The Coexpression of CD11a and CD45bright Is the Hallmark of Proliferating Myeloma Cells.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 3347-3347
Author(s):  
Catherine Pellat-Deceunynck ◽  
Nelly Robillard ◽  
Regis Bataille
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
Ex Vivo ◽  
Inverse Correlation ◽  
Labeling Index ◽  
Myeloma Cells ◽  
Hla Dr ◽  
Brdu Labeling ◽  
A Minor ◽  
Brdu Staining

Abstract To identify new potential therapeutical targets in multiple myeloma (MM), we have defined the phenotype of the subset of proliferative myeloma cells (n=66) in comparison with that of normal PC (n=25). Proliferation was evaluated by ex vivo incorporation of BrdU (labeling index, LI). Surface PC phenotype was performed in a four-color assay with CD38, CD45, CD138 and the mAb indicated. For intracellular BrdU staining, cells were first labeled with CD38, CD45 and CD138, fixed and permeabilized before BdrU staining. At least 1000 normal PC and 10000 myeloma cells were analyzed. We show that all bone marrow PC, either malignant or normal, always included a subset of proliferative PC (BrdU+) that was always located within the CD45++subpopulation. Indeed, CD45++ myeloma cells (median 12%) had a labeling index 7.5-fold higher of that of CD45+/− myeloma cells (7.1% versus 0.94%). Actually, in all cases of MM, CD45++ myeloma cells were always the most proliferative myeloma cells. As observed for myeloma cells, LI of normal PC was heterogeneous i.e., higher in the CD45++ population of PC: CD45++ PC (median 65%) had a LI 5.7-fold higher of that of CD45+/− PC. Compared to myeloma cells, LI of PC were higher in both subsets, of 20.5% and 3.6% for CD45++ and CD45+/−, respectively. Non-malignant PC from blood or tonsil were homogeneously CD45++ and did proliferate (LI> 10% and up to 45% for reactive PC). In all PC (normal, reactive, malignant), we found an inverse correlation between CD45 and Bcl-2, confirming a known inverse correlation between proliferation and Bcl-2 expression. Our data suggest that a minor cycling Bcl2lowCD45++ population of myeloma cells differentiate into a no more cycling major Bcl2high CD45+/− population of myelom a cells that accumulates. We further characterized the phenotype of the CD45++ myeloma cells population: we found that CD11a and to a less extend HLA-DR were expressed by CD45++ myeloma cells only in contrast to CD40 and CXCR4 that were expressed by all myeloma cells. Moreover, all CD45++ myeloma cells coexpressed CD11a. Thus, the-to-be-killed population of myeloma cells could be targeted through CD45 or CD11a.

Phenotypic characterization of the human myeloma cell growth fraction

Blood ◽  
2005 ◽  
Vol 105 (12) ◽  
pp. 4845-4848 ◽  
Author(s):  
Nelly Robillard ◽  
Catherine Pellat-Deceunynck ◽  
Régis Bataille
Keyword(s):  
Plasma Cells ◽  
Ex Vivo ◽  
Inverse Correlation ◽  
B Cell Lymphoma ◽  
Myeloma Cell ◽  
Phenotypic Characterization ◽  
Growth Fraction ◽  
Myeloma Cells ◽  
Brdu Labeling

Abstract In this study we quantified the proliferation rate of normal and malignant plasma cells (PCs) by ex vivo incorporation of 5-bromo-2′-deoxyuridine (BrdU; labeling index, LI) using flow cytometry. We show that all bone marrow PCs, either normal or malignant, include a subset of proliferating PCs present within the CD45bright fraction. Indeed, medullary normal and malignant PCs were always heterogeneous for CD45 expression, and proliferation was always restricted primarily to the CD45bright compartment. Moreover, an inverse correlation was found between LI or CD45 and B-cell lymphoma 2 (Bcl-2) in both malignant and normal PCs, the most proliferating CD45bright PCs have the lowest Bcl-2 expression. We investigated expression of molecules of interest in multiple myeloma (MM)—that is, CD138, CD19, CD20, CD27, CD28, CD56, and CD11a—to further characterize the CD45bright fraction. Among all of these molecules, only CD11a was exclusively expressed by CD45bright proliferating myeloma cells. In conclusion, proliferating myeloma cells are characterized by the specific CD45bright CD11apos Bcl-2low phenotype. (Blood. 2005;105:4845-4848)

Ex Vivo Multiplexed Signal Pathway Inhibitor Treatment Reveals Differential Sensitivity of Myeloma and Non-Myeloma Bone Marrow Cell Populations.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 2860-2860
Author(s):  
Virginia Espina ◽  
Alessandra Romano ◽  
Amy VanMeter ◽  
Gregory Orloff ◽  
Alexander I. Spira ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
Bone Marrow Aspirate ◽  
Ex Vivo ◽  
Signal Pathway ◽  
Bone Marrow Microenvironment ◽  
Cell Populations ◽  
Myeloma Cells ◽  
Targeted Inhibitors ◽  
Target Effects

Abstract Abstract 2860 Poster Board II-836 Multiple Myeloma (MM) is a heterogeneous disease, and despite significant advances in therapy it remains incurable. A major challenge facing clinicians is to determine which new agent or which combinations of agents will prove efficacious for individual patients. Interaction of MM cells with bone marrow microenvironment cells has a pathogenetic role in the disease and confers tumor cell resistance to conventional therapies. The new generation of therapies is designed to target signal transduction pathways, particularly tyrosine kinase signaling. The phosphorylation, or activation state, of kinase-driven signal networks contains important information concerning disease pathogenesis and the ongoing state of kinase-associated therapeutic targets. Reverse-phase protein microarray (RPMA) is a reproducible, high-throughput system for protein signal pathway profiling. Phosphorylation state of kinase-associated therapeutic targets provides direct information regarding the target and off-target effects of treatment. We describe herein an ex vivo study of signal pathway inhibitor treatments of fresh bone marrow aspirate samples from patients undergoing standard of care hematological work up for multiple myeloma at any stage or treatment course. Bone marrow aspirates (n=37 to date), not required for diagnosis, were immediately subdivided and treated ex vivo with a panel of molecular targeted inhibitors and combinations, and/or exogenous ligands (SCF, IGF-1), and/or cytokines (IL-6), that target a wide range of cellular pathways (e.g. proteosome, angiogenesis related, protein degradation, cell proliferation/survival, insulin response, and protein translation). Up to 48 different treatment conditions can be studied for each patient sample from a bone marrow aspirate volume of 5.0mL. The objectives were a) measure the signal pathway perturbations caused by the inhibitor/ligand treatment in individual bone marrow aspirate samples, b) compare the relative sensitivity of tumor and non tumor bone marrow cells treated in admixture under identical conditions to identify predictive/prognostic protein-based biomarkers. We specifically measured the post-translational modifications (i.e. phosphorylation) of the drug target substrates to provide direct information regarding the target and off-target effects of treatment. After incubation for 4 hours the bone marrow aspirate samples were placed in a preservative that suppresses fluctuations in kinase pathway proteins. MM CD138 positive cells were separated from the non-CD138+ bone marrow microenvironment cells via immunomagnetic sorting. RPMA was used to quantitatively map 75 cell signaling pathway endpoints in CD138+ and non-CD138+ cell populations that were treated simultaneously prior to cell sorting. To date the accrual has included untreated patients with MGUS, smoldering myeloma, treatment naïve, or advanced stage, pre-treated myeloma. Individual patients differed widely in response to ex vivo treatment and combination of drug treatments. Dexamethasone showed differential effects on the myeloma cells compared to non-myeloma cells from the same patient. For all the untreated patients studied to date, dexamethasone significantly suppressed the phosphoAKT or NF-KB endpoints, while increasing the pro-apoptotic signaling proteins such as cleaved caspases. In contrast, all the patients heavily pre-treated with dexamethasone in combination with other agents did not show any differential effects of the dexamethasone treatment on the CD138+ cells compared to the non-CD138+ cells in the bone marrow microenvironment. Many molecular targeted inhibitors induced an up-regulation of the target pathway even in combination therapies such as Rapamycin plus an IGF-1R inhibitor. Sorafinib, a molecular targeted tyrosine kinase inhibitor which blocks the MAPK (Raf/Mek/ERK) pathway, induced a compensatory up-regulation of ERK T202/Y204 in all patients except one who was in remission after treatment. Multiplexed phosphoprotein cell signaling analysis before or after treatment with ex vivo inhibitors may predict patient-specific therapeutic response and/or off target effects. Selected agents that show efficacy in ex vivo studies may be considered for further in vivo clinical studies in which signal profiling of the bone marrow cellular populations could be performed before and after molecular targeted therapy. Disclosures: No relevant conflicts of interest to declare.

Reactive Plasmacytoses Are Expansions of Plasmablasts Retaining the Capacity to Differentiate Into Plasma Cells

Blood ◽  
1999 ◽  
Vol 94 (2) ◽  
pp. 701-712 ◽  
Author(s):  
Gaëtan Jego ◽  
Nelly Robillard ◽  
Denis Puthier ◽  
Martine Amiot ◽  
Françoise Accard ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Plasma Cell ◽  
Plasma Cells ◽  
Myeloma Cells ◽  
Survival Factor ◽  
Hla Dr ◽  
Reactive Plasma ◽  
Reactive Plasmacytosis ◽  
A Minor

Circulating plasma cells in 10 cases of reactive plasmacytosis had a shared phenotype with early plasma cell (CD19+CD38+ CD138+ CD40+CD45+ CD11a+ CD49e−CD56−). In most cases, a minor subpopulation of CD28+ plasma cells was also detected. Reactive plasma cells were highly proliferative, suggesting the presence of circulating progenitors (plasmablasts). After CD138+ plasma cell removal, highly proliferative CD138− plasmablasts differentiated into CD138+ plasma cells within a few days. This differentiation, which was associated with increased CD38 and decreased HLA-DR expression, was further confirmed by a large increase in intracellular Ig content (associated with Ig secretion) and was concomitant with extensive secretion of interleukin-6 (IL-6). The addition of neutralizing anti–IL-6 and anti-CD126 (IL-6 receptor) monoclonal antibodies totally prevented Ig secretion and cell differentiation by inducing apoptosis of plasmablasts, which indicates that IL-6 is an essential survival factor for plasmablasts. This report provides the first characterization of normal plasmablasts and shows that their phenotype is not exactly that of multiple myeloma cells.

Reactive Plasmacytoses Are Expansions of Plasmablasts Retaining the Capacity to Differentiate Into Plasma Cells

Blood ◽  
1999 ◽  
Vol 94 (2) ◽  
pp. 701-712 ◽  
Author(s):  
Gaëtan Jego ◽  
Nelly Robillard ◽  
Denis Puthier ◽  
Martine Amiot ◽  
Françoise Accard ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Plasma Cell ◽  
Plasma Cells ◽  
Myeloma Cells ◽  
Survival Factor ◽  
Hla Dr ◽  
Reactive Plasma ◽  
Reactive Plasmacytosis ◽  
A Minor

Abstract Circulating plasma cells in 10 cases of reactive plasmacytosis had a shared phenotype with early plasma cell (CD19+CD38+ CD138+ CD40+CD45+ CD11a+ CD49e−CD56−). In most cases, a minor subpopulation of CD28+ plasma cells was also detected. Reactive plasma cells were highly proliferative, suggesting the presence of circulating progenitors (plasmablasts). After CD138+ plasma cell removal, highly proliferative CD138− plasmablasts differentiated into CD138+ plasma cells within a few days. This differentiation, which was associated with increased CD38 and decreased HLA-DR expression, was further confirmed by a large increase in intracellular Ig content (associated with Ig secretion) and was concomitant with extensive secretion of interleukin-6 (IL-6). The addition of neutralizing anti–IL-6 and anti-CD126 (IL-6 receptor) monoclonal antibodies totally prevented Ig secretion and cell differentiation by inducing apoptosis of plasmablasts, which indicates that IL-6 is an essential survival factor for plasmablasts. This report provides the first characterization of normal plasmablasts and shows that their phenotype is not exactly that of multiple myeloma cells.

scholarly journals A Novel Class of Bifunctional Immunotherapeutic That Exploit a Universal Antibody Binding Terminus (uABT) to Recruit Endogenous Antibodies to Cells Expressing CD38 Demonstrates Anti-Multiple Myeloma Activity in Vitro and Ex Vivo against Patient Tumor Cells

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 4411-4411
Author(s):  
Ann Marie Rossi ◽  
Anna Bunin ◽  
Lawrence Iben ◽  
Matthew Welsch ◽  
Tanya Berbasova ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
Ex Vivo ◽  
Employment Equity ◽  
Myeloma Cells ◽  
Immune Effector ◽  
Effector Cells ◽  
Immune Effector Cells ◽  
Human Igg1 ◽  
Equity Ownership

Background: Antibody recruiting molecules (ARM) are novel, immunotherapeutic bifunctional molecules composed of two active termini connected by a linker. One of the termini binds to a target molecule on a cancer cell. The other terminus, called universal antibody binding terminus (uABT), recruit all endogenous IgG antibodies independent of their antigen binding specificity. As a result, the target cell is "opsonized" by antibodies which then bring the immune effector cells to eliminate the target through various antibody-dependent destruction mechanisms. Kleo Pharmaceuticals has developed a series of CD38-ARM mlecules which target human CD38 highly expressed by multiple myeloma cells. CD38-ARM compounds are able to mediate ADCC without depleting CD38 expressing immune effector cells like existing therapeutic antibodies such as Daratumumab. Methods: Cyclized peptides containing natural and non-natural amino-acid that selectively bind to human CD38 were identified using Peptidream Flexizyme-based, cell free Peptide Discovery Translation System. These peptides were linked to uABT antibody binder via a linker to generate the final CD38-ARM molecules Binding of CD38-ARM was tested by ternary complex formation between CD38 expressing cells, CD38-ARM and labelled human IgG1. To confirm the activity of CD38-ARM, surrogate CD16a binding and signaling assays were performed using the NFAT Promega system. Antibody dependent cellular cytotoxicity (ADCC) assays using purified NK cells from multiple donors with polymorphism variants (V/V, F/F, and V/F) of CD16a were performed to confirm activity. Live cell imaging was utilized to assess the dynamics of NK-RAJI cell interactions mediated by CD38-ARM +/- IgG. We evaluated the ability of compounds to mediate complement dependent cytotoxicity (CDC). We tested the effect of CD38-ARM on human immune cell populations within PBMC and whole bone marrow (WBM) by flow cytometry. Lastly, ex vivo samples from WBM of MM patients at diagnosis or relapse were used to evaluate CD38-ARM anti-tumor activity as well as off-target effects, without the addition of external source of IgG, through multiparametric flow-cytometry (CD45, CD19, CD38, CD138, CD56, CD27, CD8, CD117). Results: The CD38-ARM were shown to have the ability to bind to CD38 with a 7nM affinity and to human IgG1 and IgG2 with affinity of 15nM and 11nM by SPR. Activity of KP compounds was observed in all assays except for CDC. In ternary assay, KP-6 had an apparent EC50 of 16nM while KP-7's EC50 was 6nM. Both KP-6 and 7 had comparable EC50s in the single digit nanamolar range in the NFAT activation assay induced by CD16a binding was confirmed using human IgG to induce, while Daratumumab had an apparent EC50 of 0.04nM. In the ADCC assay, both KP-6 & KP-7 had EC50s of 7 & 6nM respectively, while Daratumumab had an EC50 of 0.1nM. In addition, no NK cell depletion was observed when PBMC were treated with KP compounds, whereas a profound reduction in both percentages and absolute numbers in this cell subset was observed with Daratumumab treatment. Increasing dose of CD38-ARM (range 0.1uM- 25uM) were tested in ex vivo WBM samples from MM patients together with a negative control and Daratumumab. At concentrations of 10uM and 25uM, CD38-ARM induced a significant reduction of MM cells achieving results comparable to those of Daratumumab activity (p >0.05 in both cases), while sparing all other CD38+ normal cells such as NK, T lymphocytes, monocytes and granulocytes, which are always reduced in the presence of Daratumumab. Conclusions: CD38-ARMs are able to kill MM cells by ADCC without depleting CD38 expressing immune cells contrary to existing antibodies such as Daratumumab. CD38-ARMs do not activate complement, which might be involved in the infusion reaction seen with Daratumumab. Most importantly, CD38-ARMs kill multiple myeloma cells ex vivo in patient bone marrow samples as well as plasma cell leukemia in patient blood. Combined with the in vivo efficacy data presented elsewhere, this data establishes the therapeutic potential of CD38-ARM. They also represent the first demonstration of the ARM platform ability to generate therapeutic agents tailored to a specific indication, by varying target binding moiety of the molecule. Disclosures Rossi: Kleo pharmaceuticals: Employment, Equity Ownership. Bunin:Kleo pharmaceuticals: Employment, Equity Ownership. Iben:Kleo Pharmaceuticals: Employment, Equity Ownership. Welsch:Kleo pharmaceuticals: Employment, Equity Ownership. Berbasova:Kleo Pharmaceuticals: Employment, Equity Ownership. Riillo:Kleo Pharmaceuticals: Research Funding. Ohuchi:Peptidream Inc.: Employment. Alvarez:Kleo pharmaceuticals: Employment, Equity Ownership. Kawakami:Peptidream Inc.: Employment. Nagasawa:Peptidream Inc.: Employment. Spiegel:Kleo pharmaceuticals: Equity Ownership. Rastelli:Kleo pharmaceuticals: Employment, Equity Ownership.

scholarly journals Patient-Derived 3D Tissue-Engineered Bone Marrow Cultures Support Primary MM Growth

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 4705-4705
Author(s):  
Pilar De La Puente ◽  
Feda Azab ◽  
Barbara Muz ◽  
Justin King ◽  
Ravi Vij ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
Flow Cytometry ◽  
Ex Vivo ◽  
Myeloma Cells ◽  
Accessory Cells ◽  
Aggressive Tumors ◽  
Cd Markers ◽  
Better Than

Abstract INTRODUCTION: In vitro culturing of primary myeloma cells has been a major challenge because the lack of an in vitro technology capable of recreating the complicated bone marrow (BM) microenvironment which multiple myeloma (MM) cells depend on for its survival. While primary myeloma cells cannot grow ex vivo in SCID mice, they are able to growth in SCIDhu mice. However, animal models are expensive, time-consuming, and often have limited reproducibility. In addition, classic laboratory models cannot take into consideration the variability of disease in every patient, and the MM patient population is highly variable, both genetically and epigenetically, and the biological characteristics of patients are widely different, which demonstrates sensitivity of individual patients to different therapies. Typical two-dimensional (2D) models rely on a limited number of MM cell lines which cannot reflect the enormous heterogeneity and variations present in individual patients. The goal of this study is to create a patient derived three-dimensional tissue-engineered bone marrow (3DTEBM) culture system based on cross-linked MM-derived BM supernatant, including endogenous soluble growth factors and cytokines, and by incorporating mononuclear cells, including MM (CD138+ population) and accessory cells (CD138- population), from the same patient. METHODS: The 3DTEBMwas formed through calcium cross-linking of BM supernatants combined with the CD138+ and CD138- population from the same MM patient. We tested the growth of fresh primary CD138+ MM cells with CD138- cells in 2D vs patient-derived 3DTEBM cultures based on the original BM ratio (CD138-/CD138+) of the patient at 3 days by flow cytometry. We further selected a patient with not very aggressive tumor and evaluated the effect of CD138- increasing densities on CD138+ MM growth in 3DTEBM cultures at day 3 by flow cytometry. In addition, the effect of patient-derived 3DTEBM and 2D cultures with and without CD138- cells on CD markers expression of the CD138+ population was tested at day 3 by flow cytometry. Finally, the growth of frozen primary CD138+ MM cells with and without CD138- cells in 2D vs patient-derived 3DTEBM cultures was analyzed for 14 days by flow cytometry. RESULTS: We found that patient-derived 3DTEBM cultures support primary MM cell growth. CD138+ MM cells from patients with less aggressive tumors (high BM ratio CD138-/CD138+) showed better growth in 3DTEBM than in 2D systems at day 3. However, more aggressive tumors (low BM ratio CD138-/CD138+), in which MM cells are less dependent of accessory cells, the 3DTEBM cultures showed similar CD138+ MM growth as in 2D cultures. After that, we detected in the patient-derived 3DTEBM cultures a direct correlation between positive and negative fractions, with increased CD138- densities the CD138+ MM growth increased. Next, we analyzed the effect of 3D cultures in CD markers expression and we showed that CD138+ MM cells expressed loss of the plasma cells markers (CD38, CD56, and CD138), reduction of B cells markers (CD19, CD20 and CD22), and increased of the stem cell marker CD34 in 3DTEBM compared to 2D cultures. Finally, patient-derived 3DTEBM supported the growth of frozen primary CD138+ MM cells better than 2D cultures, and the addition of CD138- cells enhanced even more CD138+ MM growth after 14 days in culture. CONCLUSIONS: Patient-derived 3DTEBM cultures support primary fresh and frozen CD138+ MM growth better than classic systems, and induced de-differentiation of MM cells while increased a stem-cell like phenotype. These results highlight the importance of the BM microenvironment (BM supernatant, including endogenous soluble growth factors and cytokines, and the CD138- population from the same patient) to facilitate primary MM cell growth in vitro. Therefore, patient-derived 3DTEBM cultures allowed long-term culture of primary fresh and frozen myeloma cells ex vivo, and these findings indicate that patient-derived 3DTEBM can be utilized for studying multiple myeloma biology and for testing patient-targeted therapy. Disclosures No relevant conflicts of interest to declare.

scholarly journals Whole-genome optical mapping of bone-marrow myeloma cells reveals association of extramedullary multiple myeloma with chromosome 1 abnormalities

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Eva Kriegova ◽  
Regina Fillerova ◽  
Jiri Minarik ◽  
Jakub Savara ◽  
Jirina Manakova ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
High Risk ◽  
Copy Number ◽  
Optical Mapping ◽  
Chromosome 1 ◽  
Gene Copy ◽  
Whole Genome ◽  
Myeloma Cells ◽  
Genomic Architecture

AbstractExtramedullary disease (EMM) represents a rare, aggressive and mostly resistant phenotype of multiple myeloma (MM). EMM is frequently associated with high-risk cytogenetics, but their complex genomic architecture is largely unexplored. We used whole-genome optical mapping (Saphyr, Bionano Genomics) to analyse the genomic architecture of CD138+ cells isolated from bone-marrow aspirates from an unselected cohort of newly diagnosed patients with EMM (n = 4) and intramedullary MM (n = 7). Large intrachromosomal rearrangements (> 5 Mbp) within chromosome 1 were detected in all EMM samples. These rearrangements, predominantly deletions with/without inversions, encompassed hundreds of genes and led to changes in the gene copy number on large regions of chromosome 1. Compared with intramedullary MM, EMM was characterised by more deletions (size range of 500 bp–50 kbp) and fewer interchromosomal translocations, and two EMM samples had copy number loss in the 17p13 region. Widespread genomic heterogeneity and novel aberrations in the high-risk IGH/IGK/IGL, 8q24 and 13q14 regions were detected in individual patients but were not specific to EMM/MM. Our pilot study revealed an association of chromosome 1 abnormalities in bone marrow myeloma cells with extramedullary progression. Optical mapping showed the potential for refining the complex genomic architecture in MM and its phenotypes.

scholarly journals Ex Vivo Activity of Immunotherapeutic Approaches Targeting CD38 Against Daratumumab-Resistant Multiple Myeloma Patient Samples

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 1848-1848
Author(s):  
Maria Karvouni ◽  
Heyue Zhou ◽  
Arnika Kathleen Wagner ◽  
Qiangzhong Ma ◽  
Alamdar H. Baloch ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Nk Cells ◽  
Therapeutic Potential ◽  
Ex Vivo ◽  
Phase 1 ◽  
Employment Equity ◽  
Multiple Myeloma Patient ◽  
Myeloma Cells ◽  
Car T ◽  
Equity Ownership

Background: Multiple myeloma (MM) is a plasma cell malignancy that remains incurable. The identification of CD38, a transmembrane glycoprotein overexpressed on MM cells, led to the development of target-specific therapeutics such as the FDA approved monoclonal antibody (mAb) Daratumumab (DARA). Although a valuable treatment option for refractory/relapsed (R/R) MM patients, DARA has a limited response rate of below 50%, which highlights the clinical need for novel therapeutics. Aims: Aiming to further exploit the therapeutic potential of CD38 in the MM setting, immunotherapies based on the novel anti-CD38 mAb CD38A2 were tested. Methods: For the first approach, the CD38A2 mAb -that binds to a unique, distinct from DARA's, CD38 epitope- was conjugated with either the alkylating agent Duomycin (ADC-136) or the microtubulin binder Duostatin (ADC-129). The ADCs were compared to DARA, in cultures of primary MM cells from patients refractory to DARA treatment. In a second approach, a chimeric antigen receptor (CAR) consisting of the CD38A2 scFv and the intracellular domains of CD28 and CD3ζ was used to transduce primary T and NK cells from R/R MM patients. The functionality of the CAR-T and CAR-NK cells was assessed in cytotoxicity assays against autologous myeloma cells. Results: ADC-136 demonstrated the most potent cytotoxicity against the MM cells with an IC50 of 6pM at day 6 following a single dose treatment. ADC-129 showed cell killing with an IC50 of 30pM, while DARA did not exhibit appreciable cytotoxicity. Regarding the cell therapy approach, patients' T and NK cells were effectively transduced, showing a CD38A2-CAR expression ranging between 11-68%. In functional assays, CAR-T and CAR-NK cells were assayed against autologous myeloma cells, where they exhibited an increase in target cell cytotoxicity, compared to the untransduced cells. Summary/Conclusion: Altogether, our preliminary findings demonstrate that CD38 targeting using CD38A2-based immunotherapies could be a viable therapeutic approach in R/R MM patients previously exposed to DARA. Currently, an anti-CD38 CAR-T therapy based on CD38A2 is being evaluated in Phase 1 studies in R/R MM patients by Sorrento Therapeutics, Inc. Disclosures Zhou: Sorrento Therapeutics Inc: Employment, Equity Ownership. Ma:Sorrento Therapeutics Inc: Employment, Equity Ownership. Zhu:Sorrento Therapeutics Inc: Employment, Equity Ownership. Zhang:Sorrento Therapeutics Inc: Employment, Equity Ownership. Kaufmann:Sorrento Therapeutics, Inc.: Employment, Equity Ownership, Patents & Royalties.

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