Targeting the UPR-Transcription Factor XBP1 to Overcome Drug-Resistance in Ph+ ALL

Abstract Abstract 872 Background: The unfolded protein response (UPR) is a cellular machinery required to salvage of ER stress and to promote cell survival. The pathway consists of three different components, namely inositol-requiring enzyme 1a (IRE-1), PKR-like ER kinase (PERK) and activating transcription factor 6 (ATF6) and converges at the level of its effector molecule X-box binding protein 1 (XBP1). Previous work identified Xbp1 as a central requirement of plasma cell development and as critical mediator of cell survival in plasma cell-derived multiple myeloma. RESULTS: While the role of Xbp1 in plasma cells and plasma cell malignancies is well established, we report here the unexpected finding of a central role of Xbp1 in the survival of pre-B cell-derived Ph+ ALL cells. Surprisingly, patient-derived Ph+ ALL cells express Xbp1 (and related molecules in the IRE1 pathway) at significantly higher levels than normal bone marrow pre-B cells. In addition, we found that high expression levels of Xbp1 at diagnosis predict poor poor overall survival (OS), relapse-free survival (RFS) of leukemia patients in two clinical trials for patients with high risk acute lymphoblastic leukemia (n=207; COG P9906 trial; p=1.12e-4 and ECOG E2993; n=215; p=2.48e-5). In addition, high levels of XBP1 correlated with positive minimal residual disease (MRD) status at day 29 after onset of chemotherapy. To study the function of Xbp1 in Ph+ ALL in a genetic experiment, we developed a Ph+ ALL leukemia model based on bone marrow progenitor cells from mice carring loxP-flanked allele of Xbp1 (Xbp1fl/fl). On the basis of this model, bone marrow B cell precursors were transformed by BCR-ABL1 in the presence of IL7. Inducible Cre-mediated deletion of Xbp1 was achieved by transduction of leukemia cells with tamoxifen (4-OHT)-inducible Cre. Interestingly, 4-OHT-induced deletion of Xbp1 in Ph+ ALL-like leukemia cells caused rapid cell death within two days of induction. Xbp1-deletion resulted in extensive apoptosis, cellular senescence and cell cycle arrest owing to increased levels of p53, p21 and Arf. Interestingly, similar observations were made in an in vivo setting where Xbp1-deletion resulted in prolonged survival of NOD-SCID transplant recipient mice (n=7; p=0.007). Mechanistically, deletion of Xbp1 leads to increased expression of the pro-apoptotic molecule CHOP as in plasma cells/multiple myeloma and phosphorylation of the stress MAP kinases p38 and JNK. CLINICAL RELEVANCE: To test the potential clinical relevance of these findings, we used a recently identified small-molecule inhibitor STF-083010 (Papandreou et al., 2011), which blocks the endonuclease activity of upstream molecule IRE-1, essential for the splicing of the active form of Xbp1. STF-083010 indeed inhibited splicing of XBP1 and overall mimicked findings in genetic experiments. Importantly, targeting of Xbp1 by STF-083010 also induced cell death in three patient-derived cases of Ph+ ALL carrying the T315I mutations, which confers far-reaching TKI-resistance. CONCLUSIONS: These findings identify Xbp1 as a fundamentally novel target for the therapy of TKI-resistant Ph+ ALL. Like plasma cells and tumor cells in multiple myeloma, Ph+ ALL cells are selectively sensitive to ER stress and critically dependent on Xbp1 and likely other factors of the UPR pathway. Clinical validation of this concept could lead to improved treatment options for patients with TKI-resistant Ph+ ALL. Disclosures: No relevant conflicts of interest to declare.

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Myeloma Cell Associated Therapeutic Protein Discovery Using Single Cell RNA-Seq Data

Blood ◽

10.1182/blood-2020-142205 ◽

2020 ◽

Vol 136 (Supplement 1) ◽

pp. 4-5

Author(s):

Lijun Yao ◽

Reyka G Jayasinghe ◽

Tianjiao Wang ◽

Julie O'Neal ◽

Ruiyang Liu ◽

...

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

Cell Surface ◽

B Cell ◽

Tumor Cells ◽

Plasma Cell ◽

Tissue Specificity ◽

Plasma Cells ◽

Expression Data ◽

Intracellular Proteins

Multiple myeloma (MM) is a hematological cancer of the antibody-secreting plasma cells. Despite therapeutic advancements, MM remains incurable due to high incidence of drug-resistant relapse. In recent years, targeted immunotherapies, which take advantage of the immune system's cytotoxic defenses to specifically eliminate tumor cells expressing certain cell surface and intracellular proteins have shown promise in combating this and other B cell hematologic malignancies. A major limitation in the development of these therapies lies in the discovery of optimal candidate targets, which require both high expression in tumor cells as well as stringent tissue specificity. In an effort to identify potential myeloma-specific target antigens, we performed an unbiased search for genes with specific expression in plasma and/or B cells using single-cell RNA-sequencing (scRNAseq) of 53 bone marrow samples taken from 42 patients. By comparing >40K plasma cells to >97K immune cells across our cohort, we were able to identify a total of 181 plasma cell-associated genes, including 65 that encode cell-surface proteins and 116 encoding intracellular proteins. Of particular interest is that the plasma cells from each patient were shown to be transcriptionally distinct with unique sets of genes expressed defining each patient's malignant plasma cells. Using pathway enrichment analysis, we found significant overrepresentation of cellular processes related to B-Cell receptor (BCR) signaling, protein transport, and endoplasmic reticulum (ER) stress, involving genes such as DERL3, HERPUD1, PDIA4, PDIA6, RRBP1, SSR3, SSR4, TXNDC5, and UBE2J1. To note, our strategy successfully captured several of the most promising MM therapeutic targets currently under pre-clinical and clinical trials, including TNFRSF17(BCMA), SLAMF7, and SDC1 (CD138). Among these, TNFRSF17 showed very high plasma cell expression, with concomitant sharp exclusion of other immune cell types. To ascertain tissue specificity of candidate genes outside of the bone marrow, we analyzed gene and protein expression data from the Genotype-Tissue Expression (GTEx) portal and Human Protein Atlas (HPA). We found further support for several candidates (incl. TNFRSF17,SLAMF7, TNFRSF13B (TACI), and TNFRSF13C) as being both exclusively and highly expressed in lymphoid tissues. While several surface candidates were not found to be lymphocyte-restricted at the protein level, they remain relevant considerations as secondary targets for bi-specific immunotherapy approaches currently under development. To further investigate potential combinatorial targeting, we examine sample-level patterns of candidate co-expression and mutually-exclusive expression using correlation analysis. As the majority of our detected plasma cell-specific genes encode intracellular proteins, we investigated the potential utility of these epitopes as therapeutic targets via MHC presentation. Highly expressed candidates include MZB1, SEC11C, HLA-DOB, POU2AF1, and EAF2. We analyzed protein sequences using NetMHC and NETMHCII to predict high-affinity peptides for common class-I and class-II HLA alleles. To correlate MHC allelic preference with candidate expression in our cohort, we performed HLA-typing for 29 samples using Optitype. To support our scRNAseq-driven findings, we cross-referenced gene expression data with 907 bulk RNA-sequencing samples, including 15 from internal studies and 892 from the Multiple Myeloma Research Foundation (MMRF), as well as bulk global proteomics data from 4 MM cell lines (TIB.U266, RPMI8226, OPM2, MM1ST) and 4 patients. We see consistent trends across both cohorts, with high positive correlation (Pearson R ranging between 0.60 and 0.99) for a majority of genes when comparing scRNA and bulk RNA expression in the same samples. Our experimental design and analysis strategies enabled the efficient discovery of myeloma-associated therapeutic target candidates. In conclusion, this study identified a set of promising myeloma CAR-T targets, providing novel treatment options for myeloma patients. Disclosures Goldsmith: Wugen Inc.: Consultancy. DiPersio:Magenta Therapeutics: Membership on an entity's Board of Directors or advisory committees.

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A Role for Syntenin-1 in Multiple Myeloma Cell Survival

Blood ◽

10.1182/blood-2018-99-113594 ◽

2018 ◽

Vol 132 (Supplement 1) ◽

pp. 1008-1008

Author(s):

Tyler Moser-Katz ◽

Catherine M. Gavile ◽

Benjamin G Barwick ◽

Sagar Lonial ◽

Lawrence H. Boise

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

Cell Death ◽

Cell Survival ◽

Cell Lines ◽

Plasma Cells ◽

Surface Expression ◽

Myeloma Cell ◽

Myeloma Cells ◽

Rpmi 8226

Abstract Multiple myeloma is the second most common hematological malignancy in the U.S. with an estimated 30,700 new diagnoses in 2018. It is a clonal disease of plasma cells that, despite recent therapeutic advances, remains incurable. Myeloma cells retain numerous characteristics of normal plasma cells including reliance on survival signals in the bone marrow for long term viability. However, malignant transformation of plasma cells imparts the ability to proliferate, causing harmful bone lesions in patients, and in advanced stages independence of the bone-marrow microenvironment. Therefore, we are investigating the molecular mechanisms of myeloma cell survival that allow them to become extramedullary. We identified syntenin-1 (SDCBP) as a protein involved in myeloma cell survival and a potential therapeutic target. Syntenin-1 is an adapter protein that has been shown to regulate surface expression of several transmembrane proteins by binding with membrane phospholipids and mediating vesicular trafficking of proteins throughout the cell. Syntenin-1 regulates the surface expression of CD138, a plasma/myeloma cell marker. Syntenin-1 has been shown to regulate apoptosis in numerous cancer cell lines including breast cancer, glioma, and pancreatic cancer but its role in multiple myeloma survival has not been studied. To determine if syntenin-1 expression has an effect on myeloma cell survival, we utilized the CoMMpass dataset (IA12), a longitudinal study of myeloma patients that includes transcriptomic analysis throughout treatment. We found that patients with the highest expression of syntenin-1 mRNA (top quartile) had significantly worse overall survival, progression-free survival, and a shorter response duration than those in the bottom quartile of expression. To determine if syntenin-1 has a role in myeloma cell survival, we used short hairpin RNA to knock down syntenin-1 (shsyn) in RPMI 8226 and MM1.s myeloma cell lines. We then determined the amount of cell death using Annexin-V staining flow cytometry four days following lentiviral infection. We found increased cell death in syntenin-1-silenced cells compared to our empty vector control in both RPMI 8226 (control=42.17%, shsyn=71.53%, p=0.04) and MM1.s cell lines (control=8.57%, shsyn=29.9%, p=0.04) suggesting that syntenin-1 is important for myeloma cell survival. Syntenin-1 contains two PDZ domains that allow it to bind to receptor proteins via their corresponding PDZ-binding motifs. We therefore wanted to look at correlation of syntenin-1 expression with CD138 and CD86, two PDZ-binding domain containing proteins expressed on the surface of myeloma cells. Using the CoMMpass dataset, we found patients with high expression of syntenin-1 had a median expression of CD86 that was twice as high as the total population (P<0.0001) while syntenin-1-low patients expressed CD86 at levels that were half as much as the population (P<0.0001). In contrast, there was no clear relationship between syntenin-1 and CD138 mRNA expression. Indeed if one takes into account all patients, there is a positive correlation between CD86 and syntenin-1 expression (r=0.228, P<0.0001) while there is a negative correlation between CD138 and syntenin-1 (r=-0.1923, P<0.0001). The correlation with CD86 but not CD138 suggests a previously undescribed role for syntenin-1 in myeloma cells. Our lab has previously shown that expression of CD86 is necessary for myeloma cell survival, and signals via its cytoplasmic domain to confer drug resistance. Silencing syntenin-1 results in a decrease in CD86 surface expression. However, there is no change in CD86 transcript or total cellular CD86 protein levels in our shsyn treated cells. Moreover, knockdown of CD86 resulted in increased protein expression and transcript levels of syntenin-1. Taken together, these data suggest that syntenin-1 may regulate CD86 expression on the cell surface. Our data supports a novel role for syntenin-1 in myeloma cell viability and as a potential regulator of CD86 surface expression. The role of syntenin-1 has not previously been explored in multiple myeloma and determining its molecular function is warranted as it may be an attractive target for therapeutic treatment of the disease. Disclosures Lonial: Amgen: Research Funding. Boise:AstraZeneca: Honoraria; Abbvie: Consultancy.

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A Novel Role For Histone Deacetylase 11 (HDAC11) In Plasma Cell Differentation and Survival

Blood ◽

10.1182/blood.v122.21.1907.1907 ◽

2013 ◽

Vol 122 (21) ◽

pp. 1907-1907

Author(s):

Eva Sahakian ◽

Jason B. Brayer ◽

John Powers ◽

Mark Meads ◽

Allison Distler ◽

...

Keyword(s):

Gene Expression ◽

Multiple Myeloma ◽

Bone Marrow ◽

B Cells ◽

Plasma Cell ◽

Research Funding ◽

Plasma Cells ◽

Myeloma Cells ◽

Peripheral Blood Plasma

Abstract The role of HDACs in cellular biology, initially limited to their effects upon histones, is now appreciated to encompass more complex regulatory functions that are dependent on their tissue expression, cellular compartment distribution, and the stage of cellular differentiation. Recently, our group has demonstrated that the newest member of the HDAC family of enzymes, HDAC11, is an important regulator of IL-10 gene expression in myeloid cells (Villagra A Nat Immunol. 2009). The role of this specific HDAC in B-cell development and differentiation is however unknown. To answer this question, we have utilized a HDAC11 promoter-driven eGFP reporter transgenic mice (TgHDAC11-eGFP) which allows the monitoring of the dynamic changes in HDAC11 gene expression/promoter activity in B-cells at different maturation stages (Heinz, N Nat. Rev. Neuroscience 2001). First, common lymphoid progenitors are devoid of HDAC11 transcriptional activation as indicated by eGFP expression. In the bone marrow, expression of eGFP moderately increases in Pro-B-cells and transitions to the Pre- and Immature B-cells respectively. Expression of eGFP doubles in the B-1 stage of differentiation in the periphery. Of note, examination of both the bone marrow and peripheral blood plasma cell compartment demonstrated increased expression of eGFP/HDAC11 mRNA at the steady-state. These results were confirmed in plasma cells isolated from normal human subjects in which HDAC11 mRNA expression was demonstrated. Strikingly, analysis of primary human multiple myeloma cells demonstrated a significantly higher HDAC11 mRNA expression in malignant cells as compared to normal plasma cells. Similar results were observed in 4/5 myeloma cell lines suggesting that perhaps HDAC11 expression might provide survival advantage to malignant plasma cells. Support to this hypothesis was further provided by studies in HDAC11KO mice in which we observed a 50% decrease in plasma cells in both the bone marrow and peripheral blood plasma cell compartments relative to wild-type mice. Taken together, we have unveiled a previously unknown role for HDAC11 in plasma cell differentiation and survival. The additional demonstration that HDAC11 is overexpressed in primary human myeloma cells provide the framework for specifically targeting this HDAC in multiple myeloma. Disclosures: Alsina: Millennium: Membership on an entity’s Board of Directors or advisory committees, Research Funding. Baz:Celgene Corporation: Research Funding; Millenium: Research Funding; Bristol Myers Squibb: Research Funding; Novartis: Research Funding; Karyopharm: Research Funding; Sanofi: Research Funding.

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An In Vivo Model To Investigate the Identity of the Cell-of-Origin of Multiple Myeloma.

Blood ◽

10.1182/blood.v106.11.1534.1534 ◽

2005 ◽

Vol 106 (11) ◽

pp. 1534-1534

Author(s):

Fotios A. Asimakopoulos ◽

Harold E. Varmus

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

Transcription Factor ◽

Plasma Cell ◽

Germinal Center ◽

Plasma Cells ◽

Lymphoid Tissues ◽

Cell Of Origin ◽

Dark Zone ◽

Bone Marrow Plasma

Abstract Multiple myeloma (MM) is characterized by monoclonal expansion of bone marrow plasma cells. However, long-lived plasma cells resident in the marrow are terminally differentiated and possess a limited replicative lifespan; it is puzzling how they could be the source of aggressive and relapsing neoplasms. We postulate that the myeloma clonogenic progenitor may reside in a more immature compartment with greater self-renewal capacity, most probably a cell participating in, or having shortly exited the germinal center reaction. However, it is unclear whether critical mutations occur in the target cell prior to, or following commitment to the plasma cell fate. To investigate the nature of the MM cell-of-origin, we have created a novel flexible mouse model system that enables the delivery of stochastic, sequential, somatic mutations to precisely defined compartments of the germinal center in secondary lymphoid tissues. To this end, we have used BAC transgenic technology to express distinct types of avian leukosis virus (ALV) receptors, TVA and TVB, in the expanding centroblast of the dark zone and the committed plasmablast of the light zone, respectively. Mammalian tissues are refractory to transduction by retroviruses of the ALV family unless they ectopically express the cognate avian-derived receptors. Thus, the coding sequences for the TVA receptor, fused to a fluorescent protein tag were placed under the control of transcription factor A-myb, expressed in centroblasts of the dark zone. Similarly, sequences encoding a fluorescent-tagged TVB receptor were placed under the control of transcription factor Blimp1, expressed in the earliest committed plasmablasts as well as mature plasma cells. Analysis of the Blimp1: TVB mice showed that expression of the avian retroviral receptor in the hematopoietic system is limited to the light zone of germinal centers, extrafollicular collections of CD138+ cells in the spleen and lymph nodes as well as long-lived bone marrow plasma cells. Analysis of A-myb: TVA transgenic mice is currently underway. The system permits the introduction of a variety of molecular lesions to specific plasma cell precursors via retroviral transduction of oncogenes, shRNAs against tumor suppressor genes or inducible regulators of gene expression in an attempt to re-create the sequence of molecular lesions leading to MM in the relevant cellular context.

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Evidence for a bone marrow B cell transcribing malignant plasma cell VDJ joined to C mu sequence in immunoglobulin (IgG)- and IgA-secreting multiple myelomas.

Journal of Experimental Medicine ◽

10.1084/jem.178.3.1091 ◽

1993 ◽

Vol 178 (3) ◽

pp. 1091-1096 ◽

Cited By ~ 80

Author(s):

P Corradini ◽

M Boccadoro ◽

C Voena ◽

A Pileri

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

B Cells ◽

B Cell ◽

B Lymphocytes ◽

Plasma Cell ◽

Plasma Cells ◽

Bone Marrow Cells ◽

Indirect Evidence ◽

Specific Marker

Multiple myeloma is a B cell malignancy characterized by the expansion of plasma cells producing monoclonal immunoglobulins (Ig). It has been regarded as a tumor arising at the B, pre-B lymphocyte, or even stem cell level. Precursor cells are presumed to proliferate and differentiate giving rise to the plasma cell clonal expansion. Antigenic features and specific Ig gene rearrangement shared by B lymphocytes and myeloma cells have supported this hypothesis. However, the existence of such a precursor is based upon indirect evidence and is still an open question. During differentiation, B cells rearrange variable (V) regions of Ig heavy chain genes, providing a specific marker of clonality. Using an anchor polymerase chain reaction assay, these rearranged regions from five patients with multiple myeloma were cloned and sequenced. The switch of the Ig constant (C) region was used to define the B cell differentiation stage: V regions are linked to C mu genes in pre-B and B lymphocytes (pre-switch B cells), but to C gamma or C alpha in post-switch B lymphocytes and plasma cells (post-switch B cells). Analysis of bone marrow cells at diagnosis revealed the presence of pre-switch B cells bearing plasma cell V regions still joined to the C mu gene. These cells were not identified in peripheral blood, where tumor post-switch B cells were detected. These pre-switch B cells may be regarded as potential myeloma cell precursors.

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A novel membrane antigen selectively expressed on terminally differentiated human B cells

Blood ◽

10.1182/blood.v84.6.1922.1922 ◽

1994 ◽

Vol 84 (6) ◽

pp. 1922-1930 ◽

Cited By ~ 142

Author(s):

T Goto ◽

SJ Kennel ◽

M Abe ◽

M Takishita ◽

M Kosaka ◽

...

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

B Cells ◽

B Cell ◽

Plasma Cell ◽

Peripheral Blood ◽

Plasma Cells ◽

Affinity Constant ◽

Target Antigen ◽

Specific Marker

Abstract A monoclonal antibody (MoAb) that defines a novel terminal B-cell- restricted antigen, termed HM1.24, was developed against a human plasma cell line. The MoAb, designated anti-HM1.24, reacted with five different human myeloma cell lines, as well as with monoclonal neoplastic plasma cells obtained from the bone marrow or peripheral blood of patients with multiple myeloma or Waldenstrom's macroglobulinemia. The HM1.24 antigen was also expressed by mature Ig- secreting B cells (plasma cells and lymphoplasmacytoid cells) but not by other cells contained in the peripheral blood, bone marrow, liver, spleen, kidney, or heart of normal individuals or patients with non- plasma-cell-related malignancies. The anti-HM1.24 MoAb bound to human myeloma RPMI 8226 cells with an affinity constant of 9.2 x 10(8) M-1, indicating approximately 84,000 sites/cell. By immunoprecipitation assay under reducing conditions, this MoAb identified a membrane glycoprotein that had a molecular weight of 29 to 33 kD. Our studies indicate that the HM1.24-related protein represents a specific marker of late-stage B-cell maturation and potentially serves as a target antigen for the immunotherapy of multiple myeloma and related plasma cell dyscrasias.

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A novel membrane antigen selectively expressed on terminally differentiated human B cells

Blood ◽

10.1182/blood.v84.6.1922.bloodjournal8461922 ◽

1994 ◽

Vol 84 (6) ◽

pp. 1922-1930 ◽

Cited By ~ 1

Author(s):

T Goto ◽

SJ Kennel ◽

M Abe ◽

M Takishita ◽

M Kosaka ◽

...

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

B Cells ◽

B Cell ◽

Plasma Cell ◽

Peripheral Blood ◽

Plasma Cells ◽

Affinity Constant ◽

Target Antigen ◽

Specific Marker

A monoclonal antibody (MoAb) that defines a novel terminal B-cell- restricted antigen, termed HM1.24, was developed against a human plasma cell line. The MoAb, designated anti-HM1.24, reacted with five different human myeloma cell lines, as well as with monoclonal neoplastic plasma cells obtained from the bone marrow or peripheral blood of patients with multiple myeloma or Waldenstrom's macroglobulinemia. The HM1.24 antigen was also expressed by mature Ig- secreting B cells (plasma cells and lymphoplasmacytoid cells) but not by other cells contained in the peripheral blood, bone marrow, liver, spleen, kidney, or heart of normal individuals or patients with non- plasma-cell-related malignancies. The anti-HM1.24 MoAb bound to human myeloma RPMI 8226 cells with an affinity constant of 9.2 x 10(8) M-1, indicating approximately 84,000 sites/cell. By immunoprecipitation assay under reducing conditions, this MoAb identified a membrane glycoprotein that had a molecular weight of 29 to 33 kD. Our studies indicate that the HM1.24-related protein represents a specific marker of late-stage B-cell maturation and potentially serves as a target antigen for the immunotherapy of multiple myeloma and related plasma cell dyscrasias.

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c-Myb is required for plasma cell migration to bone marrow after immunization or infection

Journal of Experimental Medicine ◽

10.1084/jem.20150191 ◽

2015 ◽

Vol 212 (7) ◽

pp. 1001-1009 ◽

Cited By ~ 21

Author(s):

Kim L. Good-Jacobson ◽

Kristy O’Donnell ◽

Gabrielle T. Belz ◽

Stephen L. Nutt ◽

David M. Tarlinton

Keyword(s):

Bone Marrow ◽

Transcription Factor ◽

Cell Migration ◽

Virus Infection ◽

Plasma Cell ◽

Plasma Cells ◽

Critical Role ◽

Dramatic Reduction ◽

Factor C

Plasma cell migration is crucial to immunity, but little is known about the molecular regulators of their migratory programs. Here, we detail the critical role of the transcription factor c-Myb in determining plasma cell location. In the absence of c-Myb, no IgG+ antigen-specific plasma cells were detected in the bone marrow after immunization or virus infection. This was correlated with a dramatic reduction of plasma cells in peripheral blood, mislocalization in spleen, and an inability of c-Myb–deficient plasma cells to migrate along a CXCL12 gradient. Therefore, c-Myb plays an essential, novel role in establishing the long-lived plasma cell population in the BM via responsiveness to chemokine migration cues.

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Normal Plasma Cell Biology: Natural Achilles Heels as Targets in Myeloma

Blood ◽

10.1182/blood.v130.suppl_1.sci-7.sci-7 ◽

2017 ◽

Vol 130 (Suppl_1) ◽

pp. SCI-7-SCI-7

Author(s):

Katia Georgopoulos

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

Gene Family ◽

B Cell ◽

Plasma Cell ◽

Cell Biology ◽

Plasma Cells ◽

B Cell Differentiation ◽

High Affinity ◽

Bone Marrow Plasma

Abstract Normal Plasma Cell Biology: Natural Achilles Heels as Targets in MyelomaÊ Primary exposure to antigen leads to affinity maturation, selection and terminal differentiation of mature B cells to antibody-secreting plasma cells. Long-lived high-affinity plasma cells and their immediate precursors reside in the bone marrow and are responsible for rapid recall reactions and a life-time production of high affinity antibodies. Members of the IKAROS gene family have been specifically implicated in the generation of long-lived plasma cells. Mice deficient for Aiolos, the B cell specific member of the IKAROS gene family fail to produce high affinity plasma cells in the bone marrow and to sustain serum antibody titers after both primary or secondary immunization with a range of hapten concentrations. Chimera reconstitutions have demonstrated that the bone marrow plasma cell defect is B cell intrinsic. However, lack of AIOLOS does not alter expression of any of the previously described factors required for B cell differentiation into the plasma cell stage. No defect in somatic hyper-mutation, the generation of memory B cells, or short-lived high affinity plasma cells in the spleen is observed upon challenge or re-challenge. Thus this IKAROS family member is critically required for high-affinity bone plasma cells possibly by regulating their interaction with the bone marrow microenvironment, a process that is likely critical for long-term survival. High-affinity bone marrow plasma cells are the normal counterpart of the devastating B cell malignancy multiple myeloma. Multiple myeloma is treatable with immune modulatory drugs (IMiD) such as lenalidomide. Recent studies have shown that IMiD alter the specificity of the CRL4-CEREBLON (CRL4CRBN) E3 ubiquitin ligase complex. Among the new targets for the CRL4CRBN complex are two of the IKAROS family members, AIOLOS and IKAROS, whose degradation adversely affects the cellular fitness of multiple myeloma cells. Recent studies in B cell precursors have implicated the IKAROS gene family in the regulation of bone marrow stromal interactions. IKAROS and AIOLOS activities balance self-renewal, survival, and pre-B cell differentiation by engaging epigenetic and transcriptional machineries through discrete lineage-specific superenhancers. Such AIOLOS and IKAROS-dependent regulatory pathways engaged in early B cell precursors may also be involved in controlling bone marrow stromal interactions with high affinity plasma cells and their malignant counterparts thereby contributing to long-term growth and survival. Further exploitation of IKAROS family-targeted pathways in bone marrow plasma cells and multiple myeloma is warranted. Disclosures No relevant conflicts of interest to declare.

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CD28 Supports Normal Plasma Cell Survival and Interactions with Microenvironment

Blood ◽

10.1182/blood.v112.11.2713.2713 ◽

2008 ◽

Vol 112 (11) ◽

pp. 2713-2713

Author(s):

Cheryl H Rozanski ◽

Jayakumar Nair ◽

Louise Carlson ◽

Kelvin P. Lee

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

Dendritic Cells ◽

Cell Survival ◽

Plasma Cell ◽

Plasma Cells ◽

Bone Marrow Biopsies ◽

Myeloma Cells ◽

Normal Plasma

Abstract The long term generation of protective antibodies (Abs) requires the continuous survival of long-lived plasma cells that are maintained within specialized bone marrow niches by complex interactions that remain largely uncharacterized. Previous studies have shown that the T cell costimulatory receptor CD28 is expressed on normal and transformed (murine plasmacytoma, human multiple myeloma) plasma cells – however, its role in the B cell lineage remained unclear. We have recently shown that CD28 expressed on transformed human plasma cells (multiple myeloma cells) directly delivers pro-survival signals to the myeloma cells and protects them against intrinsically and extrinsically induced death (Bahlis et al, 2007). Furthermore, myeloma cells directly interact with dendritic cells (DC, both in vitro and in patient bone marrow biopsies), and the DC provide the ligands (i.e. CD80 and CD86) for myeloma-CD28. Others studies utilizing competitive bone marrow reconstitution have indirectly suggest a role for CD28 in the function and/or survival of normal murine plasma cells (Delogu et al, 2006). These observations led us to directly investigate the role of CD28 in normal plasma cell survival as well as cell-cell interactions with CD80/CD86+ bone marrow derived dendritic cells (BMDC). In vitro serum starvation experiments, direct activation of CD28 by an agonistic anti-CD28 mAb increased survival of serum-starved PC by 63% (p<0.001). Addition of BMDC improved the survival of PC by 20% over that seen with media alone, and resulted in a significant increase in IgG production (p<0.01). We and others have shown that CD28 binding to CD80/CD86 on DC also “backsignals” to the DC to produce the PC survival factor IL-6. We found that co-culture with the murine plasmacytoma cell line S194 induced 155 pg/ml of IL-6 from BMDC (p<0.01 vs. BMDC alone and S194 alone), and primary plasma cells isolated from bone marrow induced 290 pg/ml of IL-6 from BMDC (p<0.001 vs. BMDC alone). Induction of BMDC production of IL-6 by both primary and transformed PC was significantly inhibited (p<0.05) by antibody blockade of CD80 and CD86. Our data demonstrates that signaling through CD28 directly supports the survival of normal bone marrow plasma cells, and that “backsignaling” through PC-CD28 engagement of DC-CD80/CD86 induces DC to secrete the pro-survival cytokine IL-6. These findings suggest that CD28 is a key molecular bridge that connect normal plasma cells to the supportive microenvironment.

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