Therapeutic potential of genetically modified human ES cells in an Alzheimer's disease model: Contribution of IGF-1

Funding Type: 
SEED Grant
Grant Number: 
ICOC Funds Committed: 
Disease Focus: 
Blood Cancer
Stem Cell Use: 
Cancer Stem Cell
Embryonic Stem Cell
Cell Line Generation: 
Cancer Stem Cell
Public Abstract: 
Alzheimer’s disease (AD) is a progressive and irreversible disease of the brain leading to deterioration of mental function and eventual morbidity and death. The major defining characteristic of AD brains is the excessive accumulation of amyloid plaques (composed of clumps of Abeta) outside of nerve cells and tangles (composed of clumps of tau) inside nerve cells. These lesions are toxic to nerve cells and likely explain the progressive degeneration seen in AD brains. Currently available treatments for AD provide only limited symptomatic relief and are unable to prevent, stop, or cure the disease. Even if next generation drugs prove to be more effective, they are unlikely to reverse the disease progression. Thus, it may be necessary to replace dead or dying nerve cells in order to reverse the course of the disease in many AD patients. The long-term objective of this proposal is to use genetically modified human embryonic stem cells (ESCs) as an inexhaustible source for replacing lost or damaged nerve cells, supplying the host brain with protection from further damage, and working against the underlying factors that promote amyloid and tangle lesions. Such objective ultimately may lead to a strategy for therapeutic intervention in AD patients who do not respond to available pharmacological treatments. It is well known that mouse embryonic stem cells exhibit the remarkable ability to respond to damaged nerve cells and home in on these degenerative environments in brain. At present, the capacity of human embryonic stem cells (ESCs) to integrate into the diseased brain such as those with amyloid and tangle lesions is unknown. In this proposal, we will use a mouse model of AD that develops both amyloid plaques and tangles to test the idea that transplantation of ESCs might be beneficial in treating AD. Our hypothesis is that human ESCs possess the inherent capacity to home in and integrate into sites surrounding plaques and tangles, where nerve cell damage is occurring. In addition, we hypothesize that human ESCs genetically modified to produce a protective factor called IGF-1 will further enhance this capability, help host nerve cells from further damage, and block the accumulation of plaques and tangles. It is known that IGF-1 promotes ESCs to become nerve cells, protects nerve cells from damage by Abeta, and decreases the levels of Abeta in brain. Furthermore, IGF-1 levels are reduced in AD, and loss of IGF-1 promotes tangle-like lesions in mice. If the above hypotheses can be even partially demonstrated, the current proposal is expected significantly advance our long-term objective of applying genetically modified human ESCs as a therapeutic technology for AD patients who are refractory to available pharmacological treatments.
Statement of Benefit to California: 
Alzheimer’s disease (AD) is an age-related debilitating disease of the brain characterized by progressive deterioration of mental function and accounts for more than 70% of all dementias of the brain. AD inflicts more than 465,000 residents in California alone and places substantial medical, social, psychological, and financial burden on the patients, their families, and social/medical institutions. The per capita cost of caring for an AD patient in California was estimated to be more than $65,000 per year in 1998. It was also projected at the time that the cost of caring for AD patients in California (in 1998 dollars) will be ~$25.9 billion in 2000, ~$47.5 billion in 2020, and ~$75.4 billion in 2040. During the same time period, the number of AD patients in California is projected to rise from ~395,000 in 2000 to ~1.2 million in 2040. At present, no effective treatment is available for AD. First generation drugs can temporarily mask symptoms of the disease but rapidly lose effectiveness during the progression of AD. Even if next generation drugs prove to be more effective, they will only help to slow down the progression of AD but not reverse it. As such, it may be necessary use an alternate therapeutic strategy to replace dead or dying nerve cells, especially in patients that do not respond to available drugs. Human embryonic stem cells have emerged in recent years to hold enormous potential for cell replacement therapy for wide variety of neurological disorders, including AD. As California continues to be at the forefront of new and innovative technologies, the passage of Proposition 71 to fund stem cell research further extends this spirit of innovation. The research proposed in this application attempts to generate genetically modified human embryonic stem cells capable of not only replacing lost nerve cells but also delivering protective factors that prevent further degeneration of existing nerve cells in an animal model of AD. Such kind of technological coupling between stem cell therapy and gene therapy poses therapeutic potential for application in AD where irreversible nerve cell damage cannot be treated with even the best of next generation drugs. If successful, this will also help to offset the enormous social and financial burden of caring for AD patients in California. Technologies and therapeutics derived from stem cell research funded by the California Institute for Regenerative Medicine (CIRM) are in part the contractual property of the state of California, and hence its residents. In the event that such intellectual property leads to commercialization or licensing down the line, a portion of the proceeds are contracted to enter the California state general fund, ensuring that all California residents benefit from potential successes of this research.
Progress Report: 
  • SEED Grant Research Summary
  • Compelling studies suggest that cancer stem cells (CSC) arise from primitive self-renewing progenitor cells. Although many cancer therapies target rapidly dividing cells, CSC may be quiescent i.e. asleep resulting in therapeutic resistance. Recently, we demonstrated that CSC drive progression of chronic phase (CP) chronic myeloid leukemia (CML), a subject of many landmark cancer research discoveries, to a therapeutically recalcitrant myeloid blast crisis (BC) phase. CML CSC share cell surface markers with granulocyte-macrophage progenitors (GMP) and have amplified expression of the CML fusion gene, BCR-ABL. In addition, they aberrantly gain self-renewal capacity, in part, as a result Wnt/β-catenin activation. Because human embryonic stem cells (hESC) have robust regenerative capacity and can provide a potentially limitless source of tissue specific progenitor cells in vitro, they represent an ideal model system for generating and characterizing human CSC. The main goals of this research were to generate CSC from hESC to provide an experimentally amenable platform to expedite the development of sensitive diagnostics that predict progression and combined modality anti-CSC therapy.
  • To this end, we tested whether BCR-ABL expression in hESC is sufficient to induce changes characteristic of CML stem cells. Unlike mouse ESC, introduction of a novel lentiviral BCR-ABL vector into hESC did not drive myeloid differentiation nor did it induce stromal independence in vitro underscoring key differences between mouse and human hESC and the importance of in vivo models. Notably, Hues16 cells had a higher propensity to differentiate into CD34+ cells than other hESC lines particularly in AGM co-cultures and thus, were used in subsequent in vivo experiments. Moreover, this SEED grant funded Yosuke Minami in Professor Jean Wang’s lab to create a unique CML blast crisis mouse model typified by GMP expansion and resistance to a BCR-ABL inhibitor, imatinib (Minami et al, PNAS 2008;105:17967-72). In addition, a bioluminescent humanized model of blast crisis CML was created based on transplantation of GMP from patient blood into immune deficient mice (RAG2-/-gc-/-). Cells were tagged with firefly luciferase that emits a bioluminescent signal so that leukemic transplantation efficiency could be tracked in vivo (IVIS). As few as 1,000 human blast crisis CML GMP could transplant leukemia in immune deficient mice thereby providing an important model for studying the molecular events that contribute to leukemic transformation (Abrahamsson et al, PNAS 2009;106:3925-9).
  • In the second aim, we hypothesized that BCR-ABL is sufficient for generating CML from self-renewing stem cells. In these studies, Hues16 cells differentiated into CD34+ cells were lentivirally transduced with BCR-ABL leading to sustained BCR-ABL engraftment in 50% of transplanted mice. Chronic phase CD34+ cells derived from CML blood were less efficient at sustaining CML engraftment (7%) suggesting that hESC derived CD34+ cells have higher self-renewal potential and are similar to advanced phase CML progenitors.
  • Thirdly, we hypothesized that BCR-ABL was necessary but not sufficient for progression to blast crisis. Introduction of lentiviral activated beta-catenin or shRNA to GSK3beta, together with BCR-ABL did not enhance BCR-ABL engraftment compared with BCR-ABL transduction of hESC alone. These studies suggested that hESC may already have sufficient self-renewal capacity to sustain the malignant CML clone and are molecularly comparable to advanced CML progenitors that behave like CSC. In addition, through extensive cDNA sequencing of human blast crisis CML progenitors, we found that 57% of samples harbored a misspliced form of GSK3beta that promoted tumor production and could serve as a novel prognostic marker in CML clinical trials (Abrahamsson et al, PNAS 2009;106:3925-9).
  • In the final aim, we hypothesized that CML CSC are not eliminated by BCR-ABL inhibitors alone and that combined modality therapy will be required. In collaborative research involving in vitro analysis of imatinib resistant CML progenitors and more recently in a humanized mouse model of blast crisis CML, we found that dasatinib, a potent BCR-ABL inhibitor, is necessary but not sufficient for CSC eradication. Discovery of a GSK3beta deregulation, a negative regulator of both beta-catenin and sonic hedgehog (Shh) pathways (Zhang et al, Nature 2009), led us to disover that Shh combined with BCR-ABL inhibition abrogated CSC driven tumor formation (manuscript in preparation) providing the impetus for an upcoming Pfizer sponsored Shh inhibitor clinical trial for refractory hematologic malignancies.

© 2013 California Institute for Regenerative Medicine