1985 — 1989 |
Siddique, Teepu |
K07Activity Code Description: To create and encourage a stimulating approach to disease curricula that will attract high quality students, foster academic career development of promising young teacher-investigators, develop and implement excellent multidisciplinary curricula through interchange of ideas and enable the grantee institution to strengthen its existing teaching program. |
Recombinant Dna Strategies--Duchenne Muscular Dystrophy
Duchenne muscular dystrophy (DMD) is the most frequent of the childhood onset muscular dystrophies. Determination of the DMD gene(s) would allow the development of rational therapy. In addition, DNA restriction fragment length polymorphisms (RFLPs) closely linked to DMD gene(s) can be used in genetic counseling and prenatal diagnosis. The specific aims would be to identify such probes of cloned DMD gene region (Xp2.1) or closely linked RFLPs. Identification and subsequent successful cloning of DMD gene could lead to the identification of the defective gene product and its role in functioning of the dystrophic muscle cell. DMD is an X-linked disease. Females with DMD are rare and, except those with XO configurations, have balanced X chromosome translocations with the breakpoint at Xp2.1. DNA from transformed lymphoid line (Fra 21) of one such rare female patient having a balanced chromosome translocation (X:21) (p2.1:p1.2) is available in our laboratory. Lambda phage libraries prepared from total genomic Fra 21 DNA and from bivariantly flow sorted X-derivative (X der) and 21 derivative (21 der) chromosomes will contain translocation junction or breakpoint clones. Since both the X der and 21 der stain at the breakpoint with silver, indicating the presence of ribosomal DNA (rDNA), cloned 18S and 28S DNA probes in plaque lift experiments will identify breakpoint clones. The rDNA clones containing breakpoint portions of X chromosome will be selected by in situ hybridization and by dot blot analysis of cell lines with variable number of X chromosomes. Unique DNA sequences from these clones will serve in RFLP segregation analysis of multigeneration DMD and Becker muscular dystrophy (BMD) families which are available to us. Unique sequence RFLPs, adjacent to and overlapping breakpoint sequences, will be generated using appropriate restriction enzymes. These will then be screened against human liver and skeletal muscle cDNA libraries (available in our lab) to define Xp2.1 specific clones. Using hybridization and translation methodologies, Xp2.1 specific clones can be utilized to detect differences in specific transcripts and protein products in normal and dystrophic muscle.
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0.97 |
1993 — 1998 |
Siddique, Teepu |
P01Activity Code Description: For the support of a broadly based, multidisciplinary, often long-term research program which has a specific major objective or a basic theme. A program project generally involves the organized efforts of relatively large groups, members of which are conducting research projects designed to elucidate the various aspects or components of this objective. Each research project is usually under the leadership of an established investigator. The grant can provide support for certain basic resources used by these groups in the program, including clinical components, the sharing of which facilitates the total research effort. A program project is directed toward a range of problems having a central research focus, in contrast to the usually narrower thrust of the traditional research project. Each project supported through this mechanism should contribute or be directly related to the common theme of the total research effort. These scientifically meritorious projects should demonstrate an essential element of unity and interdependence, i.e., a system of research activities and projects directed toward a well-defined research program goal. P50Activity Code Description: To support any part of the full range of research and development from very basic to clinical; may involve ancillary supportive activities such as protracted patient care necessary to the primary research or R&D effort. The spectrum of activities comprises a multidisciplinary attack on a specific disease entity or biomedical problem area. These grants differ from program project grants in that they are usually developed in response to an announcement of the programmatic needs of an Institute or Division and subsequently receive continuous attention from its staff. Centers may also serve as regional or national resources for special research purposes. |
Identification of Gene For Familial Als On Chromosome 21 @ Northwestern University
We plan to fine-map and identify a gene on Chromosome 21 that causes familial amyotrophic lateral sclerosis (FALS). Defining the precise location of this gene will allow us to isolate it and study its function. Our mapping strategy is fourfold: (1) We will generate highly polymorphic DNA markers (such as (CA)n repeat(s) around the putative locus of the FALS gene on Chromosome 21 from previously mapped cosmid and yeast artificial chromosomes markers. These markers will be tested by linkage analysis in our FALS families in order to obtain the optimum genetic information from them. Crossovers between the disease (FALS) locus and the markers will help define the precise location of the FALS gene. (2) Once flanking markers are defined we will construct a band- specific genomic library by microdissection of chromosome 21. (3) We will test candidate FALS genes in our FALS families that are obtained from (a) spinal cord cDNA library that has been screened against the microdissected chromosome 21q band-specific library and (b) from subtracted cDNA libraries (made from the anterior and posterior gray matter as motor neurons undergo the most profound pathological change and are therefore prime candidate for expression of the FALS gene. Thus, if the FALS gene is represented in our clones, it should map to chromosome 21 and an appropriate polymorphism derived from it should segregate with the disease trait in our FALS families. This may provide a direct approach to the cloning of the FALS gene on Chromosome 21. (4) We have identified a Downs patient with an amyotrophic lateral sclerosis-like syndrome. This patient is a mosaic for trisomy 21, (46XY, normal 21 and marker 21) and a deleted chromosome 21 (46XY, normal 21 and (del)21). We plan to determine the extent of deletion in the deleted chromosome 21 as this may define the limits of the region containing the FALS gene. Yeast artificial chromosomes and cosmids from this region can then be used to generate markers for testing in our families. this robust approach in mapping the chromosome 21 FALS gene will allow its isolation and lead to an understanding of the fundamental defect in the degeneration of motor neurons. This information will be especially useful in formulating a rational treatment of this and related disorders that are otherwise fatal.
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1 |
1993 — 1997 |
Siddique, Teepu |
P50Activity Code Description: To support any part of the full range of research and development from very basic to clinical; may involve ancillary supportive activities such as protracted patient care necessary to the primary research or R&D effort. The spectrum of activities comprises a multidisciplinary attack on a specific disease entity or biomedical problem area. These grants differ from program project grants in that they are usually developed in response to an announcement of the programmatic needs of an Institute or Division and subsequently receive continuous attention from its staff. Centers may also serve as regional or national resources for special research purposes. |
Gene Mapping in Motor Neuron Disorders
We propose to map and identify genes in disorders of the motor neuron. We will use genetic linkage analysis to identify the chromosome localization of the genes for: (1) autosomal dominant form of familial amyotrophic lateral sclerosis in those (30% - 60%) families where the gene for this disease is not linked to chromosome 21. (2) Autosomal recessive and autosomal dominant forms of pure spastic paraplegias. Our laboratory is well suited for this task as we have DNA samples from families with these disorders and have recently identified chromosome loci for the autosomal dominant and autosomal forms of amyotrophic lateral sclerosis on chromosome 21q and 2q respectively. In addition we have preliminary evidence for linkage of recessive familial spastic paraplegia to chromosome 8. We plan to identify the gene for recessive amyotrophic lateral sclerosis on chromosome 2q. We will use genetic linkage techniques to fine map this gene, then use molecular techniques to obtain a physical map of that region and then identify candidate genes for this disease. These techniques include identifying Yeast artificial chromosomes that map to this region which will be used along with a microdissected chromosome 2 library to find closely linked polymorphic markers. Once markers within a million base pairs or so of the gene are obtained, it will be possible to prepare a physical (YAC/cosmid and pulsed field) map of the region based. Potential candidate genes can then be identified and should be contained in the microdissected library or in appropriate yeast artificial chromosome clones. cDNA libraries from the spinal cord will also be screened with clones from regional genomic libraries prepared from microdissected chromosome 2 to obtain candidate genes. Candidate genes can then be tested for their relevance by linkage analysis in our recessive ALS families. Similar techniques will be used to identify genes for amyotrophic lateral sclerosis (non chromosome 21) and the spastic paraplegias. Once the location of those genes are determined, identification of these genes and their function will allow the formulation of rational treatment and or prevention of those disorders.
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0.948 |
1993 — 1998 |
Siddique, Teepu |
P01Activity Code Description: For the support of a broadly based, multidisciplinary, often long-term research program which has a specific major objective or a basic theme. A program project generally involves the organized efforts of relatively large groups, members of which are conducting research projects designed to elucidate the various aspects or components of this objective. Each research project is usually under the leadership of an established investigator. The grant can provide support for certain basic resources used by these groups in the program, including clinical components, the sharing of which facilitates the total research effort. A program project is directed toward a range of problems having a central research focus, in contrast to the usually narrower thrust of the traditional research project. Each project supported through this mechanism should contribute or be directly related to the common theme of the total research effort. These scientifically meritorious projects should demonstrate an essential element of unity and interdependence, i.e., a system of research activities and projects directed toward a well-defined research program goal. P50Activity Code Description: To support any part of the full range of research and development from very basic to clinical; may involve ancillary supportive activities such as protracted patient care necessary to the primary research or R&D effort. The spectrum of activities comprises a multidisciplinary attack on a specific disease entity or biomedical problem area. These grants differ from program project grants in that they are usually developed in response to an announcement of the programmatic needs of an Institute or Division and subsequently receive continuous attention from its staff. Centers may also serve as regional or national resources for special research purposes. |
Identification and Transgenic Studies of Als Genes @ Northwestern University
The aim of this program project is to identify the gene for familial amyotrophic lateral sclerosis (ALS) and develop a transgenic model so as to be able to study the disease process in motor neuron disorders and test treatment modalities. Familial ALS is a genetic model of ALS, which is a degenerative disorder affecting the upper and lower motor neuron. ALS results in progressive paralysis and death. The cause of ALS is presently unknown, nor is there a treatment that will alter the course of the disease. This project will bring together investigators that linked a gene for familial ALS to chromosome 21. Investigators in the two laboratories will study the mapping and identification of the ALS gene in two separate and non overlapping subsets of familial ALS families. The mapping resources available to both centers will be utilized and exchanged. Polymorphic markers will be developed to narrow the gene region between 1 to 3 centiMorgans by linkage analysis. Current polymorphic markers and other resources being developed to map chromosome 21 will allow physical mapping of this interval. Genes expressed in the spinal cord that map to this interval will be used as candidate genes. Once the FALS gene is identified, tissue of individuals affected with familial ALS will be examined for gene mutations. Simultaneously with the mapping studies we will concentrate on developing a transgenic mouse model that will have specific gene expression in motor neurons. A transgenic model of motor neuron disease will also be developed. when the familial ALS gene is isolated, these transgenic models will allow the study of the ALS disease process and provide a vehicle for the design of experimental treatment in motor neuron disorders.
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1 |
1998 — 2002 |
Siddique, Teepu |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Multicenter Clinical Study of Als Genetics @ Northwestern University
The goal of this multicenter study is to identify genetic risk factors that predispose to amyotrophic lateral sclerosis (ALS). Our focus will be to identify risk genes that predispose to sporadic ALS or ALS observed in small families and familial aggregates. We will also continue our collaborative study of dominantly inherited familial ALS. Our primary hypothesis is that multiple genetic factors predispose to the development of ALS, and our specific aims of this multicenter study will test this hypothesis. We aim to: (i) Improve our resources for genetic analysis by collecting clinical information and DNA from sporadic ALS patients and their parents and from familial ALS families. (ii) Test certain genes that are hypothesized to predispose to ALS, in sporadic cases of and in small familial and in familial aggregates (iii) Search for gene loci in small familial aggregates, unsuitable for conventional parametric linkage analysis by, non-parametric linkage technique. (iv) Use parametric linkage analysis to search for gene linkage in large familial ALS families that do not have mutations in the gene for Cu,Zn superoxide dismutase. This is the first large-scale study of genetic risk factors in ALS, and is only possible because of collaborative arrangement between four centers, Northwestern University Medical School, Massachusetts General Hospital, Duke University Medical Center and Vanderbilt University Medical Center. This study will identify additional genes that cause or predispose to ALS, and contribute to the understanding of the biology of the motor neuron. This study will be a major contribution to our ultimate goal, which is to develop rational therapy for ALS based on the causes and mechanism of disease.
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1 |
1998 — 2002 |
Siddique, Teepu |
P01Activity Code Description: For the support of a broadly based, multidisciplinary, often long-term research program which has a specific major objective or a basic theme. A program project generally involves the organized efforts of relatively large groups, members of which are conducting research projects designed to elucidate the various aspects or components of this objective. Each research project is usually under the leadership of an established investigator. The grant can provide support for certain basic resources used by these groups in the program, including clinical components, the sharing of which facilitates the total research effort. A program project is directed toward a range of problems having a central research focus, in contrast to the usually narrower thrust of the traditional research project. Each project supported through this mechanism should contribute or be directly related to the common theme of the total research effort. These scientifically meritorious projects should demonstrate an essential element of unity and interdependence, i.e., a system of research activities and projects directed toward a well-defined research program goal. |
Genetic Studies of Motor Neuron Disease
Familial amyotrophic lateral sclerosis (FALS) and 'pure' familial spastic paraparesis (FSP) are prototypic disorders of motor neuron degeneration that affect both the upper and lower motor neurons (FALS) or primarily the upper motor neuron (FSP). These phenotypes of motor neuron degeneration are genetically heterogeneous. The long term goal of my laboratory is to identify the genetic contribution to the pathogenesis of these disorders in order to formulate interventions based on disease mechanism to prevent, postpone and treat these disorders. In this proposal we propose to (1) map the gene(s) for (i) FALS in large dominant FALS families that do not have mutations in the gene for SOD1 (Cu, Zn superoxide dismutase) and (ii) in large dominant 'pure' FSP families that do not have mutations in the gene for SOD1 (Cu, Zn superoxide dismutase) and (ii) in large dominant 'pure' FSP families that do not map to 2p, 14q or 15q (2) further refine the physical map around the 2p locus of dominant 'pure' FSP and the 2q33 and the newly discovered, 15q loci of autosomal recessive familial ALS, and (3) clone the genes responsible for the disorders at those loci. We will use our newly acquired automated genotyping setup to rapidly screen banked DNA from FALS and FSP families and assign gene loci using genetic linkage techniques. Simultaneously, we will test known genes and full length cDNA sequences of Expressed Sequence Tags (ESTs) that physically map to the loci for dominant FSP on 2p and/or cosmids mapped to YAC contigs on these regions. Once the non-SOD1 dominant FALS loci or additional FSP loci are identified, we will proceed to develop refined maps of loci, and utilize physical mapping and cloning techniques similar to those described above. Thus, such am assembly of genes that cause motor neuron degeneration will provide a crucial resource to search for points at which pathways leading from these genes converge in the pathogenesis of neuronal degeneration.
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0.948 |
2003 — 2007 |
Siddique, Teepu |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Genes and Genetic Models in Motor Neuron Disorders @ Northwestern University
DESCRIPTION (provided by applicant): Familial amyotrophic lateral sclerosis (FALS), 'pure' hereditary spastic paraparesis (HSP) and primary lateral sclerosis (PLS) are genetically heterogeneous disorders caused by motor neuron degeneration. These disorders either affect, both the upper and lower motor neurons (FALS), or primarily the upper motor neuron (HSP and PLS). The long-term goal of my laboratory is to identify the genetic contribution to the pathogenesis of these disorders. Success in this goal will lead to the formulation of interventions based on disease mechanisms to prevent, postpone and treat these and related disorders. We now plan to build on the work we accomplished in the last five years as part of a program project (NS 21442). We fulfilled all the Specific Aims such that we identified the ALSIN gene, that cause both recessive FALS (ALS2) and recessive juvenile PLS (JPLS1), on chromosome 2q33, studied mutations in SPASTIN that cause HSP (SPG5), and identified new loci for dominant ALS (on the X-chromosome) and ALS/dementia on chromosome 9q21-q22. We will further expand our gene discovery effort in recessive ALS and recessive HSP develop genetic models of FALS, JPLS, and HSP, and interrogate the products of causative genes for their protein-protein interactions to trace their signaling pathways. We plan to accomplish these goals by three main Specific Aims (1) Identify the genes for a more common form of recessive FALS (ALS5) on chromosome 15q15-q21 and a common form of recessive HSP (SPG5A) on chromosome 8q. (2) Develop genetic models of alsin for the JPLS and ALS phenotypes and of spastin for the HSP phenotype (SPG4). (3) Identify protein interactions of alsin. In the previous period we narrowed the loci for ALS5 and SPG5A. We will use high throughput sequencing and bioinformatic support to identify these genes, as we successfully did the case of the ALS2 gene. We will make knockout models for alsin and spastin, to study the pathology and pathogenesis of these disorders. Finally, the interacting partner proteins of alsin and subsequently of spastin will be interrogated by the two-hybrid systems and immunoprecipitation. Interaction will be confirmed by dual labeled confocal microscopy and FRET analysis.
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1 |
2004 |
Siddique, Teepu |
R13Activity Code Description: To support recipient sponsored and directed international, national or regional meetings, conferences and workshops. |
Consensus Conference On Primary Lateral Sclerosis @ Northwestern University
Primary lateral sclerosis (PLS) is a rare neurological disorder due to degeneration of tracts that connect neurons originating in the brain's cortex to motor neurons in the brainstem and spinal cord. Except for an extremely rare genetic form (Yang et al 2002), its causes are not known and its treatments are marginally effective. Diagnosis of PLS has been a subject of much debate over the years because the signs and symptoms of PLS may be seen in other disorders as well, particularly amyotrophic lateral sclerosis (ALS), the spastic parapareses (SP) and multiple sclerosis (MS). Although several sets of diagnostic criteria have been proposed (Charcot 1865, Stark and Moersch 1945, Pringle et al 1992, Brooks 1994), none were developed as the result of consensus among practitioners and researchers with expertise in sub-specialties to each contribute unique knowledge and apply current technologies to defining this disease. Additionally, present criteria are loosely and quite often inaccurately applied, even by neurologists, producing a confusing and frustrating experience for patients, and making meaningful clinical research impossible. Our long-term goal is to accurately identify PLS patients so meaningful clinical research can be undertaken to establish the cause(s) of PLS and develop effective treatments. As a first step, we plan to convene a group of 50-100 neuroscientists, on June 4-6, 2004 at the Chaminade Resort, Santa Cruz, CA, with expertise in clinical neurology, imaging, neuropathology, genetics and neurophysiology to accurately define PLS and chart areas for meaningful research. Our specific aims are (1) To define Primary lateral sclerosis (PLS) by: clinical, neurophysiological, imaging, neuropathological, and genetic criteria. (2) Delineate the relationship and differences between PLS and ALS, SP, and other spasticity causing diseases. (3) Establish future directions for PLS in clinical research, mouse models, and molecular-based research. (4) Publish a summary of the conference proceeding and consensus criteria for delineating PLS as a specific entity and its relationship to other motor neuron disorders in a major journal.
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1 |
2004 — 2008 |
Siddique, Teepu |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Genetics of Als @ Northwestern University
The goal of this Multicenter ROt is to identify genetic risk factors that predispose individuals to amyotrophic lateral sclerosis (ALS). This RO1 is a continuation of collaborative efforts that have been ongoing since 1993, when this group identified the first ALS gene (the SOD 1 gene). Our Amyotrophic Lateral Sclerosis Genetics Collaborative Group (ALSGCG) has greatly extended the genetic knowledge of ALS by identifying loci on chromosomes 2, 9, 15, 16, 18, 20 and X using large autosomal dominant families; we have generated preliminary data for additional loci on chromosomes 1, 2, 7, 8, 17, 18, and 19 in smaller multiplex families. The ALSGCG represents the collaborative efforts of groups at Northwestern University Feinberg School of Medicine (NUFSM), Massachusetts General Hospital (MGH), Duke University Medical Center (DUMC), and Vanderbilt University Medical School (VUMS). We have collected the largest set of multiplex and singleton resources in the world and now have sufficient family material to begin gene discovery within these regions and test candidate genes as they become available. This RO1 is carried out at four sites. Site 1 (NUFSM) will use the locational candidate approach to identify the genes arising in the smaller multiplex families. This will include refining the linkage peaks using linkage analysis and testing candidate genes using allelic association studies. Site 2 (MGH) will use the locational candidate approach to identify the genes arising in the large autosomal dominant families using approaches similar to Site 1. It will also test the potential role of other motor neuron disease genes in ALS using an allelic association approach. The sites are supported by an Administrative Assistant from Northwestern, and a Genetic Epidemiology and Bioinformatics supported programs at DUMC and VUMS. Analysis of all data from the projects, including clinical, risk factor, pedigree, and genotypic data for NUFSM (Site 1) and MGH (Site 2) is carried out at DUMSC and VUMS analyses sites. Those also provide all linkage and allelic association analyses for Sites 1 and 2.
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1 |
2006 — 2008 |
Siddique, Teepu |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Effect of Variation in Genes of Xenobiotic Responsive Proteins in Als @ Northwestern University
[unreadable] DESCRIPTION (provided by investigator): The goal of this project is to identify variants in environmentally responsive genes that modulate the genetic susceptibility of individuals to amyotrophic lateral sclerosis (ALS). Considerable progress has been made in identifying genes (SOD1 and ALSIN) and loci (chromosomes 9, 15, 16, 18, 2 and x) in the familial form of ALS but the etiology of sporadic ALS (SALS) remains unknown. To address the genetic contribution to SALS we have collected a large set of trios, case-control and discordant sib-pairs resources and now have sufficient material to test specific hypothesis. To fulfill our goal we will initially use a functional candidate approach to screen genetic variants in 410 environmentally responsive genes in two cohorts of ALS patients and controls. The associations detected with ALS will be verified in a second cohort of 500 case-controls subjects. We have used bioinformatics tools to mine a fairly comprehensive list of environmentally responsive genes from several databases including those identified by the NIEHS. We will screen variants (SNP's) in these genes using a high-throughput genotyping system and examine the SALS susceptibility and phenotype for allelic association to these genes. The initial screen will employ 250 trios and 500 case controls subjects. Promising candidates will be validated using a second set of 500 cases and controls and in discordant sibpairs. Furthermore, we will examine the genes associated with SALS for gene-gene and gene-environment interactions. Identification of gene variants associated with SALS or with the SALS sub-phenotypes (e.g. Age-at-onset, site-of-onset, bulbar/spinal etc.) will allow us to test genes in the entire pathway to which the candidate gene belongs. This project will therefore allow a focused study of environmentally responsive genes using a large resource of family and singleton ALS cohorts and will generate resources to investigate new approaches to ALS pathogenesis and in formulating rational treatment. The causes of sporadic ALS are largely unknown, and rational therapy is therefore not possible. A genetic susceptibility to environmental toxicity is considered to underlie ALS. We have a large cohort of ALS patients, siblings, parents and control subjects. We will use this unique resource to analyze the association of DNA variations in genes involved in detoxification of environmental toxins to ALS, and test the identified genes for their interaction with environmental exposures. This study will open fresh ground to investigative mechanisms of disease in ALS for formulation of rational treatment. [unreadable] [unreadable] [unreadable]
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1 |
2009 — 2010 |
Siddique, Teepu |
P01Activity Code Description: For the support of a broadly based, multidisciplinary, often long-term research program which has a specific major objective or a basic theme. A program project generally involves the organized efforts of relatively large groups, members of which are conducting research projects designed to elucidate the various aspects or components of this objective. Each research project is usually under the leadership of an established investigator. The grant can provide support for certain basic resources used by these groups in the program, including clinical components, the sharing of which facilitates the total research effort. A program project is directed toward a range of problems having a central research focus, in contrast to the usually narrower thrust of the traditional research project. Each project supported through this mechanism should contribute or be directly related to the common theme of the total research effort. These scientifically meritorious projects should demonstrate an essential element of unity and interdependence, i.e., a system of research activities and projects directed toward a well-defined research program goal. |
Envronment-Sensitive Genes in Motoneuron Degeneration @ Northwestern University At Chicago
DESCRIPTION (provided by applicant): The goal of this project is to identify functional variants of environmentally responsive genes that are biologically relevant to motor neuron degeneration of sporadic amyotrophic lateral sclerosis (SALS) and primary lateral sclerosis (PLS). Considerable progress has been made in inherited motor neuron degeneration where genes such as SOD1 and ALSIN have been discovered and suitable animal models engineered. However etiology of SALS and PLS remains undefined. Both are complex disorders and a genetic and it is widely accepted susceptibility encoded in the genome interacts with environment stressors to produce motor neuron degeneration. These investigators have recently identified two sets of genes of environmentally responsive enzymes that are etiologically relevant to motoneuron degeneration. The PON cluster of genes (PON1, 2 and 3) was associated with SALS and this finding has been validated in four other studies. Mutations in the second gene, CYP7B1, are caused in corticospinal degeneration in a form of hereditary spastic paraplegia (SPG5A). PONs are antioxidant and detoxify pesticides, while CYP7B1 is inhibited by agricultural fungicides. A detailed examination of variations in these genes and in genes related to them by metabolic pathways and coexpression for association to SALS and PLS will be accomplished. Further, their interactions with environmental exposure to pesticides, fungicides and other relevant stressors will also be examined. This is a novel opportunity for a comprehensive study of two etiologically relevant environmentally responsive genes in SALS and PLS. 1000 ALS cases, 400 PLS cases and 1000 matched controls will participate in this study. This investigation will establish genetic and environmental risks, and lead to rational treatment and prevention of disabling fatal disorders of PLS and SALS.
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1 |
2012 |
Siddique, Teepu |
R56Activity Code Description: To provide limited interim research support based on the merit of a pending R01 application while applicant gathers additional data to revise a new or competing renewal application. This grant will underwrite highly meritorious applications that if given the opportunity to revise their application could meet IC recommended standards and would be missed opportunities if not funded. Interim funded ends when the applicant succeeds in obtaining an R01 or other competing award built on the R56 grant. These awards are not renewable. |
Role of High Density Lipoprotein Particles in Amyotrophic Lateral Sclerosis @ Northwestern University At Chicago
DESCRIPTION (provided by applicant): Amyotrophic lateral sclerosis (ALS) or Lou Gehrig disease is a fatal neurodegenerative disease that primarily affects mid-life and older adults. There are two forms of disease, familial (FALS) and sporadic (SALS), respectively comprising 10% and 90% of cases, respectively. The genetic causes of FALS have been linked to mutations in several genes such as superoxide dismutase, TDP-43, FUS, and optineurin. The etiology of SALS, however, remains elusive. A few years ago our laboratory found that there were polymorphisms in genes for enzymes called paraoxonases that were associated with SALS. Because these enzymes detoxify certain pesticides and toxic agents, they became the first environmentally related genes that were linked to ALS. Further studies of the paraoxonase enzymes PON1 and PON3 in the plasma indicate that their levels are significantly elevated in SALS patients, although the activities were similar. PON1 and 3 are found on high density lipoprotein (HDL) particles that contain multiple proteins with several functions including lipid and cholesterol transport and protecting lipoproteins from deleterious oxidation. Similar particles are also found in the cerebrospinal fluid (CSF). Thus, the objectives of this proposal are to further characterize HDL-particles in the plasma and CSF of SALS patients using high throughput technologies that can quantify the levels of multiple proteins present in these particles. Thus, we can determine for the first time how HDL particle composition changes in a neurodegenerative disease and if they are causally linked to the disease process. We will also determine whether the genes for the HDL-associated proteins contain variants that are associated with risk of SALS and whether these changes are related to alterations in HDL particle composition. Using advanced gene targeting techniques, we will create ApoL1 transgenic model mice to study the effects of over expressing this HDL component. The results from this work will open paths to therapies that seek to rescue potential dysfunctional HDL states found not only in neurodegenerative disease such as ALS but also in more common cardiovascular and metabolic diseases.
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1 |
2012 — 2016 |
Siddique, Teepu |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Disease Mechanisms in Human Ubiquilinopathy @ Northwestern University At Chicago
DESCRIPTION (provided by applicant): Amyotrophic lateral sclerosis (ALS) is characterized by degeneration of large motor neurons in the brain and spinal cord, resulting in progressive wasting and paralysis of voluntary muscles, respiratory failure and ultimately death. Familial ALS (FALS) accounts for around 10% of all ALS cases, with the majority (90%) of the disease presenting as sporadic ALS (SALS). Discovery of genetic etiologies of ALS has been the driving force in ALS research for the last 20 years. These discoveries have provided fresh insight into pathogenesis and animal models. However, in spite of this effort, global treatments have not been possible for ALS because a common molecular mechanism downstream from the distinct etiologies has not become apparent. It is clear that discovery of additional etiologies and common downstream mechanisms will greatly advance the goals for a molecular understanding of ALS and in finding appropriate treatments. In this application we propose an advance in direction of those goals. We have discovered mutations in UBQLN2 as cause of X-linked dominant ALS and ALS/dementia of the FTD type. The pathology is novel, distinct and unique with ubiquilin2 positive inclusions in the spinal cord and cortex in ALS patients and additional inclusions in the hippocampus of patients with ALS/FTD. To our surprise we found similar ubiquilin2 positive inclusions in all cases of SALS and ALS/FTD and FALS cases studied thus far. None of those cases had mutations in UBQLN2, arguing for a posttranslational role for ubiquilin2 in ALS as a whole. Our initial in vitro and in vivo data suggest a dysfunction of the ubiquilin proteasome syndrome (UPS) and autophagy in the presence of mutant ubiquilin2. In addition, cognitive and behavioral deficits and decreased long term potentiation in far field recordings were observed in transgenic mice expressing a pathogenic UBQLN2 mutation (P497H). In this project, we propose four specific aims to understand the disease causing mechanism(s) of mutant ubiquilin2. We will establish pathological, cognitive and other behavioral phenotypes and their electrophysiological correlates in our current and to be developed mouse models of mutant ubiquiln2. We will also use our already established ubqln2 knockout mouse model and develop ubqln1 knockout and double knockouts of ubqln1/ubqln2 to test if mutant ubiquilin2 defects are manifestation of a loss of function of ubiquilin2 in relationto the UPS and autophagy systems or whether mutant ubiquilin2 exhibits a novel toxic property. Successful completion of this project will not only provide insight into understanding the pathogenic mechanism of X-linked ALS, but also rapidly provide the wider research community with useful reagents for future studies and for screening and testing potential therapies. The outcome of this project will also have important implications in the understanding of the pathogenesis and treatment of other neurodegenerative diseases.
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1 |
2013 — 2017 |
Siddique, Teepu |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Association of Als to Gene-Environment Mediated Changes in Hdl Proteins @ Northwestern University At Chicago
DESCRIPTION (provided by applicant): Amyotrophic lateral sclerosis (ALS) or Lou Gehrig disease is a fatal neurodegenerative disease that primarily affects mid-life and older adults. There are two forms of disease, familial (FALS) and sporadic (SALS), respectively comprising 10% and 90% of cases, respectively. The genetic causes of FALS have been linked to mutations in several genes such as superoxide dismutase, TDP-43, FUS, optineurin, ubiquilin 2 and C9ORF72. The etiology of SALS, however, remains elusive. A few years ago our laboratory found that there were polymorphisms in genes for enzymes called paraoxonases that were associated with SALS. These enzymes detoxify certain pesticides and toxic agents, and thus became the first environmentally related genes linked to ALS. Further studies of the paraoxonases and apolipoprotein L1 in the plasma indicate that their levels are significantly elevated in SALS patients. These proteins are found on specific high density lipoprotein (HDL) particles that have several functions including lipid and cholesterol transport and protecting lipoproteins from deleterious oxidation. Similar particles are also found in the cerebrospinal flui (CSF). Thus, in this proposal we further characterize the protein composition of selected HDL species in the plasma and CSF of SALS patients using high throughput technologies. We can now, for the first, time determine how certain HDL proteins change in a neurodegenerative disease and if they are linked to the disease process. We will also determine whether the genes for HDL-associated proteins contain variants that are associated with risk of SALS and whether these changes are related to alterations in HDL protein levels. Mechanisms of the deleterious effects of these changes will be studied in cell culture and genetically engineered mice. Results from this work will open paths to therapies to rescue potential dysfunctional HDL found not only in neurodegenerative disease such as ALS but also in more common cardiovascular and metabolic diseases.
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2016 — 2020 |
Siddique, Teepu |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Investigation of Als Caused by Mutant Chchd10 @ Northwestern University At Chicago
Some of the most robust advances in the understanding of amyotrophic lateral sclerosis (ALS), a multi- etiologic and fatal disorder of the nervous system, have come from the identification of the genetic basis of a subgroup of ALS cases. In this subgroup of ALS (familial ALS or FALS), the disease is inherited as a familial trait. Identification of a single mechanism of disease across all of ALS has been elusive, but notable amongst proposed common mechanisms is mitochondrial dysfunction. However, till recently, no direct robust etiological link between mitochondrial genes, nuclear or mitochondrial, had been found. Recently, others and we found that mutations in a gene called CHCHD10 causes ALS and additional phenotypes. Some other ALS genes, including VCP, HNRNPA1, SQSTM1 and OPTN are also associated with other phenotypes besides ALS. CHCHD10 and its protein product are not well studied. To investigate how a mutation in CHCHD10 may cause ALS, we have collected or developed promising reagents and assembled collaborators and laboratory personnel with specialized skills in genetic engineering, rodent models, cell biology, CRISPR/Cas9 gene editing, mitochondrial function, redox response, induced pluripotent stem cell (iPSC) derived motor neurons and primary motor neurons to carry out the proposed experiments in my laboratory and the laboratories of collaborators. We will investigate the pathology and behavior of a new transgenic mouse model we have engineered to overexpress the mutant (R15L) human CHCHD10 gene. Preliminary study shows axonal pathology in the spinal cord and brain of this mouse with periodic beaded axonal swellings harboring mitochondria. Additional more precise models will also be developed. To determine what aspect of mitochondrial function or morphology is affected by mutant CHCHD10 we will screen mitochondrial function of energetics, redox studies, calcium buffering and electron microscopy analysis. We will identify the binding protein partner(s) of CHCHD10 protein to understand the role of CHCHD10 in the mitochondrial biology. On completion, our study would provide a clearer understanding of the central defect(s) in this form of ALS and potentially a more granular understanding of mitochondrial defect generalizable across ALS. The study will also allow the identification of potential molecular targets for rational therapy and/or prevention of ALS.
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2018 — 2019 |
Siddique, Teepu |
R21Activity Code Description: To encourage the development of new research activities in categorical program areas. (Support generally is restricted in level of support and in time.) |
A Novel Approach to Therapy in Als and Als/Dementia Using New Insights From Als5/Spg11 @ Northwestern University At Chicago
Truncation mutations in SPG11 are associated with juvenile onset amyotrophic lateral sclerosis (ALS5), spastic paraplegia and other neurodegenerative conditions. We recently identified a novel pathology of aggregation of large dense core vesicles (LDCVs) in the motor neurons of a patient with a juvenile form of ALS and dementia called ALS5/SPG11. We have now verified such pathological aggregation of LDCVs also in our knock out (KO) mouse model of Spg11. We also have evidence that LDCV trafficking is impaired by mutations in the spastic paraplegia gene SPG11 which encodes a large protein called spatacsin that is a component of LDCVs. LDCVs pathology in motor neuron degeneration provides a new venue to develop novel therapies. We hypothesize that the LDCVs sequester in the soma and the cargo are constrained to perform their presumed neurotrophic and/or functions or communication functions. LDCVs are known to carry neuropeptides such as substance P, neuropeptide Y, brain-derived neurotrophins (e.g. BDNF), calcitonin gene related peptide (CGRP) and chromogranins. Release of LDCV cargo is regulated; they must be regenerated in the neuronal soma and refilled. The complete cargo of the motor neuron LDCVs has not been established. Studies of motor neuron LDCV cargo may reveal a novel peptide or neurotransmitter/neuromodulator whose significance may have been overlooked in motor neuron biology and which may broadly prevent or treat motor neuron degeneration. The pathological sequestration of LDCVs in aggregations in the soma of motor neurons suggests such a mechanism. A broader application of our research may be relevant because Chromogranin A, one of the markers by which LDCVs are recognized, is present in the pathological inclusions of spinal motor neurons in the sporadic form of human ALS. The abundant material available from our Spg11 KO mouse model and iPSCs derived motor neurons from eleven ALS5/SPG11 patients provides us a rare opportunity to satisfactorily carry out the proposed experiments in a robust and reproducible manner with techniques currently in use in our laboratory.This project is directed towards the neuron specific identification of the protein components and peptide cargo within large dense core vesicles (LDCVs) obtained from the Spg11 knock-out mouse and human (ALS5/SPG11 patients) induced pluripotent stem cell-derived neurons. In addition, we will identify molecularly proximate binding partners with the use of an engineered hybrid protein consisting of a modified biotin ligase bound to spatacsin and compare with the LDCVs cargo inventory from motor neuron or those derived from other neuronal systems of the Spg11 KO mouse. Motor neurons derived from human SPG11/ALS iPSCs will be evaluated for LDCVs aggregation, soma morphology, neurite extension and branching and cell viability. Suitable candidate cargo molecules will be tested for positive effect on these parameters of iPSC derived motor neurons. Knowledge of the cargo from neurons in specific regions of the brain may enable replacement therapies that deliver specific cargo (e.g. neuropeptides) to affected areas of the CNS and thus restore neuronal homeostasis.This exercise should provide a new inventory of therapeutics for ALS, ALS/dementia and possibly other forms of neurodegeneration.
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2019 — 2021 |
Siddique, Teepu |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
A Novel Innate Immunity Risk Factor For Amyotrophic Lateral Sclerosis @ Northwestern University At Chicago
Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disorder. 20% of ALS cases are genetic, with monogenic causes identified in about 17%. The remaining 80% of cases appear to be multifactorial, and no reproducible and/or robust genetic association or environmental factor(s), internal or external, have been identified. Although the role of innate immunity has been explored and some innate immunity factors highlighted, their role as initiators, amplifiers or secondary markers of an ongoing inflammatory response is not established in ALS. As the result of our study of gene variants of cargo proteins of the HDL particle, we identified the trypanocidal blood and CSF protein, APOL1, as a promising agent affecting ALS that is also sensitive to environmental factors.APOL1 is a secreted innate immunity factor carried by the high-density lipoprotein particle HDL3and CSF. APOL1 forms a channel in the lysosomal membrane and disrupts the blood borne parasite's lysosomal function, killing it. Most trypanasome exposures are clinically inapparent because only two of the twenty known trypanosome species, varieties of T.brucei and T.cruzei, are pathogenic to humans. Variant alleles of looAPOL1 emerged in Africa with the property of antagonistic pleiotropy, effective against resistant strains of T.brucei, but they can cause progressive kidney disease especially with SLE and AIDS as co-morbities. Our hypothesis is that some APOL1 variants are associated with ALS because elevated APOL1 levels in plasma and or CSF can be toxic to susceptible neurons. APOL1 levels are influenced by in cis or in trans factors. APOL1 gene expression is likely regulated in cis by specific DNA variants and/or by in trans due to exposure to trypanosomes or to cytokines such as the interferons or other, yet to be identified, environmental factors. We have patient samples with associated epidemiological data that we will use to determine which APOL1 variants are correlated with plasma concentrations of ApoL1 and/or mRNA expression in patient-derived lymphoblasts. We also will additional patient and control samples from our own collection and those collected prospectively from new patients. In selected samples we will run assays to determine if blood can be used to determine whether a prior trypansomal infection has occurred as well as levels of cytokines that may be elevated due to inflammatory responses related to infection. This is a novel pioneering effort to ascertain 1. Whether APOL1 genetic variants and APOL1 levels predispose to SALS. 2. The extent to which Apol1 levels are affected by in cis and possibly in trans genetic variantsacting as eQTLs. 3. The effect on Apol1 levels by in trans environmental factors of exposure to trypanosomes, or secondarily by cytokines such as TNF-?, ?-interferon and type 1 interferons, or a combination thereof. 4. To establish Apol1 in the pathology of vulnerable neurons in human ALS. Our promising preliminary data, experienced team, and rigorously defined cohort of age-matched ALS cases and controls and extensive collection of ALS tissue and ongoing environment exposure data puts us in a unique position to carry out this novel proposal to fundamentally effect the understanding of SALS and its treatment.
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