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Metoclopramide treatment blocks CD93-signaling mediated self-renewal of chronic myeloid leukemia stem cells

ABSTRACT: Purpose: The goal of this study was to compare the transcriptome of FACS-purified bone marrow BL/6 (WT) or CD93-/- (KO) LSKs and BL/6 (WT) or CD93-/- (KO) LSCs. We also compared the transcriptome of FACS-purified bone marrow LSCs isolated from BL/6 mice previously treated with MCP or Veh in vivo. Methods: Transcriptomic analysis of CD93-proficient and deficient bone marrow LSKs and CML LSCs or CML LSCs upon treatment with MCP or Veh, were assessed in biological replicates using Illumina. qRT–PCR validation was performed using SYBR Green assays. Results: We mapped around 30 million sequence reads per sample to the mouse genome (GRCm38 - mm10) and identified expressed transcripts in studied samples. RNA-seq data confirmed stable expression of known housekeeping genes. Differentially expressed genes among conditions were identified with a fold change ≥1.5 and FDR p-value <0.05. Conclusions: Our study represents the first detailed transcriptome analysis of CD93-proficient and deficient bone marrow LSKs and LSCs isolated from BM of naïve and CML mice generated by RNA-seq. technology. Our results show that CD93-signaling triggers stem cell maintenance- and cell proliferation-promoting signaling pathways in CML LSCs. In addition, we showed transcriptome analysis of FACS-purified bone marrow LSCs isolated from BL/6 (WT) mice which were previously treated with MCP or Veh in vivo. Our results show that MCP treatment suppresses the stem cell maintenance- and cell proliferation-promoting signaling pathways in CML LSCs.

ORGANISM(S): Mus musculus

PROVIDER: GSE149358 | GEO | 2021/02/07

REPOSITORIES: GEO

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Project description:MiR-142 is dynamically expressed and plays a regulatory role in hematopoiesis. Based on the simple observation that miR-142 levels are significantly lower in CD34+CD38- cells from blast crisis (BC) chronic myeloid leukemia (CML). CML patients compared with chronic phase (CP) CML patients (p=0.002), we hypothesized that miR-142 deficit plays a role in BC transformation. To test this hypothesis, we generated a miR-142 KO BCR-ABL (i.e., miR-142−/−BCR-ABL) mouse by crossing a miR-142−/− mouse with a miR-142+/+BCR-ABL mouse. While the miR-142+/+BCR-ABL mice developed and died of CP CML, the miR-142−/−BCR-ABL mice developed a BC-like phenotype in the absence of any other acquired gene mutations and died significantly sooner than miR-142+/+BCR-ABL CP controls (p=0.001). Leukemic stem cell (LSC)-enriched Lineage-Sca-1+c-Kit+ cells (LSKs) from diseased miR-142−/−BCR-ABL mice transplanted into congenic recipients, recapitulated the BC features thereby suggesting stable transformation of CP-LSCs into BC-LSCs in the miR-142 KO CML mouse. Single cell (sc) RNA-seq profiling showed that miR-142 deficit changed the cellular landscape of the miR-142−/−BCR-ABL LSKs compared with miR-142+/+BCR-ABL LSKs with expansion of myeloid-primed and loss of lymphoid-primed factions. Bulk RNA-seq analyses along with unbiased metabolomic profiling and functional metabolic assays demonstrated enhanced fatty acid β-oxidation (FAO) and oxidative phosphorylation (OxPhos) in miR-142−/−BCR-ABL LSKs vs miR-142+/+BCR-ABL LSKs. MiR-142 deficit enhanced FAO in miR-142−/−BCR-ABL LSKs by increasing the expression of CPT1A and CPT1B, that controls the cytosol-to-mitochondrial acyl-carnitine transport, a critical step in FAO. MiR-142 deficit also enhanced OxPhos in miR-142−/−BCR-ABL LSKs by increasing mitochondrial fusion and activity. As the homeostasis and activity of LSCs depend on higher levels of these oxidative metabolism processes, we then postulate that miR-142 deficit is a potentially druggable target for BC-LSCs. To this end, we developed a novel CpG-miR-142 mimic oligonucleotide (ODN; i.e., CpG-M-miR-142) that corrected the miR-142 deficit and alone or in combination with a tyrosine kinase inhibitor (TKI) significantly reduced LSC burden and prolonged survival of miR-142−/−BCR-ABL mice. The results from murine models were validated in BC CD34+CD38- primary blasts and patient-derived xenografts (PDXs). In conclusion, an acquired miR-142 deficit sufficed in transforming CP-LSCs into BC-LSCs, via enhancement of bioenergetic oxidative metabolism in absence of any additional gene mutations, and likely represent a novel therapeutic target in BC CML.

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Metoclopramide treatment blocks CD93-signaling mediated self-renewal of chronic myeloid leukemia stem cells

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