Heart Disease

Coding Dimension ID: 
295
Coding Dimension path name: 
Heart Disease
Funding Type: 
hiPSC Derivation
Grant Number: 
ID1-06557
Investigator: 
Type: 
PI
ICOC Funds Committed: 
$16 000 000
Disease Focus: 
Developmental Disorders
Genetic Disorder
Heart Disease
Infectious Disease
Alzheimer's Disease
Neurological Disorders
Autism
Respiratory Disorders
Vision Loss
Liver Disease
Cell Line Generation: 
iPS Cell
oldStatus: 
Active
Public Abstract: 

Induced pluripotent stem cells (iPSCs) have the potential to differentiate to nearly any cells of the body, thereby providing a new paradigm for studying normal and aberrant biological networks in nearly all stages of development. Donor-specific iPSCs and differentiated cells made from them can be used for basic and applied research, for developing better disease models, and for regenerative medicine involving novel cell therapies and tissue engineering platforms. When iPSCs are derived from a disease-carrying donor; the iPSC-derived differentiated cells may show the same disease phenotype as the donor, producing a very valuable cell type as a disease model. To facilitate wider access to large numbers of iPSCs in order to develop cures for polygenic diseases, we will use a an episomal reprogramming system to produce 3 well-characterized iPSC lines from each of 3,000 selected donors. These donors may express traits related to Alzheimer’s disease, autism spectrum disorders, autoimmune diseases, cardiovascular diseases, cerebral palsy, diabetes, or respiratory diseases. The footprint-free iPSCs will be derived from donor peripheral blood or skin biopsies. iPSCs made by this method have been thoroughly tested, routinely grown at large scale, and differentiated to produce cardiomyocytes, neurons, hepatocytes, and endothelial cells. The 9,000 iPSC lines developed in this proposal will be made widely available to stem cell researchers studying these often intractable diseases.

Statement of Benefit to California: 

Induced pluripotent stem cells (iPSCs) offer great promise to the large number of Californians suffering from often intractable polygenic diseases such as Alzheimer’s disease, autism spectrum disorders, autoimmune and cardiovascular diseases, diabetes, and respiratory disease. iPSCs can be generated from numerous adult tissues, including blood or skin, in 4–5 weeks and then differentiated to almost any desired terminal cell type. When iPSCs are derived from a disease-carrying donor, the iPSC-derived differentiated cells may show the same disease phenotype as the donor. In these cases, the cells will be useful for understanding disease biology and for screening drug candidates, and California researchers will benefit from access to a large, genetically diverse iPSC bank. The goal of this project is to reprogram 3,000 tissue samples from patients who have been diagnosed with various complex diseases and from healthy controls. These tissue samples will be used to generate fully characterized, high-quality iPSC lines that will be banked and made readily available to researchers for basic and clinical research. These efforts will ultimately lead to better medicines and/or cellular therapies to treat afflicted Californians. As iPSC research progresses to commercial development and clinical applications, more and more California patients will benefit and a substantial number of new jobs will be created in the state.

Progress Report: 
  • First year progress on grant ID1-06557, " Generation and Characterization of High-Quality, Footprint-Free Human Induced Pluripotent Stem Cell (iPSC) Lines From 3000 Donors to Investigate Multigenic Disease" has met all agreed-upon milestones. In particular, Cellular Dynamics International (CDI) has taken lease to approximately 5000 square feet of lab space at the Buck Institute for Research on Aging in Novato, CA. The majority of this space is located within the new CIRM-funded Stem Cell Research Building at the Buck Institute and was extensively reconfigured to meet the specific needs of this grant. All equipment, including tissue culture safety cabinets and incubators, liquid-handling robotics, and QC instrumentation have been installed and qualified. A total of 16 scientists have been hired and trained (13 in Production and 3 in Quality) and more than 20 Standard Operating Procedures (SOPs) have been developed and approved specifically for this project. These SOPs serve to govern the daily activities of the Production and Quality staff and help ensure consistency and quality throughout the iPSC derivation and characterization process. In addition, a Laboratory Information Management System (LIMS) had to be developed to handle the large amount of data generated by this project and to track all samples from start to finish. The first and most important phase of this LIMS project has been completed; additional functionalities will likely be added to the LIMS during the next year, but completion of phase 1 will allow us to enter full production mode on schedule in the first quarter of year 2. Procedures for the shipping, infectious disease testing, and processing of donor samples were successfully implemented with the seven Tissue Collectors. To date, over 700 samples have been received from these Tissue Collectors and derivation of the first 50 patient-derived iPSC lines has been completed on schedule. These cells have been banked in the Coriell BioRepository, also located at the Buck Institute. The first Distribution Banks will be available for commercial release during year 2.
Funding Type: 
Early Translational III
Grant Number: 
TR3-05593
Investigator: 
Institution: 
Type: 
PI
ICOC Funds Committed: 
$6 319 110
Disease Focus: 
Heart Disease
Stem Cell Use: 
Directly Reprogrammed Cell
oldStatus: 
Active
Public Abstract: 

Heart disease is a leading cause of mortality. The underlying pathology is typically loss of heart muscle cells that leads to heart failure. Because heart muscle has little or no regenerative capacity after birth, current therapeutic approaches are limited for the over 5 million Americans who suffer from heart failure. Our recent findings regarding direct reprogramming of a type of structural cell of the heart, called fibroblasts, into cardiac muscle-like cells using just three genes offers a novel approach to achieving cardiac regeneration. 50% of cells in the human heart are cardiac fibroblasts, providing a potential source of new heart muscle cells for regenerative therapy. We simulated a heart attack in mice by blocking the coronary artery, and have been able to reprogram existing mouse cardiac fibroblasts in to new muscle by delivering the three genes into the heart. We found a significant reduction in scar size and an improvement in cardiac function that persists after injury. The reprogramming of cells in the intact organ was more complete than in cells in a dish. We now propose to develop the optimal gene therapy approach to introduce cardiac reprogramming genes into the heart, to establish the optimal delivery approach to administer virus encoding cardiac reprogramming factors that results in improvement in cardiac function in a preclinical model of cardiac injury, and to establish the safety profile of in vivo cardiac reprogramming in a preclinical model.

Statement of Benefit to California: 

This research will benefit the state of California and its citizens by helping develop a new therapeutic approach to cardiac regeneration. Heart disease is a leading cause of death in adults and children in California, but there is no current treatment that can promote cardiac regeneration. This proposal will lay the groundwork for a clinical trial that could result in generation of new heart muscle cells from within the heart. If successful, there is potential economic benefit in terms of productive lives saved and in the commercialization of this technology.

Progress Report: 
  • Heart disease is a leading cause of mortality. The underlying pathology is typically loss of heart muscle cells that leads to heart failure. Because heart muscle has little or no regenerative capacity after birth, current therapeutic approaches are limited for the over 5 million Americans who suffer from heart failure. Our recent findings regarding direct reprogramming of a type of structural cell of the heart, called fibroblasts, into cardiac muscle-like cells using just three genes offers a novel approach to achieving cardiac regeneration. 50% of cells in the human heart are cardiac fibroblasts, providing a potential source of new heart muscle cells for regenerative therapy. We simulated a heart attack in mice by blocking the coronary artery, and have been able to reprogram existing mouse cardiac fibroblasts into new muscle by delivering the three genes into the heart. We found a significant reduction in scar size and an improvement in cardiac function that persists after injury. The reprogramming of cells in the intact organ was more complete than in cells in a dish. We now identified a combination of factors that reprogram human and pig cardiac fibroblasts and are optimizing a gene therapy approach to introduce cardiac reprogramming genes into the heart of pigs. In a pig model of cardiac injury, these factors were able to convert non-muscle cells into new muscle in the area of injury. We also found a viral vector that can preferentially infect the fibroblasts compare to the muscle cells. We are now in a position to test for functional improvement in pigs.
  • Heart disease is a leading cause of mortality. The underlying pathology is typically loss of heart muscle cells that leads to heart failure. Because heart muscle has little or no regenerative capacity after birth, current therapeutic approaches are limited for the over 5 million Americans who suffer from heart failure. Our recent findings regarding direct reprogramming of a type of structural cell of the heart, called fibroblasts, into cardiac muscle-like cells using just three genes offers a novel approach to achieving cardiac regeneration. We simulated a heart attack in mice by blocking the coronary artery, and have regenerated damaged hearts by converting existing mouse cardiac fibroblasts into new muscle by delivering the three genes into the heart. We have found that a combination of the three genes used in mice plus two additional factors were sufficient to identified to reprogram human and pig cardiac fibroblasts and are optimizing a gene therapy approach to introduce cardiac reprogramming genes into the heart of pigs. In a pig model of cardiac injury, we identified the optimal combination of factors that was able to convert non-muscle cells into new muscle in the area of injury. We have completed a pilot study of these five factors for functional improvement using MRI to measure cardiac output 3 days after injury and 2 months after treatment with the reprogramming factors. We also found a viral vector that can preferentially infect the fibroblasts compare to the muscle cells and have confirmed this activity. We are now testing for functional improvement in pigs using various viral vectors.
Funding Type: 
Research Leadership
Grant Number: 
LA1-08015
Investigator: 
Institution: 
Type: 
PI
ICOC Funds Committed: 
$6 368 285
Disease Focus: 
Heart Disease
Neurological Disorders
Pediatrics
Stem Cell Use: 
Embryonic Stem Cell
iPS Cell
Directly Reprogrammed Cell
Public Abstract: 

Tissues derived from stem cells can serve multiple purposes to enhance biomedical therapies. Human tissues engineered from stem cells hold tremendous potential to serve as better substrates for the discovery and development of new drugs, accurately model development or disease progression, and one day ultimately be used directly to repair, restore and replace traumatically injured and chronically degenerative organs. However, realizing the full potential of stem cells for regenerative medicine applications will require the ability to produce constructs that not only resemble the structure of real tissues, but also recapitulate appropriate physiological functions. In addition, engineered tissues should behave similarly regardless of the varying source of cells, thus requiring robust, reproducible and scalable methods of biofabrication that can be achieved using a holistic systems engineering approach. The primary objective of this research proposal is to create models of cardiac and neural human tissues from stem cells that can be used for various purposes to improve the quality of human health.

Statement of Benefit to California: 

California has become internationally renowned as home to the world's most cutting-edge stem cell biology and a global leader of clinical translation and commercialization activities for stem cell technologies and therapies. California has become the focus of worldwide attention due in large part to the significant investment made by the citizens of the state to prioritize innovative stem cell research as a critical step in advancing future biomedical therapies that can significantly improve the quality of life for countless numbers of people suffering from traumatic injuries, congenital disorders and chronic degenerative diseases. At this stage, additional investment in integration of novel tissue engineering principles with fundamental stem cell research will enable the development of novel human tissue constructs that can be used to further the translational use of stem cell-derived tissues for regenerative medicine applications. This proposal would enable the recruitment of a leading biomedical engineer with significant tissue engineering experience to collaborate with leading cardiovascular and neural investigators. The expected result will be development of new approaches to engineer transplantable tissues from pluripotent stem cell sources leading to new regenerative therapies as well as an enhanced understanding of mechanisms regulating human tissue development.

Funding Type: 
Tools and Technologies III
Grant Number: 
RT3-07899
Investigator: 
Type: 
PI
ICOC Funds Committed: 
$1 368 517
Disease Focus: 
Heart Disease
Stem Cell Use: 
Embryonic Stem Cell
Cell Line Generation: 
Embryonic Stem Cell
Public Abstract: 

Heart, stroke and other cardiovascular diseases are responsible for ~17 million deaths per year globally and this number is predicted to reach 23.3 million by 2030. Cardiovascular diseases impose a staggering annual cost of $300 billion on the U.S. health care system. Heart transplantation is the ultimate solution to end-stage heart failure. However, a major limitation in treating cardiac injury is the limited availability of donors; as a result, only a small fraction of patients will benefit from heart transplantation. Tissue engineering holds a great promise to create functional tissue constructs that can reestablish the structure and function of injured tissue with exciting success stories. However, many challenges regarding their development still remain. It is the goal of this project to develop a novel 3D bioprinting technology to fabricate cardiac tissues made from cell-laden hydrogels with engineered microvasculature. By integrating the advanced 3D bioprinting with stem cell technology, functional cardiac tissues will be created with biomimetic 3D microarchitecture and functional vasculature. This novel 3D-printed cardiac tissue will heal the damaged heart and improve its function to pave the way for a superior treatment option for the millions of cardiac patients in the U.S.

Statement of Benefit to California: 

Heart disease and other cardiovascular diseases are the #1 killer in California and remain a leading cause of disability and death. A major limitation in treating cardiac injury is the failure of current therapies to induce myocardium regeneration. Due to the limited availability of donors, only a fraction of individuals who could benefit from heart transplantations actually receive them. One possible avenue for remedying this situation is to artificially engineer cardiac tissues. Tissue engineering techniques have been successfully applied to engineer many types of tissue; however, many challenges regarding their development still remain. This proposal aims to make an advance in tissue engineering by developing a novel 3D bioprinting technology to fabricate tissues made from cell-laden hydrogels with engineered microvasculature. The completion of this work will be a paradigm shift and a landmark achievement in efforts towards clinical treatments of vascularized cardiac tissue using stem cells. This advanced technology can also have a significant economical impact as heart diseases impose a staggering annual cost of $300 billion on the U.S. health care system. In addition, the development of the 3D bioprinting technology and advanced biomaterials will keep California and the U.S. as a whole in the leading position in this emerging field.

Funding Type: 
Tools and Technologies III
Grant Number: 
RT3-07838
Investigator: 
Institution: 
Type: 
PI
Institution: 
Type: 
Co-PI
ICOC Funds Committed: 
$899 728
Disease Focus: 
Heart Disease
Stem Cell Use: 
Embryonic Stem Cell
Public Abstract: 

As ongoing CIRM-funded development of regenerative medicine (RM) progresses, the demand for increasing numbers of pluripotent stem cells and their differentiated derivatives has also increased. We have established a scalable suspension culture system for the production of large quantities of hESC for banking and to seed production of a number of regenerative medicine cell types, notably retinal pigmented epithelia, neural stem cells, dopaminergic neurons and cardiomyocytes, that support a number of CIRM and NIH-funded groups. In addition, we have adapted this system for the suspension production of several hESC derivative cell types, notably cardiomyocytes. While our system has provided unprecedented production capability for a number of cell products in pre-clinical and imminent clinical studies, it has proven impractical to scale up to the level that will be required for clinical trials for some hESC cell products, notably cardiomyocytes, due to high expected human doses. This project will resolve this scale-up challenge by adapting our suspension cell culture system, that is limited to 1-3L spinner culture flasks, to a more readily scalable and controllable suspension bioreactor system that utilizes “bags” capable of volumes up to 500L. Achieving this objective will remove a key barrier to progressing RM for cardiac applications as well as open the door to large clinical trials and commercialization of other regenerative medicine cell products in the years to come.

Statement of Benefit to California: 

We have developed GMP-compliant suspension cell culture processes for scalable production of hPSC and derivatives. These processes have been invaluable in our support of CIRM- and NIH-funded regenerative medicine projects, including those with RPE, NSC, DA neurons and cardiomyocytes (CM), as well as for production of GMP banks of hPSC for various projects. Our GMP-compliant suspension culture CM production process has made pre-clinical animal studies and small early clinical trials practical. However, while our current CM system is readily transferred to other groups and is meeting current production requirements, the scale requirements for anticipated high dose clinical trials is beyond the practical limitation of our spinner flask-based system. hPSC and CM are sensitive to changes in shear encountered at every scale-up step and re-optimizing conditions at each step is prohibitively expensive. Our experience using bag-based bioreactors for non-hESC products suggests that scale-up in bags will be more controllable and predictable than spinners or stir-tanks reactors. It is also a readily transferred technology. We propose to adapt our suspension hPSC and CM processes to a bag system, optimize conditions at a small scale, then demonstrate scalability at a moderate scale. Success in this project will remove a key barrier to developing many regenerative medicine products, and in particular those where high human doses are anticipated, such as CM.

Funding Type: 
Early Translational III
Grant Number: 
TR3-05556
Investigator: 
Name: 
Institution: 
Type: 
PI
Type: 
Partner-PI
ICOC Funds Committed: 
$4 766 231
Disease Focus: 
Heart Disease
Collaborative Funder: 
Germany
Stem Cell Use: 
Embryonic Stem Cell
oldStatus: 
Active
Public Abstract: 

Heart disease is the number one cause of morbidity and mortality in the US. With an estimated 1.5 million new or recurrent myocardial infarctions, the total economic burden on our health care system is enormous. Although conventional pharmacotherapy and surgical interventions often improve cardiac function and quality of life, many patients continue to develop refractory symptoms. Thus, the development of new therapies is urgently needed. “Tissue engineering” can be broadly defined as the application of novel bioengineering methods to understand complex structure-function relationships in normal or pathological conditions and the development of biological substitutes to restore, maintain, or improve function. It is different from “cell therapy”, which is designed to improve the function of an injured tissue by simply injecting suspensions of isolated cells into the injury site. To date, two main limitations of cell therapy are (1) acute donor cell death due to unfavorable seeding environment and (2) the lack of suitable cell type that genuinely resembles human cardiac cells. Our proposal seeks to use engineered tissue patches seeded with human embryonic stem cell-derived cardiomyocytes for treatment of ischemic heart disease in small and large animal models. It represents a significant development of novel techniques to address both of the main limitations of cell therapy, and will provide a new catalyst for the entire field of stem cell-based tissue engineering.

Statement of Benefit to California: 

Patients with end-stage heart failure have a 2-year survival rate of 25% by conventional medical therapy. Not commonly known to the public is that this dismal survival rate is actually worse when compared to patients with AIDS, liver cirrhosis, or stroke. Following a heart failure, the endogenous repair process is not sufficient to compensate for cardiomyocyte death. Thus, novel therapies with stem cells in combination with supportive scaffolds to form engineered cardiac tissue grafts is emerging as a promising therapeutic avenue. Engineered tissues have now been used to make new bladders for patients needing cystoplasty, bioarticial heart patches seeded with bone marrow cells, and more recently new trachea for patient with late stage tracheal cancer. Our multi-disciplinary team intends to push the therapeutic envelop by developing human tissue engineered myocardium for treatment of post-myocardial infarction heart failure. We will first test our engineered cardiac tissue in small and large animal models. We will perform extensive quality control measures to define morphological, molecular, and functional properties. At the end of 3 years, we are confident we will be able to derive a lead candidate that can move into IND-enabling preclinical development. These discoveries will benefit the millions of patients with heart failure in California and globally.

Progress Report: 
  • Despite advances in medical and device therapies, patients with end-stage heart failure have a survival rate of only 25% during the first 2 years following their diagnosis. Heart failure typically follows from damage induced by severe myocardial infarction (MI; heart attack). After a severe MI, the human heart may lose up to 1 billion heart muscle cells (cardiomyocytes). For most of these patients, heart transplantation is the only useful therapy, but there is a severe shortage of donor hearts. Recently, left ventricular assist devices (LVADs) have become available to take over the pumping function of the crucial left ventricle chamber of the heart. These devices were originally used as “bridge to transplant” (a temporary measure to keep patients alive until a new heart became available); recently some patients have received LVADs as “destination therapies” (permanent substitutes for transplanted hearts). The problems associated with these mechanical implants, however, include increased risk of stroke (blood clots that form due to the devices) and infection (the LVADs are powered from batteries that are carried outside the body and require wires to pierce the skin).
  • We are working to develop cardiac regenerative medicine using Engineered Heart Muscle (EHM). We are using human embryonic stem cells (hESCs) because they can be grown in very large quantities and, with the appropriate methods, can be triggered to differentiate into the cardiomyocytes, fibroblasts and smooth muscle that are lost after MI. Because these cells can be produced in essentially unlimited quantities, we could theoretically treat a very large number of patients who currently have no options.
  • During the first year of this project, we have a) established methods for producing the multi-billion quantities of hESC-derived cells needed to address this problem; b) developed methods to freeze and ship these cells to our collaborator in Germany for EHM assembly, and c) used these cells to generate 2 different forms of EHMs to compare their survival and function both in vitro (composition, force generated) and in vivo (after transplantation into rats that have been given MIs). We are now refining the EHM design with the goal of moving forward to testing them in animals with more human-like hearts (based on size and heart rate); this step will be essential to evaluate their safety and function before any clinical trial.
  • The project “Heart Repair with Human Tissue Engineered Myocardium” is designed to find a new option for the treatment of heart failure. Because of the shortage of donor hearts, many patients in need never receive this life-saving therapy. We are generating engineered heart muscles (EHMs) that are made from cardiomyocytes (heart muscle cells) derived from human embryonic stem cells. The ultimate goal of this work is to produce a beating human heart “patch” that can be transplanted onto damaged hearts, and help restore function.
  • Through the joint efforts of researchers in Dr. Joseph Wu’s laboratory at Stanford and Dr. Larry Couture’s team at City of Hope, we have developed a process that allows essentially unlimited generation of cardiomyocytes using a process that is fully compatible with eventual clinical use. Our collaborator at Gottingen University, Dr. Wolfram Zimmerman, uses these cells to produce EHMs, which are then shipped to Stanford. At Stanford, the EHMs are evaluated for their structure, overall health, and ability to generate force as measured in vitro. These EHMs are also transplanted into rodents that have been given heart attacks, to see if the EHMs can survive and improve heart function.
  • In the first year of this project, we compared different methods of making EHMs and the results that could be measured both in vitro and in vivo. We established a model of heart disease in rats with defective immune systems (necessary for the survival of human cells/tissues in this extremely foreign setting). We found that a specific grid-like patch design was both easier to construct than other options and was able to survive in the rat model of heart disease.
  • In the 2nd year, we focused on this patch design and performed a larger number of transplants. Using EHMs made from genetically engineered cells that give off a fluorescent signal, we were able to track the long-term survival of the EHMs (at least 7 months) without having to sacrifice many of the animals. Our analysis of the transplanted EHMs showed that they had survived transplantation and had taken on characteristics that made them closer to normal heart tissue. In addition, EHM transplantation resulted in improved heart function, as compared to rats that either received no transplants or received a control EHM transplant that contained dead cells.
  • The next phase of our project will be to evaluate the function of larger EHMs in swine model of heart disease, since these animals have hearts that are similar in size and heart rate to humans. This is a crucial step before considering translating this work into human patients.
Funding Type: 
Basic Biology V
Grant Number: 
RB5-07356
Investigator: 
ICOC Funds Committed: 
$1 124 834
Disease Focus: 
Heart Disease
Stem Cell Use: 
Embryonic Stem Cell
iPS Cell
oldStatus: 
Closed
Public Abstract: 

Cells in the body take up nutrients from their environment and metabolize them in a complex set of biochemical reactions to generate energy and replicate. Control of these processes is particularly important for heart cells, which need large amounts of energy to drive blood flow throughout the body. Not surprisingly, the nutritional requirements of heart cells are very different than those of stem cells. This proposal will investigate the metabolism of pluripotent stem cells and how this changes during differentiation to cardiac cells. We will determine which nutrients are important to make functional heart cells and use this information to optimize growth conditions for producing heart cells for regenerative medicine and basic biology applications. We accomplish this by feeding cells nutrients (sugar, fat) labeled with isotopes. As these labeled molecules are consumed, the isotopes are incorporated into different metabolites which we track using mass spectrometry. This advanced technique will allow us to see how sugars and fat are metabolized inside stem cells and cardiac cells obtained through differentiation. We will also study the electrical activity of these heart cells to ensure that adequate nutrients are provided for the generation of cells with optimal function. Ultimately, this project will lead to new methods for producing functional heart cells for regenerative medicine and may also lead to insights into how cardiac cells malfunction in heart disease.

Statement of Benefit to California: 

Heart disease is one of the leading causes of death in California. As a result, much of the regenerative medicine community in the state and the many Californians suffering from heart failure are interested in obtaining functional heart cells from stem cells. Our work will identify the most important nutrients required to coax stem cell-derived heart cells to behave like true adult heart cells. This information will make more effective cell models for researchers and companies to study how this disease affects heart cell metabolism. Since enzymes are highly targetable with drugs, the basic scientific findings from our work will be of great interest to California biotechnology companies and can stimulate job growth in the state. Our findings will also provide insight into very specific types of genetic heart disease, and this work may lead to additional grants from federal funding sources, bringing about additional revenue and job growth in California. A better understanding of how different nutrients influence heart cell function may provide guidance into new treatment strategies for heart disease. Finally, this work will highlight the importance of diet, nutrition, and healthy heart function, providing useful information relating to public health.

Progress Report: 
  • Growing up we have all heard the phrase, “You are what you eat.” Just like our bodies, stem cells require large quantities of fuel for energy and growth. The same is true for heart cells that continually drive blood flow. Therefore, understanding how stem cells and cardiac cells they generate consume and use different foods is important for characterizing their clinical potential. This grant aims to study how different nutritional fuels influence stem cell and cardiomyocyte (heart cell) growth, differentiation, and function. Using advanced methods that allow us to track how sugar (carbs), protein, and fat are consumed and produced by stem cells, we have identified key nutritional factors that affect stem cell performance and function. Surprisingly, most advanced stem cell media are lacking in several important factors. This deficiency negatively impacts stem cell metabolism in a number of ways, causing increased nutrient consumption, decreased respiration, and oxidative stress. Using this information we have developed improved stem cell growth conditions that mitigates these effects. Finally, we have compared the metabolism of stem cell-derived heart cells to parental stem cells, identifying key differences that will serve as benchmarks to functionally validate the performance and “maturity” of cardiac cells.
Funding Type: 
Basic Biology IV
Grant Number: 
RB4-06215
Investigator: 
Type: 
PI
ICOC Funds Committed: 
$1 367 604
Disease Focus: 
Heart Disease
Stem Cell Use: 
Adult Stem Cell
oldStatus: 
Active
Public Abstract: 

In the process of a heart attack, clots form suddenly on top of cholesterol-laden plaques, blocking blood flow to heart muscle. As a result, living heart tissue dies and is replaced by scar. The larger the scar, the higher the chance of premature death and disability following the heart attack. While conventional treatments aim to limit the initial injury (by promptly opening the clogged artery) and to prevent further damage (using various drugs), regenerative therapy for heart attacks seeks to regrow healthy heart muscle and to dissolve scar. To date, cell therapy with CDCs is the only intervention which has been shown to be clinically effective in regenerating the injured human heart. However, the cellular origin of the newly-formed heart muscle and the mechanisms underlying its generation remain unknown. The present grant seeks to understand those basic mechanisms in detail, relying upon state-of-the-art scientific methods and preclinical disease models. Our work to date suggests that much of the benefit is due to an indirect effect of transplanted CDCs to stimulate the proliferation of surrounding host heart cells. This represents a major, previously-unrecognized mechanism of cardiac regeneration in response to cell therapy. The proposed project will open up novel mechanistic insights which will hopefully enable us to boost the efficacy of stem cell-based treatments by bolstering the regeneration of injured heart muscle.

Statement of Benefit to California: 

Coronary artery disease is the predominant cause of premature death and disability in California. Clots form suddenly on top of cholesterol-laden plaques in the wall of a coronary artery, blocking blood flow to the heart muscle. This leads to a “heart attack”, in which living heart muscle dies and is replaced by scar. The larger the scar, the greater the chance of death and disability following the heart attack. While conventional treatments aim to limit the initial injury (by promptly opening the clogged artery) and to prevent further injury (using various drugs), regenerative therapy for heart attacks seeks to regrow healthy heart muscle and to dissolve scar. To date, cell therapy with CDCs is the only intervention that has been shown to be clinically effective in regenerating the injured human heart. However, the cellular origin of the newly-formed heart muscle and the mechanisms underlying its generation remain unknown. The present grant seeks to understand those basic mechanisms in detail, relying upon state-of-the-art scientific methods and preclinical disease models. The resulting insights will enable more rational development of novel therapeutic approaches, to the benefit of the public health of the citizens of California. Economic benefits may also accrue from licensing of new technology.

Progress Report: 
  • Key abbreviations:
  • CDCs: cardiosphere-derived cells
  • MI: myocardial infarction
  • The present award tests the hypothesis that CDCs promote regrowth of normal mammalian heart tissue through induction of adult cardiomyocyte cell cycle re-entry and proliferation (as occurs naturally in zebrafish and neonatal mice). Such a mechanism, if established, would challenge the dogma that terminally-differentiated adult cardiomyocytes cannot re-enter the cell cycle. We have employed an inducible cardiomyocyte-specific fate-mapping approach (to specifically mark resident myocytes and their progeny), coupled with novel methods of myocyte purification and rigorous quantification. We have also developed assays that enable us to exclude potential technical confounding factors. The use of bitransgenic mice is essential for our experimental design (as it enables fate mapping of resident myocytes in a mammalian model), while the use of mouse CDCs in our in vivo experiments (as opposed to human CDCs) enables us to avoid immunosuppression and its complications. To date, mouse, rat and pig models have proven to be reliable in predicting clinical effects of CDC therapy in humans, and results with human and mouse CDCs in comparable models (e.g., SCID mice for human CDCs versus wild-type mice for mouse CDCs) have not revealed any major mechanistic divergence. Our results demonstrate that induction of cardiomyocyte proliferation represents a major, previously-unrecognized mechanism of cardiac regeneration in response to cell therapy. One full-length publication describing these findings has appeared (K. Malliaras et al., EMBO Mol Med, 2013, 5:191-209), and another paper has been submitted. The work has already begun to open up novel mechanistic insights which will enable us to improve the efficacy of stem cell-based treatments and bolster cardiomyocyte repopulation of infarcted myocardium.
  • CDCs: cardiosphere-derived cells
  • MI: myocardial infarction
  • The present award tests the hypothesis that CDCs promote regrowth of normal mammalian heart tissue through induction of adult cardiomyocyte cell cycle re-entry and proliferation (as occurs naturally in zebrafish and neonatal mice). Such a mechanism, if established, would challenge the dogma that terminally-differentiated adult cardiomyocytes cannot reenter the cell cycle. We have employed an inducible cardiomyocyte-specific fate-mapping approach (to specifically mark resident myocytes and their progeny), coupled with novel methods of myocyte purification and rigorous quantification. We have also developed assays that enable us to exclude potential technical confounding factors. The use of bitransgenic mice is essential for our experimental design (as it enables fate mapping of resident myocytes in a mammalian model), while the use of mouse CDCs in our in vivo experiments (as opposed to human CDCs) enables us to avoid immunosuppression and its complications. To date, mouse, rat, and pig models have proven to be reliable in predicting clinical effects of CDC therapy in humans, and results with human and mouse CDCs in comparable models (e.g., SCID mice for human CDCs versus wild-type mice for mouse CDCs) have not revealed any major mechanistic divergence. Our results demonstrate that induction of cardiomyocyte proliferation represents a major, previously-unrecognized mechanism of cardiac regeneration in response to cell therapy. Two full-length publications describing these findings has appeared (Malliaras, K, et al., EMBO Mol Med. 2014, 6:760-777; Malliaras K, et al., EMBO Mol Med, 2013, 5:191-209). The work has already begun to open up novel mechanistic insights which will enable us to improve the efficacy of stem cell-based treatments and bolster cardiomyocyte repopulation of infarcted myocardium.
Funding Type: 
Basic Biology IV
Grant Number: 
RB4-06035
Investigator: 
Name: 
Institution: 
Type: 
PI
ICOC Funds Committed: 
$1 708 560
Disease Focus: 
Heart Disease
Stem Cell Use: 
Directly Reprogrammed Cell
Cell Line Generation: 
iPS Cell
oldStatus: 
Active
Public Abstract: 

Recently, we devised and reported a new regenerative medicine paradigm that entails temporal/transient overexpression of induced pluripotent stem cell based reprogramming factors in skin cells, leading to the rapid generation of “activated” cells, which can then be directed by specific growth factors and small molecules to “relax” back into various defined and homogenous tissue-specific precursor cell types (including nervous cells, heart cells, blood vessel cells, and pancreas and liver progenitor cells), which can be expanded and further differentiated into mature cells entirely distinct from fibroblasts.

In this proposal, combined with small molecules that can functionally replace reprogramming transcription factors as well as substantially improve reprogramming efficiency and kinetics, we aim to further develop and mechanistically characterize chemically defined, non-integrating approaches (e.g., mRNA, miRNA, episomal plasmids and/or small molecule-based) to robustly and efficiently reprogram skin fibroblast cells into expandable heart precursor cells. Specifically, we will: determine if we can use non-integrating methods to destabilize human fibroblasts and facilitate their direct reprogramming into the heart precursor cells; characterize of heart cells generated by the direct programming methods, both in the tissue culture dish and in a mouse model of heart attack; and characterize newly identified reprogramming enhancing small molecules mechanistically.

Statement of Benefit to California: 

This study will develop and mechanistically characterize a new method of generating safe patient specific heart cells that could be useful in treating heart failure which afflicts millions of Californians and accounts for billions of dollars in healthcare spending annually. Additionally, the small molecules discovered in this study could be good candidates for future drug development as well as being broadly useful for other regenerative medicine applications. These advances could also be a platform for new personalized medicine/ cell banking businesses which could bring economic growth in addition to improving the health of Californians.

Progress Report: 
  • During the reporting period, we have made very significant progress toward the following research aims: (1) Using the Oct4-based reprogramming assay system established, we were able to screen for and identify small molecules that can replace the other three genes in the Cell-Activation and Signaling-Directed (CASD) lineage conversion paradigm for reprogramming fibroblasts into cardiac lineage. (2) Using in-depth assays, we have examined the process using lineage-tracing methods and characterized those Oct4/small molecules-reprogrammed cardiac cells in vitro. (3) Most importantly, we were able to identify a baseline condition that appears to reprogram human fibroblasts into cardiac cells using defined conditions.
  • Cardiomyocyte-like cells can be reprogrammed from somatic fibroblasts by combinations of genes in vitro1 and in vivo, providing a new avenue for cardiac regenerative therapy. However, transdifferentiating human cells to generate fully functional cardiomyocytes remains a challenge. Moreover, genetic manipulations with multiple factors used in such conventional strategies pose safety, efficacy, and technical concerns that may limit their clinical potential. In the work funded by CIRM we identified and demonstrated that functional cardiomyocytes can be rapidly and efficiently generated from fibroblasts by a combination of small molecules. These cardiomyocytes express characteristic cardiac markers, have a well-organized sarcomeric structure, contract spontaneously, and respond to pharmacological modulations. They closely resemble cardiomyocytes in their global gene expression profiles, and electrophysiological properties. This novel pharmacological reprogramming approach may have important implications in cardiac regenerative medicine.
Funding Type: 
Basic Biology IV
Grant Number: 
RB4-05764
Investigator: 
Type: 
PI
ICOC Funds Committed: 
$1 334 880
Disease Focus: 
Heart Disease
Stem Cell Use: 
iPS Cell
oldStatus: 
Active
Public Abstract: 

Currently, over 350,000 patients per year with abnormal heart rhythm receive electronic pacemakers to restore their normal heart beat. Electronic pacemakers do not respond to the need for changing heart rate in situations such as exercise and have limited battery life, which can be resolved with biopacemakers. In this proposed project, we will examine methods that improve the generation of pacemaking cells from human induced pluripotent stem cells as candidates for biopacemaker.

Statement of Benefit to California: 

This proposal aims to generate pacemaking cells through facilitated differentiation from human induced pluripotent stem cells that may serve as biopacemakers. Over 350,000 patients a year in the U.S. require the implantation of an electronic pacemaker to restore their heart rhythm, with more than 3 million patients that are dependent on this device. At the cost of $58K per pacemaker implantation, the healthcare burden is greater than $20 billion a year. However, the cost associated with these electronic devices does not end with surgery for implantation. These devices require a battery change every 5 to 10 years that involve another surgical procedure. With California being the most populated state, this can be very costly to the Californians. It also does not give the patients the quality of life by having to endure repeated surgeries. The possibility of biopacemaker that requires no future battery replacements and other advantages such as physiological adaptation to the active state of the patient make biopacemakers a truly desirable alternative to electronic devices. Moreover, one in 20,000 infants or preemies with congenital sinoatrial node dysfunction are also inappropriate candidates to receive electronic pacemakers because they are physically too small and require a proportional increase in the length of pacing leads with their significant growth rate. Therefore, there is a great need for biopacemakers that may overcome the deficiencies of electronic devices.

Progress Report: 
  • This goal of this project is to improve the yield of pacemaking cells from human induced pluripotent stem cells (hiPSCs) that can be used to engineer biopacemaker. We have demonstrated that manipulation of the membrane potential of hiPSCs using small molecules can upregulate genes of the desired cell type progressing to the pacemaking cells at all differentiation stages. In the differentiation stage to mesodermal cells, treated hiPSCs exhibit a membrane potential that is further down the differentiation path than untreated control. This source was this change was examined.
  • We continued our work in improving the yield of pacemaking cells from human induced pluripotent stem cells (hiPSCs) that can be used to engineer biopacemakers. The ion channel isoform responsible for the induced membrane potential changes in hiPSCs and their differentiating cardiac progeny was determined. We focused on optimizing the duration and the timing of membrane potential manipulation in improving the efficiency of pro-pacemaking cardiac progenitor cells and pacemaking cells.

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