Towards an advanced therapy medicinal product based on mesenchymal stromal cells isolated from the umbilical cord tissue: quality and safety data

Abstract

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.

A new standardized clinical‐grade protocol for banking human umbilical cord tissue cells

“…Moreover, because of their immunomodulatory, homing, and trophic properties, as well as their ability to release cytokines and growth factors, UCT-MSCs are believed to support tissue regeneration and wound healing. [20][21][22][23] Thus, in view of their rapidly increasing applications, the idea of banking UCT-MSCs precursors, that is, UCT cells (UCTCs), appears very interesting. However, for banking purposes the use of standardized protocols for UCTCs isolation, as a prerequisite to subsequent MSCs expansion, remains a major issue, since it requires that all procedures, from tissue collection to final cell product, must be processed following the guidelines for good manufacturing practices (GMP).…”

“…For these multiple capabilities UCT‐MSCs have been employed in many preclinical and clinical trials for the treatment of different pathologies such as Type I diabetes, spinal cord injury, retinal disease, cerebral ischemia, ulcerative colitis, hepatic cirrhosis, Duchenne muscular dystrophy, amyotrophic lateral sclerosis, hereditary cerebellar ataxia, cardiomyopathy, rheumatoid arthritis, multiple sclerosis, and liver failure (http://www.clinicaltrials.com). Moreover, because of their immunomodulatory, homing, and trophic properties, as well as their ability to release cytokines and growth factors, UCT‐MSCs are believed to support tissue regeneration and wound healing …”

Clinical grade expansion of MSCs

“…In the literature, a vast number of papers have been recently published to support the idea to replace FBS with human PL [38,[41][42][43][44][45][46][47][48]. We have already demonstrated that PL has better effects than FBS on proliferation capacity of BM-MSC, therefore providing more efficient expansion, together with significant time saving [37].…”

Human Umbilical Cord Mesenchymal Stromal Cell Transplantation in Myocardial Ischemia (HUC-HEART Trial). A Study Protocol of a Phase 1/2, Controlled and Randomized Trial in Combination with Coronary Artery Bypass Grafting

“…Cell quality control will be validated using the guidelines of the International Conference on Harmonization Q2 (ICH Q2) [33]. The validation tests will include (i) cell enumeration, (ii) immunophenotypic characterization, (iii) viability determination, (iv) microbiological tests for sterility, mycoplasma, endotoxin levels within the cell supernatants and (v) karyotyping, and (vi) tri-lineage (adipogenic, chondrogenic and osteogenic) differentiation tests, as previously described [34]. As an integral part of the cGMP quality assurance system, all documents pertaining to cell bank manufacturing will be made available for review by the national authority.…”