Human neural stem cell drug product, culturing and characterization of cell lines for treatment of secondary progressive multiple sclerosis

For several years, human neural stem cells (hNSCs) have been a focus of research, with recent attention to their potential as a cellular treatment for neurodegenerative disorders. The works of Weiss and Reynolds in the 1990s established methods for in vitro cultivation of hNSCs from neural tissue biopsies taken from the subventricular zone (SVZ). Subsequent characterization and development of appropriate potency and safety tests have paved the way for clinical trials exploring the use of these heterologous stem cells as a therapeutic agent. From a regulatory standpoint, hNSCs are classified as advanced therapy medicinal products (ATMPs), requiring production in compliance with Good Manufacturing Practice (GMP) for human use. Neural stem cells possess two key characteristics. First, they can differentiate into specialized mature cells with specific biological functions, particularly giving rise to all nervous tissue cell types, including neurons, astrocytes, and oligodendrocytes. Second, they exhibit self-renewal capacity, undergoing numerous cell cycles without differentiating or becoming senescent, thereby maintaining the stem cell niche and ensuring a pool of cells for proper tissue homeostasis. The manufacturing process outlined here represents the first instance of using brain tissue samples from fetuses that died due to miscarriage for an hNSC-based medicine. This process involves selecting, isolating, and expanding hNSCs, followed by creating a tailored pharmaceutical formulation in accordance with the approved clinical protocol. The cells utilized in this clinical trial underwent microbiological safety testing, with each batch analyzed for identity, potency, and safety using morphological and functional assays. By cultivating NSCs in a medium devoid of growth factors and containing serum, researchers confirmed the presence of neural stem cells in the cultures. These cells demonstrated the ability to differentiate into all central nervous system cell types, even after extensive in vitro manipulation. Stemness was evaluated by examining the capacity of individual stem cells to generate monoclonal spheres, which is a crucial indicator of self-renewal. Neural stem cells are considered a promising therapeutic approach for neurodegenerative disorders, primarily due to their immunomodulatory and neuroprotective properties, rather than their proliferative and differentiation capabilities. In the clinical trial "Safety Study of Human Neural Stem Cells Injections for Secondary Progressive Multiple Sclerosis Patients" (NSC-SPMS EudraCT 2015-004855-37), patients with secondary progressive multiple sclerosis received intrathecal injections of hNSCs cultured at the Terni Hospital Cell factory. This neurodegenerative condition develops from the relapsing-remitting form, with many individuals initially diagnosed with RR eventually transitioning to a secondary progressive stage. This stage is characterized by a steadily worsening disability over time and lacks effective pharmacological interventions. The findings presented in this thesis demonstrate that the process for ATMP drugs can be replicated and standardized. Both in vitro and in vivo studies have shown that hNSCs are safe and effective in treating neurodegenerative diseases, producing beneficial effects on disease progression.