Neurological Disorders

Coding Dimension ID: 
303
Coding Dimension path name: 
Neurological Disorders

Human ES cell-derived MGE inhibitory interneuron transplantation for spinal cord injury

Funding Type: 
Early Translational III
Grant Number: 
TR3-05606
ICOC Funds Committed: 
$1 623 251
Disease Focus: 
Spinal Cord Injury
Neurological Disorders
Stem Cell Use: 
Embryonic Stem Cell
oldStatus: 
Active
Public Abstract: 
Transplantation of neuronal precursors into the central nervous system offers great promise for the treatment of neurological disorders including spinal cord injury (SCI). Among the most significant consequences of SCI are bladder spasticity and neuropathic pain, both of which likely result from a reduction in those spinal inhibitory mechanisms that are essential for normal bladder and sensory functions. Our preliminary data show that embryonic inhibitory neuron precursor cells integrate in the adult nervous system and increase inhibitory network activity. Therefore inhibitory nerve cell transplants could be a powerful way to establish new inhibitory circuits in the injured spinal cord that will reduce bladder spasticity and attenuate central neuropathic pain. We already have proof-of-principle data that murine inhibitory nerve cells integrate in the adult spinal cord and improve symptoms in an animal model of chronic spinal cord injury. We have also recently developed methods to create human inhibitory interneurons from embryonic stem cells. This proposal will capitalize on these recent developments and determine whether our human embryonic stem cell-derived inhibitory cells can be successfully transplanted into the grey matter of the injured spinal cord and reduce neurogenic bladder dysfunction and neuropathic pain, two major causes of suffering in chronic SCI patients. If successful, our studies will lay the groundwork for a potential novel therapy for chronic SCI.
Statement of Benefit to California: 
There are an estimated 260,000 individuals in the United States who currently live with disability associated with chronic spinal cord injury (SCI). Symptoms of chronic SCI include bladder dyssynergia reflected by incontinence coincident with asynchronous contraction of internal and external sphincters, and central neuropathic pain, both of which severely impede activities of daily living, reduce quality of life, and contribute to the very high medical costs of caring for the Californians who suffer from chronic spinal cord injury. The Geron trial for SCI, as well as other cell-based approaches, aim to treat acute SCI. This proposal considers a different potentially complementary cell-transplantation strategy that is directed to more chronic SCI with the goal of improving bladder function and reducing pain. We propose to use cell grafts of inhibitory interneurons that we have derived from human stem cells in order to provide a novel treatment. If successful, we will have defined a therapeutic option that targets the most prevalent population of spinal cord injured patients. As the country's most populous state, California has the largest number of patients with chronic SCI, approximately 12,000. The estimated economic cost to California in lost productivity and medical expenses amounts to $400,000,000 annually. The potential savings in medical care costs, and improvement in quality of life will therfore have a disproportional benefit to the state of California.
Progress Report: 
  • From the past six months of work, we report considerable progress toward our aims of investigating the safety and efficacy of human inhibitory nerve precursor (MGE) cell transplantation for the treatment of spinal cord injury-induced bladder spasticity and neuropathic pain. Our first aim details the injection of human MGE cells into the uninjured rodent spinal cord and investigation of cell fate and potential adverse side effects from their transplantation. During the reporting period, we completed histological analyses for the two-month time point post-injection, and we found that the human MGE cells, derived from human embryonic stem cells (hESCs), appropriately matured into forebrain-type inhibitory interneurons in the rodent spinal cord. Also, we initiated histological examination of animals six months post-injection and detected robust human cell survival, dispersal into the spinal cord grey matter, and neuronal maturation, but no evidence of tumor formation. In addition, we completed behavioral analyses of animals injected with hESC-derived MGE cells at two and six months post-injection. Thus far, we have not observed any adverse side effects when human MGE cells are transplanted into the uninjured animal as determined by measures of body weight, locomotion, bladder function, and pain sensitivity.
  • Since the beginning of this project, we report considerable progress toward our aims of investigating the safety and efficacy of human inhibitory nerve precursor (MGE) cell transplantation for the treatment of spinal cord injury-induced bladder spasticity and neuropathic pain. In year one of this award we completed the major objectives of Aim1, namely to explore the survival, integration, and cell fate of stem cell-derived MGE cell transplants in the uninjured rodent spinal cord. We have now obtained preliminary efficacy results from Aim 2, namely the effects of hESC-MGE cells injected in spinal cord injured animals. Behavioral testing has been obtained to assess pain thresholds for all injected animals up to the six month endpoint, and measures of bladder spasticity have been obtained at six months post cell injection. We are evaluating whether the unblinded data demonstrates amelioration of neuropathic pain and bladder spasticity. Our preliminary histological analysis shows robust human cell survival, distribution, and neuronal differentiation, and we have electrophysiological data indicating functional integration of the transplanted cells. We are on track to complete all aims by the end of the award period.

Multiple Sclerosis therapy: Human Pluripotent Stem Cell-Derived Neural Progenitor Cells

Funding Type: 
Early Translational III
Grant Number: 
TR3-05603
ICOC Funds Committed: 
$4 799 814
Disease Focus: 
Multiple Sclerosis
Neurological Disorders
Collaborative Funder: 
Australia
Stem Cell Use: 
Embryonic Stem Cell
iPS Cell
oldStatus: 
Active
Public Abstract: 
Multiple Sclerosis (MS) is a disease of the central nervous system (CNS) caused by inflammation and loss of cells that produce myelin, which normally insulates and protects nerve cells. MS is a leading cause of neurological disability among young adults in North America. Current treatments for MS include drugs such as interferons and corticosteroids that modulate the ability of immune system cells to invade the CNS. These therapies often have unsatisfactory outcomes, with continued progression of neurologic disability over time. This is most likely due to irreversible tissue injury resulting from permanent loss of myelin and nerve destruction. The limited ability of the body to repair damaged nerve tissue highlights a critically important and unmet need for MS patients. The long-term goal of our research is to develop a stem cell-based therapy that will not only halt ongoing loss of myelin but also lead to remyelination and repair of damaged nerve tissue. Our preliminary data in animal models of human MS are very promising and suggest that this goal is possible. Research efforts will concentrate on refining techniques for production and rigorous quality control of clinically-compatible transplantable cells generated from high-quality human pluripotent stem cell lines, and to verify the therapeutic activity of these cells. We will emphasize safety and development of the most therapeutically beneficial cell type for eventual use in patients with MS.
Statement of Benefit to California: 
One in seven Americans lives in California, and these people make up the single largest health care market in the United States. The diseases and injuries that affect Californians affect the rest of the US and the world. Many of these diseases involve degeneration of healthy cells and tissues, including neuronal tissue in diseases such as Multiple Sclerosis (MS). The best estimates indicate that there are 400,000 people diagnosed with MS in the USA and 2.2 million worldwide. In California, there are approximately 160,000 people with MS – roughly half of MS patients in the US live in California. MS is a life-long, chronic disease diagnosed primarily in young adults who have a virtually normal life expectancy but suffer from progressive loss of motor and cognitive function. Consequently, the economic, social and medical costs associated with the disease are significant. Estimates place the annual cost of MS in the United States in the billions of dollars. The development of a stem cell therapy for treatment of MS patients will not only alleviate ongoing suffering but also allow people afflicted with this disease to return to work and contribute to the economic stabilization of California. Moreover, a stem cell-based therapy that will provide sustained recovery will reduce recurrence and the ever-growing cost burden to the California medical community.
Progress Report: 
  • The team has been highly productive during the first year of work on this award. A major goal of the project is to evaluate the efficacy of neural progenitor cell transplantation to promote remyelination following virus induced central nervous system damage. With intracranial infection by the virus mouse hepatitis virus (MHV), mice develop paralysis due to immune mediated destruction of cells that generate myelin. Using protocols developed in the Loring laboratory, neural precursor cells (NPC) were derived from the human embryonic stem cell line H9. Mice developing paralysis due to intracranial infection with MHV were subject to intraspinal transplantation of these NPC, resulting in significant clinical recovery beginning at 2-3 weeks following transplant. This clinical effect of NPC transplantation remained out to six months, suggesting that these NPC are effective for long-term repair following demyelination. Despite this striking recovery, these human ES cell derived NPC were rapidly rejected. Several protocols for the generation of NPC for transplantation have been characterized, with the greatest clinical impact observed for NPC cultures bearing a high level of expression of TGF beta I and TGF beta II. These findings support the hypothesis that transplanted NPC reprogram the immune system within the central nervous system (CNS), leading to the activation of endogenous NPC and other repair mechanisms. Thus, it may not be necessary to induce complete immune suppression in order to promote remyelination and CNS repair following NPC transplantation for demyelinating diseases such as multiple sclerosis.

Cellular tools to study brain diseases affecting synaptic transmission

Funding Type: 
Tools and Technologies II
Grant Number: 
RT2-02061
ICOC Funds Committed: 
$1 906 494
Disease Focus: 
Autism
Neurological Disorders
Rett's Syndrome
Pediatrics
Stem Cell Use: 
Embryonic Stem Cell
iPS Cell
Cell Line Generation: 
Embryonic Stem Cell
iPS Cell
oldStatus: 
Active
Public Abstract: 
There is a group of brain diseases that are caused by functional abnormalities. The brains of patients afflicted with these diseases which include autism spectrum disorders, schizophrenia, depression, and mania and other psychiatric diseases have a normal appearance and show no structural changes. Neurons, the cellular units of the brain, function by making connections (or synapses) with each other and exchanging information in form of electric activity. Thus, it is believed that in those diseases many of these connections are not working properly. However, using current technology, there is no way to investigate individual neuronal synapses in the human brain. This is because it is not ethical to biopsy the brain of a living person if it is not for the direct benefit to the patient. Therefore, scientists cannot study synaptic function in psychiatric diseases. Because of the limited knowledge about the functional consequences in the affected brains, there is no cure for these diseases and the few existing therapies are often associated with severe side effects and cannot restore the normal function of the brain. Therefore, it is of great importance to better study the disease processes. A better knowledge on what the defects are on the cellular level will enable us then in a second step to test existing drugs and measure its effect or screen for new therapeutic drugs that can improve the process and hopefully also the disease symptoms. This proposal aims to develop a technology to overcome this limitation and ultimately provide neurons directly derived from affected patients. This will uniquely allow the study functional neuronal aspects in the patients' own neurons without the need to extract neurons from the brain. Our proposal has two steps, that we want to undertake in parallel with mouse and human cells. First, we want to find ways to optimally generate neurons from skin fibroblasts. Naturally, these artificial neurons will have to exhibit all functional properties that the neurons from the brain have. This includes their ability to form functional connections with each other that serve to exchange information between two cells. In the second step, we will generate such neuronal cells from a genetic form of a psychiatric disease and evaluate whether these cultured neuronal cells indeed exhibit changes in their functional behavior such as the formation of fewer connections or a decreased probability to activate a connection and thus limit the disease cells to communicate with other cells.
Statement of Benefit to California: 
Our proposed research is to develop a cellular tool which will enable the research community to study human brain diseases that are caused by improperly functioning connections between brain cells rather than structural abnormalities of the brain such as degeneration of neurons or developmental abnormalities. These diseases, which are typically classified as psychiatric diseases, include schizophrenia, bipolar diseases (depression, mania) autism spectrum disorders, and others. There are many people in California and world-wide that suffer from these mentally debilitating diseases. Therefore, there is a great need to develop therapies for these diseases. However, currently drug development is largely restricted to animal models and very often drug candidates that are successful in e.g. rodent animals can not be applied to human. It would thus be much better to possess a model that reflects the human disease much closer, ideally using human cells. We have experimental evidence that we can develop such a model. In particular, we will convert skin cells from patients suffering from psychiatric diseases into stem cells that are "pluripotent", which means they can differentiate into all cell types of the body including neurons. We want to explore whether these patient-derived neurons still contain the disease features that the neurons have in the brain. If we could indeed capture the disease in these cells, our technology would have a major impact on future work in this area. We believe that this approach could be applied to many neurological diseases including neurodegenerative diseases. Our technology would not only provide a unique experimental basis to begin to understand how these diseases work, but it would allow to then interfere with the identified cellular abnormalities which would secondarily result in the development of new drugs that can counteract the diseases and would hopefully also work for the patients themselves. Therefore, all those Californians that suffer from one of the above mentioned diseases will benefit from our research project, if it is successful.
Progress Report: 
  • During this first year of our project we have largely focused on testing various methods to directly differentiate human ES cells into neurons. As described in more detail below we were very successful and developed ways to differentiate human stem cell lines into neuronal cells with high purity and good maturation characteristics. For example, we can analyze the electrical currents in these cells which are important functional properties of neurons and we observed that these cells indeed behave just like neurons in the brain. More specifically, the cells were able to generate action potentials which are necessary in the brain to transmit information from one neuron to the other as well as form synapses, which are the structures that connect the different neurons with each other.Because the differentiation of different stem cell lines needs to be robust and reproducible we spent a lot of time optimizing the protocol and tested many different stem cell lines. This revealed a high degree of reproducibility and purity of the stem cell-derived nerve cells and we have tested human embryonic stem cells (i.e. stem cells derived from the embryo) as well as induced pluripotent (iPS) cells (i.e. stem cells reprogrammed from human skin cells). Reassuringly, the same method works in all these cell lines with very similar dynamics and functional properties of the nerve cells.
  • We also have made significant advances to convert human fibroblasts into nerve cells directly and without going through an intermediate iPS cell state. We have identified a neuronal factor called NeuroD1 as critical co-factor that in addition to the three factors that we had identified earlier to work in mouse. Those 4 factors together now allowed the generation of fully functional so called "induced neuronal" (iN) cells from both fetal and early postnatal human foreskin fibroblasts. We have also tested a number of small molecules to attempt to increase the reprogramming efficiency.
  • Finally, we have generated some essential components that will allow us to study Rett Syndrome using these technologies that are being developed at the same time (described above). In the last year we have generated several lines of iPS cells from Rett Syndrome patients and are in the process of fully characterizing them. We plan to soon apply our optimized differentiation protocol to these cells as well as control cells to look for any possible disease trait that distinguishes cells from patients and controls.
  • The generation of human pluripotent stem cells from discarded embryos (embryonic stem cells or ES cells) and directly from skin cells through reprogramming (induced pluripotent stem cells or iPS cells) holds great promise, and may revolutionize the study of human diseases. In particular, the principle possibility to turn these stem cells into fully functional neurons would provide a novel cell platform that provides excellent experimental access to study human neurons that are derived from healthy controls or diseased individuals. However, the goal to actually derive mature neurons from these stem cell populations has not been accomplished yet. While there have been many ways developed how to instruct these stem cells into specific neurons and even neuronal subtypes, these differentiation protocols take many months to complete and are laborious and most importantly, do not yield fully mature neurons. We have recently discovered a way to convert human newborn skin cells directly into functional neurons but the efficiencies were low and also most of these induced neuronal cells were still immature.
  • The goal of this project is to improve these methods and develop tools that actually allow the generation of mature human neurons. We proposed to approach this problem in two different and complementary ways: (1) We proposed to apply the methods that we used to convert human skin cells into neurons to both stem cell populations (ES and iPS cells). (2) We proposed to further improve the direct conversion of skin cells into induced neuronal cells by systematic evaluation of culture conditions and small molecule modulators alone and in combination. Finally, we then proposed to apply our newly derived tools to study one common autism-related childhood disease, called Rett Syndrome, which affects exclusively girls, which undergo normal development and brain maturation but after a period of months to years present with developmental retardation and in some cases severe behavioral and social deficiencies.
  • We are very happy to be able to report that we have made great progress towards the development of our proposed tools and are now beginning already to apply them to the study of Rett Syndrome as proposed. In particular, we have perfected the application of the technique to convert human stem cells into fully functional induced neuronal cells. With this approach we are ready, to investigate the detailed electric connectivity of neural circuits in induced neuronal cells in disease and non-disease condition.
  • We have also made good progress with the second approach and showed that it is possible to improve the conversion efficiencies significantly by using small molecule inhibitors and changing the environmental oxygen concentration. We are currently exploring whether these efficiencies are high enough to enable disease modeling while we continue to optimize the culture methods.
  • We have generated a new tool to study brain function on the cellular level. The differentiation of pluripotent stem cells like embryonic or induced pluripotent stem cells into functional nerve cells (neurons) remains a challenge. We here demonstrated that specific factors that normally regulate brain development can be exploited to "fast forward" the differentiation of human stem cells into neurons. Since these neurons are induced using exogenous factors we call these cells "induced neuronal cells" or in brief "iN" cells. Stem cell-derived iN cells show all principal functional properties of neurons, ie, they can communicate with each other (form synapses) and use electrical signals to convey information (ability to generate action potentials). Within just 2-3 weeks fully functional neuronal networks can be established using these human neurons.
  • We next demonstrated that different factor combinations yields different kind of neurons allowing us to reconstruct complex cell mixtures resembling those of normal neuronal cultures.
  • We also show that iN cells are useful proxies that report disease traits on the cellular level. In particular we demonstrated that a gene mutation that is associated with Schizophrenia leads to a functional defect measurable in human iN cells. This might lead to important new methods to find treatments for these devastating diseases.

Use of human iPS cells to study spinal muscular atrophy

Funding Type: 
Basic Biology III
Grant Number: 
RB3-02161
ICOC Funds Committed: 
$1 268 868
Disease Focus: 
Spinal Muscular Atrophy
Neurological Disorders
Pediatrics
Stem Cell Use: 
iPS Cell
Cell Line Generation: 
iPS Cell
oldStatus: 
Active
Public Abstract: 
Spinal muscular atrophy (SMA) is one of the most common autosomal recessive disorders that cause infant mortality. SMA is caused by loss of the Survival of Motor Neuron (SMN) protein, resulting in motor neuron (MN) degeneration in the spinal cord. Although SMN protein plays diverse roles in RNA metabolism and is expressed in all cells, it is unclear why a deficiency in SMN only causes MN degeneration. Since patient samples are rarely available, most knowledge in SMA is gained from animal model studies. While these studies have provided important information concerning the cause and mechanism of SMA, they are limited by complicated genetic manipulation. Results from different models are also not always consistent. These problems can be resolved if SMA patient’s MNs become readily available. Recent progress in the generation of induced pluripotent stem (iPS) cells from differentiated adult cells provides an opportunity to establish human cell-based models for neurodegenerative diseases. These cells, due to their self-renewal property, can provide an unlimited supply of the affected cell type for disease study in vitro. In this regard, SMA iPS cells may represent an ideal candidate for disease modeling as SMA is an early onset monogenic disease: the likelihood to generate disease-specific phenotypes is therefore higher than iPS cells derived from a late onset disease. In addition, the affected cell type, namely MNs, can readily be generated from iPS cells for the study. For these reasons, we established several SMA iPS cell lines from a type 1 patient and showed specific deficits in MNs derived from these iPS cells. Whether MNs derived from these iPS cell lines can recapitulate a whole spectrum of SMA pathology in animals and patients remains unclear. An answer to this question can ensure the suitability of using the iPS cell approach to study SMA pathogenesis in cell culture. We propose to examine cellular and functional deficits in MNs derived from these SMA iPS cells in Aim 1. The availability of these iPS cells also provides an opportunity to explore the mechanisms of selective MN degeneration in SMA. Dysregulation of some cellular genes has been implicated in SMA pathogenesis. We propose to use these iPS cell lines to address how one such gene is affected by SMN deficiency (Aim 2) and how a deficit in these genes leads to selective MN degeneration (Aim 3). Our study should provide valuable insights in the understanding of SMA pathogenesis and aid in exploring new molecular targets for drug intervention.
Statement of Benefit to California: 
Spinal muscular atrophy (SMA) is one of the most common autosomal recessive disorders in humans and the most common genetic cause of infant mortality. SMA is caused by loss of the Survival of Motor Neuron (SMN) protein, resulting in motor neuron (MN) degeneration in the spinal cord. SMA has a carrier frequency of approximately 1 in 35 and an incidence of 1 in 6000 in human population. In severe SMA cases, the disease onset initiates before 6 months of age and death within the first 2 years of life. Currently, there is no cure for SMA. Since MN samples from patients are rarely available, most knowledge in SMA is gained from animal model studies. While these studies have provided important information concerning the cause and mechanism of SMA, they are limited by complicated genetic manipulation. Results from different models are not always consistent either. Large-scale drug screening to treat SMA is also hampered by the lack of suitable cell lines for the study. These problems can potentially be resolved if SMA patient’s MNs become readily available. Our effort to derive induced pluripotent stem (iPS) cells from a SMA patient provides an unlimited supply of SMA cells to carry out studies to explore the disease mechanism in vitro. A better understanding in the disease mechanisms would benefit California by the identification of potential cellular targets for drug treatment. The knowledge gained from our study can also facilitate the use of these iPS cells as a platform for large-scale drug screening and validation. Our study should provide valuable insights in the understanding of SMA pathogenesis and aid in exploring new molecular targets for drug intervention.
Progress Report: 
  • During the past fiscal year, we have established in vitro coculture between motoneurons and myocytes. This coculture system will form the basis for the analysis of potential SMA pathogenesis induced by the motoneurons derived from SMA iPS cells. We have also started the analysis of potential cellular targets whose activity is affected by SMN deficiency.
  • Spinal muscular atrophy (SMA) is one of the most common autosomal recessive disorders that cause infant mortality. SMA is caused by loss of the Survival of Motor Neuron (SMN) protein, resulting in motor neuron (MN) degeneration in the spinal cord. Although SMN protein plays diverse roles in RNA metabolism and is expressed in all cells, it is unclear why a deficiency in SMN only causes MN degeneration. Since patient samples are rarely available, most knowledge in SMA is gained from animal model studies. While these studies have provided important insights of the cause and mechanism of SMA, they are limited by complicated genetic manipulation. Results from different models are also not always consistent. These problems can be addressed using induced pluripotent stem cells (iPSCs) derived from patient’s fibroblasts. These cells, due to their self-renewal capacity and their ability to differentiate into neuronal cells, can in theory provide an unlimited supply of the affected MNs for SMA study. We propose to examine cellular and functional deficits in MNs derived from these SMA iPS cells in Aim 1. To increase the yield of MN production, we have tested new strategies to differentiate SMA iPSCs into MNs. The improvement makes it feasible to isolate more pure populations of MNs for the study of SMA pathogenesis in vitro. The availability of these iPSC lines also provides an opportunity to explore the mechanisms of selective MN degeneration in SMA. Dysregulation of some cellular genes has been implicated in SMA pathogenesis. We continue to study the role of one particular cellular gene whose expression is reduced in SMA (Aim 2). We are taking approaches to reveal how SMN deficiency causes this change in gene expression. We are also taking a genomic approach to reveal all the affected genes and the signaling pathways in SMA MNs and understand how a deficit in these genes leads to selective MN degeneration (Aim 3). Our study should provide valuable insights in the understanding of SMA pathogenesis and aid in exploring new molecular targets for drug intervention.
  • Spinal muscular atrophy (SMA) is one of the most common genetic disorders that cause infant mortality. SMA is caused by loss of the Survival of Motor Neuron (SMN) protein, resulting in motor neuron degeneration in the spinal cord. Although SMN protein plays diverse roles in cells and is expressed in all cells, it is unclear why a deficiency in SMN only causes motoneuron degeneration. Since patient samples are rarely available, most knowledge in SMA is gained from animal model studies. While these studies have provided important information concerning the cause and mechanism of SMA, they are limited by complicated genetic manipulation. Results from different models are also not always consistent. These problems can be resolved if SMA patient’s motoneurons become readily available. The progress in the generation of stem cell lines from differentiated adult cells, termed induced pluripotent stem cells (iPSCs), provides an opportunity to establish human cell-based models for neurodegenerative diseases like SMA. We have previously established several SMA iPSC lines from a type 1 patient and showed specific deficits in motoneurons derived from these iPSCs. The availability of these iPSCs provides an opportunity to explore the mechanisms of selective motoneuron degeneration in SMA. We used motoneurons derived from SMA iPSCs to study potential defects in the formation of neuromuscular junctions. We also demonstrated a regulatory gene product affected by SMN deficiency. Several potential downstream targets of the regulatory gene product involved in neuron migration and synaptic transmission were identified. The roles of these genes in selective motoneuron degeneration observed in SMA are currently under study. One technical obstacle of using iPSC-derived motoneurons to study SMA in a dish is that motoneurons generally constitute only a fraction of the resulting cell population. The lack of capacity to isolate motoneurons hampers our study of SMA pathogenesis and the identification of potential downstream targets of SMN. We have employed a new approach, termed gene editing, to mark differentiated motoneurons with a fluorescence protein to facilitate their isolation by cell sorting. A proof-of-principle experiment was carried out and demonstrated the feasibility of this strategy. We are currently applying this strategy to mark motoneurons derived from SMA iPSCs.

The HD iPSC Consortium: Repeat Length Dependent Phenotypes for Assay Development

Funding Type: 
iPSC Consortia Award
Grant Number: 
RP1-05741
ICOC Funds Committed: 
$300 000
Disease Focus: 
Huntington's Disease
Neurological Disorders
Collaborative Funder: 
NIH
Stem Cell Use: 
iPS Cell
Cell Line Generation: 
iPS Cell
oldStatus: 
Active
Public Abstract: 
Statement of Benefit to California: 
Progress Report: 
  • Huntington’s disease (HD) is a significant neurodegenerative disease with unique genetic features. A CAG expansion in Huntington gene is correlated with severity and onset of sub-clinical and overt clinical symptoms, make it particularly suited to therapeutic development . The single genetic cause offers the opportunity to understand the pathological process triggered in all individuals with a CAG expansion, as emerging evidence suggests effects of the mutation in all cell types, though striatal neurons are most vulnerable to degeneration. Moreover, by virtue of a molecular test for the mutation, a unique opportunity exists to intervene/treat before the onset of overt clinical symptoms utilizing sub-clinical phenotypes emerging in pre-manifest individuals. Since human induced pluripotent stem cells (iPSCs) have the power to make any cell type in the human body, we are utilizing the technology to make patients iPSCs and study the effects of different number of CAG repeats on the neurons we generate from the patient iPS cells. Preliminary studies indicate that CAG length–dependent phenotypes occur at all stages of differentiation, from iPSC through to mature neurons and are likely to occur in non-neuronal cells as well, which can also be investigated using the iPSC that we are creating. The non-integrating technology (avoids integration of potentially deleterious reprogramming factors in the cell DNA) for producing iPSC lines is crucial to obtaining reproducible disease traits from patient cells.
  • The Cedars-Sinai RMI iPSC Core is part of the Huntington’s Disease (HD) consortium. In the past year the iPSC Core has made many new non-integrating induced pluripotent stem (iPSC) cell lines from HD patients with different numbers of CAG repeat expansions. The grant application proposed generation of 18 HD and Control iPSC lines. Instead we are generating 20 iPSC lines. So far we have already generated 17 iPSC lines from individuals with Huntington’s disease and controls (10 HD patients and 7 controls). In order to have the disease trait reproducible across multiple groups, three clonal iPSC lines were generated from each subject. Some of these lines have (or are in process) of expansion for distribution to consortium members. We are now in the process of making the last 3 lines as part of this grant application to generate a HD iPSC repository with total of 20 patient/control lines from subjects with multitude of CAG repeat numbers. Most of these lines have undergone rigorous battery of characterization for pluripotency determination, while some other lines are currently being validated through more characterization tests. Neural stem cell aggregates (EZ spheres) have been generated from few of the patient lines in the Svendsen lab (not supported by this grant). We have also submitted 6 patient iPSC lines to Coriell Cell Repository for larger banking and distribution of these important and resourceful lines to other academic investigators and industry. We strongly believe that this iPSC repository will enormously speed up the process of understanding the disease causing mechanisms in HD patient brain cells as well as discovering novel therapeutics or drugs that may one day be able to treat HD patients.
  • Huntington’s disease (HD) is a fatal neurodegenerative condition with no current treatment. This significant neurodegenerative disease, whose relatively simple and unique known genetic cause, a CAG expansion in the HD gene correlated with severity and onset of clinical symptoms, makes it particularly suited to therapeutic development. The Huntington’s disease (HD) iPS cell consortium, funded with NIH and CIRM support, brings together leading groups in stem cell and HD research to establish whether newly created iPS cell lines show HD related (i.e., CAG length-dependent) phenotypes. Human iPSC technology can be used to generate specific neuronal and glial cell types, permitting investigation of the effects of the genetic lesion in the susceptible human cell types in the context of HD. The monogenic nature of HD and the existence of allelic series of iPSCs with a range of CAG repeat lengths confer tremendous power to model HD. Through CIRM support this consortium has capitalized on new technologies to use non-integrating approaches for reprogramming and promising phenotypes in current HD iPS lines to develop robust and validated assays for drug development for HD.
  • Significant progress has been made through CIRM-funded support of this proposal. Notably, the Cedars-Sinai Medical Center’s Board of iPSC core housed in the Board of Governors Regenerative Medicine Institute has taken skin cells from HD patients with a wide range of CAG repeats (43 to 180), and unaffected healthy controls (21 to 33) and reprogrammed then to pluripotency using the latest non-integrating iPS cell technology. So far 18 well-characterized patient-specific iPSC lines have been generated. These new iPSC lines have been rigorously characterized by our iPSC core and available to HD research community throughout California and the world. The Svendsen lab and the other HD iPSC Consortium laboratories have already used these lines and differentiated into relevant neuronal cell types to study the disease mechanisms as well develop new treatment.
  • These cell lines will be an essential resource for academic groups and pharmaceutical companies for studying pathogenesis and for testing experimental therapeutics for HD. The ultimate goal is to develop and validate methods and assays using >96 well format for CAG repeat length-dependent phenotypes that are amenable to high content/throughput screening methods. Assays developed using these patient-specific iPSC lines and their neuronal derivatives will allow academic groups and pharmaceutical companies to study pathogenesis and test experimental therapeutics for HD, which will significantly advance both our understanding of HD and potential treatments for this devastating and currently untreatable disease.

Elucidating pathways from hereditary Alzheimer mutations to pathological tau phenotypes

Funding Type: 
Basic Biology V
Grant Number: 
RB5-07011
ICOC Funds Committed: 
$1 161 000
Disease Focus: 
Alzheimer's Disease
Neurological Disorders
Stem Cell Use: 
iPS Cell
Cell Line Generation: 
iPS Cell
oldStatus: 
Closed
Public Abstract: 
We propose to elucidate pathways of genes that lead from early causes to later defects in Alzheimer’s Disease (AD), which is common, fatal, and for which no effective disease-modifying drugs are available. Because no effective AD treatment is available or imminent, we propose to discover novel genetic pathways by screening purified human brain cells made from human reprogrammed stem cells (human IPS cells or hIPSC) from patients that have rare and aggressive hereditary forms of AD. We have already discovered that such human brain cells exhibit an unique biochemical behavior that indicates early development of AD in a dish. Thus, we hope to find new drug targets by using the new tools of human stem cells that were previously unavailable. We think that human brain cells in a dish will succeed where animal models and other types of cells have thus far failed.
Statement of Benefit to California: 
Alzheimer’s Disease (AD) is a fatal neurodegenerative disease that afflicts millions of Californians. The emotional and financial impact on families and on the state healthcare budget is enormous. This project seeks to find new drug targets to treat this terrible disease. If we are successful our work in the long-term may help diminish the social and familial cost of AD, and lead to establishment of new businesses in California using our approaches.

Molecular basis of plasma membrane characteristics reflecting stem cell fate potential

Funding Type: 
Basic Biology V
Grant Number: 
RB5-07254
ICOC Funds Committed: 
$1 003 590
Disease Focus: 
Neurological Disorders
Stem Cell Use: 
Adult Stem Cell
oldStatus: 
Closed
Public Abstract: 
Stem cells generate mature, functional cells after proteins on the cell surface interact with cues from the environment encountered during development or after transplantation. Thus, these cell surface proteins are critical for directing transplanted stem cells to form appropriate cells to treat injury or disease. A key modification regulating cell surface proteins is glycosylation, which is the addition of sugars onto proteins and has not been well studied in neural stem cells. We focus on a major unsolved problem in the neural stem cell field: do different proteins coated with sugars on the surfaces of cells in this lineage (neuron precursors, NPs and astrocyte precursors, APs) determine what types of mature cells will form? We hypothesize key players directing cellular decisions are glycosylated proteins controlling how precursors respond to extracellular cues. We will address this hypothesis with aims investigating whether (1) glycosylation pathways predicted to affect cell surface proteins differ between NPs and APs, (2) glycosylated proteins on the surface of NPs and APs serve as instructive cues governing fate or merely mark their fate potential, and (3) glycosylation pathways regulate cell surface proteins likely to affect fate choice. By answering these questions we will better understand the formation of NPs and APs, which will improve the use of these cells to treat brain and spinal cord diseases and injuries.
Statement of Benefit to California: 
The goal of this project is to determine how cell surface proteins differ between cells in the neural lineage that form two types of final, mature cells (neurons and astrocytes) in the brain and spinal cord. In the course of these studies, we will uncover specific properties of human stem cells that are used to treat neurological diseases and injuries. We expect this knowledge will improve the use of these cells in transplants by enabling more control over what type of mature cell will be formed from transplanted cells. Also, cells that specifically generate either neurons or astrocytes can be used for drug testing, which will help to predict the effects of compounds on cells in the human brain. We hope our research will greatly improve identification, isolation, and utility of specific types of human neural stem cells for treatment of human conditions. Furthermore, this project will generate new jobs for high-skilled workers and, hopefully, intellectual property that will contribute to the economic growth of California.

Paracrine and synaptic mechanisms underlying neural stem cell-mediated stroke recovery

Funding Type: 
Basic Biology V
Grant Number: 
RB5-07363
ICOC Funds Committed: 
$1 178 370
Disease Focus: 
Stroke
Neurological Disorders
Stem Cell Use: 
Embryonic Stem Cell
oldStatus: 
Closed
Public Abstract: 
Stem cell therapy holds promise for the almost million Americans yearly who suffer a stroke. Preclinical data have shown that human neural stem cells (hNSCs) aid recovery after stroke, resulting in a major effort to advance stem cell therapy to the clinic, and we are currently transitioning our hNSC product to the clinic for stroke therapy. In this proposal we will explore how these cells improve lost function. We have already shown that injected hNSCs secrete factors that promote the gross rewiring of the brain, a major component of the spontaneous recovery observed after stroke. We now intend to focus on the connections between neurons, the synapses, which are a critical part of this rewiring process. We aim to quantify the effect of hNSCs on synapse density and function, and explore whether the stem cells secrete restorative synaptogenic factors or form functional synapses with pre-existing neurons. Our pursuit is made possible by our combination of state-of-the-art imaging techniques enabling us to visualize, characterize, and quantify these tiny synaptic structures and their interaction with the hNSCs. Furthermore, by engineering the hNSCs we can identify the factors they secrete in the brain and identify those which modulate synaptic connections. Our proposed studies will provide important insight into how transplanted stem cells induce recovery after stroke, with potential applicability to other brain diseases.
Statement of Benefit to California: 
Cerebrovascular stroke is the fourth leading cause of mortality in the United States and a significant source of long-term physical and cognitive disability that has devastating consequences to patients and their families. In California alone, over 9% of adults 65 years or older have had a stroke according to a 2005 study. In the next 20 years the societal toll is projected to amount to millions of patients and 18.8 billion dollars per year in direct medical costs. To date, there is no approved therapeutic agent for the recovery phase after stroke, making the long-term care of stroke patients a tremendous socioeconomic burden that will continue to rise as our aging population increases. Our laboratory and others have demonstrated the promise of stem cell transplantation to treat stroke. We are dedicated to developing human neural stem cells (hNSCs) as a novel neuro-restorative treatment for lost motor function after stroke. The goal of our proposed work is to further understand how transplanted hNSCs improve stroke recovery, as dissecting the mechanism of action of stem cells in the stroke brain will ultimately improve the chance of clinical success. This could potentially provide significant cost savings to California, but more importantly benefit the thousands of Californians and their families who struggle with the aftermath of stroke.

Misregulated Mitophagy in Parkinsonian Neurodegeneration

Funding Type: 
Basic Biology V
Grant Number: 
RB5-06935
ICOC Funds Committed: 
$1 174 943
Disease Focus: 
Parkinson's Disease
Neurological Disorders
Stem Cell Use: 
iPS Cell
oldStatus: 
Closed
Public Abstract: 
Parkinson’s disease (PD), is one of the leading causes of disabilities and death and afflicting millions of people worldwide. Effective treatments are desperately needed but the underlying molecular and cellular mechanisms of Parkinson’s destructive path are poorly understood. Mitochondria are cell’s power plants that provide almost all the energy a cell needs. When these cellular power plants are damaged by stressful factors present in aging neurons, they release toxins (reactive oxygen species) to the rest of the neuron that can cause neuronal cell death (neurodegeneration). Healthy cells have an elegant mitochondrial quality control system to clear dysfunctional mitochondria and prevent their resultant devastation. Based on my work that Parkinson’s associated proteins PINK1 and Parkin control mitochondrial transport that might be essential for damaged mitochondrial clearance, I hypothesize that in Parkinson’s mutant neurons mitochondrial quality control is impaired thereby leading to neurodegeneration. I will test this hypothesis in iPSC (inducible pluripotent stem cells) from Parkinson’s patients. This work will be a major step forward in understanding the cellular dysfunctions underlying Parkinson’s etiology, and promise hopes to battle against this overwhelming health danger to our aging population.
Statement of Benefit to California: 
Parkinson's disease (PD), one of the most common neurodegenerative diseases, afflicts millions of people worldwide with tremendous global economic and societal burdens. About 500,000 people are currently living with PD in the U.S, and approximate 1/10 of them live in California. The number continues to soar as our population continues to age. An effective treatment is desperately needed but the underlying molecular and cellular mechanisms of PD’s destructive path remain poorly understood. This proposal aims to explore an innovative and critical cellular mechanism that controls mitochondrial transport and clearance via mitophagy in PD pathogenesis with elegant employment of bold and creative approaches to live image mitochondria in iPSC (inducible pluripotent stem cells)-derived dopaminergic neurons from Parkinson’s patients. This study is closely relevant to public health of the state of California and will greatly benefit its citizens, as it will illuminate the pathological causes of PD and provide novel targets for therapuetic intervention.

Molecular Imaging for Stem Cell Science and Clinical Application

Funding Type: 
Research Leadership 12
Grant Number: 
LA1_C12-06919
ICOC Funds Committed: 
$6 443 455
Disease Focus: 
Amyotrophic Lateral Sclerosis
Neurological Disorders
Spinal Cord Injury
oldStatus: 
Active
Public Abstract: 
Stem cells offer tremendous potential to treat previously intractable diseases. The clinical translation of these therapies, however, presents unique challenges. One challenge is the absence of robust methods to monitor cell location and fate after delivery to the body. The delivery and biological distribution of stem cells over time can be much less predictable compared to conventional therapeutics, such as small-molecule therapeutic drugs. This basic fact can cause road blocks in the clinical translation, or in the regulatory path, which may cause delays in getting promising treatments into patients. My research aims to meet these challenges by developing new non-invasive cell tracking platforms for emerging stem cell therapies. Recent progress in magnetic resonance imaging (MRI) has demonstrated the feasibility of non-invasive monitoring of transplanted cells in patients. This project will build on these developments by creating next-generation cell tracking technologies with improved detectability and functionality. Additionally, I will provide leadership in the integration of non-invasive cell tracking into stem cell clinical trials. Specifically, this project will follow three parallel tracks. (1) The first track leverages molecular genetics to develop new nucleic acid-based MRI reporters. These reporters provide instructions to program a cell’s innate machinery so that they produce special proteins with magnetic properties that impart MRI contrast to cells, and allow the cells to be seen. My team will create neural stem cell lines with MRI reporters integrated into their genome so that those neural stem cell lines, and their daughter cells, can be tracked days and months after transfer into a patient. (2) The second track will develop methods to detect stem cell viability in vivo using perfluorocarbon-based biosensors that can measure a stem cell's intracellular oxygen level. This technology can potentially be used to measure stem cell engraftment success, to see if the new cells are joining up with the other cells where they are placed. (3) The third project involves investigating the role that the host’s inflammatory response plays in stem cell engraftment. These studies will employ novel perfluorocarbon imaging probes that enable MRI visualization and quantification of places in the body where inflammation is occurring. Overall, MRI cell tracking methods will be applied to new stem cell therapies for amyotrophic lateral sclerosis, spinal cord injury, and other disease states, in collaboration with CIRM-funded investigators.
Statement of Benefit to California: 
California leads the nation in supporting stem cell research with the aim of finding cures for major diseases afflicting large segments of the state’s population. Significant resources are invested in the design of novel cellular therapeutic strategies and associated clinical trials. To accelerate the clinical translation of these potentially live saving therapies, many physicians need method to image the behavior and movement of cells non-invasively following transplant into patients. My research aims to meet these challenges by developing new cell tracking imaging platforms for emerging stem cell therapies. Recent progress in magnetic resonance imaging (MRI) has demonstrated the feasibility of non-invasive monitoring of transplanted cells in patients. This project will build on these developments by leading the integration of MRI cell tracking into stem cell clinical trials and by developing next-generation technologies with improved sensitivity and functionality. Initially, MRI cell tracking methods will be applied to new stem cell therapies for amyotrophic lateral sclerosis and spinal cord injury. In vivo MRI cell tracking can accelerate the process of deciding whether to continue at the preclinical and early clinical trial stages, and can facilitate smaller, less costly trials by enrolling smaller patient numbers. Imaging can potentially yield data about stem cell engraftment success. Moreover, MRI cell tracking can help improve safety profiling and can potentially lower regulatory barriers by verifying survival and location of transplanted cells. Overall, in vivo MRI cell tracking can help maximize the impact of the State’s investment in stem cell therapies by speeding-up clinical translation into patients. These endeavors are intrinsically collaborative and multidisciplinary. My project will create a new Stem Cell Imaging Center (SCIC) in California with a comprehensive set of ways to elucidate anatomical, functional, and molecular behavior of stem cells in model systems. The SCIC will provide scientific leadership to stem cell researchers and clinicians in the region, including a large number of CIRM-funded investigators who wish to bring state-of-the-art imaging into their clinical development programs. Importantly, the SCIC will focus intellectual talent on biological imaging for the state and the country. This project will help make MRI cell tracking more widespread clinically and position California to take a leadership role in driving this technology. An extensive infrastructure of MRI scanners already exist in California, and these advanced MRI methods would use this medical infrastructure better to advance stem cell therapies. Moreover, this project will lead to innovative new MRI tools and pharmaceutical imaging agents, thus providing economic benefits to California via the formation of new commercial products, industrial enterprises, and jobs.
Progress Report: 
  • Stem cells offer tremendous potential to treat previously intractable diseases. However, the clinical translation of these therapies presents unique challenges. One of which is the absence of robust methods to monitor cell location and fate after delivery to the body. The delivery and biological distribution of stem cells over time can be much less predictable compared to conventional therapeutics, such as small-molecule therapeutic drugs. This basic fact can cause road blocks in the clinical translation, or in the regulatory path, which may cause delays in getting promising treatments into patients. My research aims to meet these challenges by developing new non-invasive cell tracking platforms for emerging stem cell therapies. Recent progress in magnetic resonance imaging (MRI) has demonstrated the feasibility of non-invasive monitoring of transplanted cells in patients. This project will build on these developments, by creating next-generation cell tracking technologies with improved detectability and functionality. In year 1 of this project, we have begun to evaluate emerging stem cell imaging technologies called MRI reporters, or DNA-based instructions, that when placed into a cell’s genome causes the cell to produce a protein that is detectable with MRI. We have constructed human neural progenitor cell (NPC) lines that integrally contain the MRI reporter so that the primary cell and its progeny can be visualized using MRI. This technology enables long term tracking of the NPCs’ fate and movements in the body. We use an NPC cell type that is currently being used in clinical trials to treat major diseases such as ALS and spinal cord injury. Our initial MRI experiments in a model system have demonstrated MRI detection of NPCs following transfer into the brain. In other developments over the past year, we have helped build a new multi-modal in vivo molecular imaging center at the Sanford Consortium for Regenerative Medicine. This new resource is now fully functional and is able to serve a broad range of stem cell investigators at the Consortium, adjacent academic institutions, and local industry. Ongoing activities include the implementation of the most up-to-date methodologies for in vivo cell tracking using the molecular imaging instruments, as well as educating stem cell scientists at the Sanford Consortium and elsewhere in the region about the value of non-invasive imaging for accelerating their research.

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