Cancer stem cells (CSCs) are a small population of cells within a tumor that are self-sustaining and cause heterogeneous lineages of cancer cells. There are three distinct characteristics of CSCs. The first of these three characteristic is tumor initiation, which is the ability to regenerate the tumor from a limited number of cells. Secondly, self-renewal demonstrates the ability of stem cells to increase the size of the stem cell pool while maintaining its undifferentiated state. The third of these characteristics is the ability to give rise to heterogeneous progeny that is a phenotype of the primary tumor. The degree to which cancer stem cells differentiate into certain cell types and their ability to self-renew differs based on intrinsic properties of each cell individually.
The cancer stem cell hypothesis postulates that many cancers are hierarchically organized and a subpopulation of cells within the tumor possess the basic properties of stem cells, the ability to self-renew and differentiate. CSCs also have other stem cell-like properties including the active expression of telomerase, anti-apoptotic pathway activation, increased activity of membrane transporters, and an increased ability to migrate. Evidence suggests that CSCs may be responsible for tumor progression, metastasis, chemotherapy and radiotherapy resistance, and subsequent tumor recurrence. Additionally, epithelial-mesenchymal transition activation has been shown to generate CSCs and is associated with increased motility and acquisition of invasive and metastatic properties.
Moreover, glioma stem cells are resistant to ionizing radiation due to enhanced DNA damage repair. Hepatic cancer cells are resistant to the chemotherapeutic agents doxorubicin, 5-fluorouracil, and gemcitabine due to upregulation of ABCG2 and ABCB1 encoding efflux pumps. Additionally, paired breast cancer cores pre- and post- neoadjuvant chemotherapy (docetaxel or doxorubicin and cyclophosphamide) had an increased CSC population following chemotherapy, showing CSCs are intrinsically resistance to conventional chemotherapy. These resistant tumorigenic cells may drive tumor recurrence by regenerating the bulk of the tumor following therapy. This supports the need and extensive ongoing effort to find agents that target the CSC population.
Cancer stem cells were first identified in seminal work from Dr. John Dick’s laboratory studying Acute Myeloid Leukemia (AML). Using methods developed to isolate and characterize normal hematopoietic stem cells, the authors found only a subset of cells from AML patients were able to recapitulate the disease in severe combined immunodeficient (SCID) recipient mice. These leukemia stem cells were characterized as CD34+/CD38- and had extensive proliferative capacity, self-renewal potential, and were able to differentiate into leukemic blasts in vivo. Lineage tracing has shown a single leukemia stem cell can give rise to various populations of leukemic cells due to heterogeneity in self-renewal potential and these functional differences reflect a hierarchical arrangement of leukemia cells similar to the normal hematopoietic compartment.
The first evidence of CSCs in solid tumors came from work in breast cancer. Al-Hajj and colleagues identified breast CSCs from patient tumors using surface marker expression and tumor regeneration potential upon mammary fat pad implantation of these cells in nonobese diabetic/severe combined immunodeficient (NOD/SCID) mice. As few as 200 breast CSCs, defined by the phenotype CD44+/CD24-, were able to consistently form tumors upon transplantation, whereas 100 fold as many (20,000) CD44+/CD24+ cells did not form any tumors. Moreover, secondary and tertiary transplantation of the breast CSCs remained capable of generating the phenotypic heterogeneity of the initial tumor, demonstrating the selfrenewal capacity of this population as well as the ability to differentiate into bulk tumor cells. Ginestier and collaborators identified another marker for breast CSCs- cells with high levels of aldehyde dehydrogenase 1 (ALDH1) enzymatic activity. ALDH1 is a detoxifying enzyme responsible for intracellular aldehyde oxidation and may play a role in differentiation of stem cells through oxidation of retinol into retinoic acid. As few as 500 ALDH+ cells were able to generate tumors in the mammary fat pad of NOD/SCID mice and displayed CSC properties, notably the ability to self-renew, differentiate into ALDH- cells, and recapitulate the heterogeneity of the parental tumor. Subsequently, CSCs have been identified and isolated in a number of other malignancies, including brain cancers, melanoma, ovarian cancer, prostate cancer, bone sarcomas, colon cancer, multiple myeloma, pancreatic cancer, and head and neck cancers.
Cancer stem cell biomarker
The expression of cell surface CSC markers is heterogeneous. High variability in the CSC phenotype has been observed between patients with some tumors having few CSCs and in others CSCs are a substantial proportion of the tumor mass. Moreover, tumors may harbor multiple genetically or phenotypically distinct CSCs. The oncogenic events and mutations that contribute to transformation are thought to influence the CSC phenotype. This may explain why one tumor or tumor-derived cell line displays a certain CSC population (for example, CD44+/CD24- in MDA-MB 231 breast cancer cells) and another has a different CSC population (ALDH+ in SKBR3 breast cancer cells). Thus, a variety of markers must be used and functional assays tested when studying CSCs.
Multiple studies suggest Human Epidermal Growth Factor Receptor 2 (HER2) plays an important role in regulating the CSC population in HER2+ breast cancer. HER2 overexpression and ALDH expression are significantly correlated in human breast cancer patient samples. The CSC population in HER2 overexpressing breast cancer cell lines expresses the highest levels of HER2 protein without HER2 gene amplification changes. Additionally, HER2 overexpression expands the normal breast epithelial stem/early progenitor cell population, as well as the CSC population in malignant breast cells, resulting in increased tumorigenicity and invasiveness with HER2 amplification. HER2 blockade via trastuzumab or HER2/EGFR blockade via lapatinib decreases the CSC population. Neoadjuvant trastuzumab significantly increases pathologic complete response rate compared to chemotherapy alone, suggesting a reduction in the CSC population. Moreover, in contrast to chemotherapy, lapatinib reduced the CSC population in the neoadjuvant setting, although this decrease was not statistically significant.
Telomerase is expressed in both bulk cancer cells and CSCs, suggesting CSCs could be sensitive to telomerase inhibition therapy. Imetelstat has been shown to target the CSC population in a number of tumor types including glioblastoma, neuroblastoma, prostate, breast, and pancreatic cancer. The effect of telomerase inhibition on invasion and metastases was not addressed nor the effect of imetelstat in combination with standard therapies on the CSC population. Telomerase inhibitors are most effective when used in combination with standard therapies, likely due to the long lag time to achieve telomere shortening.
Another biomarker is ALDH1, a detoxifying enzyme that catalyzes the oxidation of aldehyde groups into carboxylic acids. Its ability to metabolize retinol into retinoic acid is under investigation for its role in stem cell differentiation. Found predominantly in liver cells due to its metabolic significance, overexpression has been associated with negative clinical prognosis for malignant human mammary stem cells. ALDH1 expression on the exterior of the plasma membrane has been demonstrated in both CSCs and progenitor cells, while internal ALDH expression is strictly associated with the undifferentiated stem cells16. Antibody staining techniques may be utilized to identify ALDH expression on cell surfaces, while the ALDEFLUOR assay has been used to identify cells with cytosolic ALDH expression.
EpCAM is a transmembrane glycoprotein found frequently on the basolateral surfaces of various epithelial cells, enabling cell-cell adhesion through calcium-independent pathways. Increased EpCAM activity correlates inversely with standard cadherin-mediated adhesion, which has the effect of deregulating epithelial cell growth and differentiation and increasing epithelial cell proliferation. EpCAM is being investigated as a potential target for cancer therapy due to its documented overexpression in a variety of carcinomas, including those originating in the pancreas, breast, prostate, and colon.
DLL1 is a delta ligand homolog in humans that participates in multiple Notch signaling pathways: Ligands are passed from a signaling cell to the Notch cell surface domain, which initiates a series of cleavage events that in turn release transcription factors into the intracellular space. It is perhaps best known for its role in directing progenitor cell differentiation, promoting characteristics of T-cell precursors while blocking progression into B-cells. Additionally, Notch signaling via DLL1 regulates stem cell renewal and differentiation in the lumen of normal breast tissue, a property that has been implicated in the initiation and progression of cancer cells.
Cancer stem cells reference