NSAIDs vs. Alzheimer's: Multiple modes of action?

Loads of interesting stuff is going on in Alzheimer's research right now. While the hot news is about a trial showing significant benefits from going after tau tangles, a recent paper in PLoS ONE continues to investigate the pathology of the amyloid-β peptide. As I've mentioned in previous posts, cleavage of the amyloid precursor protein to a 42-residue peptide (called Aβ1-42 in this paper) initiates the formation of peptide oligomers and eventually plaques. Recent research has indicated that these oligomers are sufficient to cause the development of Alzheimer's disease, but the mechanism by which they do so remains uncertain. Sara Sanz-Blasco and colleagues show that Aβ oligomers disrupt calcium homeostasis in neurons, damaging the mitochondria and promoting apoptosis, and that certain NSAIDs can suppress these adverse mitochondrial effects (1). PLoS ONE is open access, so go ahead, open the article up in another window, and follow along.

Although the appearance of plaques and neuronal death are classic hallmarks of Alzheimer's pathology, the relationship between these features is not well understood. For instance, it is possible the plaques themselves kill neurons or impair neural function. However, it seems equally likely that the appearance of plaques and the death of neurons are two distinct effects with a single cause. This view is supported by the oligomer toxicity study, but that study fails to resolve the question of exactly how Aβ oligomers kill neurons. Previous work has associated Aβ with derangement of cellular calcium (Ca²⁺) management — a 2005 paper by Demuro et al. (2) showed that soluble Aβ induced an increase in intracellular Ca²⁺ in a neuroblastoma cell line. Sanz-Blasco et al. therefore decided to directly test whether Aβ oligomers were increasing Ca²⁺ levels in neurons, and specifically in mitochondria. In order to do this last bit they used a low-affinity aequorin targeted specifically to mitochondria.

Allow me digress... to many of my readers that probably sounds like a terrible idea. If you're trying to detect a particular chemical in the cell, it seems like the best thing to do would be to get a high-affinity binding partner. And if figuring out whether there is any calcium in the mitochondria is what you want to do, then a high-affinity detector makes sense. However, when you're using a small amount of a sensor to detect changes in the concentration of a large amount of ligand, a low-affinity sensor is what you want.

To see why, take a look at the graph on the right. This is just a rough calculation based on a situation where the detector is at a concentration of 100 µM and the concentration of its ligand (that you're trying to detect) changes from 10 mM to 100 mM. Note that the concentration of the detector is at most 1% that of the ligand. If the dissociation constant K_D of this complex is 1 mM (blue) (a lower K_D means higher affinity), then the detector is almost saturated when you start, and the percentage occupied doesn't change very much over the course of the experiment. This means that it will be very difficult to tell the difference between, say, 50 mM ligand and 100 mM ligand, because that amounts to a signal difference of 1% of the maximum response. The situation gets a little better if the K_D is 10 mM (green). The lowest affinity detector here (K_D = 50 mM, red) actually does the best job of distinguishing between 50 mM and 100 mM ligand, because the difference in response amounts to 17% of the total dynamic range. Ideally, you want to tune the K_D of your detector in such a way that its response to changes in ligand concentration is large and linear over the range you are likely to be observing. For the last detector, this range lies between 10 and 40 mM of ligand, so that would likely be the ideal range to investigate with it.

The precise numbers are different in the present paper, but the principle is the same. The affinity you want in your detector will depend on what you are trying to detect and the circumstances under which you are trying to detect it. In this case, the researchers are trying to measure changes in calcium ions over a fairly wide range, which have a pretty high concentration in mitochondria, and they're doing it using a luminescent protein, which isn't very concentrated. As a result, a relatively low-affinity detection system is best.

So, what did they find? The results in Figure 1 show that Aβ oligomers and fragments cause an influx of calcium into the cytoplasm of cultured neurons, but preparations of Aβ fibrils did not cause this effect. Moreover, exposure of the cells to Aβ oligomers caused a clear influx of calcium into the mitochondria (Figure 3). This is a problem for a cell because Ca²⁺ overload in mitochondria can cause programmed cell death, or apoptosis. Using the classic TUNEL assay, the authors of this study showed that the Aβ oligomers caused apoptosis. In addition, they showed that treatment with the oligomers caused the release of mitochondrial cytochrome c (a step in the apoptotic pathway) and that the addition of cyclosporin A, which inhibits the release of proteins from the mitochondrion, blocked cell death (Figure 4). Together, these pieces of evidence support the idea that Aβ-induced Ca²⁺ influx into the mitochondria activates the apoptotic cascade, leading to neuronal death. These results are consistent with a very cool study published this week in Neuron (3) showing that amyloid plaques correlated with high neuronal Ca²⁺ levels in vivo (in live mice).

On its own this is pretty interesting, but Sanz-Blasco et al. push it a bit further. Because they had shown previously that some NSAIDs prevent mitochondrial Ca²⁺ uptake in a cancer cell line, they decided to find out if they would work in this instance, too. As you can see from Figure 6, the three NSAIDs tested kept the mitochondria calcium-free, even if the cells were treated with Aβ oligomers. NSAIDs also prevented cytochrome c release and cell death (Figure 8).

Some readers may recall that Kukar et al. showed that certain NSAIDs prevent oligomerization of Aβ1-42, hinting at a possible explanation of these results. However, the controls performed by Sanz-Blasco et al. show that under the conditions of these experiments the NSAIDs they used have no effect on cytosolic Ca²⁺ concentrations (Figure 7). If it is amyloid oligomers that let Ca²⁺ through plasma membranes, then this would appear to rule out structural disruption as a mechanism. Instead, Sanz-Blasco et al. propose that these NSAIDs specifically alter the polarity of the mitochondrial membrane in such a way as to prevent Ca²⁺ uptake.

If this is true, then NSAIDs may be able to perform a double-whammy on Alzheimer's disease. On the one hand, they appear to be capable of altering Aβ cleavage patterns to reduce the formation of toxic oligomeric precursors. In addition, they appear to have an ability to block mitochondrial breakdown and subsequent apoptosis directly. While this is encouraging, and speaks to the value of pursuing refinements of existing NSAIDs as possible Alzheimer's treatments, this experiment doesn't necessarily prove any therapeutic value. Even if the neurons are saved from death, the calcium flood may impair their function to such a degree that their continued survival doesn't matter. Only clinical trials and further research can firmly establish whether current or optimized NSAIDs can provide significant protection against Alzheimer's disease.

1. Sara Sanz-Blasco, Ruth A. Valero, Ignacio Rodríguez-Crespo, Carlos Villalobos, Lucía Núñez (2008). Mitochondrial Ca2+ Overload Underlies Aβ Oligomers Neurotoxicity Providing an Unexpected Mechanism of Neuroprotection by NSAIDs PLoS ONE, 3 (7), 0-0 DOI: 10.1371/journal.pone.0002718 OPEN ACCESS

2. A. Demuro, E. Mina, R. Kayed, S.C. Milton, I. Parker, C.G. Glabe (2005). Calcium Dysregulation and Membrane Disruption as a Ubiquitous Neurotoxic Mechanism of Soluble Amyloid Oligomers Journal of Biological Chemistry, 280 (17), 17294-17300 DOI: 10.1074/jbc.M500997200 OPEN ACCESS

3. K Kuchibotla, S Goldman, C Lattarulo, H Wu, B Hyman, B Backsai (2008). Aβ Plaques Lead to Aberrant Regulation of Calcium Homeostasis In Vivo Resulting in Structural and Functional Disruption of Neuronal Networks Neuron, 59 (2), 214-225 DOI: 10.1016/j.neuron.2008.06.008

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Diseases and cures in news and advertising

You've certainly seen some of the ads before, selling drugs that cure restless legs syndrome or erectile dysfunction. You've seen the news stories too, with credulous reporters blindly parroting news releases or self-promoting scientists without actually assessing evidence or costs. Companies have been accused of selling diseases at the same time they sell the cures, and the news media has historically done a poor job of educating the public about the true value of new advances in science and medicine. Today in PLoS Medicine, a pair of articles look at corporate and media disease-mongering, and the quality of medical reporting in general.

The first of these articles is an essay by Ray Moynihan and colleagues stating that disease mongering has entered the global health debate. Disease mongering—the creation or promotion of a diagnosis—is something you've probably suspected when watching those commercials, and it's a perfectly understandable strategy. Research into "blockbuster" drugs for major diseases often turns up medicines that fail at their intended task but have some other benefit (Viagra is a famous example of this phenomenon). Companies attempting to recoup the research investment have a substantial incentive to grow the market for these drugs. Even if there is a small group of patients for whom, say, restless leg syndrome is a significant problem, the attempt to expand this market to those for whom it is merely an occasional annoyance is disease mongering. When this tactic succeeds, people end up buying medication they don't really need, which has adverse economic consequences, not to mention the personal harm that may be suffered due to occasionally severe side effects or adverse interactions with other drugs.

The essay follows up a conference and special issue of PLoS Medicine from two years ago on this subject. The authors are pleased to report that several newspaper articles and websites are now taking notice, but this seems too optimistic to me. It is encouraging that we've seen greater awareness of disease mongering in recent years. The authors, however, are unable to point to any significant regulatory progress. A few newspaper articles and a couple of little-known websites are hardly enough to combat a pharmaceutical industry that is dedicated to selling drugs that may have a small natural audience. If we are to have a true national health care coverage program it will be essential to take regulatory steps to prevent the creation or expansion of diagnoses to justify the existence of drugs post facto. In light of the silence of regulatory bodies, even the modest optimism of the essay seems unfounded.

The other article comes from the creators of one of those little-known websites, which I hope will become better known, HealthNewsReview.org. The way the system there works is that three reviewers (one of whom is always this article's author, Gary Schwitzer), evaluate health news stories from television, newspapers, and wire services on the basis of ten criteria ranging from whether the story adequately discusses costs to whether independent sources were used. In this article, Schwitzer reports the results of a survey of 500 stories from US media sources, and the results are not particularly encouraging.

The frequency of particular criteria being met indicates a strong sensationalistic bent in American media. High percentages of articles established the novelty of the approach and discussed its availability. However, less than 40% of the articles did a satisfactory job of discussing the quality of the evidence, the existence of alternative options, or quantifying harms and benefits associated with a new treatment. Less than a quarter adequately discussed the cost of the new treatment. To my mind this speaks to a certain amount of hype. As Schwitzer notes, some of these problems are due to constraints of space and time—in a limited number of paragraphs it is far easier to convey that a novel cure has been discovered than to realistically describe the limitations of the study or the expense of the medicine. Moreover, many of the writers simply lack the expertise to interpret findings, or the resources to find experts to give a second opinion.

This is not entirely the fault of the media, however. Scientific journals, research corporations, and universities contribute by providing sensationalistic press releases that serve as the starting point (if not the entirety) of many of these stories. With commercial interests this practice is standard, though hardly worthy of celebration. In my opinion, however, it is inexcusable for universities and scientific journals to adopt this approach. The reporting of scholarship ought to be about truth, not buzz.

The presentation of results in this article seems somewhat incomplete. I would have especially liked to see a histogram showing the frequency with which a single article fulfilled multiple criteria. Do articles generally fall near the extremes, or do the results more resemble the familiar bell curve? Appropriate policy recommendations may differ significantly depending on this distribution. It would also be valuable to know the track records of particular publications: this data is not available in the article or on the website, yet it could be a valuable tool in putting pressure on editors and managers to give writers the space, resources, and training necessary to produce quality health reporting.

A further weakness of this study is that it does not address cable news or alternative media outlets such as websites and blogs, from which a significant proportion of Americans now get their news. This is especially incongruous in light of the fact that HealthNewsReview.org is itself an internet resource. To be fair, many news websites simply take their stories from the AP feed, which the study examined, and studying all these varied sources would have been an immense task. However, it seems obvious that a comprehensive evaluation of all media was never an achievable goal. Knowing that, it would have made more sense to select the broadest spectrum of types of sources, rather than intensively scanning several very similar sources. It is to be hoped that HealthNewsReview.org will expand its examination into these areas soon.

Sensationalism and sloppiness in scientific reporting represent a failure of journalism because they create a misinformed public. Moreover, overhyping of predictions and results that are often quite equivocal in reality contributes to mistrust of scientists and doctors. Scientists and journalists both must do a better job of communicating the real limits and implications of scientific findings, rather than casting all results in the most positive or revolutionary light. Hopefully HealthNewsReview.org and similar initiatives in other countries will help them do that.

PLoS Medicine is an open access journal, so you can check these articles out for free. Also check out the commentary from the PLoS Medicine editors.

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Purity is death

Perhaps you have already heard about the 14-year-old Washington boy who refused extended blood transfusion treatment and therefore died (some additional info can be found here and here). Obviously, this is a terrible tragedy for his family, and I certainly hope his aunt is a true believer because if she is not then she'll have no solace for the fact that her ignorant superstitious nonsense killed him. The judge in the case, who upheld the boy's right to refuse the transfusions, will come in for a great deal of much-warranted scrutiny, but the fact of the matter is that it is the Jehovah's Witnesses who deserve the scrutiny and the blame for what has happened.

I think anyone reading about this will have a knee-jerk reaction that the judge in the case made the wrong decision. The AP quotes judge Meyer as saying that the boy understood the consequences of his decision, which may well be true, in an analytical sense. Young Mr. Lindberg probably understood that he would die, but there are very few 14-year-olds, let alone 14-year-old boys, who have a good appreciation of what that means. Death isn't something you understand unless you've spent some time around it, brushed up against it. And think of the teenagers you know. Would you trust any of them with life-and-death decisions? Hell, we don't trust 14-year-olds with cars. It's also true that the boy's parents, who did not have custody of him, wanted him to take the transfusion—could his relationship with them have played a role in his decision?

So Meyer can be justly criticized on the grounds that Lindberg was not competent to make that judgment, or that it wasn't Lindberg's judgment to make. However, it should also be pointed out that this was not some one-off transfusion that would instantly cure the boy. The treatment under discussion was a long course of transfusions that would run alongside the chemotherapy. And according to the doctors the prognosis was that he had a 70% chance of surviving the ordeal, with all the discomfort and side effects to boot. Being forced to undergo the treatment against his will would certainly make this harder on the boy, and on his doctors.

That said, I personally feel that Meyer should have erred on the side of curing the boy. Lindberg's decision was dangerous and self-destructive, and this should have indicated the opposite ruling. However, I wasn't present for the hearing, and the decision Meyer did make wasn't groundless. Maybe there was something in Lindberg's demeanor suggesting greater maturity than his age would typically indicate.

You'll note that I didn't say anything about the religious sensibilities. That is because I give them no weight at all. Lots of people dislike the Jehovah's Witnesses for a variety of reasons, but I've never been bothered by them; certainly not to the degree that I am bothered by other odious "Christians" living a life of hatred at maximum volume. So this is not a statement emerging from a blanket dislike: their attitude towards transfusion reeks of ignorance, superstition, and flat-earthism. The soul, if it exists, is not bound up in any bodily organ or fluid. Certainly people who have received massive blood transfusions have not absorbed someone else's soul—or at least I'm sure that didn't happen to me.

There is a bright line with religious beliefs, especially laws of practice: they're fine as long as they don't hurt anyone. We in America do not allow cannibalism or polygamy, although these are both religious practices with long histories. Nor are we tolerant of female genital mutilation, stoning people to death for violations of the laws of Leviticus, or human sacrifice, religious practices all. This indoctrination against lifesaving medical procedures is just as dangerous and fatal, especially when the subject is a teenager lacking in perspective and a diversity of life experiences. Judge Meyer made a mistake by allowing Lindberg to finish the deed, but it was the Jehovah's Witnesses and their purity practices that killed that boy. That ought not be allowed.

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