Mini Review: Opportunities and Challenges for Use of Human Pluripotent Stem Cells in Hematopoietic Developmental Biology and Regenerative Medicine

By Thorsten M. Schlaeger, PhD, and Xiao Guan, PhD

2009-09-01

Dr. Schlaeger is Instructor at the Stem Cell Program at Children’s Hospital Boston and a member of the Harvard Stem Cell Institute.

Dr. Guan is a Postdoctoral Fellow in the Stem Cell Program at Children’s Hospital Boston.

Mouse Pluripotent Stem Cells

Human ESCs may be obtained from embryos produced by IVF, parthenogenesis, or, at least in theory, nuclear transfer,1 whereas iPSCs can be achieved by direct reprogramming of somatic cells, such as skin fibroblasts.2 Independent of their origin, these PSCs have the potential to give rise to all somatic cell types, including those of the hematopoietic compartment,3 thus allowing human tissue ontogeny to be studied in vitro. PSC-derived somatic cells may be used to model diseases such as leukemia, 4 to screen drugs,5 or as therapeutic agents in cell-based therapies.6
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The defining features of pluripotent stem cells (PSCs) are their unlimited proliferative capacity and their ability to differentiate into all somatic cell types as well as germ cells. Mouse embryonic stem cells (ESCs), the first known non-malignant PSC type, are derived from the early epiblast. They were originally isolated in 19811 and have since been used with great success as in vitro models of mammalian development. Highlights in hematopoiesis include their differentiation into hemangioblasts,2 erythroid cells,3 lympho-hematopoietic progenitors,4 and hematopoietic stem cells (HSCs).5 Mouse ESCs have also been key to numerous in vivo analyses of gene function and to proof-of-principle studies that demonstrate the feasibility of PSC-based therapies for degenerative disorders, such as immunodeficiency6 and sickle cell anemia.7 However, it is important to note that several aspects of human biology and pathology of particular relevance to hematology are only poorly replicated in mouse models. Examples include developmental programs of globin gene switching and pathologies such as Fanconi anemia (FA), Trisomy-21-associated M7 acute myelogenous leukemia, and poorly defined genetic syndromes (e.g., thrombocytopenia with absent radii). Furthermore, mouse models often fail to mirror responses to pharmacologic agents such as TNFα,IFNγ, EPO, and camptothecins.

Human PSCs

Pluripotent human ESCs, derived from the late epiblast, were first isolated in 1998.8 More recently, another breakthrough was achieved: the direct reprogramming of differentiated human somatic cells into induced-PSCs (iPSCs) by forced expression of pluripotency genes.9-11 Generation of human iPSCs is significant since it provides a straightforward source for normal as well as patient/disease-specific PSCs that are remarkably similar to embryo-derived PSCs while avoiding the ethically charged use of human embryos or eggs. Reprogramming is also a fascinating process per se, the study of which will undoubtedly continue to further our understanding of epigenetics, stem cell biology, cell fate regulation, cancer, and aging.

Using Human PSCs to Study Hematopoietic Ontogeny

Informed by prior mouse studies, several groups were able to develop protocols for directed differentiation of human PSCs into hemangioblasts,12 multi-lineage progenitors,13  lymphoid cells,14 and even HSC-like cells.15 However, several challenges remain. For example, in vitro differentiation tends to replicate normal ontogeny, thus favoring development of early embryonic or fetal cells over more mature lineages. In particular, generation of adult-type erythroid cells or bona fide HSCs, so far, has met with little success. Furthermore, many studies make liberal use of undefined reagents, such as animal sera or support cells, which may obscure a detailed mechanistic understanding of the patterns of cellular development and hinder clinical use of these protocols. Furthermore, we are only just beginning to understand why individual stem cell lines can differ dramatically in their intrinsic potential to form particular lineages, a fact that limits our ability to generalize findings obtained with any small set of lines.

Using Human PSCs to Study Hematopoietic Disorders and Malignancies

One group has already produced diseasecorrected hematopoietic progenitors from FA patients16 using iPSC technology. In addition, human ESCs or iPSCs from Trisomy-21-, SCID-, SBDS-, and thalassemia embryos or patients have been generated and await further analysis.17, 18 Human PSC technology is particularly powerful, as it allows for the modeling of diseases without specific knowledge of the underlying gene mutation(s), or of diseases that result from tissue-restricted somatic mutations (e.g., leukemia19). (It will be interesting to see if multi-step progression of proliferative disorders can be reconstructed in vitro and whether drug screening in such models will lead to improved treatment modalities.) Furthermore, PSC technology can provide genetically diverse sets of human cells for drug toxicity testing.

Therapeutic Use of Human PSCs

Much of the excitement around human PSCs comes from the credo that patient-specific cells can be generated, undergo gene repair (if necessary), be induced to differentiate into therapeutically valuable cell types, and subsequently be re-introduced into patients to replace or ameliorate the affected cells or tissue. PSC-derived HSCs could benefit those in need of a bone marrow transplant, since finding a suitable marrow donor remains a major obstacle. In contrast to conventional bone marrow transplants, HSCs from a single donor could be provided to several recipients and could be prepared devoid of lymphocytes to avoid graft-versus-host concerns. Additional clinical benefits of such HSCs include induction of tolerance to solid tissue transplants of the same donor type and graft versus autoimmunity.20 While human PSC-derived, clinical-grade blood cells are not yet a reality, Geron Corp. is forging ahead with an experimental therapy for spinal cord injury that uses allogeneic human ESC-derived oligodendrocyte progenitor cells. A major focus of this pioneering trial will be on safety since residual undifferentiated PSCs may contaminate any preparation of PSC-derived therapeutic cells and can give rise to teratomas.8 Stringent purification of the differentiated cells is therefore of paramount importance. For added safety, the cells can be encapsulated or irradiated prior to transplantation, but this will often significantly limit the effectiveness and durability of the therapy. Human PSC-derived erythroid cells constitute a cell type that can be irradiated without detrimental effects on function. The therapeutic effects of irradiated erythroid cells will necessarily be transient, as is the case with conventional blood transfusion. However, the combination of a clinical need, the absence of HLA restriction, and the possibility of avoiding a critical safety concern through irradiation make it likely that PSC-derived erythrocytes will become one of the first clinical success stories. Indeed, large-scale production is feasible.21 and a consortium headed by the Scottish National Blood Transfusion Service will attempt to produce clinical-grade universal donor erythrocytes.22­

The Road Ahead

The development of human PSC-based therapies faces several challenges, such as funding restrictions on ESC derivation and use, an uncertain intellectual property landscape, and a regulatory framework that was not developed with such therapies in mind. Scientists also struggle to find safe and efficient methods for patient-specific ESC or iPSC production as well as for subsequent large-scale differentiation into transplantable cells. Other issues include the question of whether human iPSCs are indeed equivalent to conventional ESCs (Emerging evidence suggests that substantial differences exist and much more needs to be learned.), and the lack of long-term clinical data. Nevertheless, while the obstacles undoubtedly may be daunting in the aggregate, each individual challenge can likely be overcome. After all, it took several decades from the first development of monoclonal antibodies to their broad success in clinical use. Likewise, if given sufficient time and support, human PSC technology promises to revolutionize regenerative medicine.

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