Elina Zotova

Long-term effects of Aβ42 immunisation in Alzheimer's disease: follow-up of a randomised, placebo-controlled phase I trial

BACKGROUND Immunisation of patients with Alzheimers disease with full-length amyloid-beta peptide (Abeta(42)) can clear amyloid plaques from the brain. Our aim was to assess the relation between Abeta(42) immune response, degree of plaque removal, and long-term clinical outcomes. METHODS In June, 2003, consent for long-term clinical follow-up, post-mortem neuropathological examination, or both, was sought from 80 patients (or their carers) who had entered a phase I randomised, placebo-controlled trial of immunisation with Abeta(42) (AN1792, Elan Pharmaceuticals) in September, 2000. The follow-up study was completed in September, 2006. Plaques were assessed in terms of the percentage area of the cortex with Abeta immunostaining (Abeta load) and in terms of characteristic histological features reflecting plaque removal. Survival of all 80 individuals until severe dementia or death was assessed with a Cox proportional hazard model. FINDINGS 20 participants--15 in the AN1792 group, five in the placebo group--died before follow-up started. A further 22 patients--19 in the AN1792 group, three in the placebo group--died during follow-up. Nine of the deceased patients, all in the AN1792 group, had given consent for post-mortem analysis; one of these who did not die with Alzheimers disease was excluded. In the remaining eight participants who received immunisation and who were examined neuropathologically, mean Abeta load was lower than in an unimmunised control group that was matched for age at death (2.1% [SE 0.7] in treated participants vs 5.1% [0.9] in controls; mean difference 3.0%, 95% CI 0.6-5.4; p=0.02). Although there was considerable variation in Abeta load and degree of plaque removal among immunised participants, the degree of plaque removal varied significantly with mean antibody response attained during the treatment study period (Kruskal-Wallis p=0.02). Seven of the eight immunised patients who underwent post-mortem assessment, including those with virtually complete plaque removal, had severe end stage dementia before death. In the whole cohort, there was no evidence of improved survival (hazard ratio 0.93, 95% CI 0.43-3.11; p=0.86) or of an improvement in the time to severe dementia (1.18, 0.45-3.11; p=0.73) in the AN1792 group versus the placebo group. INTERPRETATION Although immunisation with Abeta(42) resulted in clearance of amyloid plaques in patients with Alzheimers disease, this clearance did not prevent progressive neurodegeneration.

Systemic inflammation and disease progression in Alzheimer disease

Background: Acute and chronic systemic inflammation are characterized by the systemic production of the proinflammatory cytokine tumor necrosis factor α (TNF-α) that plays a role in immune to brain communication. Previous preclinical research shows that acute systemic inflammation contributes to an exacerbation of neurodegeneration by activation of primed microglial cells. Objective: To determine whether acute episodes of systemic inflammation associated with increased TNF-α would be associated with long-term cognitive decline in a prospective cohort study of subjects with Alzheimer disease. Methods: Three hundred community-dwelling subjects with mild to severe Alzheimer disease were cognitively assessed, and a blood sample was taken for systemic inflammatory markers. Each subject’s main caregiver was interviewed to assess the presence of incident systemic inflammatory events. Assessments of both patient and caregiver were repeated at 2, 4, and 6 months. Results: Acute systemic inflammatory events, found in around half of all subjects, were associated with an increase in the serum levels of proinflammatory cytokine TNF-α and a 2-fold increase in the rate of cognitive decline over a 6-month period. High baseline levels of TNF-α were associated with a 4-fold increase in the rate of cognitive decline. Subjects who had low levels of serum TNF-α throughout the study showed no cognitive decline over the 6-month period. Conclusions: Both acute and chronic systemic inflammation, associated with increases in serum tumor necrosis factor α, is associated with an increase in cognitive decline in Alzheimer disease.

Consequence of Aβ immunization on the vasculature of human Alzheimer's disease brain

A major feature of Alzheimers disease is the accumulation of amyloid-beta peptide (Abeta) in the brain both in the form of plaques in the cerebral cortex and in blood vessel as cerebral amyloid angiopathy (CAA). Experimental models and human clinical trials have shown that accumulation of Abeta plaques can be reversed by immunotherapy. In this study, we hypothesized that Abeta in plaques is solubilized by antibodies generated by immunization and drains via the perivascular pathway, detectable as an increase in cerebrovascular Abeta. We have performed a follow up study of Alzheimers disease patients immunized against Abeta42. Neuropathological examination was performed on nine patients who died between four months and five years after their first immunization. Immunostaining for Abeta40 and Abeta42 was quantified and compared with that in unimmunized Alzheimers disease controls (n = 11). Overall, compared with these controls, the group of immunized patients had approximately 14 times as many blood vessels containing Abeta42 in the cerebral cortex (P<0.001) and seven times more in the leptomeninges (P = 0.013); among the affected blood vessels in the immunized cases, most of them had full thickness and full circumference involvement of the vessel wall in the cortex (P = 0.001), and in the leptomeninges (P = 0.015). There was also a significantly higher level of cerebrovascular Abeta40 in the immunized cases than in the unimmunized cases (cortex: P = 0.009 and leptomeninges: P = 0.002). In addition, the immunized patients showed a higher density of cortical microhaemorrhages and microvascular lesions than the unimmunized controls, though none had major CAA-related intracerebral haemorrhages. The changes in cerebral vascular Abeta load did not appear to substantially influence the structural proteins of the blood vessels. Unlike most of the immunized patients, two of the longest survivors, four to five years after first immunization, had virtually complete absence of both plaques and CAA, raising the possibility that, given time, Abeta is eventually cleared from the cerebral vasculature. The findings are consistent with the hypothesis that Abeta immunization results in solubilization of plaque Abeta42 which, at least in part, exits the brain via the perivascular pathway, causing a transient increase in the severity of CAA. The extent to which these vascular alterations following Abeta immunization in Alzheimers disease are reflected in changes in cognitive function remains to be determined.

Inflammation in Alzheimer's disease: relevance to pathogenesis and therapy

Evidence for the involvement of inflammatory processes in the pathogenesis of Alzheimers disease (AD) has been documented for a long time. However, the inflammation hypothesis in relation to AD pathology has emerged relatively recently. Even in this hypothesis, the inflammatory reaction is still considered to be a downstream effect of the accumulated proteins (amyloid beta (Aβ) and tau). This review aims to highlight the importance of the immune processes involved in AD pathogenesis based on the outcomes of the two major inflammation-relevant treatment strategies against AD developed and tested to date in animal studies and human clinical trials - the use of anti-inflammatory drugs and immunisation against Aβ.

Proinflammatory cytokines, sickness behavior, and Alzheimer disease.

Background: In Alzheimer disease (AD), systemic inflammation is known to give rise to a delirium. However, systemic inflammation also gives rise to other centrally mediated symptoms in the absence of a delirium, a concept known as sickness behavior. Systemic inflammation is characterized by the systemic production of the proinflammatory cytokines tumor necrosis factor–α (TNFα) and interleukin-6 (IL-6) that mediate immune to brain communication and the development of sickness behavior. Objective: To determine if raised serum TNFα or IL-6 are associated with the presence of sickness behavior symptoms, independent of the development of delirium, in a prospective cohort study of subjects with AD. Methods: A total of 300 subjects with mild to severe AD were cognitively assessed at baseline and a blood sample taken for inflammatory markers. Cognitive assessments, including assessments to detect the development of a delirium, and blood samples were repeated at 2, 4, and 6 months. The development of neuropsychiatric symptoms in the subject with AD over the 6-month follow-up period was assessed independently by carer interview at 2, 4, and 6 months. Results: Raised serum TNFα and IL-6, but not CRP, were associated with an approximately 2-fold increased frequency of neuropsychiatric symptoms characteristic of sickness behavior. These relationships are independent of the development of delirium. Conclusions: Increased serum proinflammatory cytokines are associated with the presence of symptoms characteristic of sickness behavior, which are common neuropsychiatric features found in AD. This association was independent of the presence of delirium.

Inflammatory components in human Alzheimer's disease and after active amyloid-β42 immunization

Inflammatory processes are important in the pathogenesis of Alzheimers disease and in response to amyloid-β immunotherapy. We investigated the expression of multiple inflammatory markers in the brains of 28 non-immunized patients with Alzheimers disease and 11 patients with Alzheimers disease immunized against amyloid-β42 (AN1792): microglial ionized calcium-binding adaptor Iba-1, lysosome marker CD68, macrophage scavenger receptor A, Fcγ receptors I (CD64) and II (CD32); and also immunoglobulin IgG, complement C1q and the T lymphocyte marker CD3 using immunohistochemistry. The data were analysed with regard to amyloid-β and phospho-tau pathology, severity of cerebral amyloid angiopathy and cortical microhaemorrhages. In non-immunized Alzheimers disease cases, amyloid-β42 correlated inversely with CD32 and Iba-1, whereas phospho-tau correlated directly with all microglial markers, IgG, C1q and the number of T cells. In immunized Alzheimers disease cases, amyloid-β42 load correlated directly with macrophage scavenger receptor A-positive clusters and inversely with C1q. The severity of cerebral amyloid angiopathy and microhaemorrhages did not relate to any of the analysed markers. Overall, the levels of CD68, macrophage scavenger receptor A, CD64, CD32 and the number of macrophage scavenger receptor A-positive plaque-related clusters were significantly lower in immunized than non-immunized cases, although there was no significant difference in Iba-1 load, number of Iba-1-positive cells, IgG load, C1q load or number of T cells. Our findings indicate that different microglial populations co-exist in the Alzheimers disease brain, and that the local inflammatory status within the grey matter is importantly linked with tau pathology. After amyloid-β immunization, the microglial functional state is altered in association with reduced amyloid-β and tau pathology. The results suggest that, in the long term, amyloid-β immunotherapy results in downregulation of microglial activation and potentially reduces the inflammation-mediated component of the neurodegeneration of Alzheimers disease.

Microglial alterations in human Alzheimer's disease following Aβ42 immunization.

E. Zotova, C. Holmes, D. Johnston, J. W. Neal, J. A. R. Nicoll and D. Boche (2011) Neuropathology and Applied Neurobiology37, 513–524

Proinflammatory cytokines and the clinical features of dementia with lewy bodies.

Dementia with Lewy bodies (DLB) accounts for up to 25% of all cases of dementia over the age of 65 years.1 It is characterized clinically by progressive fluctuating cognitive decline, parkinsonism, and visual hallucinations and also includes the neuropsychiatric features of sleep disorder, delusions, depression, and other hallucinations.2 Systemic inflammation is characterized by the production of the proinflammatory cytokines interleukin 6 (IL-6) and tumor necrosis factor-a (TNF-a) that activate the central innate immune response to initiate a behavioral response known as sickness behaviour3 that includes increased anxiety, depressed mood, apathy, sleep disturbance, and decreased appetite. In animal models of neurodegeneration, systemic inflammation has been shown to result in an exaggeration of sickness behavior and long term increased neuronal cell loss due to a markedly increased central proinflammatory cytokine profile caused by microglial priming.4 In Alzheimer Disease (AD) we have shown that modest increases in serum proinflammatory cytokines are associated with an exaggeration of a wide range of neuropsychiatric symptoms, particularly symptoms of sickness behavior and hallucinations, as well as increased cognitive impairment.5 Animal models of Parkinson Disease show evidence of microglia priming.6 We therefore hypothesized that DLB subjects, like AD subjects, would be sensitive to peripheral inflammation and show a significant relationship between serum levels of the proinflammatory cytokines (IL6 and TNF-a) and impaired cognition and an increased frequency of a range of neuropsychiatric symptoms, including features of sickness behavior and hallucinations. METHODS A total of 35 community dwelling subjects (and their caregivers) with mild to severe dementia, fulfilling consensus criteria2 for probable or possible DLB, were recruited from clinical referrals to memory assessment services in Southampton, UK from November 2003 to February 2008. After consent procedures, subjects were examined (and carers interviewed) to assess the presence of systemic inflammatory events (SIEs) in the previous 2 weeks and the presence of delirium using the Confusion Assessment Method.4,7 Carers were interviewed to assess the highest frequency and severity of behavioral symptoms since initial diagnosis using the Neuropsychiatric Inventory (NPI).8 Subjects were then cognitively tested using the Alzheimer’s Disease Assessment Scale Cognitive subscale (ADAS-COG)9 and a blood sample taken for the proinflammatory cytokines IL-6 and TNF-a. Serum IL-6 and TNF-a was assayed using sandwich immunoassay multispot cytokine assay (Meso Scale Discovery). The lowest detectable limit was 0.7 pg/mL for IL-6 and 1.1 pg/mL for TNF-a. The study received approval from the South and West Hampshire Local Research Ethics Committee (ref 07/ Q1704/78). Allowing for 10% dropout, 35 subjects gave 80% power to detect a modest correlation (0.5) between 4 sets of variables (a=0.0125). Normality of continuous variables was determined by quantile-quantile plots of the residuals. Relationships between variables were assessed using a mixture of Student t test, Mann-Whitney U (MWU) test, and Pearson and Spearman correlations. Data interaction was assessed using linear regression analysis.

Systemic inflammation and blood–brain barrier abnormality in relapsing–remitting multiple sclerosis

Abstract Background Systemic inflammation can affect disease expression in multiple sclerosis. The mechanism might involve blood–brain barrier disruption. We aimed to assess the effects of systemic inflammation on disease progression in multiple sclerosis and the role of blood–brain barrier disruption. Methods We recruited adults with relapsing–remitting multiple sclerosis and healthy controls from the general population. Three-dimensional dynamic-contrast enhanced MRI was used to measure blood–brain barrier permeability in the normal-appearing white matter (NAWM). Urinary neopterin, a product of activated macrophages, was measured to provide a readout of systemic inflammation. All study activities were performed in University Hospital Southampton after ethics approval (REC 12/SC/0176). Findings 12 patients with mutliple sclerosis and 12 healthy controls were recruited. Blood–brain barrier permeability in NAWM, measured as the constant Ktrans, was significantly higher in patients than in controls (mean 0·024 ml/100g per min [SD·0·018] vs 0·006 [0·004], p=0·015). Systemic inflammatory activity, measured as the urinary neopterin:creatinine ratio (UNCR), was also significantly higher (3·35 [1·98] vs 1·36 [0·29], p=0·002). Across all participants, there was a weak positive correlation between Ktrans and UNCR ( r =0·40, p=0·031). Interpretation Our findings support the hypothesis that the effects of systemic inflammation on expression of multiple sclerosis disease are mediated by blood–brain barrier disruption. Targeting this disruption and systemic inflammation might provide novel avenues for disease-modifying therapy. Funding University of Southampton, National Institute for Health Research, Multiple Sclerosis Society.

Continued progression of neurodegeneration in Alzheimer's disease despite complete removal of the Abeta plaques?

Introduction: When 3 kDa soluble fluorescent dextran or 40 kDa ovalbumin are injected into the mouse striatum, they spread diffusely through the brain parenchyma and rapidly drain out of the brain along perivascular basement membranes surrounding capillaries and arteries. In the present study we test the hypothesis that immune complexes formed in the brain disrupt the perivascular drainage of solutes. Material and methods: Immune complexes were formed in the brain by actively immunizing mice against ovalbumin and then injecting ovalbumin intracerebrally. Twenty-four hours later dextran tracer was injected into the brain at the same site and animals killed 5 min and 3 h later. Results: At 5 min and 3 h after injection of the dextran tracer into the brains of immunized mice, immune complexes were located in the walls of arteries (as shown by Teeling et al. at this meeting). There was a significant reduction in the diffuse spread of dextran in the brain parenchyma and significantly fewer capillary and artery basement membranes contained dextran at 5 min when compared with the nonimmunized controls. Instead, the dextran tracer was concentrated in the perivenous spaces that were occupied by inflammatory cells. Conclusions: The results suggest that immune complexes disrupt and alter the pattern of perivascular drainage of solutes from the brain. This may have significance for inflammatory diseases in the brain and for immunotherapy for Alzheimer’s disease in which immune complexes may form and further block the perivascular drainage pathways that is already compromised by the deposition of amyloidbeta. This work is supported by the Alzheimer Research Trust.