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Expression data from BM-CD138+, BM B cells and PBL B cells obtained from newly diagnosed Multiple Myeloma patients

ABSTRACT: The iperactivation of self-renewal mechanisms, like Hedgehog (HH) pathway, which govern the refuel of Multiple Myeloma neoplastic clone, is critical to relapse events. The bulk of current knowledge is mainly based on the CD138+ cells molecular characterization, but in the light of the recent evidences, which attributed growing relevance to immature precursor clones, we become aware that multiple myeloma disease heterogeneity have more deepen roots. Here, we report on a high-throughput transcriptome study of gene expression profiles in newly diagnosed MM patients analyzed both in the CD138+ plasmacell fraction and in the CD19+ B cells compartment. Resulted data demonstrated that Hedgehog pathway, and in particular, a 10 HH-genes signature was heterogeneously expressed among MM patients, both in mature plasmacells and also in B cells at transcriptional level. Interestingly, this signature divide patients in two subgroups, and it seems that exists a sort of “ying-yang effect” between mature and immature cell compartments that are tightly regulate in order to express this relevant self-renewal pathway according to their maturation state.

ORGANISM(S): Homo sapiens

PROVIDER: GSE68891 | GEO | 2016/05/31

SECONDARY ACCESSION(S): PRJNA283981

REPOSITORIES: GEO

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Project description:Despite recent advances in the treatment of multiple myeloma (MM), it remains an incurable disease potentially due to the presence of resistant myeloma cancer stem cells (MM-CSC). Although the presence of clonogenic cells in MM was described more than 30 years ago, the phenotype of MM-CSC is still a matter of debate, especially with respect to the expression of syndecan- 1 (CD138). Here, we demonstrate the presence of two subpopulations - CD138++ (95-99%) and CD138low (1-5%) - in eight MM cell lines. To find out possible stem-cell-like features, we have phenotypically, genomic and functionally characterized the two subpopulations. Our results show that the minor CD138low subpopulation is morphologically identical to the CD138++ fraction and does not represent a more immature B-cell compartment (with lack of CD19, CD20 and CD27 surface expression). Moreover, both subpopulations have similar gene expression and genomic profiles. Importantly, both CD138++ and CD138low subpopulations have similar sensitivity to bortezomib, melphalan and doxorubicin. Finally, serial engraftment in SCID mice shows that CD138++ as well as CD138low cells have self-renewal potential and they are also phenotypically interconvertible. Overall, our results differ from previously published data which attribute a B-cell phenotype to MM-CSC and urge the need to explore more reliable markers to discriminate true clonogenic myeloma cells. To compare the gene expression profile of the subpopulations CD138++ and CD138low within the MM cell line, RPMI-8226. RNA was isolated from CD138++ (n=3) and CD138low (n=3) RPMI-8226 cells. RNA was hybridized to Human Gene 1.0 ST array (Affymetrix) according to Affymetrix protocols [Gutierrez, 2005, 402]. Microarray data were normalized using the Robust Multichip Analysis (RMA) algorithm implemented in the Affymetrix Expression Console. Data analysis was carried out using DNA-Chip Analyzer software (DChip). The comparison criteria used in DChip analysis were fold change E/BM-bM-^IM-%2 or B/EM-bM-^IM-%2, mean difference E-B>100 or B-E>100 and the lower 90% confidence bound of fold-change was used

Project description:Despite recent advances in the treatment of multiple myeloma (MM), it remains an incurable disease potentially due to the presence of resistant myeloma cancer stem cells (MM-CSC). Although the presence of clonogenic cells in MM was described more than 30 years ago, the phenotype of MM-CSC is still a matter of debate, especially with respect to the expression of syndecan- 1 (CD138). Here, we demonstrate the presence of two subpopulations - CD138++ (95-99%) and CD138low (1-5%) - in eight MM cell lines. To find out possible stem-cell-like features, we have phenotypically, genomic and functionally characterized the two subpopulations. Our results show that the minor CD138low subpopulation is morphologically identical to the CD138++ fraction and does not represent a more immature B-cell compartment (with lack of CD19, CD20 and CD27 surface expression). Moreover, both subpopulations have similar gene expression and genomic profiles. Importantly, both CD138++ and CD138low subpopulations have similar sensitivity to bortezomib, melphalan and doxorubicin. Finally, serial engraftment in SCID mice shows that CD138++ as well as CD138low cells have self-renewal potential and they are also phenotypically interconvertible. Overall, our results differ from previously published data which attribute a B-cell phenotype to MM-CSC and urge the need to explore more reliable markers to discriminate true clonogenic myeloma cells. To compare the Copy Number Abnormalities of the subpopulations CD138++ and CD138low within the MM cell line, RPMI-8226. DNA was isolated from CD138++ (n=3) and CD138low (n=3) RPMI-8226 cells. The detection of CNA was investigated using the CytoScan HD array (Affymetrix, Santa Clara, CA, USA) which includes more than 2.6 million copy number markers and 750.000 SNPs. Briefly, genomic DNA was digested with Nsp I restriction endonuclease, ligated to adaptors that recognize the cohesive four base pair overhangs and amplified by PCR. PCR product was purified and fragmented with DNAse I and then end-labeled using terminal deoxynucleotidyl transferase and hybridized to the CytoScan HD array. The arrays were processed using the Fluidics Station 450, GeneChip Scanner 3000 7 G and AGCC (AffymetrixGeneChip Command Console Software). For the analysis we employed the AffymetrixM-BM-. Chromosome Analysis Suite (ChAS) Software v2.0 and Nexus Copy Number Software Version 7.0 (BioDiscovery, El Segundo, CA, USA). The complete data set was analyzed by visual inspection using ChAS software. CNA were reported when the three following criteria were achieved: minimum of 50 markers per segment, 200 Kb minimum genomic sizes and 50% overlap with known copy number variants (Database of Genomic Variants).

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Expression data from BM-CD138+, BM B cells and PBL B cells obtained from newly diagnosed Multiple Myeloma patients

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