Immunotherapy of Alzheimer's disease (AD): from murine models to anti-amyloid beta (Abeta) human monoclonal antibodies

The deposition of amyloid beta (Abeta) protein is a key pathological feature in Alzheimer's disease (AD). In murine models of AD, both active and passive immunization against Abeta induce a marked reduction in amyloid brain burden and an improvement in cognitive functions. Preliminary results of a prematurely terminated clinical trial where AD patients were actively vaccinated with aggregated Abeta bear resemblance to those documented in murine models. Passive immunization of AD patients with anti-Abeta antibodies, in particular human antibodies, is a strategy that provides a more cautious management and control of any undesired side effects. Sera of all healthy adults contain anti-Abeta IgG autoimmune antibodies. Hence antigen-committed human B-cells are easily immortalized by Epstein-Barr virus (EBV) into anti-Abeta secreting cell lines. Two anti-Abeta human monoclonal antibodies which we recently prepared bind to the N-terminus of Abeta peptide and were shown to stain amyloid plaques in non-fixed brain sections from an AD patient. It is anticipated that specifically selected anti-Abeta human monoclonal antibodies could reduce and inhibit deposits of amyloid in brain while avoiding the cognitive decline that characterizes AD. In the future, this type of antibody may prove to be a promising immune therapy for the disease.     

Enhanced Th2 immunity after DNA prime-protein boost immunization with amyloid beta (1-42) plus CpG oligodeoxynucleotides in aged rats

Generation and accumulation of fibrillar amyloid beta (Abeta) is widely considered as the pathogenic basis of neurodegeneration in Alzheimer's disease (AD). Both active immunization with fibrillar Abeta and passive immunization with anti-Abeta antibodies in transgenic mouse models of AD result in prevention/dissociation of Abeta plaque formation and restoration of cognitive functions. However, similar immunization studies in humans had to be halted because 6% of the AD patients developed acute meningoencephalitis, likely due to anti-Abeta specific autoimmune Th1 cells. Hence, making Abeta immunotherapy successful requires production of strong antibody responses without Th1-type immunity. In an attempt to develop safer vaccines, we examined the influence of oligodeoxynucleotides as adjuvant on the Th1 and Th2 immune response to Abeta in aged rats. We further investigated whether a DNA prime-protein boost strategy could elicit a more robust Th2 response. The results of the present study showed that all the animals injected with either Abeta peptide alone or Abeta encoding plasmid alone or plasmid DNA prime followed by peptide boost have elicited specific anti-Abeta antibodies. When co-administered, synthetic oligodeoxynucleotides (ODN) further enhanced the anti-Abeta titres. More importantly, the IgG subclasses of the antibodies generated by DNA prime-peptide boost regimen with ODN as adjuvant were primarily of IgG2b and IgG1 isotypes, suggesting that heterologous immunization strategy along with ODN would be advantageous in eliciting more beneficial Th2-type humoral immune response.     

Generation and characterization of the humoral immune response to DNA immunization with a chimeric beta-amyloid-interleukin-4 minigene

Active immunization with fibrillar beta-amyloid peptide (Abeta(42)) as well as passive transfer of anti-Abeta antibodies significantly reduces Abeta plaque deposition, neuritic dystrophy, and astrogliosis in the brain of mutant amyloid precursor protein (APP)-transgenic mice. Although the mechanism(s) of clearance of Abeta from the brain following active or passive immunization remains to be determined, it is clear that anti-Abeta antibodies are critical for clearance. DNA immunization provides an attractive alternative to direct peptide and adjuvant approaches for inducing a humoral response to Abeta. We constructed a DNA minigene with Abeta fused to mouse interleukin-4 (pAbeta(42)-IL-4) as a molecular adjuvant to generate anti-Abeta antibodies and enhance the Th2-type of immune responses. Gene gun immunizations induced primarily IgG1 and IgG2b anti-Abeta antibodies. Fine epitope analysis with overlapping peptides of the Abeta(42) sequence identified the 1-15 region as a dominant B cell epitope. The DNA minigene-induced anti-Abeta antibodies bound to Abeta plaques in brain tissue from an Alzheimer's disease patient demonstrating functional activity of the antibodies and the potential for therapeutic efficacy.     

Immunization reverses memory deficits without reducing brain Abeta burden in Alzheimer's disease model

We have previously shown that chronic treatment with the monoclonal antibody m266, which is specific for amyloid beta-peptide (Abeta), increases plasma concentrations of Abeta and reduces Abeta burden in the PDAPP transgenic mouse model of Alzheimer's disease (AD). We now report that administration of m266 to PDAPP mice can rapidly reverse memory deficits in both an object recognition task and a holeboard learning and memory task, but without altering brain Abeta burden. We also found that an Abeta/antibody complex was present in both the plasma and the cerebrospinal fluid of m266-treated mice. Our data indicate that passive immunization with this anti-Abeta monoclonal antibody can very rapidly reverse memory impairment in certain learning and memory tasks in the PDAPP mouse model of AD, owing perhaps to enhanced peripheral clearance and (or) sequestration of a soluble brain Abeta species.     

Enhancing Th2 immune responses against amyloid protein by a DNA prime-adenovirus boost regimen for Alzheimer's disease

Accumulation of aggregated amyloid beta-protein (Abeta) in the brain is thought to be the initiating event leading to neurodegeneration and dementia in Alzheimer's disease (AD). Therefore, therapeutic strategies that clear accumulated Abeta and/or prevent Abeta production and its aggregation are predicted to be effective against AD. Immunization of AD mouse models with synthetic Abeta prevented or reduced Abeta load in the brain and ameliorated their memory and learning deficits. The clinical trials of Abeta immunization elicited immune responses in only 20% of AD patients and caused T-lymphocyte meningoencephalitis in 6% of AD patients. In attempting to develop safer vaccines, we previously demonstrated that an adenovirus vector, AdPEDI-(Abeta1-6)11, which encodes 11 tandem repeats of Abeta1-6 can induce anti-inflammatory Th2 immune responses in mice. Here, we investigated whether a DNA prime-adenovirus boost regimen could elicit a more robust Th2 response using AdPEDI-(Abeta1-6)11 and a DNA plasmid encoding the same antigen. All mice (n=7) subjected to the DNA prime-adenovirus boost regimen were positive for anti-Abeta antibody, while, out of 7 mice immunized with only AdPEDI-(Abeta1-6)11, four mice developed anti-Abeta antibody. Anti-Abeta titers were indiscernible in mice (n=7) vaccinated with only DNA plasmid. The mean anti-Abeta titer induced by the DNA prime-adenovirus boost regimen was approximately 7-fold greater than that by AdPEDI-(Abeta1-6)11 alone. Furthermore, anti-Abeta antibodies induced by the DNA prime-adenovirus boost regimen were predominantly of the IgG1 isotype. These results indicate that the DNA prime-adenovirus boost regimen can enhance Th2-biased responses with AdPEDI-(Abeta1-6)11 in mice and suggest that heterologous prime-boost strategies may make AD immunotherapy more effective in reducing accumulated Abeta.     

Efficacy of anti-Abeta antibody isotypes used for intracerebroventricular immunization in TgCRND8

We have previously demonstrated that intracerebroventricular (ICV) injection of anti-Abeta (IgG1, kappa against the 1-28 region of Abeta) reduced cerebral amyloid plaques by 50% after 1 month without producing hemorrhage or activating IL-1beta responses in Tg2576 brain [N.B. Chauhan, G.J. Siegel, Reversal of amyloid beta toxicity in Alzheimer's disease model Tg2576 by intraventricular antiamyloid beta antibody, J. Neurosci. Res. 69 (1) (2002) 10-23]. The current report compares the efficacy of IgG1, IgG2a and IgG2b isotypes of anti-Abeta against several different epitopes of Abeta in clearing cerebral Abeta after a single bolus ICV injection in TgCRND8. Consistent with earlier in vitro findings from other laboratories, these in vivo data demonstrate that all IgG1 isotype antibodies tested cleared cerebral Abeta more efficiently than did IgG2a and IgG2b antibodies without producing histotoxicity in brain, liver or kidney, while an antibody against the C-terminus of Abeta did not reduce plaques or diminish their accumulation with aging of the animals. Intriguingly, there was no significant difference between the Abeta-reducing efficiency of IgG1 anti-Abeta antibodies directed against residues 3-6, against residues 1-10 or against residues 1-28 of N-terminus Abeta.     

Epitope mapping and neuroprotective properties of a human single chain FV antibody that binds an internal epitope of amyloid-beta 1-42

Active and passive immunotherapy targeted at the amyloid-beta (Abeta) peptide has been proposed as therapeutic approach against Alzheimer's disease (AD), and efforts towards the generation and application of antibody-based reagents that are capable of preventing and clearing amyloid aggregates are currently under active investigation. Previously, we selected and characterized a new anti-Abeta1-42 phage-displayed scFv antibody, designated clone b4.4, using a non-immune human scFv antibody library and demonstrated that a peptide based on the sequence of the Ig heavy chain (VH) complementarity-determining region (HCDR3) of this antibody fragment bound to Abeta1-42)and had neuroprotective potential against Abeta1-42 mediated neurotoxicity in rat hippocampal cultured neurons. In the present study, using novel computational methods and in vitro experiments we demonstrated that b4.4 binds to the central region of Abeta1-42. We also demonstrated that this scFv antibody binds to Abeta-derived diffusible ligands (ADDLs) and neutralizes the toxicity of both fibrillar and oligomeric forms of Abeta1-42 tested in vitro in SH-SY5Y cell cultures.     

Dendrimeric Abeta1-15 is an effective immunogen in wildtype and APP-tg mice

Immunization of humans and APP-tg mice with full-length beta-amyloid (Abeta) results in reduced cerebral Abeta levels. However, due to adverse events in the AN1792 trial, alternative vaccines are required. We investigated dendrimeric Abeta1-15 (dAbeta1-15), which is composed of 16 copies of Abeta1-15 peptide on a branched lysine core and thus, includes an Abeta-specific B cell epitope but lacks the reported T cell epitope. Immunization by subcutaneous, transcutaneous, and intranasal routes of B6D2F1 wildtype mice led to anti-Abeta antibody production. Antibody isotypes were mainly IgG1 for subcutaneous or transcutaneous immunization and IgG2b for intranasal immunization, suggestive of a Th2-biased response. All Abeta antibodies preferentially recognized an epitope in Abeta1-7. Intranasal immunization of J20 APP-tg mice resulted in a robust humoral immune response with a corresponding significant reduction in cerebral plaque burden. Splenocyte proliferation against Abeta peptide was minimal indicating the lack of an Abeta-specific cellular immune response. Anti-Abeta antibodies bound monomeric, oligomeric, and fibrillar Abeta. Our data suggest that dAbeta1-15 may be an effective and potentially safer immunogen for Alzheimer's disease (AD) vaccination.     

Alzheimer's disease abeta vaccine reduces central nervous system abeta levels in a non-human primate, the Caribbean vervet

Amyloid beta (Abeta) protein immunotherapy lowers cerebral Abeta and improves cognition in mouse models of Alzheimer's disease (AD). Here we show that Caribbean vervet monkeys (Chlorocebus aethiops, SK) develop cerebral Abeta plaques with aging and that these deposits are associated with gliosis and neuritic dystrophy. Five aged vervets were immunized with Abeta peptide over 10 months. Plasma and cerebral spinal fluid (CSF) samples were collected periodically from the immunized vervets and five aged controls; one monkey per group expired during the study. By Day 42, immunized animals generated plasma Abeta antibodies that labeled Abeta plaques in human, AD transgenic mouse and vervet brains; bound Abeta1-7; and recognized monomeric and oligomeric Abeta but not full-length amyloid precursor protein nor its C-terminal fragments. Low anti-Abeta titers were detected in CSF. Abetax-40 levels were elevated approximately 2- to 5-fold in plasma and decreased up to 64% in CSF in immunized vervets. Insoluble Abetax-42 was decreased by 66% in brain homogenates of the four immunized animals compared to archival tissues from 13 age-matched control vervets. Abeta42-immunoreactive plaques were detected in frontal cortex in 11 of the 13 control animals, but not in six brain regions examined in each of the four immunized vervets. No T cell response or inflammation was observed. Our study is the first to demonstrate age-related Abeta deposition in the vervet monkey as well as the lowering of cerebral Abeta by Abeta vaccination in a non-human primate. The findings further support Abeta immunotherapy as a potential prevention and treatment of AD.     

Reduced effectiveness of Abeta1-42 immunization in APP transgenic mice with significant amyloid deposition.

Vaccinations with Abeta1-42 have been shown to reduce amyloid burden in transgenic models of Alzheimer's disease (AD). We have further tested the efficacy of Abeta1-42 immunization in the Tg2576 mouse model of AD by immunizing one group of mice with minimal Abeta deposition, one group of mice with modest Abeta deposition, and one group with significant Abeta deposition. The effects of immunization on Abeta deposition were examined using biochemical and immunohistochemical methods. In Tg2576 mice immunized prior to significant amyloid deposition, Abeta1-42 immunization was highly effective. Biochemically extracted Abeta40 and Abeta42 levels were significantly reduced and immunohistochemical plaque load was also reduced. Immunization of mice with modest amounts of pre-existing Abeta deposits selectively reduced Abeta42 without altering Abeta40, although plaque load was reduced. In contrast, in Tg2576 mice with significant pre-existing Abeta loads, Abeta1-42 immunization only minimally decreased Abeta42 levels, whereas no alteration in Abeta40 levels or in plaque load was observed. These results indicate that in Tg2576 mice, Abeta1-42 immunization is more effective at preventing additional Abeta accumulation and does not result in significant clearance of pre-existing Abeta deposits.     

Aβ42及亚单位疫苗诱导小鼠抗体产生及其中和Aβ42的细胞毒性

目的 :探讨Aβ4 2 及其亚单位肽疫苗接种小鼠后特异性抗Aβ4 2抗体的产生情况。方法 :75只 6wk龄雄性BALB/c小鼠 ,随机分为 5组 :即对照组、Aβ4 2 组 ,Aβ3 6 - 42 组、Aβ1- 15 组和FAβ1- 15组。分别用PBS MF59佐剂 ,Aβ4 2 MF59,Aβ36～ 4 2 七聚赖氨酸 (MAP) MF59,Aβ1- 15 MAP MF59,Aβ1- 15 MAP 福氏佐剂 ,免疫BALB/c小鼠 4次。用间接ELISA法 ,检测各组免疫小鼠血清和脑组织匀浆上清液中特异性抗体的滴度。将Aβ42 、Aβ3 6 - 42 和Aβ1- 15 与培养的PC12细胞共同培养 7d ,用MTT比色法检测 3种抗原肽对PC12细胞的毒性。将各组免疫小鼠的血清加入到含 2 0mg/LAβ4 2 的培养基中 ,再与培养的PC12细胞一起培养 7d ,用MTT比色法测定PC12细胞的存活率。结果 :第 2次免疫后 ,各实验组小鼠的免疫血清均有抗Aβ42 抗体产生 ,且抗体滴度随接种次数的增多而增高。同时 ,在脑组织匀浆上清液中也可检测出低滴度的抗Aβ4 2 抗体。Aβ42 可降低PC12细胞的存活率 ;而不同浓度的Aβ3 6 - 42 和Aβ1- 15 则对PC12细胞的存活率无显著影响。将 4组疫苗免疫血清和 2 0mg/LAβ42 同时加入培养基中培养PC12细胞时 ,可显著提高其存活率。结论 :Aβ42 及其亚单位疫苗 (Aβ1- 15 及Aβ36～ 4 2 )结合MF59佐剂免疫B     

Intramuscular delivery of a single chain antibody gene reduces brain Abeta burden in a mouse model of Alzheimer's disease

Anti-beta-amyloid (Abeta) immunotherapy has been well documented to effectively elicit amyloid plaque clearance and slow cognitive decline in experimental and clinical studies. However, anti-Abeta immunotherapy was associated with detrimental effects of brain inflammation and microhemorrhage, presumably induced by T-cell-mediated and/or Fc-mediated inflammatory responses. In the present study, a single chain antibody (scFv) against Abeta could effectively inhibit the aggregation of Abeta and promote the disaggregation of preformed Abeta fibrils. The recombined adeno-associated virus vectors carrying the scFv gene were produced to delivery the scFv gene. Hippocampus delivery of the scFv gene was effective in reducing the amyloid plaque in the hippocampus of an Alzheimer's disease (AD) mouse model. Further studies demonstrated that intramuscular delivery of the scFv gene was as effective as intracranial delivery in reducing the total Abeta level in the brain with a concomitant elevated Abeta level in serum. No enhanced microglial activation, discernable T lymphocyte infiltration, and increased microhemorrhage were found after intracranial and intramuscular delivery of the scFv gene. Our results suggest that intramuscular delivery of the scFv gene would be a novel peripheral noninflammatory immunological modality targeting Abeta clearance and be promising in future drug development for the prevention and treatment of AD.     

Antibodies from a DNA peptide vaccination decrease the brain amyloid burden in a mouse model of Alzheimer’s disease

The neuropathology of Alzheimer's disease(AD) is characterized by the accumulation of amyloid peptide Abeta in the brain derived from proteolytic cleavage of the amyloid precursor protein (APP). Vaccination of mice with plasmid DNA coding for the human Abeta42 peptide together with low doses of preaggregated peptide induced antibodies with detectable titers after only 2 weeks. One serum was directed against the four aminoterminal amino acids DAEF and differs from previously described ones. Both immune sera and monoclonal antibodies solubilized preformed aggregates of Abeta42 in vitro and recognized amyloid plaques in brain sections of mice transgenic for human APP. Passive immunization of transgenic AD mice caused a significant and rapid reduction in brain amyloid plaques within 24 h. The combined DNA peptide vaccine may prove useful for active immunization with few inoculations and low peptide dose which may prevent the recently described inflammatory reactions inpatients. The monoclonal antibodies are applicable for passive immunization studies and may lead to a therapy of AD.     

Evidence supporting a role for anti-Abeta antibodies in the treatment of Alzheimer's disease

Antibodies against Abeta have been suggested as potential therapeutic strategies for the treatment of Alzheimer disease (AD) for nearly 8 years. Animal studies have been very encouraging in that both active and passive immunization of transgenic mice can reduce amyloid load and reverse memory deficits found in these mice. Three mechanisms have been proposed to explain these results: (a). catalytic conversion of fibrillar Abeta to less toxic forms, (b). opsonization of Abeta deposits leading to microglial phagocytosis, or (c). promote the efflux of Abeta from the brain to the circulation. Evidence exists supporting all three mechanisms, which, it should be noted, are not mutually exclusive. Phase 2 clinical trials of active immunization with vaccines against human Abeta1-42 were halted due to an unacceptable incidence of meningoencephalitic reactions (6% of patients treated). However, a recent report from a fraction of the patients in this trial found that those patients developing antibodies which reacted with brain amyloid deposits had a significantly slower progression of cognitive loss over a period of 12 months. This supports the continued cautious testing of passive immunization and, possibly even active immunization against the Abeta peptide using preparations less likely to cause autoimmune reactions in the central nervous system.     

接种Aβ42全肽疫苗恒河猴的特异性体液免疫应答

目的 观察恒河猴接种Aβ42肽疫苗后的特异性抗体的产生. 方法 将5只雄性恒河猴分别在0、 2、 6、 10、 14、 18、 22 wk肌内注射Aβ42肽疫苗; 用ELISA法检测恒河猴血清抗Aβ42抗体水平及IgG亚类; 用Western blot检测血清抗Aβ42抗体的特异性; 免疫组化染色法观察抗血清对Tg2576转基因小鼠脑组织中Aβ斑的识别. 结果 疫苗接种后第8周, 恒河猴血清中出现明显的抗Aβ42抗体, 抗体水平随着接种次数的增加而升高, 第24周达1∶ 4 320, 以后抗体水平开始下降.产生的抗Aβ42抗体以IgG1和IgG2为主(IgG2/IgG1>1).血清抗Aβ42抗体具有高度特异性, 可识别Tg2576转基因小鼠脑组织中的Aβ斑. 结论 Aβ42肽疫苗可有效地诱导恒河猴产生特异性体液免疫应答.     

Immunological and anti-chaperone therapeutic approaches for Alzheimer disease

Alzheimer disease (AD) is the most common cause of dementia. Currently available therapies only provide symptomatic relief. A number of therapeutic approaches are under development that aim to increase the clearance of brain Abeta peptides. These include immune mediated clearance of Abeta and the inhibition of the interaction between Abeta and its pathological chaperones. Both active and passive immunization has been shown to have robust effects in transgenic mouse models of AD on amyloid reduction and behavioral improvements. However, a human trial of active immunization has been associated with significant toxicity in a minority of patients. New generation vaccines are being developed which likely will reduce the potential for cell-mediated toxicity. In addition, the recent development of anti-chaperone therapy opens a new therapeutic avenue which is unlikely to be associated with toxicity.     

Natural human antibodies to amyloid beta peptide

Properties of human, natural anti-Abeta antibodies and commercially available intravenous immunoglobulin (IVIg) have been examined in light of the beneficial effects of passive immunotherapy with IVIg for patients with mild to moderate Alzheimer's disease (AD). Anti-Abeta antibodies in IVIg recognize conformation-specific epitopes as well as linear epitopes from different regions of the Abeta peptide. Anti-Abeta antibodies in circulation, especially those with high avidity, are often masked by ligands and the avidity of these antibodies increases upon dissociation of the bound ligands from the antibodies. Such natural anti-Abeta antibodies have the capacity to prevent Abeta oligomer-induced neurotoxicity in N2A neuroblastoma cells. This neuro-protective effect may reflect the therapeutic potential of the natural anti-Abeta antibodies found in IVIg for the treatment of patients with AD.     

Microglial overexpression of the M-CSF receptor augments phagocytosis of opsonized Abeta

The role of microglia in Alzheimer's disease (AD) has come under intense scrutiny recently because microglia may clear amyloid beta (Abeta) by phagocytosis after immunization of transgenic mice. Increased expression of the macrophage colony-stimulating factor receptor (M-CSFR) is an important feature of microglia in AD and transgenic mouse models for AD. Increased expression of M-CSFR on mouse and human microglia accelerates phagocytosis of aggregated Abeta in part through macrophage scavenger receptors. We now show that Abeta phagocytosis by microglia overexpressing M-CSFR is further enhanced by antibody opsonization of Abeta. M-CSFR overexpression increased microglial phagocytosis of opsonized aggregated Abeta in culture medium, and accelerated ingestion of native Abeta from AD brain sections. M-CSFR overexpression also increased microglial expression of Fcgamma receptors, and blocking Fcgamma receptors attenuated the enhanced Abeta uptake observed after M-CSFR overexpression and antibody opsonization. Microglia in AD and in AD mouse models with increased expression of M-CSFR are likely to rapidly ingest opsonized Abeta after immunization, making high intracerebral antibody titers unnecessary.     

Passive immunotherapy against Abeta in aged APP-transgenic mice reverses cognitive deficits and depletes parenchymal amyloid deposits in spite of increased vascular amyloid and microhemorrhage

BACKGROUND: Anti-Abeta immunotherapy in transgenic mice reduces both diffuse and compact amyloid deposits, improves memory function and clears early-stage phospho-tau aggregates. As most Alzheimer disease cases occur well past midlife, the current study examined adoptive transfer of anti-Abeta antibodies to 19- and 23-month old APP-transgenic mice. METHODS: We investigated the effects of weekly anti-Abeta antibody treatment on radial-arm water-maze performance, parenchymal and vascular amyloid loads, and the presence of microhemorrhage in the brain. 19-month-old mice were treated for 1, 2 or 3 months while 23-month-old mice were treated for 5 months. Only the 23-month-old mice were subject to radial-arm water-maze testing. RESULTS: After 3 months of weekly injections, this passive immunization protocol completely reversed learning and memory deficits in these mice, a benefit that was undiminished after 5 months of treatment. Dramatic reductions of diffuse Abeta immunostaining and parenchymal Congophilic amyloid deposits were observed after five months, indicating that even well-established amyloid deposits are susceptible to immunotherapy. However, cerebral amyloid angiopathy increased substantially with immunotherapy, and some deposits were associated with microhemorrhage. Reanalysis of results collected from an earlier time-course study demonstrated that these increases in vascular deposits were dependent on the duration of immunotherapy. CONCLUSIONS: The cognitive benefits of passive immunotherapy persist in spite of the presence of vascular amyloid and small hemorrhages. These data suggest that clinical trials evaluating such treatments will require precautions to minimize potential adverse events associated with microhemorrhage.     

Detection of amyloid beta protein in the urine of Alzheimer's disease patients and healthy individuals

To seek for a new valid biomarker using non-invasive specimens for the diagnosis of Alzheimer's disease (AD) and mild cognitive impairment (MCI), we carried out the detection of amyloid beta (Abeta) protein in urine. Ten-millilitre urine samples were first sedimented with trichloroacetic acid, and the pellets were resuspended for further analysis by Western blotting with anti-Abeta antibody. The detection sensitivity of the method was 40pg/ml. Rates of subjects positive for monomeric Abeta according to their clinical dementia rating (CDR) were 11.1% for CDR 0, 62.5% for CDR 0.5, 83.3% for CDR 1, 54.5% for CDR 2 and 0% for CDR 3. A single Abeta band relative to the CDR score reflects an alteration in the production, solubility and clearance of Abeta in the brain. Thus, the method could be used as both a diagnostic and monitoring tool in assessing AD and MCI patients during disease-modifying therapies.