CHDI’s 8th Annual HD Therapeutics Conference took place April 8 – 11, 2013, in Venice, Italy. This unique conference series focuses on drug discovery and development for Huntington’s disease, and draws participants and speakers from the biotech and pharmaceutical sectors as well as academia and research institutions. The conference is intended as a forum where all participants can share ideas, learn about new disciplines, network with colleagues and build new collaborative partnerships. We are indebted to all of the conference speakers, and especially grateful to those who are able to make their presentations available here for a wider audience.
- Systems Drug Discovery and Neurodegenerative Diseases Hiroaki Kitano, PhD
- Applying Systems Biology at CHDI Jim Rosinski, PhD
- Gene Expression Changes in the Straita of Mouse Models of Huntington’s Disease Lesley Jones, PhD
- A Combined Genetic and Systems Biology Dissection of HD Pathogenesis in Mice X. William Yang, MD, PhD
- Weighted Network Analysis Applied to Huntington’s Disease Data Sets Steve Horvath, PhD, ScD
- Translating the Natural History of Human Striatal Development into Pluripotent Stem Cell Differentiation – Starting from Evolution Elena Cattaneo, PhD
- CAG Repeats in Huntingtin and Brain Development: Normal and Pathogenic Conditions Peg Nopoulos, MD
- Development and Molecular Signature of HD-central Cortico-striatal Projection Neurons Jeffery D. Macklis, MD
- Functional Characterization of Htt Protein During Early Embryogenesis Ali Brivanlou, PhD
- Chemical and Semisynthetic Strategies for Elucidating the Molecular Determinants of Htt Aggregation and Toxicity Hilal A. Lashuel, PhD
- Mechanisms Underlying Loss of Axonal Connectivity in Huntington’s Disease Gerardo Morfini, PhD
- Impaired TrkB Receptor Signaling Underlies Corticostraital Synaptic Dysfunction in Huntington’s Disease D. James Surmeier, PhD
- Allele-Specific Repression of Huntingtin with Engineered Zing Finger Protein Transcription Factors Philip Gregory, DPhil
- Allele-Selective Silencing by Targeting Mutant CAG Repeats David R. Corey, PhD
- Advancing Phosphodiesterase 10A (PDE10A) Inhibitor from Bench to Clinic Margaret M. Zaleska, PhD
- Development of Kynurenine Monooxygenase (KMO) Inhibitor CHDI-340246 for the Treatment of Huntington’s Disease: A Progress Update Ladislav Mrzljak, MD, PhD
- Development and Testing of Selective HDAC Class IIa Inhibitors for Preclinical Therapeutic Proof of Concept in HD Models Vahri Beaumont, PhD
- Advancing TrkB Modulation Toward Huntington’s Disease Therapeutics: Keeping on Trk Jonathan Bard, PhD
- The Pursuit of Disease Biomarkers for HD: The Importance of Replication and Validation Beth Borowsky, PhD
- Enroll-HD – A Hub for Biomarker Development Tiago Mestre, MD
- Alzheimer Prevention Initiative: A Landmark Trial for Prevention of Alzheimer Diease Carole Ho, MD
- PET and SPECT Imaging Biomarkers for HD: Developing a Translation Toolbox Kenneth Marek, MD
Systems Drug Discovery and Neurodegenerative Diseases
Presentation not yet available
Hiroaki Kitano, PhD – Systems Biology Institute & Okinawa Institute of of Science & Technology
Treating complex diseases requires a systematic approach that are most often benefits from systems biology. It requires in-depth understanding of molecular mechanisms of disease outbreak, progressions, and impact of therapeutic interventions. Proper theoretical framework and a set of software platforms are critical in successful drug development as well as long-term healthcare projects. This talk highlights some of our recent approaches for drug discoveries and software platform for supporting it. Discovering possible care for CNS related diseases is particularly challenging due to complexities of diseases as well as multifaceted nature of disease phenotypes. This talk introduces some of our attempts for CNS related diseases.
Applying Systems Biology at CHDI
Jim Rosinski, PhD – CHDI
Viewing diseases holistically as a systemic condition affecting all levels of an organism’s molecular, cellular, and organ biology has become a common way to approach therapeutic discovery. Last year, we proposed the strategy for bringing systems biology to CHDI and a plan for implementing systems biology techniques to Huntington’s Disease. We spoke of novel animal and cellular models of the disease as well as collaborations to apply new mathematical algorithms to our data. In the presentation, we’d like to update you on the progress we’ve made in building large scale genomic datasets, as well as analyzing and computationally modeling those sets. Specifically, we will discuss progress in applying causal modeling techniques to gene expression data as well as moving from array-based mRNA expression platforms to RNA sequencing.
We will show how these techniques can be applied to our data through an example of RNAseq profiling of the Q175 mouse model of Huntington’s Disease. This study shows that the Q175 model is very representative of the pathways we suspect drive HD. The study also clearly shows the power of RNAseq to observe aspects of the transcriptome that were difficult, if not impossible, to see using hybridization arrays. Finally, we will discuss new collaborations we’ve built to further implement our systems biology strategy.
Gene Expression Changes in the Straita of Mouse Models of Huntington’s Disease
Presentation not yet available
Lesley Jones, PhD – Cardiff University
Multiple mouse lines exist carrying the expanded CAG repeat of the HTT gene in their genomes through transgenesis or knock in technologies. All show some behavioural and pathological characteristics that recapitulate what we know about Huntington’s disease (HD) in people and we have been exploring these changes at a molecular and behavioural level in multiple HD model mouse lines. We have carried out gene expression analyses using Affymetrix Mouse gene ST arrays and real-time quantitative real time PCR validation. We are particularly interested in which changes are similar in multiple animal models, which changes are specific to certain models, and whether the common changes are reflected in changed gene expression in human caudate. We are also interested in using the gene expression profiles to stage the phenotype of the animals and in correlating such changes with behavioural changes in multiple motor, cognitive and behavioural paradigms. We detect a common set of changes in gene expression, particularly amongst down-regulated genes. We have examined the ability of gene expression changes in one mouse line to predict the status and phenotypic stage of other mouse models constructed using different genetic strategies. Some of the behavioural assessments give similar trajectories of change in multiple models and some do not. We have examined whether these differences could be associated with the observed gene expression profile differences in order to determine what underlying molecular changes could manifest the observed behavioural changes.
A Combined Genetic and Systems Biology Dissection of HD Pathogenesis in Mice
X. William Yang, MD, PhD, – University of California, Los Angeles
An important yet unanswered question in Huntington’s disease (HD) pathogenesis is how the ubiquitously expressed mutant huntingtin (mHtt) can cause an age-dependent plethora of neurological and psychiatric symptoms along with selective degeneration of cortical and striatal neurons. To address this complex question, our laboratory undertook a combined genetic and systems biology approach to examine mechanisms underlying age- and brain-region-specific pathogenic processes. These studies are based on a conditional Bacterial Artificial Chromosome transgenic mouse model of HD (BACHD) expressing full-length human mHtt under human genomic regulation. BACHD mice exhibit progressive motor and psychiatric-like behavioral deficits and late-onset selective cortical and striatal atrophy. We first performed affinity purification-mass spectrometry (AP-MS) to profile a spatiotemporal in vivo Htt interactome in the brain. Analyses of the entire AP-MS dataset using Weighted Gene Correlation Network Analyses (WGCNA) enabled us to visualize distinct Htt-correlated protein networks in the brain, which provided insights into normal Htt function in the brain and candidate proteins for mediating HD pathogenesis. In a second study, taking advantage of the conditional BAC transgenic design in BACHD, we systematically reduced mHtt expression in striatal, cortical, or both neuronal populations. These new BACHD variants not only helped us define the distinct neuronal targets of mHtt genetic reduction to ameliorate or prevent HD, but also provided a flexible model platform to apply systems biology to identify cortical and striatal gene networks that are the pathological consequences of cell-autonomous or non-cell-autonomous mHtt toxicities in vivo. Future validation of key network genes from these combined genetic/systems biology studies may yield novel therapeutic targets for HD.
Funding: This work is supported by grants from US National Institute of Health (NIH)/National Institute of Neurological Disorders and Stroke (NINDS), CHDI Foundation, Inc, and the Hereditary Disease Foundation (HDF).
Weighted Network Analysis Applied to Huntington’s Disease Data Sets
Steve Horvath, PhD, ScD – University of California, Los Angeles
This talk describes weighted network analysis strategies for analyzing genomic data sets (mainly gene expression) from different human and mouse tissues. The first strategy involves using sample network approaches for data pre-processing and for outlier detection. Sample networks are network whose nodes correspond to tissue samples (or subjects). The second strategy involves a consensus network approach for aggregating multiple gene expression data sets. Consensus modules are clusters of genes that are present across multiple data sets. We provide case studies where hub gene selection in consensus modules led to superior biological insights. The third strategy involves network based methods for evaluating whether disease modules found in human tissue are present in corresponding mouse tissues and vice versa. Several case studies are used to illustrate the utility of the resulting module preservation statistics. This is joint work with Peter Langfelder, Michael C Oldham, and William Yang.
Translating the Natural History of Human Striatal Development into Pluripotent Stem Cell Differentiation – Starting from Evolution
Presentation not yet available
Elena Cattaneo, PhD – University of Milano
The presentation will discuss the evolutionary aspects of the CAG in huntingtin and the data available about the CAG repeats having an impact on brain development. I will then move to the study of human brain development to show unpublished data on the spatio-temporal expression pattern of transcription factors that mark human fetal striatal development. We will reveal the antigenic and molecular attributes that qualify the striatal progenitors and their transition towards terminal neuronal differentiation as a necessary step in order to assess the influence of the normal and expanded CAG on brain development. We will finally review how incorporation of these informations into human pluripotent stem cell differentiation has allowed the generation of authentic and functionally active DARPP-32+/CTIP2+ medium-sized spiny neurons.
CAG Repeats in Huntingtin and Brain Development: Normal and Pathologic Conditions
Peg Nopoulos, MD – University of Iowa
The research program ‘Children at risk for Huntington’s Disease’, otherwise known as the KidsHD Program, is designed to evaluate the brain structure and function of children who come from families in which a parent is affected with HD. Children ages 6-18 years are recruited and are assessed with brain imaging (Magnetic Resonance Imaging, MRI), and measures of cognition, behavior, and motor skills. Children also provide DNA so that, for research purposes only, they can be divided into those that are gene-expanded (GE, CAG repeat >40) or gene non-expanded (GNE, CAG repeat <39). All of these children are without any signs of disease, and therefore exclude any child that may be of juvenile onset. The primary aims of the study are two-fold and evaluate the effects of Huntingtin (HTT) on the developing brain in both normal conditions (the GNE group) and in pathologic conditions (the GE group). The effects of HTT on the normal developing brain is a research question born from the notion that triplet repeat genes possess a unique mechanism for evolutionary progress and may have been important in the evolution from primate to human brain. More importantly, genes involved in evolutionary change are typically genes that govern development. In a sample of roughly 50 GNE children, length of CAG repeat is shown to be directly related to some aspects of cognition, behavior and motor skill with higher CAG repeats predicting superior performance in all 3 areas. The role of HTT in a pathologic condition is a research question based on the notion that abnormal brain development may play a key role in the pathophysiology of HD. In a group of approximately 50 GE children, there is both general (total brain tissue) and specific (striatum) decrements in volume compared to the GNE sample, and in comparison to a healthy control sample. Functional comparison of the GE group shows an important role of CAG repeat length with children in the range of 40-44 CAG repeat lengths doing comparable to controls while children with repeats > 45 showing subtle deficits in fine motor skills, impulsivity and inattention.
Development and Molecular Signature of HD-central Cortico-striatal Projection Neurons
Presentation not yet available
Jeffery D. Macklis, MD – Harvard University
Corticostriatal projection neurons (CStrPN) are the cortical efferent neurons of cortico-basal ganglia circuitry, and their degeneration is central to Huntington’s disease (HD). Little is known about their development, nor about their apparent selective vulnerability in HD versus hundreds of other subtypes of cortical projection neurons. Understanding how CStrPN and their circuitry develop will contribute to understanding their organization, function, and vulnerability to disease, and also potentially toward their pharmacologic or biologic protection, treatment, or repair of those that degenerate in HD.
“Intratelencephalic” CStrPN (CStrPNi) project to the contralateral striatum, with their axons fully within the telencephalon (“intratelencephalic”). CStrPNi are the dominant population of CStrPN, and are of particular interest because they share characteristics of both callosal projection neurons (CPN) and corticofugal projection neurons (CFuPN), sending axons contralaterally before descending into the contralateral striatum. The development of this “chimeric” population of CStrPNi has not been previously investigated, and the relationship of CStrPNi development to that of broader CPN and CFuPN population remains unclear.
We initiated investigation of the development of CStrPNi in mice by first studying their birthdates, maturation of connectivity, multiplicity of projections, and expression of known molecular developmental controls over projection neuron subtype differentiation, with the future aim of identifying molecular controls over their specification and differentiation. Investigation of molecular controls over subtype-specific CStrPNi development builds on strategies our lab has already applied to molecular development of corticospinal (CSMN)/subcerebral, CPN, and corticothalamic (CThPN)/CFuPN. We isolated mouse CStrPNi at distinct stages of development by retrograde labeling from striatum, along with exclusionary and dual cortico-cortical labeling. CStrPNi, “pure” CPN, [and CSMN, CThPN] were isolated using fluorescence-activated cell sorting (FACS), yielding nearly pure populations of CStrPNi (vs. these other populations) at critical stages during development. Identification of stage-specific gene expression by CStrPNi, by comparative microarray analysis and multiple biological “filters”, and comparison to that of CSMN, CPN, and CThPN promises to enable identification of critical combinatorial determinants that define, specify, and control differentiation and maturation of CStrPNi and cortico-basal ganglia circuitry more generally. We are now identifying top candidate CStrPNi developmental molecular controls based on both stage-specific expression and potential function.
Functional Characterization of Htt Protein During Early Embryogenesis
Presentation not yet available
Ali Brivanlou, PhD – The Rockefeller University
Huntington disease (HD) is due to a mutation that adds poly-Q repeats to the N-terminus of the Htt protein leading to a devastating neurodegenerative outcome many years after birth. However, Htt protein is expressed throughout development from the first cell, the fertilized egg, the embryo, and throughout life. While tremendous efforts have been undertaken to scrutinize the role of the gene in the brain, its function during the earliest stages of development is largely unexplored. Using Xenopus, mouse, and human embryos, as well as human and mouse embryonic stem cells (hESCs and mESCs), my laboratory aims to decipher the role of the Htt protein during early embryogenesis. Three topics will be discussed in that context.
First, RNA-seq analysis of hESCs led to the discovery of 4 previously undetected alternatively spliced forms of htt mRNA. The presence of these novel transcripts was confirmed by RT-PCR and sequencing. Alternatively spliced htt isoforms affect the coding sequence of the Htt protein, and post-translational modification sites. One of these isoform is also detected in Xenopus embryos and is evolutionarily conserved. Characterization of temporal and spatial expression of these previously unrecognized isoforms, particularly these that are specifically expressed in the embryonic nervous system, will enhance our knowledge of HD. Secondly, following on the original observation that the htt-/- mutation in mouse embryos leads to death with severe defects in gastrulation, and our own work demonstrating that gastrulation is governed by a specific set of embryonic signaling pathways, the cross-talk between Htt protein and the embryonic signaling network was examined. Interestingly, loss of Htt function in hESCs increased the signaling output of both arms of the canonical TGFβ signaling pathway, as demonstrated by the increase of phospho-Smad1 and -Smad2 levels. Additionally, and the cells have an increased sensitivity to BMP4 induction, as shown by reduction of pluripotency and induction of the differentiation markers. Interestingly loss of Htt also affected the dynamics of the pathway. This provides a molecular explanation of the lethality of the htt-/- mutant mouse embryo. Finally, global metabolomics analysis of a sibling hESC lines, one of which is carrying the HD mutation, and one that is normal, provide the novel finding that the mutant lines surprisingly display a general deficiency in energy metabolism at this early stage of development. Comparisons between htt-/- and normal mouse ES cells also showed that normal Htt plays a significant role in energy metabolism, suggesting this function might be seriously disrupted in the disease case.
The discovery of novel htt isoforms, the modulation of the TGFβ signaling pathway by Htt, the serious disruption of energy metabolism, the understanding of how cells cope with this deficiency and why they eventually fail, might trigger a better understanding of Htt function while potentially leading to the development of treatments for HD.
Chemical and Semisynthetic Strategies for Elucidating the Molecular Determinants of Htt Aggregation and Toxicity
Presentation not yet available
Hilal A. Lashuel, PhD – Ecole Polytechnique Fédérale de Lausanne
A better understanding of the molecular and cellular determinants that influence the pathology of Huntington’s disease (PD) is essential for developing effective diagnostic, preventative and therapeutic strategies to treat this devastating disease. Increasing evidence suggest that the aggregation and toxicity of the Huntingtin protein is strongly influenced by post-translational modification and sequences outside the polyQ repeat region. Several post-translational modifications have been identified within the N-terminal 17 residues of exon1 of the Huntingtin (Httex1), including acetylation, phosphorylation, SUMOylation and ubiquitination at multiple residues. However, whether these modifications promote or inhibit Htt or Httex1 aggregation and neurotoxicity in vivo remains unknown. This is in part due to the fact that many of the enzymes involved in regulating these modifications remain unknown and the preparation of homogeneously modified forms of the protein has not been possible.
In my talk, I will present new semisynthetic strategies developed in our laboratory to allow site specific introduction of post-translational modifications at single or multiple residues within the N-terminus of Httex1. Using these strategies, we have been able to produce in mg quantities site-specifically phosphorylated forms of Httex1, including pT3, pS13 or pS16. These advances allowed us for the first time to investigate the effect of phosphorylation in the structure and aggregation of Httex1 in vitro and provide valuable tools for future mechanistic studies and development of assays to quantify the level of these modifications in vivo or screen for the enzymes involved in regulating phosphorylation at these residues. Finally, I will present recent studies from our group that led to the identification of a novel aggregation sequence motifs outside the polyQ repeat region. The implications of this discovery for Htt aggregation, proteolysis and toxicity will be discussed.
Mechanisms Underlying Loss of Axonal Connectivity in Huntington’s Disease
Gerardo Morfini, PhD – University of Illinois, Chicago
Expansion of a polyglutamine (polyQ) tract in huntingtin (Htt) results in Huntington’s disease (HD), a fatal neurodegenerative disease involving dying back degeneration of selected neuronal populations in the striatum and cerebral cortex. Ample genetic evidence suggests that polyQ tract expansion confers upon Htt a toxic gain of function. However, mechanisms by which mutant Htt promotes loss of neuronal connectivity in HD remain elusive.
Pharmacological, biochemical, and cell biological experiments will be presented, which povide the basis for a novel pathogenic mechanism for HD. This mechanism involves activation of selected molecular components of the JNK pathway and deficits in axonal transport, a critical cellular process for the maintenance of axonal connectivity.
Impaired TrkB Receptor Signaling Underlies Corticostraital Synaptic Dysfunction in Huntington’s Disease
Presentation not yet available
D. James Surmeier, PhD – Northwestern University
In the earliest stages of Huntington’s disease (HD), uncontrolled choreic movements plague patients. These symptoms have been traced to dysfunction of striatal indirect pathway spiny projection neurons (iSPNs). However, the nature of this pathway selective pathology has been elusive. An important clue about pathogenesis has come from the discovery that striatal brain derived neurotrophic factor (BDNF) signaling is depressed in HD models and patients. We have found that in transgenic mouse models of HD, the ability of BDNF to signal through TrkB receptors is selectively impaired in iSPNs at the time motor dysfunction appears. This impairment resulted in an up-regulation in Kv4 channels (suppressing synaptic integration) and a loss of longterm potentiation at corticostriatal synapses at synapses in iSPNs. Synaptic plasticity in HD models could be rescued by inhibition of the BDNF co-activated p75 receptor pathway through phosphatase and tensin homolog depleted on chromosome 10 (PTEN) or by stimulating fibroblast growth factor receptors. These studies suggest that a correctable inhibition of TrkB receptor signaling results in compromised cortical drive of iSPNs in HD, leading to the cardinal hyperkinetic symptoms at disease onset.
Allele-Specific Repression of Huntingtin with Engineered Zing Finger Protein Transcription Factors
Presentation not available
Philip Gregory, DPhil – Sangamo BioSciences Inc
Huntington’s disease (HD) is an autosomal dominant neurodegenerative disease caused by CAG-trinucleotide repeat expansion in the first exon of the Huntingtin (Htt) gene. Repeat lengths of 35 or fewer CAGs are normal and usually have no associated pathophysiology, while those of 40 or more lead to HD with 100% penetrance, with longer repeat lengths correlating with earlier disease onset. The degeneration process primarily affects the basal ganglia and cerebral cortex, and the disease is characterized by a progressively worsening chorea, as well as cognitive and psychiatric dysfunctions. While neither the precise function of wild-type Htt protein nor the mechanism by which mutant Htt (which contains an expanded polyglutamine stretch) in HD pathogenesis is fully understood, results from rodent models of HD demonstrate that reducing mutant Htt levels, can prevent disease onset or delay disease progression. Thus, strategies that selectively reduce the expression of mutant and disease causing form of Htt represent the ideal therapeutic approach. Engineered zinc finger protein transcription factors (ZFP TF) can be designed to up- or down-regulate gene expression with exceptional specificity. Acting at the DNA level these factors turn virtually any gene into a potential drug target – a feature of particular significance for HD, where a genetic signature of disease has been identified that has thus far evaded classical small molecule drug intervention. This presentation will review recent data demonstrating that ZFP TFs can be designed to control Htt gene expression in cells and in pre-clinical mouse models of HD. Moreover, data demonstrating the selective regulation of the mutant Htt allele in HD patient derived cells will be presented. Together these results support the development of engineered ZFP TFs for the treatment of HD.
Allele-Selective Silencing by Targeting Mutant CAG Repeats
David R. Corey, PhD – UT Southwestern Medical Center
Mutant huntingtin (HTT) protein causes Huntington’s Disease (HD) and silencing mutant HTT using nucleic acids would eliminate the root cause of HD. Developing nucleic acid drugs is challenging, and an ideal clinical approach to gene silencing would combine the simplicity of single-stranded antisense oligonucleotides with the efficiency of RNAi. Here we describe RNAi by single-stranded silencing RNAs (ss-siRNAs) complementary to CAG repeats. ss-siRNAs are potent (>100-fold more than unmodified RNA) and allele-selective (>30-fold) inhibitors of mutant HTT expression in cells derived from HD patients. Strategic placement of mismatched bases mimics micro-RNA recognition and optimizes discrimination between mutant and wild-type alleles. ss-siRNAs require argonaute protein and function through the RNAi pathway. Intraventricular infusion of ss-siRNA produced selective silencing of the mutant HTT allele throughout the brain in a mouse HD model. These data demonstrate that chemically modified ss-siRNAs function through the RNAi pathway and provide allele-selective compounds for clinical development. We also describe development of potent and selective duplex RNAs and their modification to create a large family of active lead compounds. Anti-CAG duplex RNAs and ss-siRNAs provide a diverse collection of molecules for animal testing and optimization for in vivo efficacy.
Advancing Phosphodiesterase 10A (PDE10A) Inhibitor from Bench to Clinic
Margaret M. Zaleska, PhD – Pfizer Inc
Accumulating preclinical data continue to support the utility of PDE10A inhibition as a therapeutic target for treating the symptoms of Huntington’s Disease (HD). PDE10A is a dual substrate cyclic nucleotide phosphodiesterase expressed almost exclusively in medium spiny neurons (MSNs) of the mammalian striatum where it regulates the sensitivity of these neurons to glutamatergic input. PDE10A inhibition increases the activity of both the cAMP and cGMP signaling cascades as well as the MAP kinase pathway and results in a powerful induction of striatal gene transcription and an overall increase of striatal output. Moreover, results from numerous studies indicate a preferential effect of PDE10A inhibitors within the D2-expressing MSNs of the indirect pathway which are known to be affected in the initial phase of HD. These effects would be predicted to counter the well established impairment of cAMP signaling, transcriptional dysregulation and altered synaptic plasticity in HD. In collaboration with CHDI, we have recently demonstrated that inhibitors of PDE10A, are effective in reversing multiple parameters of aberrant excitability of MSNs, and in improving elements of corticostriatal dysfunction in brain slices derived from symptomatic R6/2 and Q175 knock-in mice. In vivo improvement of indirect pathway function was shown in a third model, the full length Htt transgenic BACHD rat, following acute PDE10 inhibition. Despite the dominant role of PDE10A in the regulation of striatal cAMP signaling, early recognition of the loss of PDE10A expression in transgenic models of HD suggested its utility as a target might be limited. However, recent studies in R6/2 and Q175 transgenic mice indicate PDE10A inhibition can elicit a robust biochemical response with enzyme levels as low as ~20%. A small pilot study using the PDE10A PET radioligand [18F]-MNI-695 suggests as much as 40-50% of the enzyme may be preserved in HD Stage I/II patients. This observation will be confirmed in a large collaborative, cross-sectional study with CHDI and the Karolinska Institute correlating PDE10A levels, CAG repeats and disease stage in pre-symptomatic, Stage I and Stage II patients.
Our broader strategy for interrogating the effects of PDE10A inhibition on corticostriatal function in early HD patients will focus on clinical studies with PF-2545920, a highly selective, brain penetrant PDE10A inhibitor and will include an Enzyme Occupancy study in healthy volunteers to establish the relationship between plasma exposure levels of PF-2545920 and target occupancy. Lastly, our collaborative efforts with the Brain and Spine Institute (ICM) will test the safety and translatability of preclinical findings in early HD patients in a proof-of-mechanism study that will include, in addition to traditional motor clinical measures and cognitive testing, the effects on corticostriatal circuits following a 28-day treatment with PF-02545920 or placebo using functional imaging, behavioral tasks, quantitative motor tests and a novel apathy battery. Findings from all above studies will aid patient cohort selection and refine dosing regimen for the proof-of-concept and subsequent clinical studies.
Development of Kynurenine Monooxygenase (KMO) Inhibitor CHDI-340246 for the Treatment of Huntington’s Disease: A Progress Update
Ladislav Mrzljak, MD, PhD – CHDI
Metabolites of the kynurenine pathway kynurenine (KYN) and kynurenic acid (KYNA) are suggested to modulate synaptic plasticity and have neuroprotective and anti-inflammatory roles in Huntington’s disease (HD) models (Zwilling et al. Cell 145: 1-12, 2011; Zadori et al. J. Neural Transm 118: 865-875, 2011; for review see Vecsei et al. Nature Rev/Drug Discovery 12: 64-82, 2013). We have developed a KMO inhibitor CHDI-340246 that potently increases the levels of KYN and KYNA in the brains of rodent HD models and their WT controls as well as in the cerebrospinal fluid of non-human primates. Acute and chronic efficacy testing of CHDI-340246 in animal models of HD will be discussed.
Development and Testing of Selective HDAC Class IIa Inhibitors for Preclinical Therapeutic Proof of Concept in HD Models
Vahri Beaumont, PhD – CHDI
HDAC4 heterozygous knock-out animals crossed to the R6/2 mouse model of Huntington’s disease (HD) led to significant improvement in motor co-ordination and neurological phenotypes, improved medium spiny neuron properties and corticostriatal synaptic function, as well as increased R6/2 lifespan. Furthermore, HDAC4 reduction delayed cytoplasmic aggregate formation in both R6/2 and the full length mHtt Q150 knock-in mouse model (data from Mielcarak et al, Professor G Bates group, King’s College London). These impressive genetic validation findings indicate that HDAC4, a member of the Class IIa histone deacetylase family, may present a novel target for pharmacological manipulation towards a therapeutic for HD.
Although widely expressed in the brain, the role of HDAC4 in the CNS is not understood. Class IIa HDACs, including HDAC4, have negligible catalytic activity when compared to Class 1 HDACs , and their enzymatic potential as bona fide deacetylases has been called into question . Additionally, a conserved N-terminal extension of roughly 600 residues is conserved across all Class IIa enzymes , and has been shown to adopt various regulatory functions, including protein-protein interactions and most notably binding to transcription factors . Thus a critical issue is whether or not occupancy of the HDAC4 catalytic domain with a small molecule will be capable of replicating, either fully or in part, the beneficial effects of HDAC4 genetic reduction in HD models.
Working with Biofocus, we have developed novel small-molecule inhibitors of the HDAC4 catalytic site with suitable selectivity, cell-based activity and pharmacokinetic properties for preclinical therapeutic proof of concept trials in HD models. I will review the pharmacological and pharmacokinetic profile of the lead preclinical candidate compounds and provide an overview of the current status of our evaluation of the efficacy of these inhibitors in HD-relevant assays and models.
1. Lahm A, Paolini C, Pallaoro M, Nardi MC, Jones P, et al. (2007) Unraveling the hidden catalytic activity of vertebrate class IIa histone deacetylases. Proc Natl Acad Sci U S A 104: 17335-17340.
2. Jones P, Altamura S, De Francesco R, Gallinari P, Lahm A, et al. (2008) Probing the elusive catalytic activity of vertebrate class IIa histone deacetylases. Bioorg Med Chem Lett 18: 1814-1819.
3. de Ruijter AJ, van Gennip AH, Caron HN, Kemp S, van Kuilenburg AB (2003) Histone deacetylases (HDACs): characterization of the classical HDAC family. Biochem J 370: 737-749.
4. Zhao X, Sternsdorf T, Bolger TA, Evans RM, Yao TP (2005) Regulation of MEF2 by histone deacetylase 4- and SIRT1 deacetylase-mediated lysine modifications. Mol Cell Biol 25: 8456-8464.
Advancing TrkB Modulation Toward Huntington’s Disease Therapeutics: Keeping on Trk
Jonathan Bard, PhD – CHDI
Brain-derived neurotrophic factor (BDNF) is a pleiotropic secreted protein that promotes neuronal cell survival by activating the TrkB receptor. Reduced levels of BDNF have been described in animal models of and human patients with Huntington’s disease (HD), and suggested to play a role in the pathogenesis of the disease; therefore, the ability to enhance TrkB signaling specifically within the cortical-striatal-thalamic pathways, and avoiding other subregions that may contribute to adverse side effects, may help to slow the onset or progression of HD.
Our previous strategy involved a computationally-driven rational design approach to identify a small molecule agonist or positive allosteric modulator of trkB receptors, which was unsuccessful. Our current efforts are centered on evaluating the potential therapeutic effects of a monoclonal TrkB agonist antibody (TrkB/mAb) developed by Pfizer [1,2], of an AAV/BDNF virus, and of inhibitors of the p75NTR low affinity neurotrophin receptor.
We demonstrate potent activity of the TrkB/mAb for selectively agonizing the TrkB receptor, resulting in activation of predictive downstream markers and leading to ex vivo neuroprotection from mutHtt-mediated neurotoxicity. I will discuss testing this TrkB/mAb in vivo in HD rodent models.
I will also describe our current efforts in the intra-parenchymal delivery of AAV-BDNF to brain regions affected in HD as a separate therapeutic intervention strategy.
CHDI’s goal is to determine the suitability for these approaches as potential therapeutics, and to identify potential translational endpoints to facilitate clinical development.
1. Tsao, D., et al., 2008, Endocrin., 149(3): 1038-48
2. Lin, J., et al., 2008, PLoS ONE, 3(4):e1900
The Pursuit of Disease Biomarkers for HD: The Importance of Replication and Validation
Beth Borowsky, PhD – CHDI
Identifying and validating biomarkers for HD clinical trials are extremely important and challenging goals of the HD community. Biomarkers have been classified in multiple ways, but essentially they can be divided into two main clusters: disease-related biomarkers and drug-related biomarkers. Prospective, longitudinal, observational studies have helped inform us on potential disease-related biomarkers, including state and trait markers. Biomarkers of pharmacologic activity, sometimes called pharmacodynamic markers or mechanism of action markers, are important to confirm that a therapeutic agent reached its intended target and had a biochemical or physiological effect. Because the use of inappropriate biomarkers in clinical trials has the risk of both promoting the continuation of trials likely to fail (potential false positive signal) and the halting of trials that may have led to a positive outcome (potential false negative), potential new biomarkers must be evaluated using the most careful and rigorous scientific principles and analysis tools. In this talk, I will present results from several different approaches we have undertaken to identify and validate disease biomarkers for HD. Results will be presented from the 36-monthTRACK-HD data-cut, the development of a HD Cognitive Assessment Battery, and the evaluation of 8OHdG as a potential HD plasma biomarker. While the results show that we have several promising biomarkers to track disease progression, we do not know which, if any, of these will prove useful as markers to predict clinical efficacy of a drug candidate. I will introduce an upcoming clinical trial of aerobic exercise in HD which will be designed to assess the ability of our most promising cognitive, motor and imaging biomarkers to respond to an intervention.
Enroll-HD – A Hub for Biomarker Development
Tiago Mestre, MD – University of Toronto
The ability to detect disease changes in a sensitive and reproducible manner and to relate those changes with the effect of a compound is paramount for drug development. Biomarker discovery and qualification is fundamental in this endeavor.
ENROLL–HD is a global research platform built to facilitate clinical research by establishing a common infrastructure, reducing bureaucracy, fostering international collaboration, establishing common standards and facilitating the harmonization of best care practices. The most visible level of the platform ENROLL-HD is the global prospective observational international cohort study that integrates two existent HD registries (the COHORT study based in North America and Australia, and REGISTRY based in Europe), builds up from those two registries and involves other regions of the globe. As such, the ENROLL-HD is actively recruiting individuals who are HD gene expansion mutation carriers, regardless of symptomatic status, and controls that are both HD gene expansion negative individuals within HD families and non-blood related healthy individuals. The ENROLL-HD is collecting relevant clinical research data, with visits occurring annually, and promotes the additional completion of extended clinical assessments and bio-banking of biological specimens. The monitorization process will ensure that the data collected is correct and medically appropriate.
Another level the ENROLL-HD platform is the participation of independent HD researchers whose interests fall into the goals of ENROLL-HD, namely, biomarkers development and validation. ENROLL-HD also provides a flexible platform that, by protocol, welcomes the application of new internal studies in specific HD sub-populations and/or control populations with the goal of developing specific biomarkers or validating results from previous studies.
Examples of studies that will be typically conducted in the ENROLL-HD platform and are currently being initiated or in planning phase are: 1) Multiple tissue biopsy study, 2) striatal selective phosphodiesterase 10 PET study, and the 3) Kinemed© CSF study.
Alzheimer Prevention Initiative: A Landmark Trial for Prevention of Alzheimer Disease
Presentation not available
Carole Ho, MD – Genentech
Genentech, the Banner Alzheimer’s Institute, the University of Antioquia and the National Institutes of Health have partnered to collaborate on a historic prevention trial in cognitively healthy individuals who are likely to develop Alzheimer Disease (AD) due to their genetic history. This study will take place in Antioquia, Colombia and will involve about 300 participants from local families that share a rare genetic mutation that typically triggers AD symptoms around age 45. This autosomal dominant form of AD is transmitted through a mutation in presenilin 1, a protein that is important in the generation of toxic amyloid species.
This landmark trial will be one of the first to assess the potential of a therapeutic to stop Alzheimer Disease before it starts and is the cornerstone of a new international collaborative led by Banner Alzheimer’s Institute, the Alzheimer’s Prevention Initiative (API).
The therapeutic agent to be studied in this trial, crenezumab, is a humanized monoclonal antibody that binds to amyloid beta (Abeta), the main constituent of amyloid plaque in the brains of patients with AD. Abeta is proposed to be causative in the development of AD. Crenezumab was selected by an international Drug Selection committee convened by Banner Alzheimer’s Institute to be used in this study.
This trial represents a unique opportunity for a more definitive test of the amyloid hypothesis in an autosomal dominant cohort, where participants who are carriers have a near 100% certainty of disease. This study also has the potential to impact sporadic AD, by testing whether prevention of disease is possible and by qualifying sensitive biomarkers correlated with clinical benefit in pre-symptomatic disease. The learnings from this collaboration will be important for other diseases where genetic or other biomarkers can identify pre-symptomatic individuals, such as Huntington’s Disease.
PET and SPECT Imaging Biomarkers for HD: Developing a Translation Toolbox
Kenneth Marek, MD – Institute for Neurodegenerative Disorders
During the past decade Positron Emission Tomography (PET) and Single Photon Emission computerized tomography (SPECT) imaging tracers targeting striatal pathophysiology have provided increasing opportunities to probe neurodegenerative disorders with striatal pathology. Advances in PET and SPECT camera sensitivity coupled with the explosion of available novel tracers have enabled PET and SPECT imaging to be used in both non-human primates and in human studies in HD. Imaging biomarkers for HD have the potential to help understand disease mechanism, identify and track degeneration even before neurologic symptoms are present, and to evaluate potential therapies to assess target engagement as well as drug effect on slowing degeneration in study subjects.
Recent pre-clinical and clinical HD data have identified several potential PET and SPECT tracer targets of both disease pathology and potential therapeutics. In particular, consistent with the marked degeneration of striatal medium spiny neurons in HD, animal data has shown a reduction in PDE10, an enzyme highly expressed in these neurons. Recent PET imaging data (using 18F MNI-659) targeting PDE10 has similarly demonstrated a marked reduction in PDE10 expression in HD subjects. Moreover the reduction in PDE10 expression appears to be highly correlated with disease severity ranging from pre-manifest to mild to moderate HD. These data have suggested that 18F MNI-659 may be a potential biomarker of HD pathology and could potentially be utilized in clinical studies to monitor disease. In addition PET and SPECT tracers targeting the adenosine 2A receptor, the metabotropic GluR5 receptor, the Histamine 3 receptor and cannabanoid receptor 1 may also be utilized to provide further insight into the complex striatal pathology that occurs in HD.
PET and SPECT studies in HD subjects have the potential to both accelerate drug discovery and to track disease pathology. Initial studies targeting striatal pathology indicate that PET studies reflect expected disease pathology. While these studies are promising, longitudinal follow-up in subjects with a range of disease severity will be required to further examine the value of these markers in understanding disease and testing new HD therapies.