Mesenchymal Stem Cells (MSCs) Generated Using mRNA Reprogramming Show Enhanced Growth Potential, Secretome, and Therapeutic Efficacy in a Demyelinating Disease Model

Jasmine K. Harris1, Christopher B. Rohde2, Matthew Angel2

1Novellus, Inc., Cambridge, MA, 2Factor Bioscience Inc., Cambridge, MA

Mol Ther, Vol 28, No 4S1, 2020

MSCs have undergone extensive clinical testing for many diseases and have consistently demonstrated safety. While the immunomodulatory properties of MSCs have been well characterized, adult-tissue-derived MSCs have shown limited therapeutic efficacy as well as significant variability among samples and limited proliferative capacity. Induced pluripotent stem cells (iPSCs) are being explored as an alternative source of MSCs to potentially overcome these limitations. We generated iPSCs from adult human dermal fibroblasts using a high-efficiency, immunosuppressant-free mRNA-based protocol. iPSCs were then differentiated into MSCs using a 21-day high-yield monolayer protocol. rtPCR analysis showed downregulation of Nanog and Oct4 and upregulation of CD73 and CD105 in the differentiated MSCs. Multipotency was confirmed by differentiation into adipocytes, osteoblasts, and chondrocytes. iPSC MSCs had approximately 13kb long telomeres compared to 7kb long telomeres of bone marrow- derived MSCs (BM MSCs), as measured by Southern analysis of terminal restriction fragments. When serially passaged, iPSC MSCs underwent >70 population doublings before senescence, compared to <20 population doublings for BM MSCs. Comparative secretome analysis showed overexpression of multiple neuroprotective and anti- inflammatory factors, including CXCL1, VEGF-A, and CXCL5. When administered to an experimental autoimmune encephalomyelitis (EAE) mouse model, iPSC MSCs delayed disease progression and improved clinical score (p < 0.001), while BM MSCs showed no difference from the disease-only control. Hierarchical clustering analysis revealed greater similarity between BM MSCs and fibroblasts than iPSC-derived MSCs, suggesting that the limited therapeutic efficacy of BM MSCs may be due to loss of an undifferentiated MSC gene-expression pattern.