Second, neither dogs nor mice/rats show much neuron loss with age. In AD the neuron loss leads to severe cognitive deterioration requiring institutionalization. Animals in this state would succumb (or, more humanely, be euthanized).

Third and most importantly, there is a critical difference in the amyloid deposits found in dogs versus Alzheimer humans and APP transgenic mice. They are not senile plaques, but only diffuse deposits (which the authors are careful to indicate in the text, although still referring to the deposits as plaques). The diffuse deposits do not have dystrophic neurites, microglial activation, astrocyte reactivity, or a host of other clear manifestations of pathology. ...  Read more

Second, neither dogs nor mice/rats show much neuron loss with age. In AD the neuron loss leads to severe cognitive deterioration requiring institutionalization. Animals in this state would succumb (or, more humanely, be euthanized).

Third and most importantly, there is a critical difference in the amyloid deposits found in dogs versus Alzheimer humans and APP transgenic mice. They are not senile plaques, but only diffuse deposits (which the authors are careful to indicate in the text, although still referring to the deposits as plaques). The diffuse deposits do not have dystrophic neurites, microglial activation, astrocyte reactivity, or a host of other clear manifestations of pathology. Diffuse deposits appear, histologically, to be quite benign. Hence it is conceivable that clearing them may have little impact on cognitive function, because they are relatively innocuous to begin with.

The absence of changes on the dot blots with A11 are intriguing, but it would be important to know whether this form of Aβ increases over the lifespan of the dogs and whether this measurement correlates with cognitive function in the dogs (it evidently did not). Lacking these data, it is difficult to put much interpretive zeal into the absence of an effect.

On the other hand, this may be one more piece of evidence to suggest that removing amyloid alone may not be sufficient to produce memory improvement in AD cases. Data reported by James Nicoll at the NYAS meeting last march showed that despite complete clearance of amyloid, patients from the Phase 1 trials of the Elan vaccine (who received 5-10 inoculations) had no cognitive benefits compared to control AD cases. Although the numbers are very small (two), this suggests that additional approaches, perhaps focusing on neuronal survival and/or tau pathology, may be essential to thwart progression of the disease.

We all anxiously await the results from the Phase 2 trial of bapineuzumab from Elan-Wyeth to inform us more regarding the role of amyloid in the cognitive symptoms of AD.

View all comments by Dave Morgan

We agree with Dr. Morgan that there is a positive correlation between the extent of diffuse Aβ and learning and memory error scores in aged dogs (Cummings, 1996; Head et al., 1998). The current study was originally designed to selectively reduce Aβ in order to test the hypothesis that Aβ causes cognitive dysfunction in aged dogs. It would appear then that reducing Aβ in dogs can have a cognitive benefit over the long term through maintenance of function. However, immediate effects of Aβ removal on cognition appear to be minimal. It is difficult to tease out with the current experimental design whether the cognitive impairments we see when Aβ is already present in old beagles is directly due to the Aβ or the consequence of Aβ being present for extended periods of time (in our case, Aβ can show up in canine prefrontal cortex as young as eight years of age, and thus may have been present in the...  Read more

We agree with Dr. Morgan that there is a positive correlation between the extent of diffuse Aβ and learning and memory error scores in aged dogs (Cummings, 1996; Head et al., 1998). The current study was originally designed to selectively reduce Aβ in order to test the hypothesis that Aβ causes cognitive dysfunction in aged dogs. It would appear then that reducing Aβ in dogs can have a cognitive benefit over the long term through maintenance of function. However, immediate effects of Aβ removal on cognition appear to be minimal. It is difficult to tease out with the current experimental design whether the cognitive impairments we see when Aβ is already present in old beagles is directly due to the Aβ or the consequence of Aβ being present for extended periods of time (in our case, Aβ can show up in canine prefrontal cortex as young as eight years of age, and thus may have been present in the study dogs for between one and four years (Head et al., 2000).

Alternatively, as Dr. Morgan points out, cognitive dysfunction in aged dogs may be due to events completely independent of Aβ. The perfect test of the Aβ hypothesis in the canine model is to prevent accumulation of Aβ rather than reverse it. It is also possible that if we combined immunotherapy with an intervention that restores neuron health, and the effects are truly additive, this outcome would provide further proof that Aβ causes both neuronal dysfunction and cognitive impairments.

As in all other animal models studied to date, there is not the overwhelming neuron loss as occurs in the AD brain. However, we would like to point out that there is evidence of select neuron loss in vulnerable brain regions in the canine. Aged canines exhibit hilar neuron loss in the hippocampus (Siwak-Tapp et al., 2006) and significant atrophy (prefrontal cortex and hippocampus) seen by MRI in association with Aβ and cognition in vulnerable brain regions (Tapp et al., 2004).

Given the apparent protective effects of immunotherapy and Aβ reduction on frontal function in canines, it is difficult to completely discount diffuse Aβ (without associated tau pathology and glial reaction) as being innocuous. Similar “less worsening” of clinical outcomes was reported in the AN1792 clinical trial (Gilman et al., 2005) along with Aβ reduction but with no effect on neurofibrillary tangle pathology (Nicoll et al., 2003; Ferrer et al., 2004; Masliah et al., 2005). Further, in middle-aged individuals with Down syndrome, a primary pathology is diffuse Aβ with neuritic pathology developing almost a decade later (Hof et al., 1995; Hyman, 1995). If diffuse Aβ is the precursor to downstream pathology (development of neuritic plaques, neurofibrillary tangles, neuron loss, etc.), or makes neurons vulnerable to further insults, then this is the time when we would want to intervene in human clinical trials.

With respect to our oligomer data, we concur with Dr. Morgan. We have some preliminary evidence to suggest that Aβ oligomers, specifically, do increase with age in the canine temporal cortex but this is obviously an area that needs further work. Measuring oligomers in a system that does not overexpress APP and accumulates endogenous levels of Aβ can be a challenge, although not impossible.

As is Dr. Morgan, we are also looking forward to what are hopefully going to be positive outcomes from the new clinical trials with immunotherapy. Immunotherapy still is one of the most promising approaches for slowing or potentially halting AD progression. Our work using the canine model illustrates the need to test Aβ-reducing therapies in different animal model systems, each with their own unique advantages, including those that do not overexpress mutant human APP (such as non-human primates), as the outcomes may provide some novel insights into the development of future treatments.

References:
Cummings BJ, Head E, Afagh AJ, Milgram NW, Cotman CW. Beta-amyloid accumulation correlates with cognitive dysfunction in the aged canine. Neurobiol Learn Mem. 1996 Jul;66(1):11-23. Abstract

Ferrer I, Boada Rovira M, Sánchez Guerra ML, Rey MJ, Costa-Jussá F. Neuropathology and pathogenesis of encephalitis following amyloid-beta immunization in Alzheimer's disease. Brain Pathol. 2004 Jan;14(1):11-20. Abstract

Gilman S, Koller M, Black RS, Jenkins L, Griffith SG, Fox NC, Eisner L, Kirby L, Rovira MB, Forette F, Orgogozo JM, AN1792(QS-21)-201 Study Team. Clinical effects of Abeta immunization (AN1792) in patients with AD in an interrupted trial. Neurology. 2005 May 10;64(9):1553-62. Abstract

Head E, Callahan H, Muggenburg BA, Cotman CW, Milgram NW. Visual-discrimination learning ability and beta-amyloid accumulation in the dog. Neurobiol Aging. 1998 Sep-Oct;19(5):415-25. Abstract

Head E, McCleary R, Hahn FF, Milgram NW, Cotman CW. Region-specific age at onset of beta-amyloid in dogs. Neurobiol Aging. 2000 Jan-Feb;21(1):89-96. Abstract

Hof PR, Bouras C, Perl DP, Sparks DL, Mehta N, Morrison JH. Age-related distribution of neuropathologic changes in the cerebral cortex of patients with Down's syndrome. Quantitative regional analysis and comparison with Alzheimer's disease. Arch Neurol. 1995 Apr;52(4):379-91. Abstract

Hyman BT, West HL, Rebeck GW, Lai F, Mann DM. Neuropathological changes in Down's syndrome hippocampal formation. Effect of age and apolipoprotein E genotype. Arch Neurol. 1995 Apr;52(4):373-8. Abstract

Masliah E, Hansen L, Adame A, Crews L, Bard F, Lee C, Seubert P, Games D, Kirby L, Schenk D. Abeta vaccination effects on plaque pathology in the absence of encephalitis in Alzheimer disease. Neurology. 2005 Jan 11;64(1):129-31. Abstract

Nicoll JA, Wilkinson D, Holmes C, Steart P, Markham H, Weller RO. Neuropathology of human Alzheimer disease after immunization with amyloid-beta peptide: a case report. Nat Med. 2003 Apr;9(4):448-52. Abstract

Siwak-Tapp CT, Head E, Muggenburg BA, Milgram NW, Cotman CW. Region specific neuron loss in the aged canine hippocampus is reduced by enrichment. Neurobiol Aging. 2008 Jan;29(1):39-50. Abstract

Tapp PD, Siwak CT, Gao FQ, Chiou JY, Black SE, Head E, Muggenburg BA, Cotman CW, Milgram NW, Su MY. Frontal lobe volume, function, and beta-amyloid pathology in a canine model of aging. J Neurosci. 2004 Sep 22;24(38):8205-13. Abstract

View all comments by David Cribbs
View all comments by Elizabeth Head

Amyloid-β: The Beginning of the End and the End of the Beginning
The paper by Head and colleagues (2008) should serve as a major warning for those hoping that removing amyloid will be effective in the treatment of Alzheimer disease (AD) or age-related cognitive impairment. So far, aside from transgenic mice engineered to overproduce amyloid, the record for amyloid immunotherapy has been one of abject failure: 1) trial suspension (human) and 2) no improvement (dogs). At minimum, these studies indicate that our current mouse models of AD are inadequate and, being amyloidocentric, naturally respond to amyloidocentric therapies. By contrast, rather than being driven by a transgene, the amyloid in aging or AD is there for a physiological or pathological reason (Nunomura et al., 2001), and removal of amyloid will not remove these precipitating factors. Moreover, we suspect that the production of amyloid, in response to a primary disease etiology, may...  Read more

Amyloid-β: The Beginning of the End and the End of the Beginning
The paper by Head and colleagues (2008) should serve as a major warning for those hoping that removing amyloid will be effective in the treatment of Alzheimer disease (AD) or age-related cognitive impairment. So far, aside from transgenic mice engineered to overproduce amyloid, the record for amyloid immunotherapy has been one of abject failure: 1) trial suspension (human) and 2) no improvement (dogs). At minimum, these studies indicate that our current mouse models of AD are inadequate and, being amyloidocentric, naturally respond to amyloidocentric therapies. By contrast, rather than being driven by a transgene, the amyloid in aging or AD is there for a physiological or pathological reason (Nunomura et al., 2001), and removal of amyloid will not remove these precipitating factors. Moreover, we suspect that the production of amyloid, in response to a primary disease etiology, may serve a beneficial function such that the removal of amyloid would be detrimental and exacerbate any underlying pathological processes (Perry et al., 2000; Rottkamp et al., 2002; Smith et al., 2002; Castellani et al., 2006; Lee et al., 2006a). We previously noted the existence of amyloid spin doctors (Castellani et al., 2007) and wonder whether the ever-increasing inconsistencies in the amyloid hypothesis (Lee et al., 2006b), together with recent therapeutic failures, will finally lead to the realization that it is time to stop spinning. Perhaps the time has come for this to be the beginning of the end for amyloid and the end of the beginning in our quest to understand and treat this disease.

References:
Castellani R, Lee HG, Perry G, Smith MA, Zhu X (2007) ARF Comment: Amyloid spin doctors.

Castellani RJ, Lee HG, Zhu X, Nunomura A, Perry G, Smith MA (2006) Neuropathology of Alzheimer disease: pathognomonic but not pathogenic. Acta Neuropathol (Berl) 111, 503-509. Abstract

Head E, Pop V, Vasilevko V, Hill M, Saing T, Sarsoza F, Nistor M, Christie LA, Milton S, Glabe C, Barrett E, Cribbs D (2008) A Two-Year Study with Fibrillar {beta}-Amyloid (A{beta}) Immunization in Aged Canines: Effects on Cognitive Function and Brain A{beta}. J Neurosci 28, 3555-3566. Abstract

Lee HG, Zhu X, Nunomura A, Perry G, Smith MA (2006a) Amyloid-beta vaccination: testing the amyloid hypothesis?: heads we win, tails you lose! Am J Pathol 169, 738-739. Abstract

Lee HG, Zhu X, Nunomura A, Perry G, Smith MA (2006b) Amyloid beta: the alternate hypothesis. Curr Alzheimer Res 3, 75-80. Abstract

Nunomura A, Perry G, Aliev G, Hirai K, Takeda A, Balraj EK, Jones PK, Ghanbari H, Wataya T, Shimohama S, Chiba S, Atwood CS, Petersen RB, Smith MA (2001) Oxidative damage is the earliest event in Alzheimer disease. J Neuropathol Exp Neurol 60, 759-767. Abstract

Perry G, Nunomura A, Raina AK, Smith MA (2000) Amyloid-beta junkies. Lancet 355, 757. Abstract

Rottkamp CA, Atwood CS, Joseph JA, Nunomura A, Perry G, Smith MA (2002) The state versus amyloid-beta: the trial of the most wanted criminal in Alzheimer disease. Peptides 23, 1333-1341. Abstract

Smith MA, Joseph JA, Atwood CS, Perry G (2002) Dangers of the amyloid-beta vaccination. Acta Neuropathol (Berl) 104, 110. Abstract

View all comments by Rudy Castellani
View all comments by Hyoung-gon Lee
View all comments by Akihiko Nunomura
View all comments by George Perry
View all comments by Mark A. Smith
View all comments by Xiongwei Zhu

Previously it was shown that the NTB composite z-scores were regressed on the geometric mean antibody titers (2) and the AN1792 vaccine reduced the number of Ab plaques, but not the level of soluble Aβ in brain (4). Interestingly, Head and coauthors reported that reduced levels of Aβ in the prefrontal cortex correlated with cognitive improvement, but vaccination did not reduce the binding of A11 antibodies to Aβ42 species in dot-blot assays.

Unfortunately, the authors did not show the levels of the different forms of Aβ42 (monomers, oligomers, and fibrils) by Western blot or combination of Western blot and immunoprecipitation, and did not demonstrate directly the association of these forms of β amyloid with memory impairment in aged beagles. This is important, because the lack of a significant therapeutic benefit of the AN1792 vaccine (2,5,6) may be connected to its inability to decrease the levels of soluble (oligomeric) Aβ in the brains (4). Recently the same kind of results were reported when a therapeutic vaccination strategy was tested in APP/Tg 2576 mice immunized with a novel peptide epitope vaccine (3).

In sum, data from this canine study, results generated with Aβ-immunotherapy of APP/Tg mice (3,7), and new data coming from AN1792 trials (4,8) and data reported by James Nicoll just this month at the NYAS meeting, 2008 suggest that early intervention in the disease process, pre-symptomatic if possible, is likely to be significantly more beneficial than attempting to intervene in the disease process after clinical diagnosis of the disease. Such a protective vaccination strategy may not only be more effective, but also safer than the therapeutic vaccination approach (both active and passive), because it could significantly reduce the probability of adverse events, such as microhemorrhages and subsequent possible infiltration of lymphocytes into the brain.

The rationale for focusing on the development of protective vaccine for treatment of pre-symptomatic people rather than AD patients is also based on the data demonstrating the pathologic correlates of mild cognitive impairment and early-stage AD. More specifically, Morris and Price (9) have reported that at the time dementia was only minimally apparent clinically, AD was already firmly established histopathologically. Moreover, they reported that entorhinal neuronal number, tissue volume and cortical synaptic integrity decreased notably by the time very mild AD is clinically expressed.

Recent breakthroughs in the development of biomarkers for AD provide hope that patients can be accurately identified while they are still in the preclinical stages of AD (10-13) which should facilitate delivery of new approaches, including anti-Aβ immunotherapy, before extensive neuronal damage and cerebral amyloid angiopathy has occurred in the brain.

References:
1. Head E, Barrett EG, Murphy MP, Das P, Nistor M, Sarsoza F, Glabe CC, Kayed R, Milton S, Vasilevko V, Milgram NW, Agadjanyan MG, Cribbs DH, Cotman CW. Immunization with fibrillar Abeta(1-42) in young and aged canines: Antibody generation and characteristics, and effects on CSF and brain Abeta. Vaccine. 2006 Apr 5;24(15):2824-34. Abstract

2. Gilman S, Koller M, Black RS, Jenkins L, Griffith SG, Fox NC, Eisner L, Kirby L, Rovira MB, Forette F, Orgogozo JM, AN1792(QS-21)-201 Study Team. Clinical effects of Abeta immunization (AN1792) in patients with AD in an interrupted trial. Neurology. 2005 May 10;64(9):1553-62. Abstract

3. Petrushina I, Ghochikyan A, Mktrichyan M, Mamikonyan G, Movsesyan N, Davtyan H, Patel A, Head E, Cribbs DH, Agadjanyan MG. Alzheimer's disease peptide epitope vaccine reduces insoluble but not soluble/oligomeric Abeta species in amyloid precursor protein transgenic mice. J Neurosci. 2007 Nov 14;27(46):12721-31. Abstract

4. Patton RL, Kalback WM, Esh CL, Kokjohn TA, Van Vickle GD, Luehrs DC, Kuo YM, Lopez J, Brune D, Ferrer I, Masliah E, Newel AJ, Beach TG, Castaño EM, Roher AE. Amyloid-beta peptide remnants in AN1792-immunized Alzheimer's disease patients: a biochemical analysis. Am J Pathol. 2006 Sep;169(3):1048-63. Abstract

5. Bayer AJ, Bullock R, Jones RW, Wilkinson D, Paterson KR, Jenkins L, Millais SB, Donoghue S. Evaluation of the safety and immunogenicity of synthetic Abeta42 (AN1792) in patients with AD. Neurology. 2005 Jan 11;64(1):94-101. Abstract

6. Fox NC, Black RS, Gilman S, Rossor MN, Griffith SG, Jenkins L, Koller M, AN1792(QS-21)-201 Study. Effects of Abeta immunization (AN1792) on MRI measures of cerebral volume in Alzheimer disease. Neurology. 2005 May 10;64(9):1563-72. Abstract

7. Lesné S, Koh MT, Kotilinek L, Kayed R, Glabe CG, Yang A, Gallagher M, Ashe KH. A specific amyloid-beta protein assembly in the brain impairs memory. Nature. 2006 Mar 16;440(7082):352-7. Abstract

8. Boche D, Nicoll JA. The role of the immune system in clearance of abeta from the brain. Brain Pathol. 2008 Apr;18(2):267-78. Abstract

9. Morris JC, Price AL. Pathologic correlates of nondemented aging, mild cognitive impairment, and early-stage Alzheimer's disease. J Mol Neurosci. 2001 Oct;17(2):101-18. Abstract

10. de Jong D, Jansen RW, Kremer BP, Verbeek MM. Cerebrospinal fluid amyloid beta42/phosphorylated tau ratio discriminates between Alzheimer's disease and vascular dementia. J Gerontol A Biol Sci Med Sci. 2006 Jul;61(7):755-8. Abstract

11. Fagan, A. M., Roe, C. M., Xiong, C., Mintun, M. A., Morris, J. C. & Holtzman, D. M. in 8th International Conference "Alzheimer's and Parkinson's Disease: Progress and New Perspectives" (Salzburg, Austria, 2007).

12. Fagan AM, Roe CM, Xiong C, Mintun MA, Morris JC, Holtzman DM. Cerebrospinal fluid tau/beta-amyloid(42) ratio as a prediction of cognitive decline in nondemented older adults. Arch Neurol. 2007 Mar;64(3):343-9. Abstract

13. Klunk WE, Engler H, Nordberg A, Wang Y, Blomqvist G, Holt DP, Bergström M, Savitcheva I, Huang GF, Estrada S, Ausén B, Debnath ML, Barletta J, Price JC, Sandell J, Lopresti BJ, Wall A, Koivisto P, Antoni G, Mathis CA, Långström B. Imaging brain amyloid in Alzheimer's disease with Pittsburgh Compound-B. Ann Neurol. 2004 Mar;55(3):306-19. Abstract

View all comments by Michael G. Agadjanyan

The intimate association of vascular disease and AD suggests that in many patients with early signs of dementia, amyloid disruption therapy may be complicated by impaired perfusion and drainage. Such conditions may explain the outcomes observed following the first human vaccination trial in which senile plaques were disrupted, but the amyloid remnants did not exit the brain. That makes the data of Nikolic et al. exciting, as these experiments suggest another means to actually eliminate and/or effectively alleviate the effects of toxic Aβ molecules.

Successful disruption of amyloid plaques will mitigate and perhaps eliminate the cascade of consequential destruction associated with these deposits. But great uncertainties remain regarding the ultimate risks posed by the chronic...  Read more

The intimate association of vascular disease and AD suggests that in many patients with early signs of dementia, amyloid disruption therapy may be complicated by impaired perfusion and drainage. Such conditions may explain the outcomes observed following the first human vaccination trial in which senile plaques were disrupted, but the amyloid remnants did not exit the brain. That makes the data of Nikolic et al. exciting, as these experiments suggest another means to actually eliminate and/or effectively alleviate the effects of toxic Aβ molecules.

Successful disruption of amyloid plaques will mitigate and perhaps eliminate the cascade of consequential destruction associated with these deposits. But great uncertainties remain regarding the ultimate risks posed by the chronic disturbance of the immobilized amyloid sequestered within plaques. The relative stress resilience of animal model and human neurons may be quite different, making the extrapolation of laboratory results to clinical outcomes such as improvements in cognitive status a challenging endeavor.

We cannot yet delineate precise pathological role(s) of Aβ in AD dementia, nor can we assign with certainty a normal-state function for this evolutionarily conserved molecule. Inducing or infusing high titers of antibody in the periphery to effect a CNS compartment change is still in an experimental stage. Present immunization trials employing this stratagem must be supported by further analyses beyond antibody titers and plaque removal.

View all comments by Tyler A. Kokjohn
View all comments by Alex E. Roher

In addition, Phase 2 clinical trials with Dimebon indicate that alternate drugs based on the non-amyloid theory have more success. I believe the field should stop animal experimentation for amyloid-based studies, especially the use of dogs, who should not be subjected to this fruitless research. Finally, requesting additional funding to continue research based on the "golden amyloid" theory appears to me to be wasting public money and blocking alternate...  Read more

In addition, Phase 2 clinical trials with Dimebon indicate that alternate drugs based on the non-amyloid theory have more success. I believe the field should stop animal experimentation for amyloid-based studies, especially the use of dogs, who should not be subjected to this fruitless research. Finally, requesting additional funding to continue research based on the "golden amyloid" theory appears to me to be wasting public money and blocking alternate research and clinical trials that are able to provide necessary tools to fight this devastating disease in the near future.

View all comments by Gjumrakch Aliev