Project description:We report the gene expression profiling of mouse BC CML model which were knockdown with either shControl or shAMD1 constructs. By obtaining RNA from primary BC CML cells from those mice (n: 3 for each shRNA),we find that thousands gene were dysregulated by AMD1 knockdown.
Project description:To gain insight into how miR-142 deficit drives a BC-like transformation, we performed RNA-seq on bone marrow (BM) Lin-Sca-1+c-Kit+ cells (LSKs) harvested from normal miR-142+/+ (wt) and miR-142−/− (miR-142 KO) mice, as well as from leukemic miR-142+/+ BCR-ABL (CP CML) and miR-142−/− BCR-ABL (BC CML) mice, two weeks after BCR-ABL induction. We then performed gene expression profiling analysis using data obtained from RNA-seq of 24 samples of LSK cells from 4 mouse strains (KO vs WT, KO CML vs CML).
Project description:To assess how the miR-142 deficit affected the BM LSC-enriched LSK landscape, we conducted a single cell RNA-seq analysis of BM miR-142+/+ BCR-ABL (CP CML) and miR-142−/− BCR-ABL (BC CML) LSKs which were harvested from the respective mice, 2 weeks after BCR-ABL induction.
Project description:Microarray analysis of primary effusion lymphoma cell lines BC-1 and BC-3 Total RNA from BC-1 and BC-3 cell lines was processed for analysis in one replicate on Human Gene 1.0 ST arrays to obtain data on whether genes are expressed and to compare to existing microarray data.
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.
Project description:A comparison of global gene expression between rigorously defined stem and progenitor cells from patients with chronic myeloid leukaemia (CML) in chronic (CP), accelerated (AP) and blastic (BC) phase and similar populations isolated from normal volunteers. Cryopreserved CD34+ enriched cell populations obtained from patients with CML in CP, AP or BC at diagnosis prior to treatment -- or from normal volunteers -- were thawed and flow sorted into rigorously defined sub-populations (HSC, MPP, CMP, GMP and MEP -- using surface phenotype as described below). Total RNA was obtained and global gene expression measured following hybridisation to Affymetrix HuGene-1_0-st-v1 gene-chips. In total, 3 normal patient samples were compared with 6 CP, 4 AP and 2 BC samples. HSC, CMP, GMP and MEP populations were obtained from all specimens, MPP populations were obtained from normal, AP and BC specimens but not CP specimens.
Project description:Tyrosine kinase inhibitor (TKI) treatment of chronic myeloid leukemia (CML) is guided by the pre-defined European Leukemia Net (ELN) or other response criteria. This allows patient stratification only during but not prior to treatment initiation. Gene expression profiling (GEP)-based response prediction might become a valuable tool for patient stratification in CML, but so far published data for response prediction are conflicting. We generated an imatinib response predicting gene signature by GEP from peripheral blood samples of pre-treated CML patients in late chronic phase.
Project description:A comparison of global gene expression between rigorously defined stem and progenitor cells from patients with chronic myeloid leukaemia (CML) in chronic (CP), accelerated (AP) and blastic (BC) phase and similar populations isolated from normal volunteers.
Project description:We examine the oncogenic function of the Far Upstream Element Binding Protein 1 (FUBP1) in leukemia-initiating cells. For this perpose we performed knockdown of Fubp1 in CML cells in our mouse model. Here, we transduce donor bone marrow cell with the oncogene BCR-ABL1 and Fubp1 or scrambled shRNA and transplanted in recipient mice. Lineage- BCR-ABL1+ shRNA+ cells were then sorted from diseased mice and analyzed by RNA sequencing. We compare the gene expression of the Fubp1 shRNA expressing CML cells versus the control (scrambled shRNA expressing) cells in order to identify the relevant Fubp1 target genes.