Abstract
BackgroundCD133+ stem cells represent a promising subpopulation for innovative cell-based therapies in cardiovascular regeneration. Several clinical trials have shown remarkable beneficial effects following their intramyocardial transplantation. Yet, the purification of CD133+ stem cells is typically performed in centralized clean room facilities using semi-automatic manufacturing processes based on magnetic cell sorting (MACS®). However, this requires time-consuming and cost-intensive logistics.MethodsCD133+ stem cells were purified from patient-derived sternal bone marrow using the recently developed automatic CliniMACS Prodigy® BM-133 System (Prodigy). The entire manufacturing process, as well as the subsequent quality control of the final cell product (CP), were realized on-site and in compliance with EU guidelines for Good Manufacturing Practice. The biological activity of automatically isolated CD133+ cells was evaluated and compared to manually isolated CD133+ cells via functional assays as well as immunofluorescence microscopy. In addition, the regenerative potential of purified stem cells was assessed 3 weeks after transplantation in immunodeficient mice which had been subjected to experimental myocardial infarction.ResultsWe established for the first time an on-site manufacturing procedure for stem CPs intended for the treatment of ischemic heart diseases using an automatized system. On average, 0.88 × 106 viable CD133+ cells with a mean log10 depletion of 3.23 ± 0.19 of non-target cells were isolated. Furthermore, we demonstrated that these automatically isolated cells bear proliferation and differentiation capacities comparable to manually isolated cells in vitro. Moreover, the automatically generated CP shows equal cardiac regeneration potential in vivo.ConclusionsOur results indicate that the Prodigy is a powerful system for automatic manufacturing of a CD133+ CP within few hours. Compared to conventional manufacturing processes, future clinical application of this system offers multiple benefits including stable CP quality and on-site purification under reduced clean room requirements. This will allow saving of time, reduced logistics and diminished costs.
Highlights
CD133+ stem cells represent a promising subpopulation for innovative cell-based therapies in cardiovascular regeneration
We evaluated the recently developed CEmarked CliniMACS Prodigy® bone marrow (BM)-133 System (Prodigy) (Miltenyi Biotec GmbH, Bergisch Gladbach, Germany) in order to assess its suitability for the automatic manufacturing of a CD133+ cell product (CP) [20,21,22]
The data were collected during the Good Manufacturing Practice (GMP) inspection process, which led to the certification of GMP in compliance with EU guidelines, quality control (QC) and media fill validation were performed by the Institute for Clinical Chemistry and Laboratory Medicine (ILAB, Rostock University Medical Center) and the Institute for Medical Microbiology, Virology and Hygiene (IMIKRO, Rostock University Medical Center), which are accredited for compliance with clinical laboratory quality standards (ILAB: DIN EN ISO 15189, DIN EN ISO 22870, license D-ML-13193_02_00; IMIKRO: DIN EN ISO 15189, DIN EN ISO/IEC 17025)
Summary
CD133+ stem cells represent a promising subpopulation for innovative cell-based therapies in cardiovascular regeneration. Several clinical trials showed an increased left ventricular ejection fraction (LVEF) and an improved regional perfusion following intramyocardial application of cluster of differentiation (CD) 133+ stem cells [5,6,7,8,9] These adult cells represent a subset of CD34+ progenitors and are accessible from patient-derived bone marrow (BM) posing little ethical conflicts [10,11,12]. CD133+ stem cells for clinical use are purified using manual or semi-automatic devices based on the conventional magnetic cell sorting (MACS®) technique Such purification requires highlevel clean room facilities [19] and time-consuming logistics (Additional file 1: Figure S1a). The development of an automatic system is urgently needed to guarantee stable product quality, to simplify the manufacturing procedure and to reduce the risk of contamination (Additional file 1: Figure S1b)
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