Abstract
BackgroundExtracellular vesicles (EVs) produced by human bone marrow-derived mesenchymal stromal cells (hBM-MSCs) are currently investigated for their clinical effectiveness towards immune-mediated diseases. The large amounts of stem cell-derived EVs required for clinical testing suggest that bioreactor production systems may be a more amenable alternative than conventional EV production methods for manufacturing products for therapeutic use in humans.MethodsTo characterize the potential utility of these systems, EVs from four hBM-MSC donors were produced independently using a hollow-fiber bioreactor system under a cGMP-compliant procedure. EVs were harvested and characterized for size, concentration, immunophenotype, and glycan profile at three separate intervals throughout a 25-day period.ResultsBioreactor-inoculated hBM-MSCs maintained high viability and retained their trilineage mesoderm differentiation capability while still expressing MSC-associated markers upon retrieval. EVs collected from the four hBM-MSC donors showed consistency in size and concentration in addition to presenting a consistent surface glycan profile. EV surface immunophenotypic analyses revealed a consistent low immunogenicity profile in addition to the presence of immuno-regulatory CD40 antigen. EV cargo analysis for biomarkers of immune regulation showed a high abundance of immuno-regulatory and angiogenic factors VEGF-A and IL-8.ConclusionsSignificantly, EVs from hBM-MSCs with immuno-regulatory constituents were generated in a large-scale system over a long production period and could be frequently harvested with the same quality and quantity, which will circumvent the challenge for clinical application.
Highlights
Intrinsic properties of extracellular vesicles (EVs) have rendered them attractive biocompatible therapeutic nanovesicles under their native form or engineered as gene/vaccine/drug delivery systems [1, 2]
Significantly, Extracellular vesicle (EV) from Human bone marrow-derived mesenchymal stromal/stem cells (hBM-Mesenchymal stromal/stem cell (MSC)) with immuno-regulatory constituents were generated in a large-scale system over a long production period and could be frequently harvested with the same quality and quantity, which will circumvent the challenge for clinical application
EVs with immuno-regulatory constituents were generated in a large-scale system over a long production period and could be frequently harvested with the same quality and quantity, which will circumvent the challenge for clinical application
Summary
Intrinsic properties of extracellular vesicles (EVs) have rendered them attractive biocompatible therapeutic nanovesicles under their native form or engineered as gene/vaccine/drug delivery systems [1, 2]. Through the transfer of their bioactive payload (cytokines, growth factors, signaling lipids, mRNAs, and regulatory miRNAs) and/or through the binding via their membrane-bound molecules (receptors, lipids, integrins, and glycans), EVs can regulate cell/tissue responses, and more broadly, have immune-modulating effects [6,7,8,9,10] Their native membrane constituents and intrinsic abilities to be transferred from one cell to another may play a role in their enhanced bioavailability and lower immunogenicity and prospects as a new biological nanoplatforms for drug delivery or diagnostic purpose [11, 12]. The hollow-fiber bioreactor from FiberCell Systems allows for seeding large amounts of adherent cells based on its hollow-fiber technology which increases the cell seeding surface area (medium-sized cartridge offers 4000 cm of surface area) [18, 19] This configuration allows for a scale-up of continuous EV production sampled over time from the EV-rich cell-conditioned medium produced by millions of cells, allowing anticipated clinical doses to be achieved. The large amounts of stem cell-derived EVs required for clinical testing suggest that bioreactor production systems may be a more amenable alternative than conventional EV production methods for manufacturing products for therapeutic use in humans
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