Anti-CD38 Pretargeted Radioimmunotherapy Demonstrates Therapeutic Efficacy In a Human Multiple Myeloma Mouse Xenograft Model

Abstract Abstract 1842 Multiple myeloma (MM) remains incurable despite improved response rates and improved progression free survival in the era of therapy with novel agents, including bortezomib, thalidomide, and lenalidomide. Disease persistence is presumably due to residual malignant plasma cell clones that evade or develop resistance to available therapies. The efficacy of radioimmunotherapy (RIT) in the treatment of hematologic malignancies is well established and the radiosensitivity of malignant plasma cells has been demonstrated in both preclinical and clinical settings. The ectoenzyme receptor CD38 is a plasma cell antigen that exhibits relatively specific, stable and uniform expression (95–100%) at a high epitope density on myeloma cells, making it an attractive target for antibody based therapies, including RIT. Pretargeted RIT (PRIT), using a multi-step streptavidin (SA)-biotin targeting system enhances the therapeutic index of delivered radiation. We have generated an anti-CD38 antibody (Ab)-SA synthetic chemical conjugate (OKT10-SA). The OKT10-SA construct binds with high avidity to myeloma cells while retaining full biotin-binding capability for radiolabeled DOTA-biotin. Blood, tumor and nonspecific organ uptakes of OKT10-SA were directly measured in biodistribution experiments involving athymic nude mice bearing human MM xenograft tumors. Groups of 5 mice with s.c. L363 human MM (IgG) xenograft tumors received 1.4 nmol (300 μg) of either OKT10-SA (anti-CD38 SA) or an IgG1 isotype matched control Ab BHV1-SA (bovine herpes virus-1) 22 hrs prior to synthetic biotin-acetyl-galactosamine clearing agent (CA; 5.8 nmol [50 μg]) and 24 hrs prior to trace labeled 111In-DOTA-biotin (1 μg). The CA removed >95% of both unbound OKT10-SA and BHV1-SA from the mouse circulation within 30 minutes of administration. Animals were euthanized and comprehensive tissue biodistributions were assessed 2, 24, 48 and 96 hrs after 111In-DOTA-biotin injection. Tumors excised from mice pretargeted with OKT10-SA contained 13.1 ± 1.9 % of the injected dose of 111In-DOTA-biotin per gram (% ID/g) after 2 hrs and 8.8 ± 2.8 % ID/g after 24 hrs compared to 2.4 ± 0.6 % ID/g after 2 hrs and 0.9 ± 0.4 % ID/g after 24 hrs in tumors excised from control mice pretargeted with BHV1-SA. Tumor-to-normal organ ratios of absorbed radioactivity were 8:1; 10:1; 8:1; and 6:1 respectively for blood, lung, liver and kidney in mice pretargeted with OKT10-SA; compared to 0.6:1; 0.9:1; 0.8:1 and 0.4:1 respectively, in control mice pretargeted with BHV1-SA. Therapy studies were then performed in athymic nude mice (n=9-10/group) bearing s.c. L363 human MM xenograft tumors. Reagent concentrations and time-points for administration of OKT10-SA, BHV1-SA and CA were identical to those reported for the biodistribution studies. The high energy beta particle emitter 90Yttrium (t1/2 = 64 hrs) was used as the therapeutic radionuclide. 90Y-DOTA-biotin (2 μg) was labeled with 400 μCi, 800 μCi, or 1200 μCi per mouse in 3 OKT10-SA groups and 3 control groups (untreated control; 800 μCi or 1200 μCi 90Y-DOTA-biotin following BHV1-SA). All mice in the untreated control and BHV1-SA control groups experienced exponential MM tumor growth and 78% of the untreated control animals required euthanasia within 17 days. All mice pretargeted with OKT10-SA demonstrated tumor shrinkage by day 6 at all dose levels (see figure). After 17 days, 90% of the OKT10-SA treated animals in the 400 μCi and 1200 μCi groups and 100% of the animals in the 800 μCi remained alive. One animal treated with 1200 μCi was euthanized on day 10 due to weight loss, however the remaining 9 animals from that group were 106 ±9% of initial body weight on day 17. Objective remissions were observed within 6 days in 100% of the mice treated with OKT10-SA followed by 1200 μCi of 90Y-DOTA-biotin, including 100% complete remissions (no detectable tumor in OKT10-SA treated mice compared to tumors that were 5240 ± 2495% of initial tumor volume in untreated control animals) by day 17. These studies represent the first application of both PRIT and CD38 targeted radioimmunotherapy in MM. Favorable OKT10-SA biodistribution findings correlate with early evidence of therapeutic efficacy. Tumor responses in this MM xenograft tumor model are encouraging, but long term toxicity and survival results are not yet mature. Future studies combining PRIT and novel agents are planned in xenograft and SCID-hu myeloma models. Disclosures: Gopal: Millenium. Wood:BD Biosciences: Research Funding.

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Anti-CD38 Pretargeted Radioimmunotherapy Eradicates Multiple Myeloma Xenografts In a Murine Model

Blood ◽

10.1182/blood.v122.21.882.882 ◽

2013 ◽

Vol 122 (21) ◽

pp. 882-882 ◽

Cited By ~ 2

Author(s):

Damian J. Green ◽

Jon C. Jones ◽

Mark D. Hylarides ◽

Donald K. Hamlin ◽

D Scott Wilbur ◽

...

Keyword(s):

Multiple Myeloma ◽

Plasma Cell ◽

Nude Mice ◽

Dose Range ◽

High Energy ◽

Beta Particle ◽

Control Group ◽

High Avidity ◽

Myeloma Cells ◽

Athymic Nude Mice

Abstract Novel therapies, including immunomodulatory agents (thalidomide, lenalidomide and pomalidomide) and proteasome inhibitors (bortezomib, carfilzomib) have improved response rates and prolonged progression free survival for patients with multiple myeloma (MM). Despite these advances the disease remains incurable. MM persistence is presumably due to residual malignant plasma cell clones that evade or develop resistance to available therapies. The efficacy of radioimmunotherapy (RIT) in the treatment of hematologic malignancies is well established and the radiosensitivity of malignant plasma cells has been demonstrated in both preclinical and clinical settings. CD38 is a plasma cell antigen that exhibits relatively specific, stable and uniform expression (95-100%) at a high epitope density on myeloma cells, making it an attractive target for antibody based therapies, including RIT. Recently, the unmodified CD38 mAb daratumumab has demonstrated anti-MM tumor cell activity both in vitro and in mouse xenografts, validating the antigen as a desirable target. Pretargeted RIT (PRIT), using a multi-step streptavidin (SA)-biotin targeting system enhances the therapeutic index of delivered radiation. We have generated an anti-CD38 antibody (Ab)-SA chemical conjugate (OKT10-SA). The OKT10-SA construct binds with high avidity to myeloma cells while retaining full biotin-binding capability for radiolabeled DOTA-biotin. Blood, tumor and nonspecific organ uptakes of OKT10-SA directly measured in biodistribution experiments involving athymic nude mice bearing human MM xenograft tumors (Green, et al. ASH 2011) demonstrated tumor-to-normal organ ratios of absorbed radioactivity that were 8:1; 10:1; 8:1; and 6:1 respectively for blood, lung, liver and kidney in mice pretargeted with OKT10-SA; compared to 0.6:1; 0.9:1; 0.8:1 and 0.4:1 respectively, in control mice pretargeted with the IgG1 isotype matched control Ab BHV1-SA (bovine herpes virus-1). In therapy studies athymic nude mice (n=9-10/group) bearing s.c. NCI-H929 human MM xenograft tumors received 1.4 nmol (300 µg) of either OKT10-SA (anti-CD38 SA) or BHV1-SA 22 hrs prior to synthetic biotin-acetyl-galactosamine clearing agent (CA; 5.8 nmol [50 µg]) and 24 hrs prior to 90Y-DOTA-biotin (2 µg) labeled with 800 µCi per mouse. Previously, a dose range of 800 µCi to 1200 µCi was identified as optimal when the high energy beta particle emitter 90Yttrium (t1/2 = 64 hrs) was used as the therapeutic radionuclide in a MM xenograft model (Green, et al. ASH 2011). All mice in the BHV1-SA control group experienced exponential MM tumor growth and 100% of these control animals required euthanasia within 37 days. All mice pretargeted with OKT10-SA demonstrated tumor shrinkage by day 6. After 70 days, 100% of the OKT10-SA treated animals remained alive and objective remissions were observed within 6 days in 100% of the mice treated with OKT10-SA followed by 800 µCi of 90Y-DOTA-biotin, including 100% complete remissions (no detectable tumor in OKT10-SA treated mice) compared to tumors that were 2982 ± 2834% of initial tumor volume [p<0.0001, Student’s t-test] in untreated control animals by day 23 [Figure]. Hepatic and renal function was assessed 160 days after mice received OKT10-SA followed by 90Y-DOTA-biotin (800 µCi, n=5) and no significant toxicity was observed. The measured serum creatinine was <0.4 mg/dl in all animals, the average BUN, ALT and AST values were 48 + 33 mg/dl, 46.8 +40 and 120.4 + 116.7 U/L respectively. The observed anti-tumor activity in this model was not attributable to an effector response mediated by the CD38 mAb in the absence of radiolabeled DOTA-biotin. Mice bearing MM xenografts (n=10) treated with OKT10-SA administered alone (without subsequent 90Y-DOTA-biotin infusion) demonstrated no objective tumor responses and 100% required euthanasia by day 17 for tumors that were 7837 ± 3492% of their initial volume. In a matched cohort of mice (n=10) treated concurrently with identical doses of OKT10-SA followed by CA and 90Y-DOTA-biotin (800µCi), 100% of the treated animals achieved a complete remission by day 17 and 70% of these animals were alive and tumor free at day 80. Tumor responses in this MM xenograft tumor model are encouraging and results support further evaluation of both PRIT and CD38 targeted RIT in MM. Studies combining PRIT and novel agents are ongoing in both xenograft and SCID-hu MM models. Disclosures: No relevant conflicts of interest to declare.

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Frequent Detection of Minimal Residual Disease (MRD) in Autografts from Patients with Multiple Myeloma during Remission Using a New Culture Method.

Blood ◽

10.1182/blood.v104.11.4877.4877 ◽

2004 ◽

Vol 104 (11) ◽

pp. 4877-4877

Author(s):

Yossi Cohen ◽

Yzhar Hardan ◽

Arnon Nagler ◽

Dov Zipori

Keyword(s):

Multiple Myeloma ◽

Plasma Cell ◽

Residual Disease ◽

Culture Method ◽

Expansion Method ◽

Myeloma Cell ◽

Current Treatment ◽

Myeloma Cells ◽

The Moment

Abstract Current treatment of multiple myeloma includes autologous stem cell transplantation. However, it is unknown at the moment what is the extent of graft contamination with clonotypic myeloma cells. In order to evaluate the extent of residual contamination of the graft with myeloma cells, we used our new myeloma cell culture and expansion method developed in the Weizmann Institute of Science for the detection of MRD. We observed readily growing residual myeloma cells in 6 of seven cases, confirmed by clonal markers (FACS, PCR and FISH). However, there was some variability in the pattern of growth; one case of plasma cell leukemia and two cases with t(4;14) showed earlier and more pronounced growth, whereas one case with systemic amyloidosis and another case with MGUS failed to grow in this culture. We are currently arranging a multicenter study for further assessment of these findings and aim to answer the question whether the culture can distinguish between multiple myeloma and other plasma cell dyscrasias. Another goal is to correlate the pattern of in vitro growth of multiple myeloma cells, with clinical and chromosomal characteristics. Figure Figure

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Exposure to Novel Agents Contribute to Extramedullary Spread of Multiple Myeloma Cells and Altered Expression of Adhesion Molecules: A Clinicopathologic Study of 91 Consecutive Autopsy Cases

Blood ◽

10.1182/blood.v120.21.1832.1832 ◽

2012 ◽

Vol 120 (21) ◽

pp. 1832-1832

Author(s):

Sohtaro Mine ◽

Shotaro Hagiwara ◽

Amane Tagashira ◽

Toru Igari ◽

Akiyoshi Miwa

Keyword(s):

Multiple Myeloma ◽

Adhesion Molecules ◽

Conflicts Of Interest ◽

Remission Rate ◽

Novel Agents ◽

Altered Expression ◽

Myeloma Cells ◽

Factors Associated ◽

Patient Profile ◽

Duration Of Illness

Abstract Abstract 1832 Background Recent development of novel agents such as bortezomib, lenalidomide, and thalidomide improved the remission rate and survival of multiple myeloma. However, the end stage of myeloma is still uncontrollable. The extramedullary disease (EMD) progression of myeloma was frequently observed in heavily treated patients, whereas few data exist about its incidence and predictive factors. It is well known that adhesion molecules play a role in disease progression and drug resistance. We investigated factors associated with the EMD progression based on the autopsy cases. In addition, the effect of the exposure to novel agents on the expression of adhesion molecules on myeloma cells was analyzed. Methods We reviewed autopsy reports and medical records of 91 multiple myeloma cases between 1979 and 2012 at the National Medical Center for Global Health and Medicine in Tokyo, Japan. The sites of myeloma cell invasion, infection, hemorrhage and renal complications were studied gross and microscopically. Patient profile, duration of illness, type of monoclonal gammopathy, clinical stage and history of treatment were studied. Durie & Salmon's criteria was used for diagnosis and staging. Factors associated with EMD were statistically analyzed using Student's t-test and the chi-square test. NCAM, VCAM, ICAM, and LFA-1 immunostaining in the bone marrow was performed. Results In 91 autopsy cases, 62.6% was male. Mean age and the duration of illness was 63.0 (38–85) years old and 40.6 months (1–156). Eighteen patients (19.8%), 23 (25.3%), and 15 (16.4%) were treated with novel agents, SCT and both respectively. EMD progression of myeloma cells was observed in 65 patients (71.4%). Frequent sites of EMD were spleen (48.9%), liver (37.8%), kidney (31.1%), lymph nodes (28.6%), lung (25.6%), pancreas (20.0%), and gastro-intestinal tract (18.9%). The incidence of EMD was significantly higher in patients treated with novel agents than in patients without novel agents (94.4% vs. 65.8%, p=0.016). The risk factors of EMD were novel agents, longer duration of illness, and adverse cytogenetic abnormalities. In the cases with novel agents, the expression of NCAM was significantly low (11.1% vs. 57.4%, p=0.049) compared to the cases without novel agents. However, the expression of VCAM was significantly higher in the cases with novel agents than the cases without novel agents (40.0% vs. 0%, p=0.01). Conclusion Multi-organ involvement of myeloma is not rare in autopsy cases of the disease. Exposure to novel agents may contribute to extramedullary spread of myeloma cells and altered expression of adhesion molecules. Further study on other adhesion molecules is needed. Disclosures: No relevant conflicts of interest to declare.

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Enumeration and characterization of circulating multiple myeloma cells in patients with plasma cell disorders

British Journal of Haematology ◽

10.1111/bjh.15003 ◽

2017 ◽

Vol 180 (1) ◽

pp. 71-81 ◽

Cited By ~ 13

Author(s):

Brad Foulk ◽

Mike Schaffer ◽

Steve Gross ◽

Chandra Rao ◽

Denis Smirnov ◽

...

Keyword(s):

Multiple Myeloma ◽

Plasma Cell ◽

Myeloma Cells ◽

Plasma Cell Disorders

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Comparison of Newly Diagnosed and Relapsed Refractory Multiple Myeloma Using Transcriptional Profiling of Myeloma Cells.

Blood ◽

10.1182/blood.v106.11.1555.1555 ◽

2005 ◽

Vol 106 (11) ◽

pp. 1555-1555

Author(s):

Shaji Kumar ◽

Philip R. Greipp ◽

Jessica L. Haug ◽

Michael Kline ◽

Wee Joo Chng ◽

...

Keyword(s):

Multiple Myeloma ◽

Drug Resistance ◽

Differential Expression ◽

Median Survival ◽

Plasma Cell ◽

Plasma Cells ◽

Expression Profiles ◽

Significant Proportion ◽

Newly Diagnosed ◽

Myeloma Cells

Abstract Background: Multiple myeloma (MM) is a plasma cell malignancy that is incurable with current approaches. The median survival for patients with MM is around four years and a significant proportion of patients experience a course characterized by multiple relapses treated with different therapies. The median survival for patients relapsing after the initial therapy is nearly 18 months and successive treatment strategies result in decreasing response durations, likely reflecting acquired drug resistance. In order to better understand the biological changes associated with advanced, relapsed, refractory MM, we compared gene expression profiles (GEP) of malignant plasma cells isolated from patients with relapsed refractory MM and compared them to plasma cells from patients with newly diagnosed MM. Methods: In order to obtain two relatively homogenous group of patients, we compared samples from 44 patient with newly diagnosed MM enrolled in the ECOG E1A00 clinical trial (comparing thalidomide and dexamethasone to dexamethasone alone) to 44 patients with relapsed refractory MM enrolled in a phase II trial of Velcade (SUMMIT), where most patients had four or more previous relapses. Plasma cells from bone marrow aspirates were separated by magnetic bead selection of CD138 positive cells and studied using Affymetrix HG-U133A chips using standard methodology. The arrays were analyzed using Genespring 7.2 software following GCRMA normalization and genes with differential expression between the two datasets were examined. Differentially expressed genes were further analyzed using Ingenuity Pathways Analysis program. Results: A total of 864 genes were identified which were at least two fold and significantly different between the newly diagnosed and relapsed patients. Using Ingenuity software, 437 of these genes were mapped to different biological networks. Examination of the canonical pathways demonstrated several important cellular pathways differentially regulated between the two groups. Several important mediators of the cytokines, receptors and respective signaling pathways appear to be down regulated in the relapsed group and included IGF-1, HGF, SDF-1 alpha, gp130 and importantly the MEK/ERK pathway. Additionally expression of adhesion molecules such as VCAM1 and PECAM was decreased in the relapsed group compared to newly diagnosed pts. There appear to increased tissue hypoxia in the relapsed marrow as indicated by up regulation of HIF-1 alpha as well increased levels of Placental growth factor. Myeloma cells from relapsed disease were characterized by decreased expression of mcl1, FLIP1, and bcl-xL and increased caspase 8 relative to newly diagnosed group. Also seen was decreased expression of the glucocorticoid and interferon receptors in the relapsed setting. Conclusion: Comparison of the GEP between MM cells from newly diagnosed and relapsed pts demonstrates important differences that have potential biological relevance. The plasma cell in the relapsed setting appears to be more independent of the tumor microenvironment. Additionally, differential expression of some of the genes provides clues to mechanisms of drug resistance commonly observed in the relapsed pts. We are in the process of validating some of the key findings from these analyses.

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Angiogenesis in Multiple Myeloma (MM): Angiogenic Switch or Reflexion of Plasma Cell Number? A Gene Expression Based Survey in Primary Myeloma Cells and the Bone Marrow Microenvironment.

Blood ◽

10.1182/blood.v108.11.3405.3405 ◽

2006 ◽

Vol 108 (11) ◽

pp. 3405-3405

Author(s):

Dirk Hose ◽

John DeVos ◽

Christiane Heiß ◽

Jean-Francois Rossi ◽

Angela Rösen-Wolff ◽

...

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

Plasma Cell ◽

Empirical Bayes ◽

Plasma Cells ◽

De Novo ◽

Cell Number ◽

Bone Marrow Microenvironment ◽

Myeloma Cells ◽

Angiogenic Switch

Abstract BACKGROUND. Angiogenesis is a hallmark of active multiple myeloma. However, two etiologic hypotheses have been proposed: an angiogenic switch (i.e. differential or de novo expression of pro/antiangiogenic genes in MM), and, alternatively an effect of increased plasma cell number. AIM of this study was to investigate the angiogenic signature of multiple myeloma cells (MMC), normal bone marrow plasma cells (BMPC), the bone marrow microenvironment (BMME) and cellular subfractions therein. PATIENTS AND METHODS. 128 newly diagnosed MM-patients (65 training (TG) / 63 independent validation group (VG)) and 14 normal donors (ND) were included. Bone marrow aspirates were CD138-purified by activated magnetic cell sorting. Whole bone marrow (n=49) and FACSAria sorted subfractions thereof (n=5) were investigated. RNA was in-vitro transcribed and hybridised to Affymetrix HG U133 A+B GeneChip (TG) and HG U133 2.0 plus arrays (VG). Expression data were gcrma-normalised and the empirical Bayes algorithm used. p-Values were adjusted using the Benjamini-Hochberg method (Bioconductor). iFISH was performed on purified MM-cells using probesets for chromosomes 1q21, 9q34, 11q23, 11q13, 13q14, 15q22, 17p13, 19q13, 22q11 and the translocations t(4;14) and t(11;14). HGF expression was verified by real time RT-PCR and western blotting. Based on Medline review, we established a list of 89 pro- and 56 antiangiogenic genes and investigated their expression according to the stage of disease: BMPC vs. MGUS, SD stage I (asymptomatic myeloma) vs. SD stage II/III (symptomatic myeloma requiring therapy). RESULTS. BMPC express pro- (e.g. VEGFA) and antiangiogenic genes (e.g. TIMP2). Only one pro-angiogenic gene (hepatocyte growth factor, HGF) is significantly overexpressed in MMC compared to BMPC. HGF has previously been linked with myeloma progression and induction of angiogenesis. Six antiangiogenic genes (TIMP2, SERPINF1, COL18A1, PF4, THBS1, CXCL14) are downregulated in MMC compared with BMPC. Compared to healthy donors, the BMME of MM shows a significant downregulation of PLAU (urokinase, antiangiogenic) and upregulation of TNF(proangiogenic). CONCLUSION. Upregulation of HGF-expression, downregulation of TIMP2, SERPINF1, COLA18A1, PF4, THBS1 and CXCL14 expression in MMC as well as downregulation of PLAU and upregulation of TNFα in the BMME seem to indicate an “angiogenic switch”. However, given the relatively low number of differentially expressed genes (7/145) and the expression of angiogenic genes by BMPC, an effect caused by an increasing number of plasma cells might be evenly important.

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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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The Role of the Gut Microbiome in Pathogenesis, Biology, and Treatment of Plasma Cell Dyscrasias

Frontiers in Oncology ◽

10.3389/fonc.2021.741376 ◽

2021 ◽

Vol 11 ◽

Author(s):

Marcin Jasiński ◽

Jarosław Biliński ◽

Grzegorz W. Basak

Keyword(s):

Multiple Myeloma ◽

Gut Microbiota ◽

Plasma Cell ◽

Response To Treatment ◽

Faecal Microbiota ◽

Myeloma Cells ◽

Risk Of Progression ◽

Plasma Cell Dyscrasias ◽

Refractory Malignancy

In response to emerging discoveries, questions are mounting as to what factors are responsible for the progression of plasma cell dyscrasias and what determines responsiveness to treatment in individual patients. Recent findings have shown close interaction between the gut microbiota and multiple myeloma cells. For instance, that malignant cells shape the composition of the gut microbiota. We discuss the role of the gut microbiota in (i) the development and progression of plasma cell dyscrasias, and (ii) the response to treatment of multiple myeloma and highlight faecal microbiota transplantation as a procedure that could modify the risk of progression or sensitize refractory malignancy to immunotherapy.

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Characterization of Human Multiple Myeloma Cells with High-Resolution Magic Angle Spinning Magnetic Resonance Spectroscopy

Blood ◽

10.1182/blood.v112.11.5122.5122 ◽

2008 ◽

Vol 112 (11) ◽

pp. 5122-5122

Author(s):

Albert Oriol ◽

Ignasi Barba ◽

Angels Barbera ◽

Carles Arús ◽

Jose-Luis Garcia-Dorado ◽

...

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

Magnetic Resonance ◽

High Resolution ◽

Plasma Cell ◽

Plasma Cell Leukemia ◽

Resonance Spectroscopy ◽

Metabolic Profiles ◽

Cell Leukemia ◽

Myeloma Cells

Abstract Advancements in the pathogenetic pathways in multiple myeloma have led to the identification of several primary and secondary genetic lesions and ultimately to a multiple myeloma genetic classification with prognostic implications. Although disregulation of cyclin activity has been recognized as a key event leading to the multiple myeloma phenotype, little is known about the metabolic consequences of this phenomenon. We have studied intact multiple myeloma cells by high resolution magnetic resonance spectroscopy to establish the metabolomic profiles of different native multiple myeloma cells as compared to other lymphoproliferative disorders. Multiple myeloma cells obtained from bone marrow aspirates (n =15), blood (n =3) or other biologic tissues (n =2) from 20 multiple myeloma patients and separated by density gradient centrifugation were evaluated and metabolic profiles were correlated with cytogenetic characteristics of the disease and patients clinical characteristics. Twelve patients were females (60%) with a median age of 65 years (range 50–82). Multiple myeloma monoclonal proteins were IgG (N=9), IgA (N=5) or BJ (N=6). Five of them (25%) had renal insufficiency. Nine patients (45%) had predominantly extramedullar diesase including four cases of plasma cell leukemia. IgH translocations were identified in 5 samples (25%), hyperploidy in 2 (10%), and other or no genetic lesions in 13 (65%), del13 was present in 9 samples (45%) and p53 alterations in 5 (25%). Bone marrow samples from thirteen patients with conventional multiple myeloma presented a relatively constant metabolic pattern with predominantly lipidic signals and a metilen to metil ratio ranging from 1.9 to 4.9 (median 2.9). No differences in this pattern were observed among subgroups of primary translocations or involvement of Rb and p53 genes. Four patients with plasma cell leukemia and three with predominant extranodal disease presented either non detectable lipid signals (N=3) or a higher metilen to metil ratio ranging from 2.8 to 3.9 (median 3.5). In fact, extranodal or leukemic disease was significantly associated to undetectable lipids (P < 0.031) or the composite variable undetectable lipids or metilen to metil ratio > 3 (P < 0.043). Furthermore, after a median follow-up of 18 months, absence of lipids in the metabolic profile was also associated to a shorter survival (median 0.45 years, 95%CI 0–1.03 versus 3 years, 95%CI 0.95–5.06, P < 0.022). These results suggest that metabolic profiles of different multiple myeloma genetic subtypes share common and reletively constant characteristics, while cells obtained from patients with plasma cell leukemia or predominantly extramedullar disease present a clearly distinct profile, probably reflecting the metabolic effect of clonal evolution at a genetic level.

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Activation of Rac1 through FARP1 Induces Dexamethasone Resistance in Multiple Myeloma Cells

Blood ◽

10.1182/blood.v124.21.2248.2248 ◽

2014 ◽

Vol 124 (21) ◽

pp. 2248-2248

Author(s):

Atsuko Yamazaki ◽

Masahiro Takeuchi ◽

Tatsuzo Mishina ◽

Miki Yamazaki ◽

Chika Kawajiri ◽

...

Keyword(s):

Multiple Myeloma ◽

Cell Line ◽

Transforming Growth Factor Beta ◽

Transforming Growth Factor ◽

Conflicts Of Interest ◽

Novel Agents ◽

Tumor Cell Death ◽

Myeloma Cells ◽

Dexamethasone Resistance ◽

Number Of Cells

Abstract Recently, novel agents such as bortezomib and lenalidomide have been introduced for multiple myeloma (MM) treatment and have improved patients' survival drastically. However, dexamethasone remains a mainstay in the treatment of MM. Dexamethasone effectively induces tumor cell death when used for the initial treatment of MM. In addition, dexamethasone has a synergistic effect with novel agents and is hence used in combination with such agents. However, prolonged dexamethasone exposure may lead to drug resistance. To elucidate the mechanism of dexamethasone resistance, we generated a dexamethasone-resistant subline of the MM cell line RPMI8226. We cultured RPMI8226 cells with 1 µM dexamethasone for 7 weeks and established the dexamethasone-resistant cell line Dex-R. This cell line showed no difference in survival in the presence or absence of 1 µM dexamethasone. We then examined differences in gene expression between RPMI8226 and Dex-R cells using cDNA microarray. Expression of the FARP1 gene, which is a transforming growth factor beta (TGF-b) target gene in myeloma cells, was increased approximately 50-fold in Dex-R cells compared to that in RPMI8226 cells. In some myeloma patients who become chemoresistant, myeloma cells show high levels of FARP1 expression at the initial stage. FARP1 has a Rho-GEF domain and can associate with proteins on the cell membrane through the FERM domain. In the nervous system, FARP1 is involved in synaptogenesis via the activation of Rac1. Based on these observations, we hypothesize that Dex-R cells acquires dexamethasone resistance with an increase in the level of FARP1 expression via the activation of Rac1. To verify this hypothesis, we established inducible FARP1 knockdown Dex-R cells using the TET-ON lentiviral system. We cultivated these cells for 24 h with doxycycline and added 1 µM dexamethasone. A total of 48 h after adding dexamethasone, we measured cell viability using the MTS assay. We cultured Dex-R cells with a Rac1 inhibitor (NSC23766) and added dexamethasone 12 h later. FARP1 expression decreased to approximately 10% in FARP1 knockdown cells 24 h after the addition of doxycycline. Without dexamethasone, there was no difference in survival in the presence or absence of doxycycline. However, when cells were cultured with dexamethasone, the growth of FARP1 knockdown Dex-R cells was significantly inhibited compared with that of the control (Fig 1). Next, we examined the change in dexamethasone resistance on the addition of the Rac1 inhibitor. The number of cells increased after 96 h without dexamethasone. On the other hand, the number of cells significantly decreased when cultured with dexamethasone (Fig 2). These data suggest that resistance to dexamethasone in Dex-R cells was mitigated by the inhibition of Rac1. We conclude that the activation of Rac1 through FARP1 is one mechanism of dexamethasone resistance in MM. Disclosures No relevant conflicts of interest to declare.

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