Discovering Potent Molecules with Human ESCs to Treat Heart Disease

Discovering Potent Molecules with Human ESCs to Treat Heart Disease

Funding Type: 
SEED Grant
Grant Number: 
RS1-00169
Award Value: 
$688,274
Disease Focus: 
Heart Disease
Stem Cell Use: 
Embryonic Stem Cell
Status: 
Closed
Public Abstract: 
This work is directly relevant to human embryonic stem cell (hESC) research because it brings new ideas about novel compounds to affect cardiomyogenesis. The work addresses an urgent need to develop new agents to treat cardiovascular disease. We will develop potent and selective drug-like molecules as cardiomyocyte differentiation agents. Heart disease is the leading cause of mortality and decline in the quality of life in the developed world. The ability of hESCs to form cardiomyocytes has spawned hope that these cells may be used to replace damaged myocardium. Despite their ability to form cardiomyocytes, efficient and controlled cardiomyogenesis in ESC cultures has not been achieved due to the unavailability of differentiation agents and an incomplete understanding of the pathways that regulate cardiac development. Success has been achieved in developing a robust and dependable high-throughput assay to study the effects of small molecules on cardiomyocyte differentiation. Powerful cell-based assays were developed and provided readouts that led to high-content results because multiple signals were probed. The assay is capable of capturing fast or long-acting biology because of the time-course readouts. Cell-based assays are superior to molecular screens because the cell-based assay delivers active compounds or “hits” that are permeable and non-cytotoxic. Moreover, refined “hits” can be used as probes to reveal novel signaling pathways and proteins that control differentiation, in a process termed chemical biology. By taking advantage of knowledge of the current “hits” we will rapidly synthesize novel drug-like compounds in a low-risk approach to. The “hits” will be refined and improved through an efficient synthetic process we use in our lab called “Dynamic Medicinal Chemistry”. Even after miniaturization and automation, screening is still expensive. A key to improve the screening process is to use pharmacologically active, drug-like compounds to provide rich target-relevant information. Intelligently designing libraries for screening by incorporating drug-like features into “lead” library design will improve the attrition rate and lead to more pharmacologically relevant compounds for future studies. This proposal is directly responsive to the California Institute for Regenerative Medicine SEED Grant Program because it provides for developing and testing new agents of use in cardiomyoenesis of hESCs. Importantly, it brings new investigators and a collaborative approach to the stem cell field. The agents discovered and developed may hold great promise as the groundwork for future medications development for a new class of damaged myocardium replacement agents. The theoretical rationale for the work is the use of high-content screening coupled with drug-like new agent discovery approaches. The work will also be of use in the elucidation of key biochemical targets and novel signaling pathways important in hESC cardiomyogenesis.
Statement of Benefit to California: 
In 2002, in the State of California, approximately 697,000 adult Californians died from heart disease. The cost as measured by loss of lifelong earnings was more than $79 billion. Setting aside the pain and suffering, the economic impact of cardiovascular disease to the State of California is staggering. Despite recent advances in cardiovascular medications development, new approaches and novel drug-like compounds are urgently needed to treat cardiovascular disease in California and elsewhere. The poor prognosis for heart disease for Californians underscores the critical need to develop alternative therapeutic strategies. The demonstrated ability of human embryonic stem cells (hESCs) to form cardiomyocytes has spawned widespread hope that these cells may be used as a source to replace damaged myocardium in humans. Despite their ability to form cardiomyocytes, efficient and controlled cardiomyogenesis in hESC culture has not been achieved due to the unavailability of differentiation agents and also because of an incomplete understanding of the pathways that regulate cardiac cell development. Using a high-throughput whole cell assay with image analysis, we have identified four small molecules that promote cardiomyogenesis in human ESCs. This proposal is directly responsive to the California Institute for Regenerative Medicine SEED Grant Program because it provides for developing and testing new agents of use in cardiomyogenesis of hESCs. It also brings new investigators and new collaborative approaches to the field. The promising agents discovered already constitute an excellent starting point and further refinement and development of these compounds may hold great promise as the groundwork for future medications development for a new class of damaged myocardium differentiation agents. The theoretical rationale for the work is the use of high-content screening coupled with drug-like new agent discovery approaches. The work will be of use in the elucidation of key biochemical targets and novel signaling pathways important in hESC cardiomyogenesis. The compounds discovered in our whole hESC-based assays thus far are not potent enough to be developed as drug candidates. But these compounds hold great promise as agents that could be refined further into drug leads. If the leads become drugs, promise of a new class of medication to treat cardiovascular disease may become a reality. Such drugs would decrease cardiovascular disease and decrease health care costs in California. This will likely have a significant economic impact to the State of California. The proposed work represents essential translational research required for new drug development.
Progress Report: 

Year 1

The original goals of the proposal were to apply medicinal chemistry to generate more potent and drug-like analogs of small molecules that stimulate differentiation of cardiomyocytes from embryonic stem cell (ESC) and potentially other progenitor cell types found in adult human heart. During the grant period, we over-achieved each Aim and provided large numbers of drug-like small molecules for cardiomyocyte differentiation studies. In addition, other related information was gained that has considerably expanded our understanding related to developing regenerative medicines. 1. Synthetic Chemistry: From an initial screen of thousands of compounds, six 'hits' were identified. Almost 1300 compounds were synthesized as analogs of these “hits” with the goal of generating more effective novel compounds as possible therapeutics for heart disease. 2. Assay development and screening: Novel synthetic chemical analogs were studied in cell-based assays to evaluate potency of stimulating cardiac cell development relative to the starting 'hit' compounds. The biological data contributed to structure activity relationship (SAR) studies, and provided valuable information about parts of the molecules important for cardiomyocyte stem cell differentiation and for other important pharmaceutical properties. The iterative feedback from the biological testing helped to guide the next generation designs of new and ever more effective compounds. 3. Chemical optimization. Focused structure activity relationship (SAR) studies for 4 chemical series from the ESC cardiogenesis differentiation screen were done. SAR for 2 additional chemical classes was done but those agents proved less potent. In addition to SAR, considerable information was obtained leading to improved solubility and membrane permeability of compounds in development, which became a focus of the chemical optimizations. In summary, the work has already led to one or more promising drug-like compounds ready for efficacy testing in animal models and thus, efforts have greatly accelerated the timeline of getting compounds to human patients.

Year 2

The original goals of the proposal were to apply medicinal chemistry to generate more potent and drug-like analogs of small molecules that stimulate differentiation of cardiomyocytes from embryonic stem cells (ESCs) and potentially other progenitor cell types found in adult human heart. During the grant period, we over-achieved each Aim and provided large numbers of drug-like small molecules for cardiomyocyte differentiation studies. In addition, other related information was gained that has considerably expanded our understanding related to developing regenerative medicines. 1. Synthetic Chemistry: From an initial screen of thousands of compounds, six ‘hits’ were identified. Almost 1400 compounds were synthesized as analogs of these “hits” with the goal of generating more effective novel compounds as possible therapeutics for heart disease. 2. Assay development and screening: Novel synthetic chemical analogs were studied in cell-based assays to evaluate potency of stimulating cardiac cell development relative to the starting ‘hit’ compounds. The biological data contributed to structure activity relationship (SAR) studies, and provided valuable information about parts of the molecules important for cardiomyocyte stem cell differentiation and for other important pharmaceutical properties. The iterative feedback from the biological testing helped to guide the next generation design of new and ever more effective compounds. 3. Chemical optimization. Focused structure activity relationship (SAR) studies for 4 chemical series from the ESC cardiogenesis differentiation screen were done. SAR for 2 additional chemical classes was done but those agents proved less potent. In addition to SAR, considerable information was obtained leading to improved solubility and membrane permeability of compounds in development, which became a focus of the chemical optimizations. The most potent compounds increased stem cell differentiation to cardiomyocytes 5-10 fold. The compounds were non-toxic, reasonably tractable to make, stable and were water-soluble and hence relatively easy to handle. 4. A number of biological signaling pathways were identified as affiliated with cardiomyocyte differentiation. One such pathway also is involved in anti-cancer activities. Thus, our efforts in identifying cardiomyocyte differentiation agents led us to study novel biology associated with cancer. One “hit” of this signaling pathway was chosen to do synthetic chemistry and “hit” to lead refinement. Approximately 100 compounds were synthesized and tested for inhibition of this signaling pathway. In summary, the work has already led to a number of promising drug-like compounds ready for efficacy testing in animal models and thus, efforts have greatly accelerated the timeline of getting compounds to human patients. A total of 1500 compounds were synthesized to optimize the potency and properties of cardiomyocyte differentiation agents. The most potent stimulated production of human cardiomyocytes 5-10-fold compared to vehicle-stimulated cells.

© 2013 California Institute for Regenerative Medicine