236 results on '"Martino Introna"'
Search Results
2. Functional Activity of Cytokine-Induced Killer Cells Enhanced by CAR-CD19 Modification or by Soluble Bispecific Antibody Blinatumomab
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Silvia Zaninelli, Silvia Panna, Sarah Tettamanti, Giusi Melita, Andrea Doni, Francesca D’Autilia, Rut Valgardsdottir, Elisa Gotti, Alessandro Rambaldi, Josée Golay, and Martino Introna
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chimeric antigen receptor ,bispecific antibody ,cytokine-induced killer ,B-cell neoplasia ,CAR signaling ,Immunologic diseases. Allergy ,RC581-607 - Abstract
Strategies to increase the anti-tumor efficacy of cytokine-induced killer cells (CIKs) include genetic modification with chimeric antigen receptors (CARs) or the addition of soluble T-cell engaging bispecific antibodies (BsAbs). Here, CIKs were modified using a transposon system integrating two distinct anti-CD19 CARs (CAR-MNZ and CAR-BG2) or combined with soluble CD3xCD19 BsAb blinatumomab (CIK + Blina). CAR-MNZ bearing the CD28-OX40-CD3ζ signaling modules, and CAR-BG2, designed on the Tisagenlecleucel CAR sequence (Kymriah®), carrying the 4-1BB and CD3ζ signaling elements, were employed. After transfection and CIK expansion, cells expressed CAR-CD19 to a similar extent (35.9% CAR-MNZ and 17.7% CAR-BG2). In vitro evaluations demonstrated robust proliferation and cytotoxicity (~50% cytotoxicity) of CARCIK-MNZ, CARCIK-BG2, and CIK + Blina against CD19+ target cells, suggesting similar efficacy. All effectors formed an increased number of synapses, activated NFAT and NFkB, and secreted IL-2 and IFN-ɣ upon encountering targets. CIK + Blina displayed strongest NFAT and IFN-ɣ induction, whereas CARCIK-BG2 demonstrated superior synapse formation. All the effectors have shown therapeutic activity in vivo against the CD19+ Daudi tumor model, with CARCIK cells showing a more durable response compared to CIK + Blina, likely due to the short half-life of Blina in this model.
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- 2024
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3. A phase I study of autologous mesenchymal stromal cells for severe steroid-dependent nephrotic syndrome
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Marina Vivarelli, Manuela Colucci, Mattia Algeri, Federica Zotta, Francesco Emma, Ines L’Erario, Marco Busutti, Stefano Rota, Chiara Capelli, Martino Introna, Marta Todeschini, Federica Casiraghi, Annalisa Perna, Tobia Peracchi, Andrea De Salvo, Nadia Rubis, Franco Locatelli, Giuseppe Remuzzi, and Piero Ruggenenti
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Clinical trials ,Nephrology ,Medicine - Abstract
BACKGROUND Severe forms of idiopathic nephrotic syndrome (INS) require prolonged immunosuppressive therapies and repeated courses of high-dose glucocorticoids. Mesenchymal stromal cells (MSCs) have promising immunomodulatory properties that may be employed therapeutically to reduce patient exposure to medications and their side effects.METHODS We performed a phase I open-label trial assessing safety and feasibility of autologous bone marrow–derived MSCs (BM-MSCs) in children and young adults with severe forms of steroid-dependent nephrotic syndrome. Following autologous BM-MSC preparation and infusion, oral immunosuppression was tapered. Safety, efficacy, and immunomodulatory effects in vivo were monitored for 12 months.RESULTS Sixteen patients (10 children, 6 adults) were treated. Adverse events were limited and not related to BM-MSC infusions. All patients relapsed during follow-up, but in the 10 treated children, time to first relapse was delayed (P = 0.02) and number of relapses was reduced (P = 0.002) after BM-MSC infusion, compared with the previous 12 months. Cumulative prednisone dose was also reduced at 12 months compared with baseline (P < 0.05). No treatment benefit was observed in adults.In children, despite tapering of immunosuppression, clinical benefit was mirrored by a significant reduction in total CD19+, mature, and memory B cells and an increase in regulatory T cells in vivo up to 3–6 months following BM-MSC infusionCONCLUSION Treatment with autologous BM-MSCs is feasible and safely reduces relapses and immunosuppression at 12 months in children with severe steroid-dependent INS. Immunomodulatory studies suggest that repeating MSC infusions at 3–6 months may sustain benefit.TRIAL REGISTRATION EudraCT 2016-004804-77.FUNDING AIFA Ricerca Indipendente 2016-02364623.
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- 2023
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4. Potency assays and biomarkers for cell-based advanced therapy medicinal products
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Chiara Capelli, Carolina Cuofano, Chiara Pavoni, Simona Frigerio, Daniela Lisini, Sara Nava, Michele Quaroni, Valentina Colombo, Francesco Galli, Svetlana Bezukladova, Paola Panina-Bordignon, Giuseppe Gaipa, Patrizia Comoli, Giulio Cossu, Gianvito Martino, Andrea Biondi, Martino Introna, and Josée Golay
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advanced therapy medicinal product (ATMP) ,potency ,CAR (chimeric antigen receptor) ,T cell therapy ,stem cell ,tissue regeneration ,Immunologic diseases. Allergy ,RC581-607 - Abstract
Advanced Therapy Medicinal Products (ATMPs) based on somatic cells expanded in vitro, with or without genetic modification, is a rapidly growing area of drug development, even more so following the marketing approval of several such products. ATMPs are produced according to Good Manufacturing Practice (GMP) in authorized laboratories. Potency assays are a fundamental aspect of the quality control of the end cell products and ideally could become useful biomarkers of efficacy in vivo. Here we summarize the state of the art with regard to potency assays used for the assessment of the quality of the major ATMPs used clinic settings. We also review the data available on biomarkers that may substitute more complex functional potency tests and predict the efficacy in vivo of these cell-based drugs.
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- 2023
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5. Kidney transplant tolerance associated with remote autologous mesenchymal stromal cell administration
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Federica Casiraghi, Norberto Perico, Eliana Gotti, Marta Todeschini, Marilena Mister, Monica Cortinovis, Valentina Portalupi, Anna Rita Plati, Flavio Gaspari, Alessandro Villa, Martino Introna, Elena Longhi, and Giuseppe Remuzzi
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immunoregulation ,immunosuppression withdrawal ,kidney transplantation ,mesenchymal stromal cells ,tolerance ,Medicine (General) ,R5-920 ,Cytology ,QH573-671 - Abstract
Abstract Here we report the case of successful immune tolerance induction in a living‐donor kidney transplant recipient remotely treated with autologous bone marrow‐derived mesenchymal stromal cells (MSC). This case report, which to the best of our knowledge is the first in the world in this setting, provides evidence that the modulation of the host immune system with MSC can enable the safe withdrawal of maintenance immunosuppressive drugs while preserving optimal long‐term kidney allograft function.
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- 2020
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6. Tolerance to Bone Marrow Transplantation: Do Mesenchymal Stromal Cells Still Have a Future for Acute or Chronic GvHD?
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Martino Introna and Josée Golay
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mesenchymal stromal cell ,graft versus host disease ,hematopoietic stem cell transplantation ,immunosuppressive drugs ,inflammation ,Immunologic diseases. Allergy ,RC581-607 - Abstract
Mesenchymal Stromal Cells (MSCs) are fibroblast-like cells of mesodermal origin present in many tissues and which have the potential to differentiate to osteoblasts, adipocytes and chondroblasts. They also have a clear immunosuppressive and tissue regeneration potential. Indeed, the initial classification of MSCs as pluripotent stem cells, has turned into their identification as stromal progenitors. Due to the relatively simple procedures available to expand in vitro large numbers of GMP grade MSCs from a variety of different tissues, many clinical trials have tested their therapeutic potential in vivo. One pathological condition where MSCs have been quite extensively tested is steroid resistant (SR) graft versus host disease (GvHD), a devastating condition that may occur in acute or chronic form following allogeneic hematopoietic stem cell transplantation. The clinical and experimental results obtained have outlined a possible efficacy of MSCs, but unfortunately statistical significance in clinical studies has only rarely been reached and effects have been relatively limited in most cases. Nonetheless, the extremely complex pathogenetic mechanisms at the basis of GvHD, the fact that studies have been conducted often in patients who had been previously treated with multiple lines of therapy, the variable MSC doses and schedules administered in different trials, the lack of validated potency assays and clear biomarkers, the difference in MSC sources and production methods may have been major factors for this lack of clear efficacy in vivo. The heterogeneity of MSCs and their different stromal differentiation potential and biological activity may be better understood through more refined single cell sequencing and proteomic studies, where either an “anti-inflammatory” or a more “immunosuppressive” profile can be identified. We summarize the pathogenic mechanisms of acute and chronic GvHD and the role for MSCs. We suggest that systematic controlled clinical trials still need to be conducted in the most promising clinical settings, using better characterized cells and measuring efficacy with specific biomarkers, before strong conclusions can be drawn about the therapeutic potential of these cells in this context. The same analysis should be applied to other inflammatory, immune or degenerative diseases where MSCs may have a therapeutic potential.
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- 2020
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7. Protective Effects of Human Nonrenal and Renal Stromal Cells and Their Conditioned Media in a Rat Model of Chronic Kidney Disease
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Barbara Imberti, Domenico Cerullo, Daniela Corna, Cinzia Rota, Monica Locatelli, Anna Pezzotta, Martino Introna, Chiara Capelli, Claudia Elisa Carminati, Ton J. Rabelink, Danielle G. Leuning, Carlamaria Zoja, Marina Morigi, Giuseppe Remuzzi, Ariela Benigni, and Valerie Luyckx
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Medicine - Abstract
Mesenchymal stromal cells (MSCs) are emerging as a novel therapeutic option for limiting chronic kidney disease progression. Conditioned medium (CM) containing bioactive compounds could convey similar benefits, avoiding the potential risks of cell therapy. This study compared the efficacy of nonrenal and renal cell-based therapy with the corresponding CM in rats with renal mass reduction (RMR). Infusions of human kidney stromal cells (kPSCs) and CM-kPSCs, but not umbilical cord (uc) MSCs or CM-ucMSCs, reduced proteinuria and preserved podocyte number and nephrin expression in RMR rats. Glomerular fibrosis, microvascular rarefaction, and apoptosis were reduced by all treatments, while the peritubular microvascular loss was reduced by kPSCs and CM-kPSCs treatment only. Importantly, kPSCs and CM-kPSCs reduced NG2-positive pericytes, and all therapies reduced α-smooth muscle actin expression, indicating reduced myofibroblast expansion. Treatment with kPSCs also significantly inhibited the accumulation of ED1-positive macrophages in the renal interstitium of RMR rats. These findings demonstrate that the CM of ucMSCs and kPSCs confers similar renoprotection as the cells. kPSCs and CM-kPSCs may be superior in attenuating chronic renal injury as a cell source.
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- 2020
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8. Therapeutic potential of stromal cells of non-renal or renal origin in experimental chronic kidney disease
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Cinzia Rota, Marina Morigi, Domenico Cerullo, Martino Introna, Ornella Colpani, Daniela Corna, Chiara Capelli, Ton J. Rabelink, Danielle G. Leuning, Daniela Rottoli, Ariela Benigni, Carlamaria Zoja, and Giuseppe Remuzzi
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Mesenchymal stromal cell therapy ,Renal perivascular cells ,Conditioned medium ,Renal repair ,Chronic kidney disease ,Medicine (General) ,R5-920 ,Biochemistry ,QD415-436 - Abstract
Abstract Background Mesenchymal stromal cell (MSC)-based therapy is a promising strategy for preventing the progression of chronic kidney disease (CKD), with the potential to induce tissue regeneration. In search of the best cellular source we compared, in the rat model of adriamycin (ADR) nephropathy, the regenerative potential of human stromal cells of non-renal origin, such as bone marrow (bm) MSCs and umbilical cord (uc) MSCs, with that of newly discovered stromal cells of renal origin, the kidney perivascular cells (kPSCs) known to exhibit tissue-specific properties. Methods The therapeutic effect of repeated infusions of human bmMSCs, ucMSCs, kPSCs (1.5 × 106 cells/rats) or conditioned medium from ucMSCs was studied in athymic rats with ADR-induced nephropathy (7.9 mg/kg). The ability of the three stromal cell populations to engraft the damaged kidney was evaluated by detecting the presence of human nuclear antigenpos cells. Glomerular podocyte loss and endothelial damage, sclerotic lesions and inflammation were assessed at 14 and 28 days. In-vitro experiments with a transwell system were performed to investigate the effects of different stromal cell populations on parietal epithelial cells (PECs) activated or not with albumin or angiotensin II for 24 h. Results Infusions of non-renal and renal stromal cells resulted in a comparable engraftment into the kidney, in the peritubular areas and around the glomerular structures. All three cell populations limited podocyte loss and glomerular endothelial cell injury, and attenuated the formation of podocyte and PEC bridges. This translated into a reduction of glomerulosclerosis and fibrosis. Human ucMSCs had an anti-inflammatory effect superior to that of the other stromal cells, reducing macrophage infiltration and inducing polarisation towards the M2 macrophage phenotype. Conditioned medium from ucMSCs shared the same renoprotective effects of the cells. Consistent with in-vivo data, bmMSCs and kPSCs, but even more so ucMSCs, limited proliferation, migratory potential and extracellular matrix production of activated PECs, when cultured in a transwell system. Conclusions Our data indicate that either non-renal or renal stromal cells induce renal tissue repair, highlighting ucMSCs and their conditioned medium as the most reliable clinical therapeutic tool for CKD patients.
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- 2018
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9. Multiple intracerebroventricular injections of human umbilical cord mesenchymal stem cells delay motor neurons loss but not disease progression of SOD1G93A mice
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Francesca Sironi, Antonio Vallarola, Martina Bruna Violatto, Laura Talamini, Mattia Freschi, Roberta De Gioia, Chiara Capelli, Azzurra Agostini, Davide Moscatelli, Massimo Tortarolo, Paolo Bigini, Martino Introna, and Caterina Bendotti
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Max 6 ,Amyotrophic lateral sclerosis ,Mesenchymal stem cells ,Umbilical cord ,Transgenic SOD1G93A mice ,Motor neuron ,Gliosis ,Biology (General) ,QH301-705.5 - Abstract
Stem cell therapy is considered a promising approach in the treatment of amyotrophic lateral sclerosis (ALS) and mesenchymal stem cells (MSCs) seem to be the most effective in ALS animal models. The umbilical cord (UC) is a source of highly proliferating fetal MSCs, more easily collectable than other MSCs. Recently we demonstrated that human (h) UC-MSCs, double labeled with fluorescent nanoparticles and Hoechst-33258 and transplanted intracerebroventricularly (ICV) into SOD1G93A transgenic mice, partially migrated into the spinal cord after a single injection. This prompted us to assess the effect of repeated ICV injections of hUC-MSCs on disease progression in SOD1G93A mice. Although no transplanted cells migrated to the spinal cord, a partial but significant protection of motor neurons (MNs) was found in the lumbar spinal cord of hUC-MSCs-treated SOD1G93A mice, accompanied by a shift from a pro-inflammatory (IL-6, IL-1β) to anti-inflammatory (IL-4, IL-10) and neuroprotective (IGF-1) environment in the lumbar spinal cord, probably linked to the activation of p-Akt survival pathway in both motor neurons and reactive astrocytes. However, this treatment neither prevented the muscle denervation nor delayed the disease progression of mice, emphasizing the growing evidence that protecting the motor neuron perikarya is not sufficient to delay the ALS progression.
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- 2017
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10. Human mesenchymal stromal cells transplanted into mice stimulate renal tubular cells and enhance mitochondrial function
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Luca Perico, Marina Morigi, Cinzia Rota, Matteo Breno, Caterina Mele, Marina Noris, Martino Introna, Chiara Capelli, Lorena Longaretti, Daniela Rottoli, Sara Conti, Daniela Corna, Giuseppe Remuzzi, and Ariela Benigni
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Science - Abstract
Mesenchymal stromal cells drive renal regeneration following injury. Here, the authors show that human mesenchymal stromal cells, when transplanted into mice with acute kidney injury, stimulate renal tubular cell growth and enhance mitochondrial function via SIRT3.
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- 2017
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11. Long-Term Clinical and Immunological Profile of Kidney Transplant Patients Given Mesenchymal Stromal Cell Immunotherapy
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Norberto Perico, Federica Casiraghi, Marta Todeschini, Monica Cortinovis, Eliana Gotti, Valentina Portalupi, Marilena Mister, Flavio Gaspari, Alessandro Villa, Sonia Fiori, Martino Introna, Elena Longhi, and Giuseppe Remuzzi
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mesenchymal stromal cells ,kidney transplantation ,tolerance ,memory CD8+ T cells ,regulatory T cells ,B cells ,Immunologic diseases. Allergy ,RC581-607 - Abstract
We report here the long-term clinical and immunological results of four living-donor kidney transplant patients given autologous bone marrow-derived mesenchymal stromal cells (MSCs) as part of a phase 1 study focused on the safety and feasibility of this cell therapy. According to study protocols implemented over time, based on initial early safety findings, the patients were given MSC at day 7 posttransplant (n = 2) or at day −1 pretransplant (n = 2) and received induction therapy with basiliximab and low-dose rabbit anti-thymocyte globulin (RATG) or RATG alone, and were maintained on low-dose ciclosporin (CsA)/mycophenolate mofetil (MMF). All MSC-treated patients had stable graft function during the 5- to 7-year follow-up, without increased susceptibility to infections or neoplasm. In three MSC recipients, but not historical control patients, circulating memory CD8+ T cell percentages remained lower than basal, coupled with persistent reduction of ex vivo donor-specific cytotoxicity. Two patients showed a long-lasting increase in the regulatory T cell/memory CD8+ T cell ratio, paralleled by high circulating levels of naïve and transitional B cells. In one of these two patients, CsA was successfully discontinued, and currently the low-dose MMF monotherapy is on the tapering phase. The study shows that MSC therapy is safe in the long term and could promote a pro-tolerogenic environment in selected patients. Extensive immunomonitoring of MSC-treated kidney transplant recipients could help selection of patients for safe withdrawal of maintenance immunosuppressive drugs (NCT00752479 and NCT02012153).
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- 2018
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12. Mesenchymal stromal cells from human umbilical cord prevent the development of lung fibrosis in immunocompetent mice.
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Gianluca Moroncini, Chiara Paolini, Fiorenza Orlando, Chiara Capelli, Antonella Grieco, Cecilia Tonnini, Silvia Agarbati, Eleonora Mondini, Stefania Saccomanno, Gaia Goteri, Silvia Svegliati Baroni, Mauro Provinciali, Martino Introna, Nicoletta Del Papa, and Armando Gabrielli
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Medicine ,Science - Abstract
Lung fibrosis is a severe condition resulting from several interstial lung diseases (ILD) with different etiologies. Current therapy of ILD, especially those associated with connective tissue diseases, is rather limited and new anti-fibrotic strategies are needed. In this study, we investigated the anti-fibrotic activity in vivo of human mesenchymal stromal cells obtained from whole umbilical cord (hUC-MSC). Adult immunocompetent C57BL/6 mice (n. = 8 for each experimental condition) were injected intravenously with hUC-MSC (n. = 2.5 × 105) twice, 24 hours and 7 days after endotracheal injection of bleomycin. Upon sacrifice at days 8, 14, 21, collagen content, inflammatory cytokine profile, and hUC-MSC presence in explanted lung tissue were analyzed. Systemic administration of a double dose of hUC-MSC significantly reduced bleomycin-induced lung injury (inflammation and fibrosis) in mice through a selective inhibition of the IL6-IL10-TGFβ axis involving lung M2 macrophages. Only few hUC-MSC were detected from explanted lungs, suggesting a "hit and run" mechanism of action of this cellular therapy. Our data indicate that hUC-MSC possess strong in vivo anti-fibrotic activity in a mouse model resembling an immunocompetent human subject affected by inflammatory ILD, providing proof of concept for ad-hoc clinical trials.
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- 2018
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13. Longitudinal tracking of triple labeled umbilical cord derived mesenchymal stromal cells in a mouse model of Amyotrophic Lateral Sclerosis
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Martina Bruna Violatto, Chiara Santangelo, Chiara Capelli, Roberta Frapolli, Raffaele Ferrari, Leopoldo Sitia, Massimo Tortarolo, Laura Talamini, Sara Previdi, Davide Moscatelli, Mario Salmona, Martino Introna, Caterina Bendotti, and Paolo Bigini
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Mesenchymal stromal cells ,Umbilical cord ,Amyotrophic Lateral Sclerosis ,Cell tracking ,Nanotechnology ,In vivo imaging ,Biology (General) ,QH301-705.5 - Abstract
The translational potential of cell therapy to humans requires a deep knowledge of the interaction between transplanted cells and host tissues. In this study, we evaluate the behavior of umbilical cord mesenchymal stromal cells (UC-MSCs), labeled with fluorescent nanoparticles, transplanted in healthy or early symptomatic transgenic SOD1G93A mice (a murine model of Amyotrophic Lateral Sclerosis). The double labeling of cells with nanoparticles and Hoechst-33258 enabled their tracking for a long time in both cells and tissues. Whole-body distribution of UC-MSCs was performed by in-vivo and ex-vivo analyses 1, 7, 21 days after single intravenous or intracerebroventricular administration. By intravenous administration cells were sequestered by the lungs and rapidly cleared by the liver. No difference in biodistribution was found among the two groups. On the other hand, UC-MSCs transplanted in lateral ventricles remained on the choroid plexus for the whole duration of the study even if decreasing in number. Few cells were found in the spinal cord of SOD1G93A mice exclusively. No migration in brain parenchyma was observed. These results suggest that the direct implantation in brain ventricles allows a prolonged permanence of cells close to the damaged areas and makes this method of tracking reliable for future studies of efficacy.
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- 2015
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14. Direct Reprogramming of Human Bone Marrow Stromal Cells into Functional Renal Cells Using Cell-free Extracts
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Evangelia Papadimou, Marina Morigi, Paraskevas Iatropoulos, Christodoulos Xinaris, Susanna Tomasoni, Valentina Benedetti, Lorena Longaretti, Cinzia Rota, Marta Todeschini, Paola Rizzo, Martino Introna, Maria Grazia de Simoni, Giuseppe Remuzzi, Michael S. Goligorsky, and Ariela Benigni
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Medicine (General) ,R5-920 ,Biology (General) ,QH301-705.5 - Abstract
The application of cell-based therapies in regenerative medicine is gaining recognition. Here, we show that human bone marrow stromal cells (BMSCs), also known as bone-marrow-derived mesenchymal cells, can be reprogrammed into renal proximal tubular-like epithelial cells using cell-free extracts. Streptolysin-O-permeabilized BMSCs exposed to HK2-cell extracts underwent morphological changes—formation of “domes” and tubule-like structures—and acquired epithelial functional properties such as transepithelial-resistance, albumin-binding, and uptake and specific markers E-cadherin and aquaporin-1. Transmission electron microscopy revealed the presence of brush border microvilli and tight intercellular contacts. RNA sequencing showed tubular epithelial transcript abundance and revealed the upregulation of components of the EGFR pathway. Reprogrammed BMSCs integrated into self-forming kidney tissue and formed tubular structures. Reprogrammed BMSCs infused in immunodeficient mice with cisplatin-induced acute kidney injury engrafted into proximal tubuli, reduced renal injury and improved function. Thus, reprogrammed BMSCs are a promising cell resource for future cell therapy.
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- 2015
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15. The specific Bruton tyrosine kinase inhibitor acalabrutinib (ACP-196) shows favorable in vitro activity against chronic lymphocytic leukemia B cells with CD20 antibodies
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Josée Golay, Greta Ubiali, and Martino Introna
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Diseases of the blood and blood-forming organs ,RC633-647.5 - Published
- 2017
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16. Phenotypical and Functional Characteristics of in Vitro-Expanded Adipose-Derived Mesenchymal Stromal Cells from Patients with Systematic Sclerosis
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Chiara Capelli, Eleonora Zaccara, Paola Cipriani, Paola Di Benedetto, Wanda Maglione, Romina Andracco, Gabriele Di Luca, Francesca Pignataro, Roberto Giacomelli, Martino Introna, Claudio Vitali, and Nicoletta Del Papa M.D., U.O.C.
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Medicine - Abstract
Mesenchymal stromal cells (MSCs) have received attention as an ideal source of regenerative cells because of their multipotent differentiation potential. Adipose tissue is an attractive source of MSCs. Recent studies have shown that autologous fat grafting may be effective in the treatment of systemic sclerosis (SSc), but no specific study exists that aimed at investigating whether adipose tissue-derived stromal cells (ADSCs) from SSc patients maintain normal phenotypic and functional characteristics. The purpose of the current study was to investigate whether ADSCs from patients with SSc (SSc-ADSCs) are phenotypically and functionally identical to those from healthy controls (HC-ADSCs). Adipose tissue samples were obtained from 10 patients with SSc and from 8 HCs. Both MSC populations were evaluated for their capacity to (a) express specific MSC surface antigens by flow cytometry analysis, (b) proliferate, (c) differentiate along the adipogenic and osteogenic lineages, (d) suppress in vitro lymphocyte proliferation induced by a mitogenic stimulus, and (e) support endothelial cell (EC) tube formation. ADSCs from SSc patients and HCs showed similar surface phenotype and multilineage differentiation capabilities. In PBMC proliferation inhibition assays, no significant differences were observed between SSc- and HC-ADSCs. Using ADSC/EC cocultures, both SSc- and HC-ADSCs improved tube formation by both HC- and SSc-ECs. This effect was enhanced under hypoxic conditions in all of the cocultures. SSc-ADSCs exhibited the same phenotypic pattern, proliferation and differentiation potentials, and immunosuppressive properties as those from HCs. The proangiogenic activity shown by SSc-ADSCs, namely, under hypoxic conditions, suggests that autologous ADSC grafting may represent a possible therapeutic option for SSc.
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- 2017
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17. Innovative Clinical Perspectives for CIK Cells in Cancer Patients
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Martino Introna and Fabio Correnti
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CIK ,NK ,GvHD ,Biology (General) ,QH301-705.5 ,Chemistry ,QD1-999 - Abstract
Cytokine-induced killer (CIK) cells are T lymphocytes that have acquired, in vitro, following extensive manipulation by Interferon gamma (IFN-γ), OKT3 and Interleukin 2 (IL-2) addition, the expression of several Natural Killer (NK) cell-surface markers. CIK cells have a dual “nature”, due to the presence of functional TCR as well as NK molecules, even if the antitumoral activity can be traced back only to the NK-like structures (DNAM-1, NKG2D, NKp30 and CD56). In addition to antineoplastic activity in vitro and in several in-vivo models, CIK cells show very limited, if any, GvHD toxicity as well as a strong intratumoral homing. For all such reasons, CIK cells have been proposed and tested in many clinical trials in cancer patients both in autologous and allogeneic combinations, up to haploidentical mismatching. Indeed, genetic modification of CIK cells as well as the possibility of combining them with specific monoclonal antibodies will further expand the possibility of their clinical utilization.
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- 2018
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18. Ibrutinib interferes with the cell-mediated anti-tumor activities of therapeutic CD20 antibodies: implications for combination therapy
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Fabio Da Roit, Patrick J. Engelberts, Ronald P. Taylor, Esther C.W. Breij, Giuseppe Gritti, Alessandro Rambaldi, Martino Introna, Paul W.H.I. Parren, Frank J. Beurskens, and Josée Golay
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Diseases of the blood and blood-forming organs ,RC633-647.5 - Abstract
The novel Bruton tyrosine kinase inhibitor ibrutinib and phosphatidyl-4-5-biphosphate 3-kinase-δ inhibitor idelalisib are promising drugs for the treatment of chronic lymphocytic leukemia and B-cell non-Hodgkin lymphoma, either alone or in combination with anti-CD20 antibodies. We investigated the possible positive or negative impact of these drugs on all known mechanisms of action of both type I and type II anti-CD20 antibodies. Pretreatment with ibrutinib for 1 hour did not increase direct cell death of cell lines or chronic lymphocytic leukemia samples mediated by anti-CD20 antibodies. Pre-treatment with ibrutinib did not inhibit complement activation or complement-mediated lysis. In contrast, ibrutinib strongly inhibited all cell-mediated mechanisms induced by anti-CD20 antibodies rituximab, ofatumumab or obinutuzumab, either in purified systems or whole blood assays. Activation of natural killer cells, and antibody-dependent cellular cytotoxicity by these cells, as well as phagocytosis by macrophages or neutrophils were inhibited by ibrutinib with a half maximal effective concentration of 0.3–3 μM. Analysis of anti-CD20 mediated activation of natural killer cells isolated from patients on continued oral ibrutinib treatment suggested that repeated drug dosing inhibits these cells in vivo. Finally we show that the phosphatidyl-4-5-biphosphate 3-kinase-δ inhibitor idelalisib similarly inhibited the immune cell-mediated mechanisms induced by anti-CD20 antibodies, although the effects of this drug at 10 μM were weaker than those observed with ibrutinib at the same concentration. We conclude that the design of combined treatment schedules of anti-CD20 antibodies with these kinase inhibitors should consider the multiple negative interactions between these two classes of drugs.
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- 2015
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19. Frequent occurrence of non-malignant genetic alterations in clinical grade mesenchymal stromal cells expanded for cell therapy protocols
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Chiara Capelli, Olga Pedrini, Gisella Cassina, Orietta Spinelli, Silvia Salmoiraghi, Josée Golay, Alessandro Rambaldi, Ursula Giussani, and Martino Introna
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Diseases of the blood and blood-forming organs ,RC633-647.5 - Published
- 2014
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20. The Polo-Like Kinase 1 (PLK1) inhibitor NMS-P937 is effective in a new model of disseminated primary CD56+ acute monoblastic leukaemia.
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Alessia Casolaro, Josee Golay, Clara Albanese, Roberta Ceruti, Veronica Patton, Sabrina Cribioli, Alice Pezzoni, Marco Losa, Gemma Texido, Ursula Giussani, Francesco Marchesi, Nadia Amboldi, Barbara Valsasina, Silvia Bungaro, Gianni Cazzaniga, Alessandro Rambaldi, Martino Introna, Enrico Pesenti, and Rachele Alzani
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Medicine ,Science - Abstract
CD56 is expressed in 15-20% of acute myeloid leukaemias (AML) and is associated with extramedullary diffusion, multidrug resistance and poor prognosis. We describe the establishment and characterisation of a novel disseminated model of AML (AML-NS8), generated by injection into mice of leukaemic blasts freshly isolated from a patient with an aggressive CD56(+) monoblastic AML (M5a). The model reproduced typical manifestations of this leukaemia, including presence of extramedullary masses and central nervous system involvement, and the original phenotype, karyotype and genotype of leukaemic cells were retained in vivo. Recently Polo-Like Kinase 1 (PLK1) has emerged as a new candidate drug target in AML. We therefore tested our PLK1 inhibitor NMS-P937 in this model either in the engraftment or in the established disease settings. Both schedules showed good efficacy compared to standard therapies, with a significant increase in median survival time (MST) expecially in the established disease setting (MST = 28, 36, 62 days for vehicle, cytarabine and NMS-P937, respectively). Importantly, we could also demonstrate that NMS-P937 induced specific biomarker modulation in extramedullary tissues. This new in vivo model of CD56(+) AML that recapitulates the human tumour lends support for the therapeutic use of PLK1 inhibitors in AML.
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- 2013
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21. Mesenchymal Stromal Cells Do Not Increase the Risk of Viral Reactivation Nor the Severity of Viral Events in Recipients of Allogeneic Stem Cell Transplantation
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Giovanna Lucchini, Erica Dander, Fabio Pavan, Irene Di Ceglie, Adriana Balduzzi, Paolo Perseghin, Giuseppe Gaipa, Alessandra Algarotti, Martino Introna, Alessandro Rambaldi, Attilio Rovelli, Andrea Biondi, Ettore Biagi, and Giovanna D'Amico
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Internal medicine ,RC31-1245 - Abstract
Mesenchymal stromal cells (MSC) are tested in clinical trials to treat graft versus host disease (GvHD) after stem cell transplantation (SCT). In vitro studies demonstrated MSC's broad immunosuppressive activity. As infections represent a major risk after SCT, it is important to understand the role of MSC in this context. We analyzed 24 patients (pts) receiving MSC for GvHD in our Unit between 2009 and 2011. We recorded viral reactivations as measured in whole blood with polymerase chain reaction for 100 days following MSC administration. In patients with a documented viral reactivation in the first 3 days following MSCs infusion the frequency of virus-specific IFNgamma-producing cells was determined through enzyme-linked immunospot assay. In our cohort of patients viral reactivation after MSC infusion occurred in 45% of the cases, which did not significantly differ from the incidence in a historical cohort of patients affected by steroid resistant GvHD and treated with conventional immunosuppression. No patient presented severe form of infection. Two cases could be checked for immunological response to viral stimulus and demonstrated virus specific T-cytotoxic lymphocyte activity. In our experience MSC infusion did not prove to trigger more frequent or severer viral reactivations in the post transplantation setting.
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- 2012
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22. Pleiotropic anti-myeloma activity of ITF2357: inhibition of interleukin-6 receptor signaling and repression of miR-19a and miR-19b
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Katia Todoerti, Valentina Barbui, Olga Pedrini, Marta Lionetti, Gianluca Fossati, Paolo Mascagni, Alessandro Rambaldi, Antonino Neri, Martino Introna, Luigia Lombardi, and Josée Golay
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Diseases of the blood and blood-forming organs ,RC633-647.5 - Abstract
Background The histone deacetylase inhibitor ITF2357 has potent cytotoxic activity in multiple myeloma in vitro and has entered clinical trials for this disease.Design and Methods In order to gain an overall view of the activity of ITF2357 and identify specific pathways that may be modulated by the drug, we performed gene expression profiling of the KMS18 multiple myeloma cell line treated with the drug. The modulation of several genes and their biological consequence were verified in a panel of multiple myeloma cell lines and cells freshly isolated from patients by using polymerase chain reaction analysis and western blotting.Results Out of 38,500 human genes, we identified 140 and 574 up-regulated genes and 102 and 556 down-modulated genes at 2 and 6 h, respectively, with a significant presence of genes related to transcription regulation at 2 h and to cell cycling and apoptosis at 6 h. Several of the identified genes are particularly relevant to the biology of multiple myeloma and it was confirmed that ITF2357 also modulated their encoded proteins in different multiple myeloma cell lines. In particular, ITF2357 down-modulated the interleukin-6 receptor α (CD126) transcript and protein in both cell lines and freshly isolated patients’ cells, whereas it did not significantly modify interleukin-6 receptor β (CD130) expression. The decrease in CD126 expression was accompanied by decreased signaling by interleukin-6 receptor, as measured by STAT3 phosphorylation in the presence and absence of inter-leukin-6. Finally, the drug significantly down-modulated the MIRHG1 transcript and its associated microRNA, miR-19a and miR-19b, known to have oncogenic activity in multiple myeloma.Conclusions ITF2357 inhibits several signaling pathways involved in myeloma cell growth and survival.
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- 2010
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23. Repeated infusions of donor-derived cytokine-induced killer cells in patients relapsing after allogeneic stem cell transplantation: a phase I study
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Martino Introna, Gianmaria Borleri, Elena Conti, Marta Franceschetti, Anna Maria Barbui, Raewyn Broady, Erica Dander, Giuseppe Gaipa, Giovanna D’Amico, Ettore Biagi, Matteo Parma, Enrico M. Pogliani, Orietta Spinelli, Donatella Baronciani, Anna Grassi, Josée Golay, Tiziano Barbui, Andrea Biondi, and Alessandro Rambaldi
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Diseases of the blood and blood-forming organs ,RC633-647.5 - Abstract
Background and Objectives Cytokine-induced killer (CIK) cells have shown anti-leukemic activity and little graft-versus-host disease (GVHD) in several animal models. The safety of these cells in autologous settings has been shown. We performed a phase I study of allogeneic (donor’s) CIK cells in patients relapsing after allogeneic haematopoietic stem cell transplantation (HSCT).Design and Methods Eleven patients with acute myelogenous leukemia (n=4), Hodgkin’s disease (n=3), chronic myelomonocytic leukemia, (n=1), pre-B acute lymphoblastic leukemia (n=1) and myelodysplasia (n=2), all of whom had relapsed after sibling (n=6) or matched unrelated donor (n=5) HSCT, entered this study.Results Before CIK administration, six patients had received other salvage treatments including chemotherapy (n=5), radiotherapy (n=1) and unmanipulated donor lymphocytes (n=6) without any significant tumor response. The median number of CIK infusions was two (range 1–7) and the median number of total CIK cells was 12.4 × 106/kg (range 7.2–87.4). The infusions were well tolerated and no acute or late infusion-related reactions were recorded. Acute GVHD (grade I and II) was observed in four patients, 30 days after the last CIK infusion, and progressed into extensive chronic GVHD in two cases. Disease progression and death occurred in six patients. One patient had stable disease, one had hematologic improvement and three achieved complete responses.Interpretation and Conclusions This study shows that the production of allogeneic CIK cells is feasible under clinical-grade conditions, well tolerated and may contribute to clinical responses.
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- 2007
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24. Optimization of therapeutic T cell expansion in G-Rex device and applicability to large-scale production for clinical use
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Elisa Gotti, Sarah Tettamanti, Silvia Zaninelli, Carolina Cuofano, Irene Cattaneo, Maria Caterina Rotiroti, Sabrina Cribioli, Rachele Alzani, Alessandro Rambaldi, Martino Introna, Josée Golay, Gotti, E, Tettamanti, S, Zaninelli, S, Cuofano, C, Cattaneo, I, Rotiroti, M, Cribioli, S, Alzani, R, Rambaldi, A, Introna, M, and Golay, J
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Cytotoxicity, Immunologic ,Cancer Research ,Transplantation ,T-Lymphocytes ,Immunology ,T cell ,Cell Biology ,Killer Cells, Natural ,bioreactor ,Mice ,Cytokine-Induced Killer Cells ,Oncology ,Precursor B-Cell Lymphoblastic Leukemia-Lymphoma ,Animals ,Immunology and Allergy ,immunotherapy ,cytokine-induced killer cell ,Genetics (clinical) ,Cell Proliferation - Abstract
Our center performs experimental clinical studies with advanced therapy medicinal products (ATMPs) based on polyclonal T cells, all of which are currently expanded in standard T-flasks. Given the need to increase the efficiency and safety of large-scale T cell expansion for clinical use, we have optimized the method to expand in G-Rex devices both cytokine-induced killer cells (CIKs) from peripheral or cord blood and blinatumomab-expanded T cells (BETs). We show that the G-Rex reproducibly allowed the expansion of >30 × 106 CD3+ cells/cm2 of gas-permeable membrane in a mean of 10 to 11 days in a single unit, without manipulation, except for addition of cytokines and sampling of supernatant for lactate measurement every 3 to 4 days. In contrast, 21 to 24 days, twice-weekly cell resuspension and dilution into 48 to 72 T-flasks were required to complete expansions using the standard method. We show that the CIKs produced in G-Rex (CIK-G) were phenotypically very similar, for a large panel of markers, to those expanded in T-flasks, although CIK-G products had lower expression of CD56 and higher expression of CD27 and CD28. Functionally, CIK-Gs were strongly cytotoxic in vitro against the NK cell target K562 and the REH pre-B ALL cell line in the presence of blinatumomab. CIK-Gs also showed therapeutic activity in vivo in the Ph+ pre-B ALL-2 model in mice. The expansion of both CIKs and BETs in G-Rex was validated in good manufacturing practices (GMP) conditions, and we plan to use G-Rex for T cell expansion in future clinical studies.
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- 2022
25. Safety, tolerability, and activity of mesenchymal stem cells versus placebo in multiple sclerosis (MESEMS): a phase 2, randomised, double-blind crossover trial
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Antonio Uccelli, Alice Laroni, Rehiana Ali, Mario Alberto Battaglia, Morten Blinkenberg, Lou Brundin, Michel Clanet, Oscar Fernandez, James Marriott, Paolo Muraro, Seyed Massood Nabavi, Roberto S Oliveri, Ernst Radue, Cristina Ramo Tello, Irene Schiavetti, Johann Sellner, Per Soelberg Sorensen, Maria Pia Sormani, Jens Thomas Wuerfel, Mark S Freedman, Naser Aghdami, Eduardo Agüera-Morales, David Allan, Leila Arab, Mario Battaglia, Isabelle Berry, Bruno Bonetti, Chiara Capelli, Lucio Castellan, Maria Cellerino, Maria Teresa Cencioni, Giancarlo Comi, David Courtman, Francesco Dazzi, Anne Fischer-Nielsen, Victoria Fernandez, Mark S. Freedman, Roberto Furlan, Mario Gimona, Francesca Gualandi, Qingdong Guan, Ellen Iacobaeus, Matilde Inglese, Martino Introna, Guillermo Izquierdo, Shahedeh Karimi, Katarina Le Blanc, Sandra Loaiza, Shahrukh Mallik, Stephen Marley, Ruth Ann Marrie, James Marriot, Gianvito Martino, David Miller, Paolo A. Muraro, Richard Nicholas, Giovanni Orengo, Renuka Palanicawande, Matteo Pardini, Ernst W Radue, Carolina Rush, Luc Sensebe, Dirk Strunk, David Szwajcer, and Claire Thalamas
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Adult ,medicine.medical_specialty ,Multiple Sclerosis ,Adolescent ,Placebo ,Young Adult ,Double-Blind Method ,Internal medicine ,Clinical endpoint ,Humans ,Medicine ,Adverse effect ,Cross-Over Studies ,Expanded Disability Status Scale ,business.industry ,Surrogate endpoint ,Multiple sclerosis ,Brain ,Mesenchymal Stem Cells ,Middle Aged ,medicine.disease ,Crossover study ,Malformations of Cortical Development ,Tolerability ,Neurology (clinical) ,business - Abstract
Summary Background Mesenchymal stem cells (MSCs), also known as mesenchymal stromal cells, have been proposed as a promising therapeutic option for people with multiple sclerosis on the basis of their immunomodulatory and neuroprotective properties. The MEsenchymal StEm cells for Multiple Sclerosis (MESEMS) study was devised to evaluate the safety, tolerability, and activity of autologous MSCs derived from bone marrow and infused intravenously in patients with active multiple sclerosis. Methods MESEMS is a randomised phase 2 trial done at 15 sites in nine countries. Patients (aged 18–50 years) with active relapsing-remitting or progressive multiple sclerosis were included if they had a disease duration of 2–15 years since onset of multiple sclerosis and an Expanded Disability Status Scale score of 2·5–6·5. Patients were randomly assigned (1:1), according to a crossover design, to receive a single intravenous dose of autologous bone marrow-derived MSCs followed by placebo at week 24, or to receive placebo followed by autologous MSCs at week 24, with a follow-up visit at week 48. Primary objectives were to test safety and activity of MSC treatment. The primary safety endpoint was to assess the number and severity of adverse events within each treatment arm. The primary efficacy endpoint was the number of gadolinium-enhancing lesions (GELs) counted over week 4, 12, and 24 compared between treatment groups. The primary efficacy endpoint was assessed in the full analyis set, after all participants completed the week 24 visit. Efficacy endpoints were evaluated using a predefined statistical testing procedure. Safety was monitored throughout the study by recording vital signs and adverse events at each visit. Findings From July 16, 2012, until July 31, 2019, 144 patients were randomly assigned to first receive early intravenous infusion of autologous bone marrow-derived MSCs (n=69) or placebo (n=75). MSC treatment did not meet the primary endpoint of efficacy on the total number of GELs accumulated from baseline to week 24 (rate ratio [RR] 0·94, 95% CI 0·58–1·50; p=0·78). 213 adverse events were recorded, similarly distributed between groups (93 cases recorded in 35 [51%] of 69 patients treated first with MSCs vs 120 cases in 42 [56%] of 75 patients infused first with placebo). The most frequent adverse events reported were infection and infestations, with a total of 54 (25%) of 213 adverse events (18 [19%] of 93 in the early-MSC group and 36 [30%] of 120 in the delayed-MSC group). Nine serious adverse events were reported in seven patients treated with placebo versus none in the MSC group. All serious adverse events were considered to be unrelated to the treatment infusion. No deaths were reported during the study. Interpretation Bone marrow-derived MSC treatment was safe and well tolerated but did not show an effect on GELs, an MRI surrogate marker of acute inflammation, in patients with active forms of multiple sclerosis, at week 24. Thus, this study does not support the use of bone marrow-derived MSCs to treat active multiple sclerosis. Further studies should address the effect of MSCs on parameters related to tissue repair. Funding Fondazione Italiana Sclerosi Multipla (FISM), the European Committee for Treatment and Research in Multiple Sclerosis (ECTRIMS), and the Multiple Sclerosis International Federation (MSIF) for centralised activities. Individual trials participating in the MESEMS network are funded by the following agencies: FISM and Compagnia di San Paolo (Italy); The Danish Multiple Sclerosis Society, The Toyota Foundation, and Danish Blood Donors’ Research Foundation (Denmark); the Spanish Health Research Institute Carlos 3 and the Andalusian Public Foundation Progreso y Salud (Spain); the Royan Institute for Stem Cell Biology and Technology (Iran); the Spinal Cord Injury and Tissue Regeneration Centre Salzburg, Paracelsus Medical University, and Salzburg (Austria); the Fondation pour l’aide a la recherche sur la sclerose en plaques (ARSEP), French Muscular Dystrophy Association (AFM)-Telethon (France); the UK Multiple Sclerosis Society and the UK Stem Cell Foundation (UK); and the Multiple Sclerosis Society of Canada and The Multiple Sclerosis Scientific Research Foundation and Research Manitoba (Canada).
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- 2021
26. Third-party bone marrow–derived mesenchymal stromal cell infusion before liver transplantation: A randomized controlled trial
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Stefania Camagni, Giuseppe Remuzzi, Caterina Mele, Michele Colledan, M. Zambelli, Matteo Breno, Stefano Fagiuoli, Josée Golay, Marilena Mister, Norberto Perico, Marina Buzzi, Pamela Yossenaidy Rodriguez Ordonez, Chiara Capelli, Manuel Alfredo Podestà, Federica Casiraghi, Matteo Cescon, Lorenzo Maroni, Valentina Rosa Bertuzzo, Alessandro Villa, Marta Todeschini, Antonio Daniele Pinna, Martino Introna, Casiraghi, F, Perico, N, Podesta, M, Todeschini, M, Zambelli, M, Colledan, M, Camagni, S, Fagiuoli, S, Pinna, A, Cescon, M, Bertuzzo, V, Maroni, L, Introna, M, Capelli, C, Golay, J, Buzzi, M, Mister, M, Ordonez, P, Breno, M, Mele, C, Villa, A, Remuzzi, G, Casiraghi, Federica, Perico, Norberto, Podestà, Manuel A, Todeschini, Marta, Zambelli, Marco, Colledan, Michele, Camagni, Stefania, Fagiuoli, Stefano, Pinna, Antonio D, Cescon, Matteo, Bertuzzo, Valentina, Maroni, Lorenzo, Introna, Martino, Capelli, Chiara, Golay, Josee T, Buzzi, Marina, Mister, Marilena, Ordonez, Pamela Y R, Breno, Matteo, Mele, Caterina, Villa, Alessandro, and Remuzzi, Giuseppe, Matteo Ravaioli
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medicine.medical_specialty ,Stromal cell ,medicine.medical_treatment ,Liver transplantation ,clinical research/practice ,Mesenchymal Stem Cell Transplantation ,Gastroenterology ,law.invention ,Randomized controlled trial ,Bone Marrow ,law ,Internal medicine ,medicine ,Humans ,Immunology and Allergy ,Pharmacology (medical) ,Transplantation ,tolerance ,liver transplantation ,business.industry ,Mesenchymal stem cell ,Mesenchymal Stem Cells ,Hepatology ,practice ,stem cell ,Clinical trial ,medicine.anatomical_structure ,clinical research ,hepatology ,Bone marrow ,Stem cell ,business ,liver transplantation/hepatology ,Immunosuppressive Agents - Abstract
Mesenchymal stromal cells (MSC) have emerged as a promising therapy to minimize the immunosuppressive regimen or induce tolerance in solid organ transplantation. In this randomized open-label phase Ib/IIa clinical trial, 20 liver transplant patients were randomly allocated (1:1) to receive a single pre-transplant intravenous infusion of third-party bone marrow-derived MSC or standard of care alone. The primary end-point was the safety profile of MSC administration during the one-year follow-up. Nineteen patients completed the study, and none of those who received MSC experienced infusion-related complications. The incidence of serious and non-serious adverse events was similar in the two groups. Circulating Treg/memory Treg and tolerant NK subset of CD56bright NK cells increased slightly over baseline, albeit not to a statistically significant extent, in MSC-treated patients but not in the control group. Graft function and survival, as well as histologic parameters and intragraft expression of tolerance-associated transcripts in 1-year protocol biopsies were similar in the two groups. In conclusion, pre-transplant MSC infusion in liver transplant recipients was safe and induced mild positive changes in immunoregulatory T and NK cells in the peripheral blood. This study opens the way for a trial on possible tolerogenic efficacy of MSC in liver transplantation. ClinicalTrials.gov identifier: NCT02260375.
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- 2021
27. Immune Reconstitution Following Infusion of Autologous Blinatumomab Expanded T-Cells (BET) in CD20+ Indolent Non-Hodgkin Lymphomas and Chronic Lymphocytic Leukemia Receiving Front-Line Treatment with Fludarabine-Cyclophosphamide-Rituximab or Bendamustine-Rituximab
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Giuseppe Gritti, Federico Lussana, Silvia Ferrari, Anna Maria Barbui, Francesco Landi, Giulia Quaresmini, Gianmaria Borleri, Muriel Paganessi, Chiara Pavoni, Elisa Gotti, Chiara Capelli, Josee Golay, Martino Introna, and Alessandro Rambaldi
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Immunology ,Cell Biology ,Hematology ,Biochemistry - Published
- 2022
28. Final Results of Phase I/II Study of Donor-Derived CAR T Cells Engineered with Sleeping Beauty in Pediatric and Adult Patients with B-Cell Acute Lymphoblastic Leukemia Relapsed Post-HSCT
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Federico Lussana, Chiara Francesca Magnani, Giuseppe Gaipa, Stefania Galimberti, Giuseppe Gritti, Daniela Belotti, Sara Napolitano, Chiara Buracchi, Gian Maria Borleri, Benedetta Rambaldi, Giuliana Rizzuto, Anna Grassi, Muriel Paganessi, Silvia Ferrari, Sarah Tettamanti, Giovanni Gazzaniga, Chiara Capelli, Elisa Gotti, Martino Introna, Adriana Balduzzi, Maria Grazia Valsecchi, Giuseppe Dastoli, Alessandro Rambaldi, and Andrea Biondi
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Immunology ,Cell Biology ,Hematology ,Biochemistry - Published
- 2022
29. Carcik-CD19 Cells Expand In Vivo Toward a CD8+ Memory Phenotype and Their Persistence Is Associated with a Longer Duration of Response
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Benedetta Rambaldi, Stefania Galimberti, Giuliana Rizzuto, Chiara Francesca Magnani, Chiara Buracchi, Giulia Risca, Martina Paredi, Daniela Belotti, Alex Moretti, Marianna Ponzo, Sarah Tettamanti, Gian Maria Borleri, Cristian Meli, Muriel Paganessi, Silvia Zaninelli, Elisa Gotti, Chiara Capelli, Martino Introna, Federico Lussana, Giuseppe Gritti, Silvia Ferrari, Anna Grassi, Sara Napolitano, Adriana Balduzzi, Maria Grazia Valsecchi, Giuseppe Dastoli, Alessandro Rambaldi, Andrea Biondi, and Giuseppe Gaipa
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Immunology ,Cell Biology ,Hematology ,Biochemistry - Published
- 2022
30. Sleeping Beauty–engineered CAR T cells achieve antileukemic activity without severe toxicities
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Gianmaria Borleri, Adriana Balduzzi, Benedetta Cabiati, Michele Quaroni, Martino Introna, Grazia Fazio, Sara Napolitano, Chiara Buracchi, Stefania Cesana, Giovanni Cazzaniga, Giuseppe Gaipa, Giuseppe Gritti, Giada Matera, Silvia Zaninelli, Ettore Biagi, Andrea Biondi, Stefania Galimberti, Federico Lussana, Fabrizio Benedicenti, Attilio Rovelli, Giuseppe Dastoli, Eugenio Montini, Sarah Tettamanti, Valentina Colombo, Andrea Calabria, Maria Grazia Valsecchi, Alessandro Rambaldi, Silvia Ferrari, Chiara F. Magnani, Daniela Belotti, Magnani, C, Gaipa, G, Lussana, F, Belotti, D, Gritti, G, Napolitano, S, Matera, G, Cabiati, B, Buracchi, C, Borleri, G, Fazio, G, Zaninelli, S, Tettamanti, S, Cesana, S, Colombo, V, Quaroni, M, Cazzaniga, G, Rovelli, A, Biagi, E, Galimberti, S, Calabria, A, Benedicenti, F, Montini, E, Ferrari, S, Introna, M, Balduzzi, A, Valsecchi, M, Dastoli, G, Rambaldi, A, and Biondi, A
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Adult ,Male ,0301 basic medicine ,Oncology ,medicine.medical_specialty ,Adolescent ,CD3 ,medicine.medical_treatment ,Hematopoietic stem cell transplantation ,Immunotherapy, Adoptive ,CD19 ,03 medical and health sciences ,0302 clinical medicine ,Refractory ,Precursor B-Cell Lymphoblastic Leukemia-Lymphoma ,Internal medicine ,Leukemias ,medicine ,Humans ,Clinical Trials ,Child ,Hematology ,biology ,Cancer gene therapy ,business.industry ,Hematopoietic Stem Cell Transplantation ,Infant ,Cancer ,General Medicine ,Immunotherapy ,Allografts ,medicine.disease ,Clinical trial ,030104 developmental biology ,Child, Preschool ,030220 oncology & carcinogenesis ,biology.protein ,Female ,Clinical Medicine ,business - Abstract
BACKGROUND: Chimeric antigen receptor (CAR) T cell immunotherapy has resulted in complete remission (CR) and durable response in highly refractory patients. However, logistical complexity and high costs of manufacturing autologous viral products limit CAR T cell availability. METHODS: We report the early results of a phase I/II trial in B cell acute lymphoblastic leukemia (B-ALL) patients relapsed after allogeneic hematopoietic stem cell transplantation (HSCT) using donor-derived CD19 CAR T cells generated with the Sleeping Beauty (SB) transposon and differentiated into cytokine-induced killer (CIK) cells. RESULTS: The cellular product was produced successfully for all patients from the donor peripheral blood (PB) and consisted mostly of CD3(+) lymphocytes with 43% CAR expression. Four pediatric and 9 adult patients were infused with a single dose of CAR T cells. Toxicities reported were 2 grade I and 1 grade II cytokine-release syndrome (CRS) cases at the highest dose in the absence of graft-versus-host disease (GVHD), neurotoxicity, or dose-limiting toxicities. Six out of 7 patients receiving the highest doses achieved CR and CR with incomplete blood count recovery (CRi) at day 28. Five out of 6 patients in CR were also minimal residual disease negative (MRD(–)). Robust expansion was achieved in the majority of the patients. CAR T cells were measurable by transgene copy PCR up to 10 months. Integration site analysis showed a positive safety profile and highly polyclonal repertoire in vitro and at early time points after infusion. CONCLUSION: SB-engineered CAR T cells expand and persist in pediatric and adult B-ALL patients relapsed after HSCT. Antileukemic activity was achieved without severe toxicities. TRIAL REGISTRATION: ClinicalTrials.gov NCT03389035. FUNDING: This study was supported by grants from the Fondazione AIRC per la Ricerca sul Cancro (AIRC); Cancer Research UK (CRUK); the Fundación Científica de la Asociación Española Contra el Cáncer (FC AECC); Ministero Della Salute; Fondazione Regionale per la Ricerca Biomedica (FRRB).
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- 2020
31. Kidney transplant tolerance associated with remote autologous mesenchymal stromal cell administration
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Martino Introna, E. Longhi, Eliana Gotti, Norberto Perico, Valentina Portalupi, Federica Casiraghi, Giuseppe Remuzzi, Marta Todeschini, Alessandro Villa, Anna Rita Plati, Marilena Mister, Monica Cortinovis, and Flavio Gaspari
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0301 basic medicine ,Adult ,Male ,Stromal cell ,immunoregulation ,kidney transplantation ,Mesenchymal Stem Cell Transplantation ,Kidney transplant ,Immune tolerance ,03 medical and health sciences ,0302 clinical medicine ,Immune system ,Human Clinical Article ,Medicine ,Humans ,Transplantation, Homologous ,lcsh:QH573-671 ,Kidney transplantation ,Kidney ,lcsh:R5-920 ,tolerance ,business.industry ,lcsh:Cytology ,Mesenchymal stem cell ,Mesenchymal Stem Cells ,Cell Biology ,General Medicine ,medicine.disease ,Autologous bone ,030104 developmental biology ,medicine.anatomical_structure ,immunosuppression withdrawal ,Cancer research ,Transplantation Tolerance ,business ,mesenchymal stromal cells ,lcsh:Medicine (General) ,030217 neurology & neurosurgery ,Developmental Biology - Abstract
Here we report the case of successful immune tolerance induction in a living‐donor kidney transplant recipient remotely treated with autologous bone marrow‐derived mesenchymal stromal cells (MSC). This case report, which to the best of our knowledge is the first in the world in this setting, provides evidence that the modulation of the host immune system with MSC can enable the safe withdrawal of maintenance immunosuppressive drugs while preserving optimal long‐term kidney allograft function., This report provides the evidence that autologous BM‐MSC infusion induces a pro‐tolerogenic immune environment, thus allowing complete discontinuation of antirejection drugs in kidney transplantation
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- 2020
32. Immunotherapy for HER2-Positive Breast Cancer: Clinical Evidence and Future Perspectives
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Elisa Agostinetto, Filippo Montemurro, Fabio Puglisi, Carmen Criscitiello, Giampaolo Bianchini, Lucia Del Mastro, Martino Introna, Carlo Tondini, Armando Santoro, and Alberto Zambelli
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immune checkpoint inhibitors ,Cancer Research ,Oncology ,CAR-T cells ,antibody–drug conjugates ,HER2-positive breast cancer ,immunotherapy ,vaccines ,skin and connective tissue diseases - Abstract
Breast cancer is the most common malignancy among women worldwide, and HER2-positive breast cancer accounts for approximately 15% of all breast cancer diagnoses. The advent of HER2-targeting therapies has dramatically improved the survival of these patients, significantly reducing their risk of recurrence and death. However, as a significant proportion of patients ultimately develop resistance to these therapies, it is extremely important to identify new treatments to further improve their clinical outcomes. Immunotherapy has revolutionized the treatment and history of several cancer types, and it has already been approved as a standard of care for patients with triple-negative breast cancer. Based on a strong preclinical rationale, immunotherapy in HER2-positive breast cancer represents an intriguing field that is currently under clinical investigation. There is a close interplay between HER2-targeting therapies (both approved and under investigation) and the immune system, and several new immunotherapeutic strategies, including immune checkpoint inhibitors, CAR-T cells and therapeutic vaccines, are being studied in this disease. In this narrative review, we discuss the clinical evidence and the future perspectives of immunotherapy for patients with HER2-positive breast cancer.
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- 2022
33. A comprehensive report of long-term stability data for a range ATMPs: A need to develop guidelines for safe and harmonized stability studies
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Chiara Capelli, Simona Frigerio, Daniela Lisini, Sara Nava, Giuseppe Gaipa, Daniela Belotti, Benedetta Cabiati, Silvia Budelli, Lorenza Lazzari, Jessica Bagnarino, Matteo Tanzi, Patrizia Comoli, Norberto Perico, Martino Introna, Josée Golay, Capelli, C, Frigerio, S, Lisini, D, Nava, S, Gaipa, G, Belotti, D, Cabiati, B, Budelli, S, Lazzari, L, Bagnarino, J, Tanzi, M, Comoli, P, Perico, N, Introna, M, and Golay, J
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Cryopreservation ,Cancer Research ,Transplantation ,Cell Survival ,Immunology ,Cell Differentiation ,Cell Biology ,stability study ,potency assay ,Immunophenotyping ,Good Manufacturing Practice ,Oncology ,Immunology and Allergy ,shelf life ,advanced therapy medicinal product ,quality control ,Genetics (clinical) - Abstract
Background aims: Advanced therapy medicinal products (ATMPs) are novel drugs based on genes, cells or tissues developed to treat many different diseases. Stability studies of each new ATMP need to be performed to define its shelf life and guarantee efficacy and safety upon infusion, and these are presently based on guidelines originally drafted for standard pharmaceutical drugs, which have properties and are stored in conditions quite different from cell products. The aim of this report is to provide evidence-based information for stability studies on ATMPs that will facilitate the interlaboratory harmonization of practices in this area. Methods: We have collected and analyzed the results of stability studies on 19 different cell-based experimental ATMPs, produced by five authorized cell factories forming the Lombardy “Plagencell network” for use in 36 approved phase I/II clinical trials; most were cryopreserved and stored in liquid nitrogen vapors for 1 to 13 years. Results: The cell attributes collected in stability studies included cell viability, immunophenotype and potency assays, in particular immunosuppression, cytotoxicity, cytokine release and proliferation/differentiation capacity. Microbiological attributes including sterility, endotoxin levels and mycoplasma contamination were also analyzed. All drug products (DPs), cryopreserved in various excipients containing 10% DMSO and in different primary containers, were very stable long term at
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- 2021
34. Protective Effects of Human Nonrenal and Renal Stromal Cells and Their Conditioned Media in a Rat Model of Chronic Kidney Disease
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Marina Morigi, Claudia Elisa Carminati, Chiara Capelli, Cinzia Rota, Ariela Benigni, Carlamaria Zoja, Daniëlle G. Leuning, Domenico Cerullo, Martino Introna, Ton J. Rabelink, Giuseppe Remuzzi, Barbara Imberti, Valerie A. Luyckx, Monica Locatelli, Daniela Corna, and Anna Pezzotta
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0301 basic medicine ,medicine.medical_specialty ,Stromal cell ,030232 urology & nephrology ,Biomedical Engineering ,renal repair ,lcsh:Medicine ,Podocyte ,Cell therapy ,Nephrin ,03 medical and health sciences ,0302 clinical medicine ,Fibrosis ,Internal medicine ,medicine ,Animals ,Humans ,Renal Insufficiency, Chronic ,stromal cells ,Transplantation ,biology ,business.industry ,Mesenchymal stem cell ,lcsh:R ,Cell Biology ,medicine.disease ,Rats ,Disease Models, Animal ,030104 developmental biology ,medicine.anatomical_structure ,Endocrinology ,conditioned medium ,biology.protein ,renal perivascular cells ,Original Article ,business ,Myofibroblast ,chronic kidney disease ,Kidney disease - Abstract
Mesenchymal stromal cells (MSCs) are emerging as a novel therapeutic option for limiting chronic kidney disease progression. Conditioned medium (CM) containing bioactive compounds could convey similar benefits, avoiding the potential risks of cell therapy. This study compared the efficacy of nonrenal and renal cell-based therapy with the corresponding CM in rats with renal mass reduction (RMR). Infusions of human kidney stromal cells (kPSCs) and CM-kPSCs, but not umbilical cord (uc) MSCs or CM-ucMSCs, reduced proteinuria and preserved podocyte number and nephrin expression in RMR rats. Glomerular fibrosis, microvascular rarefaction, and apoptosis were reduced by all treatments, while the peritubular microvascular loss was reduced by kPSCs and CM-kPSCs treatment only. Importantly, kPSCs and CM-kPSCs reduced NG2-positive pericytes, and all therapies reduced α-smooth muscle actin expression, indicating reduced myofibroblast expansion. Treatment with kPSCs also significantly inhibited the accumulation of ED1-positive macrophages in the renal interstitium of RMR rats. These findings demonstrate that the CM of ucMSCs and kPSCs confers similar renoprotection as the cells. kPSCs and CM-kPSCs may be superior in attenuating chronic renal injury as a cell source.
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- 2020
35. Targeting CD33 in Chemoresistant AML Patient-Derived Xenografts by CAR-CIK Cells Modified with an Improved SB Transposon System
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Chiara F. Magnani, Marta Serafini, Maria Grazia Valsecchi, Tamás Raskó, Vincenzo Perriello, Ettore Biagi, Sarah Tettamanti, Gaia Alberti, Martino Introna, Zsuzsanna Izsvák, Maria Caterina Rotiroti, Felix Lundberg, Giuseppe Dastoli, Claudia Cappuzzello, Silvia Arcangeli, Chiara Buracchi, Amit Pande, Andrea Biondi, Stefania Galimberti, Rotiroti, M, Buracchi, C, Arcangeli, S, Galimberti, S, Valsecchi, M, Perriello, V, Rasko, T, Alberti, G, Magnani, C, Cappuzzello, C, Lundberg, F, Pande, A, Dastoli, G, Introna, M, Serafini, M, Biagi, E, Izsvák, Z, Biondi, A, and Tettamanti, S
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Myeloid ,Cell Transplantation ,THP-1 Cells ,medicine.medical_treatment ,CD33 ,Sialic Acid Binding Ig-like Lectin 3 ,Adoptive ,Drug Resistance ,Transposases ,Mice, SCID ,Immunotherapy, Adoptive ,Cell therapy ,Mice ,0302 clinical medicine ,AML ,Mice, Inbred NOD ,Drug Discovery ,Receptors ,Medicine ,Cell Engineering ,0303 health sciences ,Receptors, Chimeric Antigen ,Leukemia ,Cytokine-induced killer cell ,Gene Transfer Techniques ,CAR ,Leukemia, Myeloid, Acute ,medicine.anatomical_structure ,Treatment Outcome ,030220 oncology & carcinogenesis ,Molecular Medicine ,Heterografts ,Original Article ,Immunotherapy ,cytokine-induced killer cells ,Context (language use) ,Acute ,SCID ,03 medical and health sciences ,Experimental ,Genetics ,Animals ,Humans ,Molecular Biology ,B cell ,030304 developmental biology ,Pharmacology ,Leukemia, Experimental ,business.industry ,Chimeric Antigen ,Genetic Therapy ,Sleeping Beauty transposon system ,Xenograft Model Antitumor Assays ,Chimeric antigen receptor ,Drug Resistance, Neoplasm ,non-viral gene transfer ,Cancer research ,Sleeping Beauty transposon ,Feasibility Studies ,Inbred NOD ,Neoplasm ,business ,cytokine-induced killer cell - Abstract
The successful implementation of chimeric antigen receptor (CAR)-T cell therapy in the clinical context of B cell malignancies has paved the way for further development in the more critical setting of acute myeloid leukemia (AML). Among the potentially targetable AML antigens, CD33 is insofar one of the main validated molecules. Here, we describe the feasibility of engineering cytokine-induced killer (CIK) cells with a CD33.CAR by using the latest optimized version of the non-viral Sleeping Beauty (SB) transposon system "SB100X-pT4." This offers the advantage of improving CAR expression on CIK cells, while reducing the amount of DNA transposase as compared to the previously employed "SB11-pT" version. SB-modified CD33.CAR-CIK cells exhibited significant antileukemic activity invitro and invivo in patient-derived AML xenograft models, reducing AML development when administered as an "early treatment" and delaying AML progression in mice with established disease. Notably, by exploiting an already optimized xenograft chemotherapy model that mimics human induction therapy in mice, we demonstrated for the first time that CD33.CAR-CIK cells are also effective toward chemotherapy resistant/residual AML cells, further supporting its future clinical development and implementation within the current standard regimens.
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- 2020
36. Utility of routine evaluation of sterility of cellular therapy products with or without extensive manipulation: Best practices and clinical significance
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Mara Magri, Chiara Capelli, Marco Passera, Elisa Gotti, Josée Golay, Francesca Vailati, Gianmaria Borleri, Martino Introna, Alessandro Rambaldi, Claudio Farina, and Olga Pedrini
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Cancer Research ,medicine.medical_specialty ,Time Factors ,Sterility ,medicine.drug_class ,medicine.medical_treatment ,Immunology ,Antibiotics ,Cell- and Tissue-Based Therapy ,Context (language use) ,Hematopoietic stem cell transplantation ,030204 cardiovascular system & hematology ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,Internal medicine ,medicine ,Humans ,Immunology and Allergy ,Good manufacturing practice ,Genetics (clinical) ,Transplantation ,business.industry ,Mesenchymal stem cell ,Hematopoietic Stem Cell Transplantation ,Sterilization ,Mesenchymal Stem Cells ,Cell Biology ,Hematopoietic Stem Cells ,Culture Media ,medicine.anatomical_structure ,Oncology ,chemistry ,Blood Component Removal ,Bone marrow ,ATMP ,business ,030215 immunology - Abstract
Background We analyzed the results of routine sterility testing performed in our center over the last 10 years, in the context both hematopoietic stem cell transplantation (HSCT) and Advanced Therapeutic Medicinal Products (ATMPs). Methods For sterility tests 14-day cultures were performed in culture media detecting aerobic and anaerobic microorganisms. Results In this study, 22/1643 (1.3%) of apheretic products for autologous or allogeneic HSCT were contaminated, whereas 14/73 bone marrow (BM) harvests (17.8%) were positive. In 22 cases, the contaminated HSCs were infused to patients, but there was no evidence of any adverse impact of contamination on the hematologic engraftment or on infections. Indeed none of the five positive hemocultures detected in patients following infusion could be linked to the contaminated stem cell product. Our Cell Factory also generated 286 ATMPs in good manufacturing practice (GMP) conditions since 2007 and all final products were sterile. In three cases of mesenchymal stromal cell expansions, the starting BM harvests were contaminated, but the cell products at the end of expansion were sterile, presumably thanks to the presence of an antibiotic in the culture medium. Discussion The decreased rate of contamination of cell harvests observed with time suggests that routine sterility testing and communication of the results to the collecting centers may improve clinical practices. Furthermore, we recommend the use of antibiotics in the medium for ATMP expansion, to decrease the likelihood of expanding microorganisms within clean rooms. Finally we discuss the costs of sterility testing of ATMPs by GMP-approved external laboratories.
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- 2018
37. Donor-Derived CAR T Cells Engineered with Sleeping Beauty in Pediatric and Adult Patients with Acute Lymphoblastic Leukemia Relapsed Post-HSCT
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Giovanni Cazzaniga, Chiara F. Magnani, Gianluca Cavallaro, Giuseppe Gaipa, Giuseppe Dastoli, Gian Maria Borleri, Andrea Biondi, Sarah Tettamanti, Silvia Ferrari, Giuliana Rizzuto, Adriana Balduzzi, Federico Lussana, Maria Grazia Valsecchi, Giuseppe Gritti, Benedetta Rambaldi, Alessandro Rambaldi, Chiara Buracchi, Martino Introna, Sara Napolitano, Daniela Belotti, Silvia Zaninelli, and Stefania Galimberti
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Oncology ,medicine.medical_specialty ,Adult patients ,business.industry ,Lymphoblastic Leukemia ,Immunology ,Cell Biology ,Hematology ,Biochemistry ,Internal medicine ,medicine ,Donor derived ,Car t cells ,business - Abstract
Introduction Allogeneic Chimeric Antigen Receptor (CAR) T cells engineered with non-viral methods offer a modality to reduce costs and logistical complexity of the viral process and allow lymphodepleted patients to access CAR T cell treatment. We recently proposed the use of Sleeping Beauty (SB) transposon to engineer donor-derived T cells differentiated according to the cytokine-induced killer (CIK) cell protocol (Magnani CF et al. J Clin Invest. 2021). We report here outcomes on B-cell acute lymphoblastic leukemia (B-ALL) patients, relapsing after transplantation, treated with donor-derived anti-CD19 CAR T cells (CARCIK-CD19). Methods We conducted an academic, multi-center, phase I/II dose-escalation trial in patients relapsed after allogeneic hematopoietic stem cell transplantation (HSCT). The infusion product was manufactured in-house starting from 50 mL of peripheral blood from the HSCT donor by electroporation with GMP-grade plasmids. All patients underwent lymphodepletion with Fludarabine (30 mg/m 2/day x 4 days) and Cyclophosphamide (500 mg/m 2/day x 2 days), before proceeding to CARCIK-CD19 infusion. We used the Bayesian Optimal Interval (BOIN) design to define a four-dose escalation scheme. Primary objectives were to define the Maximum Tolerated Dose (MTD), safety, and feasibility. Secondary objectives included the assessment of complete hematologic response (CR), duration of response (DOR), progression-free (PFS), event-free (EFS), and overall survival (OS). This study was registered at ClinicalTrials.gov, NCT03389035. Results From January 2018 to June 2021, a total of 32 patients were screened, 26 enrolled (6 children and 20 adults) and 21 infused (4 children and 17 adults). Reasons for not receiving infusion included consent withdrawal (N=1), disease progression not controlled by bridging therapy (N=3), acquisition of myeloid phenotype (N=1). The median number of prior therapies was 4 (range, 1-7) with a median time interval from HSCT to relapse of 9 months. The median BM blasts was 60% (range, 5-100%) at enrollment and 7% (range, 0-96%) post lymphodepletion. Of the 21 patients infused, CARCIK-CD19 were obtained by HLA-identical sibling (n=6, 29%), matched unrelated (n= 7, 33%), and haploidentical donors (n=8, 38%). Three patients (14%) received the first dose level of 1x10 6 CARCIK-CD19 cells/Kg, three (14%) the second of 3x10 6, and three (14%) the third of 7.5x10 6 whereas 12 patients (57%) received the fourth and last planned dose level of 15x10 6 cells/Kg, as no dose limiting toxicity (DLT) was observed. CRS was observed in six patients (three grade I and three grade II) and immune effector cell-associated neurotoxicity in two patients at the highest dose. Although 9 out of 21 had experienced acute or chronic graft-versus-host disease (GvHD) after the previous HSCT, secondary GvHD was never induced by CARCIK-CD19. Complete response was achieved by 13 out of 21 patients (61.9%, 95%CI=38-82%) and by 11 out of 15 patients treated with the 2 highest doses (73.3%, 95%CI=45-92%). Eleven of these responders were MRD-negative. Notably, the type of donor did not influence the achievement of CR 28 days post-infusion. At a median follow up of 21.6 months (range, 1.0-38.4 months), 10 patients (47.6%) are alive in CR (9 in the 2 highest dose levels). Overall, the median OS and EFS were 9.7 and 3.2 months, respectively, with a median DOR of 4.0 months (range, 1.0-23.5 months). Patients in CR at 28-days had a 6-months relapse-free survival of 48.4% (SE=14.9). EFS at 6 months was 26.5% (SE=9.9) and OS was 67.6% (SE=11.1). Among the 13 patients who achieved CR, two children underwent consolidation with a second allo-HSCT in complete remission. Adult patients did not receive any additional anti-leukemic therapies unless a relapse occurred, and four of them remained in remission and alive (+24, +9, +6, and +4 months). Robust CARCIK-CD19 cell expansion was achieved in most patients and CARCIK-CD19 cells were measurable for up to 22 months. Conclusions SB-engineered CAR T cells induce sustained responses in B-ALL patients relapsed after HSCT irrespective of the donor type and without severe toxicities. Disclosures Lussana: Incyte: Honoraria; Pfizer: Honoraria; Astellas Pharma: Honoraria; Amgen: Honoraria. Gritti: Takeda: Consultancy; Roche: Consultancy; Kite Gilead: Consultancy; IQvia: Consultancy; Italfarmaco: Consultancy; Clinigen: Consultancy. Biondi: Incyte: Consultancy, Other: Advisory Board; Bluebird: Other: Advisory Board; Novartis: Honoraria; Amgen: Honoraria; Colmmune: Honoraria.
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- 2021
38. International Forum on GMP-grade human platelet lysate for cell propagation: summary
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Katharina Schallmoser, Michaela Öller, M. Waidmann, Markus Rojewski, Karen Bieback, Martino Introna, Olafur E. Sigurjonsson, Daniela Fioravanti, Sandra Mjoll Jonsdottir-Buch, Luca Pierelli, Jo Anna Reems, Miquel Lozano, Denese C. Marks, Johanna Nystedt, Sandra Laner-Plamberger, H. Montazeri, Ramin Lotfi, A. Falanga, Jan Pierce, Eva Rohde, Richard Schäfer, Dirk Strunk, Amber Preslar, H. Schennach, Minoko Takanashi, C. Capelli, P. Iudicone, O. Lόpez-Villar, Thierry Burnouf, G. Gstraunthaler, Hubert Schrezenmeier, Tamam Bakchoul, Yen Siew Loh, Strunk, D, Lozano, M, Marks, D, Loh, Y, Gstraunthaler, G, Schennach, H, Rohde, E, Laner-Plamberger, S, Öller, M, Nystedt, J, Lotfi, R, Rojewski, M, Schrezenmeier, H, Bieback, K, Schäfer, R, Bakchoul, T, Waidmann, M, Jonsdottir-Buch, S, Montazeri, H, Sigurjonsson, O, Iudicone, P, Fioravanti, D, Pierelli, L, Introna, M, Capelli, C, Falanga, A, Takanashi, M, Lόpez-Villar, O, Burnouf, T, Reems, J, Pierce, J, Preslar, A, and Schallmoser, K
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0301 basic medicine ,Lysis ,business.industry ,Cell ,Human platelet ,platelet lysate ,Hematology ,General Medicine ,030204 cardiovascular system & hematology ,03 medical and health sciences ,030104 developmental biology ,0302 clinical medicine ,medicine.anatomical_structure ,no keywords for this article ,Immunology ,medicine ,Platelet lysate ,Platelet ,business - Published
- 2017
39. Human mesenchymal stromal cells transplanted into mice stimulate renal tubular cells and enhance mitochondrial function
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Lorena Longaretti, Giuseppe Remuzzi, Caterina Mele, Marina Morigi, Chiara Capelli, Ariela Benigni, Martino Introna, Matteo Breno, Cinzia Rota, Daniela Rottoli, Marina Noris, Daniela Corna, Sara Conti, and Luca Perico
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0301 basic medicine ,SIRT3 ,Science ,General Physics and Astronomy ,Mice, SCID ,Mesenchymal Stem Cell Transplantation ,Article ,General Biochemistry, Genetics and Molecular Biology ,Mice ,03 medical and health sciences ,Adenosine Triphosphate ,0302 clinical medicine ,Sirtuin 3 ,medicine ,Animals ,Humans ,lcsh:Science ,Cell Proliferation ,Multidisciplinary ,biology ,Cell growth ,Regeneration (biology) ,Mesenchymal stem cell ,Acute kidney injury ,Mesenchymal Stem Cells ,General Chemistry ,Acute Kidney Injury ,NAD ,medicine.disease ,Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1-alpha ,Mitochondria ,Cell biology ,Kidney Tubules ,030104 developmental biology ,Mitochondrial biogenesis ,030220 oncology & carcinogenesis ,Immunology ,Sirtuin ,biology.protein ,Female ,lcsh:Q ,NAD+ kinase ,Cisplatin - Abstract
Mesenchymal stromal cells (MSCs) are renoprotective and drive regeneration following injury, although cellular targets of such an effect are still ill-defined. Here, we show that human umbilical cord (UC)-MSCs transplanted into mice stimulate tubular cells to regain mitochondrial mass and function, associated with enhanced microtubule-rich projections that appear to mediate mitochondrial trafficking to create a reparative dialogue among adjacent tubular cells. Treatment with UC-MSCs in mice with cisplatin-induced acute kidney injury (AKI) regulates mitochondrial biogenesis in proximal tubuli by enhancing PGC1α expression, NAD+ biosynthesis and Sirtuin 3 (SIRT3) activity, thus fostering antioxidant defenses and ATP production. The functional role of SIRT3 in tubular recovery is highlighted by data that in SIRT3-deficient mice with AKI, UC-MSC treatment fails to induce renoprotection. These data document a previously unrecognized mechanism through which UC-MSCs facilitate renal repair, so as to induce global metabolic reprogramming of damaged tubular cells to sustain energy supply., Mesenchymal stromal cells drive renal regeneration following injury. Here, the authors show that human mesenchymal stromal cells, when transplanted into mice with acute kidney injury, stimulate renal tubular cell growth and enhance mitochondrial function via SIRT3.
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- 2017
40. The specific Bruton tyrosine kinase inhibitor acalabrutinib (ACP-196) shows favorable in vitro activity against chronic lymphocytic leukemia B cells with CD20 antibodies
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Greta Ubiali, Josée Golay, and Martino Introna
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CD20 ,biology ,Chronic lymphocytic leukemia ,Hematology ,medicine.disease ,In vitro ,03 medical and health sciences ,Leukemia ,chemistry.chemical_compound ,0302 clinical medicine ,chemistry ,Antigen ,immune system diseases ,hemic and lymphatic diseases ,030220 oncology & carcinogenesis ,Ibrutinib ,Immunology ,biology.protein ,medicine ,Cancer research ,Bruton's tyrosine kinase ,Acalabrutinib ,030215 immunology - Abstract
Acalabrutinib is a new, irreversible Bruton tyrosine kinase (BTK) inhibitor, reported to be more selective than ibrutinib.[1][1]–[3][2] BTK inhibitors are often combined with anti-CD20 antibodies in the clinic, but ibrutinib interferes with some of the cell-mediated mechanisms of action of anti
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- 2017
41. Induction of Mouse Lung Injury by Endotracheal Injection of Bleomycin
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Mauro Provinciali, Silvia Agarbati, Armando Gabrielli, Martino Introna, A. Grieco, Chiara Paolini, Chiara Capelli, Fiorenza Orlando, Gianluca Moroncini, and C. Tonnini
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Pathology ,medicine.medical_specialty ,General Chemical Engineering ,Pulmonary Fibrosis ,Lung injury ,Bleomycin ,Mesenchymal Stem Cell Transplantation ,General Biochemistry, Genetics and Molecular Biology ,Umbilical Cord ,03 medical and health sciences ,chemistry.chemical_compound ,Mice ,Fibrosis ,In vivo ,Pulmonary fibrosis ,Medicine ,Animals ,0303 health sciences ,General Immunology and Microbiology ,business.industry ,General Neuroscience ,030305 genetics & heredity ,Mesenchymal stem cell ,Mesenchymal Stem Cells ,Lung Injury ,respiratory system ,medicine.disease ,Pathophysiology ,respiratory tract diseases ,Mice, Inbred C57BL ,Trachea ,Disease Models, Animal ,chemistry ,Systemic administration ,Female ,business - Abstract
Pulmonary fibrosis is a hallmark of several human lung diseases with a different etiology. Since current therapies are rather limited, mouse models continue to be an essential tool for developing new antifibrotic strategies. Here we provide an effective method to investigate in vivo antifibrotic activity of human mesenchymal stromal cells obtained from whole umbilical cord (hUC-MSC) in attenuating bleomycin-induced lung injury. C57BL/6 mice receive a single endotracheal injection of bleomycin (1.5 U/kg body weight) followed by a double infusion of hUC-MSC (2.5 x 105) into the tail vein, 24 h and 7 days after the bleomycin administration. Upon sacrifice at days 8, 14, or 21, inflammatory and fibrotic changes, collagen content, and hUC-MSC presence in explanted lung tissue are analyzed. The injection of bleomycin into the mouse trachea allows the direct targeting of the lungs, leading to extensive pulmonary inflammation and fibrosis. The systemic administration of a double dose of hUC-MSC results in the early blunting of the bleomycin-induced lung injury. Intravenously infused hUC-MSC are transiently engrafted into the mouse lungs, where they exert their anti-inflammatory and antifibrotic activity. In conclusion, this protocol has been successfully applied for the preclinical testing of hUC-MSC in an experimental mouse model of human pulmonary fibrosis. However, this technique can be easily extended both to study the effect of different endotracheally administered substances on the pathophysiology of the lungs and to validate new anti-inflammatory and antifibrotic systemic therapies.
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- 2019
42. Manufacturing Mesenchymal Stromal Cells for the Treatment of Graft-versus-Host Disease: A Survey among Centers Affiliated with the European Society for Blood and Marrow Transplantation
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Maria Ester Bernardo, Katarina Le Blanc, Yves Beguin, Martino Introna, Cristina Trento, Jane F. Apperley, Chiara Bonini, Francesco Dazzi, Fermín Sánchez-Guijo, Selim Kuçi, Ulrike Köhl, Helene Roelofs, Giuseppe Gaipa, Lorenza Lazzari, Jürgen Kuball, Dirk Strunk, Antonio Galleu, Martin Bornhäuser, Maria Antonietta Avanzini, Christian Chabannon, Arnon Nagler, Adomas Bukauskas, Willem E. Fibbe, Ann Van Campenhout, Trento, Cristina, Bernardo, Maria Ester, Nagler, Arnon, Kuçi, Selim, Bornhäuser, Martin, Köhl, Ulrike, Strunk, Dirk, Galleu, Antonio, Sanchez-Guijo, Fermin, Gaipa, Giuseppe, Introna, Martino, Bukauskas, Adoma, Le Blanc, Katarina, Apperley, Jane, Roelofs, Helene, Van Campenhout, Ann, Beguin, Yve, Kuball, Jürgen, Lazzari, Lorenza, Avanzini, Maria Antonietta, Fibbe, Willem, Chabannon, Christian, Bonini, Chiara, and Dazzi, Francesco
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0301 basic medicine ,Oncology ,medicine.medical_specialty ,Immunology ,Cellular therapy ,Mesenchymal stromal cells ,Graft vs Host Disease ,Umbilical cord ,Graft-versus-host disease ,Article ,Cell therapy ,03 medical and health sciences ,Release criteria ,hematología ,Internal medicine ,Surveys and Questionnaires ,medicine ,Humans ,Transplantation ,Hematology ,business.industry ,3205.04 Hematología ,Mesenchymal stem cell ,1103 Clinical Sciences ,Mesenchymal Stem Cells ,medicine.disease ,Clinical trial ,Europe ,Manufacturing ,030104 developmental biology ,medicine.anatomical_structure ,Product specification ,células del estroma mesenquimatoso ,enfermedad injerto contra huésped ,Bone marrow ,business - Abstract
Highlights • Seventeen EBMT centers participated to a questionnaire on MSC manufacturing. • 88% of centers manufacture MSC from bone marrow and only 2 centers from umbilical cord. • Human platelet lysate has replaced bovine serum as culture medium supplement. • Release criteria extensively differ among centers. • The results highlight the need to harmonize MSC manufacturing., The immunosuppressive properties of mesenchymal stromal cells (MSC) have been successfully tested to control clinical severe graft-versus host disease and improve survival. However, clinical studies have not yet provided conclusive evidence of their efficacy largely because of lack of patients’ stratification criteria. The heterogeneity of MSC preparations is also a major contributing factor, as manufacturing of therapeutic MSC is performed according to different protocols among different centers. Understanding the variability of the manufacturing protocol would allow a better comparison of the results obtained in the clinical setting among different centers. In order to acquire information on MSC manufacturing we sent a questionnaire to the European Society for Blood and Marrow Transplantation centers registered as producing MSC. Data from 17 centers were obtained and analyzed by means of a 2-phase questionnaire specifically focused on product manufacturing. Gathered information included MSC tissue sources, MSC donor matching, medium additives for ex vivo expansion, and data on MSC product specification for clinical release. The majority of centers manufactured MSC from bone marrow (88%), whilst only 2 centers produced MSC from umbilical cord blood or cord tissue. One of the major changes in the manufacturing process has been the replacement of fetal bovine serum with human platelet lysate as medium supplement. 59% of centers used only third-party MSC, whilst only 1 center manufactured exclusively autologous MSC. The large majority of these facilities (71%) administered MSC exclusively from frozen batches. Aside from variations in the culture method, we found large heterogeneity also regarding product specification, particularly in the markers used for phenotypical characterization and their threshold of expression, use of potency assays to test MSC functionality, and karyotyping. The initial data collected from this survey highlight the variability in MSC manufacturing as clinical products and the need for harmonization. Until more informative potency assays become available, a more homogeneous approach to cell production may at least reduce variability in clinical trials and improve interpretation of results.
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- 2018
43. Donor-Derived CAR T Cells Engineered with Sleeping Beauty Achieve Anti-Leukemic Activity without Severe Toxicity
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Chiara F. Magnani, Silvia Ferrari, Giuliana Rizzuto, Sarah Tettamanti, Federico Lussana, Fabrizio Benedicenti, Valentina Colombo, Ettore Biagi, Giovanni Cazzaniga, Attilio Rovelli, Stefania Galimberti, Silvia Zaninelli, Maria Grazia Valsecchi, Giuseppe Gritti, Benedetta Cabiati, Martino Introna, Andrea Biondi, Sara Napolitano, Gian Maria Borleri, Chiara Buracchi, Eugenio Montini, Daniela Belotti, Alessandro Rambaldi, Giuseppe Dastoli, Giuseppe Gaipa, and Adriana Balduzzi
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medicine.medical_specialty ,education.field_of_study ,Cyclophosphamide ,business.industry ,medicine.medical_treatment ,T cell ,Immunology ,Population ,Cell Biology ,Hematology ,Hematopoietic stem cell transplantation ,medicine.disease ,Biochemistry ,Fludarabine ,Cytokine release syndrome ,medicine.anatomical_structure ,Internal medicine ,Clinical endpoint ,Medicine ,business ,education ,CD8 ,medicine.drug - Abstract
Background Significant efforts over the past few years led Chimeric Antigen Receptor (CAR) T cell therapy to success in relapsed and refractory (r/r) B-cell malignancies. Still logistical complexity, high costs and toxicities are currently the main barriers to the use of CAR T cell therapy. We therefore propose non-viral engineering of an allogeneic T cell population according to cytokine induced killer (CIK) cell protocol of differentiation. Methods We reported the updated results of our phase I/II trial in B-cell acute lymphoblastic leukemia (B-ALL) patients relapsed after allogeneic hematopoietic stem cell transplantation (HSCT) using donor-derived CD19 CAR T cells generated with the Sleeping Beauty (SB) transposon and differentiated into CIK (CARCIK-CD19) according to the method enclosed in the filed patent EP20140192371. After lymphodepletion with Fludarabine (30 mg/m2/day) x 4 days and Cyclophosphamide (500 mg/m2/day) x 2 days, CARCIK-CD19 were infused following a four-dose escalation scheme (1x106, 3x106, 7.5x106 and 15x106 transduced CAR+ T cells/kg) according to the Bayesian Optimal Interval Design (BOIN). During the cell manufacturing period, bridging anti leukemic therapy from patient registration to the beginning of the lymphodepletion, was allowed. The primary endpoint was to define the Maximum Tolerated Dose (MTD) and the safety assessment. Key secondary endpoints included the assessment of complete hematologic response (CR), defined as < 5% bone marrow (BM) blasts, circulating blasts < 1%, no clinical evidence of extramedullary disease, as well as the characterization of CARCIK-CD19 persistence in PB and BM (NCT03389035). Results The cellular product was produced successfully for all patients starting from the donor-derived peripheral blood (PB) and consisted mostly of CD3+ lymphocytes (mean 98.85% ±SD 1.19%) with a mean of 38.6% CAR expression (range 15.10%-73.17%). From January 2018 to July 2020, a total of 24 patients were screened, and 15 were enrolled (4 children and 11 adults) and infused with a single dose of CARCIK-CD19 (n=3 HLA identical sibling, n=4 MUD, n=8 haploidentical donor). The leukemic burden in the BM post lymphodepletion/pre-infusion ranged from 0% to 96%. Robust expansion was achieved in the majority of the patients. The maximal expansion reached about 1x106 transgene copies per μg DNA and 70% of CAR+ T cells in PB. CD8+ T cells represented the predominant circulating CAR+ T cell subset. Persistence of central memory CAR+ T cells was observed after infusion and CAR T cells were measurable up to 9 months. CARCIK-CD19 were characterized by a high profile of safety in all treated patients. Toxicities reported were two grade I and two grade II cytokine release syndrome (CRS) cases at the highest dose in the absence of graft-versus-host disease (GvHD), neurotoxicity, or dose-limiting toxicities. Seven out of 9 patients, receiving the highest doses, achieved CR and CRi at day 28. MRD-negative status for all responders was achieved by 6 out of 9 patients (1 currently in evaluation). The two patients in CR but with MRD+ relapsed with a CD19+ disease at +2.3 and +1.9 months post infusion, respectively. Among the 6 patients who achieved MRD-negative CR, two children underwent consolidation with a second allo-HSCT and are still alive and disease free (+17 and +13 months), two adult patients died of subsequent CD19+ disease relapse and two adult patients are still alive and disease free (+14 and +12 months) without additional therapies. The distribution profile of integration sites (IS) showed no preference for gene dense or promoter regions, and no particular differences between pre- and post- infusion sample IS. Samples harvested at early time points after infusion showed a highly polyclonal repertoire. At later time points (≥ 28 days after infusion) the repertoire of IS showed a marked reduction towards oligoclonality, in absence of specific dominant clones. Conclusions We can conclude that SB-engineered CAR T cells expand and persist in pediatric and adult B-ALL patients relapsed after HSCT. Sustained response was achieved without severe toxicities. All analyzed samples appear to have a highly polyclonal IS repertoire and no signs of genotoxicity by transposon insertions could be observed. Disclosures Gritti: IQVIA: Consultancy; Amgen: Honoraria; Autolus: Consultancy; Italfarmaco: Consultancy; F. Hoffmann-La Roche Ltd: Honoraria; Jannsen: Other: Travel Support; Takeda: Honoraria; Kite: Consultancy. Rambaldi:Sanofi: Honoraria, Other: TRAVEL, ACCOMMODATIONS, EXPENSES (paid by any for-profit health care company); Omeros: Honoraria, Other: TRAVEL, ACCOMMODATIONS, EXPENSES (paid by any for-profit health care company); Amgen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: TRAVEL, ACCOMMODATIONS, EXPENSES (paid by any for-profit health care company). Research grant from Amgen Inc.; Pfizer: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: TRAVEL, ACCOMMODATIONS, EXPENSES (paid by any for-profit health care company). Advisory board and speaker fees from Pfizer.; Novartis: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: TRAVEL, ACCOMMODATIONS, EXPENSES (paid by any for-profit health care company); Gilead: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Travel support from Gilead.; Roche: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: Support of parent study and funding of editorial support. Received travel support., Research Funding; University of Milan: Current Employment; BMS/Celgene: Honoraria, Other: TRAVEL, ACCOMMODATIONS, EXPENSES (paid by any for-profit health care company); Astellas: Honoraria, Other: TRAVEL, ACCOMMODATIONS, EXPENSES (paid by any for-profit health care company).
- Published
- 2020
44. Therapeutic potential of stromal cells of non-renal or renal origin in experimental chronic kidney disease
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Martino Introna, Ariela Benigni, Domenico Cerullo, Ornella Colpani, Daniela Rottoli, Giuseppe Remuzzi, Carlamaria Zoja, Daniela Corna, Ton J. Rabelink, Daniëlle G. Leuning, Marina Morigi, Chiara Capelli, and Cinzia Rota
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0301 basic medicine ,Male ,Medicine (miscellaneous) ,Mesenchymal stromal cell therapy ,Podocyte ,Umbilical Cord ,0302 clinical medicine ,Chronic kidney disease ,lcsh:QD415-436 ,Kidney ,lcsh:R5-920 ,Renal perivascular cells ,Glomerulosclerosis, Focal Segmental ,Podocytes ,Graft Survival ,Antigens, Nuclear ,medicine.anatomical_structure ,Molecular Medicine ,Stem cell ,lcsh:Medicine (General) ,Stromal cell ,Transplantation, Heterologous ,Bone Marrow Cells ,Mesenchymal Stem Cell Transplantation ,Biochemistry, Genetics and Molecular Biology (miscellaneous) ,Nephropathy ,lcsh:Biochemistry ,03 medical and health sciences ,Rats, Nude ,Renal repair ,medicine ,Animals ,Humans ,Regeneration ,Renal Insufficiency, Chronic ,Conditioned medium ,Cell Proliferation ,business.industry ,Macrophages ,Research ,Mesenchymal stem cell ,Epithelial Cells ,Mesenchymal Stem Cells ,Cell Biology ,medicine.disease ,Coculture Techniques ,Rats ,Disease Models, Animal ,030104 developmental biology ,Doxorubicin ,Culture Media, Conditioned ,Cancer research ,Bone marrow ,business ,030217 neurology & neurosurgery ,Biomarkers ,Kidney disease - Abstract
Background Mesenchymal stromal cell (MSC)-based therapy is a promising strategy for preventing the progression of chronic kidney disease (CKD), with the potential to induce tissue regeneration. In search of the best cellular source we compared, in the rat model of adriamycin (ADR) nephropathy, the regenerative potential of human stromal cells of non-renal origin, such as bone marrow (bm) MSCs and umbilical cord (uc) MSCs, with that of newly discovered stromal cells of renal origin, the kidney perivascular cells (kPSCs) known to exhibit tissue-specific properties. Methods The therapeutic effect of repeated infusions of human bmMSCs, ucMSCs, kPSCs (1.5 × 106 cells/rats) or conditioned medium from ucMSCs was studied in athymic rats with ADR-induced nephropathy (7.9 mg/kg). The ability of the three stromal cell populations to engraft the damaged kidney was evaluated by detecting the presence of human nuclear antigenpos cells. Glomerular podocyte loss and endothelial damage, sclerotic lesions and inflammation were assessed at 14 and 28 days. In-vitro experiments with a transwell system were performed to investigate the effects of different stromal cell populations on parietal epithelial cells (PECs) activated or not with albumin or angiotensin II for 24 h. Results Infusions of non-renal and renal stromal cells resulted in a comparable engraftment into the kidney, in the peritubular areas and around the glomerular structures. All three cell populations limited podocyte loss and glomerular endothelial cell injury, and attenuated the formation of podocyte and PEC bridges. This translated into a reduction of glomerulosclerosis and fibrosis. Human ucMSCs had an anti-inflammatory effect superior to that of the other stromal cells, reducing macrophage infiltration and inducing polarisation towards the M2 macrophage phenotype. Conditioned medium from ucMSCs shared the same renoprotective effects of the cells. Consistent with in-vivo data, bmMSCs and kPSCs, but even more so ucMSCs, limited proliferation, migratory potential and extracellular matrix production of activated PECs, when cultured in a transwell system. Conclusions Our data indicate that either non-renal or renal stromal cells induce renal tissue repair, highlighting ucMSCs and their conditioned medium as the most reliable clinical therapeutic tool for CKD patients. Electronic supplementary material The online version of this article (10.1186/s13287-018-0960-8) contains supplementary material, which is available to authorized users.
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- 2018
45. Donor-Derived CD19 CAR Cytokine Induced Killer (CIK) Cells Engineered with Sleeping Beauty Transposon for Relapsed B-Cell Acute Lymphoblastic Leukemia (B-ALL)
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Sarah Tettamanti, Silvia Rigamonti, Chiara Buracchi, Fabrizio Benedicenti, Adriana Balduzzi, Giovanni Cazzaniga, Alessandro Rambaldi, Benedetta Cabiati, Silvia Ferrari, Attilio Rovelli, Giuseppe Gritti, Andrea Biondi, Giada Matera, Giuseppe Dastoli, Silvia Zaninelli, Grazia Fazio, Martino Introna, Gian Maria Borleri, Giuseppe Gaipa, Chiara F. Magnani, Sara Napolitano, Eugenio Montini, Federico Lussana, Stefania Cesana, Valentina Colombo, and Daniela Belotti
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biology ,business.industry ,Genetic enhancement ,medicine.medical_treatment ,Immunology ,Cell Biology ,Hematology ,Immunotherapy ,medicine.disease ,Sleeping Beauty transposon system ,Biochemistry ,CD19 ,Transplantation ,Cytokine release syndrome ,Cytokine ,Acute lymphocytic leukemia ,biology.protein ,medicine ,Cancer research ,business - Abstract
Background Immunotherapy using patient-derived CAR T cells has achieved complete remission and durable response in highly refractory populations. However, logistical complexity and high costs of manufacturing autologous viral products limit CAR T cell availability. Allogeneic Cytokine Induced Killer (CIK) cells, a T-cell population characterized by the enrichment of CD3+CD56+ cells, have demonstrated a high profile of safety in acute lymphoblastic leukemia (ALL) patients (Introna M et al. Biol Blood Marrow Transplant. 2017). CIK cells could be easily engineered by the non-viral Sleeping Beauty (SB) transposon for the clinical application (Magnani CF et al, Hum Gene Ther. 2018, Biondi A et al. J Autoimmun. 2017). Methods CIK cells were generated from 50 ml of donor-derived peripheral blood (PB) by electroporation with the GMP-grade CD19.CAR/pTMNDU3 and pCMV-SB11 plasmids according to the method enclosed in the filed patent EP20140192371. After lymphodepletion with Fludarabine (30 mg/m2/day) x 4 days and Cyclophosphamide (300 mg/m2/day) x 2 days, CARCIK-CD19 were infused in pediatric and adult B-cell ALL (B-ALL) patients relapsed after allogeneic hematopoietic stem cell transplantation (HSCT). The clinical trial follows a four-dose escalation scheme (1x106, 3x106, 7.5x106 and 15x106 transduced CAR+ T cells/kg) using the novel Bayesian Optimal Interval Design (BOIN). During the cell manufacturing period, bridging anti leukemic therapy from patient registration to the beginning of the lymphodepletion, was allowed. The primary endpoint was to define the Maximum Tolerated Dose (MTD) and a safety assessment. Key secondary endpoints included the assessment of complete hematologic response (CR), defined as < 5% bone marrow (BM) blasts, circulating blasts < 1%, no clinical evidence of extramedullary disease, as well as the characterization of CARCIK-CD19 persistence in PB and BM (NCT03389035). Results We manufactured eighteen batches by seeding a median of 126.8x106 allogeneicPBMCs. At the end of expansion, the mean harvesting was 6.46x109 nucleated cells (range 1.39 - 16.00x109). Manufactured cells were mostly CD3+ lymphocytes (mean 98.90% ±SE 0.30%). Of these, 43.57%±3.73% were CAR+, 47.07%±2.74% were CD56+, 80.44%±2.53% were CD8+. The quality requirements for batch release were met in 17 productions. As of the data cut-off date (July 19, 2019), 4 pediatric and 7 adult patients were infused with a single dose of CARCIK-CD19 (n=2 HLA identical sibling, n=4 MUD, n=5 haploidentical donor). The leukemic burden in the BM post lymphodepletion/pre-infusion ranged from 0% to 96%. CARCIK-CD19 were characterized by a high profile of safety in all treated patients. Toxicities reported were a grade I cytokine release syndrome and an infusion-related DMSO-associated seizure, with absence of dose-limiting toxicities, neurotoxicity and graft-versus-host disease (GvHD) in any of the treated patients. Four out of 5 patients, receiving the highest doses, achieved CR and CRi at day 28. The 3 patients in CR were also MRD- (by flow cytometry and RT-PCR) while the CRi was MRD+ and relapsed at day+49. Robust expansion was achieved in the majority of the patients as defined by detectable CAR T-cell detection (vector copy number VCN, range 4645-977992 transgene copies/ug) and flow (range 0.5-30%) in PB. The median time to peak engraftment was 14 days. The magnitude of expansion was correlated with the CD19+ burden in the BM at the time of the infusion (P value = 0.0006, R square 0.7469). CD8+ T cells represented the predominant CARCIK-CD19 T-cell subset (78.88%±5.33% d14 n=6) along with CD3+CD56+ CIK cells and CD4+ T cells to a lesser extent. The majority of CAR T cells had a central and effector memory phenotype. CAR T cells were measurable by VCN up to 6 months with a mean persistence of 70.5 ± 14.85 days (follow up ranging from 28 days to 1 year). No major difference was observed by integration analyses of the patients' PB and the cell products. The vector integration sites reflected the classical random distribution of SB without any tendency for gene dense regions. Conclusions Our ongoing phase I/II trial demonstrates that SB-engineered CARCIK-CD19 cells are able to expand and persist in pediatric and adult B-ALL patients relapsed after HSCT, with important implications for a non-viral technology. These encouraging results prompted us to expand our study. Disclosures Gritti: Autolus Ltd: Honoraria; Roche: Other: Not stated; Abbvie: Other: Not stated; Becton Dickinson: Other: Not stated. Rambaldi:Celgene: Membership on an entity's Board of Directors or advisory committees, Other: travel support, Speakers Bureau; Roche: Membership on an entity's Board of Directors or advisory committees, Other: travel support, Research Funding, Speakers Bureau; Jazz: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau, travel support; Pfizer: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Gilead: Membership on an entity's Board of Directors or advisory committees, Other: travel support, Speakers Bureau; Amgen: Membership on an entity's Board of Directors or advisory committees, Other: travel support, Research Funding, Speakers Bureau; Novartis: Membership on an entity's Board of Directors or advisory committees, Other: travel support, Speakers Bureau; Italfarmaco: Membership on an entity's Board of Directors or advisory committees, Other: travel support, Research Funding, Speakers Bureau; Omeros: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau.
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- 2019
46. Human neutrophils express low levels of FcγRIIIA, which plays a role in PMN activation
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Gerta Musaraj, Josée Golay, Martino Introna, Orietta Spinelli, Rut Valgardsdottir, and Damiano Giupponi
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Trogocytosis ,medicine.drug_class ,Neutrophils ,Phagocytosis ,Immunology ,Plenary Paper ,Gene Expression ,Monoclonal antibody ,Biochemistry ,Flow cytometry ,Western blot ,medicine ,Humans ,RNA, Messenger ,Receptor ,Opsonin ,Cells, Cultured ,medicine.diagnostic_test ,biology ,Chemistry ,Receptors, IgG ,hemic and immune systems ,Cell Biology ,Hematology ,Molecular biology ,Leukemia, Lymphocytic, Chronic, B-Cell ,Tissue Donors ,Immunoglobulin G ,biology.protein ,Antibody ,Gene Deletion - Abstract
We have identified a rare healthy FcγRIIIB (CD16B)-null donor completely lacking FCGR3B RNA and protein expression and dissected the role of the different neutrophil Fcγ receptors in the response to therapeutic anti-CD20 monoclonal antibodies. We observed that polymorphonuclear neutrophils (PMNs) from FcγRIIIB wild-type (WT) individuals or the null donor were more effectively activated by chronic lymphocytic leukemia (CLL) B-cell targets opsonized with glycoengineered anti-CD20 antibodies compared with fully core-fucosylated anti-CD20 antibodies, suggesting the presence and role of FcγRIIIA (CD16A) on PMNs. Indeed, we demonstrated by reverse-transcription polymerase chain reaction, flow cytometry, and western blot analysis that PMNs from FcγRIIIB WT donors and the null individual express low levels of FcγRIIIA on their surfaces. FcγRIIIA is a functional and activating molecule on these cells, because anti-CD16 F(ab′)2 antibodies alone were able to activate highly purified PMNs from the FcγRIIIB-null donor. Use of blocking anti-CD16 and anti-CD32 antibodies showed that FcγRIIIA is also a major mediator of phagocytosis of CD20-opsonized beads by FcγRIIIB WT and null PMNs. In contrast, trogocytosis of antibody-opsonized CLL B cells by PMNs was mediated primarily by FcγRIIIB in WT PMNs and by FcγRIIA in null PMNs. We conclude that FcγRIIIA is an important player in PMN functions, whereas FcγRIIIB is dispensable for activation and phagocytosis. We discuss the clinical implications of these findings.
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- 2018
47. Cord blood-derived cytokine-induced killer cells combined with blinatumomab as a therapeutic strategy for CD19
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Benedetta Mazzanti, Riccardo Saccardi, Sabrina Cribioli, Josée Golay, Martino Introna, Elisa Gotti, Simona Martinelli, Rachele Alzani, Clara Albanese, Alessandro Rambaldi, and Bruna Pasini
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0301 basic medicine ,Cytotoxicity, Immunologic ,Male ,Cancer Research ,Immunology ,Antigens, CD19 ,Mice, Transgenic ,Mice, SCID ,CD8-Positive T-Lymphocytes ,Immunotherapy, Adoptive ,CD19 ,03 medical and health sciences ,Mice ,0302 clinical medicine ,Antineoplastic Agents, Immunological ,Cytokine-Induced Killer Cells ,Mice, Inbred NOD ,Neoplasms ,Antibodies, Bispecific ,medicine ,Immunology and Allergy ,Cytotoxic T cell ,Animals ,Humans ,Genetics (clinical) ,Transplantation ,Cytokine-induced killer cell ,biology ,Chemistry ,Infant, Newborn ,CD28 ,Cell Biology ,medicine.disease ,Fetal Blood ,Combined Modality Therapy ,Killer Cells, Natural ,Leukemia ,030104 developmental biology ,Treatment Outcome ,Oncology ,Cord blood ,Cancer research ,biology.protein ,Blinatumomab ,Female ,K562 Cells ,CD8 ,030215 immunology ,medicine.drug - Abstract
Background Cytokine-induced killer cells (CIKs) are an advanced therapeutic medicinal product (ATMP) that has shown therapeutic activity in clinical trials but needs optimization. We developed a novel strategy using CIKs from banked cryopreserved cord blood units (CBUs) combined with bispecific antibody (BsAb) blinatumomab to treat CD19+ malignancies. Methods CB-CIKs were expanded in vitro and fully characterized in comparison with peripheral blood (PB)–derived CIKs. Results CB-CIKs, like PB-CIKs, were mostly CD3+ T cells with mean 45% CD3+CD56+ and expressing mostly TCR(T cell receptor)αβ with a TH1 phenotype. CB-CIK cultures had, however, a larger proportion of CD4+ cells, mostly CD56−, as well as a greater proportion of naive CCR7+CD45RA+ and a lower percentage of effector memory cells, compared with PB-CIKs. CB-CIKs were very similar to PB-CIKs in their expression of a large panel of co-stimulatory and inhibitory/exhaustion markers, except for higher CD28 expression among CD8+ cells. Like PB-CIKs, CB-CIKs were highly cytotoxic in vitro against natural killer (NK) cell targets and efficiently lysed CD19+ tumor cells in the presence of blinatumomab, with 30–60% lysis of target cells at very low effector:target ratios. Finally, both CB-CIKs and PB-CIKs, combined with blinatumomab, showed significant therapeutic activity in an aggressive PDX Ph+ CD19+ acute lymphoblastic leukemia model in NOD-SCID mice, without sign of toxicity or graft-versus-host disease. The improved expansion protocol was finally validated in good manufacturing practice conditions, showing reproducible expansion of CIKs from cryopreserved cord blood units with a median of 28.8 × 106 CIK/kg. Discussion We conclude that CB-CIKs, combined with bispecific T-cell–engaging antibodies, offer a novel, effective treatment strategy for leukemia.
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- 2017
48. Longitudinal tracking of triple labeled umbilical cord derived mesenchymal stromal cells in a mouse model of Amyotrophic Lateral Sclerosis
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Massimo Tortarolo, Laura Talamini, Davide Moscatelli, Chiara Santangelo, Roberta Frapolli, Sara Previdi, Raffaele Ferrari, Chiara Capelli, Martino Introna, Paolo Bigini, Caterina Bendotti, Leopoldo Sitia, Mario Salmona, and Martina Bruna Violatto
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Biodistribution ,Pathology ,medicine.medical_specialty ,Mesenchymal stromal cells ,Mice, Transgenic ,Biology ,Mesenchymal Stem Cell Transplantation ,Umbilical cord ,Cell therapy ,Lateral ventricles ,Parenchyma ,medicine ,Amyotrophic Lateral Sclerosis ,Cell tracking ,In vivo imaging ,Nanotechnology ,Cell Biology ,Developmental Biology ,Medicine (all) ,Animals ,Humans ,Tissue Distribution ,lcsh:QH301-705.5 ,Cell Size ,Injections, Intraventricular ,Medicine(all) ,Staining and Labeling ,Mesenchymal stem cell ,Mesenchymal Stem Cells ,General Medicine ,Anatomy ,Spinal cord ,Mice, Inbred C57BL ,Disease Models, Animal ,medicine.anatomical_structure ,lcsh:Biology (General) ,Organ Specificity ,Injections, Intravenous ,Choroid plexus - Abstract
The translational potential of cell therapy to humans requires a deep knowledge of the interaction between transplanted cells and host tissues. In this study, we evaluate the behavior of umbilical cord mesenchymal stromal cells (UC-MSCs), labeled with fluorescent nanoparticles, transplanted in healthy or early symptomatic transgenic SOD1G93A mice (a murine model of Amyotrophic Lateral Sclerosis). The double labeling of cells with nanoparticles and Hoechst-33258 enabled their tracking for a long time in both cells and tissues. Whole-body distribution of UC-MSCs was performed by in-vivo and ex-vivo analyses 1, 7, 21days after single intravenous or intracerebroventricular administration. By intravenous administration cells were sequestered by the lungs and rapidly cleared by the liver. No difference in biodistribution was found among the two groups. On the other hand, UC-MSCs transplanted in lateral ventricles remained on the choroid plexus for the whole duration of the study even if decreasing in number. Few cells were found in the spinal cord of SOD1G93A mice exclusively. No migration in brain parenchyma was observed. These results suggest that the direct implantation in brain ventricles allows a prolonged permanence of cells close to the damaged areas and makes this method of tracking reliable for future studies of efficacy.
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- 2015
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49. Direct Reprogramming of Human Bone Marrow Stromal Cells into Functional Renal Cells Using Cell-free Extracts
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Marina Morigi, Paola Rizzo, Marta Todeschini, Maria Grazia De Simoni, Paraskevas Iatropoulos, Martino Introna, Christodoulos Xinaris, Susanna Tomasoni, Evangelia Papadimou, Valentina Benedetti, Ariela Benigni, Michael S. Goligorsky, Cinzia Rota, Lorena Longaretti, and Giuseppe Remuzzi
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Stromal cell ,Cellular differentiation ,Cell ,Mice, SCID ,In Vitro Techniques ,Biology ,Kidney ,Mesenchymal Stem Cell Transplantation ,Biochemistry ,Article ,Kidney Tubules, Proximal ,Cell therapy ,Mice ,03 medical and health sciences ,0302 clinical medicine ,Mice, Inbred NOD ,Genetics ,medicine ,Animals ,Humans ,lcsh:QH301-705.5 ,Cell Line, Transformed ,030304 developmental biology ,0303 health sciences ,lcsh:R5-920 ,Cell-Free System ,Gene Expression Profiling ,Mesenchymal stem cell ,Cell Differentiation ,Mesenchymal Stem Cells ,Cell Biology ,Cellular Reprogramming ,3. Good health ,Cell biology ,medicine.anatomical_structure ,lcsh:Biology (General) ,Cell culture ,Immunology ,Female ,Transcriptome ,lcsh:Medicine (General) ,Reprogramming ,030217 neurology & neurosurgery ,Developmental Biology - Abstract
Summary The application of cell-based therapies in regenerative medicine is gaining recognition. Here, we show that human bone marrow stromal cells (BMSCs), also known as bone-marrow-derived mesenchymal cells, can be reprogrammed into renal proximal tubular-like epithelial cells using cell-free extracts. Streptolysin-O-permeabilized BMSCs exposed to HK2-cell extracts underwent morphological changes—formation of “domes” and tubule-like structures—and acquired epithelial functional properties such as transepithelial-resistance, albumin-binding, and uptake and specific markers E-cadherin and aquaporin-1. Transmission electron microscopy revealed the presence of brush border microvilli and tight intercellular contacts. RNA sequencing showed tubular epithelial transcript abundance and revealed the upregulation of components of the EGFR pathway. Reprogrammed BMSCs integrated into self-forming kidney tissue and formed tubular structures. Reprogrammed BMSCs infused in immunodeficient mice with cisplatin-induced acute kidney injury engrafted into proximal tubuli, reduced renal injury and improved function. Thus, reprogrammed BMSCs are a promising cell resource for future cell therapy., Highlights • BMSCs cross lineage boundaries toward renal cells via cell-extract reprogramming • Reprogrammed BMSCs acquire proximal tubular-like epithelial cell properties • Reprogrammed BMSCs integrate into proximal tubuli and protect mice from AKI, In this article, Remuzzi, Papadimou, and colleagues show that human BMSCs can be directly reprogrammed into renal proximal tubular-like epithelial cells using epithelial cell extracts. BMSC reprogramming is demonstrated by the acquisition of epithelial features such as morphologic, ultrastructural, antigenic, genetic makeup, and functional properties (organ-forming and renal damage repair capabilities).
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- 2015
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50. Disposal of the residual autologous HSC units: Results of a survey carried out two years after the publication of a national policy in Italy
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Carlo Petrini, Paolo Perseghin, Francesca Bonifazi, Patrizia Accorsi, Martino Introna, Letizia Lombardini, and Daniele Laszlo
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Male ,Economic growth ,medicine.medical_treatment ,Health Policy ,Hematopoietic Stem Cell Transplantation ,Hematology ,Hematopoietic stem cell transplantation ,030204 cardiovascular system & hematology ,Residual ,03 medical and health sciences ,0302 clinical medicine ,Italy ,Political science ,Surveys and Questionnaires ,medicine ,National Policy ,Humans ,Female ,Medical Waste Disposal ,Autografts ,Health policy ,030215 immunology - Published
- 2017
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