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Systemic Lupus Erythematosus (SLE) is a rare, heterogeneous, potentially life-threatening autoimmune disease. Presence of kidney or other major organ (brain, heart or lung) involvement are predictors of poor outcome and in a subset of patients resistant to 1st or 2nd line conventional treatment. The 10-year mortality remains around 10-15 %. Chimeric Antigen Receptors (CAR) are molecules that allow to redirect the engineered immune cells towards specific target antigens and to simultaneously boost their activation. Following breakthrough results observed in the treatment of hematological malignancies, conventional CAR T-cell therapy has recently been applied to refractory SLE patients. Compared to the use of monoclonal antibodies, anti-CD19 CAR T-cells allow to achieve deeper depletion of autoreactive B cells, notably at site of inflamed tissues and lymphoid organs (i.e. lymph node), to suppress interferon signature and to restore the immune tolerance with the reemergence of naïve B-cells with a new repertoire. All clinical data reported in SLE patients so far showed that autologous anti-CD19 CAR T-cell treatment allowed impressive short- and longer-term resolution of lupus nephritis and other severe disease-related manifestations, without major toxicities and only mild cytokine-release syndrome. These clinical effects persisted after B-cell reconstitution and were associated with normalization of double-stranded DNA antibodies and complement levels in drug-free patients until three years after the procedure. Overall, these pioneering experiences show unique clinical and immunological response to CAR T-cell therapy in SLE, and the need for extended follow-up to determine its long-term efficacy., Competing Interests: Declaration of competing interest We have no conflict of interest., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

First-line treatments of autoimmune systemic diseases (ARD) are based on the use of various types of immunosuppressive or immunomodulatory drugs, either alone or in association, according to standardized reference protocols. Prolonged use of these drugs in severe or refractory ARD is associated with high morbidity and increased mortality. Innovative cell therapies represent a new promising approach for patients with ARDs, with the recent clinical use of: a) mesenchymal stromal cells (MSCs), based on their immunomodulatory, antifibrotic and pro-angiogenic properties and b) Chimeric Antigen Receptors (CAR) T cell therapies T lymphocytes, where genetically modified expression of a chimeric antigen receptor (CAR-T cells). Therapeutic use of MSC or CAR-T cells, remains indications of exception in patients with severe ARDs resistant to prior standard therapies with new prerequisite and organisation of health-care pathways as compared to traditional drugs, not only for the Cell and Gene Therapy (CGT) product definition and delivery process, but also for the patient clinical management before and after administration of the CGT product. The aim of this workshop under the auspices of the French Speaking Society of Bone Marrow and Cell transplantation (SFGM-TC) working group on autoimmune diseases (MATHEC) is to describe: a) the prerequisite for French hospitals to set-up the specific health-care pathways for MSC or CART therapy in ARDs patients, in accordance with regulatory and safety needs to perform academic or industry sponsored clinical trials, and b) the care-pathway for ARD patients treated with CGT, highlighting the importance of working in tandem between the ARD and the CAR-T cell specialist all along the indication, procedures and follow-up of ARDs. Patient safety considerations are central to guidance on patient selection to be validated collectively at the multidisciplinary team meeting (MDTM) based on recent (less than 3 months) thorough patient evaluation. MSC and CAR-T procedural aspects and follow-up are then carried out within appropriately experienced and SFGM-TC accredited centres in close collaboration with the ADs specialist., (Copyright © 2024 Société Française du Cancer. Published by Elsevier Masson SAS. All rights reserved.)

Abstract: Follicular lymphoma (FL) is an indolent yet incurable germinal center B-cell lymphoma retaining a characteristic follicular architecture. FL tumor B cells are highly dependent on direct and indirect interactions with a specific and complex tumor microenvironment (TME). Recently, great progress has been made in describing the heterogeneity and dynamics of the FL TME and in depicting how tumor clonal and functional heterogeneity rely on the integration of TME-related signals. Specifically, the FL TME is enriched for exhausted cytotoxic T cells, immunosuppressive regulatory T cells of various origins, and follicular helper T cells overexpressing B-cell and TME reprogramming factors. FL stromal cells have also emerged as crucial determinants of tumor growth and remodeling, with a key role in the deregulation of chemokines and extracellular matrix composition. Finally, tumor-associated macrophages play a dual function, contributing to FL cell phagocytosis and FL cell survival through long-lasting B-cell receptor activation. The resulting tumor-permissive niches show additional layers of site-to-site and kinetic heterogeneity, which raise questions about the niche of FL-committed precursor cells supporting early lymphomagenesis, clonal evolution, relapse, and transformation. In turn, FL B-cell genetic and nongenetic determinants drive the reprogramming of FL immune and stromal TME. Therefore, offering a functional picture of the dynamic cross talk between FL cells and TME holds the promise of identifying the mechanisms of therapy resistance, stratifying patients, and developing new therapeutic approaches capable of eradicating FL disease in its different ecosystems., (© 2024 American Society of Hematology. Published by Elsevier Inc. All rights are reserved, including those for text and data mining, AI training, and similar technologies.)

The rapidly growing field of mesenchymal stromal cell (MSC) basic and translational research requires standardization of terminology and functional characterization. The International Standards Organization's (ISO) Technical Committee (TC) on Biotechnology, working with extensive input from the International Society for Cells and Gene Therapy (ISCT), has recently published ISO standardization documents that are focused on biobanking of MSCs from two tissue sources, Wharton's Jelly, MSC(WJ) and Bone Marrow, MSC(M)), for research and development purposes and development. This manuscript explains the path towards the consensus on the following two documents: the Technical Standard ISO/TS 22859 for MSC(WJ) and the full ISO Standard 24651 for MSC(M) biobanking. The ISO standardization documents are aligned with ISCT's MSC committee position and recommendations on nomenclature because there was active input and incorporation of ISCT MSC committee recommendations in the development of these standards. The ISO standardization documents contain both requirements and recommendations for functional characterization of MSC(WJ) and MSC(M) using a matrix of assays. Importantly, the ISO standardization documents have a carefully defined scope and are meant for research use of culture expanded MSC(WJ) and MSC(M). The ISO standardization documents can be updated in a revision process and will be systematically reviewed after 3-5 years as scientific insights grow. They represent international consensus on MSC identity, definition, and characterization; are rigorous in detailing multivariate characterization of MSCs and represent an evolving-but-important first step in standardization of MSC biobanking and characterization for research use and development., Competing Interests: Declaration of Competing Interest The authors have no commercial, proprietary or financial interest in the products or companies described in this article., (Copyright © 2023 International Society for Cell & Gene Therapy. Published by Elsevier Inc. All rights reserved.)

Purpose: We aimed to investigate the molecular basis of a novel recognizable neurodevelopmental syndrome with scalp and enamel anomalies caused by truncating variants in the last exon of the gene FOSL2, encoding a subunit of the AP-1 complex., Methods: Exome sequencing was used to identify genetic variants in all cases, recruited through Matchmaker exchange. Gene expression in blood was analyzed using reverse transcription polymerase chain reaction. In vitro coimmunoprecipitation and proteasome inhibition assays in transfected HEK293 cells were performed to explore protein and AP-1 complex stability., Results: We identified 11 individuals from 10 families with mostly de novo truncating FOSL2 variants sharing a strikingly similar phenotype characterized by prenatal growth retardation, localized cutis scalp aplasia with or without skull defects, neurodevelopmental delay with autism spectrum disorder, enamel hypoplasia, and congenital cataracts. Mutant FOSL2 messenger RNAs escaped nonsense-mediated messenger RNA decay. Truncated FOSL2 interacts with c-JUN, thus mutated AP-1 complexes could be formed., Conclusion: Truncating variants in the last exon of FOSL2 associate a distinct clinical phenotype by altering the regulatory degradation of the AP-1 complex. These findings reveal a new role for FOSL2 in human pathology., Competing Interests: Conflict of Interest L.A.P.-J. is founding partner and scientific advisor of qGenomics Laboratories. All other authors declare no conflicts of interest., (Copyright © 2022 American College of Medical Genetics and Genomics. All rights reserved.)

The therapeutic potential of culture-adapted adipose-derived stromal cells (ASCs) is largely related to their production of immunosuppressive factors that are inducible in vitro by priming with inflammatory stimuli, in particular tumor necrosis factor-α (TNFα) and interferon-γ (IFNγ). In vivo, obesity is associated with chronic inflammation of white adipose tissue, including accumulation of neutrophils, infiltration by IFNγ/TNFα-producing immune cells, and ASC dysfunction. In the current study, we identified in obese patients a simultaneous upregulation of CD40Lin the adipose tissue stroma vascular fraction (AT-SVF), correlated with the Th1 gene signature, and an overexpression of CD40 by native ASCs. Moreover, activated CD4 + T cells upregulated CD40 on culture-expanded ASCs and triggered their production of IL-8 in a CD40L-dependent manner, leading to an increased capacity to recruit neutrophils. Finally, activation of ASCs by sCD40L or CD40L-expressing CD4 + T cells relies on both canonical and non-canonical NF-κB pathways, and IL-8 was found to be coregulated with NF-κB family members in AT-SVF. These data identify the CD40-CD40L axis as a priming mechanism of ASCs, able to modulate their cross talk with neutrophils in an inflammatory context, and their functional capacity for therapeutic applications., Competing Interests: Conflict-of-interest Disclosure Authors have no competing interests to declare., (Copyright © 2022 International Society for Cell & Gene Therapy. Published by Elsevier Inc. All rights reserved.)

The differentiation of B cells into plasmablasts (PBs) and then plasma cells (PCs) is associated with extensive cell reprogramming and new cell functions. By using specific inhibition strategies (including a novel morpholino RNA antisense approach), we found that early, sustained upregulation of the proviral integrations of Moloney virus 2 (PIM2) kinase is a pivotal event during human B-cell in vitro differentiation and then continues in mature normal and malignant PCs in the bone marrow. In particular, PIM2 sustained the G1/S transition by acting on CDC25A and p27Kip1 and hindering caspase 3-driven apoptosis through BAD phosphorylation and cytoplasmic stabilization of p21Cip1. In PCs, interleukin-6 triggered PIM2 expression, resulting in antiapoptotic effects on which malignant PCs were particularly dependent. In multiple myeloma, pan-PIM and myeloid cell leukemia-1 (MCL1) inhibitors displayed synergistic activity. Our results highlight a cell-autonomous function that links kinase activity to the newly acquired secretion ability of the PBs and the adaptability observed in both normal and malignant PCs. These findings should finally prompt the reconsideration of PIM2 as a therapeutic target in multiple myeloma., (© 2022 by The American Society of Hematology.)

The Cellular Therapy Coding and Labeling Advisory Group of the International Council for Commonality in Blood Banking Automation and the International Society for Cell & Gene Therapy mesenchymal stromal cell (MSC) committee are providing specific recommendations on abbreviating tissue sources of culture-adapted MSCs. These recommendations include using abbreviations based on the ISBT 128 terminology model that specifies standard class names to distinguish cell types and tissue sources for culture-adapted MSCs. Thus, MSCs from bone marrow are MSC(M), MSCs from cord blood are MSC(CB), MSCs from adipose tissue are MSC(AT) and MSCs from Wharton's jelly are MSC(WJ). Additional recommendations include using these abbreviations through the full spectrum of pre-clinical, translational and clinical research for the development of culture-adapted MSC products. This does not apply to basic research focused on investigating the developmental origins, identity or functionalities of endogenous progenitor cells in different tissues. These recommendations will serve to harmonize nomenclature in describing research and development surrounding culture-adapted MSCs, many of which are destined for clinical and/or commercial translation. These recommendations will also serve to align research and development efforts on culture-adapted MSCs with other cell therapy products., (Copyright © 2021 International Society for Cell & Gene Therapy. Published by Elsevier Inc. All rights reserved.)

CAR-T cells have recently made a stunning entry on the arena of immunotherapy of B-cell lymphomas. This new treatment approach represents the culmination of 30 years of efforts to understand the role of T cells in the antitumor response. However, this technology is still in its infancy and suffers from a number of limitations. Many areas for improvement, based in particular on the possibilities of additional genetic manipulations of CAR-T cells, aim at reducing their toxicity, increasing their persistence in vivo, preventing the risk of tumor escape, recruiting other immune effectors, or extending their application to other cancers. Further studies of the dynamic interaction between the patient and these live drugs will allow elucidating the mechanisms determining the antitumor response in this context and thus developing more efficiently the future CAR-T cells., (Copyright © 2021 Société Française du Cancer. Published by Elsevier Masson SAS. All rights reserved.)

Follicular lymphoma (FL) originates in the lymph nodes (LNs) and infiltrates bone marrow (BM) early in the course of the disease. BM FL B cells are characterized by a lower cytological grade, decreased proliferation, and a specific phenotypic and subclonal profile. Mesenchymal stromal cells (MSCs) obtained from FL BM display a specific gene expression profile (GEP), including enrichment for a lymphoid stromal cell signature, and an increased capacity to sustain FL B-cell growth. However, the mechanisms triggering the formation of the medullar FL permissive stromal niche have not been identified. In the current work, we demonstrate that FL B cells produce extracellular vesicles (EVs) that can be internalized by BM-MSCs, making them more efficient to support FL B-cell survival and quiescence. Accordingly, EVs purified from FL BM plasma activate transforming growth factor β-dependent and independent pathways in BM-MSCs and modify their GEP, triggering an upregulation of factors classically associated with hematopoietic stem cell niche, including CXCL12 and angiopoietin-1. Moreover, we provide the first characterization of BM FL B-cell GEP, allowing the definition of the landscape of molecular interactions they could engage with EV-primed BM-MSCs. This work identifies FL-derived EVs as putative mediators of BM stroma polarization and supports further investigation of their clinical interest for targeting the crosstalk between BM-MSCs and malignant B cells., (© 2021 by The American Society of Hematology.)

The International Society for Cell & Gene Therapy mesenchymal stromal cell (MSC) committee has been an interested observer of community interests in all matters related to MSC identity, mechanism of action, potency assessment and etymology, and it has regularly contributed to this conversation through a series of MSC pre-conferences and committee publications dealing with these matters. Arising from these reflections, the authors propose that an overlooked and potentially disruptive perspective is the impact of in vivo persistence on potency that is not predicted by surrogate cellular potency assays performed in vitro and how this translates to in vivo outcomes. Systemic delivery or extravascular implantation at sites removed from the affected organ system seems to be adequate in affecting clinical outcomes in many pre-clinical murine models of acute tissue injury and inflammatory pathology, including the recent European Medicines Agency-approved use of MSCs in Crohn-related fistular disease. The authors further propose that MSC viability and metabolic fitness likely dominate as a potency quality attribute, especially in recipients poised for salutary benefits as defined by emerging predictive biomarkers of response., (Copyright © 2021 International Society for Cell & Gene Therapy. Published by Elsevier Inc. All rights reserved.)

Systemic sclerosis (SSc) is a rare chronic autoimmune disease characterized by vasculopathy, dysregulation of innate and adaptive immune responses, and progressive fibrosis. SSc remains an orphan disease, with high morbity and mortality in SSc patients. The mesenchymal stromal cells (MSC) demonstrate in vitro and in vivo pro-angiogenic, immuno-suppressive, and anti-fibrotic properties and appear as a promising stem cell therapy type, that may target the key pathological features of SSc disease. This review aims to summarize acquired knowledge in the field of :1) MSC definition and in vitro and in vivo functional properties, which vary according to the donor type (allogeneic or autologous), the tissue sources (bone marrow, adipose tissue or umbilical cord) or inflammatory micro-environment in the recipient; 2) preclinical studies in various SSc animal models , which showed reduction in skin and lung fibrosis after MSC infusion; 3) first clinical trials in human, with safety and early efficacy results reported in SSc patients or currently tested in several ongoing clinical trials., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Ever since their discovery in 2001, adipose mesenchymal stromal cells (ASC) have profoundly modified clinical indications and our practice of plastic surgery, thereby placing our discipline at the forefront of regenerative medicine. These cells act through paracrine signaling by synthesizing immunosuppressive and pro-angiogenic factors. They are of key importance with regard to the regenerative properties of autologously grafted adipose tissue (AT). Taken together, they make up the stromal vascular fraction (SVF) comprising all AT cells except for adipocytes. As our knowledge evolves, we are moving from fat grafting towards SVF grafting, of which the essential sought-after effect is tissue regeneration. The objective of the present review is to synthesize present-day information on ASCs and their immunomodulatory properties and, from a practical standpoint, to indicate present-day and future steps towards establishment of clinical routine, particularly through application of techniques favoring mechanical digestion of adipose tissue., (Copyright © 2018 Elsevier Masson SAS. All rights reserved.)

PREDICTIVE BIOMARKERS OF RESPONSE TO IMMUNE CHECKPOINT INHIBITORS: PD-1 checkpoint inhibitors are becoming the reference treatment for several types of cancers. Many patients show remarkable efficacy and low toxicity. However, some patients have a better outcome than others with PD-1 checkpoint inhibitors. So, it is crucial to identify biomarkers of response. We review here the available data of several potential biomarkers of efficacy. The expression of PD-L1, detected by immunohistochemistry on tumor cells and immune cells is a good predictive biomarker of response for some cancers; however, this method is not standardized, and there are different antibodies, different cut-off values, and different targets (tumor or microenvironment). Moreover, the expression of PD-L1 is dynamic and heterogeneous within the tumor: expression is discordant between primary tumor and metastasis or between biopsy and surgical specimen. Peripheral blood lymphocytes also can be informative, especially the baseline neutrophil to lymphocyte ratio which is easy to measure in daily practice. High rate of neoantigens is also associated with improved response. Therefore, mutation burden can be predictive of response and this explains why tumors with microsatellites instability have an enhanced response. Similarly, genetic signatures are linked with resistance or response to treatment. Gut microbiota is associated with improved antitumor immune response although the underlying mechanism is not well understood so far. Lastly, it seems that cytokines, mediators of immunity may play a role in the response to immunotherapy and so, constitute an interesting biomarker. Several potential biomarkers are identified but none is prospectively validated so far., (© 2018 Société Française du Cancer. Publié par Elsevier Masson SAS. Tous droits réservés.)

Obinutuzumab is a type II anti-CD20 monoclonal antibody that enhances antibody-dependent cellular cytotoxicity better than rituximab. Given promising results with lenalidomide and rituximab, this phase 1b study assessed the safety and efficacy of lenalidomide combined with obinutuzumab (GALEN). Patients age ≥18 years with relapsed or refractory (R/R) follicular lymphoma (FL) after rituximab-containing therapy received escalating doses (10 [n = 7], 15 [n = 3], 20 [n = 6], and 25 mg [n = 3]) of daily oral lenalidomide on days 1 to 21 of cycle 1 and on days 2 to 22 of cycles 2 to 6 (28-day cycles). Obinutuzumab 1000 mg IV was administered on days 8, 15, and 22 (cycle 1) and on day 1 (cycles 2-6). Dose was escalated in a 3 + 3 design based on dose-limiting toxicity (DLT) during cycle 1 to establish the maximum tolerated dose (MTD). We observed 164 adverse events (AEs), of which 139 were grade 1/2. The most common AEs were constipation (52.6%), neutropenia (47.4%), and asthenia (36.8%); 64.3% (9 of 14) of the grade 3/4 AEs were neutropenia/neutrophil decrease, but without any febrile neutropenia. Four DLTs occurred in 2 patients, all deemed unrelated to treatment. MTD was not reached. Twelve patients (63.2%) responded: 8 complete, 3 unconfirmed complete, and 1 partial response. Oral lenalidomide plus obinutuzumab is well tolerated and effective in R/R FL. The recommended dose of lenalidomide was established at 20 mg based on the risk of grade 3/4 neutropenia from cycle 2. This trial was registered at www.clinicaltrials.gov as #NCT01582776., (© 2018 by The American Society of Hematology.)

Follicular lymphoma (FL) is the most frequent indolent lymphoma and is characterized by the accumulation of germinal center-derived malignant B cells engaged in a bidirectional crosstalk with their supportive microenvironment in invaded lymph nodes (LNs) and bone marrow (BM). T follicular helper (T FH ) cells and infiltrating stromal cells have been shown to favor FL B-cell growth, but the mechanisms of their protumoral effect and how the LN/BM microenvironment is converted into a lymphoma-permissive cell niche remain poorly understood. We demonstrated here that FL-infiltrating LN and BM stromal cells overexpressed CXCL12 in situ. Interleukin-4 high (IL-4 hi ) FL-T FH cells, unlike FL B cells themselves, triggered CXCL12 upregulation in human stromal cell precursors. In agreement, expression of CXCL12 was associated with IL-4 expression and signaling within the FL BM and LN niches. This IL-4/CXCL12 axis was amplified in activated lymphoid stromal cells as shown in our in vitro model of human lymphoid stroma differentiation and in an inducible mouse model of ectopic lymphoid organ formation. Finally, CXCL12 triggered primary FL B-cell activation, migration, and adhesion, a process antagonized by BTK and PI3K inhibitors. These data identified the IL-4/CXCL12 loop as a previously unrecognized pathway involved in lymphoid stroma polarization and as a potential therapeutic target in FL patients., (© 2017 by The American Society of Hematology.)

In diffuse large B-cell lymphoma (DLBCL), the number of circulating monocytes and neutrophils represents an independent prognostic factor. These cell subsets include monocytic and granulocytic myeloid-derived suppressor cells (M- and G-MDSCs) defined by their ability to suppress T-cell responses. MDSCs are a heterogeneous population described in inflammatory and infectious diseases and in numerous tumors including multiple myeloma, chronic lymphocytic leukemia, and DLBCL. However, their mechanisms of action remain unclear. We broadly assessed the presence and mechanisms of suppression of MDSC subsets in DLBCL. First, a myeloid suppressive signature was identified by gene expression profiling in DLBCL peripheral blood. Accordingly, we identified, in a cohort of 66 DLBCL patients, an increase in circulating G-MDSC (Lin(neg)HLA-DR(neg)CD33(pos)CD11b(pos)) and M-MDSC (CD14(pos)HLA-DR(low)) counts. Interestingly, only M-MDSC number was correlated with the International Prognostic Index, event-free survival, and number of circulating Tregs. Furthermore, T-cell proliferation was restored after monocyte depletion. Myeloid-dependent T-cell suppression was attributed to a release of interleukin-10 and S100A12 and increased PD-L1 expression. In summary, we identified expanded MDSC subsets in DLBCL, as well as new mechanisms of immunosuppression in DLBCL., (© 2016 by The American Society of Hematology.)

The mechanisms regulating megakaryopoiesis and platelet production (thrombopoiesis) are still incompletely understood. Identification of a progenitor with enhanced thrombopoietic capacity would be useful to decipher these mechanisms and to improve our capacity to produce platelets in vitro. Differentiation of peripheral blood CD34(+) cells in the presence of bone marrow-human mesenchymal stromal cells (MSCs) enhanced the production of proplatelet-bearing megakaryocytes (MKs) and platelet-like elements. This was accompanied by enrichment in a MK precursor population exhibiting an intermediate level of CD41 positivity while maintaining its expression of CD34. Following sorting and subculture with MSCs, this CD34(+)CD41(low) population was able to efficiently generate proplatelet-bearing MKs and platelet-like particles. Similarly, StemRegenin 1 (SR1), an antagonist of the aryl hydrocarbon receptor (AhR) transcription factor known to maintain CD34 expression of progenitor cells, led to an enriched CD34(+)CD41(low) fraction and to an increased capacity to generate proplatelet-producing MKs and platelet-like elements ultrastructurally and functionally similar to circulating platelets. The effect of MSCs, like that of SR1, appeared to be mediated by an AhR-dependent mechanism because both culture conditions resulted in repression of its downstream effector CYP1B1. This newly described isolation of a precursor exhibiting strong MK potential could be exploited to study normal and abnormal thrombopoiesis and for in vitro platelet production., (© 2016 by The American Society of Hematology.)

Follicular lymphoma (FL) results from the accumulation of malignant germinal center (GC) B cells leading to the development of an indolent and largely incurable disease. FL cells remain highly dependent on B-cell receptor (BCR) signaling and on a specific cell microenvironment, including T cells, macrophages, and stromal cells. Importantly, FL BCR is characterized by a selective pressure to retain surface immunoglobulin M (IgM) BCR despite an active class-switch recombination process, and by the introduction, in BCR variable regions, of N-glycosylation acceptor sites harboring unusual high-mannose oligosaccharides. However, the relevance of these 2 FL BCR features for lymphomagenesis remains unclear. In this study, we demonstrated that IgM(+) FL B cells activated a stronger BCR signaling network than IgG(+) FL B cells and normal GC B cells. BCR expression level and phosphatase activity could both contribute to such heterogeneity. Moreover, we underlined that a subset of IgM(+) FL samples, displaying highly mannosylated BCR, efficiently bound dendritic cell-specific intercellular adhesion molecule-3-grabbing nonintegrin (DC-SIGN), which could in turn trigger delayed but long-lasting BCR aggregation and activation. Interestingly, DC-SIGN was found within the FL cell niche in situ. Finally, M2 macrophages induced a DC-SIGN-dependent adhesion of highly mannosylated IgM(+) FL B cells and triggered BCR-associated kinase activation. Interestingly, pharmacologic BCR inhibitors abolished such crosstalk between macrophages and FL B cells. Altogether, our data support an important role for DC-SIGN-expressing infiltrating cells in the biology of FL and suggest that they could represent interesting therapeutic targets., (© 2015 by The American Society of Hematology.)

In follicular lymphoma (FL), follicular helper T cells (TFH) have been depicted as one of the main components of the malignant B-cell niche and a promising therapeutic target. Although defined by their capacity to sustain FL B-cell growth together with specific gene expression and cytokine secretion profiles, FL-TFH constitute a heterogeneous cell population. However, specific markers reflecting such functional heterogeneity are still lacking. In this study, we demonstrate that CD10 identifies a subset of fully functional germinal center TFH in normal secondary lymphoid organs. Importantly, this subset is amplified in the FL context, unlike in other B-cell lymphomas with a follicular growth pattern. Furthermore, whereas FL-TFH produce high levels of interleukin (IL)-21 and low levels of IL-17 irrespectively of their CD10 expression, CD10(pos) FL-TFH specifically exhibit an IL-4(hi)IFN-γ(lo)TNF-α(hi) cytokine profile associated with a high capacity to sustain directly and indirectly malignant B-cell survival. Altogether, our results highlight the important role of this novel functional subset in the FL cell niche., (© 2015 by The American Society of Hematology.)

Over the last decade, the clinical use of adipose-derived stromal/stem cells (ASC) in regenerative medicine is rapidly increasing. ASC belong to the mesenchymal stromal cells initially obtained from the bone marrow. Their limited differentiation capacity in vivo into functional mature cells has led to a reassessment of their mechanisms of action. One of the major clinical interests appears related to paracrine effects through a temporary production of trophic and immunomodulatory factors. Our purpose is to provide a review on the latest knowledge in the field of ASC, mechanisms of action, mainly immunomodulatory/immunosuppressive properties, methods of obtention, with a focus on clinical perspectives particularly in the field of cellular therapy and fat grafting technique in plastic surgery., (Copyright © 2014 Elsevier Masson SAS. All rights reserved.)

Follicular lymphoma (FL) is a B-cell neoplasm resulting from the transformation of germinal center (GC) B cells. Although t(14;18) and ectopic B-cell lymphoma 2 (BCL2) expression constitute the genetic hallmark of FL, t(14;18)(pos) B cells bearing genotypic and phenotypic features of FL cells can be found in the blood of most healthy individuals. Nevertheless, the localization of these FL-like cells (FLLCs) in nonmalignant GC-rich tissues and the functional consequences of BCL2 overexpression have not been evaluated thus far. Among 85 reactive lymph node (RLN) samples, 14% were found to contain high levels of t(14;18) by quantitative polymerase chain reaction. In t(14;18)(hi) RLNs, CD20(pos)BCL2(pos)CD10(pos) FLLCs consistently accumulated within the GC, essentially as nonproliferative CXCR4(neg) centrocytes. Moreover, they displayed a reduced response to proliferative stimuli in vitro. Altogether, our findings provide new insights into in situ FLLC functional properties and suggest that these cells have not acquired the ultimate genetic events leading to FL transformation., (© 2014 by The American Society of Hematology.)

Antigen receptor-mediated nuclear factor κB (NF-κB) activation relies on the formation of a large multi-protein complex that contains CARMA1, BCL10, and MALT1 (CBM complex). This signalosome is pirated in the activated B-cell-like subgroup of diffuse large B-cell lymphoma (ABC DLBCL) to drive aberrant NF-κB activation, thereby promoting cell survival and propagation. Using an unbiased proteomic approach, we screened for additional components of the CBM in lymphocytes. We found that the linear ubiquitin chain assembly complex (LUBAC), which was previously linked to cytokine-mediated NF-κB activation, dynamically integrates the CBM and marshals NF-κB optimal activation following antigen receptor ligation independently of its catalytic activity. The LUBAC also participates in preassembled CBM complex in cells derived from ABC DLBCL. Silencing the LUBAC reduced NF-κB activation and was toxic in ABC DLBCL cell lines. Thus, our findings reveal a role for the LUBAC during lymphocyte activation and in B-cell malignancy.

Operationally tolerant patients (TOL) display a higher number of blood B cells and transcriptional B cell signature. As they rarely develop an allo-immune response, they could display an abnormal B cell differentiation. We used an in vitro culture system to explore T-dependent differentiation of B cells into plasma cells. B cell phenotype, apoptosis, proliferation, cytokine, immunoglobulin production and markers of differentiation were followed in blood of these patients. Tolerant recipients show a higher frequency of CD20(+) CD24(hi) CD38(hi) transitional and CD20(+) CD38(lo) CD24(lo) naïve B cells compared to patients with stable graft function, correlating with a decreased frequency of CD20(-) CD38(+) CD138(+) differentiated plasma cells, suggestive of abnormal B cell differentiation. B cells from TOL proliferate normally but produce more IL-10. In addition, B cells from tolerant recipients exhibit a defective expression of factors of the end step of differentiation into plasma cells and show a higher propensity for cell death apoptosis compared to patients with stable graft function. This in vitro profile is consistent with down-regulation of B cell differentiation genes and anti-apoptotic B cell genes in these patients in vivo. These data suggest that a balance between B cells producing IL-10 and a deficiency in plasma cells may encourage an environment favorable to the tolerance maintenance., (© Copyright 2013 The American Society of Transplantation and the American Society of Transplant Surgeons.)

Cultured mesenchymal stromal cells (MSCs) possess immune regulatory properties and are already used for clinical purposes, although preclinical data (both in vitro and in vivo in animal models) are not always homogeneous and unequivocal. However, the various MSC-based clinical approaches to treat immunological diseases would be significantly validated and strengthened by using standardized immune assays aimed at obtaining shared, reproducible and consistent data. Thus, the MSC Committee of the International Society for Cellular Therapy has decided to put forward for general discussion a working proposal for a standardized approach based on a critical view of literature data., (Copyright © 2013 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.)

In the past decade, the therapeutic value of mesenchymal stromal cells (MSCs) has been studied in various indications, thereby taking advantage of their immunosuppressive properties. Easy procurement from bone marrow, adipose tissue or other sources and conventional in vitro expansion culture have made their clinical use attractive. Bridging the gap between current scientific knowledge and regulatory prospects on the transformation potential and possible tumorigenicity of MSCs, the Cell Products Working Party and the Committee for Advanced Therapies organized a meeting with leading European experts in the field of MSCs. This meeting elucidated the risk of potential tumorigenicity related to MSC-based therapies from two angles: the scientific perspective and the regulatory point of view. The conclusions of this meeting, including the current regulatory thinking on quality, nonclinical and clinical aspects for MSCs, are presented in this review, leading to a clearer way forward for the development of such products., (Copyright © 2013 International Society for Cellular Therapy. All rights reserved.)

Accumulating evidence indicates that infiltrating stromal cells contribute directly and indirectly to tumor growth in a wide range of cancers. In follicular lymphoma (FL), malignant B cells are found admixed with heterogeneous lymphoid-like stromal cells within invaded lymph nodes and BM. In addition, mesenchymal stromal cells (MSCs) support in vitro FL B-cell survival, in particular after their engagement toward lymphoid differentiation. We show here that BM-MSCs obtained from patients with FL (FL-MSCs) display a specific gene expression profile compared with MSCs obtained from healthy age-matched donors (HD-MSCs). This FL-MSC signature is significantly enriched for genes associated with a lymphoid-like commitment. Interestingly, CCL2 could be detected at a high level within the FL-cell niche, is up-regulated in HD-MSCs by coculture with malignant B cells, and is overexpressed by FL-MSCs, in agreement with their capacity to recruit monocytes more efficiently than HD-MSCs. Moreover, FL-MSCs and macrophages cooperate to sustain malignant B-cell growth, whereas FL-MSCs drive monocyte differentiation toward a proangiogenic and lipopolysaccharide-unresponsive phenotype close to that of tumor-associated macrophages. Altogether, these results highlight the complex role of FL stromal cells that promote direct tumor B-cell growth and orchestrate FL-cell niche, thus emerging as a potential therapeutic target in this disease.

Background Aims: Mesenchymal stromal cells (MSC) are the focus of research in regenerative medicine aiming at the regulatory approval of these cells for specific indications. To cope with the regulatory requirements for somatic cell therapy, novel approaches that do not interfere with the natural behavior of the cells are necessary. In this context in vivo magnetic resonance imaging (MRI) of labeled MSC could be an appropriate tool. Cell labeling for MRI with a variety of different iron oxide preparations is frequently published. However, most publications lack a comprehensive assessment of the non-interference of the contrast agent with the functionality of the labeled MSC, which is a prerequisite for the validity of cell-tracking via MRI., Methods: We studied the effects of iron oxide-poly(l-lactide) nanoparticles in MSC with flow cytometry, transmission electron microscopy (TEM), confocal laser scanning microscopy (CLSM), Prussian blue staining, CyQuant® proliferation testing, colony-forming unit-fibroblast (CFU-F) assays, flow chamber adhesion testing, immunologic tests and differentiation tests. Furthermore iron-labeled MSC were studied by MRI in agarose phantoms and Wistar rats., Results: It could be demonstrated that MSC show rapid uptake of nanoparticles and long-lasting intracellular persistence in the endosomal compartment. Labeling of the MSC with these particles has no influence on viability, differentiation, clonogenicity, proliferation, adhesion, phenotype and immunosuppressive properties. They show excellent MRI properties in agarose phantoms and after subcutaneous implantation in rats over several weeks., Conclusions: These particles qualify for studying MSC homing and trafficking via MRI.

Clinical-grade human mesenchymal stromal cells (MSCs) have been expanded in vitro for tissue engineering or immunoregulatory purposes without standardized culture conditions or release criteria. Although human MSCs show poor susceptibility for oncogenic transformation, 2 recent studies described their capacity to accumulate chromosomal instability and to give rise to carcinoma in immunocompromised mice after long-term culture. We thus investigated the immunologic and genetic features of MSCs expanded with fetal calf serum and fibroblast growth factor or with platelet lysate in 4 cell-therapy facilities during 2 multicenter clinical trials. Cultured MSCs showed a moderate expression of human leukocyte antigen-DR without alteration of their low immunogenicity or their immunomodulatory capacity. Moreover, some transient and donor-dependent recurring aneuploidy was detected in vitro, independently of the culture process. However, MSCs with or without chromosomal alterations showed progressive growth arrest and entered senescence without evidence of transformation either in vitro or in vivo.

Accumulating evidence indicates that the cellular microenvironment plays a key role in follicular lymphoma (FL) pathogenesis, both within tumor lymph nodes (LNs) and in infiltrated bone marrow where ectopic LN-like reticular cells are integrated within malignant B-cell nodular aggregates. In normal secondary lymphoid organs, specific stromal cell subsets provide a highly specialized microenvironment that supports immune response. In particular, fibroblastic reticular cells (FRCs) mediate immune cell migration, adhesion, and reciprocal interactions. The role of FRCs and their postulated progenitors, that is, bone marrow mesenchymal stem cells (MSCs), in FL remains unexplored. In this study, we investigated the relationships between FRCs and MSCs and their capacity to sustain malignant B-cell growth. Our findings strongly suggest that secondary lymphoid organs contain MSCs able to give rise to adipocytes, chondrocytes, osteoblasts, as well as fully functional B-cell supportive FRCs. In vitro, bone marrow-derived MSCs acquire a complete FRC phenotype in response to a combination of tumor necrosis factor-alpha and lymphotoxin-alpha1beta2. Moreover, MSCs recruit primary FL cells that, in turn, trigger their differentiation into FRCs, making them able to support malignant B-cell survival. Altogether, these new insights into the cross talk between lymphoma cells and their microenvironment could offer original therapeutic strategies.

Identification of growth factors in neoplasias may be a target for future therapies by blocking either growth factor receptor interaction or the induced pathway. Using gene expression profiling, we identified overexpression of 2 receptors for a proliferation-inducing ligand (APRIL) and B-cell activating factor (BAFF) in malignant plasma cells compared with normal plasma cells. APRIL and BAFF are involved in a variety of tumor and autoimmune diseases, including B-cell malignancies. We confirmed the expression of BAFF and APRIL receptors (B-cell maturation antigen [BCMA], transmembrane activator and calcium modulator and cyclophilin ligand interactor [TACI], and BAFF-R) in a majority of 13 myeloma cell lines and in the purified primary myeloma cells of 11 patients. APRIL and BAFF were potent survival factors for exogenous cytokine-dependent myeloma cell lines and were autocrine growth factors for the RPMI8226 and L363 autonomously growing cell lines. These factors activated nuclear factor (NF)-kappaB, phosphatidylinositol-3 (PI-3) kinase/AKT, and mitogen-activated protein kinase (MAPK) kinase pathways and induced a strong up-regulation of the Mcl-1 and Bcl-2 antiapoptotic proteins in myeloma cells. BAFF or APRIL was also involved in the survival of primary myeloma cells cultured with their bone-marrow environment, and protected them from dexamethasone (DEX)-induced apoptosis. Finally, the serum levels of BAFF and APRIL were increased about 5-fold in patients with multiple myeloma (MM) as compared with healthy donors. Altogether, these data suggest that APRIL/BAFF inhibitors may be of clinical value in MM.

Plasma cells (PCs), the end point of B-cell differentiation, are a heterogeneous cell compartment comprising several cell subsets from short-lived highly proliferative plasmablasts to long-lived nondividing fully mature PCs. Whereas the major transcription factors driving the differentiation of B cells to PCs were recently identified, the subtle genetic changes that underlie the transition from plasmablasts to mature PCs are poorly understood. We recently described an in vitro model making it possible to obtain a large number of cells with the morphologic, phenotypic, and functional characteristics of normal polyclonal plasmablastic cells (PPCs). Using Affymetrix microarrays we compared the gene expression profiles of these PPCs with those of mature PCs isolated from tonsils (TPCs) and bone marrow (BMPCs), and with those of B cells purified from peripheral blood (PBB cells) and tonsils (TBCs). Unsupervised principal component analysis clearly distinguished the 5 cell populations on the basis of their differentiation and proliferation status. Detailed statistical analysis allowed the identification of 85 PC genes and 40 B-cell genes, overexpressed, respectively, in the 3 PC subsets or in the 2 B-cell subsets. In addition, several signaling molecules and antiapoptotic proteins were found to be induced in BMPCs compared with PPCs and could be involved in the accumulation and prolonged survival of BMPCs in close contact with specialized stromal microenvironment. These data should help to better understand the molecular events that regulate commitment to a PC fate, mediate PC maintenance in survival niches, and could facilitate PC immortalization in plasma cell dyscrasias.

A new way to identify tumor-specific genes is to compare gene expression profiles between malignant cells and their autologous normal counterparts. In patients with multiple myeloma, a major plasma cell disorder, normal plasma cells are not easily attainable in vivo. We report here that in vitro differentiation of peripheral blood B lymphocytes, purified from healthy donors and from patients with multiple myeloma, makes it possible to obtain a homogeneous population of normal plasmablastic cells. These cells were identified by their morphology, phenotype, production of polyclonal immunoglobulins, and expression of major transcription factors involved in B-cell differentiation. Oligonucleotide microarray analysis shows that these polyclonal plasmablastic cells have a gene expression pattern close to that of normal bone marrow-derived plasma cells. Detailed analysis of genes statistically differentially expressed between normal and tumor plasma cells allows the identification of myeloma-specific genes, including oncogenes and genes coding for tumor antigens. These data should help to disclose the molecular mechanisms of myeloma pathogenesis and to define new therapeutic targets in this still fatal malignancy. In addition, the comparison of gene expression between plasmablastic cells and B cells provides a new and powerful tool to identify genes specifically involved in normal plasma cell differentiation.

In multiple myeloma (MM), the growth of primary plasma cells depends not only on interleukin-6 (IL-6), but also on additional unidentified signals delivered by the bone marrow environment. Using Atlas complementary DNA (cDNA) arrays comprising 268 genes coding for intercellular signaling molecules, this study identified genes that are overexpressed in myeloma cells compared to autologous B-lymphoblastoid cell lines. These genes encode the oncogenic Tyro3 tyrosine kinase receptor, the heparin-binding epidermal growth factor-like growth factor (HB-EGF) that is an epithelial autocrine tumor growth factor, the thrombin receptor (TR) that is linked to HB-EGF and syndecan-1 processing and to cell invasion, chemokine receptors CCR1 and CCR2, the Wnt pathway actor Frizzled-related protein (FRZB), and the Notch receptor ligand Jagged 2. These data, obtained with the Atlas cDNA array, were confirmed by reverse transcriptase-polymerase chain reaction or protein analysis or both. Furthermore, Tyro3, HB-EGF, TR, and FRZB gene expression was documented in purified primary malignant plasma cells from patients with plasma cell leukemia or MM. HB-EGF and FRZB were poorly expressed in purified polyclonal plasma cells. Finally, HB-EGF was proved to be an essential autocrine growth factor for the XG-1 myeloma cells. This study shows the potency and the biologic relevance of cDNA arrays used to analyze simultaneously a large panel of intercellular signaling genes and, by identifying several genes overexpressed in malignant plasma cells, opens new fields of investigation in MM biology. (Blood. 2001;98:771-780)

Agonist antihuman gp130 transducer monoclonal antibodies (MoAbs) were used in SCID mice to grow myeloma cells whose survival and proliferation is dependent on gp130 transducer activation. The agonist anti-gp130 MoAbs neither bound to murine gp130 nor activated murine cells and, as a consequence, did not induce interleukin-6 (IL-6)-related toxicities in mice. They have a 2-week half-life in vivo when injected in the peritoneum. The agonist antibodies made possible the in vivo growth of exogenous IL-6-dependent human myeloma cells as well as that of freshly explanted myeloma cells from 1 patient with secondary plasma cell leukemia. Tumors occurred 4 to 10 weeks after myeloma cell graft and weighed 3 to 5 g. They grew as solid tumors in the peritoneal cavity and metastasized to the different peritoneal organs: liver, pancreas, spleen, and intestine. Tumoral cells were detected in blood and bone marrow of mice grafted with the XG-2 myeloma cells. Tumoral cells grown in SCID mice had kept the phenotypic characteristics of the original tumoral cells and their in vitro growth required the presence of IL-6 or agonist anti-gp130 MoAbs. Myeloma cells from 4 patients with medullary involvement persisted for more than 1 year as judged by detectable circulating human Ig. However, no tumors were detected, suggesting a long-term survival of human myeloma cells without major proliferation. These observations paralleled those made in in vitro cultures as well as the tumor growth pattern in these patients. This gp130 transducer-dependent SCID model of multiple myeloma should be useful to study various therapeutical approaches in multiple myeloma in vivo.

Bone marrow dendritic cells (DC) from patients with multiple myeloma (MM) were recently reported to be infected with Kaposi's sarcoma-associated herpesvirus (KSHV). Because immunotherapy strategies using DC are very promising in this disease, we looked for KSHV DNA in clinical-grade DC generated in vitro from MM patients. Adherent apheresis cells from MM patients were maintained for 7 days in clinical-grade X-VIVO 15 culture medium supplemented with granulocyte-macrophage colony-stimulating factor, interleukin-4, or interleukin-13. Tumor necrosis factor alpha was added for the last 2 days. We obtained a cell population with a DC phenotype able to endocytose fluorescein isothiocyanate (FITC)-dextran and efficiently activate resting allogenic T lymphocytes. To detect KSHV DNA, we used polymerase chain reaction (PCR) followed by Southern blotting of PCR product with a sensitivity detecting a few copies of viral DNA. All the PCR were repeated in a blinded fashion three times, on 1 mug and 0.2 mug of genomic DNA, in two different laboratories. Clinical-grade DC from 10 (91%) of 11 patients were not infected with KSHV. The apheresis cells and the purified CD34(+) cells from the same patients were also negative. A very weak PCR band was detected with DC from one patient, but the initial apheresis cells were negative. The detection of KSHV infection in 1 (9%) of 11 MM patients probably represents background seroprevalence. It seems likely that functional and clinical-grade DC from MM patients can safely be used in clinical trials.

Defects in immune response are often reported in patients with multiple myeloma (MM). Because dendritic cells (DCs) are key effectors in promoting cellular immunity and are potential vectors for immunotherapy, we have evaluated the ability of MM patients' apheresis cells to generate DCs in short-term cultures. We report here the obtaining of a virtually pure population of DCs (89.7% +/- 6%, n = 18) after culturing adherent apheresis cells for 7 days with granulocyte-macrophage colony-stimulating factor (GM-CSF ) and interleukin-4 (IL-4). These cells exhibited all the phenotypic characteristics (CD1a+, HLA-DR+, CD80+, CD40+, CD14-) and the MLR stimulating capacity of mature DCs. The number of DCs reached 12. 1% of the initial apheresis cell number put into culture. As DC precursors involved in this model were CD34(-) cells, the unabsorbed cells resulting from clinical-grade CD34 purification were a reliable source of DCs, even after freezing. The proliferation of DC precursors could be increased 10-fold by adding IL-3 and tumor necrosis factor-alpha together with GM-CSF and IL-4. Thus, CD34- apheresis cells from patients with MM offer an interesting source for generating pure, functional, and potentially proliferating DCs.