Translation of a standardized manufacturing protocol for mesenchymal stromal cells: A systematic comparison of validation and manufacturing data

TLDR

Expansion protocols for mesenchymal stromal cells (MSCs) are well documented, yet data on translating validated GMP protocols from laboratory to clinical manufacturing remain scarce. This study validates and translates a standardized pre‑clinical MSC isolation and expansion protocol for use in a clinical trial aimed at alveolar bone reconstruction. The authors compared key parameters from 22 large‑scale bone‑marrow MSC expansions used for validation with 11 expansions produced for the MAXILLO‑1 trial, using metrics such as CFU‑F, CD34⁺, MNCs, and WBCs to predict yield and avoid costly failures. Manufacturing the autologous MSC product MAXILLO‑1‑MSC was achieved within 21 days, with BM aspirates and cells transported within 24 h, meeting all regulatory quality criteria; a transient mosaic chromosomal finding did not impair differentiation or surface markers, the mosaic proportion declined over long‑term culture, and cells ceased growth after 38.4 population doublings, demonstrating feasibility for bone regeneration even across long distances.

Abstract

Many data are available on expansion protocols for mesenchymal stromal cells (MSCs) for both experimental settings and manufacturing for clinical trials. However, there is a lack of information on translation of established protocols for Good Manufacturing Practice (GMP) from validation to manufacturing for clinical application. We present the validation and translation of a standardized pre-clinical protocol for isolation and expansion of MSCs for a clinical trial for reconstitution of alveolar bone.Key parameters of 22 large-scale expansions of MSCs from bone marrow (BM) for validation were compared with 11 expansions manufactured for the clinical trial "Jaw bone reconstruction using a combination of autologous mesenchymal stromal cells and biomaterial prior to dental implant placement (MAXILLO1)" aimed at reconstruction of alveolar bone.Despite variations of the starting material, the robust protocol led to stable performance characteristics of expanded MSCs. Manufacturing of the autologous advanced therapy medicinal product MAXILLO-1-MSC was possible, requiring 21 days for each product. Transport of BM aspirates and MSCs within 24 h was guaranteed. MSCs fulfilled quality criteria requested by the national competent authority. In one case, the delivered MSCs developed a mosaic in chromosomal finding, showing no abnormality in differentiation capacity, growth behavior or surface marker expression during long-term culture. The proportion of cells with the mosaic decreased in long-term culture and cells stopped growth after 38.4 population doublings.Clinical use of freshly prepared MSCs, manufactured according to a standardized and validated protocol, is feasible for bone regeneration, even if there was a long local distance between manufacturing center and clinical site. Several parameters, such as colony forming units fibroblasts (CFU-F), percentage of CD34+ cells, cell count of mononuclear cells (MNCs) and white blood cells (WBCs), of the BM may serve as a predictive tool for the yield of MSCs and may help to avoid unnecessary costs for MSC manufacturing due to insufficient cell expansion rates.