Vaccination against amyloid is a promising approach for the development of Alzheimer’s disease (AD) therapeutics. Approximately half of the investigational new therapeutics in human clinical trials for AD are active or passive immunotherapeutics.
Active vaccination involves the injection of an antigen and relies on the production of antibodies in the vaccinated patient. Four human clinical trials of active vaccination currently are under way. Passive immunization is also a promising strategy that involves the production of antibodies outside of the patient and injection of these antibodies. There are currently 12 clinical trials of passive immunization. You can check for Alzheimer therapeutics in human clinical trials by visiting www.clinicaltrials.gov and searching for key words “Alzheimer’s and immunotherapy.”
Thinking out of the box
The development of vaccinations as a strategy for treating or preventing Alzheimer’s is an example of thinking out of the box. Vaccinations commonly are associated with infectious diseases, like influenza, small pox and polio, which appear to have little in common with neurodegenerative diseases, like Alzheimer’s. Moreover, the brain is an immunoprivileged site with little access to antibodies, so it seems unlikely antibodies would be protective in the brain.
Researchers were pleasantly surprised when Dale Schenk and co-workers at Elan Inc. reported that vaccination of transgenic mouse models of AD against the amyloid Aß peptide prevented amyloid deposition in young animals and removed pre-existing amyloid deposits in older animals. Subsequent work showed that immunization against Aß prevented or reversed many other pathological features and prevented cognitive dysfunction in transgenic mice and non-human primates. This vaccine (Elan AN1792) was tested in human clinical trials, where it showed similar beneficial effects of removing amyloid deposits and slowing cognitive decline in patients with significant levels of anti-Aß antibodies, but the clinical trial was halted because 6 percent of the patients developed meningoencephalitis, an inflammatory side effect.
Second-generation vaccines and passive immunization
To circumvent the unwanted inflammatory side effects, second-generation active vaccines have been developed and passive immunization strategies have been explored. The second-generation vaccines use small pieces of the amyloid Aß sequence to avoid activating the T-cells responsible for meningoencephalitis, while passive immunization bypasses the human immune response by directly supplying antibodies. These newer strategies have shown the same beneficial effects in transgenic mice and passive immunization has shown some promise in a subset of patients in human trials, but they have raised new questions about their effectiveness and potential new side effects. Elan/Wyeth reported preliminary results from clinical trials of their monoclonal antibody, Bapineuzimab, that demonstrated only a small benefit in a subgroup of patients who lack the apoE4 genotype. They also failed to observe an improved benefit with an increased dose of antibody and reported side effects, like a buildup of fluid in the brain. Results of active vaccination human clinical trials with second-generation vaccines remain to be reported.
Third-generation vaccines and antibodies: Thinking perpendicular to the box
Both second-generation vaccines and antibodies suffer from a common problem. They both target linear amino acid sequences found in normal human proteins (the amyloid precursor protein) and in the amyloid deposits themselves. Making antibodies against normal human proteins can cause autoimmune side effects, in which the immune system is attacking normal human cells in addition to the Alzheimer’s pathology. Fortunately, it is difficult to make antibodies against self-proteins because of immune suppression of auto antibodies. Third-generation vaccines seek to overcome these problems of autoimmune side effects and autoimmune suppression by using antibodies that target structures specific to the amyloid aggregates and that do not react with normal human proteins.
Cure Alzheimer’s Fund has been supporting two projects that seek to develop third-generation immunotherapeutics. Dr. Charles Glabe’s laboratory is developing active vaccines and monoclonal antibodies that recognize conformations of the amyloid peptide that only occur in the pathological amyloid oligomer aggregates, while Dr. Rob Moir’s lab is working on cross-linked amyloid peptides (CAPs) that are only found in disease-related aggregates. Dr. Glabe’s strategy relies on the fact that when the Aß peptide aggregates into ß-sheet oligomers, it creates new antibody recognition sites, known as epitopes, that are not found on native proteins. The surprising finding is that these oligomer-specific antibodies recognize amyloid oligomers from other diseases that involve amyloids formed from sequences unrelated to Aß. This means the same antibodies also may be effective for other amyloid-related neurodegenerative diseases, like Parkinson’s disease.
The explanation for why the antibodies are specific for amyloid oligomers that involve several individual peptide strands arranged in a sheet and yet recognize these sheets when they are formed from other amino acid sequences is simple and elegant (Figure 1). It is now known that most pathological amyloids aggregate into simple and very regular structures where the peptide strands are arranged in parallel and where the amino acid sequence is in exact register. This is like a sheet of paper upon which the same sentence is written on each line. The individual amino acids line up and down the sheet in homogeneous tracts, known as “steric zippers.” The steric zippers do not occur in normal protein structures and the oligomer-specific antibodies are thought to recognize these steric zipper patterns on the surface of the sheets. Since all proteins are made up using the same 20 amino acids, any sequence in this parallel, in-register structure gives rise to the same steric zippers regardless of the linear sequence, which can explain why the antibodies recognize the oligomers formed by different proteins.
Dr. Moir’s group is working on CAPs, where Aß is cross-linked by oxidation of a tyrosine residue at position 10 of the peptides’ sequence. Aß is oxidized after it is produced from the amyloid precursor protein as a consequence of the abnormally high level of oxidative activity in a brain with AD and the peptides’ propensity to bind redox active metals. Excessive CAPs generation is associated with the disease state and is not a normal feature of Aß biology. The cross-linking at tyrosine 10 that gives rise to CAPs may serve to align the peptides in a parallel, in-register fashion and promote the generation of still-larger oligomeric aggregates that display steric zippers on their surface.
Dr. Moir and Dr. Rudy Tanzi’s labs found that natural antibodies to CAPs are reduced in the blood of patients with AD. More recently, evidence published by Tony Weiss-Coray’s group at Stanford University supports the idea that antibodies that recognize steric zippers and CAPs may be important for protecting against Alzheimer’s disease. The levels of these antibodies that target the zippers and CAPs were among the highest in young, normal humans; levels dropped with aging and with AD. Furthermore, the results of a recent study supported by Baxter Biosciences of patients that received human antibodies purified from normal individuals (IVIg) reported that antibody treatment reduced the risk of being diagnosed with AD by 42 percent over the five-year study period. This is one of the most remarkable reports of prevention of AD by any therapy. Although the normal human antibodies that target amyloid primarily recognize the steric zippers and CAPs, these antibodies are present at relatively low levels. It is reasonable to imagine that an even greater protective effect might be achieved by boosting the levels of these protective antibodies by either active vaccination or passive immunization.
Figure 1 shows how the same steric zipper patterns are formed on parallel, in-register oligomers from completely different sequences. A segment of the Aß sequences is shown in the upper left corner and a random sequence is shown in the upper right. Each amino acid is designated by a capital letter. Typical antibodies recognize the linear sequence (from left to right) indicated in the horizontal boxes, which is unique to each sequence. When the peptides aggregate to form pathological oligomers, they line up in a parallel, in-register fashion, shown below. This gives rise to steric zippers that run up and down the sheet perpendicular to the sequence, shown in vertical boxes. Aggregation-dependent, disease-specific antibodies recognize the steric zippers from many different amyloid sequences. Zippers from F and V amino acids are shown in boxes, but there are potentially 20 different zippers; one for each of the 20 amino acids.
The fact that a completely random sequence can form the same type of steric zipper as is found in Aß amyloid in Alzheimer’s disease means we can use a non-human, random peptide sequence as a vaccine to produce a protective immune response that has a very low potential for autoimmune side effects. Vaccines based on non-human peptides, like diphtheria and pertussis toxin, are so safe they routinely are given to infants. There is no reason to expect that a vaccine for AD that targets the disease-specific steric zippers wouldn’t be as safe and free of side effects. A goal of the research funded by Cure Alzheimer’s Fund is to do the preclinical investigations that are a necessary prelude to getting these third-generation vaccines and monoclonal antibodies that target disease-specific epitopes into human clinical trials.