Targeting Notch, Hedgehog, and Wnt pathways in cancer stem cells: clinical update Cancer stem cells (CSCs) have been increasingly identified in many malignancies. Although the origin and plasticity of these cells remain controversial, tumour heterogeneity and the presence of small populations of cells with stem-like characteristics is established in most malignancies. CSCs display many features of embryonic or tissue stem cells, and typically demonstrate persistent activation of one or more highly conserved signal transduction pathways involved in development and tissue homeostasis, including the Notch, Hedgehog (HH), and Wnt pathways. CSCs generally have slow growth rates and are resistant to chemotherapy and/or radiotherapy. Thus, new treatment strategies targeting these pathways to control stem-cell replication, survival and differentiation are under development. Herein, we provide an update on the latest advances in the clinical development of such approaches, and discuss strategies for overcoming CSC-associated primary or acquired resistance to cancer treatment. Given the crosstalk between the different embryonic developmental signalling pathways, as well as other pathways, designing clinical trials that target CSCs with rational combinations of agents to inhibit possible compensatory escape mechanisms could be of particular importance. We also share our views on the future directions for targeting CSCs to advance the clinical development of these classes of agents. The theory that malignancies arise from a small subset of stem-cell-like cancer cells has received increasing attention during the past decade. These cells, referred to as cancer stem cells (CSCs) or cancer-initiating cells (CICs), have been identified in many malignancies and are hypothesized to form the clonogenic core of tumour tissues. The origin of CSCs in human tumours is, however, not fully understood. Such cells could potentially originate from a more-differentiated cancer cell that acquires self-renewal properties, perhaps as a result of epithelial-to-mesenchymal transition (EMT). Alternatively, CSCs might derive from a normal tissue stem cell that undergoes transformation as a result of oncogenic somatic mutations, under the influence of extrinsic microenvironmental factors. Although the co-occurrence of subpopulations of cancer cells with different tumorigenic properties within individual tumours is no longer in question, the CSC hypothesis remains controversial. This controversy arises as a consequence of the technical and logistical challenges in isolating and identifying CSCs from human solid tumours that contain heterogeneous cell populations, and the limited number of validated surrogate assays currently available to substantively confirm stem-cell-like properties. These cells tend to comprise a small fraction of total tumour mass and are, therefore, difficult to unequivocally identify histologically. Moreover, tumour dissociation from normal tissues and subsequent flow cytometric analysis of tumour cells is not always possible with human biospecimens. Furthermore, markers that identify CSCs vary across different tumour types, and no clear-cut and clinically validated assay isTargeting Notch, Hedgehog, and Wnt pathways in cancer stem cells: clinical update Cancer stem cells (CSCs) have been increasingly identified in many malignancies. Although the origin and plasticity of these cells remain controversial, tumour heterogeneity and the presence of small populations of cells with stem-like characteristics is established in most malignancies. CSCs display many features of embryonic or tissue stem cells, and typically demonstrate persistent activation of one or more highly conserved signal transduction pathways involved in development and tissue homeostasis, including the Notch, Hedgehog (HH), and Wnt pathways. CSCs generally have slow growth rates and are resistant to chemotherapy and/or radiotherapy. Thus, new treatment strategies targeting these pathways to control stem-cell replication, survival and differentiation are under development. Herein, we provide an update on the latest advances in the clinical development of such approaches, and discuss strategies for overcoming CSC-associated primary or acquired resistance to cancer treatment. Given the crosstalk between the different embryonic developmental signalling pathways, as well as other pathways, designing clinical trials that target CSCs with rational combinations of agents to inhibit possible compensatory escape mechanisms could be of particular importance. We also share our views on the future directions for targeting CSCs to advance the clinical development of these classes of agents. The theory that malignancies arise from a small subset of stem-cell-like cancer cells has received increasing attention during the past decade. These cells, referred to as cancer stem cells (CSCs) or cancer-initiating cells (CICs), have been identified in many malignancies and are hypothesized to form the clonogenic core of tumour tissues. The origin of CSCs in human tumours is, however, not fully understood. Such cells could potentially originate from a more-differentiated cancer cell that acquires self-renewal properties, perhaps as a result of epithelial-to-mesenchymal transition (EMT). Alternatively, CSCs might derive from a normal tissue stem cell that undergoes transformation as a result of oncogenic somatic mutations, under the influence of extrinsic microenvironmental factors. Although the co-occurrence of subpopulations of cancer cells with different tumorigenic properties within individual tumours is no longer in question, the CSC hypothesis remains controversial. This controversy arises as a consequence of the technical and logistical challenges in isolating and identifying CSCs from human solid tumours that contain heterogeneous cell populations, and the limited number of validated surrogate assays currently available to substantively confirm stem-cell-like properties. These cells tend to comprise a small fraction of total tumour mass and are, therefore, difficult to unequivocally identify histologically. Moreover, tumour dissociation from normal tissues and subsequent flow cytometric analysis of tumour cells is not always possible with human biospecimens. Furthermore, markers that identify CSCs vary across different tumour types, and no clear-cut and clinically validated assay is