Van T. Hoang

Immortality and the base of multicellular life: Lessons from cnidarian stem cells

Cnidarians are phylogenetically basal members of the animal kingdom (>600 million years old). Together with plants they share some remarkable features that cannot be found in higher animals. Cnidarians and plants exhibit an almost unlimited regeneration capacity and immortality. Immortality can be ascribed to the asexual mode of reproduction that requires cells with an unlimited self-renewal capacity. We propose that the basic properties of animal stem cells are tightly linked to this archaic mode of reproduction. The cnidarian stem cells can give rise to a number of differentiated cell types including neuronal and germ cells. The genomes of Hydra and Nematostella, representatives of two major cnidarian classes indicate a surprising complexity of both genomes, which is in the range of vertebrates. Recent work indicates that highly conserved signalling pathways control Hydra stem cell differentiation. Furthermore, the availability of genomic resources and novel technologies provide approaches to analyse these cells in vivo. Studies of stem cells in cnidarians will therefore open important insights into the basic mechanisms of stem cell biology. Their critical phylogenetic position at the base of the metazoan branch in the tree of life makes them an important link in unravelling the common mechanisms of stem cell biology between animals and plants.

The rarity of ALDH+ cells is the key to separation of normal versus leukemia stem cells by ALDH activity in AML patients

To understand the precise disease driving mechanisms in acute myeloid leukemia (AML), comparison of patient matched hematopoietic stem cells (HSC) and leukemia stem cells (LSC) is essential. In this analysis, we have examined the value of aldehyde dehydrogenase (ALDH) activity in combination with CD34 expression for the separation of HSC from LSC in 104 patients with de novo AML. The majority of AML patients (80 out of 104) had low percentages of cells with high ALDH activity (ALDH+ cells; <1.9%; ALDH‐rare AML), whereas 24 patients had relatively numerous ALDH+ cells (≥1.9%; ALDH‐numerous AML). In patients with ALDH‐rare AML, normal HSC could be separated by their CD34+ALDH+ phenotype, whereas LSC were exclusively detected among CD34+ALDH− cells. For patients with ALDH‐numerous AML, the CD34+ALDH+ subset consisted mainly of LSC and separation from HSC was not feasible. Functional analyses further showed that ALDH+ cells from ALDH‐numerous AML were quiescent, refractory to ARA‐C treatment and capable of leukemic engraftment in a xenogenic mouse transplantation model. Clinically, resistance to chemotherapy and poor long‐term outcome were also characteristic for patients with ALDH‐numerous AML providing an additional risk‐stratification tool. The difference in spectrum and relevance of ALDH activity in the putative LSC populations demonstrates, in addition to phenotypic and genetic, also functional heterogeneity of leukemic cells and suggests divergent roles for ALDH activity in normal HSC versus LSC. By acknowledging these differences our study provides a new and useful tool for prospective identification of AML cases in which separation of HSC from LSC is possible.

Identifying leukemia stem cells – Is it feasible and does it matter?

Present evidence indicates that acute myeloid leukemia (AML) is a stem cell disease. Leukemia stem cells (LSC) might originate from malignant transformation of normal hematopoietic stem cells (HSC), or alternatively, from progenitors in which the acquired mutations have re-installed a dysregulated self-renewal program. Since LSC, similar to their normal counterparts, divide extreme slowly, this might account for the ineffectiveness of conventional chemotherapy in inducing long-term cure. The present review will focus on the detection of LSC, their cellular and molecular biology, their genetic heterogeneity and on correlative studies that have demonstrated the clinical significance of estimating LSC burden. For long-term cure of AML, it is of importance to define LSC candidates and to understand their biology compared to normal HSC. Finally, we will discuss the perspectives of developing treatment strategies for eradication of LSC.

Differences between healthy hematopoietic progenitors and leukemia cells with respect to CD44 mediated rolling versus adherence behavior on hyaluronic acid coated surfaces

We previously demonstrated that leukemia cell lines expressing CD44 and hematopoietic progenitor cells (HPC) from umbilical cord blood (CB) showed rolling on hyaluronic acid (HA)-coated surfaces under physiological shear stress. In the present study, we quantitatively assessed the interaction of HPC derived from CB, mobilized peripheral blood (mPB) and bone marrow (BM) from healthy donors, as well as primary leukemia blasts from PB and BM of patients with acute myeloid leukemia (AML) with HA. We have demonstrated that HPC derived from healthy donors showed relative homogeneous rolling and adhesion to HA. In contrast, highly diverse behavioral patterns were found for leukemia blasts under identical conditions. The monoclonal CD44 antibody (clone BU52) abrogated the shear stress-induced rolling of HPC and leukemia blasts, confirming the significance of CD44 in this context. On the other hand, the immobile adhesion of leukemia blasts to the HA-coated surface was, in some cases, not or incompletely inhibited by BU52. The latter property was associated with non-responsiveness to induction chemotherapy and subsequently poor clinical outcome.

Identification of leukemia stem cells in acute myeloid leukemia and their clinical relevance

Acute myeloid leukemia (AML) is considered to be a disease of stem cells. A rare defective stem cell population is purported to drive tumor growth. Similarly to their normal counterparts, leukemic stem cells (LSC) divide extreme slowly. This may explain the ineffectiveness of conventional chemotherapy in combatting this disease. Novel treatment strategies aimed at disrupting the binding of LSC to stem cell niches within the bone marrow might render the LSC vulnerable to chemotherapy and thus improving treatment outcome. This review focuses on the detection of LSC, our current knowledge about their cellular and molecular biology, and LSC interaction with the niche. Finally, we discuss the clinical relevance of LSC and prospective targeted treatment strategies for patients with AML.

Identification and Separation of Normal Hematopoietic Stem Cells and Leukemia Stem Cells from Patients with Acute Myeloid Leukemia

Mounting evidences indicate that leukemic cells in patients with acute myeloid leukemia (AML) are derived from leukemia stem cells (LSC). In analogy to normal hematopoietic stem cells (HSC), LSC remain mostly dormant and are hence resistant to conventional chemotherapy. Residual, physiological HSC exist alongside with LSC, with heterogeneous dominance of LSC over HSC in individual patients. We have devised a flow cytometric method for the identification and separation of these two stem cell populations based on surface antigen markers such as CD34, CD38, lineage aberrant markers, and aldehyde dehydrogenase (ALDH) enzyme activity.

Reduced hematopoietic stem cell frequency predicts outcome in acute myeloid leukemia

In patients with acute myeloid leukemia and low percentages of aldehyde-dehydrogenase-positive cells, non-leukemic hematopoietic stem cells can be separated from leukemic cells. By relating hematopoietic stem cell frequencies to outcome we detected poor overall- and disease-free survival of patients with low hematopoietic stem cell frequencies. Serial analysis of matched diagnostic and follow-up samples further demonstrated that hematopoietic stem cells increased after chemotherapy in patients who achieved durable remissions. However, in patients who eventually relapsed, hematopoietic stem cell numbers decreased dramatically at the time of molecular relapse demonstrating that hematopoietic stem cell levels represent an indirect marker of minimal residual disease, which heralds leukemic relapse. Upon transplantation in immune-deficient mice cases with low percentages of hematopoietic stem cells of our cohort gave rise to leukemic or no engraftment, whereas cases with normal hematopoietic stem cell levels mostly resulted in multi-lineage engraftment. Based on our experimental data, we propose that leukemic stem cells have increased niche affinity in cases with low percentages of hematopoietic stem cells. To validate this hypothesis, we developed new mathematical models describing the dynamics of healthy and leukemic cells under different regulatory scenarios. These models suggest that the mechanism leading to decreases in hematopoietic stem cell frequencies before leukemic relapse must be based on expansion of leukemic stem cells with high niche affinity and the ability to dislodge hematopoietic stem cells. Thus, our data suggest that decreasing numbers of hematopoietic stem cells indicate leukemic stem cell persistence and the emergence of leukemic relapse.