- Email: [email protected]
DEVELOPING TARGETED MONOCLONAL ANTIBODY THERAPIES TO DETECT AND REVERSE CARDIAC AMYLOIDOSIS
Poster Presentations: Wednesday, July 27, 2016
protein-ligand associations. Allo decreased expression of OLR1 and PCSK9 that promote cholesterol efflux and reduce inflammation. Allo increased the gene expression of CXCL16, LRP6, HMGSC1, and Cyp51 involved in stimulation of the PI3K/Akt/ Erk and Wnt pathways and cholesterol synthesis pathway. STAB2 and Idi2 increased and to reduce inflammation signals. Allo also decreased PCSK9 and AKR1D1 genes to promote cholesterol efflux while decreased ANGPTL3 provides a mechanism for reducing plasma cholesterol levels. Allo increased LXRb, LDLR, SORL1, ABCA1, Cyp46a1, and decreased NF-kB protein expression in 3xTgAD male mice. Conclusions: Collectively, increased expression of ABCA1, APOE, LDL-R and SORL1 support the hypothesis that Allo treatment promotes the clearance of cholesterol and Ab. Allo activation of LXR provides a plausible therapeutic strategy to increase Ab degradation and clearance by utilizing ABCA1 and LDL-R to remove Ab. Earlier analysis indicated that Allo increases LXR, PXR, and HMG-CoA-R protein expression (Chen et al., 2011). The results suggest that Allo plays a role in cholesterol homeostasis by directly or indirectly activating LXR and PXR leading to the induction of beneficial genes and pathways to reduce Alzheimer’s pathology. Additional genes promoting cholesterol homeostasis were altered with Allo treatment to protect the brain from inflammation, apoptosis, decrease Ab, and to regulate cholesterol clearance. Research supported by NIA U01 AG031115; NIA U01 AG047222; and UF1 AG046148 to RDB.
P4-027
COMBING EVIDENCE ACROSS MULTIPLE COHORTS FOR SYSTEMS-BASED TARGET DISCOVERY: THE AMP-AD KNOWLEDGE PORTAL
Kristen Dang1, Thanneer M. Perumal1, Mariet Allen2, Corey Funk3, Minghui Wang4, Jishu Xu5, Ben Logsdon1, Lei Yu6, Sarah Schuyler7, Stephen H. Friend1, David A. Bennett6, Bin Zhang4, Eric Schadt4, Philip de Jager5,8,9, Nathan D. Price10, Nilufer Ertekin-Taner2, Lara M. Mangravite1, 1 Sage Bionetworks, Seattle, WA, USA; 2Mayo Clinic, Jacksonville, FL, USA; 3Institute Systems Biology, Seattle, WA, USA; 4Icahn School of Medicine at Mount Sinai, New York, NY, USA; 5Broad Institute, Cambridge, MA, USA; 6Rush University Medical Center, Chicago, IL, USA; 7Mt Sinai, New York, NM, USA; 8Harvard Medical School, Boston, MA, USA; 9 Brigham and Women’s Hospital, Boston, MA, USA; 10Institute for Systems Biology, Seattle, WA, USA. Contact e-mail: thanneer.perumal@sagebase. org Background: Accumulating failures in Alzheimer’s disease drug
development highlight a critical need to improve understanding of the etiology of this disease. The AMP-AD consortium is seeking to advance this understanding and support target discovery through genomic analysis of human disease. To date, the consortium has generated genomic data from blood and postmortem brain samples collected across more than ten sources. The heterogeneity in this collection provides an unprecedented opportunity to identify mechanisms that are robustly observed across multiple representations of this complex disease. Here, we evaluate the continuity in diseaseassociated differential gene expression in RNA sequencing data collected across four sources. Methods: All genomic data generated on brain collections used in AMP-AD was deposited in a publicly accessible to date, centralized repository including RNA sequencing data generated from postmortem brain tissue from four sources: the ROS and MAP cohort studies (N¼547), the Mayo Brain Bank (N¼550), and the Mt. Sinai Brain Bank (N¼448). Experimental design differed across studies in sample ascertainment, criteria for diagnosis, data generation and process-
P1025
ing methods. For 10 samples from each source, data generation was repeated across sequencing centers. Gene-level differential expression was calculated separately within each source by linear regression using VOOM normalized data. When multiple brain regions from the same individual were available, data were jointly analyzed using a mixed effects model. Meta-analysis was performed using Fisher’s method with Brown’s correction. Results: Despite major sources of study-specific technical and biological variation, differences in gene expression between samples classified as Alzheimer’s disease versus control significantly overlapped across studies. 11696 genes were differentially expressed in at least one study (FDR¼5%). Meta-analysis across all studies identified 9186 differentially expressed genes. 582 genes were differentially expressed in more than 75% of studies and these were enriched for genes previously demonstrated to be involved in neurodegenerative processes. Conclusions: Combined analysis identified a highconfidence set of genes that were differentially expressed in relation to Alzheimer’s disease in a consistent manner across a diverse collection of studies. Future analytical efforts that leverage this heterogeneous collection have the potential to identify robust disease mechanisms in support of target discovery.
P4-028
DEVELOPING TARGETED MONOCLONAL ANTIBODY THERAPIES TO DETECT AND REVERSE CARDIAC AMYLOIDOSIS
Natalie J. Galant1,2, Jeffrey N. Higaki3, Avi Chakrabartty1,2, 1University Health Network, Toronto, ON, Canada; 2University of Toronto, Toronto, ON, Canada; 3Prothena Biosciences, San Francisco, CA, USA. Contact e-mail: [email protected] Background: Transthyretin (TTR, or prealbumin) is an abundant
blood serum protein which normally forms soluble, stable tetrameric complexes. Under conditions which are not fully understood, the TTR tetramer structure dissociates into aggregation- and misfolding-prone monomers. These monomers aggregate and accumulate as amyloid fibrils and plaques (amyloidosis) throughout the body-particularly in the heart. TTR-related cardiac amyloidosis (TTR-CA) progresses through dysrhythmia and systolic heart failure, leading eventually to overall heart failure and death (Ruberg, 2012). Monoclonal antibodies (mAbs) which specifically recognize and bind to the disease-associated form of TTR via ’cryptotopes’ have been developed. Cryptotopes are antigens which are normally deeply buried within a protein but exposed in its altered conformation. Understanding the mAbs mechanism of fibril inhibition is investigated. Methods: Four mAbs obtained from Prothena Biosciences (So. South Francisco) were subject to end-point immunogold and initial aggregation kinetics electron microscopy studies. Results: Cryptotopes within pathological TTR misfolding intermediates and end-point insoluble TTR fibrils are exposed and effective mAb targets. Conclusions: Monoclonal antibodies targeting pathological protein conformations are potentially effective pharmacotherapies for preventing the progression of protein misfolding diseases using the exemplar of TTR amyloidosis. Ruberg, F. L., Berk, J. L. (2012). Transthyretin (TTR) cardiac amyloidosis. Circulation. 126(10): 1286-1300. P4-029
GENE EXPRESSION CHANGES IN TGAD AND NON-TRANSGENIC LITTERMATE MICE AS A FUNCTION OF SCYLLO-INOSITOL TREATMENT
Qingda Hu1, Aaron Y. Lai2, Mary E. Brown2, JoAnne McLaurin2, 1 Sunnybrook Health Sciences Centre, Toronto, ON, Canada; 2Sunnybrook
protein-ligand associations. Allo decreased expression of OLR1 and PCSK9 that promote cholesterol efflux and reduce inflammation. Allo increased the gene expression of CXCL16, LRP6, HMGSC1, and Cyp51 involved in stimulation of the PI3K/Akt/ Erk and Wnt pathways and cholesterol synthesis pathway. STAB2 and Idi2 increased and to reduce inflammation signals. Allo also decreased PCSK9 and AKR1D1 genes to promote cholesterol efflux while decreased ANGPTL3 provides a mechanism for reducing plasma cholesterol levels. Allo increased LXRb, LDLR, SORL1, ABCA1, Cyp46a1, and decreased NF-kB protein expression in 3xTgAD male mice. Conclusions: Collectively, increased expression of ABCA1, APOE, LDL-R and SORL1 support the hypothesis that Allo treatment promotes the clearance of cholesterol and Ab. Allo activation of LXR provides a plausible therapeutic strategy to increase Ab degradation and clearance by utilizing ABCA1 and LDL-R to remove Ab. Earlier analysis indicated that Allo increases LXR, PXR, and HMG-CoA-R protein expression (Chen et al., 2011). The results suggest that Allo plays a role in cholesterol homeostasis by directly or indirectly activating LXR and PXR leading to the induction of beneficial genes and pathways to reduce Alzheimer’s pathology. Additional genes promoting cholesterol homeostasis were altered with Allo treatment to protect the brain from inflammation, apoptosis, decrease Ab, and to regulate cholesterol clearance. Research supported by NIA U01 AG031115; NIA U01 AG047222; and UF1 AG046148 to RDB.
P4-027
COMBING EVIDENCE ACROSS MULTIPLE COHORTS FOR SYSTEMS-BASED TARGET DISCOVERY: THE AMP-AD KNOWLEDGE PORTAL
Kristen Dang1, Thanneer M. Perumal1, Mariet Allen2, Corey Funk3, Minghui Wang4, Jishu Xu5, Ben Logsdon1, Lei Yu6, Sarah Schuyler7, Stephen H. Friend1, David A. Bennett6, Bin Zhang4, Eric Schadt4, Philip de Jager5,8,9, Nathan D. Price10, Nilufer Ertekin-Taner2, Lara M. Mangravite1, 1 Sage Bionetworks, Seattle, WA, USA; 2Mayo Clinic, Jacksonville, FL, USA; 3Institute Systems Biology, Seattle, WA, USA; 4Icahn School of Medicine at Mount Sinai, New York, NY, USA; 5Broad Institute, Cambridge, MA, USA; 6Rush University Medical Center, Chicago, IL, USA; 7Mt Sinai, New York, NM, USA; 8Harvard Medical School, Boston, MA, USA; 9 Brigham and Women’s Hospital, Boston, MA, USA; 10Institute for Systems Biology, Seattle, WA, USA. Contact e-mail: thanneer.perumal@sagebase. org Background: Accumulating failures in Alzheimer’s disease drug
development highlight a critical need to improve understanding of the etiology of this disease. The AMP-AD consortium is seeking to advance this understanding and support target discovery through genomic analysis of human disease. To date, the consortium has generated genomic data from blood and postmortem brain samples collected across more than ten sources. The heterogeneity in this collection provides an unprecedented opportunity to identify mechanisms that are robustly observed across multiple representations of this complex disease. Here, we evaluate the continuity in diseaseassociated differential gene expression in RNA sequencing data collected across four sources. Methods: All genomic data generated on brain collections used in AMP-AD was deposited in a publicly accessible to date, centralized repository including RNA sequencing data generated from postmortem brain tissue from four sources: the ROS and MAP cohort studies (N¼547), the Mayo Brain Bank (N¼550), and the Mt. Sinai Brain Bank (N¼448). Experimental design differed across studies in sample ascertainment, criteria for diagnosis, data generation and process-
P1025
ing methods. For 10 samples from each source, data generation was repeated across sequencing centers. Gene-level differential expression was calculated separately within each source by linear regression using VOOM normalized data. When multiple brain regions from the same individual were available, data were jointly analyzed using a mixed effects model. Meta-analysis was performed using Fisher’s method with Brown’s correction. Results: Despite major sources of study-specific technical and biological variation, differences in gene expression between samples classified as Alzheimer’s disease versus control significantly overlapped across studies. 11696 genes were differentially expressed in at least one study (FDR¼5%). Meta-analysis across all studies identified 9186 differentially expressed genes. 582 genes were differentially expressed in more than 75% of studies and these were enriched for genes previously demonstrated to be involved in neurodegenerative processes. Conclusions: Combined analysis identified a highconfidence set of genes that were differentially expressed in relation to Alzheimer’s disease in a consistent manner across a diverse collection of studies. Future analytical efforts that leverage this heterogeneous collection have the potential to identify robust disease mechanisms in support of target discovery.
P4-028
DEVELOPING TARGETED MONOCLONAL ANTIBODY THERAPIES TO DETECT AND REVERSE CARDIAC AMYLOIDOSIS
Natalie J. Galant1,2, Jeffrey N. Higaki3, Avi Chakrabartty1,2, 1University Health Network, Toronto, ON, Canada; 2University of Toronto, Toronto, ON, Canada; 3Prothena Biosciences, San Francisco, CA, USA. Contact e-mail: [email protected] Background: Transthyretin (TTR, or prealbumin) is an abundant
blood serum protein which normally forms soluble, stable tetrameric complexes. Under conditions which are not fully understood, the TTR tetramer structure dissociates into aggregation- and misfolding-prone monomers. These monomers aggregate and accumulate as amyloid fibrils and plaques (amyloidosis) throughout the body-particularly in the heart. TTR-related cardiac amyloidosis (TTR-CA) progresses through dysrhythmia and systolic heart failure, leading eventually to overall heart failure and death (Ruberg, 2012). Monoclonal antibodies (mAbs) which specifically recognize and bind to the disease-associated form of TTR via ’cryptotopes’ have been developed. Cryptotopes are antigens which are normally deeply buried within a protein but exposed in its altered conformation. Understanding the mAbs mechanism of fibril inhibition is investigated. Methods: Four mAbs obtained from Prothena Biosciences (So. South Francisco) were subject to end-point immunogold and initial aggregation kinetics electron microscopy studies. Results: Cryptotopes within pathological TTR misfolding intermediates and end-point insoluble TTR fibrils are exposed and effective mAb targets. Conclusions: Monoclonal antibodies targeting pathological protein conformations are potentially effective pharmacotherapies for preventing the progression of protein misfolding diseases using the exemplar of TTR amyloidosis. Ruberg, F. L., Berk, J. L. (2012). Transthyretin (TTR) cardiac amyloidosis. Circulation. 126(10): 1286-1300. P4-029
GENE EXPRESSION CHANGES IN TGAD AND NON-TRANSGENIC LITTERMATE MICE AS A FUNCTION OF SCYLLO-INOSITOL TREATMENT
Qingda Hu1, Aaron Y. Lai2, Mary E. Brown2, JoAnne McLaurin2, 1 Sunnybrook Health Sciences Centre, Toronto, ON, Canada; 2Sunnybrook