RESUMO
Introduction: Acute myeloid leukemia (AML) is a highly heterogeneous malignancy caused by various genetic alterations and characterized by the accumulation of immature myeloid blasts in the bone marrow (BM). This abnormal growth of AML cells disrupts normal hematopoiesis and alters the BM microenvironment components, establishing a niche supportive of leukemogenesis. Bone marrow stromal cells (BMSCs) play a pivotal role in giving rise to essential elements of the BM niche, including adipocytes and osteogenic cells. Animal models have shown that the BM microenvironment is significantly remodeled by AML cells, which skew BMSCs toward an ineffective osteogenic differentiation with an accumulation of osteoprogenitors. However, little is known about the mechanisms by which AML cells affect osteogenesis. Methods: We studied the effect of AML cells on the osteogenic commitment of normal BMSCs, using a 2D co-culture system. Results: We found that AML cell lines and primary blasts, but not normal hematopoietic CD34+ cells, induced in BMSCs an ineffective osteogenic commitment, with an increase of the early-osteogenic marker tissue non-specific alkaline phosphatase (TNAP) in the absence of the late-osteogenic gene up-regulation. Moreover, the direct interaction of AML cells and BMSCs was indispensable in influencing osteogenic differentiation. Mechanistic studies identified a role for AML-mediated Notch activation in BMSCs contributing to their ineffective osteogenic commitment. Inhibition of Notch using a γ-secretase inhibitor strongly influenced Notch signaling in BMSCs and abrogated the AML-induced TNAP up-regulation. Discussion: Together, our data support the hypothesis that AML infiltration produces a leukemia-supportive pre-osteoblast-rich niche in the BM, which can be partially ascribed to AML-induced activation of Notch signaling in BMSCs.
Assuntos
Leucemia Mieloide Aguda , Células-Tronco Mesenquimais , Animais , Osteogênese , Células da Medula Óssea/metabolismo , Leucemia Mieloide Aguda/patologia , Medula Óssea/metabolismo , Células-Tronco Mesenquimais/metabolismo , Microambiente TumoralRESUMO
In acute myeloid leukemia (AML), malignant stem cells hijack the normal bone marrow niche where they are largely protected from the current therapeutic approaches. Thus, eradicating these progenitors is the ultimate challenge in the treatment of this disease. Specifically, the development of chimeric antigen receptors (CARs) against distinct mesenchymal stromal cell subpopulations involved in the maintenance of leukemic stem cells within the malignant bone marrow microenvironment could represent a new strategy to improve CAR T-cell therapy efficacy, which is still unsuccessful in AML. As a proof of concept, we generated a novel prototype of Tandem CAR, with one specificity directed against the leukemic cell marker CD33 and the other against the mesenchymal stromal cell marker CD146, demonstrating its capability of simultaneously targeting two different cell types in a 2D co-culture system. Interestingly, we could also observe an in vitro inhibition of CAR T cell functionality mediated by stromal cells, particularly in later effector functions, such as reduction of interferon-gamma and interleukin-2 release and impaired proliferation of the CAR+ effector Cytokine-Induced Killer (CIK) cells. Taken together, these data demonstrate the feasibility of a dual targeting model against two molecules, which are expressed on two different target cells, but also highlight the immunomodulatory effect on CAR CIK cells exerted by stromal cells, confirming that the niche could be an obstacle to the efficacy of CAR T cells. This aspect should be considered in the development of novel CAR T cell approaches directed against the AML bone marrow niche.