Turning Pluriptoent Human Embryonic Stem Cells into a Large Supply of Plastic CNS Derivatives for Cell Therapy

San Diego Regenerative Medicine Institute and Xcelthera announce the publication of Dr. Parsons’ original research article supported by the National Institutes of Health, titled “An Engraftable Human Embryonic Stem Cell Neuronal Lineage-Specific Derivative Retains Embryonic Chromatin Plasticity for Scale-Up CNS Regeneration”, in Journal of Regenerative Medicine & Tissue Engineering.

Human stem cell transplantation represents a promising therapeutic approach closest to provide a cure to restore the lost nerve tissue and function for a wide range of devastating and untreatable neurological disorders. However, to date, lack of a clinically-suitable source of engraftable human stem/progenitor cells with adequate neurogenic potential has been the major setback in developing effective cell-based therapy as a treatment option for restoring the damaged or lost central nervous system (CNS) structure and circuitry. The traditional sources of engraftable human stem cells with neural potential for transplantation therapies have been multipotent human neural stem cells (hNSCs) isolated directly from the human fetal CNS. However, cell therapy based on CNS tissue-derived hNSCs has encountered supply restriction and difficulty to use in the clinical setting due to their limited expansion ability and declining plasticity with aging, potentially restricting the tissue-derived hNSC as an adequate source for graft material.

Alternatively, the pluripotent human embryonic stem cells (hESCs) proffer cures for a wide range of neurological disorders by supplying the diversity of human neuronal cell types in the developing CNS for regeneration and repair. We must bear in mind that the pluripotent hESC itself cannot be used for therapeutic applications. It has been recognized that pluripotent hESCs must be transformed into fate-restricted derivatives before use for cell therapy. However, realizing the therapeutic potential of hESC derivatives has been hindered by the current state of the art for generating functional cells through multi-lineage differentiation of pluripotent cells, which is uncontrollable, inefficient, instable, highly variable, difficult to reproduce and scale-up, and often causes phenotypic heterogeneity and instability, hence, a high risk of tumorigenicity following transplantation. Under protocols presently employed in the field, the prototypical neuroepithelial-like nestin-positive hNSCs, either isolated from CNS in vivo or derived from pluripotent cells in vitro via conventional multi-lineage differentiation, appear to exert their therapeutic effects primarily by their non-neuronal progenies through producing trophic and/or neuroprotective molecules to rescue endogenous dying host neurons, but not related to regeneration from the graft or host remyelination.

We recently reported that pluripotent hESCs maintained under a defined platform can be uniformly converted into a cardiac or neural lineage by small molecule induction. This technology breakthrough enables well-controlled generation of a large supply of neuronal lineage-specific derivatives across the spectrum of developmental stages direct from the pluripotent state of hESCs with small molecule induction. Having achieved uniformly conversion of pluripotent hESCs to a neuronal lineage, in this study, the expression and intracellular distribution patterns of a set of chromatin modifiers in the hESC neuronal lineage-specific derivative hESC-I hNuPs were examined and compared to the two prototypical neuroepithelial-like hNSCs either derived from hESCs in vitro or isolated directly from the human fetal CNS in vivo. These hESC-I hNuPs expressed high levels of active chromatin modifiers, including acetylated histone H3 and H4, HDAC1, Brg-1, and hSNF2H, retaining an embryonic acetylated globally active chromatin state. Consistent with this observation, several repressive chromatin remodeling factors regulating histone H3K9 methylation, including SIRT1, SUV39H1, and Brm, were inactive in hESC-I hNuPs. These Nurr1-positive hESC-I hNuPs, which did not express the canonical hNSC markers, yielded neurons efficiently (> 90%) and exclusively, as they did not differentiate into glial cells, such as astrocytes, and oligodendrocytes. Following engraftment in the brain, hESC-I hNuPs yielded well-dispersed and well-integrated human neurons at a high prevalence. No graft overgrowth, formation of teratomas or neoplasms, or appearance of non-neuronal cell types was observed following engraftment. Transplanted wild type mice developed hyper-active behavior, such as fast movement and fast spin, which also suggested that transplanted human neuronal cells had survived and integrated into the mouse brain to function and control mouse behavior. By contrast, the prototypical neuroepithelial-like nestin-positive hNSCs derived either from hESCs or CNS can spontaneously differentiate into a mixed population of cells containing undifferentiated hNSCs, neurons (10-30%), astrocytes, and oligodendrocytes in vitro and in vivo. These observations suggest that, unlike the prototypical neuroepithelial-like nestin-positive hNSCs, these in vitro neuroectoderm-derived Nurr1-positive hESC-I hNuPs are a more neuronal lineage-specific and plastic human stem cell derivative, providing an engraftable human embryonic neuronal progenitor in high purity and large supply with adequate neurogenic potential for scale-up CNS regeneration as stem cell therapy to be translated to patients in clinical trials.