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Keywords:

  • Alzheimer's disease;
  • amyloid-beta protein;
  • immunotherapy;
  • protofibrils;
  • transgenic mice

Abstract

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. BIOMARKERS
  5. BIOCHEMICAL ANALYSES OF Aβ FIBRILLIZATION AND SOLUBLE Aβ AGGREGATES
  6. DEVELOPMENT OF CONFORMATION-SELECTIVE ANTIBODIES AND A DIAGNOSTIC ELISA
  7. A NEW TRANSGENIC MODEL OF ALZHEIMER'S DISEASE: CHARACTERIZATION AND PHARMACOLOGICAL STUDIES
  8. CONCLUSION
  9. Acknowledgment
  10. REFERENCES

The identification of disease-causing mutations in Alzheimer's disease has contributed greatly to the understanding of the pathogenesis of this disease. The amyloid-β (Aβ) peptide has come into focus and is believed to be central to the pathogenesis of Alzheimer's disease. With only symptomatic treatment available, efforts to develop new therapeutics aimed at lowering the amount of Aβ peptides in the affected brain have intensified. In particular, immunotherapy against Aβ peptides has attracted considerable interest, as it offers the possibility to generate highly specific molecules targeting highly specific moieties. Due to intense research efforts and massive investments at universities and in the pharmaceutical industry, the outlook for patients and their relatives has never been brighter.


INTRODUCTION

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. BIOMARKERS
  5. BIOCHEMICAL ANALYSES OF Aβ FIBRILLIZATION AND SOLUBLE Aβ AGGREGATES
  6. DEVELOPMENT OF CONFORMATION-SELECTIVE ANTIBODIES AND A DIAGNOSTIC ELISA
  7. A NEW TRANSGENIC MODEL OF ALZHEIMER'S DISEASE: CHARACTERIZATION AND PHARMACOLOGICAL STUDIES
  8. CONCLUSION
  9. Acknowledgment
  10. REFERENCES

Alzheimer's disease (AD) has an insidious onset and a progressive course. Due to multiple cognitive dysfunctions and behavioral changes, afflicted patients become dependent upon care, which is often provided by close relatives. AD is the most common dementia disorder, accounting for 50–60% of all dementia cases, and as demographics change the costs for dementia care will continue to increase. The only available drug therapies are acetylcholine esterase inhibitors and memantine, an N-methyl-D-aspartate (NMDA)-receptor inhibitor. Discoveries in the 1970s that suggested AD is caused by atrophy of cholinergic pathways from the basal forebrain and acetylcholine deficiency led to the development of acetylcholine esterase inhibitors. These drugs are purely aimed at treating AD symptoms and do not influence the underlying neurodegenerative processes, which are now known to involve multiple neurotransmitter systems and neuronal populations and not only cholinergic neurons. Therefore, the clinical effects of acetylcholine esterase inhibitors are transient and modest, and better medical therapy of AD is needed. The understanding of AD pathogenesis has increased greatly since the early 1990s, and this might lead to the development of new types of drugs.

AD is currently perceived as a protein aggregation disorder in which amyloid-β (Aβ) peptides initiate and drive the pathogenesis that leads to the dysfunctions and demise of neurons and, finally, to dementia. Several mutations result in early-onset AD, either by increasing Aβ1 or by elevating the Aβ42/Aβ40 ratio. Increased Aβ levels likely accelerate aggregation. Moreover, the Arctic (E693G) mutation2 in the Aβ domain of the amyloid precursor protein (APP) results in Aβ peptides prone to form Aβ protofibrils, i.e., large soluble Aβ aggregates. The production, aggregation, and clearance of Aβ are thus all attractive and feasible targets for drug development. Enzymes like β- or γ-secretase, which regulate the processing of APP and Aβ production, could be inhibited with small molecules synthesized by organic chemists. However, it has proved difficult to make substances that penetrate into the brain, are specific for the target, and are not toxic.3 Increased clearance of Aβ is an alternative approach, and in this arena biological therapeutics have been at the forefront. Immunotherapy comprises active and passive immunization. A vaccine represents active immunotherapy that induces an immune response, while passive immunotherapy means delivery of antibodies. Recent advances in protein engineering and the production of recombinant proteins make it feasible to produce tailor-made antibodies at a reasonable price for therapeutic use. Examples of successful immunotherapies are Rituxan® (Genentech, South San Francisco, Calif., USA) for lymphoma and other types of cancers and Remicade® (Centocor Ortho Biotech, Inc., Horsham, Pa., USA) for rheumatoid arthritis.

In 1999 Schenk et al.4 showed that active vaccination with the Aβ peptide could possibly be used to treat AD. Using vaccination with Aβ fibrils, they were able to both prevent and clear amyloid deposits in transgenic mice with platelet-derived growth factor promoter expressing amyloid precursor protein (PDAPP). The marked therapeutic effects in the absence of side effects in animals led to the rapid initiation of clinical trials with AD patients. Although initially promising, the tests were halted in phase II clinical trials because some patients developed meningoencephalitis. This inflammatory condition could be reversed with corticosteroids. Another drawback seen in this study was that high antibody titers were reached in only approximately 20% of the patients.5 This was not unexpected, since the immune system in the elderly is known to be difficult to trigger. Findings in postmortem brain from several patients enrolled in this study, however, showed vaccination was efficacious. Markedly little amyloid deposition was found in the brains of these subjects,6 and levels of tau in the cerebrospinal fluid (CSF) were reduced,5 indicating suppressed neurodegenerative processes. The potential side effects and the low number of responders to active vaccination led researchers to focus instead on passive immunotherapy with recombinant Aβ antibodies. Several antibodies targeting the N- and C-terminus as well as the mid-domain of Aβ are currently under investigation in clinical trials.7

In recent years there has been increased interest in soluble oligomeric assemblies of Aβ, rather than senile plaques, as the culprit in AD. In the late 1990s the Arctic mutation was found in a family from northern Sweden. Studies of this mutation led to the realization that one of its pathogenic effects was to generate toxic soluble Aβ oligomers, i.e., protofibrils8–10; therefore, an attempt was made to target these soluble Aβ aggregates with immunotherapy. The development of a conformation-dependent antibody with the ability to recognize a unique structure in the Aβ protofibril was undertaken. In contrast, most other research groups have developed therapeutic Aβ antibodies that bind to a linear epitope in Aβ. Such antibodies will thereby also clear Aβ monomers and possibly interfere with normal physiological functions of Aβ.11

BIOMARKERS

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. BIOMARKERS
  5. BIOCHEMICAL ANALYSES OF Aβ FIBRILLIZATION AND SOLUBLE Aβ AGGREGATES
  6. DEVELOPMENT OF CONFORMATION-SELECTIVE ANTIBODIES AND A DIAGNOSTIC ELISA
  7. A NEW TRANSGENIC MODEL OF ALZHEIMER'S DISEASE: CHARACTERIZATION AND PHARMACOLOGICAL STUDIES
  8. CONCLUSION
  9. Acknowledgment
  10. REFERENCES

In order to develop new drugs for AD, it is important to evaluate efficacy in patients. The efficacy of symptomatic drugs is determined only with rating scales of patients' cognitive functions or activities of daily living. The use of functional rating scales alone for the evaluation of new disease-modifying drugs can be misleading. Clinical trials need to be complemented with biomarkers, quantitative measures that reflect biological processes in the AD brain that are intended to be targeted by the drug and, ideally, are also relevant to the pathogenesis. Unfortunately, the biomarkers available for AD are limited to Aβ and tau in CSF, structural imaging of brain atrophy, and positron emission tomography with labeled glucose or amyloid ligands.12 Most of these markers have early ceiling effects and are therefore not valuable for measuring the delay in disease progression. Thus, another aim of developing conformation-dependent antibodies that could recognize Aβ protofibrils was to use these for the development of in vitro assays to measure Aβ oligomers in CSF in order to improve the differential diagnosis of dementia disorders and to measure disease progression.8

BIOCHEMICAL ANALYSES OF Aβ FIBRILLIZATION AND SOLUBLE Aβ AGGREGATES

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. BIOMARKERS
  5. BIOCHEMICAL ANALYSES OF Aβ FIBRILLIZATION AND SOLUBLE Aβ AGGREGATES
  6. DEVELOPMENT OF CONFORMATION-SELECTIVE ANTIBODIES AND A DIAGNOSTIC ELISA
  7. A NEW TRANSGENIC MODEL OF ALZHEIMER'S DISEASE: CHARACTERIZATION AND PHARMACOLOGICAL STUDIES
  8. CONCLUSION
  9. Acknowledgment
  10. REFERENCES

Considerable knowledge has been gained about the formation and dissolution of Aβ aggregates as well as their regulation by various physiochemical parameters.13 Synthetic peptides, both wild type (Aβwt) and with the Arctic mutation (AβArc), were investigated and compared at different conditions in vitro. A size-exclusion chromatography/high-performance liquid chromatography system was used, and since Aβ42 peptides are very hydrophobic, this system needed to be optimized in order to allow the detection of Aβ protofibrils. Certain fatty acids, such as docosahexaenoic acid, can stabilize Aβ protofibrils.14,15 The finding of stabilized Aβ protofibril preparations was valuable for preparing antigen for immunization. Furthermore, conditions in which incubation of synthetic Aβ peptides results in pure Aβ monomer and Aβ fibril preparation were established. The quality of the Aβ antigen preparations was verified both with size-exclusion chromatography/high-performance liquid chromatography and electron microscopy and tested for cellular toxicity with the MTT assay. It was shown that synthetic large soluble Aβ oligomers (protofibrils) were toxic to PC-12 cells but not to monomeric Aβ and Aβ fibrils. The basic knowledge on how to handle Aβ peptides and generate pure preparations of monomers, protofibrils, and fibrils was crucial for the development of antibodies and the diagnostic enzyme-linked immunosorbent assay (ELISA).

DEVELOPMENT OF CONFORMATION-SELECTIVE ANTIBODIES AND A DIAGNOSTIC ELISA

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. BIOMARKERS
  5. BIOCHEMICAL ANALYSES OF Aβ FIBRILLIZATION AND SOLUBLE Aβ AGGREGATES
  6. DEVELOPMENT OF CONFORMATION-SELECTIVE ANTIBODIES AND A DIAGNOSTIC ELISA
  7. A NEW TRANSGENIC MODEL OF ALZHEIMER'S DISEASE: CHARACTERIZATION AND PHARMACOLOGICAL STUDIES
  8. CONCLUSION
  9. Acknowledgment
  10. REFERENCES

Mice were immunized with Aβ protofibrils made from AβArc, in some cases stabilized with docosahexaenoic acid, to generate conformation-selective antibodies. Freund's adjuvant was used, and after a minimum of four boosts, when titers of anti-Aβ antibodies were high in sera, the mice were sacrificed and splenocytes prepared. After fusing them with Sp2/0 mouse myeloma cells, the hybridoma culture supernatants were tested for the presence of anti-Aβ antibodies. Positive clones were subcloned to certify monoclonality, and these were then further evaluated for their conformation selectivity using an inhibition ELISA. In this method, the antibody-antigen interactions take place in solution and at low concentrations, which makes it more suitable for affinity measurements than, for example, a direct ELISA in which the antigen is in large excess. Binding of the antibodies to low-molecular-weight (LMW)-Aβ or Aβ protofibril coat was inhibited by the addition of serially diluted Aβ in different conformations. The antigen concentration required to inhibit half of the maximum signal in the inhibition ELISA was defined as IC50, which can be used as an estimate of the antibody's affinity for the antigen investigated.16

Several monoclonal antibodies were characterized with the ELISA, but one was particularly interesting, mAb158 (IgG2a). As seen in Figure 1a, inhibition of mAb158's binding to LMW-Aβ coated wells with Aβ protofibrils resulted in IC50 values in the low nanomolar range, at least 200-fold lower than for LMW-Aβ and Aβ1–16. Thus, binding of mAb158 to Aβ was dependent upon conformation. As a comparison, the commercially available antibody 6E10 had equal affinity for Aβ protofibrils, AβArc protofibrils, LMW-Aβ, and Aβ1–16 and thus did not show any preference for a certain Aβ conformation (Figure 1b). In addition to mAb158, the high-affinity IgG1 antibody mAb1C3 was obtained. In contrast to mAb158, mAb1C3 was more similar to 6E10, with small differences in IC50 for the different Aβ conformations (Figure 1c). To further characterize the antibodies' Aβ-binding patterns, synthetic Aβ protofibrils and LMW-Aβ were analyzed in a low-denaturing Western blot in which most of the Aβ in the protofibril preparation remained aggregated. This experiment revealed that mAb158 bound to a smear of Aβ aggregates in the range of 50–200 kDa but not to monomeric Aβ, whereas 6E10 and mAb1C3 bound both to the aggregate smear and to monomeric Aβ, again proving mAb158 is conformation specific (Figure 1e). Further studies revealed mAb158, in contrast to other conformation-dependent antibodies,17,18 did not recognize oligomers and fibrils of other amyloidogenic proteins. Moreover, it did not bind APP.

image

Figure 1. mAb158 is a high-affinity amyloid-β (Aβ) conformation-dependent antibody. (a–c) Binding of mAb158, 6E10, and mAb1C3 to different Aβ conformations was analyzed by inhibition ELISA on low-molecular-weight Aβ (LMW-Aβ) coated wells with the molar (mol/L) concentration of Aβ displayed on the x-axis. (a) mAb158 displayed 200-fold higher affinity for Aβ in the protofibril conformation compared with LMW-Aβ and the N-terminal Aβ1–16 fragment. (b) 6E10 and (c) mAb1C3 did not show any conformation selectivity, and Aβ17–40 did not inhibit binding of any of the antibodies. (d) Small differences in IC50 values (nmol/L) were obtained when inhibiting mAb158's binding to LMW-Aβ coat and Aβ protofibril coat, respectively. (e) The binding of mAb158 to different Aβ species was also analyzed with a low-denaturing Western blot, in which mAb158 bound to a smear of Aβ aggregates in the 50–200 kDa range but not to LMW-Aβ, whereas 6E10 and mAb1C3 bound both to the smear and to LMW-Aβ. Reproduced with permission from Englund et al. (2007).8

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The characteristics of mAb158 enabled specific measurements of Aβ protofibrils by using mAb158 as both capturing and detecting antibody. To validate the conformation specificity of the ELISA (Figure 2a), titrated synthetic LMW-Aβ, Aβ protofibrils, and Aβ1–16 were used. The hydrophilic Aβ1–16 peptide was used because it is not expected to aggregate, and an ELISA composed of two identical antibodies requires at least a dimer of a protein to produce a signal. As predicted, the nonaggregating Aβ1–16 was not detected with the mAb158 sandwich-ELISA, even at micromolar concentrations (Figure 2a). To further validate the specificity of the protofibril ELISA, an increasing excess of Aβ1–16 to a fixed concentration of Aβ protofibrils (50 pmol/L when expressed in monomer units) was added and analyzed with both the mAb158 ELISA and a 6E10-6E10 sandwich ELISA (Figure 2b). A 500,000-fold molar excess of Aβ1–16, when compared with Aβ protofibrils, did not disturb the measurements with the mAb158 sandwich ELISA, as expected because Aβ1–16 binds poorly to the capture antibody. In contrast, a 500-fold excess of Aβ1–16 was enough to decrease the signal in the 6E10-6E10 ELISA, in which Aβ1–16, as shown in Figure 1b, binds with high affinity to the capture antibody (Figure 2b) and thereby occupies the binding sites of the capture antibody and inhibits detection of Aβ protofibrils.

image

Figure 2. Amyloid-β (Aβ) protofibril-specific sandwich ELISA. (a) Analysis of synthetic Aβ protofibrils, with and without the Arctic mutation, compared with low-molecular-weight Aβ (LMW-Aβ) and Aβ1–16 with the mAb158 sandwich ELISA shows the Aβ protofibril specificity of this method. (b) Analysis of 50 pmol/L synthetic Aβ protofibrils with the mAb158 sandwich ELISA was not affected by the addition of 500,000-fold molar excess of Aβ1–16, whereas a 500-fold excess of Aβ1–16 was enough to disturb the analysis with a 6E10-6E10 sandwich ELISA. Reproduced with permission from Englund et al. (2007).8

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The mAb158 sandwich ELISA measured Aβ protofibrils down to approximately 1 pmol/L and enabled Aβ protofibril levels to be assayed in biological samples. Aβ protofibril levels were significantly higher in APP-ArcSwe human embryonic kidney (HEK) cell culture media as compared to APP-Swe when HEK cells were transiently transfected, and Aβ protofibrils were not found in mock media as would be expected (Figure 3). Moreover, Aβ protofibrils were found in brains from tg-ArcSwe and tg-Swe mice homogenized in Tris-buffered saline (TBS) with the mAb158 sandwich ELISA. Similar to the findings with cell culture media, Aβ protofibril levels differed significantly between the groups and were higher in the brains of mice carrying the Arctic mutation (Figure 3). Background signals were detected in the nontransgenic mice, although they were significantly lower than those in transgenic mice, a phenomenon observed by others when measuring Aβ in rodent brain homogenates.19

image

Figure 3. The mAb158 ELISA detects Aβ protofibrils with and without the Arctic mutation equally. (a) Levels of Aβ protofibrils were approximately 2.5-fold higher in cell culture media from human embryonic kidney 293 cells transiently transfected with APP-ArcSwe (n = 11) than in cells transfected with APP-Swe (n = 8). Conditioned media from mock cells (n = 3) gave no signal. (b) Aβ protofibril levels in the TBS-soluble fraction of nontransgenic mouse brain homogenates (n = 6) were compared with tg-Swe (n = 3) and tg-ArcSwe (n = 6). (c) Similar to the cell culture media, Aβ protofibril levels of tg-ArcSwe mice were 3.5-fold higher than those in APP-Swe mice. Reproduced with permission from Englund et al. (2007).8

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Aβ protofibrils have been assayed in the CSF of transgenic tg-ArcSwe mice but not in the CSF of nontransgenic mice (Figure 4). Furthermore, Aβ protofibrils have been detected in human brain tissue homogenized in TBS. Aβ protofibrils are found only in brain from AD patients, suggesting it is a specific marker. Thus far, Aβ protofibrils have not been detected in human CSF with the mAb158 ELISA. Different approaches are now used to lower the detection limit of the assay in order to evaluate Aβ protofibrils as a CSF biomarker for AD in the near future.

image

Figure 4. Aβ protofibrils in AD brain and in cerebrospinal fluid of tg-ArcSwe. (a) Aβ protofibrils were found in both human brain homogenates and in CSF from transgenic mice. Significant levels of Aβ protofibril were found in the TBS-soluble fraction of postmortem brain of AD, but not in frontotemporal lobe dementia (FTD) or normal control. Aβ protofibrils were only detected in brains from deceased AD patients. (b) Aβ protofibrils were also found in the CSF of tg-ArcSwe mice (= 3), but not in nontransgenic mice (= 3), with the mAb158 sandwich ELISA.

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A NEW TRANSGENIC MODEL OF ALZHEIMER'S DISEASE: CHARACTERIZATION AND PHARMACOLOGICAL STUDIES

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. BIOMARKERS
  5. BIOCHEMICAL ANALYSES OF Aβ FIBRILLIZATION AND SOLUBLE Aβ AGGREGATES
  6. DEVELOPMENT OF CONFORMATION-SELECTIVE ANTIBODIES AND A DIAGNOSTIC ELISA
  7. A NEW TRANSGENIC MODEL OF ALZHEIMER'S DISEASE: CHARACTERIZATION AND PHARMACOLOGICAL STUDIES
  8. CONCLUSION
  9. Acknowledgment
  10. REFERENCES

A disease model is the critical link between test tubes or cell culture experiments and clinical research. The ability to experiment with drugs on patients and gain access to biopsies or other samples is limited. Studies with animal models are therefore essential for drug development. They should replicate important features of human disease, and as clinical experiences become available, be improved to enhance their predictive validity. The latter refers to the ability to forecast the outcome of a clinical study based on experiences from pharmacological and toxicological studies in animal models of human disease.

APP transgenic models (e.g., Tg2576, PDAPP) have been state-of-the-art to evaluate new drugs targeting Aβ.13 Pharmacological effects on Aβ metabolism, i.e., production and clearance of Aβ monomers, are determined by measuring Aβ and/or senile plaque burden – the final end-product of Aβ fibril formation. A limitation with these models is the inability to measure soluble Aβ aggregates, which are likely to be central to AD pathogenesis. A better transgenic model was developed by engineering such that Aβ peptides more prone to aggregate than wild-type Aβ were produced. Indeed, using both the Arctic (E693G) and Swedish (KM670/671NL) APP mutations, which enhance APP processing six- to eight-fold, a founder line with high expression of Arctic APP and unique Aβ phenotypes was identified. In these, punctate Aβ-immunopositive staining was seen inside neurons in brain tissues of young mice. At the time, there were only a few reports of a similar observation, which is referred to as intraneuronal Aβ, i.e., assemblies of Aβ inside neurons. In tg-ArcSwe mice, the new model developed, immunopositive Aβ granules were found already in very young mice, i.e., 1–2 months of age. Intraneuronal Aβ increased with age until senile plaques emerged, at around 5–6 months of age, and then declined. This suggested intraneuronal Aβ was intermediary in the process of Aβ aggregation.20 There were also increased levels of soluble Aβ protofibrils in young tg-ArcSwe mice as compared with other transgenic models lacking the Arctic mutation.9 When mice grew older, the frequency of senile plaques increased rapidly in an age-dependent manner, while levels of soluble Aβ protofibrils increased only modestly.21 Aβ plaques were found mainly in the hippocampus and cerebral cortex, brain regions typically affected in AD, and they were surrounded by affected nerve endings that were phospho-tau immunoreactive and by gliosis, a sign of local inflammatory reaction in the brain.20 These responses by nerve endings and cells in the vicinity of senile plaques are also seen in postmortem AD brain.

In further investigations, learning and memory in young and aged tg-ArcSwe mice showed modest impairment at an early age in comparison with that observed in nontransgenic mice. The Morris water maze, which challenges animals to locate a hidden platform beneath the water surface by remembering locations of landmarks surrounding the pool, was used in this study. The test is known to depend on functioning of the hippocampus, a brain region afflicted early in AD pathogenesis, resulting in reduced memory of recent events. Aβ protofibril levels were found to inversely correlate with spatial learning ability (Figure 5).21 In addition, new ways to functionally test transgenic mice have been developed. Large groups of mice have been studied in novel object recognition with the T-maze and the IntelliCage (TSE Systems, Inc., Chesterfield, Mo., USA). The IntelliCage is a new type of behavioral test in which mice are tested in a social setting in their home environment. Animals carry a microchip and are continuously tracked by computer.

image

Figure 5. Young tg-ArcSwe with Aβ protofibrils and intraneuronal Aβ are poor learners. (a) tg-ArcSwe mice show intraneuronal Aβ already at 1–2 months of age but this phenotype diminishes when senile plaques begin to appear at 6 months (inset: arrowheads point to intraneuronal Aβ aggregates, and arrow points to a senile plaque). (b) Spatial learning deficiency in young tg-ArcSwe mice as measured by the ability to find a submerged platform in the Morris water maze. (c) The ability to learn (= improvement) correlated inversely with Aβ protofibril levels, suggesting these interfere with the learning process. Reproduced with permission from Lord et al. (2006)20 (a) and Lord et al. (2009)21 (b and c).

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Amyloid deposition in brain was also investigated with a procedure of sequential biochemical extraction. Senile plaques are very difficult to dissolve in human AD brain and harsh reagents, while amyloid plaques in transgenic models like Tg2576 are soluble in water. Interestingly, in the tg-ArcSwe model, amyloid plaques were highly insoluble and similar to those in human AD brain. Moreover, through experiments with electron microscopy, new amyloid ligands, and luminescent conjugated probes, amyloid fibrils from the tg-ArcSwe mouse model were found to grow differently, resulting in a denser and more compact structure of senile plaques as compared to mice carrying only the Swedish mutation. 22

mAb158 TREATMENT OF TRANSGENIC MICE MODELING ALZHEIMER'S DISEASE

tg-ArcSwe mice have been used to evaluate the efficacy of the conformation-selective antibody, mAb158, which targets soluble Aβ protofibrils. Antibody mAb158 has been tested in several therapeutic regimens. The treatment was efficacious and safe and did not cause side effects observed with other types of immunotherapy, such as microhemorrhages due to disruption of vessels with cerebrovascular amyloid.23

The resilience of Aβ deposits in tg-ArcSwe might suggest this animal model is superior for testing drugs. APP transgenic models have been criticized for their lack of predictive validity, since a wide range of drugs are efficacious in the mice but not in AD patients. Indeed, recent investigations have shown existing plaques cannot be cleared with immunotherapy in tg-ArcSwe mice, while this is possible in Tg2576 mice. Clinical trials with immunotherapy, i.e., tailor-made antibodies against insoluble Aβ, are currently ongoing, and results from these studies will show their effects in patients in terms of senile plaque clearance and functional improvement. An alternative strategy, in which antibodies against soluble Aβ are used, will soon be tested in AD patients.

CONCLUSION

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. BIOMARKERS
  5. BIOCHEMICAL ANALYSES OF Aβ FIBRILLIZATION AND SOLUBLE Aβ AGGREGATES
  6. DEVELOPMENT OF CONFORMATION-SELECTIVE ANTIBODIES AND A DIAGNOSTIC ELISA
  7. A NEW TRANSGENIC MODEL OF ALZHEIMER'S DISEASE: CHARACTERIZATION AND PHARMACOLOGICAL STUDIES
  8. CONCLUSION
  9. Acknowledgment
  10. REFERENCES

Genetic studies suggest enhanced Aβ levels alone are sufficient to cause AD, but clinicopathological studies in which the frequency and location of senile plaques have been compared with cognitive dysfunctions in postmortem AD brain samples suggest senile plaques are not the main cause of dementia. It is therefore commonly believed that soluble Aβ aggregates are the main culprits of disease. Aβ aggregation and fibril formation is a complex multistep process, and it has been difficult to model the individual steps in this process both in vitro and in vivo. The ability to analyze protofibril formation and the further generation of fibrils in vitro has allowed significant progress to be made. This was a prerequisite for the development of a stable antigen and also for the screening of antibody candidates. The antibody identified, mAb158, and the transgenic model, tg-ArcSwe, are both unique in the world. These tools have made it possible to develop the idea to target Aβ protofibrils specifically with immunotherapy. Several immunotherapeutic strategies are now being tested in clinical trials. It is difficult to predict which one of these will be most successful. The addition of a unique approach, the selective targeting of a soluble Aβ aggregate, Aβ protofibrils, has contributed significantly to the field. The strategy has obvious advantages, such as the low risk of disrupting plaques and amyloid-laden vessels, minimal interference with Aβ monomers and thereby also with normal physiological functions linked to APP processing and Aβ metabolism.

REFERENCES

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. BIOMARKERS
  5. BIOCHEMICAL ANALYSES OF Aβ FIBRILLIZATION AND SOLUBLE Aβ AGGREGATES
  6. DEVELOPMENT OF CONFORMATION-SELECTIVE ANTIBODIES AND A DIAGNOSTIC ELISA
  7. A NEW TRANSGENIC MODEL OF ALZHEIMER'S DISEASE: CHARACTERIZATION AND PHARMACOLOGICAL STUDIES
  8. CONCLUSION
  9. Acknowledgment
  10. REFERENCES