Development of a human umbilical cord-derived mesenchymal stromal cell-based advanced therapy medicinal product to treat immune and/or inflammatory diseases
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ABSTRACT:
SUBMITTER: Mebarki M
PROVIDER: S-EPMC8590372 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:IntroductionStandardization of mesenchymal stromal cells (MSCs) manufacturing is urgently needed to enable translational activities and ultimately facilitate comparison of clinical trial results. In this work we describe the adaptation of a proprietary method for isolation of a specific umbilical cord tissue-derived population of MSCs, herein designated by its registered trademark as UCX®, towards the production of an advanced therapy medicinal product (ATMP).MethodsThe adaptation focused on different stages of production, from cell isolation steps to cell culturing and cryopreservation. The origin and quality of materials and reagents were considered and steps for avoiding microbiological and endotoxin contamination of the final cell product were implemented. Cell isolation efficiency, MSCs surface markers and genetic profiles, originating from the use of different medium supplements, were compared. The ATMP-compliant UCX® product was also cryopreserved avoiding the use of dimethyl sulfoxide, an added benefit for the use of these cells as an ATMP. Cells were analyzed for expansion capacity and longevity. The final cell product was further characterized by flow cytometry, differentiation potential, and tested for contaminants at various passages. Finally, genetic stability and immune properties were also analyzed.ResultsThe isolation efficiency of UCX® was not affected by the introduction of clinical grade enzymes. Furthermore, isolation efficiencies and phenotype analyses revealed advantages in the use of human serum in cell culture as opposed to human platelet lysate. Initial decontamination of the tissue followed by the use of mycoplasma- and endotoxin-free materials and reagents in cell isolation and subsequent culture, enabled the removal of antibiotics during cell expansion. UCX®-ATMP maintained a significant expansion potential of 2.5 population doublings per week up to passage 15 (P15). They were also efficiently cryopreserved in a DMSO-free cryoprotectant medium with approximately 100% recovery and 98% viability post-thaw. Additionally, UCX®-ATMP were genetically stable upon expansion (up to P15) and maintained their immunomodulatory properties.ConclusionsWe have successfully adapted a method to consistently isolate, expand and cryopreserve a well-characterized population of human umbilical cord tissue-derived MSCs (UCX®), in order to obtain a cell product that is compliant with cell therapy. Here, we present quality and safety data that support the use of the UCX® as an ATMP, according to existing international guidelines.
| S-EPMC4055140 | biostudies-literature
Project description:Umbilical cord-derived mesenchymal stem/stromal cells (UC-MSCs) emerge as a perspective for therapeutic use in immune and inflammatory diseases. Indeed, immunomodulatory and anti-inflammatory properties, associated to fewer ethical, availability, and safety issues, position UC-MSCs as a promising active substance to develop medicinal products. Since 2007, UC-MSC-based products are classified as advanced therapy medicinal products (ATMP) according to the European Regulation 1394/2007/EC. This new regulatory status required a total adaptation of stakeholders wishing to develop UC-MSC-based ATMPs. Cell production in tissue and cell banks has been replaced by the manufacturing of a medicine, in authorized establishments, according to the good manufacturing practices (GMP) specific to ATMPs. After a brief description of UC-MSCs, we described in this review their recent use in a large panel of immune and inflammatory pathologies, including early and late phase clinical trials. Despite the use of the same product, we noticed an important heterogeneity in terms of indication, posology and study design. Then, we discussed regulatory and manufacturing challenges for stakeholders, especially in terms of process harmonization and cells characterization. Our aim was to point that despite MSCs use for several decades, the development of an UC-MSC-based ATMP remains at this day a real challenge for both academic institutions and pharmaceutical companies.
| S-EPMC7907784 | biostudies-literature
Project description:Mesenchymal stromal cells (MSCs) are multipotent progenitors that can be isolated from different sources, such as the bone marrow, adipose tissue and umbilical cord. The therapeutic potential of MSCs is related to a plethora of immunomodulatory, anti-inflammatory and pro-repair actions, which are at least partially dependent on their secretome. Among the components of MSCs' secretome, EVs have received considerable attention because MSC-EVs exert similar therapeutic properties as their parent cells. Among the different MSC sources for EV production, human umbilical cord MSCs (hUCMSCs) show advantages such as tissue availability, high proliferative profile of the cells and potential beneficial therapeutic effects in a variety of different diseases, such as stroke This study aimed to provide novel insights for future hUCMSC-EVs research and treatment selection. We investigated the influence of the culture and harvesting conditions on the EV proteomic profile, productivity, surface markers expression and evaluated their in vivo biodistribution and toxicity.
2024-05-04 | PXD038850 | Pride
Project description:Umbilical cord mesenchymal stromal cells (UCMSCs) have shown an ability to modulate the immune system through the secretion of paracrine mediators, such as extracellular vesicles (EVs). However, the culture conditions that UCMSCs are grown in can alter their secretome and thereby affect their immunomodulatory potential. UCMSCs are commonly cultured at 21% O2 in vitro, but recent research is exploring their growth at lower oxygen conditions to emulate circulating oxygen levels in vivo. Additionally, a pro-inflammatory culture environment is known to enhance UCMSC anti-inflammatory potential. Therefore, this paper examined EVs from UCMSCs grown in normal oxygen (21% O2), low oxygen (5% O2) and pro-inflammatory conditions to see the impact of culture conditions on the EV profile. EVs were isolated from UCMSC conditioned media and characterised based on size, morphology and surface marker expression. EV protein cargo was analysed using a proximity-based extension assay. Results showed that EVs had a similar size and morphology. Differences were found in EV protein cargo, with pro-inflammatory primed EVs showing an increase in proteins associated with chemotaxis and angiogenesis. This showed that the UCMSC culture environment could alter the EV protein profile and might have downstream implications for their functions in immunomodulation.
| S-EPMC7140705 | biostudies-literature
Project description:Mesenchymal stromal cells (MSCs) administered intravenously (IV) have shown efficacy in preclinical models of various diseases. This is despite the cells not reaching the site of injury due to entrapment in the lungs. The immunomodulatory properties of MSCs are thought to underlie their therapeutic effects, irrespective of whether they are sourced from bone marrow, adipose tissue, or umbilical cord. To better understand how MSCs affect innate immune cell populations in the lung, we evaluated the distribution and phenotype of neutrophils, monocytes, and macrophages by flow cytometry and histological analyses after delivering human umbilical cord-derived MSCs (hUC-MSCs) IV into immunocompetent mice. After 2 hr, we observed a significant increase in neutrophils, and proinflammatory monocytes and macrophages. Moreover, these immune cells localized in close proximity to the MSCs, suggesting an active role in their clearance. By 24 hr, we detected an increase in anti-inflammatory monocytes and macrophages. These results suggest that the IV injection of hUC-MSCs leads to an initial inflammatory phase in the lung shortly after injection, followed by a resolution phase 24 hr later.
| S-EPMC10497368 | biostudies-literature
Project description:Autoimmune conditions, such as rheumatoid arthritis, are characterised by a loss of immune tolerance, whereby the immune cells attack self-antigens causing pain and inflammation. These conditions can be brought into remission using pharmaceutical treatments, but often have adverse side effects and some patients do not respond favourably to them. Human umbilical cord mesenchymal stromal cells (UCMSCs) present a promising alternative therapeutic due to their innate anti-inflammatory properties which can be strengthened using pro-inflammatory conditions. Their therapeutic mechanism of action has been attributed to paracrine signalling, by which nanosized acellular particles called 'extracellular vesicles' (EVs) are one of the essential components. Therefore, this research analysed the anti-inflammatory properties of UCMSC-EVs 'primed' with pro-inflammatory cytokines and at baseline with no inflammatory cytokines (control). Both control and primed EVs were co-cultured with un-pooled peripheral blood mononuclear cells (PBMCs; n = 6) from healthy donors. Neither control nor primed EVs exerted a pro-inflammatory effect on PBMCs. Instead, the primed EVs showed the immunosuppressive potential by increasing the expression of the anti-inflammatory protein FoxP3 in PBMCs. This may be attributed to the upregulated miRNAs identified in primed EVs in comparison to control EVs (miR-139-5p, miR-140-5p, miR-214-5p). These findings aid in understanding how UCMSC-EVs mediate immunosuppression and support their potential use in treating autoimmune conditions.
| S-EPMC10579155 | biostudies-literature
Project description:Many cell types, including cancer cells, release tissue factor (TF)-exposing extracellular vesicles (EVs). It is unknown whether MSC-EVs pose a thromboembolism risk due to TF expression. Knowing that MSCs express TF and are procoagulant, we hypothesize that MSC-EVs also might. Here, we examined the expression of TF and the procoagulant activity of MSC-EVs and the impact of EV isolation methods and cell culture expansion on EV yield, characterization, and potential risk using a design of experiments methodology. MSC-EVs were found to express TF and have procoagulant activity. Thus, when MSC-derived EVs are employed as a therapeutic agent, one might consider TF, procoagulant activity, and thromboembolism risk and take steps to prevent them.
| S-EPMC10252357 | biostudies-literature
Project description:MSCs derived from the umbilical cord tissue, termed UCX, were investigated for their immunomodulatory properties and compared to bone marrow-derived MSCs (BM-MSCs), the gold-standard in immunotherapy. Immunogenicity and immunosuppression were assessed by mixed lymphocyte reactions, suppression of lymphocyte proliferation and induction of regulatory T cells. Results showed that UCX were less immunogenic and showed higher immunosuppression activity than BM-MSCs. Further, UCX did not need prior activation or priming to exert their immunomodulatory effects. This was further corroborated in vivo in a model of acute inflammation. To elucidate the potency differences observed between UCX and BM-MSCs, gene expression related to immune modulation was analysed in both cell types. Several gene expression profile differences were found between UCX and BM-MSCs, namely decreased expression of HLA-DRA, HO-1, IGFBP1, 4 and 6, ILR1, IL6R and PTGES and increased expression of CD200, CD273, CD274, IL1B, IL-8, LIF and TGFB2. The latter were confirmed at the protein expression level. Overall, these results show that UCX seem to be naturally more potent immunosuppressors and less immunogenic than BM-MSCs. We propose that these differences may be due to increased levels of immunomodulatory surface proteins such as CD200, CD273, CD274 and cytokines such as IL1β, IL-8, LIF and TGFβ2.
| S-EPMC4443932 | biostudies-literature
Project description:BackgroundThe spleen plays an important role in erythrocyte turnover, adaptive immunity, antibody production, and the mobilization of monocytes/macrophages (Mφ) following tissue injury. In response to trauma, the spleen initiates production of inflammatory cytokines, which in turn recruit immune cells to the inflamed tissue, exacerbating damage. Our previous work has shown that intravenous mesenchymal stromal cell (MSC) infusion has potent immunomodulatory effects following spinal cord injury (SCI), associated with the transplanted cells homing to and persisting within the spleen. Therefore, this work aimed to characterize the relationship between the splenic inflammatory response and SCI pathophysiology, emphasizing splenic involvement in MSC-mediated effects.MethodsUsing a rodent model of cervical clip-compression SCI, secondary tissue damage and functional recovery were compared between splenectomised rodents and those with a sham procedure. Subsequently, 2.5 million MSCs from the term human umbilical cord matrix cells (HUCMCs) were infused via tail vein at 1-h post-SCI and the effects were assessed in the presence or absence of a spleen.ResultsSplenectomy alone had no effect on lesion volume, hemorrhage, or inflammation. There was also no significant difference between the groups in functional recovery and those in lesion morphometry. Yet, while the infusion of HUCMCs reduced spinal cord hemorrhage and increased systemic levels of IL-10 in the presence of a spleen, these effects were lost with splenectomy. Further, HUCMC infusion was shown to alter the expression levels of splenic cytokines, suggesting that the spleen is an important target and site of MSC effects.ConclusionsOur results provide a link between MSC function and splenic inflammation, a finding that can help tailor the cells/transplantation approach to enhance therapeutic efficacy.
| S-EPMC6091078 | biostudies-literature
Project description:BACKGROUND:Mesenchymal stromal cells (MSCs) are used in over 800 clinical trials mainly due to their immune inhibitory activity. Umbilical cord (UC), the second leading source of clinically used MSCs, is usually cut in small tissue pieces. Subsequent cultivation leads to a continuous outgrowth of MSC explant monolayers (MSC-EMs) for months. Currently, the first MSC-EM culture takes approximately 2 weeks to grow out, which is then expanded and applied to patients. The initiating tissue pieces are then discarded. However, when UC pieces are transferred to new culture dishes, MSC-EMs continue to grow out. In case the functional integrity of these cells is maintained, later induced cultures could also be expanded and used for cell therapy. This would drastically increase the number of available cells for each patient. To test the functionality of MSC-EMs from early and late induction time points, we compared the first cultures to those initiated after 2 months by investigating their clonality and immunomodulatory capacity. METHODS:We analyzed the clonal composition of MSC-EM cultures by umbilical cord piece transduction using integrating lentiviral vectors harboring genetic barcodes assessed by high-throughput sequencing. We investigated the transcriptome of these cultures by microarrays. Finally, the secretome was analyzed by multiplexed ELISAs, in vitro assays, and in vivo in mice. RESULTS:DNA barcode analysis showed polyclonal MSC-EMs even after months of induction cycles. A transcriptome and secretome analyses of early and late MSC cultures showed only minor changes over time. However, upon activation with TNF-α and IFN-γ, cells from both induction time points produced a multitude of immunomodulatory cytokines. Interestingly, the later induced MSC-EMs produced higher amounts of cytokines. To test whether the different cytokine levels were in a therapeutically relevant range, we used conditioned medium (CM) in an in vitro MLR and an in vivo killing assay. CM from late induced MSC-EMs was at least as immune inhibitory as CM from early induced MSC-EMs. CONCLUSION:Human umbilical cord maintains a microenvironment for the long-term induction of polyclonal and immune inhibitory active MSCs for months. Thus, our results would offer the possibility to drastically increase the number of therapeutically applicable MSCs for a substantial amount of patients.
| S-EPMC6755709 | biostudies-literature