Racke MM, Boone LI, Hepburn DL, Parsadainian M, Bryan MT, Ness DK, Piroozi KS, Jordan WH, Brown DD, Hoffman WP, Holtzman DM, Bales KR, Gitter BD, May PC, Paul SM, Demattos RB. Exacerbation of cerebral amyloid angiopathy-associated microhemorrhage in amyloid precursor protein transgenic mice by immunotherapy is dependent on antibody recognition of deposited forms of amyloid beta. J Neurosci. 2005 Jan 19;25(3):629-36. PubMed.
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Comments
Massachusetts General Hospital
This very nice study confirms the finding of Matthias Jucker and colleagues [1] that anti-Aβ antibody treatment can provoke CAA-related hemorrhage, in particular, antibodies that bind to deposited rather than soluble Aβ. The mechanism remains unclear; a logical possibility is that the same mechanisms that clear Aβ deposits can also "punch holes" in the amyloid-laden vessel wall.
For now, immunization-related hemorrhage remains a phenomenon of transgenic mice rather than human disease. Hemorrhagic stroke was not reported in the Elan-Wyeth vaccine studies [2], and the microhemorrhages seen on pathological examination of these brains [3,4] appear related to the underlying CAA rather than the vaccine itself. Human CAA may instead respond to inflammation by vascular dysfunction and reversible white matter changes [5]. Clearly, much remains to be learned about the role of CAA in determining the safety and efficacy of immune-based therapies for AD.
References:
Pfeifer M, Boncristiano S, Bondolfi L, Stalder A, Deller T, Staufenbiel M, Mathews PM, Jucker M. Cerebral hemorrhage after passive anti-Abeta immunotherapy. Science. 2002 Nov 15;298(5597):1379. PubMed.
Orgogozo JM, Gilman S, Dartigues JF, Laurent B, Puel M, Kirby LC, Jouanny P, Dubois B, Eisner L, Flitman S, Michel BF, Boada M, Frank A, Hock C. Subacute meningoencephalitis in a subset of patients with AD after Abeta42 immunization. Neurology. 2003 Jul 8;61(1):46-54. PubMed.
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. PubMed.
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. PubMed.
Eng JA, Frosch MP, Choi K, Rebeck GW, Greenberg SM. Clinical manifestations of cerebral amyloid angiopathy-related inflammation. Ann Neurol. 2004 Feb;55(2):250-6. PubMed.
Michigan State University
This article points out an important consideration with respect to passive immunotherapy against Aβ and its possible application to Alzheimer patients, namely, the increased risk for vascular hemorrhage with N-terminal-specific antibodies. Equally important, Racke et al. demonstrate that a mid-domain anti-Aβ antibody, which fails to decorate amyloid deposits in brain, does not increase this hemorrhage risk.
Coupled with the observation of Pfeifer et al. (2002) using N-terminal-specific antibodies in old APP23 mice, and our recent observations with C-terminal antibodies in old Tg2576 mice (Wilcock et al., 2004), this may be a common sequel of antibody-mediated amyloid removal. Our work also found 90 percent reduction in parenchymal, but fourfold elevation of vascular amyloid deposits with antibody therapy, implying a redistribution of the material.
A number of important questions need to be addressed. First, given the observation of Hock et al., (2003), that the only patients in the truncated Elan trial with a cognitive benefit had antibodies that decorated brain amyloid deposits, it may be that antibodies binding soluble Aβ, such as m266, may not benefit cognition. However, if the risk of hemorrhage is increased with antibodies against deposited Aβ, that will cause a serious dilemma regarding any form of Aβ immunotherapy.
A second issue regards the possibility that the increased microhemorrhage may apply to any therapy clearing Aβ. If our observation that vascular amyloid increases generalizes to other agents clearing preexisting plaques (e.g., plaque busters, zinc chelators), then these agents may have the same consequences.
A third question is if the problem is primarily due to the rapid rate of amyloid removal by these agents. The dose of antibody used by Racke et al. was 3-5 times higher than that used by others before (50 mg/kg). If the problem is saturation of efflux mechanisms at the vessels and resultant buildup of vascular amyloid, then a lower dose and more protracted exposure may reduce the amyloid loads without a concomitant rise in vascular deposits and increased risk of hemorrhage. Interestingly, one feature of the Dutch mutation in the Aβ peptide is a dramatic reduction in vascular efflux (Monro et al., 2002). While some of these issues are tractable in mouse models, others can only be addressed by carefully controlled trials in AD patients. Such trials are already in progress. Hopefully, detailed work in mouse models educated by the results of human trials will permit development of safe and effective Aβ immunotherapy.
References:
Pfeifer M, Boncristiano S, Bondolfi L, Stalder A, Deller T, Staufenbiel M, Mathews PM, Jucker M. Cerebral hemorrhage after passive anti-Abeta immunotherapy. Science. 2002 Nov 15;298(5597):1379. PubMed.
Wilcock DM, Rojiani A, Rosenthal A, Subbarao S, Freeman MJ, Gordon MN, Morgan D. 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. J Neuroinflammation. 2004 Dec 8;1(1):24. PubMed.
Hock C, Konietzko U, Streffer JR, Tracy J, Signorell A, Müller-Tillmanns B, Lemke U, Henke K, Moritz E, Garcia E, Wollmer MA, Umbricht D, de Quervain DJ, Hofmann M, Maddalena A, Papassotiropoulos A, Nitsch RM. Antibodies against beta-amyloid slow cognitive decline in Alzheimer's disease. Neuron. 2003 May 22;38(4):547-54. PubMed.
Monro OR, Mackic JB, Yamada S, Segal MB, Ghiso J, Maurer C, Calero M, Frangione B, Zlokovic BV. Substitution at codon 22 reduces clearance of Alzheimer's amyloid-beta peptide from the cerebrospinal fluid and prevents its transport from the central nervous system into blood. Neurobiol Aging. 2002;23(3):405-12. PubMed.
New York University School of Medicine
The study by Racke and colleagues is the third report showing cerebral bleeding in different AD mouse models following passive immunization with monoclonal antibodies with high affinity for Aβ plaques and congophilic angiopathy (Pfeifer et al., 2002; Wilcock et al., 2004b; Racke et al., 2005). This effect was not observed by other investigators employing another anti-Aβ antibody administered intracerebroventricularly in the Tg2576 model (Chauhan and Siegel, 2003). Also, microhemorrhages have not been reported following active immunizations, although it is unlikely that this has been assessed in most studies. We have recently sampled mouse brain sections from Tg2576 mice immunized with our Aβ derivatives (Sigurdsson et al., 2001; Sigurdsson et al., 2004), and we have yet to detect any microhemorrhages, although many of the animals immunized with K6Aβ1-30 had high anti-Aβ titer (Sigurdsson et al., unpublished observation).
Racke and colleagues did not assess amyloid burden or behavior (Racke et al., 2005), whereas Pfeifer’s study resulted in a modest reduction in plaque burden (Pfeifer et al., 2002). Also, Wilcock et al. observed that while causing microhemorrhages, their antibody treatment improved cognition, reduced plaque burden, but increased congophilic angiopathy (Wilcock et al., 2004b). That increase may be related to plaque clearance via the vasculature. In the three autopsy studies from the AN-1796 trial, vascular amyloid remained while plaques appeared to have been cleared in certain brain regions (Nicoll et al., 2003; Ferrer et al., 2004; Masliah et al., 2005). Eli Lilly’s antibody, 266, which binds soluble Aβ but does not recognize plaques and did not produce bleeding, has previously been shown to reduce plaque burden (DeMattos et al., 2001) and also to improve cognition acutely without affecting amyloid burden (Dodart et al., 2002). Elan was not able to replicate the effect of 266 on amyloid burden (Seubert et al., 2003), questioning its long-term efficacy.
So which type of antibody do you pick for costly clinical development? The Zurich AN-1792 trial indicated that AD patients who developed antibodies against amyloid plaques showed less cognitive decline than did other patients. However, even if 266 may potentially be less efficacious than antibodies binding to Aβ aggregates, it seems appropriate to start therapy with that antibody if it appears to be safer. Although, if 266 needs to be given more often than other monoclonals to improve cognition, it may be more likely to elicit an anti-idiotypic response with subsequent vasculitis and glomerulonephritis. If that is ineffective, then antibodies with high affinity for Aβ fibrils could be tested.
What about the isotype? IgG2a recognizing amino acids 3 to 7 of Aβ has been shown to promote plaque clearance after intraperitoneal injection, but IgG1 or IgG2b against the same epitope were not effective (Bussiere et al., 2004). However, different anti-Aβ IgG1 has been shown to result in plaque clearance when given intracranially (Wilcock et al., 2003; Lombardo et al., 2003) or intraperitoneally (Pfeifer et al., 2002; Wilcock et al., 2004a). Preferably, all these issues should have been sorted out in animal studies by direct comparison of these different types of antibodies, but clinical trials have already started on at least one of these approaches (AAB-001 by Elan/Wyeth), which is understandable because no effective therapy exists.
In addition to monoclonals, clinical trials of ivIg therapy are ongoing. This preparation contains pooled human immunoglobulins including autoantibodies against Aβ. If those trials will improve cognition in AD patients, future studies will have to determine if that effect was caused by the known antiinflammatory effect of ivIg and/or by the autoantibodies against Aβ.
Alternatively, active immunization with Aβ derivatives may be the way to go. This approach should produce several isotypes of antibodies against different epitopes and perhaps conformations, as well. Our Aβ derivative vaccination approaches have all improved cognition while eliciting different antibody responses and having various effects on amyloid burden (Scholtzova et al., 2002; Sigurdsson et al., 2004), indicating that a modest immune response is sufficient to improve cognition in AD mouse models. Prior active and passive immunization studies have also observed cognitive improvements without obvious correlation with certain Aβ measurements (Janus et al., 2000; Morgan et al., 2000; Dodart et al., 2002; Kotilinek et al., 2002).
It is fair to say at this point that low molecular weight species of Aβ, including oligomers and perhaps monomers, are likely to affect cognition. However, although extensive plaque deposition is a defense mechanism to isolate excess Aβ that cannot be cleared, it is likely that it will eventually affect neuronal connectivity, and plaque-associated glial activation should also have some toxic effects. Any immunotherapy should, therefore, at least slow the progression of plaque deposition.
All these different approaches are still worth pursuing. Because a self-antigen is being targeted, the immunotherapy may have to be individually tailored as a regular drug treatment. For example, an Aβ-derived immunogen may be chosen based on haplotype screening to provide first a T cell-independent IgM response with antibodies recognizing different epitopes and conformations of Aβ, which may prove to be more efficacious than targeting a single entity. If ineffective, a different Aβ derivative could then be administered that would be predicted to elicit a slightly stronger immune response. This process could then be continued until cognition improves.
See also:
Scholtzova H, Wisniewski T, Ahlawat S, Watanabe M, Quartermain D, Frangione B, and Sigurdsson EM. Safety of potential vaccines for Alzheimer's disease. Society for Neuroscience Abstracts, 227.1.2002.
Seubert P, Games D, Khan K, Buttini M, Bard F, Guido T, Grajeda H, Barbour R, Nguyen M, Kling K, Vasquez N, Schenk D, Hagen M, and Eldridge J. Comparative efficacy of different immunotherapeutic approaches in reducing AD-like neuropathology. Society for Neuroscience Abstracts, 133.3, 2003.
References:
Bussière T, Bard F, Barbour R, Grajeda H, Guido T, Khan K, Schenk D, Games D, Seubert P, Buttini M. Morphological characterization of Thioflavin-S-positive amyloid plaques in transgenic Alzheimer mice and effect of passive Abeta immunotherapy on their clearance. Am J Pathol. 2004 Sep;165(3):987-95. PubMed.
Chauhan NB, Siegel GJ. Intracerebroventricular passive immunization with anti-Abeta antibody in Tg2576. J Neurosci Res. 2003 Oct 1;74(1):142-7. PubMed.
DeMattos RB, Bales KR, Cummins DJ, Dodart JC, Paul SM, Holtzman DM. Peripheral anti-A beta antibody alters CNS and plasma A beta clearance and decreases brain A beta burden in a mouse model of Alzheimer's disease. Proc Natl Acad Sci U S A. 2001 Jul 17;98(15):8850-5. Epub 2001 Jul 3 PubMed.
Dodart JC, Bales KR, Gannon KS, Greene SJ, DeMattos RB, Mathis C, DeLong CA, Wu S, Wu X, Holtzman DM, Paul SM. Immunization reverses memory deficits without reducing brain Abeta burden in Alzheimer's disease model. Nat Neurosci. 2002 May;5(5):452-7. PubMed.
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. PubMed.
Janus C, Pearson J, McLaurin J, Mathews PM, Jiang Y, Schmidt SD, Chishti MA, Horne P, Heslin D, French J, Mount HT, Nixon RA, Mercken M, Bergeron C, Fraser PE, St George-Hyslop P, Westaway D. A beta peptide immunization reduces behavioural impairment and plaques in a model of Alzheimer's disease. Nature. 2000 Dec 21-28;408(6815):979-82. PubMed.
Kotilinek LA, Bacskai B, Westerman M, Kawarabayashi T, Younkin L, Hyman BT, Younkin S, Ashe KH. Reversible memory loss in a mouse transgenic model of Alzheimer's disease. J Neurosci. 2002 Aug 1;22(15):6331-5. PubMed.
Lombardo JA, Stern EA, McLellan ME, Kajdasz ST, Hickey GA, Bacskai BJ, Hyman BT. Amyloid-beta antibody treatment leads to rapid normalization of plaque-induced neuritic alterations. J Neurosci. 2003 Nov 26;23(34):10879-83. PubMed.
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. PubMed.
Morgan D, Diamond DM, Gottschall PE, Ugen KE, Dickey C, Hardy J, Duff K, Jantzen P, DiCarlo G, Wilcock D, Connor K, Hatcher J, Hope C, Gordon M, Arendash GW. A beta peptide vaccination prevents memory loss in an animal model of Alzheimer's disease. Nature. 2000 Dec 21-28;408(6815):982-5. PubMed.
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. PubMed.
Pfeifer M, Boncristiano S, Bondolfi L, Stalder A, Deller T, Staufenbiel M, Mathews PM, Jucker M. Cerebral hemorrhage after passive anti-Abeta immunotherapy. Science. 2002 Nov 15;298(5597):1379. PubMed.
Racke MM, Boone LI, Hepburn DL, Parsadainian M, Bryan MT, Ness DK, Piroozi KS, Jordan WH, Brown DD, Hoffman WP, Holtzman DM, Bales KR, Gitter BD, May PC, Paul SM, Demattos RB. Exacerbation of cerebral amyloid angiopathy-associated microhemorrhage in amyloid precursor protein transgenic mice by immunotherapy is dependent on antibody recognition of deposited forms of amyloid beta. J Neurosci. 2005 Jan 19;25(3):629-36. PubMed.
Sigurdsson EM, Knudsen E, Asuni A, Fitzer-Attas C, Sage D, Quartermain D, Goni F, Frangione B, Wisniewski T. An attenuated immune response is sufficient to enhance cognition in an Alzheimer's disease mouse model immunized with amyloid-beta derivatives. J Neurosci. 2004 Jul 14;24(28):6277-82. PubMed.
Sigurdsson EM, Scholtzova H, Mehta PD, Frangione B, Wisniewski T. Immunization with a nontoxic/nonfibrillar amyloid-beta homologous peptide reduces Alzheimer's disease-associated pathology in transgenic mice. Am J Pathol. 2001 Aug;159(2):439-47. PubMed.
Wilcock DM, DiCarlo G, Henderson D, Jackson J, Clarke K, Ugen KE, Gordon MN, Morgan D. Intracranially administered anti-Abeta antibodies reduce beta-amyloid deposition by mechanisms both independent of and associated with microglial activation. J Neurosci. 2003 May 1;23(9):3745-51. PubMed.
Wilcock DM, Rojiani A, Rosenthal A, Levkowitz G, Subbarao S, Alamed J, Wilson D, Wilson N, Freeman MJ, Gordon MN, Morgan D. Passive amyloid immunotherapy clears amyloid and transiently activates microglia in a transgenic mouse model of amyloid deposition. J Neurosci. 2004 Jul 7;24(27):6144-51. PubMed.
Wilcock DM, Rojiani A, Rosenthal A, Subbarao S, Freeman MJ, Gordon MN, Morgan D. 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. J Neuroinflammation. 2004 Dec 8;1(1):24. PubMed.
University of California, Irvine
Are conformation-dependent antibodies the key to a safe and effective AD immunotherapy?
Although immunotherapy for AD has become very attractive and its effectiveness is being applauded, there are lingering safety concerns as scientists continue to work to develop an effective and safe therapy. The recent paper by Racke et al. [1] confirms and extends the results from Mathias Jucker’s group [2] that indicate that passive immunization results in an increase in cerebral hemorrhage in addition to reducing amyloid deposition. Some scientists criticized these results because of the lack of control antibodies, and it was proposed that the cerebral hemorrhage observed in response to passive immunization was related to the particular mouse model used (APP23) rather than the actual passive immunization.
To date, three different groups using three different transgenic mouse models and different antibodies showed that passive immunization itself caused increased cerebral hemorrhage. Interestingly, all of the antibodies associated with increased cerebral hemorrhage bind plaques at either the N-terminus [1,2] or C-terminus [2], while antibodies against the middle region of the Aβ sequence, which do not bind plaques, do not increase hemorrhage. The mid-region antibodies can be considered conformation-dependent in the sense that their epitope is buried within the insoluble fibril structure and is exposed in the soluble monomer. Importantly, in fibrils, both the N-terminus (1-13) and C-terminus (39-42) are exposed at the surface of the fibril, while the middle region of the sequence is buried [3,4]. Therefore, m266 and 4G8 (which bind at approximately the same region, residues 13-28), which appear to be conformation-specific (their epitopes are buried within fibrils), may be the solution, since they would not bind amyloid fibril deposits.
The results of Racke et al. also indicate that in order for an active vaccine to be both safe and effective, it would have to lead to the production of antibodies that do not bind to insoluble amyloid deposits. Conformation-dependent antibodies may provide a solution to this problem [5,6]. Vaccination with a molecular mimic of amyloid oligomers gives rise to conformation-dependent antibodies that are specific for amyloid oligomers, which represent an intermediate in the fibril formation pathway and may be the primary toxic species of amyloids [7]. This oligomer-specific antibody does not bind extensively to insoluble amyloid deposits in AD brain; rather, it recognizes a much more restricted subset of soluble oligomers [7], suggesting that it would not cause problems with cerebral hemorrhage. In addition, this antibody neutralizes the toxicity of amyloid oligomers, which may represent a therapeutic benefit beyond removing amyloid deposits. Another attractive feature of conformation-dependent antibodies, such as the oligomer-specific antibody, is that they recognize an epitope that is specifically associated with pathogenesis. They do not recognize the normal, native protein structure. This suggests that anti-oligomer antibodies would be less likely to cause autoimmune complications.
Overall, the results of this paper make it appear increasingly likely that antibodies that bind to amyloid deposits (whether passive or active) may be more likely to have problems with cerebral hemorrhage than are other antibodies. The solution may be in conformation-dependent antibodies that specifically target restricted epitopes. It is still too early to judge the therapeutic benefits of conformation-specific antibodies. At this point, we think these antibodies represent an attractive opportunity to develop an effective immunotherapy for AD with a lower risk of hemorrhage or inflammatory complications.
References:
Racke MM, Boone LI, Hepburn DL, Parsadainian M, Bryan MT, Ness DK, Piroozi KS, Jordan WH, Brown DD, Hoffman WP, Holtzman DM, Bales KR, Gitter BD, May PC, Paul SM, Demattos RB. Exacerbation of cerebral amyloid angiopathy-associated microhemorrhage in amyloid precursor protein transgenic mice by immunotherapy is dependent on antibody recognition of deposited forms of amyloid beta. J Neurosci. 2005 Jan 19;25(3):629-36. PubMed.
Pfeifer M, Boncristiano S, Bondolfi L, Stalder A, Deller T, Staufenbiel M, Mathews PM, Jucker M. Cerebral hemorrhage after passive anti-Abeta immunotherapy. Science. 2002 Nov 15;298(5597):1379. PubMed.
Török M, Milton S, Kayed R, Wu P, McIntire T, Glabe CG, Langen R. Structural and dynamic features of Alzheimer's Abeta peptide in amyloid fibrils studied by site-directed spin labeling. J Biol Chem. 2002 Oct 25;277(43):40810-5. PubMed.
Kheterpal I, Williams A, Murphy C, Bledsoe B, Wetzel R. Structural features of the Abeta amyloid fibril elucidated by limited proteolysis. Biochemistry. 2001 Oct 2;40(39):11757-67. PubMed.
Dumoulin M, Dobson CM. Probing the origins, diagnosis and treatment of amyloid diseases using antibodies. Biochimie. 2004 Sep-Oct;86(9-10):589-600. PubMed.
Glabe CG. Conformation-dependent antibodies target diseases of protein misfolding. Trends Biochem Sci. 2004 Oct;29(10):542-7. PubMed.
Kayed R, Sokolov Y, Edmonds B, McIntire TM, Milton SC, Hall JE, Glabe CG. Permeabilization of lipid bilayers is a common conformation-dependent activity of soluble amyloid oligomers in protein misfolding diseases. J Biol Chem. 2004 Nov 5;279(45):46363-6. PubMed.
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