Preclinical Tumorigenicity Study of an Advanced Therapy Medicinal Product for Diffuse Cartilage Lesions in an Osteoarthritic Environment.

BACKGROUND: Advanced therapy medicinal products require rigorous preclinical testing to exclude tumorigenicity. Human articular cartilage cells expanded at low density with human platelet lysate show enhanced proliferation, matrix production, and immunomodulatory properties, supporting their use for diffuse cartilage lesions in osteoarthritic joints. This study evaluated tumorigenicity and biodistribution of cartilage cell spheroids generated using two platelet lysate sources. METHODS: Cartilage cells were expanded at low density with two platelet lysates and assembled into spheroids. Cytogenetic stability was assessed by metaphase karyotyping following expansion. Immunodeficient mice received subcutaneous implantation and were monitored for 180 days. Human colon carcinoma cells and mouse fibroblasts were used as controls. Clinical follow-up, full organ histopathology, and immunohistochemistry were performed to detect human cell persistence. RESULTS: Expanded cartilage cells showed predominantly normal karyotypes, with rare low-level mosaic chromosomal alterations not detected at the previous passage. Cartilage cell spheroids were well tolerated in vivo, with complete survival and no evidence of tumorigenicity, inflammation, or human cell persistence at implantation sites or distant organs. Control experiments confirmed the sensitivity of the model, and no systemic toxicity was observed. CONCLUSIONS: Spheroids derived from cartilage cells are non-tumorigenic, non-migratory, and biologically safe in immunodeficient mice. These findings support their development as cell-based cartilage therapies and align with regulatory recommendations for non-clinical safety evaluation.