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TGFβ1-mediated functional inhibition of mesenchymal stromal cells in MDS and AML

ABSTRACT: Mesenchymal stromal cells (MSC) are involved in the pathogenesis of myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML), but how they contribute to the expansion of malignant cells and hematopoietic failure is poorly understood. To further characterize the pathological phenotype we performed RNA sequencing of MSC from patients with MDS and AML. Data analysis revealed a specific molecular signature with a significant overlap of genes commonly deregulated in all MDS subtypes and in AML. Pathway analysis revealed a strong enrichment of genes related to osteogenesis, senescence, inflammatory processes and inhibitory cytokines thereby reflecting the structural and functional deficits of MDS- and AML-derived MSC on a molecular level. Further analysis identified TGFß1 as the most probable extrinsic trigger factor for this altered gene expression. Following exposure to TGFß1 healthy MSC adopted a phenotype reminiscent of that observed in primary patient-derived MSC which was characterized by impaired growth potential, osteogenic differentiation capacities, a specific gene expression profile and diminished stromal hematopoietic support. These suppressive effects of TGFß1 on MSC functionality were abrogated by SD-208, an established inhibitor of TGFß receptor signalling. Blockade of TGFß signalling by SD-208 also restored the osteogenic differentiation capacity of patient-derived MSC, thus confirming the role of TGFß1 in the bone marrow microenvironment of patients with MDS and AML. Our findings establish TGFß1 as a relevant trigger causing functional inhibition of MSC in MDS and AML and identify SD-208 as a candidate for the reversal of these effects

ORGANISM(S): Homo sapiens

PROVIDER: GSE107490 | GEO | 2018/11/29

REPOSITORIES: GEO

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Project description:Ineffective hematopoiesis is a hallmark of myelodysplastic syndromes (MDS). Hematopoietic alterations in MDS patients strictly correlate with microenvironment dysfunctions, eventually affecting also the mesenchymal stromal cells (MSCs) compartment. Stromal cells are indeed epigenetically reprogrammed to cooperate with leukemic cells and propagate the disease as "tumor unit"; therefore, changes on MSCs epigenetic profile might contribute to the hematopoietic perturbations typical of MDS. Here, we unveil that the histone variant macroH2A1 (mH2A1) regulates the crosstalk between epigenetics and inflammation in MDS-MSCs, potentially affecting their hematopoietic support ability. We show that the mH2A1 splicing isoform mH2A1.1 accumulates in MDS-MSCs, correlating with the expression of the Toll-like receptor 4 (TLR4), an important pro-tumor activator of MSC phenotype associated to a pro-inflammatory behavior. MH2A1.1-TLR4 axis was further investigated in HS-5 stromal cells after ectopic mH2A1.1 overexpression (mH2A1.1-OE). Proteomic data confirmed the activation of a pro-inflammatory signature associated to TLR4 and nuclear factor kappa B (NFkB) activation. Moreover, mH2A1.1-OE proteomic profile identified several upregulated proteins associated to DNA and histones hypermethylation, including S-adenosylhomocysteine hydrolase, a strong inhibitor of DNA methyltransferase and of the methyl donor S-adenosyl-methionine (SAM). HPLC analysis confirmed higher SAM/SAH ratio along with a metabolic reprogramming. Interestingly, an increased LDHA nuclear localization was detected both in mH2A1.1-OE cells and MDS-MSCs, probably depending on MSC inflammatory phenotype. Finally, coculturing healthy mH2A1-OE MSCs with CD34+ cells, we found a significant reduction in the number of CD34+ cells, which was reflected in a decreased number of colony forming units (CFU -Cs). These results suggest a key role of mH2A1.1 in driving the crosstalk between epigenetic signaling, inflammation and cell metabolism networks in MDS-MSCs.

Project description:Heterogeneity and variable survival outcomes of Acute Myeloid Leukemia (AML), suggest that yet undiscovered genes and pathways contribute to AML. Mesenchymal stem cells (MSC), an important component of bone marrow/ stromal microenvironment has been shown to contribute to development and progression of various cancers. Current study was aimed at characterizing MSC from AML bone marrow (BM) and evaluate their therapeutic potential in controlling survival of AML leukemic blasts. MSCs were isolated and cultured from BM of high-risk AML patients. MSC were also obtained from BM of bi-lineage leukemia (ETP-ALL) patients as control MSC from precursor stage of leukemia. MSC derived from uninvolved BM (UnBM) of lymphoma patients were used as normal MSC control. Leukemic blasts were derived from BM of AML and ETP-ALL patients. Patient derived AML-MSC exhibited characteristic profile of MSC Type-II CD90+CD45lo expressing high percent of TLR3 than TLR4 receptors, indicating pro-tumorigenic nature. Gene expression profiles of freshly derived leukemia blasts, AML cell line and AML MSC exhibited deregulated and overactivated Aurora Kinase pathway and inflammasome innate immune pathway. These two pathways are known to be upregulated in many solid and hematological cancers. Further we observed few tumor suppressor genes were downregulated in these groups. In vitro, AML MSC supported survival of leukemic blasts and increased chemoresistance to standard anticancer drugs. Hence AML MSC and ETP-ALL MSC were treated with immunomodulatory drug cocktail (IMiD) containing TLR3 antibody, TLR4 ligand and CXCR4 antagonist peptide (Gift from Kyoto University, Japan), designated as MSC-IMiD. It was observed that MSC-IMiD reduced chemoresistance of leukemic blasts to certain extent in vitro. Further we evaluated if leukemia BM derived MSC, MSC culture supernatant or MSC-IMiD can provide therapeutic benefit to control growth of AML xenograft in mouse model. Human gene expression profile was mapped in human-mouse xenograft made as subcutaneous tumor in immunodeficient NOD-SCID mice model from AML cell line KG1. Comparison was done of all treated groups with tumor bearing control gene expression data. It was interesting to note that MSC and MSC-IMiD could reduce tumor growth in 50% of mice tested. Mice treated with MSC culture supernatant exhibited reduction in tumor growth at par with that seen in Adriamycin treated positive control group. Deregulation of Aurora kinase pathway, inflammasome pathway genes and tumor suppressor genes was found to be reversed in residual tumor tissue of mice treated with MSC and MSC culture supernatant. This gene expression profiling study has helped understanding pathways involved in chemoresistance and immune suppression observed in AML. Moreover, this study has provided leads that MSC or its conditioned media can be explored further to control abnormal myelopoiesis in AML and develop targeted therapy. Funding source: Grant ID: 53/13/2013/CMB/BMS. Grantee: Jyoti Kode Grant title: Understanding the cross-talk between mesenchymal stromal cells and leukemic stem cells in Acute Myeloid Leukemia: Implications in disease biology and therapy. Name of the funding source: Indian Council of Medical Research, Government of India, India.

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TGFβ1-mediated functional inhibition of mesenchymal stromal cells in MDS and AML

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