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Disease modifying therapy by targeting generic protein secondary structure of pathological oligomers at any stages of Alzheimer's disease models [Meeting Abstract]

Goni, F; Herline, K; Marta-Ariza, M; Prelli, F; Wisniewski, T
Background: We have previously demonstrated the feasibility of eliciting a unique antibody response independent of the primary and tertiary structure of proteins/peptides and specifically recognizing generic secondary beta-sheet structure dominant on misfolded proteins of oligomeric form; a hallmark of toxic prion-like "infectious" pathology in neurodegenerative diseases and specifically in the dual Abeta and tau Alzheimer's disease (AD) (Goni et al 2013 J. Neuroinflammation). As a putative antigen we have used a small peptide with a non-self primary structure, only beta-sheet secondary structure and no tertiary structure do to its size. The highly polymerized peptide (pBri) preserved a stable 90% beta-sheet secondary structure without any other outstanding epitope that could be recognized by an immune system. Young animals of AD models APP/PS1, 3xTg and SwDI as well as wild type animals, all developed polyclonal antibodies IgM and IgG that recognized oligomers of Abeta and tau and significantly ameliorated pathology. CD-1 animals were used to produced monoclonal antibodies (mAbs) that proved unequivocally the recognition was not to a sequence dependent epitope but to a generic beta-sheet secondary structure of pathologic oligomers but not fibrils or native proteins (Goni et al 2017 Scientific reports). The mAbs were characterized as specific anti-beta-sheet secondary structure monoclonal antibodies (AbetaComAbs). Either the stable IgM or the engineered IgG AbetaComAbs could reverse pathology and produce cognitive rescue in 3xTg animals (Goni et al 2018, Alz Res Therapy; Herline et al 2018 Alz Res Therapy).
Objective(s): To demonstrate that small engineered peptides with similar beta-sheet dominant structure but different primary sequences would elicit similar anti-beta-sheet responses and ameliorate pathology on 3xTg animals. To demonstrate that old 3xTg animals with already flourished pathology would develop the same type of anti-beta-sheet polyclonal IgM and IgG response and compared to AbetaComAbs infused animals produced comparable cognitive rescue and reduced oligomeric pathology. Finally, to demonstrate the specific reaction to the secondary structure but not the primary/tertiary structure does not produce antibody dependent cerebral microhemorrhages in an old SwDI CAA model of AD.
Method(s): Groups of 8 to 12 male and female 16 months old 3xTg AD and SwDI animals were inoculated i.p. weekly for 9 weeks with either 100 mug of the stable IgMk GW-23B7 or the derived TWF9 IgG2ak AbetaComAbs in 100 muL of sterile saline or with 100 muL of vehicle alone. Other groups of young 3xTg were inoculated with several tailored made mutations of the original pBri; whereas groups of 17 months old 3xTg were inoculated with the highly polymerized original pBri. The pBri immunizations and the AbetaComAbs were repeated in aged 16 month old SwDI animals prone to develop cerebral amyloid angiopathy due to the nature of the model, exacerbated by eventual reaction to Abeta amyloid epitopes existing in vessels of the brain. All animals were subjected to radial arm maze, locomotor tests or Barnes maze after the treatment and before sacrifice. Histochemical and biochemical analysis were performed on brains from treated and control animals.
Result(s): No adverse reactions were demonstrated during any one of the different treatments. Old animals inoculated with the pBri produced a fare anti-beta-sheet response; however, lower than in young animals. There was a significant cognitive rescue compared to controls and a significant reduction of both Abeta and tau oligomers. The mutated peptides were not as efficient to elicit anti-beta-sheet polyclonal response but still produce a significant amelioration on cognitive decline close to the results obtained by the original pBri. All SwDI animals either pBri immunized or AbetaComAbs infused -stable IgMk or IgG2aK- did not show any signs of vascular complication. In all cases the main biochemical improvement was the significant decrease on the number of oligomers of both Abeta and tau as assessed by specific blots of solubilized brain extractions, ELISAs and MSD measurements. Immunohistochemically the extracellular Abeta in plaques was significantly decreased whereas the intracellular PHF-tau remained fairly the same.
Conclusion(s): The development of a specific antibody response -polyclonal or monoclonal- to strictly the secondary structure of a protein or peptide frozen in a stable beta-sheet state is successfully achieved by the immunization with a highly polymerized, beta-sheet only, immunogen with no other visible epitope. That specific response is completely independent of the primary structure of pathological conformers; thus, it interferes only with oligomeric pathologic conformers that show the dominant beta-sheet secondary structure. Either treatment can produce , in various degrees, amelioration of AD pathology or evident cognitive rescue depending on the time of start of treatment. The mechanism is likely related to reductions of the levels of soluble oligomeric forms of Abeta and Tau; the species most closely linked to cognitive deficits in AD patients and the prionlike propagation. These results are extremely encouraging for the further testing of potential combinations of immunizations and AbetaComAbs in clinical trials with disease modifying potential and minimal risk of autoimmune complications
ISSN: 2426-0266
CID: 3900672

Immunotherapy to improve cognition and reduce pathological species in an Alzheimer's disease mouse model

Herline, Krystal; Prelli, Frances; Mehta, Pankaj; MacMurray, Claire; Goñi, Fernando; Wisniewski, Thomas
BACKGROUND:Alzheimer's disease (AD) is characterized by physiologically endogenous proteins amyloid beta (Aβ) and tau undergoing a conformational change and accumulating as soluble oligomers and insoluble aggregates. Tau and Aβ soluble oligomers, which contain extensive β-sheet secondary structure, are thought to be the most toxic forms. The objective of this study was to determine the ability of TWF9, an anti-β-sheet conformation antibody (aβComAb), to selectively recognize pathological Aβ and phosphorylated tau in AD human tissue compared with cognitively normal age-matched controls and to improve the performance of old 3xTg-AD mice with advanced pathology in behavioral testing after acute treatment with TWF9. METHODS:In this study, we used immunohistochemistry, immunoprecipitation, and enzyme-linked immunosorbent assay (ELISA) to characterize TWF9 specificity. We further assessed cognitive performance in old (18-22 months) 3xTg-AD mice using both a Barnes maze and novel object recognition after intraperitoneal administration of TWF9 (4 mg/kg) biweekly for 2 weeks before the start of behavioral testing. Injections continued for the duration of the behavioral testing, which lasted 2 weeks. RESULTS:Histological analysis of TWF9 in formalin-fixed paraffin-embedded human control and AD (ABC score: A3B3C3) brain tissue revealed preferential cytoplasmic immunoreactivity in neurons in the AD tissue compared with controls (p < 0.05). Furthermore, ELISA using oligomeric and monomeric Aβ showed a preferential affinity for oligomeric Aβ. Immunoprecipitation studies showed that TWF9 extracted both phosphorylated tau (p < 0.01) and Aβ (p < 0.01) from fresh frozen brain tissues. Results show that treated old 3xTg-AD mice have an enhanced novel object recognition memory (p < 0.01) and Barnes maze performance (p = 0.05) compared with control animals. Overall plaque burden, neurofibrillary tangles, microgliosis, and astrocytosis remained unchanged. Soluble phosphorylated tau was significantly reduced in TWF9-treated mice (p < 0.05), and there was a trend for a reduction in soluble Aβ levels in the brain homogenates of female 3xTg-AD mice (p = 0.06). CONCLUSIONS:This study shows that acute treatment with an aβComAb can effectively improve performance in behavioral testing without reduction of amyloid plaque burden, and that peripherally administered IgG can affect levels of pathological species in the brain.
PMID: 29914551
ISSN: 1758-9193
CID: 3157282

Anti-β-sheet conformation monoclonal antibody reduces tau and Aβ oligomer pathology in an Alzheimer's disease model

Goñi, Fernando; Martá-Ariza, Mitchell; Herline, Krystal; Peyser, Daniel; Boutajangout, Allal; Mehta, Pankaj; Drummond, Eleanor; Prelli, Frances; Wisniewski, Thomas
BACKGROUND:Oligomeric forms of amyloid-β (Aβ) and tau are increasing being recognized as key toxins in the pathogenesis of Alzheimer's disease (AD). METHODS:We developed a novel monoclonal antibody (mAb), GW-23B7, that recognizes β-sheet secondary structure on pathological oligomers of neurodegenerative diseases. RESULTS:The pentameric immunoglobulin M kappa chain (IgMκp) we developed specifically distinguishes intra- and extracellular pathology in human AD brains. Purified GW-23B7 showed a dissociation constant in the nanomolar range for oligomeric Aβ and did not bind monomeric Aβ. In enzyme-linked immunosorbent assays, it recognized oligomeric forms of both Aβ and hyperphosphorylated tau. Aged triple-transgenic AD mice with both Aβ and tau pathology infused intraperitoneally for 2 months showed IgMκp in the soluble brain homogenate, peaking at 24 h postinoculation. Treated mice exhibited significant cognitive rescue on radial arm maze testing compared with vehicle control-infused mice. Immunohistochemically, treatment resulted in a significant decrease of extracellular pathology. Biochemically, treatment resulted in significant reductions of oligomeric forms of Aβ and tau. CONCLUSIONS:These results suggest that GW-23B7, an anti-β-sheet conformational mAb humanized for clinical trials, may be an effective therapeutic agent for human AD.
PMID: 29378642
ISSN: 1758-9193
CID: 2933312

Potential Novel Approaches to Understand the Pathogenesis and Treat Alzheimer's Disease

Drummond, Eleanor; Goñi, Fernando; Liu, Shan; Prelli, Frances; Scholtzova, Henrieta; Wisniewski, Thomas
There is growing genetic and proteomic data highlighting the complexity of Alzheimer's disease (AD) pathogenesis. Greater use of unbiased "omics" approaches is being increasingly recognized as essential for the future development of effective AD research, that need to better reflect the multiple distinct pathway abnormalities that can drive AD pathology. The track record of success in AD clinical trials thus far has been very poor. In part, this high failure rate has been related to the premature translation of highly successful results in animal models that mirror only limited aspects of AD pathology to humans. We highlight our recent efforts to increase use of human tissue to gain a better understanding of the AD pathogenesis subtype variety and to develop several distinct therapeutic approaches tailored to address this diversity. These therapeutic approaches include the blocking of the Aβ/apoE interaction, stimulation of innate immunity, and the simultaneous blocking of Aβ/tau oligomer toxicity. We believe that future successful therapeutic approaches will need to be combined to better reflect the complexity of the abnormal pathways triggered in AD pathogenesis.
PMID: 29562516
ISSN: 1875-8908
CID: 3001492

Vaccination strategies

Wisniewski, Thomas; Goñi, Fernando
Currently all prion diseases are without effective treatment and are universally fatal. It is increasingly being recognized that the pathogenesis of many neurodegenerative diseases, such as Alzheimer disease (AD), includes "prion-like" properties. Hence, any effective therapeutic intervention for prion disease could have significant implications for other neurodegenerative diseases. Conversely, therapies that are effective in AD might also be therapeutically beneficial for prion disease. AD-like prion disease has no effective therapy. However, various vaccine and immunomodulatory approaches have shown great success in animal models of AD, with numerous ongoing clinical trials of these potential immunotherapies. More limited evidence suggests that immunotherapies may be effective in prion models and in naturally occurring prion disease. In particular, experimental data suggest that mucosal vaccination against prions can be effective for protection against orally acquired prion infection. Many prion diseases, including natural sheep scrapie, bovine spongiform encephalopathy, chronic wasting disease, and variant Creutzfeldt-Jakob disease, are thought to be acquired peripherally, mainly by oral exposure. Mucosal vaccination would be most applicable to this form of transmission. In this chapter we review various immunologically based strategies which are under development for prion infection.
PMID: 29887149
ISSN: 0072-9752
CID: 3154922

Production of Monoclonal Antibodies to Pathologic beta-sheet Oligomeric Conformers in Neurodegenerative Diseases

Goni, Fernando; Marta-Ariza, Mitchell; Peyser, Daniel; Herline, Krystal; Wisniewski, Thomas
We describe a novel approach to produce conformational monoclonal antibodies selected to specifically react with the beta-sheet secondary structure of pathological oligomeric conformers, characteristic of many neurodegenerative diseases. Contrary to past and current efforts, we utilize a mammalian non-self-antigen as an immunogen. The small, non-self peptide selected was covalently polymerized with glutaraldehyde until it reached a high beta-sheet secondary structure content, and species between 10-100kDa that are immunogenic, stable and soluble (p13Bri). Inoculation of p13Bri in mice elicited antibodies to the peptide and the beta-sheet secondary structure conformation. Hybridomas were produced and clones selected for their reactivity with at least two different oligomeric conformers from Alzheimer's, Parkinson and/or Prion diseases. The resulting conformational monoclonals are able to detect pathological oligomeric forms in different human neurodegenerative diseases by ELISA, immunohistochemistry and immunoblots. This technological approach may be useful to develop tools for detection, monitoring and treatment of multiple misfolding disorders.
PMID: 28852189
ISSN: 2045-2322
CID: 2679032

Targeting Pathological Proteins in Alzheimer's Disease [Meeting Abstract]

Herline, Krystal; Goni, Fernando; Wisniewski, Thomas
ISSN: 1554-6578
CID: 2645092

Dual Role of Inorganic Polyphosphate (POLYP) in the Regulation of Mitochondria-Dependent Cell Death [Meeting Abstract]

Torregrosa, Maria de la Encarnacion Solesio; Marta-Ariza, Mitchell; Goni, Fernando; Pavlov, Evgeny V
ISSN: 1542-0086
CID: 2597582

Anti-oligomer immunotherapy to improve cognition and reduce pathological species in a mouse model of both amyloid and tau pathology [Meeting Abstract]

Herline, K; Goni, F; Wisniewski, K
Aims The objective for this study was to determine the ability of an anti-oligomer antibody to improve performance using behavioral testing and to reduce pathological species with acute treatment in 3xTg- AD mice. Method Cognitive performance in old (18-22 months) 3xTg mice was assessed with both radial arm maze and novel object recognition after intraperitoneal administration of 100 micrograms of anti-pathological conformation IgG biweekly for two weeks before start of behavioral testing. Injections continued for the duration of the behavioral testing. Control mice received saline injections. Results Results show treated old 3xTg mice have a significantly enhanced novel object recognition memory compared to vehicle treated mice. Histological analysis in the hippocampus was performed to analyze plaque burden, phosphorylated tau, and astrocytosis. Overall plaque burden, phosphorylated tau, and astrocytosis remained unchanged. However, an additional histological analysis of plaque burden was performed using an antibody specific for the serine 8 phosphorylation site on amyloid beta. This analysis revealed a lower serine 8 modified amyloid beta burden in plaques of treated mice. Preliminary data reveals that phosphorylated amyloid beta burden correlates with behavioral performance in radial arm maze. Biochemical analysis of amyloid oligomers and phosphorylated tau are on-going. Conclusion This study shows 1) that acute treatment with an anti-oligomeric antibody can effectively rescue performance in behavioral testing without needing to affect overall amyloid plaque burden 2) and that this peripherally administered IgG can affect levels of pathological species in the brain. Overall, this IgG antibody has potential for being used as a therapeutic intervention for AD
ISSN: 1660-2862
CID: 2554352

Anti-conformation monoclonal antibody effective in pre-clinical treatment of full Alzheimer's disease animal models by targeting pathological oligomeric forms of A beta and modified Tau [Meeting Abstract]

Goni, F; Marta-Ariza, M; Herline, K; Boutajangout, A; Mehta, P; Drummond, E; Prelli, F; Wisniewski, T
ISSN: 1521-4141
CID: 2283602