Cell Contact with Endothelial Cells Favors the In Vitro Maintenance of Human Chronic Myeloid Leukemia Stem and Progenitor Cells.

Chronic Myeloid Leukemia (CML) originates in a leukemic stem cell that resides in the bone marrow microenvironment, where they coexist with cellular and non-cellular elements. The vascular microenvironment has been identified as an important element in CML development since an increase in the vascularization has been suggested to be related with poor prognosis; also, using murine models, it has been reported that bone marrow endothelium can regulate the quiescence and proliferation of leukemic stem and progenitor cells. This observation, however, has not been evaluated in primary human cells. In this report, we used a co-culture of primitive (progenitor and stem) CML cells with endothelial colony forming cells (ECFC) as an in vitro model to evaluate the effects of the vascular microenvironment in the leukemic hematopoiesis. Our results show that this interaction allows the in vitro maintenance of primitive CML cells through an inflammatory microenvironment able to regulate the proliferation of progenitor cells and the permanence in a quiescent state of leukemic stem cells.

Evolución temporal de una línea celular leucémica que compite con la hematopoyesis saludable / Temporal evolution of a leukemic line that competes with healthy hematopoiesis

Introducción: La hipótesis del cáncer de las células madre se ha convertido en uno de los paradigmas más importantes en la investigación biomédica. Durante los últimos años se ha ido acumulando evidencia de la existencia de poblaciones similares a células madre en diferentes tipos de cáncer, especialmente en las leucemias. Objetivos: Mostrar mediante la modelación matemática y la simulación computacional cómo los cambios en los parámetros que describen las tasas de proliferación y las propiedades de autorrenovación pueden influir en la dinámica de las poblaciones de células sanas y leucémicas. Métodos: Se utilizó un modelo matemático que es una extensión de los modelos de hematopoyesis sana. Se resolvió el modelo mediante herramientas computacionales basadas en métodos numéricos, lo que permitió realizar simulaciones con diferentes parámetros e intervalos de tiempo. Resultados: Al imponer ciertas condiciones iniciales y resolver matemáticamente el modelo se obtuvo la evolución temporal de las variables de estado del sistema hematopoyético. Partiendo de un estado conocido del sistema hematopoyético se predijo el comportamiento en el tiempo de las variables de estado. Se particularizó para cuatro escenarios clínicamente relevantes. Conclusiones: El análisis del modelo dio como resultado diferentes escenarios de crecimiento de células leucémicas, entre los cuales la proliferación aumentada de células malignas es el más prominente. Sin embargo, diferentes escenarios son posibles, como la inducción de apoptosis o la autorrenovación mejorada(AU)

Introduction: The stem cell cancer hypothesis has become one of the most important paradigms in biomedical research. In recent years, evidence has accumulated for the existence of stem cell-like populations in different types of cancer, especially in leukemias. Objectives: To show, through mathematical modeling and computational simulation, how changes in the parameters that describe proliferation rates and self-renewal properties can influence the dynamics of healthy and leukemic cell populations. Methods: A mathematical model was used which is an extension of the healthy hematopoiesis models. The model was solved using computational tools based on numerical methods, this allowed to carry out countless simulations with different parameters and time intervals. Results: By imposing certain initial conditions and mathematically solving the model, the temporal evolution of the state variables of the hematopoietic system was obtained, that is, starting from a known state of the hematopoietic system, the behavior over time of the state variables of the system was predicted. It was particularized for four clinically relevant scenarios. Conclusions: The analysis of the model results in different growth scenarios of leukemic cells, among which the increased proliferation of malignant cells is the most prominent. However, different scenarios are possible, such as apoptosis induction or enhanced self-renewal(AU)

Cell signaling pathways as molecular targets to eliminate AML stem cells.

Acute myeloid leukemia (AML) remains the most lethal of leukemias and a small population of cells called leukemic stem cells (LSCs) has been associated with disease relapses. Some cell signaling pathways play an important role in AML survival, proliferation and self-renewal properties and are abnormally activated or suppressed in LSCs. This includes the NF-κB, Wnt/ß-catenin, Hedgehog, Notch, EGFR, JAK/STAT, PI3K/AKT/mTOR, TGF/SMAD and PPAR pathways. This review aimed to discuss these pathways as molecular targets for eliminating AML LSCs. Herein, inhibitors/activators of these pathways were summarized as a potential new anti-AML therapy capable of eliminating LSCs to guide future researches. The clinical use of cell signaling pathways data can be useful to enhance the anti-AML therapy.

Evaluation of the primitive fraction by functional in vitro assays at the RNA and DNA level represents a novel tool for complementing molecular monitoring in chronic myeloid leukemia.

Quantification of BCR-ABL1 mRNA levels in peripheral blood of chronic myeloid leukemia patients is a strong indicator of response to tyrosine-kinase inhibitors (TKI) treatment. However, additional prognostic markers are needed in order to better classify patients. The hypothesis of leukemic stem cells (LSCs) heterogeneity and persistence, suggests that their functional evaluation could be of clinical interest. In this work, we assessed the primitive and progenitor fractions in patients at diagnosis and during TKI treatment using functional in vitro assays, defining a "functional leukemic burden" (FLB). We observed that the FLB was reduced in vivo in both fractions upon treatment. However, different FLB levels were observed among patients according to their response to treatment, suggesting that quantification of the FLB could complement early molecular monitoring. Given that FLB assessment is limited by BCR-ABL1 mRNA expression levels, we developed a novel detection method of primitive cells at the DNA level, using patient-specific primers and direct nested PCR in colonies obtained from functional in vitro assays. We believe that this method could be useful in the context of discontinuation trials, given that it is unknown whether the persistent leukemic clone represents LSCs, able to resume the leukemia upon TKI removal.

CDKIs p18(INK4c) and p57(Kip2) are involved in quiescence of CML leukemic stem cells after treatment with TKI.

Chronic Myeloid Leukemia (CML) is sustained by a small population of cells with stem cell characteristics known as Leukemic Stem Cells that are positive to BCR-ABL fusion protein, involved with several abnormalities in cell proliferation, expansion, apoptosis and cell cycle regulation. Current treatment options for CML involve the use of Tirosine Kinase Inhibitor (Imatinib, Nilotinib and Dasatinib), that efficiently reduce proliferation proliferative cells but do not kill non proliferating CML primitive cells that remain and contributes to the persistence of the disease. In order to understand the role of Cyclin Dependent Kinase Inhibitors in CML LSC permanence after TKI treatment, in this study we analyzed cell cycle status, the levels of several CDKIs and the subcellular localization of such molecules in different CML cell lines, as well as primary CD34(+)CD38(-)lin(-) LSC and HSC. Our results demonstrate that cellular location of p18(INK4c) and p57(Kip2) seems to be implicated in the antiproliferative activity of Imatinib and Dasatinib in CML cells and also suggest that the permanence of quiescent stem cells after TKI treatment could be associated with a decrease in p18(INK4c) and p57(Kip2) nuclear location. The differences in p18(INK4c)and p57(Kip2)activities in CML and normal stem cells suggest a different cell cycle regulation and provide a platform that could be considered in the development of new therapeutic options to eliminate LSC.