GMP manufacturing, quality characterization, and GLP-based preclinical evaluation of human nasal turbinate–derived stem cells for the treatment of rheumatoid arthritis

Vollständiger Abstract

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Abstract Background Human nasal turbinate-derived stem cells (hNTSCs) are tissue-derived adult stem cells that are isolated from surgically discarded nasal turbinate tissue and exhibit robust proliferative capacity, and stable phenotypic properties. Unlike conventional MSCs, hNTSCs, although classified as tissue-derived adult stem cells, exhibit MSC-like phenotypes but display distinct gene expression patterns and functional properties that may be advantageous for indications of inflammatory and autoimmune disease. Rheumatoid arthritis (RA) is a chronic autoimmune disorder characterized by persistent synovitis, progressive joint destruction, and incomplete responses to existing disease-modifying antirheumatic drugs and biologics. Cell-based immunomodulatory therapies represent promising alternative strategies; however, the clinical translation of novel stem cell platforms requires not only the demonstration of therapeutic efficacy but also establishment of a reproducible manufacturing process, rigorous quality characterization, and comprehensive nonclinical safety evaluation.In this study, we developed a clinical-grade injectable hNTSC product for treating RA by integrating a GMP-equivalent manufacturing process, systematic quality control (QC) testing, and GLP-compliant nonclinical safety, biodistribution, and efficacy assessments. This integrated regulatory science framework supports the potential of hNTSCs as a distinct and clinically translatable adult stem cell platform for the treatment of rheumatoid arthritis. Methods Clinical-grade hNTSCs were manufactured from surgically obtained nasal turbinate tissue using a GMP-equivalent process that included serial cell expansion, multitiered cell banking, and final drug product formulation. Quality control and characterization included flow cytometry (FACS)-based analyses for cell identity and purity, as well as in vitro differentiation assays to assess biological characteristics. Additional quality assessments included cell viability, genetic stability, sterility, and mycoplasma, endotoxin, and adventitious virus testing. Biodistribution was evaluated using a validated TaqMan-based quantitative PCR (qPCR) assay targeting human ALU sequences to quantify hNTSC-derived DNA in mouse tissues. GLP-compliant nonclinical studies included single-dose intravenous biodistribution, repeated-dose toxicity with a recovery period, tumorigenicity assessment, and efficacy evaluation in a collagen-induced arthritis (CIA) mouse model. Results The GMP-equivalent manufacturing process consistently produced hNTSC injectable products that met predefined release criteria across multiple cell banks and batches, demonstrating process robustness and product reproducibility. Flow cytometry confirmed that the identity and purity of the hNTSCs were satisfactory, and differentiation assays demonstrated the maintenance of key biological characteristics. The validated qPCR assay enabled sensitive and reliable detection of hNTSC-derived DNA in biological matrices. Biodistribution analysis revealed predominant localization in the lung at early time points following intravenous administration, followed by a time-dependent decline, with minimal or no detectable signals in most tissues at later time points. Repeated-dose toxicity and tumorigenicity studies revealed no treatment-related mortality, systemic toxicity, or tumor formation. In the CIA mouse model, treatment with hNTSCs significantly attenuated clinical arthritis severity and joint destruction, accompanied by improvements in bone structural parameters and histopathological findings. Conclusions This study demonstrates the successful establishment of a GMP-equivalent manufacturing process, comprehensive quality characterization incorporating flow cytometry–based identity and purity testing and differentiation-based functional assessment, and GLP-compliant nonclinical evaluation for hNTSC injectable cell therapy. These findings support the further clinical development of hNTSCs as a novel cell-based therapy for the treatment of rheumatoid arthritis.