Abstract
BackgroundDuring acute myeloid leukemia (AML) growth, the bone marrow (BM) niche acquires significant vascular changes that can be offset by therapeutic blast cytoreduction. The molecular mechanisms of this vascular plasticity remain to be fully elucidated. Herein, we report on the changes that occur in the vascular compartment of the FLT3-ITD+ AML BM niche pre and post treatment and their impact on leukemic stem cells (LSCs).MethodsBM vasculature was evaluated in FLT3-ITD+ AML models (MllPTD/WT/Flt3ITD/ITD mouse and patient-derived xenograft) by 3D confocal imaging of long bones, calvarium vascular permeability assays, and flow cytometry analysis. Cytokine levels were measured by Luminex assay and miR-126 levels evaluated by Q-RT-PCR and miRNA staining. Wild-type (wt) and MllPTD/WT/Flt3ITD/ITD mice with endothelial cell (EC) miR-126 knockout or overexpression served as controls. The impact of treatment-induced BM vascular changes on LSC activity was evaluated by secondary transplantation of BM cells after administration of tyrosine kinase inhibitors (TKIs) to MllPTD/WT/Flt3ITD/ITD mice with/without either EC miR-126 KO or co-treatment with tumor necrosis factor alpha (TNFα) or anti-miR-126 miRisten.ResultsIn the normal BM niche, CD31+Sca-1high ECs lining arterioles have miR-126 levels higher than CD31+Sca-1low ECs lining sinusoids. We noted that during FLT3-ITD+ AML growth, the BM niche lost arterioles and gained sinusoids. These changes were mediated by TNFα, a cytokine produced by AML blasts, which induced EC miR-126 downregulation and caused depletion of CD31+Sca-1high ECs and gain in CD31+Sca-1low ECs. Loss of miR-126high ECs led to a decreased EC miR-126 supply to LSCs, which then entered the cell cycle and promoted leukemia growth. Accordingly, antileukemic treatment with TKI decreased the BM blast-produced TNFα and increased miR-126high ECs and the EC miR-126 supply to LSCs. High miR-126 levels safeguarded LSCs, as shown by more severe disease in secondary transplanted mice. Conversely, EC miR-126 deprivation via genetic or pharmacological EC miR-126 knock-down prevented treatment-induced BM miR-126high EC expansion and in turn LSC protection.ConclusionsTreatment-induced CD31+Sca-1high EC re-vascularization of the leukemic BM niche may represent a LSC extrinsic mechanism of treatment resistance that can be overcome with therapeutic EC miR-126 deprivation.Graphic abstract
Highlights
During acute myeloid leukemia (AML) growth, the bone marrow (BM) niche acquires significant vascu‐ lar changes that can be offset by therapeutic blast cytoreduction
Utilizing the FMS-like tyrosine kinase 3 (FLT3) gene internal tandem duplication (ITD) (FLT3-ITD) knockin mouse and FLT3-ITD+ AML patient-derived xenograft (PDX) models that recapitulate features of human FLT3-ITD+ AML, we report here on previously unrecognized non-genetic, extrinsic mechanisms of treatment resistance in Leukemic stem cell (LSC) that involve the vascular compartment of the leukemic BM niche and that are mediated by a Tumor necrosis factor alpha (TNFα)-miR-126 axis in the BM endothelial cells (ECs)
In the normal mouse BM niche, we found that ECs expressed at least a log-fold higher level of miR-126 than normal long-term (LT) Hematopoietic stem cell (HSC) (LSK Fit3−CD150+CD48−) and other non-hematopoietic stromal cells, including osteoblasts (OBs; CD45−Ter119−CD31−CD166+Sca-1−) and mesenchymal stromal progenitors (MSPs; CD45−Ter119−CD31−CD166−Sca-1+) [30] (Additional file 1: Fig. S8a, b)
Summary
During acute myeloid leukemia (AML) growth, the bone marrow (BM) niche acquires significant vascu‐ lar changes that can be offset by therapeutic blast cytoreduction. We report on the changes that occur in the vascular compartment of the FLT3ITD+ AML BM niche pre and post treatment and their impact on leukemic stem cells (LSCs). We noted that during FLT3-ITD+ AML growth, the BM niche lost arterioles and gained sinusoids These changes were mediated by TNFα, a cytokine produced by AML blasts, which induced EC miR-126 downregu‐ lation and caused depletion of C D31+Sca-1high ECs and gain in CD31+Sca-1low ECs. Loss of miR-126high ECs led to a decreased EC miR-126 supply to LSCs, which entered the cell cycle and promoted leukemia growth. While these reported mechanisms have been mainly reported as intrinsic to malignant cells, it is possible that they include mechanisms that are extrinsic to malignant cells, such as those involving the microenvironment and that protect malignant cells during treatment exposure [7]
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