Human pluripotent stem cells (hPSC) have the capacity to differentiate into every cell in the adult body, and they are thus a highly promising source of differentiated cells for the investigation and treatment of numerous human diseases. For example, neurodegenerative disorders are an increasing healthcare problem that affect the lives of millions of Americans, and Parkinson's Disease (PD) in particular exacts enormous personal and economic tolls. Expanding hPSCs and directing their differentiation into dopaminergic neurons, the cell type predominantly lost in PD, promises to yield cells that can be used in cell replacement therapies. However, developing technologies to create the enormous numbers of safe and healthy dopaminergic neurons required for clinical development and implementation represents a bottleneck in the field, because the current systems for expanding and differentiating hPSCs face numerous challenges including difficulty in scaling up cell production, concerns with the safety of some materials used in the current cell culture systems, and limited reproducibility of such systems.
An emerging principle in stem cell engineering is that basic advances in stem cell biology can be translated towards the creation of “synthetic stem cell niches” that emulate the properties of natural microenvironments and tissues. We have made considerable progress in engineering bioactive materials to support hESC expansion and dopaminergic differentiation. For example, basic knowledge of how hESCs interact with the matrix that surrounds them has led to progress in synthetic, biomimetic hydrogels that have biochemical and mechanical properties to support hESC expansion. Furthermore, biology often presents biochemical signals that are patterned or structured at the nanometer scale, and our application of materials chemistry has yielded synthetic materials that imitate the nanostructured properties of endogenous ligands and thereby promise to enhance the potency of growth factors and morphogens for cell differentiation.
We propose to build upon this progress to create general platforms for hPSC expansion and differentiation through two specific aims: 1) To determine whether a fully defined, three dimensional (3D) synthetic matrix for expanding immature hPSCs can rapidly and scaleably generate large cell numbers for subsequent differentiation into potentially any cell , and 2) To investigate whether a 3D, synthetic matrix can support differentiation into healthy, implantable human DA neurons in high quantities and yields. This blend of stem cell biology, neurobiology, materials science, and bioengineering to create “synthetic stem cell niche” technologies with broad applicability therefore addresses critical challenges in regenerative medicine.
This proposal will develop novel tools and capabilities that will strongly enhance the scientific, technological, and economic development of stem cell therapeutics in California. The most important net benefit will be for the treatment of human diseases.
Efficiently expanding immature hPSCs in a scaleable, safe, and economical manner is a greatly enabling capability that would impact many downstream medical applications. The development of platforms for scaleable and safe cell differentiation will benefit therapeutic efforts for Parkinson’s Disease. Furthermore, the technologies developed in this proposal are designed to be tunable, such that they can be readily adapted to numerous downstream applications.
The resulting technologies have strong potential to benefit human health. Furthermore, this proposal directly addresses several research targets of this RFA – the development and validation of stem cell scale-up technologies including
novel cell expansion methods and bioreactors for both human pluripotent cells and differentiated cell types – indicating that CIRM believes that the proposed capabilities are a priority for California’s stem cell effort. While the potential applications of the proposed technology are broad, we will apply it to a specific and urgent biomedical problem: developing systems for generating clinically relevant quantities of dopaminergic neurons from hPSCs, part of a critical path towards developing therapies for Parkinson’s disease. This proposal would therefore work towards developing capabilities that are critical for hPSC-based regenerative medicine applications in the nervous system to clinically succeed.
The principal investigator and co-investigator have a strong record of translating basic science and engineering into practice through interactions with industry, particularly within California. Finally, this collaborative project will focus diverse research groups with many students on an important interdisciplinary project at the interface of science and engineering, thereby training future employees and contributing to the technological and economic development of California.
This proposal focuses on the development of a synthetic three dimensional (3D) extracellular matrix (ECM) to support human pluripotent stem cell (hPSC) expansion and differentiation into functional dopaminergic (DA) neurons for the treatment of Parkinson’s disease (PD). Current DA differentiation methods face many challenges, including limited efficiency, reproducibility and scalability, as well as the use of animal-derived products. The applicant proposes to address these translational bottlenecks in two Specific Aims: (1) to engineer and optimize a fully defined, synthetic, 3D ECM that supports the rapid and scalable expansion of hPSCs; and (2) to engineer and optimize a second, transplantable ECM that supports the differentiation of hPSCs into healthy DA neurons.
The reviewers agreed that this proposal addresses a significant bottleneck in the clinical translation of cell therapies. Traditional sources of neurons to treat PD provide limited cell numbers and while hPSCs are a promising alternative, current differentiation methods fall short in a number of areas. The applicant proposes a series of creative and innovative approaches to address these shortcomings and develop defined, scalable methods for generating large numbers of implantable DA neurons. Reviewers were optimistic that this proposal could have a major impact on the treatment of not only PD but other neurological diseases.
Reviewers described the research plan as focused, thorough and carefully designed. They appreciated the strong preliminary data, which demonstrates the development of peptides to support hPSC proliferation, neuronal and glial differentiation and to enhance the efficiency of DA neuron differentiation. Reviewers noted that while endless combinations of ECM modifications could be tested, the applicant has carefully narrowed down the variables. They also appreciated the applicant will compare human embryonic stem cells with induced pluripotent stem cells. Reviewers noted that potential pitfalls and alternative approaches are discussed and the project timeline is well defined. The reviewers had one significant criticism of the research plan, which was that insufficient attention is paid to developing methods which optimize both the percentage of DA neurons generated and their subtype. Reviewers cautioned that the generation of grafts containing the appropriate DA neuron subtype, uncontaminated by non-DA neurons, is as important as generating large numbers of cells. Clinical trials have suggested that an uncharacterized population of mesencephalic neurons may release too much dopamine, resulting in debilitating dyskinesias. Reviewers recommended that the applicant incorporate analysis of neuronal types and subtypes following differentiation.
The reviewers described the Principal Investigator (PI) as an expert in stem cell biology and tissue engineering with an impressive list of publications related to this proposal. They appreciated that the PI has worked and published successfully with the Co-Investigator for many years and generally found the research team well qualified. While reviewers agreed that the out-of-state consulting investigator appears to have the appropriate in vivo cell transplant expertise, they suggested it should be possible to find a qualified collaborator within California.
Overall, the reviewers were enthusiastic about this innovative proposal to develop novel technologies for hPSC expansion and differentiation into DA neurons. They agreed that it addresses a significant translational bottleneck and could have a major impact on PD as well as the whole field of regenerative medicine.