Developing an Advanced Therapy Medicinal Product (ATMP) for the treatment of GvHD

Abstract

Mesenchymal stem/ stromal cells (MSCs) have been proved to be capable to modulate the immune system through direct interactions target cell-MSC and secretion of soluble molecules that are induced or upregulated following cross-talk with target cells. In this study, a full biological characterization of ImmuneSafe® (IS) features including identity, potency and safety which constitute the Critical Quality Attributes (CQAs) of the product was performed with purpose of providing tools that will assure the consistency and robustness of the manufacturing process or demonstrating product/ process comparability after a particular change in the manufacturing process. In order to achieve this goal a (bio) assay panel was developed and applied to IS in two different steps of manufacturing process. IS CQAs allowed a robust and reproducible characterization of the product, demonstrating their potential to be used throughout the production stage. Similar studies were also performed with similar cell types, such as human skin fibroblasts and MSCs differentiated in adipocytes and osteocytes, which were then benchmarked with IS. The results showed that none of these cell types demonstrated a comparable level of therapeutic potency to IS. The patient enrolling protocol for IS clinical trial to treat GvHD will include the administration of immunosuppressive drugs (methylprednisolone or prednisolone) concomitantly with IS administration. The impact of these drugs on IS identity was evaluated through immunophenotype characterization and potency was evaluated through the activation of the different immunomodulatory pathways. The results showed that immunosuppressive drugs tested, methylprednisolone and prednisolone, did not seem to have a beneficial or detrimental interaction with IS. MSCs are commonly stored in cryopreservation conditions before the deliver to the patient. However, recent studies have shown that banked fresh thawed MSCs have impaired immunomodulatory properties compared to MSCs in culture. IS response to an inflammatory microenvironment was compared in different times of release and fresh thawed cells revealed to have several responses compromised under pro-inflammatory environment. Additionally, IS secretome was also affected, since the production of several cytokines were decreased or even switched off, as well as the immunosuppressive activity of the product. For these reasons the implementation of a release culture step was found to be advantageous in order to maximize the therapeutic potency of IS. Another important issue for cell-based therapies is the product delivery to the hospital. Cells should be formulated in a suitable excipient for intravenous infusion capable of maintaining the cell viability and therapeutic potential during the purposed product shelf-life. For this purpose IS was formulated in a saline solution and identity and potency tests were performed. Hypothermosol was capable of maintaining at least 70% of initial cell number population with 80% of viability, as well as its identity and potency features within a 48h-window. These studies enable a comprehensive IS characterization and the set-up of the assays to be used in the manufacturing process under GMP conditions. Additionally, no detrimental effects on the therapeutic potency of IS were associated with the interactions with the immunosuppressive drugs that will be used in the clinical trial, thus indicating the clinical results will not probably be affected by the background therapy applied to patients. The process of product release was also optimized to guarantee a cell product with maximized immunomodulatory properties and a 48-hour shelf-life was determined, which is a critical issue for the planning of IS logistics for the clinical trial.