ALS Research Projects: Grants on the Rise

To date, the Robert Packard Center for ALS Research at Johns Hopkins has awarded 106 research grants and has several new grants pending, at a total cost of close to $21.2 million. The Center awards ALS research grants throughout the year on a flexible time frame, with the idea that rapid funding fosters innovative ideas. 

Borchelt | Brown | Cleveland | Cleveland | Farah | Gao | Gitler | Julien | Kabashi | Kaspar & Bergels | | Manfredi | Miller | P²ALS | Pandey | Robberecht | Taylor | Wang | Wong | Wong | Xu | Yang

David Borchelt, PhD

University of Florida
Propagation of toxic SOD1 conformations in ALS

 In many, if not most cases of ALS, the disease appears to spread along anatomically connected pathways. Whether this apparent spreading actually represents the physical spread of some entity that ultimately leads to loss of neuromuscular connectivity in life sustaining functions is presently unclear. We aim to investigate whether the spread of misfolded, toxic conformations of the superoxide dismutase 1 (SOD1) protein throughout the central nervous system has a role in the pathogenesis of ALS. We intend to use various techniques to first induce a focal accumulation of SOD1 and then determine whether these disease-specific proteins can propagate out from this point and spread within the spinal cord. These data will help to better understand the ALS disease process and may also have important therapeutic implications.

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Solange Brown, MD, PhD

Johns Hopkins University
Inhibitory Cortical Circuits in Presymptomatic ALS

One of the hallmarks of ALS is the death of upper motor neurons in the cerebral cortex. However, it is not known whether more subtle changes in cortical circuits precede upper motor neuron degeneration. We propose to study the excitatory and inhibitory neurons of the motor cortex, and their patterns of connectivity, to determine whether functional changes in these circuits represent early events in the development of the disease. The goal of this work is to identify primary events in the disease process that can suggest new preventative strategies and therapeutic targets in ALS. Furthermore, combining this work with the studies of lower motor neurons will give a fuller picture of why motor neurons are particularly vulnerable in ALS.

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Don Cleveland, PhD

Ludwig Institute for Cancer Research/UCSD
Transcriptome analyses on tissue from C9orf72 ALS patients

Funding for this project was made possible through a collaboration with Packard Center, Project ALS and P²ALS

We propose to use RNA sequencing approaches to determine whether expression of the C9orf72 expansion affects the RNAs within lumbar spinal cord in comparison to parallel analyses of normal spinal cords. We will identify (using DNA blotting) the length of the expansions in each of the three c9orf72 casesand determine whether there is a clear C9orf72 signature. Our RNA expression analysis will be done much as we have already done for RNAs from normal cells and tissues or those depleted of TDP-43 (Polymenidou et al, Nat. Neurosci. 2011). If C9orf72 signatures RNA signatures are identified, we would propose to extend this analysis to RNAs from the cortex and from cerebellum.

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Don Cleveland, PhD

Ludwig Institute for Cancer Research/UCSD
Mice expressing C9orf72 RNA for antisense oligonucleotide therapy development and decoding disease mechanism

Funding for this project was made possible through a collaboration between the Packard Center and ALSA

The most frequent cause of inherited ALS was discovered in October 2011 to be a large expansion of a hexanucleotide repeat in the C9ORF72 gene. When that large repeat is copied into the messenger RNA which carries the information encoded by the gene, the mRNA aberrantly accumulates. In other diseases caused by expanded repeats like myotonic dysrophy, factors required for processing the mRNA with the expansion – and needed for processing RNAs encoded by other genes – get trapped on the expanded repeat sequence, and correspondingly fail to mediate processing of the other RNAs. Building on that example and in partnership with Isis Pharmaceuticals, we have designed a gene silencing approach to develop a drug - called an antisense oligonucleotide or ASO - that will selectively destroy the ALS-causing mRNA with the expanded repeat. . We will build a mouse model expressing the expanded human C9ORF72 mRNA and use this to validate the efficacy of the ASO drug in development. The mice may also represent models that will develop ALS-like disease and which can be used to determine exactly what goes wrong in the presence of the aberrant mRNA.

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Mohamed H. Farah, PhD

Johns Hopkins University
New tools for the assessment of pathology and therapeutic responses in models of motor neuron diseases

This renewal project will continue the work of Packard scientist Dr. John W. (Jack) Griffin, who passed away untimely earlier this year. Dr. Griffin identified the lateral thoracic nerve/cutaneus maximus muscle (LTN/CMM; Griffin et al., 2010) as a system that has unique advantages for visualization and electrophysiological analyses of motor nerve and neuromuscular junction degeneration. We propose to carry out detailed morphological studies and electrophysiological analysis of the LTN/CMM system in a G93A SOD1 mouse at early stages of axon terminal degeneration. The goal is to thoroughly characterize the CMM/LTN system in SOD1 mouse model and identify the earliest degenerative events in order for the method to be widely adopted in future studies to aid in much higher throughput of pathogenetic analysis and therapy development in the field.

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Fen-Biao Gao, PhD 

University of Massachusetts
Identification of Genetic Modifiers of GGGGCC Repeats Toxicity in Drosophila

A repeat expansion in the C9ORF72 gene is the most common causal mutation in amyotrophic lateral sclerosis and frontotemporal dementia. However, no one knows exactly how this mutation causes disease. We will use Drosophila as a model system to investigate neurotoxicity of these repeat expansions. We hope these studies will contribute to the development of potential therapeutic interventions.

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Aaron Gitler, PhD

University of Pennsylvania
Novel Yeast and Zebrafish Models to Explore the Role of FUS/TLS in ALS

In motor neurons of both humans and many animal models of ALS, it's not uncommon to find abnormal clumps, or aggregations, of several proteins. Gitler and Granato propose to study the biology of one of them, a protein that's generated by the FUS/TLS gene. Using yeast cells as easy-to-study model systems, they hope to learn how and why the FUS/TLS protein forms clump within cells and how that's related to motor neuron death. Other work involves studying FUS/TLS in zebrafish, which are both fast-dividing and transparent, enabling easy study.

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Jean-Pierre Julien, PhD

Laval University, Quebec
Intrabody-based therapy for blocking TDP-43 interaction with NF-?B p65

We recently discovered that an upregulation of TDP-43 in ALS acts as co-activator of nuclear factor-?B (NF-?B), which can induce hyperactivity of NF-?B resulting in exaggerated innate immune responses and increased neuronal vulnerability to toxic environment. Here we propose to develop a therapeutic approach that uses intracellular antibodies (intrabodies) that work in the nucleus to block the specific interaction between TDP-43 and a small piece of NF-?B known as p65. First, we will use a hybrid of two different cell culture lines to generate monoclonal antibodies against the N-terminal domain and RRM1 domains of TDP-43. Then, from the appropriate hybrid cell lines we will derive small pieces of DNA to generate intrabodies using an adeno-associated virus to block TDP-43 and p65 interaction in the nucleus, thereby alleviating ALS.

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Edor Kabashi, PhD

Brain and Spinal Cord Institute, Paris
Identifying molecular partners of mutant TDP-43 & FUS

Our goal is to identify novel protein partners of mutant TDP-43 and FUS that could contribute to mechanisms associated with motor neuron degeneration using the transgenic zebrafish models we previously developed. Identification of specific protein partners that can selectively induce neurodegeneration has been impeded by the similar toxic profiles of TDP-43 and FUS. The discovery of these partners can shed light to a better understanding of neurodegenerative pathways elicited in ALS and may provide both novel avenues for genetic factors in motor neuron diseases and new targets for therapeutic intervention in ALS patients.

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Brian Kaspar, PhD & Dwight Bergles, PhD

Nationwide Children's Hospital & Johns Hopkins
Development of Gene Delivery to Oligodendrocytes

Funding for this project was made possible through a collaboration with P²ALS and ALSA

Inherited forms of amyotrophic lateral sclerosis (ALS) have been linked to mutations in genes that are widely expressed by both neurons and their supporting glial cells. In mouse models of ALS, deletion of mutant genes from glial cells can significantly prolong life, indicating that these non-neuronal cells are important contributors to this disease. Our recent studies in G93A SOD1 mice show that oligodendrocytes, a class of glia that form myelin sheaths around axons, degenerate in the spinal cord, and that selective removal of mutant SOD1 from oligodendrocytes prolongs life. These studies indicate that oligodendrocytes are an important target for therapeutic manipulation in vivo. Although it is possible to manipulate gene expression in oligodendrocytes in mice using complex transgenic manipulation, such approaches are not applicable for therapeutic intervention in humans. We intend to use a new technique known as molecular or directed evolution to engineer an adeno-associated virus (AAV) that will readily infect oligodendrocytes but not other cells. Generation of this vector will help define the mechanisms responsible for oligodendrocyte degeneration in ALS, and enable development of new strategies to minimize the toxic effects of these mutant proteins in vivo.

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Giovanni Manfredi, MD, PhD

Weill Medical College, Cornell University
Mitochondrial involvement in TDP-43 mutant mice

Mitochondria provide neurons with the energy necessary for their function and survival. In ALS, mitochondria degenerate, and fail to provide adequate energy supplies to cells, thereby participating to their death. Our current knowledge on mitochondrial dysfunction in ALS is mostly limited to familial forms of the disease with SOD1 mutations, but new evidence links mitochondrial abnormalities with another form of familial ALS associated with TDP-43 mutations. In this application, we propose to apply our expertise on mitochondrial biology to understanding how mitochondrial dysfunction is involved in the disease, in a TDP-43 transgenic mouse model. We will characterize the bioenergetics and dynamics of neuronal mitochondria and their ability to control the levels of calcium, a fundamental ion that regulates vital cellular functions.

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Timothy Miller, MD, PhD

Washington University in St. Louis
Determining and blocking miRNA changes in activated microglia in vitro and in ALS mice

Cells in the spinal cord known as microglia can become activated during amyotrophic lateral sclerosis. These cells are also involved in inflammation and appear to accelerate disease progression. We have determined part of the genetic program underlying the activation of inflammatory pathways in microglia. We will attempt to block this genetic program as a way to slow down progression of disease in an animal model of ALS, and then translate this type of therapy to treatment of human ALS.

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Udai Pandey, PhD

Louisiana State University
A Drosophila Model to Investigate the Role of FUS in ALS

ALS is marked by abnormal accumulations of a protein named FUS/TLS. In creating fruit flies (Drosophila melanogaster) that carry the human FUS/TLS gene, we've found an economical, easily-studied model of ALS. Our goal with this Packard grant is to understand how the FUS/TLS gene promotes the disease in Drosophila. What we discover, we assume, will shed clearer light on both familial and sporadic ALS in patients.

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Wim Robberecht, MD, PhD

University of Leuven, Belgium
Role of Oligodendrocytes in the Pathogenesis of Mutant TDP-43 Associated ALS

Oligodendrocytes have recently been shown to contribute to the pathogenesis of ALS. TDP-43 plays a role both in familial and sporadic ALS. In this project, we intend to investigate whether expression of mutant or wild-type TDP-43 in oligodendrocytes or in oligodendrocyte precursor cells is sufficient to cause motor neuron degeneration. We also intend to compare the potential motor neuron degeneration in oligodendrocytes to the phenotype induced by selective expression of mutant or wild-type TDP-43 in motor neurons.

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J. Paul Taylor, MD, PhD

St. Jude Children's Research Hospital
The role of TDP-43 in RNA transport granules and the impact of disease mutations

Previous studies have shown that TDP-43 is part of the protein and nucleic acid complexes that transport molecules from the nucleus to the cytoplasm. Mutations to TDP-43 affect the cell's ability to move molecules into the cytoplasm. We hypothesize that these TDP-43 granules facilitate delivery and local translation of mRNAs in motor neurons. Specifically, we are studying the function of several proteins implicated in ALS (TDP-43, hnRNPA2B1 and hnRNPA1) that bind to mRNA immediately after it is transcribed by ribosomes. We intend to discover how these proteins are directly perturbed by disease mutation or indirectly impacted by mutations in VCP or C9orf72. This project is one of four thematically related projects investigating the role of perturbed RNA metabolism in ALS pathogenesis.

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Jiou Wang, MD, PhD

Johns Hopkins University
Using C. elegans to Investigate Molecular Mechanisms of ALS

Funding for this project was made possible by the Muscular Dystrophy Association

Our laboratory seeks to use the small, soil-living, transparent roundworm – Caenorhabditis elegans – as a model organism to clarify how disease genes cause ALS in humans. Like the fruitfly, C. elegans is used by researchers to investigate how both healthy and mutated genes affect function. With its fast growth rate and the amount of knowledge we already hold about its genome, the tiny animal – it's about the width of a pencil lead – provides a unique system to study aging related diseases.

Recently, we genetically engineered C. elegans to model one specific form of inherited ALS (that with mutant SOD1 genes). Using the C. elegans model, we were able to conduct unbiased, large-scale screens to search for uncommon genes that might slightly raise the risk of having the disease. Now we aim to establish new C. elegans models of ALS linked to those mutations in patients. We hope that our studies will reveal important disease pathways in humans, and thus provide potential drug targets.

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Philip C. Wong, PhD

Johns Hopkins University
Determining the pathogenic mechanism(s) of C9ORF72-linked ALS-FTD

Funding for this project was made possible through a collaboration between the Packard Center and ALSA; Packard Center funding made possible by A Midwinter Night's Dream

The hexanucleotide GGGGCC repeat expansion in a gene termed C9ORF72 is the major cause of ALS and FTD, but how such expansion causes malfunction of nerve cells in these illnesses remains unknown. We plan initially to develop mouse model systems to test if losing the activity of C9ORF72 is responsible for death of motor neurons and secondly, whether toxicity arises through the presence of a toxic RNA derived from the diseased C9ORF72 gene that determines malfunction of motor nerve cells. Outcomes from studies proposed in this application will clarify how the repeat expansion in the C9ORF72 gene causes motor nerve cell loss in a large proportion of cases of ALS. These efforts will have important implications for design of therapy and provide useful mouse model systems for testing therapies that would eventually benefit patients with ALS.

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Philip C. Wong, PhD

Johns Hopkins School of Medicine
Generation and Characterization of Wildtype and ALS-linked FUS/TLS mice

Funding for this project was made possible by the Muscular Dystrophy Association

The discovery of mutations in a gene encoding an RNA binding protein (FUS/TLS) linked to both sporadic and familial ALS provides the opportunity to further our understanding of the underlying cause of this illness. In this proposal, we plan to study the effects of an ALS-associated mutant FUS/TLS protein in transgenic mice and identify any resulting motor neuron disease. If disease results, this mouse model will help clarify the behavioral and pathological signs associated with ALS onset and disease progression. These transgenic mice will also help with the design of therapeutic strategies to attenuate this devastating disease.

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Zuoshang Xu, PhD

University of Massachusetts
Modeling ALS by TDP-43 hypomorphism in mice

TDP-43 is a key causal gene that is involved in a majority of ALS cases, including the sporadic and some familial cases. To understand the role of TDP-43 in neurodegeneration and to design therapeutic strategies, a critical question to resolve is whether TDP-43 causes neurodegeneration by a gain of toxicity, e.g. from forming toxic protein aggregates, or from loss of its function, e.g. from mutations or being absorbed into the aggregates. To answer this question, both overexpression and loss-of-function models are necessary. This proposal will establish a transgenic mouse model for ALS by decreasing the level of TDP-43. This model will be important for understanding the disease mechanism and designing therapeutic strategies.

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Yongjie Yang, PhD

Tufts University
Role of Astrocyte Exocytosis in the Pathogenesis of ALS

This project aims to investigate the role of chemicals secreted by astrocytes in pathogenesis of motor neuron degeneration by employing novel mouse genetic tools. Results from this study will demonstrate whether these secreted compounds contribute to toxicity in astrocytes with a mutation in SOD1 and provide clues how to identify the toxic factors from astrocytes. Ultimately these results will help develop astrocyte-based neuroprotective strategies against motor neuron degeneration in ALS.

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