Development of an automated workflow for autologous 2D-iPSCs generation via non-integrating Sendai virus vector.

Abstract

Induced pluripotent stem cells (iPSCs) represent a transformative platform for regenerative and personalized medicine. Autologous iPSC therapies offer superior immunological compatibility, particularly for pediatric patients and indications requiring durable engraftment. However, individualized production remains constrained by substantial logistical and economic challenges. A critical barrier to clinical translation is the absence of closed, automated, and GMP-compliant culture systems that can support standardized and reproducible workflows. A previously manual iPSC generation protocol was adapted to the CliniMACS Prodigy platform to establish a fully automated, closed workflow under GMP-compliant conditions. Key steps-including vessel surface coating, density gradient centrifugation (DGC), and Sendai virus-mediated reprogramming-were integrated into a single-device process. Peripheral blood mononuclear cells were isolated, reprogrammed, and expanded within the system using existing configurations. Process optimization demonstrated that vessel coating prior to DGC enhanced cell adhesion and improved workflow reproducibility. The resulting iPSCs exhibited stable colony morphology, expressed pluripotency markers, and retained robust trilineage differentiation potential, including cardiomyocyte induction. To our knowledge, this study provides the first demonstration of a fully automated, closed-system workflow for autologous iPSC production. The approach offers a practical pathway toward standardized, cost-effective manufacturing to support future clinical applications in cell therapy and regenerative medicine.