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Novel phospho-Tau specific monoclonal antibodies for the treatment of Tau-mediated pathology
S480
Poster Presentations P2
Background: Research over the last decade has highlighted the role that metals may play in the pathogenesis of Alzheimer’s disease (AD). To date, the primary end point for these studies has been ß-amyloid. Recent literature supports the notion that metals may also interact with tau and be important for the formation of neurofibrillary tangles (NFTs). In this study we surveyed metal levels in the rTg(tauP301L)4510 mouse model and utilised clioquinol (CQ) to assess the effect of metal modulation on tangle pathology. Methods: Metal levels were assessed by ICPMS in cohorts of mice at either w2 months (n ¼ 6 WT; n ¼ 6 Tg) or w5 months (n ¼ 12 WT; n ¼ 11 Tg) of age. The effect of oral clioquinol (6 months of treatment, 30mg/kg) was assessed in a separate cohort of Tg (n ¼ 25) and WT mice (n ¼ 30). Drug treatment began at w6 months of age and prior to culling animals were assessed in the open field, rotarod, y-maze and water maze. Young rTg4510 mice, prior to the formation of any pathology, had significantly altered brain metal content, including elevations in zinc (25%, p ¼ 0.01) and copper (31%, p ¼ 0.02) and a trend to elevated iron (20%, p ¼ 0.08). In contrast, aged rTg4510 mice had significantly decreased levels of zinc (18%, p ¼ 0.01) and iron levels were also decreased (20%, p ¼ 0.06). Behavioural analysis of older mice did not reveal any effect of CQ on performance in the water maze, Y-maze or open field. In the rotarod, however, the Tg animals spent a significantly shorter amount of time on the rotarod (3.5 sec, Tg; 35 sec, WT; p < 0.0001), and clioquinol treatment of the Tg animals significantly improved their performance (10.2 sec, Tg+clioquinol; p ¼ 0.01 compared to Tg). Further metal analyses and histological studies are currently being finalised and will be discussed. Conclusions: We have demonstrated that there is a dysregulation in metal ion homeostasis in the rTg4510 model of tauopathy, and that there are also age-dependent alterations in metal levels that occur coincident with the evolution of neurofibrillary pathology. These data support a potential interaction between tau/NFTs and various metal ions and provide further insight into novel therapeutic approaches for the treatment of tauopathies.
P2-525
NOVEL PHOSPHO-TAU SPECIFIC MONOCLONAL ANTIBODIES FOR THE TREATMENT OF TAUMEDIATED PATHOLOGY
Oskar Adolfsson1, Natalia Crespo-Biel2, Maria Pihlgren1, Clara Theunis2, Valerie Gafner1, Peter Borghgraef2, Maria Pilar Lopez Deber1, Herman Devijver2, David T. Hickman1, Anna Lucia Buccarello1, Nathalie Chuard1, 1AC Immune SA, Lausanne, Switzerland; 2KULeuven, Leuven, Belgium. Background: The brain of patients with Alzheimer’s disease (AD) is histopathologically characterized by two hallmarks, beta-amyloid plaques and neurofibrillar tangles (NFT) composed of hyper-phosphorylated Tau. Tau pathology in the absence of overt beta-amyloid pathology is characteristic of a subset of patients with frontotemporal dementia (FTD). Progression and spreading of tau pathology throughout the brain correlates with cognitive demise and disease progression in AD. Immunotherapy for various neurodegenerative diseases recently emerged as a promising approach for clearing pathological protein conformers, presumed to cause these disorders. Active immunization targeting phosphorylated Tau have raised phospho-Tau antibody responses, which reduced pathological Tau aggregation in brain of transgenic mice (Asuni et al., 2007). As a specific therapeutic approach we developed monoclonal antibodies directed against pathological phospho-epitopes of protein Tau (pTau). Methods: Liposome-based vaccines were generated to display synthetic pTau peptides, formulated similarly to previously described beta-amyloid peptide vaccines (Muhs et al., 2007). Mice, selected for the most specific polyclonal immune responses, were boosted and their splenocytes used to generate hybridomas by classical fusion techniques. Screening of IgG producing hybridomas identified monoclonal antibodies for further characterization. Binding specificity was verified by pTau specific ELISA, by affinity by SPR BiaCore and by specificity for human pTau using western-blot and dot-blot assays. Furthermore, the specificity for Tau pathology was demonstrated immunohistochemically
on brain sections of Tau transgenic mice and section from human tauopathy subjects. Results: Selected anti pTau IgG antibodies were identified with high binding affinity to pTau (down to Kd 0.12 nM) without appreciable reaction with the non-phosphorylated Tau peptides, and without non-specific binding or cross-reaction to different non-target phospho-epitopes. Biochemical analysis showed also specific binding to linearized and natively folded full-length, phosphorylated human pTau protein. On brain sections of tau. P301LxGSK3ß mice (biGT) with proven tauopathy (Terwel et al., 2008) the specificity of the Mabs for tauopathy was demonstrated, and moreover confirmed on brain sections from human subjects suffering from various with tauopathies including AD. Abundant labeling of neurofibrillary tangles and neuropil threads in hippocampus and cortex was evident. Conclusions: Phospho-Tau monoclonal antibodies specifically reacting with phosphorylated protein Tau were successfully derived and characterized. The data demonstrate that targeting pTau in combination with detection of aggregated Tau in NFT could be a promising therapeutic approach as indicated by the specificity and binding activity.
P2-526
RENAL FUNCTION OUTCOME AFTER LITHIUM THERAPY FOR COGNITIVE DECLINE
Ivan Aprahamian1, Breno Diniz2, Franklin Santos2, Marcia Radanovic2, Orestes Forlenza2, 1Psychogeriatric Unit, Laboratory of Neuroscience (LIM 27), Department and Institute of Psychiatry, S~ao Paulo, Brazil; 2Laboratory of Neuroscience - LIM 27, Department and Institute of Psychiatry Faculty of Medicine, University of S~ao Paulo, S~ao Paulo, Brazil. Background: Experimental studies conducted in neuronal culture and animal models provide evidence of the neurotrophic and neuroprotective properties of lithium, with alleged impact on the management of certain neurodegenerative conditions, namely Alzheimer’s disease (AD). Recent clinical trials support the feasibility and the potential benefits of this approach. However, chronic lithium use is associated with clinically relevant adverse effects, particularly renal dysfunction. Objective: To evaluate longterm safety of lithium therapy on parameters of renal function of elderly patients with mild cognitive impairment (MCI). Methods: Subjects included in this study are participants of a randomized clinical trial with lithium addressing disease-modification in the MCI-AD continuum. Study design: double blind, placebo controlled trial of lithium (N ¼ 22) versus placebo (N ¼ 22) for the treatment of patients with MCI for three years. Identical tablets holding 150, 300, 450 and 600 mg of lithium carbonate or placebo were administered orally (b.i.d.). Lithium doses were adjusted to keep serum levels between 0.2 and 0.5mEq/L. Safety and tolerability of treatments were longitudinally evaluated at three-month intervals by clinical examination, scores on the UKU side-effect scale, and laboratory tests addressing renal function (and other parameters not included in the present assessment). Statistical analysis was performed with SPSS 18. Results: Baseline serum concentrations of creatinine were not significantly different between the two groups at baseline (lithium, 0.84 6 0.20; placebo, 0.87 6 0.23; p ¼ 0.76), nor at the endpoint of three years (lithium, 0.93 6 0.27; placebo, 0.96 6 0.38; p ¼ 0.19). Kaplan-Meyer curves did not indicate significant changes in renal function according to treatment groups over time (p ¼ 0.31); intra-group comparisons (p ¼ 0.94) indicated long-term stability of such parameters upon follow-up. Conclusions: Low-dose, chronic lithium therapy appears to be clinically safe and not associated with significant changes in renal function in older adults with MCI. The present findings support the use of lithium salts as a candidate drug for disease-modification in AD. P2-527
TARGETING HYPERPHOSPHORYLATED TAU PROTEIN WITH A MONOCLONAL ANTIBODY AT AN ADVANCED STAGE OF TAU PATHOLOGY IMPROVES COGNITION IN A MOUSE MODEL
Allal Boutajangout1, Hameetha Banu Rajamohamed Sait1, Veronica Gonzalez1, Einar Sigurdsson1, 1NYU School of Medicine, New York, New York, United States.
Poster Presentations P2
Background: Research over the last decade has highlighted the role that metals may play in the pathogenesis of Alzheimer’s disease (AD). To date, the primary end point for these studies has been ß-amyloid. Recent literature supports the notion that metals may also interact with tau and be important for the formation of neurofibrillary tangles (NFTs). In this study we surveyed metal levels in the rTg(tauP301L)4510 mouse model and utilised clioquinol (CQ) to assess the effect of metal modulation on tangle pathology. Methods: Metal levels were assessed by ICPMS in cohorts of mice at either w2 months (n ¼ 6 WT; n ¼ 6 Tg) or w5 months (n ¼ 12 WT; n ¼ 11 Tg) of age. The effect of oral clioquinol (6 months of treatment, 30mg/kg) was assessed in a separate cohort of Tg (n ¼ 25) and WT mice (n ¼ 30). Drug treatment began at w6 months of age and prior to culling animals were assessed in the open field, rotarod, y-maze and water maze. Young rTg4510 mice, prior to the formation of any pathology, had significantly altered brain metal content, including elevations in zinc (25%, p ¼ 0.01) and copper (31%, p ¼ 0.02) and a trend to elevated iron (20%, p ¼ 0.08). In contrast, aged rTg4510 mice had significantly decreased levels of zinc (18%, p ¼ 0.01) and iron levels were also decreased (20%, p ¼ 0.06). Behavioural analysis of older mice did not reveal any effect of CQ on performance in the water maze, Y-maze or open field. In the rotarod, however, the Tg animals spent a significantly shorter amount of time on the rotarod (3.5 sec, Tg; 35 sec, WT; p < 0.0001), and clioquinol treatment of the Tg animals significantly improved their performance (10.2 sec, Tg+clioquinol; p ¼ 0.01 compared to Tg). Further metal analyses and histological studies are currently being finalised and will be discussed. Conclusions: We have demonstrated that there is a dysregulation in metal ion homeostasis in the rTg4510 model of tauopathy, and that there are also age-dependent alterations in metal levels that occur coincident with the evolution of neurofibrillary pathology. These data support a potential interaction between tau/NFTs and various metal ions and provide further insight into novel therapeutic approaches for the treatment of tauopathies.
P2-525
NOVEL PHOSPHO-TAU SPECIFIC MONOCLONAL ANTIBODIES FOR THE TREATMENT OF TAUMEDIATED PATHOLOGY
Oskar Adolfsson1, Natalia Crespo-Biel2, Maria Pihlgren1, Clara Theunis2, Valerie Gafner1, Peter Borghgraef2, Maria Pilar Lopez Deber1, Herman Devijver2, David T. Hickman1, Anna Lucia Buccarello1, Nathalie Chuard1, 1AC Immune SA, Lausanne, Switzerland; 2KULeuven, Leuven, Belgium. Background: The brain of patients with Alzheimer’s disease (AD) is histopathologically characterized by two hallmarks, beta-amyloid plaques and neurofibrillar tangles (NFT) composed of hyper-phosphorylated Tau. Tau pathology in the absence of overt beta-amyloid pathology is characteristic of a subset of patients with frontotemporal dementia (FTD). Progression and spreading of tau pathology throughout the brain correlates with cognitive demise and disease progression in AD. Immunotherapy for various neurodegenerative diseases recently emerged as a promising approach for clearing pathological protein conformers, presumed to cause these disorders. Active immunization targeting phosphorylated Tau have raised phospho-Tau antibody responses, which reduced pathological Tau aggregation in brain of transgenic mice (Asuni et al., 2007). As a specific therapeutic approach we developed monoclonal antibodies directed against pathological phospho-epitopes of protein Tau (pTau). Methods: Liposome-based vaccines were generated to display synthetic pTau peptides, formulated similarly to previously described beta-amyloid peptide vaccines (Muhs et al., 2007). Mice, selected for the most specific polyclonal immune responses, were boosted and their splenocytes used to generate hybridomas by classical fusion techniques. Screening of IgG producing hybridomas identified monoclonal antibodies for further characterization. Binding specificity was verified by pTau specific ELISA, by affinity by SPR BiaCore and by specificity for human pTau using western-blot and dot-blot assays. Furthermore, the specificity for Tau pathology was demonstrated immunohistochemically
on brain sections of Tau transgenic mice and section from human tauopathy subjects. Results: Selected anti pTau IgG antibodies were identified with high binding affinity to pTau (down to Kd 0.12 nM) without appreciable reaction with the non-phosphorylated Tau peptides, and without non-specific binding or cross-reaction to different non-target phospho-epitopes. Biochemical analysis showed also specific binding to linearized and natively folded full-length, phosphorylated human pTau protein. On brain sections of tau. P301LxGSK3ß mice (biGT) with proven tauopathy (Terwel et al., 2008) the specificity of the Mabs for tauopathy was demonstrated, and moreover confirmed on brain sections from human subjects suffering from various with tauopathies including AD. Abundant labeling of neurofibrillary tangles and neuropil threads in hippocampus and cortex was evident. Conclusions: Phospho-Tau monoclonal antibodies specifically reacting with phosphorylated protein Tau were successfully derived and characterized. The data demonstrate that targeting pTau in combination with detection of aggregated Tau in NFT could be a promising therapeutic approach as indicated by the specificity and binding activity.
P2-526
RENAL FUNCTION OUTCOME AFTER LITHIUM THERAPY FOR COGNITIVE DECLINE
Ivan Aprahamian1, Breno Diniz2, Franklin Santos2, Marcia Radanovic2, Orestes Forlenza2, 1Psychogeriatric Unit, Laboratory of Neuroscience (LIM 27), Department and Institute of Psychiatry, S~ao Paulo, Brazil; 2Laboratory of Neuroscience - LIM 27, Department and Institute of Psychiatry Faculty of Medicine, University of S~ao Paulo, S~ao Paulo, Brazil. Background: Experimental studies conducted in neuronal culture and animal models provide evidence of the neurotrophic and neuroprotective properties of lithium, with alleged impact on the management of certain neurodegenerative conditions, namely Alzheimer’s disease (AD). Recent clinical trials support the feasibility and the potential benefits of this approach. However, chronic lithium use is associated with clinically relevant adverse effects, particularly renal dysfunction. Objective: To evaluate longterm safety of lithium therapy on parameters of renal function of elderly patients with mild cognitive impairment (MCI). Methods: Subjects included in this study are participants of a randomized clinical trial with lithium addressing disease-modification in the MCI-AD continuum. Study design: double blind, placebo controlled trial of lithium (N ¼ 22) versus placebo (N ¼ 22) for the treatment of patients with MCI for three years. Identical tablets holding 150, 300, 450 and 600 mg of lithium carbonate or placebo were administered orally (b.i.d.). Lithium doses were adjusted to keep serum levels between 0.2 and 0.5mEq/L. Safety and tolerability of treatments were longitudinally evaluated at three-month intervals by clinical examination, scores on the UKU side-effect scale, and laboratory tests addressing renal function (and other parameters not included in the present assessment). Statistical analysis was performed with SPSS 18. Results: Baseline serum concentrations of creatinine were not significantly different between the two groups at baseline (lithium, 0.84 6 0.20; placebo, 0.87 6 0.23; p ¼ 0.76), nor at the endpoint of three years (lithium, 0.93 6 0.27; placebo, 0.96 6 0.38; p ¼ 0.19). Kaplan-Meyer curves did not indicate significant changes in renal function according to treatment groups over time (p ¼ 0.31); intra-group comparisons (p ¼ 0.94) indicated long-term stability of such parameters upon follow-up. Conclusions: Low-dose, chronic lithium therapy appears to be clinically safe and not associated with significant changes in renal function in older adults with MCI. The present findings support the use of lithium salts as a candidate drug for disease-modification in AD. P2-527
TARGETING HYPERPHOSPHORYLATED TAU PROTEIN WITH A MONOCLONAL ANTIBODY AT AN ADVANCED STAGE OF TAU PATHOLOGY IMPROVES COGNITION IN A MOUSE MODEL
Allal Boutajangout1, Hameetha Banu Rajamohamed Sait1, Veronica Gonzalez1, Einar Sigurdsson1, 1NYU School of Medicine, New York, New York, United States.