Different CD44 splicing patterns define prognostic subgroups in multiple myeloma

CD44 variant isoforms (CD44v) are generated by alternative splicing of the nuclear RNA resulting in the expression of additional protein domains in the extracellular region of the CD44 standard molecule (CD44s). In multiple myeloma (MM), CD44 mediates binding of tumor cells to stroma and regulates interleukin-6 production. To evaluate the role of CD44v isoforms in MM, CD44v expression was analyzed by immunohistochemical staining of 64 bone marrow biopsies from 38 MM patients. Expression of variant isoforms containing the 9v domain was observed in 36% of cases and was associated with an advanced stage (P ‼ .02; n = 61), a progressive disease (P ‼ .001; n = 61), and a shorter overall survival (P ‼ .02; n = 36). In contrast, 3v, 4v, 6v, or 10v isoforms were detected only in a small percentage of the patients. To analyze the exon composition in RNA-transcripts, reverse transcriptase polymerase chain reaction analyses followed by Southern hybridization with exon-specific probes were performed in fluorescence-activated cell sorted myeloma plasma cells. Tumors expressing the 9v domain showed complex, 9v-containing transcripts in combinations with the 3v, 7v, 8v, and 10v exons. Identical transcripts were detected in several myeloma cell lines and in a Ki-1 B-immunoblastic lymphoma. Similar to high-grade non-Hodgkin's lymphoma and gastric and renal cell carcinoma, overexpression of 9v-containing isoforms in MM is related to an unfavorable clinical presentation and represents a new prognostic parameter.

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CD28-Mediated Regulation of Multiple Myeloma Cell Proliferation and Survival.

Blood ◽

10.1182/blood.v106.11.355.355 ◽

2005 ◽

Vol 106 (11) ◽

pp. 355-355

Author(s):

Kelvin P. Lee ◽

Nizar J. Bahlis ◽

Anne M. King ◽

Despina Kolonias ◽

Louise M. Carlson ◽

...

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

Cell Proliferation ◽

T Cells ◽

Cell Survival ◽

Plasma Cells ◽

Survival Function ◽

Myeloma Cell ◽

Serum Starvation ◽

Bone Marrow Biopsies

Abstract Although interactions with bone marrow stromal cells are essential for multiple myeloma (MM) cell survival, the specific molecular and cellular elements involved are largely unknown due to the complexity of the bone marrow microenvironment. The CD28 receptor, which costimulates survival signals in T cells, is also expressed on normal plasma cells and myeloma cells. In MM, CD28 expression correlates significantly with disease progression, also suggesting a pro-survival function. In contrast to T cells however, activation and function of CD28 in myeloma and plasma cells is almost entirely undefined. We found that direct activation of myeloma cell CD28 by anti-CD28 mAb alone induced activation of NFkappaB, suppressed MM cell proliferation and protected against serum starvation and dexamethasone-induced cell death. We hypothesized that the specific CD80/CD86 expressing stromal cell partner of this interaction is a professional antigen presenting cells, in particular dendritic cells. Histological studies demonstrated DC were extensively interdigitated throughout the myeloma infiltrates in patient bone marrow biopsies. In vitro coculture with DC also elicited CD28-mediated effects on MM survival and proliferation, and could be blocked by CD28Ig. Our findings suggest a previously undescribed myeloma:DC cell-cell interaction involving CD28 that may play an important role in myeloma cell survival within the bone marrow stroma. These data also suggest that CD28 may represent a therapeutic target in the treatment of multiple myeloma.

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Ruxolitinib Regulates the Autophagy Machinery in Multiple Myeloma Cells

Anti-Cancer Agents in Medicinal Chemistry ◽

10.2174/1871520620666200218105159 ◽

2020 ◽

Vol 20 (18) ◽

pp. 2316-2323 ◽

Cited By ~ 3

Author(s):

Alican Kusoglu ◽

Bakiye G. Bagca ◽

Neslihan P.O. Ay ◽

Guray Saydam ◽

Cigir B. Avci

Keyword(s):

Multiple Myeloma ◽

Cell Line ◽

B Lymphocyte ◽

Plasma Cells ◽

Annexin V ◽

Myeloma Cell ◽

Control Group ◽

Multiple Myeloma Cell ◽

Ic50 Values ◽

Stat Pathway

Background: Ruxolitinib is a selective JAK1/2 inhibitor approved by the FDA for myelofibrosis in 2014 and nowadays, comprehensive investigations on the potential of the agent as a targeted therapy for haematological malignancies are on the rise. In multiple myeloma which is a cancer of plasma cells, the Interleukin- 6/JAK/STAT pathway is emerging as a therapeutic target since the overactivation of the pathway is associated with poor prognosis. Objective: In this study, our purpose was to discover the potential anticancer effects of ruxolitinib in ARH-77 multiple myeloma cell line compared to NCI-BL 2171 human healthy B lymphocyte cell line. Methods: Cytotoxic effects of ruxolitinib in ARH-77 and NCI-BL 2171 cells were determined via WST-1 assay. The autophagy mechanism induced by ruxolitinib measured by detecting autophagosome formation was investigated. Apoptotic effects of ruxolitinib were analyzed with Annexin V-FITC Detection Kit and flow cytometry. We performed RT-qPCR to demonstrate the expression changes of the genes in the IL-6/JAK/STAT pathway in ARH-77 and NCI-BL 2171 cells treated with ruxolitinib. Results: We identified the IC50 values of ruxolitinib for ARH-77 and NCI-BL 2171 as 20.03 and 33.9μM at the 72nd hour, respectively. We showed that ruxolitinib induced autophagosome accumulation by 3.45 and 1.70 folds in ARH-77 and NCI-BL 2171 cells compared to the control group, respectively. Treatment with ruxolitinib decreased the expressions of IL-6, IL-18, JAK2, TYK2, and AKT genes, which play significant roles in MM pathogenesis. Conclusion: All in all, ruxolitinib is a promising agent for the regulation of the IL-6/JAK/STAT pathway and interferes with the autophagy mechanism in MM.

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Loss of Spleen Visualization on Whole-Body Diffusion-Weighted Imaging in Patients with Multiple Myeloma Is Associated with Poor Prognosis and High Tumor Burden

Blood ◽

10.1182/blood-2021-147250 ◽

2021 ◽

Vol 138 (Supplement 1) ◽

pp. 3797-3797

Author(s):

Toshiki Terao ◽

Youichi Machida ◽

Ukihide Tateishi ◽

Takafumi Tsushima ◽

Kentaro Narita ◽

...

Keyword(s):

Multiple Myeloma ◽

Diffusion Weighted Imaging ◽

Poor Prognosis ◽

Tumor Volume ◽

Plasma Cells ◽

Myeloma Cell ◽

Stage Iii ◽

Whole Body ◽

Cell Infiltration ◽

Group A

Abstract Introduction Multiple myeloma (MM) is caused by the proliferation of monoclonal malignant plasma cells in the bone marrow (BM). Imaging has played a major role in visualizing myeloma lesions, assessing tumor volume, and predicting the prognosis. Recently, we reported that the total diffusion volume (tDV), assessed using a pretreatment whole-body diffusion-weighted imaging (WB-DWI), was associated with a high BM plasma cells (BMPCs) and a poor prognosis in patients with MM (Terao. et al., Eur Radiol 2021). During that study, we unexpectedly found the frequent absence of a spleen signal in patients with MM and its reappearance after treatment. Therefore, this study aimed to investigate the association between spleen visualization changes on WB-DWI and myeloma tumor load and prognosis in patients with MM. Methods The data of 96 consecutive patients with symptomatic newly-diagnosed MM (NDMM) at Kameda Medical Center from January 2016 to December 2020, 15 consecutive patients with smoldering MM (sMM), and two autopsied spleens of patients with PC dysplasia were retrospectively reviewed. All patients underwent at least one WB-DWI prior to treatment. The detail of WB-DWI was previously reported (Terao. et al., Eur Radiol 2021). "Loss of spleen visualization" (LSV) was defined as a visual loss of the spleen in maximum intensity projection on the WB-DWI (Fig1). The spleen-to-spinal cord (SC) ratio (SSR) was used in each regions-of-interest (ROI) to compare the signal intensity. The spleen ROIs were defined as non-overlapping ROIs of 30-50 pixels. The SC ROI was the largest ROI without overhanging in the image depicting the maximum size of the spleen. This study was approved by the institutional review board and conducted in accordance with the Declaration of Helsinki. All participants provided informed consent. Results The median patient age was 75.5 years and 81 patients (84.4%) were 65 years or older. Almost all patients (n=91) received proteasome inhibitors (PIs) as remission induction therapy and 33 patients received autologous stem-cell transplantation (ASCT). LSV was observed on the WB-DWI of 56/96 (58.3%) patients with NDMM and in one patient with sMM (1/15, 6.7%). Patients with NDMM and LSV had a higher median BMPC infiltration as assessed by CD138-immunohistochemistry (80.0% vs. 50.0%, p<0.001), a higher median tDV (540.2 mL vs. 137.0 mL, p=0.003), higher rate of ISS stage III (p<0.01), a lower SSR (0.36 vs. 0.96; p<0.001), and lower tDV (540.2 mL vs. 137.0 mL; p=0.003) than those without LSV. The three-year PFS (p=0.27) and three-year OS (p=0.021) were lower in patients with NDMM with LSV (PFS: 51.2% and OS: 72.5%) than in patients without LSV (PFS: 63.4% and OS: 100%). Next, we investigated the spleen signal change of patients who underwent WB-DWI twice or more during treatment (n=74). Of 42 out of the 74 patients with LSV at diagnosis, the spleen during treatment became visible on 31/42 (73.8%) patients. Representative patients with various spleen signal changes during treatment are shown in Figure 3 as group A (n=32; patients without LSV at diagnosis and during treatment), group B (n=31; patients who had LSV at diagnosis but the spleen reappeared after treatment), and group C (n=11; patients who had LSV at diagnosis, and despite treatment response, did not regain the spleen signal). Patients in group C showed significantly worse three-year PFS and OS (not available due to early events) than those in group A and B, even after excluding patients who did not achieve partial response or worse (n=11) (Fig1). In the multivariate analysis, the group C retained its prognostic significance for both PFS (hazard ratio [HR], 1.98, 95% confidence interval [CI] 1.00-3.90, p = 0.049) and OS (HR, 5.16, 95% CI 1.27-21.0, p = 0.022) even after adjustment for age over 70 years and the revised-ISS stage III. At last, to investigate the pathological cause of LSV, we reviewed two patients who underwent autopsies, who had both received WB-DWI within 3 months before their deaths (Fig1). One patient showed diffuse myeloma cell infiltration in the spleen and the other showed the amyloid deposition without myeloma cell infiltration. Conclusion This study showed that LSV and a low SSR on pretreatment WB-DWI are correlated with a high tumor volume and poor prognosis. As patients with LSV during treatment had very poor prognosis, the relationships between LSV and other variables should be investigated. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

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Targeting the methyltransferase SETD8 impairs tumor cell survival and overcomes drug resistance independently of p53 status in multiple myeloma

Clinical Epigenetics ◽

10.1186/s13148-021-01160-z ◽

2021 ◽

Vol 13 (1) ◽

Author(s):

Laurie Herviou ◽

Sara Ovejero ◽

Fanny Izard ◽

Ouissem Karmous-Gadacha ◽

Claire Gourzones ◽

...

Keyword(s):

Multiple Myeloma ◽

Plasma Cells ◽

Myeloma Cell ◽

Histone H4 ◽

Myeloma Cells ◽

Functional Studies ◽

Replicative Stress ◽

Poor Outcome ◽

Subsequent Decrease ◽

Lysine Methyltransferase

Abstract Background Multiple myeloma (MM) is a malignancy of plasma cells that largely remains incurable. The search for new therapeutic targets is therefore essential. In addition to a wide panel of genetic mutations, epigenetic alterations also appear as important players in the development of this cancer, thereby offering the possibility to reveal novel approaches and targets for effective therapeutic intervention. Results Here, we show that a higher expression of the lysine methyltransferase SETD8, which is responsible for the mono-methylation of histone H4 at lysine 20, is an adverse prognosis factor associated with a poor outcome in two cohorts of newly diagnosed patients. Primary malignant plasma cells are particularly addicted to the activity of this epigenetic enzyme. Indeed, the inhibition of SETD8 by the chemical compound UNC-0379 and the subsequent decrease in histone H4 methylation at lysine 20 are highly toxic in MM cells compared to normal cells from the bone marrow microenvironment. At the molecular level, RNA sequencing and functional studies revealed that SETD8 inhibition induces a mature non-proliferating plasma cell signature and, as observed in other cancers, triggers an activation of the tumor suppressor p53, which together cause an impairment of myeloma cell proliferation and survival. However, a deadly level of replicative stress was also observed in p53-deficient myeloma cells treated with UNC-0379, indicating that the cytotoxicity associated with SETD8 inhibition is not necessarily dependent on p53 activation. Consistent with this, UNC-0379 triggers a p53-independent nucleolar stress characterized by nucleolin delocalization and reduction of nucleolar RNA synthesis. Finally, we showed that SETD8 inhibition is strongly synergistic with melphalan and may overcome resistance to this alkylating agent widely used in MM treatment. Conclusions Altogether, our data indicate that the up-regulation of the epigenetic enzyme SETD8 is associated with a poor outcome and the deregulation of major signaling pathways in MM. Moreover, we provide evidences that myeloma cells are dependent on SETD8 activity and its pharmacological inhibition synergizes with melphalan, which could be beneficial to improve MM treatment in high-risk patients whatever their status for p53.

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Expression of functional interleukin-15 receptor and autocrine production of interleukin-15 as mechanisms of tumor propagation in multiple myeloma

Blood ◽

10.1182/blood.v95.2.610 ◽

2000 ◽

Vol 95 (2) ◽

pp. 610-618 ◽

Cited By ~ 87

Author(s):

Inge Tinhofer ◽

Ingrid Marschitz ◽

Traudl Henn ◽

Alexander Egle ◽

Richard Greil

Keyword(s):

Multiple Myeloma ◽

Cell Survival ◽

Cell Lines ◽

Plasma Cells ◽

Constitutive Expression ◽

Myeloma Cell ◽

Myeloma Cells ◽

Cytotoxic Treatment ◽

Interleukin 15 ◽

Induced Apoptosis

Interleukin-15 (IL-15) induces proliferation and promotes cell survival of human T and B lymphocytes, natural killer cells, and neutrophils. Here we report the constitutive expression of a functional IL-15 receptor (IL-15R) in 6 of 6 myeloma cell lines and in CD38high/CD45low plasma cells belonging to 14 of 14 patients with multiple myeloma. Furthermore, we detected IL-15 transcripts in all 6 myeloma cell lines, and IL-15 protein in 4/6 cell lines and also in the primary plasma cells of 8/14 multiple myeloma patients. Our observations confirm the existence of an autocrine IL-15 loop and point to the potential paracrine stimulation of myeloma cells by IL-15 released from the cellular microenvironment. Blocking autocrine IL-15 in cell lines increased the rate of spontaneous apoptosis, and the degree of this effect was comparable to the pro-apoptotic effect of depleting autocrine IL-6 by antibody targeting. IL-15 was also capable of substituting for autocrine IL-6 in order to promote cell survival and vice versa. In short-term cultures of primary myeloma cells, the addition of IL-15 reduced the percentage of tumor cells spontaneously undergoing apoptosis. Furthermore, IL-15 lowered the responsiveness to Fas-induced apoptosis and to cytotoxic treatment with vincristine and doxorubicin but not with dexamethasone. These data add IL-15 to the list of important factors promoting survival of multiple myeloma cells and demonstrate that it can be produced and be functionally active in an autocrine manner.

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THE BONE MARROW ON STERNAL ASPIRATION IN MULTIPLE MYELOMA

Blood ◽

10.1182/blood.v3.9.987.987 ◽

1948 ◽

Vol 3 (9) ◽

pp. 987-1018 ◽

Cited By ~ 52

Author(s):

EDWIN D. BAYRD

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

Plasma Cell ◽

Plasma Cells ◽

Myeloma Cell ◽

Multinucleated Cells ◽

Cytoplasmic Bodies ◽

Plasma Cell Type ◽

Well Differentiated ◽

Diagnostic Pitfalls

Abstract Generalizing, it can be said that the pathologic cells seen in smears of the bone marrow in multiple myeloma resemble the plasma cell and vary from the very anaplastic and immature cell to the well-differentiated and almost characteristic plasma cell. The feature which the "myeloma" cell shares with the plasma cell is the abundant, granular, basophilic cytoplasm which tends to be fragile and undergo the same degenerative changes in each; namely, the formation of Russell bodies and vacuolization. Fairly frequently a perinuclear clear area or Hof is present and the nucleus tends to be eccentrically placed. Cytoplasmic extensions or pseudopodia may also be seen in either case, but they occur more often and more dramatically in instances of multiple myeloma. Multinucleated cells are commonly seen. In addition, myeloma-plasma cells will often have a large clear nucleolus and a leptochromatic nucleus and will exhibit a tendency to the formation of isolated areas of condensed chromatin. Cytoplasmic extrusions, free cytoplasmic bodies, occasionally complete with Russell bodies and vacuoles are almost universally present. All cases were of the plasma cell type; there was no exception. In these cases, the myeloma-plasma cell constituted from 2.5 to 96 per cent of the leukocytic elements present. The opinion was expressed that all so-called types of multiple myeloma are merely variations in differentiation of this same cell. It was noted that anaplasia, hypernucleation and lack of plasma cell predominance in certain cases were diagnostic pitfalls. Additional evidence was adduced to confirm the reticulo-endothelial origin of the myeloma-plasma cell. It was further observed that certain prognostically valuable information could be gleaned from a careful review of the cytologic characteristics in these cases.

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CD28-mediated regulation of multiple myeloma cell proliferation and survival

Blood ◽

10.1182/blood-2006-03-012542 ◽

2007 ◽

Vol 109 (11) ◽

pp. 5002-5010 ◽

Cited By ~ 83

Author(s):

Nizar J. Bahlis ◽

Anne M. King ◽

Despina Kolonias ◽

Louise M. Carlson ◽

Hong Yu Liu ◽

...

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

Cell Proliferation ◽

Cell Survival ◽

Plasma Cells ◽

Cell Interaction ◽

Myeloma Cell ◽

Serum Starvation ◽

Costimulatory Receptor ◽

Multiple Myeloma Cell

Abstract Although interactions with bone marrow stromal cells are essential for multiple myeloma (MM) cell survival, the specific molecular and cellular elements involved are largely unknown, due in large part to the complexity of the bone marrow microenvironment itself. The T-cell costimulatory receptor CD28 is also expressed on normal and malignant plasma cells, and CD28 expression in MM correlates significantly with poor prognosis and disease progression. In contrast to T cells, activation and function of CD28 in myeloma cells is largely undefined. We have found that direct activation of myeloma cell CD28 by anti-CD28 mAb alone induces activation of PI3K and NFκB, suppresses MM cell proliferation, and protects against serum starvation and dexamethasone (dex)–induced cell death. Coculture with dendritic cells (DCs) expressing the CD28 ligands CD80 and CD86 also elicits CD28-mediated effects on MM survival and proliferation, and DCs appear to preferentially localize within myeloma infiltrates in primary patient samples. Our findings suggest a previously undescribed myeloma/DC cell-cell interaction involving CD28 that may play an important role in myeloma cell survival within the bone marrow stroma. These data also point to CD28 as a potential therapeutic target in the treatment of MM.

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Amphiregulin Is Produced by Myeloma Cells and Is Involved in Tumor-Environment Interactions in Multiple Myeloma.

Blood ◽

10.1182/blood.v104.11.4306.4306 ◽

2004 ◽

Vol 104 (11) ◽

pp. 4306-4306

Author(s):

Karène Mahtouk ◽

Dirk Hose ◽

Thierry Reme ◽

John De Vos ◽

Michel Jourdan ◽

...

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

Growth Factors ◽

Plasma Cells ◽

Myeloma Cell ◽

Erbb Receptors ◽

Heparin Binding ◽

Myeloma Cells ◽

Heparin Binding Growth Factors ◽

Egf Family

Abstract Multiple myeloma (MM) is characterized by the accumulation of clonal malignant plasma cells in the bone marrow. One of the hallmarks of plasma cells is the expression of the heparan-sulfate proteoglycan syndecan-1. In epithelial cells, syndecan-1 plays a major role as a coreceptor for heparin-binding growth factors and chemokines. This stresses that heparin-binding growth factors may play a major role in the biology of MM cells. Recently we have demonstrated that heparin-binding EGF-like growth factor (HB-EGF), one of the ten members of the Epidermal Growth Factor (EGF) family, is produced by the tumor microenvironment and is able to trigger myeloma cell growth. As amphiregulin (AREG) is another member of the EGF family that also binds heparan-sulphate chains, we investigated its role in MM. We looked for AREG expression on a panel of 7 normal plasmablastic cells (PPCs), 7 normal bone marrow plasma cells (BMPCs), purified MM cells from 65 patients and 20 myeloma cell lines (HMCLs), with Affymetrix U133A+B microarrays. We showed that primary MM cells overexpress AREG compared to normal BMPCs and PPCs. We then investigated the expression of the ErbB receptors with real-time RT-PCR. Myeloma cells variably expressed the 4 ErbB receptors. Normal BMPCs also expressed ErbB1 and ErbB2 unlike PPCs that did not express any ErbB receptors. We demonstrated that the high AREG expression by primary myeloma cells may have a dual effect. On the one hand, AREG stimulated IL-6 production and growth of bone-marrow stromal cells that highly express the AREG ErbB1 receptor. On the other hand, AREG could promote HMCL proliferation, suggesting that a functional autocrine loop involving AREG and ErbB receptors is involved in MM cell growth. Finally, we looked for the effect of ErbB inhibitors on MM cells of 14 patients cultured for 6 days together with their bone marrow environment. A pan-ErbB inhibitor (PD-169540, Pfizer) and an ErbB1-inhibitor (IRESSA, Astrazeneca) induced strong MM cell apoptosis in respectively 71% of patients (10 of 14) and 29% of patients (4 of 14). Of major interest, when PD169540 or IRESSA were combined with dexamethasone, they induced a dramatic myeloma cell death (respectively 92% and 69% inhibition of MM cell survival), while non-myeloma cells were unaffected. Thus ErbB activation is critical to trigger MM-cell survival in short-term culture. In conclusion, our findings provide evidence for a major role of AREG and HB-EGF in the biology of multiple myeloma and identify ErbB receptors as putative therapeutic targets. These data emphasize the interest of clinical evaluation of specific-ErbB-inhibitors in patients with MM, either used alone or in combination with dexamethasone.

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PTEN Gene Deletions in Patients with Multiple Myeloma.

Blood ◽

10.1182/blood.v104.11.4889.4889 ◽

2004 ◽

Vol 104 (11) ◽

pp. 4889-4889

Author(s):

Xiao Ying Qi ◽

A. Keith Stewart ◽

Hong Chang

Keyword(s):

Multiple Myeloma ◽

Plasma Cells ◽

Progression Free Survival ◽

Myeloma Cell ◽

Suppressor Gene ◽

High Dose Chemotherapy ◽

Pten Gene ◽

Allelic Loss ◽

High Dose ◽

13Q Deletion

Abstract PTEN, a tumor suppressor gene, negatively regulates the anti-apoptotic action of akt phosphorylation. Allelic loss or mutation of this gene has been detected in many solid tumors and more recently in human myeloma cell lines (HMCLs). Expression of PTEN has resulted in growth inhibition and apoptosis of a HMCL, suggesting that it may play a role in the pathogenesis of multiple myeloma (MM). However, the PTEN status in tumor cells from patients with MM has not been determined. Using a triple staining method combining staining for cytoplasmic light chains and fluorescence in situ hybridization (FISH) with chromosome 10-centromere and PTEN-gene specific probes, we analyzed clonal plasma cells from 71 patients with MM, 10 with plasma cell leukemia (PCL) and 10 HMCLs. Hemizygous PTEN deletions were detected in 4 of 71 (5.6%) MM patients, 2 of 10 (20%) PCLs, and 2 of 10 (20%) HMCLs. The percentages of clonal plasma cells containing PTEN deletions ranged from 21–90% (median, 56%). Three of the 4 patients with PTEN deletions were detected at diagnosis with stage III disease (Duire-Salmon) and 1 was detected at relapse. Two patients had IgG kappa, 1 IgG lambda and 1 free lambda light chain. To correlate the PTEN status with other known genetic abnormalities in MM, we investigated 4 MM and 2 PCLs with PTEN deletions using FISH for chromosome13q, p53 status, translocations t(11;14), t(4;14) and t(14;16). One MM had a 13q deletion, 1 PCL had a t(11;14), and the other PCL had a t(14;16), a 13q deletion and a p53 deletion. All 4 MM patients with hemizygous PTEN deletions received melphalan based high-dose chemotherapy and autologous stem cell support. Their median overall survival (OS) was 48.1months, and progression free survival (PFS) was 42.8 months as compared to patients without PTEN deletions (OS, not reached, PFS, 25.8 months) (p=0.51 for OS, p=0.67 for PFS). Our results indicate that PTEN deletions are uncommon in MM patients and therefore unlikely represent a primary event for MM. PTEN deletions appear to occur in the advance stage of the disease, and are more frequently involved in PCL or HMCLs suggesting that deletions of PTEN may be associated with disease progression in a subset of MM.

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Evaluation of NCAM and DKK1 Expression in Multiple Myeloma and MGUS by Gene and Tissue Microarrays: Expression Accompanies Progression.

Blood ◽

10.1182/blood.v108.11.3399.3399 ◽

2006 ◽

Vol 108 (11) ◽

pp. 3399-3399

Author(s):

Sushil Gupta ◽

Yongsheng Huang ◽

James Stewart ◽

Fenghuang Zhan ◽

Bart Barlogie ◽

...

Keyword(s):

Gene Expression ◽

Multiple Myeloma ◽

Plasma Cells ◽

Molecular Classification ◽

Tissue Microarrays ◽

Bone Lesions ◽

Bone Marrow Biopsies ◽

Dkk1 Expression ◽

Significant Difference ◽

Expression Microarrays

Abstract Introduction: Expression NCAM1, a cell adhesion molecule involved in neuron-neuron adhesion, is also expressed by multiple myeloma (MM PC). Osteolytic bone lesions are a hallmark of MM and elevated expression of NCAM in MM PC has been correlated with this process (Ely and Knowles, 2003). We have previously reported that MM PC express DKK1 and MM blocks osteoblast differentiation in a DKK1-specific manner, suggesting that secretion of the Wnt signaling inhibitor plays a role in MM bone disease. Herein we used gene expression microarrays and tissue microarrays (TMAs) to investigate the simultaneous expression of DKK1 and NCAM in MM and MGUS. Methods: The study population consisted of 198 newly diagnosed MM and 44 MGUS. RNA from CD138-selected plasma cells was hybridized to Affymetrix U133Plus microarrays and data processed with Affymetrix Microarray Suite GCOS1.1 software. TMAs were constructed from formalin-fixed, paraffin embedded bone marrow biopsies. Serial sections of TMA were immunostained for CD138, NCAM and DKK1. TMAs were scanned using ScanScope using 20x lens, assessed using TMA lab software (Aperio Technologies) and scored as an average number of cells in the context of CD138 staining. Results: When put in context of a recently described molecular classification (Zhan et al., 2006), NCAM and DKK1 were found significantly co-over-expressed (DKK1+/NCAM+) in HYPERDIPLOID MM (P < 0.01); NCAM+/DKK1− was typical for MMSET-spike MM (P<0.001); NCAM−/DKK1+ DKK1 was characteristic of MM in MAF-spike disease (P < 0.001), CCND1 spikes with co-expression of CD20 (P<0.01), and the so-called MYELOID subgroup (P<0.001). In contrast, virtually all cases of MGUS were DKK1−/NCAM-. On TMAs, DKK1 expression varied in CD138-positive cells in MM, and in osteocytes and megakaryocytes in MM and MGUS, but was clearly negative in osteoblasts/lining cells. NCAM expression, was also variably expressed in PC of MM cases and in virtually 100% of osteoblasts/lining cells in both MM and MGUS. Osteocytes were distinctly negative for NCAM in both diseases. Of 195 analyzable MM biopsies, PC were DKK1+ in 1% to > 90% of PC in 191 (98%); of these, 94 were also NCAM+ in 5% to > 90% of PC; 1 case was positive for NCAM+/DKK− and 3 were NCAM−/DKK−. There was no significant difference in clinical parameters and survival in a comparison of NCAM+/DKK+ and NCAM−/DKK+ groups. DKK1 gene expression was higher in DKK1+/NCAM+ than in DKK1+/NCAM− MM (P=0.006); NCAM gene expression was higher in DKK1+/NCAM+ than in DKK+/NCAM− MM (P<0.0001) and NCAM was the number one SAM-defined gene over-expressed in DKK1+/NCAM+ relative to DKK1+/NCAM− disease. Consistent with GEP data, DKK1+/NCAM+ MM was over-represented in HYPERDIPLOID MM (32% v. 10%; P<0.001), while DKK1+/NCAM− disease was overrepresented in MAF-spike (15% v. 2%; P=0.003) and MYELOID subgroups (32% v. 18%; P=0.03). DKK1+/NCAM+ MM was completely absent in CCND1-spike /CD20-negative disease, with only 4% of DKK1+/NCAM− cases in this subgroup. No difference was observed for the other subtypes. Conclusion: DKK1 is expressed in all cases of MM with half also expressing NCAM. Neither gene is expressed in MGUS PC. These data suggests that expression of these genes in plasma cells accompanies disease progression.

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