Stem cells, cancer, and cancer stem cells
Stem cell biology has come of age. Unequivocal proof that stem cells exist in the haematopoietic system has given way to the prospective isolation of several tissue-specific stem and progenitor cells, the initial delineation of their properties and expressed genetic programmes, and the beginnings of their utility in regenerative medicine. Perhaps the most important and useful property of stem cells is that of self-renewal. Through this property, striking parallels can be found between stem cells and cancer cells: tumours may often originate from the transformation of normal stem cells, similar signalling pathways may regulate self-renewal in stem cells and cancer cells, and cancer
cells may include ‘cancer stem cells’ — rare cells with indefinite potential for self-renewal that drive tumorigenesis.
Stem cells are defined as cells that have the ability to perpetuate themselves through selfrenewal and to generate mature cells of a particular tissue through differentiation. In most tissues, stem cells are rare. As a result, stem cells must be identified prospectively and purified carefully in order to study their properties. Although it seems reasonable to propose that each tissue arises from a tissue-specific stem cell, the rigorous identification and isolation of these somatic stem cells has been accomplished only in a few instances. For example, haematopoietic stem cells (HSCs) have been isolated from mice and humans, and have been shown to be responsible for the generation and regeneration of the blood-forming and immune (haematolymphoid) systems (Fig. 1). Stem cells from a variety of organs might have the potential to be used for therapy in the future, but HSCs — the vital elements in bone-marrow transplantation — have already been used extensively in therapeutic settings.
The recent discovery that bone marrow, as well as purified HSCs, can give rise to non-haematopoietic tissues suggests that these cells may have greater differentiation potential than was assumed previously. Definitive experiments are needed to determine whether the cells from the bone marrow that are capable of giving rise to different non-haematopoietic lineages are indeed HSCs or another population. If further studies support the idea of HSC plasticity, this will undoubtedly open new frontiers for understanding the developmental potential of HSCs, as well as expand their therapeutic application.
As the characteristics of HSCs, their differentiation potential and clinical applications have been covered in earlier reviews, here we discuss emerging evidence that stem cell biology could provide new insights into cancer biology. In particular, we focus on three aspects of the relationship between stem cells and tumour cells: first, the similarities in the mechanisms that regulate self-renewal of normal stem cells and cancer cells; second, the possibility that tumour cells might arise from normal stem cells; and third, the notion that tumours might contain ‘cancer stem cells’ — rare cells with indefinite proliferative potential that drive the formation and growth of tumours. Through much of this review we focus on the haematopoietic system because both normal stem cells and cancer cells from this tissue are well characterized. Moreover, cancers of the haematopoietic system (that is, leukaemias) provide the best evidence that normal stem cells are the targets of transforming mutations, and that cancer cell proliferation is driven by cancer stem cells.
Self-renewal of haematopoietic stem cells
One of the most important issues in stem cell biology is understanding the mechanisms that regulate self-renewal. Self-renewal is crucial to stem cell function, because it is required by many types of stem cells to persist for the lifetime of the animal. Moreover, whereas stem cells from different organs may vary in their developmental potential, all stem cells must self-renew and regulate the relative balance between self-renewal and differentiation. Understanding the regulation of normal stem cell self-renewal is also fundamental to understanding the regulation of cancer cell proliferation, because cancer can be considered to be a disease of unregulated self-renewal.
In the haematopoietic system, stem cells are heterogeneous with respect to their ability to self-renew. Multipotent progenitors constitute 0.05% of mouse bone-marrow cells, and can be divided into three different populations: longterm self-renewing HSCs, short-term self-renewing HSCs, and multipotent progenitors without detectable self-renewal potential. These populations form a lineage in which the long-term HSCs give rise to short-term HSCs, which in turn give rise to multipotent progenitors. As HSCs mature from the long-term self-renewing pool to multipotent progenitors, they progressively lose their potential to self-renew but become more mitotically active. Whereas long-term HSCs give rise to mature haematopoietic cells for the lifetime of the mouse, short-term HSCs and multipotent progenitors reconstitute lethally irradiated mice for less than eight weeks.
Although the phenotypic and functional properties of HSCs have been extensively characterized, the fundamental question of how self-renewal is regulated remains unanswered. In most cases, combinations of growth factors that can induce potent proliferation cannot prevent the differentiation of HSCs in long-term cultures. Although progress has been made in identifying culture conditions that maintain HSC activity in culture, it has proved exceedingly difficult to identify combinations of defined growth factors that cause a significant expansion in culture in the number of progenitors with transplantable HSC activity.
Pathways regulating stem cell self-renewal and oncogenesis
Because normal stem cells and cancer cells share the ability to selfrenew, it seems reasonable to propose that newly arising cancer cells appropriate the machinery for self-renewing cell division that is normally expressed in stem cells. Evidence shows that many pathways that are classically associated with cancer may also regulate normal stem cell development (Fig. 2). For example, the prevention of apoptosis by enforced expression of the oncogene bcl-2 results in increased numbers of HSCs in vivo, suggesting that cell death has a role in regulating the homeostasis of HSCs.
Other signalling pathways associated with oncogenesis, such as the Notch, Sonic hedgehog (Shh) and Wnt signalling pathways, may also regulate stem cell self-renewal. Notch activation in HSCs in culture using the ligand Jagged-1 have consistently increased the amount of primitive progenitor activity that can be observed in vitro and in vivo, suggesting that Notch activation promotes HSC self-renewal, or at least the maintenance of multipotentiality. Shh signalling has also been implicated in the regulation of self-renewal by the finding that populations highly enriched for human HSCs (CD34+Lin–CD38–) exhibit increased self-renewal in response to Shh stimulation in vitro, albeit in combination with other growth factors. The involvement of Notch and Shh in the self-renewal of HSCs is especially interesting in light of studies that implicate these pathways in the regulation of self-renewal of stem cells from other tissues as well.
One particularly interesting pathway that has also been shown to regulate both self-renewal and oncogenesis in different organs is the Wnt signalling pathway (Fig. 2). Wnt proteins are intercellular signalling molecules that regulate development in several organisms and contribute to cancer when dysregulated. The expression of Wnt proteins in the bone marrow suggests that they may influence HSCs as well. Using highly purified mouse bone-marrow HSCs, we have shown that overexpression of activated b-catenin (a downstream activator of the Wnt signalling pathway) in long-term cultures of HSCs expands the pool of transplantable HSCs determined by both phenotype (Thy1.1loLin–/loSca1+c-kit+) and function (ability to reconstitute the haematopoietic system in vivo). Moreover, ectopic expression of Axin, an inhibitor of Wnt signalling, leads to inhibition of HSC proliferation, increased death of HSCs in vitro, and reduced reconstitution in vivo (T.R. et al., submitted). In separate studies, soluble Wnt proteins from conditioned supernatants have also been shown to influence the proliferation of haematopoietic progenitors from mouse fetal liver and human bone marrow.
Studies of epidermal and gut progenitors suggest that the Wnt signalling pathway may contribute to the regulation of stem cell/progenitor cell self-renewal in other tissues. Cultured human keratinocytes with higher proliferative potential have increased levels of b-catenin compared with keratinocytes with lower proliferative capacity. Moreover, retroviral transduction of activated b-catenin results in increased epidermal stem cell self-renewal and decreased differentiation. In vivo data from transgenic mice suggest that activation of the Wnt signalling pathway in epidermal stem cells leads to epithelial cancers. Furthermore, mice lacking TCF-4, one of the transcriptional mediators of the Wnt signalling pathway, quickly exhaust the undifferentiated progenitors in the crypts of the gut epithelium during fetal development, suggesting that this pathway is required for the maintenance or self-renewal of gut epithelial stem cells.
Cumulatively, the above findings suggest that Wnt signalling may promote stem cell self-renewal in a variety of different epithelia in addition to HSCs. The molecular mechanisms by which Wnt signaling influences stem cells remain to be elucidated. It will also be important to determine whether the Wnt, Notch and Shh pathways interact to regulate stem and progenitor cell self-renewal.