Preclinical Evaluation of the Safety and Efficacy of Cryopreserved Bone Marrow Mesenchymal Stromal Cells for Corneal Repair

Abstract

Purpose Mesenchymal stromal cells (MSCs) have been shown to enhance tissue repair as a cell-based therapy. In preparation for a phase I clinical study, we evaluated the safety, dosing, and efficacy of bone marrow–derived MSCs after subconjunctival injection in preclinical animal models of mice, rats, and rabbits. Methods Human bone marrow–derived MSCs were expanded to passage 4 and cryopreserved. Viability of MSCs after thawing and injection through small-gauge needles was evaluated by vital dye staining. The in vivo safety of human and rabbit MSCs was studied by subconjunctivally injecting MSCs in rabbits with follow-up to 90 days. The potency of MSCs on accelerating wound healing was evaluated in vitro using a scratch assay and in vivo using 2-mm corneal epithelial debridement wounds in mice. Human MSCs were tracked after subconjunctival injection in rat and rabbit eyes. Results The viability of MSCs after thawing and immediate injection through 27- and 30-gauge needles was 93.1% ± 2.1% and 94.9% ± 1.3%, respectively. Rabbit eyes demonstrated mild self-limiting conjunctival inflammation at the site of injection with human but not rabbit MSCs. In scratch assay, the mean wound healing area was 93.5% ± 12.1% in epithelial cells co-cultured with MSCs compared with 40.8% ± 23.1% in controls. At 24 hours after wounding, all MSC-injected murine eyes had 100% corneal wound closure compared with 79.9% ± 5.5% in controls. Human MSCs were detectable in the subconjunctival area and peripheral cornea at 14 days after injection. Conclusions Subconjunctival administration of MSCs is safe and effective in promoting corneal epithelial wound healing in animal models. Translational Relevance These results provide preclinical data to support a phase I clinical study.

Cryopreservation does not change the performance and characteristics of allogenic mesenchymal stem cells highly over-expressing a cytoplasmic therapeutic transgene for cancer treatment

“…The successful development of 'off-the-shelf ' GDEPT is dependent on cells retaining their therapeutic functions post freeze-thaw. Majority of the studies examined impact of cryopreservation on functionality of MSC such as their immunomodulatory [46,51], anti-inflammatory properties [17,58] and regenerative properties [59,60]. In contrast to the defined pharmacokinetics of single agent, unmodified MSC therapeutics demonstrate conditional, complex and multifactorial paracrine effect in vitro and in vivo [55][56][57].…”

Pre-Clinical Evaluation of Efficacy and Safety of Human Limbus-Derived Stromal/Mesenchymal Stem Cells with and without Alginate Encapsulation for Future Clinical Applications

“…In order to determine the toxicity or safety profile of the drug or cell product, preclinical testing and compliance with various regulatory requirements are required. MSCs derived from bone marrow have been shown to be safe and effective for corneal repair in a recent study by Putra et al [ 49 ]. The aforementioned study was carried out in advance of the Phase I clinical trial.…”

Impact of Cryopreservation and Freeze-Thawing on Therapeutic Properties of Mesenchymal Stromal/Stem Cells and Other Common Cellular Therapeutics

“…Although some studies support the notion that fresh low-passage MSCs can provide a more efficacious therapy [ 9 •, 10 , 12 ], there are also some preclinical data that propose that thawed MSCs are somewhat equal to fresh MSCs [ 104 – 110 ]. Indeed, the biggest puzzle to solve is the current lack of understanding considering MSCs’ MoA in humans upon infusion [ 34 , 35 ].…”