Evaluation of a cell-based osteogenic formulation compliant with good manufacturing practice for use in tissue engineering

Abstract

Background: Proper bony tissue regeneration requires mechanical stabilization, an osteogenic biological activity and appropriate scaffolds. The latter two elements can be combined in a hydrogel format for effective delivery, so it can readily adapt to the architecture of the defect. Results: We evaluated a Good Manufacturing Practice-compliant formulation composed of bone marrow-derived mesenchymal stromal cells in combination with bone particles (Ø = 0.25 to 1 µm) and fibrin, which can be readily translated into the clinical setting for the treatment of bone defects, as an alternative to bone tissue autografts. Remarkably, cells survived with unaltered phenotype (CD73+, CD90+, CD105+, CD31-, CD45-) and retained their osteogenic capacity up to 48 h after being combined with hydrogel and bone particles, thus demonstrating the stability of their identity and potency. Moreover, in a subchronic toxicity in vivo study, no toxicity was observed upon subcutaneous administration in athymic mice and signs of osteogenesis and vascularization were detected two months after administration. Conclusions: The preclinical data gathered in the present work, in compliance with current quality and regulatory requirements, demonstrated the feasibility of formulating an osteogenic cell-based tissue engineering product with a defined profile including identity, purity and potency (in vitro and in vivo), and the stability of these attributes, which complements the preclinical package required prior to move towards its use of prior to its clinical use. Background Failure of the physiologic reaction to acute or chronic bone disorders (e.g. fractures, non-unions, large defects after trauma or tumors) typically requires surgical intervention and implantation of bone grafts. Autografts and heterologous transplants of bony tissue, as well as the use of implants made of biomaterials, are the most common approaches in today's orthopedics field (1). Among them, the bony autograft sourced from the iliac crest is the preferred treatment option for a wide range of orthopedic conditions, including the management of complex fractures or in non-union defects (1, 2). The use of autologous tissue as a vector for bony regeneration fulfils three key requirements: 1) introduces cells with osteogenic potential, 2) offers structural support (osteoconduction), and 3)

Preclinical Development of a Therapy for Chronic Traumatic Spinal Cord Injury in Rats Using Human Wharton’s Jelly Mesenchymal Stromal Cells: Proof of Concept and Regulatory Compliance

“…Proliferation of MSC,WJ cultured in presence or absence of FK506 (Fujisawa Pharmaceuticals, Osaka, Japan) was monitored by using the ATP-based CellTiter-Glo ® Luminescent Cell Viability Assay (Promega, Madison, WI, USA) as reported elsewhere [ 18 ]. Briefly, cells were cultured in multiwell format plates in the presence or absence of 10 ng/mL FK506 diluted in DMSO and, at the time of analysis, luminescent reagent was added to each well at 1:1 ( v / v ) ratio with respect to culture medium and mixed for 2 min using an orbital shaker.…”

Preservation of critical quality attributes of mesenchymal stromal cells in 3D bioprinted structures by using natural hydrogel scaffolds

“…An experimental matrix was defined to test suitable combinations of hydrogels, which were chosen based on prior preclinical and clinical experience in the generation of (non‐3D printed) osteogenic MSC‐based TEP, as reported elsewhere (García de Frutos et al, 2020; Prat et al, 2018; Vivas et al, 2020; Vives et al, 2021). Particularly: clinical grade fibrin (Tissucol Duo; Baxter) as a result from mixing fibrinogen and 1:100 diluted thrombin at 5 UI/mL final concentration in saline solution (Viaflo Plasmalyte 148; Baxter) supplemented with 2% human serum albumin (HSA; Albutein®; Grifols); clinical grade HA (Bioiberica) dissolved either at 1% or 5% (w/v) in warm saline solution and sterilized by filtration with a 0.45 µm Millex‐HV filter (Merck Millipore) or by autoclaving, respectively; and research grade polymer precursors of gelatin and alginate methacryloyl (GelMA and AlgMA, respectively) (Sigma‐Aldrich) were altered to a 40% methacrylation degree.…”