Plasma Cells Of Multiple Myeloma Patients Research Articles

Identification of Genes Associated with Increased Bone Marrow Angiogenesis in Multiple Myeloma.

Introduction: Increased bone marrow angiogenesis is a characteristic feature of multiple myeloma (MM) and correlates with disease progression. Increased bone marrow angiogenesis at the time of diagnosis, measured in terms of microvessel density (MVD), is a powerful adverse prognostic factor for patients with MM. To better understand the biological basis of this phenomenon and to understand the mechanisms responsible for increased MVD in MM we compared the gene expression profiles of plasma cells from newly diagnosed MM patients with low and high MVD.Methods: Bone marrow biopsy sections from pts with newly diagnosed MM were studied using CD34 immunostaining and graded as low, intermediate, or high by MVD as previously described. 19 pts each, with low or high MVD were included for this study. RNA was isolated from CD138+ plasma cells of MM patients and analyzed using Affymetrix HG-U133A arrays. To examine differential expression, GC-RMA normalized data was analyzed using GeneSpring 7.2 software and genes with ≥ 2-fold differential expression between high and low MVD samples were identified. The Welch T-test was used to determine the significance of differential expression.Results and Conclusion: Expression of 42 transcripts was increased ≥ 2-fold in samples with high MVD in comparison to samples with low MVD. Of these transcripts, 14 were found to be significantly increased (P<0.05) using the Welch T-test (see Table 1). Expression of 16 transcripts was decreased ≥ 2-fold in samples with high MVD in comparison to samples with low MVD. Of these transcripts, 6 were found to be significantly decreased (P<0.05) using the Welch T-test (see Table 1). Genes differentially expressed include those involved in inhibiting apoptosis, facilitating IL-6 signaling, ion transport, extracellular matrix interaction, and regulating gene expression. Classical angiogenesis genes including VEGF, FGF, and IGF were not found to be differentially expressed (> 2-fold).In conclusion, the list of differentially expressed genes reveals many functions relevant to MM disease pathology including proliferation, apoptosis, regulation of gene expression, cytokine signaling, and adhesion. Current studies evaluating the expression and function of these genes in relation to MM may identify factors critical for angiogenesis and MM progression and provide insight for therapeutic intervention.GeneDescriptionFold ChangeCOL1A2collagen, type I, alpha 23.008MCL1myeloid cell leukemia sequence 1 (BCL2-related)2.793209183_s_atchromosome 10 open reading frame 102.541VIL2villin 22.478IL6STinterleukin 6 signal transducer (gp130)2.404CLIC2chloride intracellular channel 22.364DEPDC6DEP domain containing 62.35PBXIP1pre-B-cell leukemia transcription factor interacting protein 12.337CLIC4chloride intracellular channel 42.334CYP51A1cytochrome P450, family 51, subfamily A, polypeptide 12.095LIMS1LIM and senescent cell antigen-like domains 12.091YWHAEtyrosine 3-monooxygenase/tryptophan 5-monooxygenase activation protein, epsilon polypeptide2.042OAS12′,5′-oligoadenylate synthetase 1, 40/46kDa2.031S100A11S100 calcium binding protein A112.018222378_atunknown transcribed sequences0.496SLC5A3mitochondrial ribosomal protein S60.463213089_atunknown transcribed sequences0.406PDE4Bphosphodiesterase 4B, cAMP-specific0.454SVILsupervillin0.386RIPXrap2 interacting protein x0.383

Stromal Cell IL-8 Production Is Increased in MM and May Impact Both Tumor Cells and the Microenvironment.

Background: Multiple myeloma (MM) is an incurable hematological malignancy characterized by the expansion of a plasma cell clone that localizes to the bone marrow. Stromal cells residing in the bone marrow respond to signals from MM cells and other cell types by producing cytokines and other proteins that stimulate tumor cell growth, survival, adhesion, migration, and drug resistance. We have examined the proteins produced by stromal cells in response to stimulation by bone marrow from patients diagnosed with monoclonal gammopathy of undetermined significance (MGUS), smoldering multiple myeloma (SMM), and MM. Furthermore, we have begun analyzing the properties of one of these proteins, the pro-angiogenic chemokine IL-8, in MM.Methods: Bone marrow aspirates from patients with MGUS (n=3), SMM (n=7), and MM (n=6) were cultured for 48 hours, and the culture supernatants were incubated with stromal cells for an additional 48 hours. Protein levels were analyzed using antibody array and ELISA. Microvessel density (MVD) was determined as a measure of angiogenesis in patient bone marrow samples using CD34 staining. Flow cytometry analysis of MM cell lines and patient bone marrow samples was performed using monoclonal antibodies against IL-8 receptors CXCR1 and CXCR2.Results and Conclusion: We observed a significant increase in stromal cell IL-8 production stimulated by bone marrow cells from patients with active myeloma and a subset of SMM patients (16.67 ± 9.82 ng/ml) in comparison to bone marrow of patients with MGUS and all other SMM patients (0.55 ± 0.17 ng/ml; P=0.0004). Use of an IL-1 inhibitor and recombinant IL-1β demonstrated that IL-8 production was dependent upon IL-1β signaling. Increased BM microvessel density correlated with stimulation of stromal cell IL-8 production (P=0.0005). Furthermore, the majority of MM cell lines (7/9) and MM patient plasma cells were found to express IL-8 receptors CXCR1 and CXCR2. In addition to its function as a pro-angiogenic factor, IL-8 may directly influence MM cells through its CXCR1 and CXCR2 receptors. We conclude that stromal cell IL-8 production parallels MM disease activity, is IL-1β induced, correlates with bone marrow angiogenesis, and may influence MM disease via impact upon both the microenvironment and tumor cells.

Muc-1 Core Protein Is Expressed on Multiple Myeloma Cells and Is Induced by Dexamethasone

AbstractMonoclonal antibodies (MoAbs) that selectively identify Muc-1 core protein (MoAbs DF3-P, VU-4H5) determinants were used to identify the Muc-1 glycoform present on 7 multiple myeloma (MM) cell lines, 5 MM patient plasma cells, 12 MM patient B cells, as well as 32 non-MM cell lines and normal hematopoietic cells. Flow cytometry studies demonstrated that all MM cell lines, MM patient plasma cells, and MM patient B cells expressed Muc-1 core protein epitopes. Circulating B cells from 4 normal donors also expressed Muc-1 core protein. In contrast, Muc-1 core protein was absent on 28 of 32 non-MM neoplastic cell lines, 17 of which expressed Muc-1. Splenic and tonsillar B cells, CD34+ stem cells, resting T cells, and bone marrow plasma cells obtained from normal donors both lacked Muc-1 glycoforms. We next studied the effects of estrogen, progesterone, and glucocorticoid receptor agonists and antagonists on Muc-1 expression, because consensus sequences for the response elements of these steroids are present on the Muc-1 gene promoter. These studies showed that dexamethasone (Dex) induced Muc-1 expression on MM cell lines, as determined by both flow cytometry and Western blot analyses. Dex also induced upregulation of Muc-1 on prostate and ovarian cancer cell lines. Time and dose-response studies demonstrated that Dex induced maximal cell surface Muc-1 expression by 24 hours at concentrations of 10−8 mol/L. Dex induced Muc-1 upregulation could be blocked with a 10-fold excess of the glucocorticoid receptor antagonist RU486, confirming that Dex was acting via the glucocorticoid receptor. No changes in Muc-1 expression were observed on MM cells treated with estrogen and progesterone receptor agonists and antagonists or with RU486. These studies provide the framework for targeting Muc-1 core protein in vaccination and serotherapy trials in MM. In addition, the finding that Muc-1 expression on MM cells can be augmented by Dex at pharmacologically achievable levels suggests their potential utility in enhancing treatments targeting Muc-1 in MM.

Muc-1 Core Protein Is Expressed on Multiple Myeloma Cells and Is Induced by Dexamethasone

Monoclonal antibodies (MoAbs) that selectively identify Muc-1 core protein (MoAbs DF3-P, VU-4H5) determinants were used to identify the Muc-1 glycoform present on 7 multiple myeloma (MM) cell lines, 5 MM patient plasma cells, 12 MM patient B cells, as well as 32 non-MM cell lines and normal hematopoietic cells. Flow cytometry studies demonstrated that all MM cell lines, MM patient plasma cells, and MM patient B cells expressed Muc-1 core protein epitopes. Circulating B cells from 4 normal donors also expressed Muc-1 core protein. In contrast, Muc-1 core protein was absent on 28 of 32 non-MM neoplastic cell lines, 17 of which expressed Muc-1. Splenic and tonsillar B cells, CD34(+) stem cells, resting T cells, and bone marrow plasma cells obtained from normal donors both lacked Muc-1 glycoforms. We next studied the effects of estrogen, progesterone, and glucocorticoid receptor agonists and antagonists on Muc-1 expression, because consensus sequences for the response elements of these steroids are present on the Muc-1 gene promoter. These studies showed that dexamethasone (Dex) induced Muc-1 expression on MM cell lines, as determined by both flow cytometry and Western blot analyses. Dex also induced upregulation of Muc-1 on prostate and ovarian cancer cell lines. Time and dose-response studies demonstrated that Dex induced maximal cell surface Muc-1 expression by 24 hours at concentrations of 10(-8) mol/L. Dex induced Muc-1 upregulation could be blocked with a 10-fold excess of the glucocorticoid receptor antagonist RU486, confirming that Dex was acting via the glucocorticoid receptor. No changes in Muc-1 expression were observed on MM cells treated with estrogen and progesterone receptor agonists and antagonists or with RU486. These studies provide the framework for targeting Muc-1 core protein in vaccination and serotherapy trials in MM. In addition, the finding that Muc-1 expression on MM cells can be augmented by Dex at pharmacologically achievable levels suggests their potential utility in enhancing treatments targeting Muc-1 in MM.