth uses a humanized version of an N-terminal Aβ antibody that recognizes forms of Aβ peptide in amyloid plaques; Lilly uses a mid-region antibody (m266) that captures soluble Aβ and tests the peripheral sink hypothesis of Aβ clearance (DeMattos et al., 2002). But rather than running a sizable and long Phase 2 trial (230 patients/19 months in the case of Elan/Wyeth), and scrutinizing its results for hints for efficacy (see ARF ICAD story), Lilly decided to sidestep clinical efficacy entirely. It instead ran a small and short (52 patients/3 months) Phase 2 trial that was heavily geared toward fluid biomarkers, and used those results as the basis to move into Phase 3 by 2009. Safety and biomarkers, but not efficacy, are also what drove Lilly's Phase 2 testing of its γ-secretase inhibitor, which is currently enrolling for Phase 3 (see Fleisher et al., 2008). Here are the details:

At ICAD, Eric Siemers and Ron DeMattos of Eli Lilly and Company in Indianapolis separately presented results of the trial and its attendant biomarker research, respectively. The Phase 2 trial tested four different doses of LY2062430, a therapeutic antibody, in people with mild to moderate AD in order to determine which doses and which interval between doses to use in Phase 3. The patients received 12 weekly infusions of placebo or antibody arranged to contain either 100 mg every four weeks, 100 mg every week, or 400 mg every four weeks, or 400 mg every week. There were 10 or 11 patients per group. The injections and most assessments ended at week 12, but patients will be followed for a year. At ICAD, Siemers showed interim data from day 112/week 16, i.e., partway into the follow-up period. The study was recently locked for definitive analysis, Siemers said.

The trial assessed its primary outcome of safety by looking for host immune responses against the Lilly antibody and for fluid or inflammatory signals by MRI and CSF, in addition to the usual adverse event monitoring. It also measured ADAS-Cog at the beginning and at 12 weeks, did plasma and CSF research, and a little neuroimaging.

On safety, Siemers said that no side effect cropped up that could be attributed to the antibody. None of the 42 treated patients showed evidence of edema, microhemorrhage, inflammation, nor were there laboratory abnormalities or troubling signs of immune response. Five of the patients, four of them dosed weekly, showed antibody titers to LY2062430, but these titers had no detectable effects. “The safety data could not be better,” Siemers told this reporter.

If the antibody is so safe, is it doing anything at all? In plasma, it caused both Aβ40 and 42 concentration to shoot up several orders of magnitude, as was also seen in the Phase 1 trial of this compound (see ARF Keystone story). The scientists showed a graph with different-sized saw tooth zigzag lines of plasma Aβ levels curving up in response to each injection, and they calculated the pharmacodynamics to conclude that monthly injections would be acceptable for Phase 3.

A tiny fraction, some 0.1 percent, of the antibody crosses into the CSF. There, too, total Aβ40 and 42 concentrations increase up to four orders of magnitude in a dose-dependent way, Siemers reported. “We expected this data,” Siemers said. But there was a surprise, as well. The Lilly scientists developed assays to distinguish in CSF between free Aβ and Aβ bound to antibody, doing so separately for Aβ40 and 42. At ICAD, they reported that the unbound, i.e., free, CSF Aβ40 decreased with greater doses of antibody, whereas the free CSF Aβ42 increased with rising doses of antibody. According to the poster, the Aβ40 drop happened in the 3,000 picogram/ml range, the Aβ42 rise in the 200 picogram/ml range. (CSF Aβ42 concentrations are low in people with AD.)

What does this mean? “We had to think about this,” Siemers said. “To us it suggests that the plaques are beginning to dissolve and Aβ42 slowly leaves the brain.” Siemers said further that at the time of spinal tap, all injected antibody is saturated with Aβ, so that additional Aβ42 coming off the plaques would stay free. Siemers briefly mentioned brain imaging with IMPY in this trial. However, experts in amyloid imaging later noted that IMPY is too poor a marker of amyloid to support firm conclusions, adding that Lilly will hopefully be able to use PIB or one of the 18F amyloid imaging agents that are being tested clinically.

For his part, DeMattos described in more detail fluid biomarker research aimed at characterizing how the humanized m266 antibody acts on soluble Aβ. In essence, DeMattos was looking for markers that would be able to track whether equilibria between different pools of Aβ in the body’s periphery and CNS shift after treatment. DeMattos developed an acid urea gel technique that allowed him to resolve full-length Aβ from within a mixture of modified or truncated species. He then used the 3D6 and the 21F12 antibodies to label the peptides on Western blots, on which, he noted, Aβ species run to their true size. This approach visualized a pattern of different truncated Aβ species extracted from human AD brain. It confirmed published work by others. DeMattos estimated that about 75 percent of Aβ extracted from human AD brain and ending at amino acid 42 is truncated. In DeMattos’s hands, a small fraction of that appeared to be 3-42 pyroglutamate derivatives of Aβ (see ARF related Keystone story), which is thought to be an early aggregating form of Aβ.

Soluble Aβ in brain interstitial fluid is generally thought to be in equilibrium with plaque Aβ, which draws Aβ out of solution. The hope is that m266 will do the opposite, i.e., draw Aβ out of plaques and toward clearance. It’s unproven at present that increasing free CSF Aβ42 will do that, but the new biomarker data hint that this may happen, DeMattos said. When applied to the plasma samples taken during the Phase 2 trial, the acid urea technique detected an accumulation of not only full-length Aβ but also a range of N-truncated fragments that probably originate in the brain. This includes, for example, the pyroglutamate forms. They were never before seen in biological fluids and are considered specific to plaques, DeMattos said. It also includes what DeMattos called “fragment 2,” an as yet uncharacterized Aβ snippet that might be plaque-specific as well. “We see a robust dose-dependent and time-dependent accumulation of this fragment 2 series. We think it is being liberated from plaque. We think we are shifting the equilibrium,” DeMattos said.

Curiously, DeMattos also reported seeing a higher concentration of pyroglutamate Aβ fragments in plasma of some of 16 healthy controls, who had received a single injection of m266. They were people who were also positive for IMPY imaging, suggesting to DeMattos that they had amyloid in their brain and that the bolus of m266 might have drawn some of it out. Overall, the increase in free Aβ42 to DeMattos means that m266 is dissolving various different forms of Aβ out of plaques, from where they make their way to the CSF and then to the blood. The ICAD presentations included no data on whether all that Aβ bound up in plasma is then being degraded and excreted at pace. “The importance to us of this biomarker data is that we know the drug hits its target,” DeMattos said. Added Siemers “We have made a decision to go to Phase 3 with this antibody, and will be starting in 2009.”—Gabrielle Strobel.

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References

News Citations

Paper Citations

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.
Fleisher AS, Raman R, Siemers ER, Becerra L, Clark CM, Dean RA, Farlow MR, Galvin JE, Peskind ER, Quinn JF, Sherzai A, Sowell BB, Aisen PS, Thal LJ. Phase 2 safety trial targeting amyloid beta production with a gamma-secretase inhibitor in Alzheimer disease. Arch Neurol. 2008 Aug;65(8):1031-8. PubMed.

Chicago: APP Roles in Caspase Gene Regulation, Muscle Function?

18 Aug 2008

One way to get the lowdown on someone is to check out the person’s closest friends and colleagues. Similarly, some Alzheimer disease researchers are teasing out the biological function of amyloid precursor protein (APP) by studying “whom” it associates with, and puzzling out what these partners might be doing in neurons. In a plenary talk at the International Conference on Alzheimer’s Disease (ICAD) held last month in Chicago, Joseph Buxbaum of Mount Sinai School of Medicine in New York presented new data suggesting that an APP interactor, Fe65, associates with teashirt (a zinc-finger transcription factor) and SET (a histone acetyltransferase inhibitor) to form a gene-silencing complex that binds caspase-4. In the study, AD appears linked with higher RNA levels of caspase-4 and decreased expression of Fe65 and teashirt, raising the intriguing possibility that APP processing could impinge on transcriptional regulation of an inflammatory caspase. Well-known for their role in apoptosis, some caspases, including caspase-4, also play a role in the maturation of cytokines that mediate inflammation (for reviews, see Nadiri et al., 2006 and Martinon and Tschopp, 2007), a process linked to AD in beneficial and harmful ways (for a recent example, see ARF related news story). Another study at ICAD—presented on a poster from the lab of Ulrike Müller at the University of Heidelberg in Germany—addressed APP’s physiological function using a different approach. Müller’s group studied the role of individual APP proteolytic fragments in mice lacking full-length APP and related family members.

Infamous as the cell-surface protein that gets snipped to form the Aβ peptides gumming up the brains of AD patients, APP has physiological functions about which comparatively little is known. Clues surfaced when an earlier study (Cao and Südhof, 2001) suggested that the APP cleavage product containing its cytoplasmic tail (aka AICD, or APP intracellular domain) may play a role in transcriptional activation by stably associating with the adaptor protein Fe65, and Tip60, a histone acetyltransferase (see ARF related news story). A number of candidate gene targets for this complex have subsequently been identified, but no such gene has since received broad-based replication and consensus, leaving the issue shrouded in mystery. Buxbaum and colleagues stepped up to the challenge by trying to identify other Fe65 interactors. Fe65 has two phosphotyrosine binding modules, the second of which (PTB2) binds APP (Fiore et al., 1995). To get at what might bind the first (PTB1), Yuji Kajiwara, an M.D./Ph.D. student in Buxbaum’s lab, did a yeast two-hybrid screen and pulled out teashirt (Tsh), a Drosophila homeotic protein. Mammals have three Tsh proteins, all of which were found to bind Fe65 in confocal microscopy experiments and immunoprecipitations from primary neuronal cultures, Buxbaum reported at ICAD. Based on the yeast two-hybrid data, he said Tsh3 was the most interesting to follow up, and this protein was used in most of the transcription studies presented subsequently.

In cells transfected with APP-Gal4 and various combinations of tagged Fe65 and Tsh3, the researchers showed that Tsh3 acts as a repressor. Fe65 profoundly activated gene expression by AICD, and Tsh3 knocked it down, Buxbaum reported in Chicago. Tsh3 did not seem to be a general inhibitor of transcription, as it only behaved as a repressor when bound to Fe65. Further experiments suggested that Tsh3’s repressive effects were mediated by recruitment of histone deacetylases (HDAC 1/2)—enzymes that promote gene silencing by removing acetyl groups from core histones, thereby making the associated DNA more compact and less accessible to transcription factors. Fe65 also binds the nucleosome assembly factor SET, and this interaction appears necessary for the transcriptional activation mediated by Fe65 (Telese et al., 2005). SET is a subunit of the inhibitors of acetyltransferases (INHAT) complex, which binds histones and blocks their access to acetyltransferases. Histone acetyltransferases (HATs), which oppose the function of HDACs, promote gene transcription by transferring acetyl groups onto histones, thereby loosening chromatin structure and increasing exposure of the associated DNA to transcription factors. (For a freely accessible review on HATs and HDACs in neurodegeneration, see Saha and Pahan, 2005.) Buxbaum boiled down his take on the proposed Fe65/Tsh/HDAC/SET complex (see figure below) in a phone conversation with this reporter. “You're taking off acetyl groups that are there (HDAC), and you're blocking further addition of acetyl groups (SET),” he said. “It's kind of a double mechanism to make sure the genome is shut down.”

The APP interactor, Fe65 (unlabeled green protein with two phosphotyrosine binding (PTB1 and PTB2) and WW domains), associates with SET and Tsh in proposed gene-silencing complex targeting caspase-4. Image credit: Joseph Buxbaum

Shifting attention to what this gene-silencing complex might target, Buxbaum and colleagues found a promising candidate—caspase-4—using a macroarray containing several hundred genes enriched for AD-related factors. The macroarray was probed using RNA from human neuroglioma cells stably transfected with Fe65 and Tsh3. In these same cells, the researchers showed that high expression of Fe65 and Tsh3 inhibited caspase expression. Conversely, knocking down expression of Fe65, Tsh3, or both using antisense RNAs led to increased caspase-4 expression, Buxbaum reported. Chromatin immunoprecipitations using nuclear extracts from these cells demonstrated direct physical interaction between Tsh and Fe65 proteins and the caspase-4 promoter.

Do these interactions have anything to do with AD? In Chicago, Buxbaum presented several lines of evidence to support a “yes” claim. First, data from postmortem human brain samples seem to indicate that Tsh/Fe65 and caspase-4 expression are anti-correlated—that is, Tsh3 expression drops while caspase-4 expression rises in early AD. These hundred or so samples came from the Mount Sinai Brain Bank and included controls and varying degrees of disease severity by several measures, including CDR scores and plaque pathology. “Caspase-4 goes from low to maximally expressed as you go from no to some neuritic plaques,” and remains high as disease progresses, Buxbaum said.

The other piece of evidence presented at ICAD to suggest that the Fe65/teashirt interaction is relevant to Alzheimer’s came from a search for Tsh gene variants in AD. Genome-wide scans turned up two AD-associated single-nucleotide polymorphisms (SNPs) in Tsh3 (p-values 0.00525 and 0.0095), and one in Tsh1 (p-value = 0.000205). For both Tsh3-associated SNPs, the risk alleles were associated with reduced Tsh expression, Buxbaum said, adding that separate cohorts will be analyzed to see if these effects—as yet unpublished—can be replicated.

The obvious next step for evaluating caspase-4 regulation by Fe65/Tsh is to analyze this activity in AD mouse models, but there’s a caveat: mice don’t have caspase-4. This could be interesting in and of itself, Buxbaum noted, as it might explain in part why most current AD mice show little neurodegeneration compared with what happens in human disease. Buxbaum’s group is collaborating with Greg Elder, also at Mount Sinai School of Medicine, to make a transgenic mouse containing the entire human caspase-4 gene. Thus far this has proven non-trivial, as the caspase genes are clustered in the human genome, making it hard to cleanly isolate caspase-4 with all its genetic elements. If the researchers succeed in making the humanized caspase-4 mouse, they will cross it to one of the APP-overexpressing transgenic lines. If they see increased neurodegeneration and/or more tau pathology in the caspase-4-expressing APP mice, these animals might serve as better disease models, Buxbaum said, noting recent studies (Colton et al., 2006 and Wilcock et al., 2008) in which knockout of an immune modulator—nitric oxide synthase 2—seemed to make mouse models better at recapitulating human disease (cerebral amyloid angiopathy and AD). Other studies (for example, Pompl et al., 2003) have looked more generally at caspases in AD, showing their expression goes up at the early stages of disease, but these findings are merely correlative. Caspase-4/APP transgenic mice would enable researchers to more clearly address the role of inflammatory caspases in disease progression—for instance, by seeing if the mice are improved by methods that regulate caspase-mediated inflammation.

Teasing Out APP Function in Mice: Knock-ins, Knockouts Galore
Uncovering biological roles for APP has proved challenging in part due to the presence of its close relatives APLP1 and APLP2. Ulrike Müller’s group had shown previously that these proteins in mice have overlapping and non-redundant functions (see ARF related news story). For instance, mice lacking APP survive, but APP/APLP2 double knockouts die after birth, as do mice with both APLP1 and APLP2 genes knocked out. On the other hand, APP/APLP1 double knockouts survive and appear phenotypically normal.

For a more detailed look at APP’s biological function, Müller and colleagues used a knock-in approach whereby they inserted genes encoding proteolytic fragments of APP into the mouse genome precisely where endogenous APP had been knocked out. They generated two lines of knock-in mice—one expressing the secreted APP ectodomain (APPsα) and another expressing a C-terminal truncation lacking the YENPTY motif essential for APP interactions with phosphotyrosine-binding domains of other proteins including Fe65. Based on brain and body weight assessments and various learning and motor tests with these mice, the researchers concluded that the APP C-terminus is dispensable and that APPsα is sufficient to mediate the physiological functions of APP (Ring et al., 2007). They wondered whether APLP2 and APLP1 might compensate for the loss of APP function in the APPsα-expressing knock-in (APPsα-KI) mice. To this end, analysis of APPsα-KI/APLP2-KO mice was described in the ICAD poster by first author Sascha Weyer and colleagues.

As it turns out, expression of APPsα rescued the postnatal lethality of the APP/APLP2 double knockouts. Sixty percent of the new mice, which lack APLP2 and express no full-length APP but only its secreted ectodomain, survived into adulthood. This finding was reported last year (see ARF SfN meeting report) and with expanded analysis at ICAD. The surviving mice had motor impairments by several tests including grip strength analysis, open field, rotarod, and beam walking. The researchers also found some neuromuscular junction abnormalities—fragmented post-synaptic acetylcholine receptor clusters that were associated with increased frequency and amplitude of spontaneous miniature endplate potentials—in APPsα/APPsα APLP2-/- mice, compared with APPsα-/+ APLP2-/- animals. The authors conclude that APPsα is important for muscle function through interaction with cell-surface APP interactors. Though these interactions may play a role in ensuring survival, the remaining deficits would seem to indicate that the APP C-terminus—which binds Fe65 and other cytoplasmic proteins—could still mediate key APP functions.—Esther Landhuis.

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References

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Fiore F, Zambrano N, Minopoli G, Donini V, Duilio A, Russo T. The regions of the Fe65 protein homologous to the phosphotyrosine interaction/phosphotyrosine binding domain of Shc bind the intracellular domain of the Alzheimer's amyloid precursor protein. J Biol Chem. 1995 Dec 29;270(52):30853-6. PubMed.
Telese F, Bruni P, Donizetti A, Gianni D, D'Ambrosio C, Scaloni A, Zambrano N, Rosenfeld MG, Russo T. Transcription regulation by the adaptor protein Fe65 and the nucleosome assembly factor SET. EMBO Rep. 2005 Jan;6(1):77-82. PubMed.
Saha RN, Pahan K. HATs and HDACs in neurodegeneration: a tale of disconcerted acetylation homeostasis. Cell Death Differ. 2006 Apr;13(4):539-50. PubMed.
Colton CA, Vitek MP, Wink DA, Xu Q, Cantillana V, Previti ML, Van Nostrand WE, Weinberg JB, Weinberg B, Dawson H. NO synthase 2 (NOS2) deletion promotes multiple pathologies in a mouse model of Alzheimer's disease. Proc Natl Acad Sci U S A. 2006 Aug 22;103(34):12867-72. PubMed.
Wilcock DM, Lewis MR, Van Nostrand WE, Davis J, Previti ML, Gharkholonarehe N, Vitek MP, Colton CA. Progression of amyloid pathology to Alzheimer's disease pathology in an amyloid precursor protein transgenic mouse model by removal of nitric oxide synthase 2. J Neurosci. 2008 Feb 13;28(7):1537-45. PubMed.
Pompl PN, Yemul S, Xiang Z, Ho L, Haroutunian V, Purohit D, Mohs R, Pasinetti GM. Caspase gene expression in the brain as a function of the clinical progression of Alzheimer disease. Arch Neurol. 2003 Mar;60(3):369-76. PubMed.
Ring S, Weyer SW, Kilian SB, Waldron E, Pietrzik CU, Filippov MA, Herms J, Buchholz C, Eckman CB, Korte M, Wolfer DP, Müller UC. The secreted beta-amyloid precursor protein ectodomain APPs alpha is sufficient to rescue the anatomical, behavioral, and electrophysiological abnormalities of APP-deficient mice. J Neurosci. 2007 Jul 18;27(29):7817-26. PubMed.

Chicago: Flurizan Postmortem

20 Aug 2008

At the International Conference for Alzheimer’s Disease, 26-31 July in Chicago, investigators reported results on a range of different clinical trials. There were no breakthroughs but the field is shifting. While some new drug candidates are entering the pipeline, others have dropped out and saw their postmortem discussion at ICAD. Here is a closing summary on one of them:

On July 2, the sponsors of Flurizan/tarenflurbil had announced in four terse sentences that this γ-secretase-modulating agent had fallen flat in its definitive Phase 3 trial and was finished as a development product (see ARF related news story). At ICAD, Robert Green of Boston University gave a detailed presentation about the disappointing trial. The Phase 2 trial had seemed to give some hint of clinical efficacy for the highest dose in patients with mild AD (see ARF related news story on trial design), and formal publication of the trial in Lancet recently had fanned hopes further that Flurizan might do some good at this disease stage (Wilcock et al., 2008).

According to the Phase 3 results, it did not. On both primary efficacy endpoints, the ADAS-Cog and the ACDS activities of daily living scales, the treatment and placebo curve overlapped almost completely. There was no effect whatsoever in the group as a whole. And while the overall side effect profile was similar between placebo and treatment groups, anemia, infections, and gastrointestinal ulcers did appear more often in people on Flurizan, Green reported.

According to Green, some 20 percent of patients stayed stable or improved. However, neither pre-specified nor post-hoc analyses, of ApoE genotype or other factors, brought up any correlations as to why that might be.

If the trial failed and the drug is dead, then why write about it at all? This 18-month trial had redeeming features that inspire confidence that the field as a whole will be able to show efficacy and safety once better disease-modifying drugs roll around, Green said. “This is the largest Phase 3 trial in AD ever completed. It was a model case of a public-private partnership to test drugs as we are entering the era of disease-modifying drugs,” Green said.

The trial was designed and conducted well, and it proceeded smoothly as 133 U.S. centers followed the protocol without a troubling degree of site variation, Green said. The 1,649 participants were randomized into well-matched groups, the placebo group declined as expected, and the trial was powered sufficiently. Dropout rates were 33 percent for placebo and 39 percent for Flurizan, and the investigators followed the pre-specified analysis plan. “It’s important to acknowledge that this was an extremely successful trial in terms of its conduct. It showed that patients decline enough over 18 months to see a signal. That had been questioned before. This is very important—we can see the effect of a disease-modifying compound,” Green said.

It seems that the field assigns blame for this failure to Flurizan itself. This compound had shown effects in vitro and in vivo in mice (e.g., Eriksen et al., 2003; Kukar et al., 2007). However, not enough of it reached its intended target in the human brain. Other scientists noted that the human exposure of Flurizan never reached concentrations close to the observed IC50 in vitro. Therefore, it is reasonable to speculate that γ-secretase in the brain of the patients was never effectively modulated. Neither the Phase 2 nor the Phase 3 trial presented CSF data to show clearly how much of the drug enters the brain and that it alters Aβ levels there. In fact, a Phase 1 trial conducted by Douglas Galasko and colleagues at the University of California, San Diego, observed no significant reductions in plasma and CSF Aβ42 levels of healthy volunteers treated with the highest dose of Flurizan that was then used in the Phase 3 trial (Galasko et al., 2007). “In hindsight, it is fair to assume that Flurizan, given its poor potency and brain permeability, failed because of insufficient pharmacodynamics,” said Sascha Weggen from the Heinrich-Heine-University in Düsseldorf, Germany.

For this reason, scientists questioned whether this trial tested the amyloid hypothesis and the therapeutic concept of γ-secretase modulation at all. A number of other companies large and small are continuing research on their own, hopefully more potent, γ-secretase modulators. “It is important to point out that Flurizan was just a first-generation γ-secretase modulator. One could argue that its poor pharmacological properties and marginal efficacy in preclinical studies made it an ill-advised first try, but its good safety profile also made it an opportunity that needed to be explored. Now we need to develop improved compounds that efficiently modulate γ-secretase activity in brain of preclinical animal models and humans,” Weggen added. For his part, Green was philosophical. To reporters, he cited a quote by Woody Allen: “If you are not failing every now and then, it’s a sign you are not doing anything very innovative.”—Gabrielle Strobel.

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Wilcock GK, Black SE, Hendrix SB, Zavitz KH, Swabb EA, Laughlin MA, . Efficacy and safety of tarenflurbil in mild to moderate Alzheimer's disease: a randomised phase II trial. Lancet Neurol. 2008 Jun;7(6):483-93. PubMed.
Eriksen JL, Sagi SA, Smith TE, Weggen S, Das P, McLendon DC, Ozols VV, Jessing KW, Zavitz KH, Koo EH, Golde TE. NSAIDs and enantiomers of flurbiprofen target gamma-secretase and lower Abeta 42 in vivo. J Clin Invest. 2003 Aug;112(3):440-9. PubMed.
Kukar T, Prescott S, Eriksen JL, Holloway V, Murphy MP, Koo EH, Golde TE, Nicolle MM. Chronic administration of R-flurbiprofen attenuates learning impairments in transgenic amyloid precursor protein mice. BMC Neurosci. 2007;8:54. PubMed.
Galasko DR, Graff-Radford N, May S, Hendrix S, Cottrell BA, Sagi SA, Mather G, Laughlin M, Zavitz KH, Swabb E, Golde TE, Murphy MP, Koo EH. Safety, tolerability, pharmacokinetics, and Abeta levels after short-term administration of R-flurbiprofen in healthy elderly individuals. Alzheimer Dis Assoc Disord. 2007 Oct-Dec;21(4):292-9. PubMed.

Chicago: Translational and Basic Science of Tau Advances

21 Aug 2008

Beneath the fanfare of Rember^TM—the first tau-based Alzheimer disease treatment with publicly reported Phase 2 trial results (see ARF ICAD story)—the International Conference on Alzheimer’s Disease, held last month in Chicago, featured some noteworthy preclinical and basic science tau developments. This story highlights a few of them, but all readers are cordially invited to use the comment box below to add their own notes and observations about other tau-related highlights at ICAD. Einar Sigurdsson of New York University School of Medicine presented data that may strengthen the case for tau-based immunotherapy: in his new AD mouse model, a phospho-tau immunogen not only cleared tau pathology, but also seemed to prevent cognitive decline. Cathy Andorfer of Mayo Clinic in Jacksonville, Florida, also reported on prevention of cognitive decline, neurodegeneration, and tau pathology in an aggressive tauopathy mouse model—but her team used a brain-penetrant microtubule stabilizer under development as a cancer drug at Bristol-Myers Squibb, Wallingford, Connecticut. Meanwhile, molecular insight into a possible role for tau’s non-microtubule binding domain in Aβ-mediated neurodegeneration came from a presentation by Jürgen Götz of the University of Sydney, Australia. (For Andorfer and Götz, see Part 2 and Part 3.)

Preventive Tau Vaccine Improves Cognition in Tauopathy Mice
In an earlier study demonstrating the feasibility of a phospho-tau vaccine approach, Sigurdsson and colleagues immunized transgenic mice expressing the human tau mutant P301L, which develop rapid and aggressive tau pathology. With immunizations starting at two months of age, the tau peptide (a 30-amino-acid fragment phosphorylated at two sites) triggered a specific antibody response that cleared tau aggregates and improved motor performance when assessed at five and eight months (see ARF related news story). However, the real proof of principle is whether the tau-based immunotherapy could prevent cognitive decline. For this, the P301L mice are ill-suited, Sigurdsson said, as older animals develop motor impairments that would interfere with common tests of rodent cognition such as those that involve maze running.

To skirt this problem, the researchers set out to test their immunization protocol in hTau mice, which express all six wild-type isoforms of human tau on a mouse tau knockout background (Andorfer et al., 2003). These mice better resemble AD in that they develop tau pathology primarily in the cortex and hippocampus with very little in areas important for motor activity (e.g., brain stem, spinal cord). For the immunization studies, hTau mice would pose other challenges, though. The onset of disease varies considerably in them, such that some animals show extensive pathology by nine months, while others may not until 15-18 months. This slow development of pathology would also drag out the evaluation of immunotherapy.

While gearing up for the hTau studies, Allal Boutajangout of Sigurdsson’s group was trying to address a separate issue—the effect of mutant presenilin-1 (PS1) on tau pathology—by crossing the hTau mice with a transgenic line expressing mutant PS1 (M146L). These double transgenics were breeding faster than the hTau mice, Sigurdsson said, so Boutajangout and colleagues ended up testing their phospho-tau immunogen in the hTau/PS1 mice, using the hTau animals as internal controls. (They decided this before learning that mutant PS1 accelerates tau pathology, serendipitously enabling quicker completion of the immunotherapy studies.)

As with the studies in P301L mice, Sigurdsson’s team began immunizing at two to three months, boosted two weeks later, and continued with monthly injections. Starting at seven to eight months, the mice faced three cognitive tests and were sacrificed at eight to nine months. The phospho-tau peptide stimulated an IgM/IgG response and reduced tau aggregates by 56 percent in the pyriform cortex in immunized mice, Sigurdsson reported. Importantly, the reduction of tau aggregates correlated with cognitive improvements assessed by three tests—radial arm maze, closed field symmetrical maze, and object recognition test. The cognitive differences did not stem from movement defects, as all groups did equally well on motor tests. The researchers have not yet looked at markers of inflammation¬—a potential side effect that has plagued human immunotherapy trials—but would like to in the future, Sigurdsson told this reporter.

Because hTau/PS1 mice typically develop their first hints of tau pathology around two to three months of age, the vaccine in those studies essentially functioned as a prophylactic. Sigurdsson said his group has begun testing their immunotherapy protocol in older animals—starting injections at eight months to see if the treatment can reverse tau pathology and cognitive deficits. They are also looking to immunize with different tau epitopes, and are doing studies to further tease out the mechanisms, e.g., how do the anti-tau antibodies reach and bind to brain neurons, and clear tau aggregates? For a different approach to treating tauopathy mice, see Part 2 of this series.—Esther Landhuis.

This is Part 1 of a three-part series on tau. See also Part 2 and Part 3.

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Chicago: Brain-Penetrant Microtubule Stabilizer in Tauopathy Mice

22 Aug 2008

Another preclinical study presented at the International Conference on Alzheimer’s Disease, held 26-31 July in Chicago, capitalized on tau’s microtubule-binding property. This feature has inspired the notion that pathologic forms of tau—such as those that develop into the hallmark AD tangles—destabilize microtubules and thereby cause neurodegeneration. Consistent with this idea, previous work (Michaelis et al., 2004; Zhang et al., 2005) has demonstrated that microtubule-stabilizing agents such as the cancer drug paclitaxel (aka Taxol) can fend off Aβ-mediated neurodegeneration. The Zhang et al. study, led by John Trojanowski at University of Pennsylvania School of Medicine, Philadelphia, also showed that very low doses of paclitaxel could reverse motor deficits characteristic of PrPT44 mice, which overexpress the shortest human tau isoform in the central nervous system.

Cathy Andorfer and her collaborators at Mayo Clinic, Jacksonville, Florida, and Bristol-Myers Squibb, Wallingford, Connecticut, were intrigued by this finding. As were Sigurdsson and colleagues (see Part 1 of this series), these investigators worried that the motor impairments—deriving from predominantly spinal cord expression of transgenic tau in PrPT44 mice—would confound cognitive tests needed to show drug efficacy. Furthermore, taxols penetrate the brain poorly and thus seemed unlikely to reach the clinic as an AD treatment. Researchers at Bristol-Myers Squibb had identified an epothilone (another microtubule-binding cancer drug) that efficiently crosses the blood-brain barrier, and wanted to test this compound in a tauopathy mouse model.

They and Andorfer chose rTg4510 mice, a well-characterized transgenic line that expresses a tetracycline-inducible tau mutant (P301L) primarily in the cortex and hippocampus, and hence lacks a motor phenotype. These mice accumulate abnormal tau in AD-relevant areas as early as 1.5 to two months of age, with mature tangles and measurable cognitive defects by four to five months and gross brain shrinkage by 10 months (see ARF related news story). Andorfer and colleagues initially tested the mice at 2.5 months of age using the Morris water maze to establish baseline memory function, and thereafter began weekly injections of either vehicle or one of two epothilone doses (each 10-100-fold below the dose used in oncology). At 4.5 months, the mice were retested in the water maze and sacrificed for histopathologic analysis a month later.

Mice treated with the lower (1 mg/kg) epothilone dose were able to learn the location of the submerged platform—a spatial memory task that stymied the vehicle-treated and high-dose (10 mg/kg) groups. This treatment effect held up in post-training probe trials. The cognitive benefits were associated with a significant reduction in the hippocampal CA1 neuronal loss typical of rTg4510 mice. Low-dose epothilone treatment also noticeably affected tau pathology, reducing hyperphosphorylated and conformationally abnormal tau, as well as Gallyas silver-positive neurofibrillary tangles. The latter results in particular came as a surprise, Andorfer said, because no significant pathological changes were seen in the earlier paclitaxel study (Zhang et al., 2005). Coupled with the absence of obvious adverse consequences such as weight loss or motor dysfunction, these findings support the hypothesis that tau dysfunction drives microtubule dynamics through a destabilization effect that epothilones and related drugs appear to counter. Bristol-Myers Squibb collaborator Charlie Albright said that further mouse studies are underway to better understand the drug’s mechanism, which will, in turn, provide new insight into how abnormal tau wreaks havoc in the brain in AD and related diseases. For more on that, see Part 3 of this series.—Esther Landhuis.

This is Part 2 of a three-part series on tau. See also Part 1 and Part 3.

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Michaelis ML, Ansar S, Chen Y, Reiff ER, Seyb KI, Himes RH, Audus KL, Georg GI. {beta}-Amyloid-induced neurodegeneration and protection by structurally diverse microtubule-stabilizing agents. J Pharmacol Exp Ther. 2005 Feb;312(2):659-68. PubMed.
Zhang B, Maiti A, Shively S, Lakhani F, McDonald-Jones G, Bruce J, Lee EB, Xie SX, Joyce S, Li C, Toleikis PM, Lee VM, Trojanowski JQ. Microtubule-binding drugs offset tau sequestration by stabilizing microtubules and reversing fast axonal transport deficits in a tauopathy model. Proc Natl Acad Sci U S A. 2005 Jan 4;102(1):227-31. PubMed.

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Chicago: Tau Projection Domain May Block Excitotoxicity in Mice

25 Aug 2008

The development of tau-based drugs for Alzheimer disease and other tauopathies depends on a deeper understanding of how tau begins to run amok in the first place. Molecular details for how this might occur in AD have emerged from mouse studies that have begun to zero in on how specific domains of the tau protein contribute to Aβ-induced neurodegeneration. Questions about which forms of tau help or harm neurons have long hampered the development of tau-based therapeutics. A recent study (Roberson et al., 2007) laid aside such concerns and showed, astonishingly, that AD transgenic mice could be protected from Aβ-induced cognitive decline and premature lethality simply by halving their levels of tau protein. By crossing tau knockout mice with human APP-overexpressing transgenics, Lennart Mucke’s group at the University of California, San Francisco, showed that lowering tau not only helped neurons resist the toxic effects of Aβ but also seemed to protect the brain from other excitotoxic insults (see ARF related news story).

Jürgen Götz and colleagues at the University of Sydney, Australia, wondered which regions of the tau protein mediated those powerful effects. To tackle this question, Lars Ittner in the group took tau’s “projection” domain (aka Δtau)—a 255-amino acid N-terminal segment that lacks the domain associated with microtubule binding and pathological aggregation—and expressed it in mice that express mutant human APP and lack full-length tau. These animals were generated by crossing APP23 mice with tau knockouts. At the International Conference on Alzheimer’s Disease in Chicago last month, Götz reported that expression of the Δtau fragment seemed to do just as well as the sweeping tau reductions at protecting mice from early death induced by APP overexpression. By 10 months of age, nearly half of APP23 animals die, but those expressing either Δtau or hemizygous levels of full-length tau enjoyed an 80-90 percent survival rate. APP23 mice with both tau manipulations got a complete rescue, with 100 percent surviving at 10 months. On a non-APP overexpressing background, Götz reported, both Δtau expression and tau reduction (tau-/-) reduced the mean severity of pentylenetetrazole-induced seizures by 30-40 percent compared to controls with normal APP and tau. In APP23 mice, each tau gene alteration increased latency for more severe seizures by about twofold. These findings extend another recent report from the Mucke lab (Palop et al., 2007) and tap into growing interest in abnormal network activity in AD.

How might the Δtau fragment protect against excitotoxicity? Götz reminded the audience that excitotoxicity is mediated by formation of a complex between an NMDA receptor (NR) and post-synaptic density protein PSD-95. This complex is stabilized by phosphorylation of the NR subunit NR2B by the kinase Fyn. The researchers examined these interactions and found that NMDA receptor surface expression dropped by about 75 percent in tau-/- mice relative to wild-type. This was associated with reduced assembly of the NR/PSD-95 complex, as neurons from both tau knockout and Δtau transgenic mice had no NMDA receptor subunit 1 (NR1) co-immunoprecipitating with PSD-95. Furthermore, both tau-/- and Δtau-expressing mice had lower levels of phosphorylated NR2B, which typically stabilizes the NR/PSD-95 interaction.

To see whether Fyn abnormalities might be responsible for the NR/PSD-95 defects, the researchers looked at Fyn-tau interactions and found less Fyn interacting with endogenous tau in Δtau transgenic mice. In both tau-/- and Δtau-expressing mice, Fyn was found at much higher levels in cell bodies than dendrites of CA1 neurons and at lower levels in synaptosomal preparations from hippocampi of these mice. Considering that Δtau is targeted to axons and excluded from dendrites in the Δtau mice, these findings suggest that Δtau traps Fyn in cell bodies, Götz said. This would prevent Fyn from phosphorylating NR2B and thereby destabilize the NR/PSD-95 interaction that mediates excitotoxicity. Consistent with this proposed model, Götz reported in Chicago that the peptide PSD-95 inhibitor Tat-NR2B9c was able to reduce Aβ toxicity in primary neuronal cultures in a dose-dependent fashion.—Esther Landhuis.

This concludes our three-part series on tau news from ICAD. See Part 1 and Part 2.

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Roberson ED, Scearce-Levie K, Palop JJ, Yan F, Cheng IH, Wu T, Gerstein H, Yu GQ, Mucke L. Reducing endogenous tau ameliorates amyloid beta-induced deficits in an Alzheimer's disease mouse model. Science. 2007 May 4;316(5825):750-4. PubMed.
Palop JJ, Chin J, Roberson ED, Wang J, Thwin MT, Bien-Ly N, Yoo J, Ho KO, Yu GQ, Kreitzer A, Finkbeiner S, Noebels JL, Mucke L. Aberrant excitatory neuronal activity and compensatory remodeling of inhibitory hippocampal circuits in mouse models of Alzheimer's disease. Neuron. 2007 Sep 6;55(5):697-711. PubMed.

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Chicago: Does Saying “I Do” Lower Late-life Dementia Risk?

26 Aug 2008

The newest potential protective factor for Alzheimer’s might be wrapped around your ring finger. So suggests a prospective, population-based study presented last month at the International Conference on Alzheimer’s Disease (ICAD) in Chicago. Having analyzed data on more than 1,400 people from eastern Finland, Krister Håkansson of Växjö University and Karolinska Institutet, Växjö, and Stockholm, Sweden, reported that those who were married or living with a significant other in mid-life had a 50 percent lower risk of developing late-life dementia compared to those living alone. Taking a closer look at subgroups who maintained their solo-dwelling status over at least two decades, Håkansson found that singles and divorcées had a two- to threefold higher risk of dementia relative to the marrieds, whereas widows were more than six times as likely to develop AD. Among widows carrying the AD genetic risk factor ApoE4, the frequency of AD cases shot up to roughly 14-fold relative to non-ApoE4 carriers living together. In providing hard numbers to suggest that getting hitched and staying together has long-term brain benefits, this study supports the general hypothesis that social engagement may help people compensate for neurodegeneration seen with AD and related diseases.

Twin studies have placed AD heritability estimates between 60 and 80 percent, with the remaining variance stemming from environmental influences (Gatz et al., 2006). Among such non-genetic risk factors is an active social life (see ARF related news story). This generally means higher participation in activities that involve physical or mental exercise, factors that protect against AD in people (Teri et al., 2003) and mice (see ARF related news story). In light of the broader claim that intellectual and social enrichment helps guard against age-related dementia, Håkansson had a hunch that these protective effects could be more specifically linked to mid-life marital status. “It’s hard to imagine any form of social and intellectual stimulation more intensive than couple relations,” he said.

The 1,449 participants in Håkansson’s analysis came from the population-based Cardiovascular Risk Factors, Aging, and Dementia (CAIDE) study, which assessed middle-aged individuals between 1972 and 1987, and again about 21 years later, for signs of dementia. Owing to an unusually high number of male deaths from Finland’s world-leading cardiovascular mortality rate and its war with Russia during World War II, the sample included many early widows, most of whom (105/111) were still unmarried decades later.

At re-examination in 1998, 143 participants in Håkansson’s study were diagnosed with some form of cognitive impairment—among those, 82 with mild cognitive impairment (MCI) and 48 with AD. Compared to those living with a spouse or significant other at mid-life, singles had a doubled risk of late-life dementia. If those singles continued to live without a partner through follow-up 21 years later, their rate of developing cognitive impairment went up to nearly three times that of couples living together, Håkansson said. The study adjusted for other mid-life factors, including education, body mass index, cholesterol, blood pressure, occupation, physical activity, smoking habits, depression, ApoE status, gender, and age at follow-up.

A closer look at the circumstances behind singlehood yielded some intriguing findings as well. If living without a partner were regarded as a risk factor for late-life dementia, one might presume that all-life singles would be worse off than those who had been previously married. Yet Håkansson found the opposite. Middle-aged widows who remained alone into late-life were almost three times as likely as other singles—and more than six times as likely as people living with spouses or significant others—to develop Alzheimer disease. AD prevalence among widows carrying the ApoE4 allele was about 14 times higher than that of non-ApoE4 carriers living together. To Håkansson, these findings fit the socio-genetic disease model. He likened singlehood induced by losing one’s partner early in life to a traumatic event that, if sustained, seems to add to the AD risk conferred by the ApoE4 gene.—Esther Landhuis.

References

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Paper Citations

Gatz M, Reynolds CA, Fratiglioni L, Johansson B, Mortimer JA, Berg S, Fiske A, Pedersen NL. Role of genes and environments for explaining Alzheimer disease. Arch Gen Psychiatry. 2006 Feb;63(2):168-74. PubMed.
Teri L, Gibbons LE, McCurry SM, Logsdon RG, Buchner DM, Barlow WE, Kukull WA, LaCroix AZ, McCormick W, Larson EB. Exercise plus behavioral management in patients with Alzheimer disease: a randomized controlled trial. JAMA. 2003 Oct 15;290(15):2015-22. PubMed.

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Chicago: More News From Phase 2s

27 Aug 2008

At the International Conference for Alzheimer’s Disease, held 26-31 July in Chicago, several investigators presented data on compounds in Phase 2. Below is a summary of an approach targeting a neurotransmitter receptor, two approaches targeting Aβ, and one testing a nutrient supplement. In addition to these, Phase 2 data of AL-108, a neuroprotective compound administered through the nose, presented data that Alzforum had summarized recently after a similar conference presentation in Boston (see ARF related news story).

Boosting Neurotransmission: SB-742457
SB-742457 by GlaxoSmithKline is one of a range of small-molecule drugs that aim to right neurochemical disturbances in AD. The cholinergic and also the serotonergic neurotransmitter systems, among others, are impaired as Alzheimer’s develops and progresses, and they present opportunities for renewed efforts at propping up neurotransmission. These drugs are not billed as targeting primarily what many consider to be causal AD pathways of Aβ amyloidosis and tauopathy. Instead they are more modestly presented as improving symptoms of memory loss and behavior in AD and other forms of dementia, perhaps as part of future combination drug regimens.

SB-742457 is a new antagonist of the 5-HT receptor 6, a largely CNS-specific type of serotonine receptor subfamily (Upton et al., 2008). At ICAD, GSK researchers presented clinical data on two Phase 2 trials, as well as preclinical data on cognition models. Gareth Maher-Edwards of GSK described a dose-finding trial in 371 people with mild to moderate AD who received one of three doses of this antagonist or placebo for 24 weeks, and then had its efficacy assessed by means of the CIBIC+ scale to look for global function and the ADAS-Cog to look for cognitive change. Dropout rates ranged from 13 to 20 percent depending on the dose. Maher-Edwards reported a statistically significant improvement over baseline in CIBIC+ at the highest dose, also a trend without statistical significance for a slight improvement in ADAS-Cog.

A second, smaller trial of 197 patients compared this highest dose of 35 mg/day dose of the serotonin receptor antagonist to 5 mg/day of donepezil, a widely prescribed cholinesterase inhibitor that is standard therapy in AD, and placebo. Using the same outcome measures, SB-742457 showed small treatment effects of roughly similar size to donepezil. The placebo groups did not decline by six months in either of these two trials, so the small reported drug effects are essentially improvements over baseline consistent with a symptomatic effect. People who started the trials with lower MMSE values, i.e., the more impaired patients, appeared to respond a little better, and the drug appeared to be safe and well tolerated, Maher-Edwards said. He added that four deaths in the study were due to traffic accidents, cancer, and a heart attack but unrelated to the study drug.

Rather than proceed to Phase 3, GSK is continuing to explore this drug in Phase 2 with two more six-month trials. An 89-center, 672-patient trial testing a single dose of SB-742457 in some 670 patients in addition to donepezil is recruiting at present, and a slightly smaller trial comparing SB-742457 to donepezil is set to get going later this year. Besides the usual cognitive, global, and safety measures, these trials will look for pharmacogenomic effects, but they have no fluid or imaging biomarker components.

Other neurochemical targets under active study include nicotinic and muscarinic acetylcholine receptors, as well as 5-HT4 receptors (for coverage of active clinical programs on the latter two, see ARF recent Keystone story) and the serotonine 1A receptors.—Gabrielle Strobel.

This is Part 1 of a three-part series. See also Part 2

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Chicago: More Phase 2 News—PBT2 and IVIg

28 Aug 2008

Besides Elan/Wyeth’s and Eli Lilly and Company’s antibodies, two additional anti-amyloid approaches—PBT2 and IVIg—came out with Phase 2 data at ICAD. Jeffrey Cummings of the University of California, Los Angeles, presented results on PBT2. The Alzforum has reported this data this past March, after the drug’s developer, Australian Prana Biotechnology Ltd., first announced results (see ARF related drug news story). The full publication appears in the September Lancet Neurology, now available online (Lannfelt et al., 2008).

At ICAD, some scientists cautioned that the drug’s reported signals in two cognitive tests, i.e., trail making B and category fluency, should not be overinterpreted, as only 29 patients had received the higher, presumably effective, dose. For his part, Cummings said that he saw the main significance of this trial in the drug’s demonstrable ability to lower human CSF Aβ42 levels. This has been difficult to achieve with previous therapeutic agents, including, most recently, Flurizan. Cummings also noted that the trial investigators monitored very closely for any hints of subacute myelo-optic neuropathy, a serious side effect associated with clioquinol in the past, but found no indications of this with PBT2. “We feel the safety and tolerability of PBT2 is comparable to placebo,” Cummings said. Prana at present is looking for funding to support a planned Phase 2b trial.

Norman Relkin of Weill Medical College, Cornell University, New York, presented interim nine-month data from an ongoing 18-month Phase 2 trial of IVIg (aka Gammagard), a preparation of pooled human antibodies made by Baxter Biosciences. While more specific vaccines are being developed, this product drew Relkin’s attention because it has a decades-long safety record in humans for use in other conditions. Relkin and colleagues had previously conducted an open-label, Phase 1 dose-ranging study in people with AD, and reported a symptomatic benefit (Relkin et al., 2008; also ARF related news story). One question that arose in this earlier study was how long this benefit could last. The AD patients in this study had a break in treatment after six months, after which they lost the improvement they had gained during the previous six months on IVIg. Relkin and colleagues address whether this decline was due to the washout of IVIg, or whether this preparation simply does not help longer than six months, in a subsequent placebo-controlled, randomized Phase 2 trial at Weill that is still ongoing. Both trials are small—eight patients in the open-label study, 24 patients in the Phase 2—owing in part to the drug’s cost and availability (see below).

In the Phase 2 trial, four patients each received one of four doses of IVIg for 24 weeks, and eight patients received infusions of saline placebo. Evaluations occurred at 12 weeks and 24 weeks; after that the patients on placebo switched over to one of four doses of IVIg for another 12 months. From this point on, then, the trial had no true placebo group anymore, and the former placebo group was partially blinded in that the rater did not know what dose they were on. The primary outcome measures were the ADAS-Cog, ADCS-CGIC; secondary outcome measures included the ADL and some other cognitive and global measures. (This trial also incorporated CSF biomarker measures, as well as imaging scans using FDG PET to assess brain activity, PIB-PET to measure amyloid levels, and PK11195-PET to measure microglial activation in a subgroup of patients. Those data are not available yet, Relkin said.)

At ICAD, Relkin presented nine-month interim data on the primary and secondary readouts. On all outcomes, the curves between placebo and IVIg separated on the three-, six-, and nine-month time points. The results presented at a press briefing and on a poster were confusing at first glance because the slope graphs indicated a nine-month time point for the placebo group even though those people at this point had been on various doses of drug for the past three months. In an e-mail, Relkin clarified that the main point to take away from this interim analysis is that the symptomatic benefit that was apparent in the Phase 1 study stayed stable for nine months in this second study. “This Phase 2 study was the first time we treated AD patients with IVIg for longer than six months without interruption. The interim analysis presented at ICAD was the first demonstration of the persistence of benefits at nine months when IVIg treatment was uninterrupted,” Relkin wrote. Relkin clearly stated that this small trial was not powered to measure efficacy; hence, numerical comparisons between the groups are less meaningful than the general observation that treated patients maintained their benefit for nine months so far.

The trial confirmed the best dose as being 0.4g/kg infused every two weeks, and this information will help in the conduct of a subsequent Phase 3 trial. Starting this fall, the Alzheimer’s Disease Cooperative Study will enroll 360 patients at some 35 sites into an 18-month trial that incorporates CSF, MRI, FDG PET and PIB-PET. This trial will be pivotal for the prospects of this treatment in AD.

“The NIA-sponsored ADCS is the most appropriate organization to test IVIg as a potential treatment for AD, since IVIg is not patentable and consequently of little interest to the pharmaceutical industry. The partnership between the ADCS, with its expertise in AD clinical trials, and Baxter has made it possible to design a much more comprehensive and exciting Phase 3 trial than would otherwise have been possible,” Relkin said. If the trial were to be positive, the issue of IVIg’s limited supply would come to the fore. At present, there is a relative shortage of this product, which is made from the blood of several thousand donors. According to Relkin, its sole manufacturer Baxter Bioscience sells all it can prepare within a few weeks for other indications. Scale-up to satisfy demand for a potential new AD indication would require a major investment in manufacturing, and issues related to this have slowed down the planning for the ADCS Phase 3 trial, said the ADCS director Paul Aisen of the University of California, San Diego.

Cost is another issue. A small number of families are working with neurologists besides Relkin to purchase IVIg and treat AD in a loved one outside of formal trials. With supply so limited, this can be expensive. Here is how Relkin explained the issue: “The cost of IVIg is dependent on market factors. There are often considerable mark-ups when it is purchased outside of a hospital setting. Minimum base cost when purchased in bulk by hospitals is about $50 per gram, and we typically give between about 20-30 grams per infusion depending on the subject’s weight. That translates into about $36,000 per year, but with mark-ups that can easily rise to $50,000-$75,000 per year or in extreme cases as high as $100,000 per year. Since supply is limited, for now the high cost helps to discourage premature off-label use that might otherwise deplete the supply available to treat other diseases.”

“If approved, IVIg supply could become a major issue. We are working on strategies to increase the efficiency and ease of delivery, as well as ways to identify best responders predictively (FDG PET looks promising for this purpose). The IVIg manufacturers are now considering plans to increase production. If successful, these initiatives could help extend the available supply to more patients and/or reduce overall expenditures,” Relkin wrote by e-mail.—Gabrielle Strobel.

This is Part 2 of a three-part series. See also Part 1.

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References

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Lannfelt L, Blennow K, Zetterberg H, Batsman S, Ames D, Harrison J, Masters CL, Targum S, Bush AI, Murdoch R, Wilson J, Ritchie CW, . Safety, efficacy, and biomarker findings of PBT2 in targeting Abeta as a modifying therapy for Alzheimer's disease: a phase IIa, double-blind, randomised, placebo-controlled trial. Lancet Neurol. 2008 Sep;7(9):779-86. PubMed.
Relkin NR, Szabo P, Adamiak B, Burgut T, Monthe C, Lent RW, Younkin S, Younkin L, Schiff R, Weksler ME. 18-Month study of intravenous immunoglobulin for treatment of mild Alzheimer disease. Neurobiol Aging. 2009 Nov;30(11):1728-36. PubMed.

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Chicago: Phase 2 News—Therapeutic Breakfast Food?

29 Aug 2008

While the glare of the media focuses on the trials and tribulations of the next potential billion-dollar pharmaceutical treatment of AD, other investigators have been quietly cooking up alternative approaches. One of those was on the menu at the International Conference for Alzheimer’s Disease (ICAD), held 26-31 July in Chicago. Philip Scheltens of Vrije University Medical Center in Amsterdam, the Netherlands, is well established in the AD field for his extensive brain imaging research in Alzheimer disease and related dementias. At ICAD, however, Scheltens presented data for a proof-of-concept human trial of a multi-nutrient drink. The drink is a product of Numico Research in Wageningen, The Netherlands, which in July 2007 was bought by Danone (Dannon to Americans). The drink is a defined mixture of ingredients said to be needed for membrane synthesis and synapse formation. According to Scheltens, it yielded a promising signal in a controlled 12-week trial of 212 people with early AD.

The idea behind this approach is that the synapse loss that characterizes AD might slow down if the ailing brain had an ample supply of synaptic membrane constituents, such as phosphatides, more readily available. These membrane phosphatides, in turn, are synthesized from molecular ingredients that were shown in previous research to influence learning and memory in various animal models. Much of the prior research over the past decade that has led up to the present trial comes from the laboratory of Richard Wurtman at the Massachusetts Institute for Technology (see, for example, Cansev et al., 2008; Wurtman et al., 2006; Wang et al., 2005; Holguin et al., 2008).

Scheltens and colleagues tested a drink formulated to contain ingredients in six functional categories previously studied by Wurtman’s and other groups: uridine-5’-monophosphate (UMP), omega-3 fatty acids (see also below for update on DHA trial), choline, phospholipids, B vitamins, and antioxidants. This mixture is trade-named Fortasyn^TM Connect, and the drink goes by Souvenaid^TM.

Conducted at 28 sites in The Netherlands, Belgium, Germany, and the U.S., the trial was randomized, double-blind, and controlled with a second drink that tasted the same as Souvenaid. “It tastes good; people liked it,” Scheltens said. One hundred six patients who were recently diagnosed with probable AD as per NINCDS-ADRDA criteria drank Souvenaid, and 106 placebo, once a day for 12 weeks. After that, they could choose to continue the blinded study; 160 patients finished the full 24 weeks. At ICAD, Scheltens presented the 12-week data. Primary outcome measures were a delayed verbal memory task (WMS-r) and ADAS-Cog-13, an extended ADAS-Cog battery. Secondary outcomes included functional, quality of life and behavioral measures, as well as plasma levels of nutrients and homocysteine, and safety measures. This initial trial included no CSF measures to show brain penetration and exposure data, nor brain imaging. The patients in this trial did not take acetylcholinesterase inhibitors or Namenda. The trial is not on clinicaltrials.gov but was listed with the Dutch Trial Register.

The groups were well matched, Scheltens reported, and Souvenaid produced no worrisome safety signals. A number of side effects were more common in the Souvenaid group, though none of the differences reached statistical significance. Some 18 percent in both control and Souvenaid groups had diarrhea, constipation, or nausea. Upwards of 92 percent of patients complied with the regimen. In plasma at 12 weeks, Souvenaid increased the concentration of the omega-3 fatty acid DHA (see more below), and reduced that of homocysteine, a metabolite thought to hasten cognitive decline.

Also at 12 weeks, Souvenaid improved performance of the WMS-r memory test. This effect was statistically significant, and the effect size was larger in a pre-specified subgroup of people who had an MMSE of greater than 23 at study onset, i.e., very mild AD, than in the more impaired group. The ADAS-Cog curves overlapped, showing no effect in the treatment group and no decline in the control group. A pre-specified analysis looking at baseline MMSE value as a predictor showed a greater numerical effect in the milder cases, but this did not reach statistical significance.

Analysis of the secondary outcomes as well as of the 24-week data is still to come, Scheltens said. “We know Souvenaid has a quick effect,” he added. “Whether it lasts past 12 weeks we do not know yet.” For now, he emphasized that Souvenaid seems to be safe and well tolerated, and has shown a signal of efficacy that serves as a proof of concept to support further clinical study of this product.

Fortasyn^TM is not to be mixed up with Ketasyn^TM, another breakfast shake drink meant to shore up starving neurons in AD, but containing different ingredients (see ARF related conference story).

Also at ICAD, there were tidbits of news on one component of the Souvenaid mixture, i.e., the polyunsaturated omega-3 fatty acids. One of those is docosahexaenoic acid (DHA), which in its own right has a growing body of epidemiological and experimental evidence suggesting it may protect against age-related cognitive decline (see, for example, Calon et al., 2004; Freund-Levy et al., 2006; Freund-Levy et al., 2007). Several presentations from different groups converged around the theme that DHA, and perhaps other nutrient-based approaches, show the most robust efficacy signal early on, that is at the MCI or very early AD stage, but help little by the time a person has moderate AD.

On DHA, the Alzheimer’s Disease Cooperative Study reported that enrollment of its DHA trial has been completed in 10 months. This trial tests a recombinant version of DHA generated in microscopic algae by Martek Biosciences, which is widely used as an ingredient of infant formula. Previously, this form of DHA had been shown to reduce accumulation of both amyloid and tau pathology in triple-transgenic mice, among other models (see Green et al., 2007).—Gabrielle Strobel.

This concludes a three-part series. See also Part 1 and Part 2.

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References

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Cansev M, Wurtman RJ, Sakamoto T, Ulus IH. Oral administration of circulating precursors for membrane phosphatides can promote the synthesis of new brain synapses. Alzheimers Dement. 2008 Jan;4(1 Suppl 1):S153-68. PubMed.
Wurtman RJ, Ulus IH, Cansev M, Watkins CJ, Wang L, Marzloff G. Synaptic proteins and phospholipids are increased in gerbil brain by administering uridine plus docosahexaenoic acid orally. Brain Res. 2006 May 9;1088(1):83-92. PubMed.
Wang L, Pooler AM, Albrecht MA, Wurtman RJ. Dietary uridine-5'-monophosphate supplementation increases potassium-evoked dopamine release and promotes neurite outgrowth in aged rats. J Mol Neurosci. 2005;27(1):137-45. PubMed.
Holguin S, Martinez J, Chow C, Wurtman R. Dietary uridine enhances the improvement in learning and memory produced by administering DHA to gerbils. FASEB J. 2008 Nov;22(11):3938-46. PubMed.
Calon F, Lim GP, Yang F, Morihara T, Teter B, Ubeda O, Rostaing P, Triller A, Salem N, Ashe KH, Frautschy SA, Cole GM. Docosahexaenoic acid protects from dendritic pathology in an Alzheimer's disease mouse model. Neuron. 2004 Sep 2;43(5):633-45. PubMed.
Freund-Levi Y, Eriksdotter-Jönhagen M, Cederholm T, Basun H, Faxén-Irving G, Garlind A, Vedin I, Vessby B, Wahlund LO, Palmblad J. Omega-3 fatty acid treatment in 174 patients with mild to moderate Alzheimer disease: OmegAD study: a randomized double-blind trial. Arch Neurol. 2006 Oct;63(10):1402-8. PubMed.
Freund-Levi Y, Basun H, Cederholm T, Faxén-Irving G, Garlind A, Grut M, Vedin I, Palmblad J, Wahlund LO, Eriksdotter-Jönhagen M. Omega-3 supplementation in mild to moderate Alzheimer's disease: effects on neuropsychiatric symptoms. Int J Geriatr Psychiatry. 2008 Feb;23(2):161-9. PubMed.
Green KN, Martinez-Coria H, Khashwji H, Hall EB, Yurko-Mauro KA, Ellis L, LaFerla FM. Dietary docosahexaenoic acid and docosapentaenoic acid ameliorate amyloid-beta and tau pathology via a mechanism involving presenilin 1 levels. J Neurosci. 2007 Apr 18;27(16):4385-95. PubMed.

Chicago: Studies Probe Diminishing Placebo Decline, Part 1

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Chicago: Studies Probe Diminishing Placebo Decline, Part 2

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Chicago: ICAD Offered Mix Bag of Setbacks and Hopeful News

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Chicago: Trial Design Bedevils Search for New AD Drugs, Part 1

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Chicago: The Phase 2 Problem and the Way to a Surrogate, Part 2

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Chicago: Out of the Blue—A Tau-based Treatment for AD?

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Chicago: Dimebon Safe for 18 Months

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Chicago: Bapineuzumab’s Phase 2—Was the Data Better Than the Spin?

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Chicago: Lilly’s Antibody Appears to Do No Harm, But Will It Help?

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