(T1330-10-64) Advancing Neuro-Regenerative Therapies: Scalable Manufacturing and Purification of Extracellular Vesicles for Treating Neurodegenerative Diseases

Purpose: Extracellular vesicles (EVs) encompass a range of nano to micro sized particles that originate from endosomes and are released through the exocytosis pathway. Interest in EVs has grown because of their significant roles in cell-to-cell communication, facilitating the transport of essential biological materials such as miRNA, lncRNA, mRNA, proteins, and DNA. EVs have gained attention as natural nanoparticles that facilitate the transfer of coding and noncoding RNAs, as well as proteins between cells and organs. It is these functions which make exosomes ideal for potential therapy in nerve injury, multiple sclerosis, Alzheimer's disease, and Parkinson's disease. Schwann cells (SCs) function as the myelinating glial cells of the peripheral nervous system (PNS) and retain the capacity to alternate between radically different phenotypes in response to cell-extrinsic signals changing their support role through these phenotypes. Even though EVs are being investigated for their therapeutic potential, SC EVs have been overlooked and have a potential therapeutic value in injury and diseases of the PNS. However, there are still significant hurdles to overcome in manufacturing and purifying EVs for therapeutic purposes.

Methods: Using hTERT-immortalized Schwann cells obtained from ATCC, that have tested negative for viruses and mycoplasma, we established a process for manufacturing and purifying EVs using a bioreactor and assessing the therapeutic value of SC EVs from this process. We began with the development of methods to grow SCs in a xeno-free proprietary growth medium (SchwaMed®) at large. Exosomes were isolated using ultracentrifugation following pre-established protocols in EriVan's lab. Our approach to assessing the therapeutic potential of SC EVs from this process was to examine neurite outgrowth in dissociated dorsal root ganglia (DRG).

Results: Our studies evaluated the ability of SC hTERT cell line to grow in culture in adherent format in DMEM supplemented with 10% fetal bovine serum, mixing at 37 °C and 5% CO₂ (Figure 1. A-B). SCs were cultured as suspension in a stirred tank bioreactor in sphere format (Schwanosphere®) for the first time in a proprietary growth medium (SchwaMed®) (Figure 1. C-D). Besides, other data suggest that Schwann cells are the cell type most responsible for creating a permissive environment for nerve regeneration. The result are being processed for the presentation.

Conclusion: We have shown that Schwann cells can be successfully grown in a bioreactor at scale using our proprietary techniques as a sphere culture. Using this manufacturing we can produce extracellular vesicles that can increase the neurite outgrowth in a dissociated dorsal root model.

A-B, adherent SC culture. C-D suspension sphere culture of SCs