This project is aimed at testing the feasibility of using human embryonic stem cell (hESC)-derived neural stem/progenitor cells (hNPCs) expressing constitutively active MEF2C (MEF2CA) transcriptional factor as a cell therapy candidate for stroke. A rodent stroke model is being tested for behavioral and histological improvement after transplantation with hNPCs stably expressing MEF2CA (designated here as stable-MEF2CA hNPCs). Currently, we are optimizing this therapeutic candidate for most efficient and tolerated dose, and generating scalable quantities of hESC-derived stable-MEF2CA hNPCs. These cells are transplanted into the brain of the spontaneously hypertensive rat (SHR) model of focal stroke. We will then evaluate behavioral and histological improvements, and perform preliminary safety assessments. Leading up to these experiments, during the past year, we completed a direct comparison of two different types of MEF2CA-programmed candidate stem cell products under two expression systems (stable transformant vs. transient AAV-infection) and found that the stable-MEF2CA hNPC line was superior. Specifically, we performed twelve-week behavioral assessments in the living animals and then sacrificed them to conduct histological evaluations of postmortem brain samples. The results from these experiments indicate significant improvement compared to saline-injected animals. Moreover, there was no significant difference in behavioral improvement or in vivo histological analysis between AAV infection-mediated MEF2CA expression vs. stable-MEF2CA hNPCs. In light of these results, and given that the stable-MEF2CA hNPC line consists of homogenous cells that are more amenable for scale-up for future clinical trials, we concluded with CIRM Program approval that we should move forward with the stable-MEF2CA line for the reminder of the studies. In order to determine the optimal therapeutic dose of cells for transplantation, we are currently conducting dose-response experiments by transplanting three doses of stable-MEF2CA-hNPCs into the striatum of our rat stroke model. At the time of this report we are completing these transplantation experiments and are beginning behavioral assessments in these animals.
Reporting Period:
Year 2
This project tested the feasibility of using human embryonic stem cell (hESC)-derived neural stem/progenitor cells (hNPCs) expressing constitutively active MEF2C (MEF2CA) transcription factor as a cell therapy candidate for stroke. A rodent stroke model (spontaneously hypertensive rat (SHR) model of focal stroke) was used for testing behavioral and histological improvement after transplantation with hNPCs stably expressing MEF2CA (designated here as stable-MEF2CA hNPCs). We first compared two different types of MEF2CA-programmed candidate stem cell products under two expression systems (stable transformant vs. transient AAV-infection) and found that the stable-MEF2CA hNPC line was superior. In order to determine the optimal therapeutic dose of cells for transplantation, we next conducted dose-response experiments by transplanting three doses (500K, 1 million, or 3 million cells per animal) of stable-MEF2CA-hNPCs into the brain of our rat model after stroke. We found that the dose of 3 million cells exhibited limiting toxicity. In contrast, the animals that received either the 1 million or 500K dose of cells showed significant behavioral improvements over a 12-week observation period, along with histological evidence of graft survival and differentiation into neurons in the absence of noticeable inflammation, overt toxicity, tumor formation/hyperproliferation, or presence of grafted cells in ectopic locations in the body. Since we did not observe any clear advantage using a higher dose (1 million cells/animal), we conclude that a dose of 500K cells in rodents is most effective, and an equivalent dose estimated based on this rodent dose will serve as a starting point for future testing of this therapeutic candidate. In summary, we have achieved each of the success criteria that had been set for all the milestones with excellent results that support further therapeutic development of our product.
Grant Application Details
Application Title:
Programming Human ESC-derived Neural Stem Cells with MEF2C for Transplantation in Stroke
Public Abstract:
The goal of this project is to produce a stem cell-based therapy for stroke (also known as an ischemic cerebral infarct). Stroke is the third leading cause of death in the USA, and a leading cause of disability among adults. Currently, there are no effective treatments once a stroke has occurred (termed completed stroke). In this proposal, we aim to develop human stem cells for therapeutic transplantation to treat stroke. Potential benefits will outweigh risks because only patients with severe strokes that have compromised activities of daily living to an extreme degree will initially be treated. Using a novel approach, we will generate stem cells that do not form tumors, but instead only make new nerve cells. We will give drugs to avoid rejection of the transplanted cells. Thus, the treatment should be safe. We will first test the cells in stroke models in rodents (mice and rats) in preparation for a human clinical trial. We will collect comprehensive data on the mice and rats to determine if the stem cells indeed become new nerve cells to replace the damaged tissue and to assess if the behavior of the mice and rats has improved. If successfully developed and commercialized, this approach has the potential for revolutionizing stroke therapy.
Statement of Benefit to California:
The goal of this project is to produce a stem cell-based therapy for stroke (also known as an ischemic cerebral infarct). Stroke is the third leading cause of death in the State of California, and a leading cause of disability among adults. Currently, there are no effective treatments once a stroke has occurred (termed completed stroke), and the quality of life is severely compromised in those that survive the malady. In this proposal, we aim to develop human stem cells for therapeutic transplantation to treat stroke. Using a novel approach, we will generate stem cells that do not form tumors, but instead only make new nerve cells. If successfully developed and commercialized, this approach could provide a therapeutic candidate for the unmet medical need, which would have a tremendous impact on the quality of life for the patient, his or her family, and for the economic and emotional burden on the State of California and its citizens.
