Background & Aim: The Cellular Immuno-Therapy for COVID-19 related ARDS (CIRCA-19) was a phase 1, single site, dose escalation trial using a 3+3+3 design to determine the safety and maximum feasible tolerated dose of intravenously delivered, freshly cultured UC-MSCs. Nine patients, each receiving repeated unit doses of UC-MSCs over 3 consecutive days, were enrolled into 3 dose panels: Panel 1: 25×106 cells/dose (cumulative dose: 75×106 MSCs);Panel 2: 50×106 cells/dose (cumulative dose: 150×106 MSCs);Panel 3: up to 90×106 cells/dose (cumulative dose: 270×106 MSCs). Methods, Results & Conclusion: UC-MSCs were isolated from cords of healthy term pregnancies delivered by C-section. Cords were mechanically and enzymatically digested, and UC-MSCs were propagated in xeno-free conditions for 2 weeks prior to cryopreservation in a cord specific cell bank. One fully validated cell bank was used in CIRCA-19 that was free of adventitious agents (HBV, HCV, HSV1/2, Parvo B19 and Retroviruses), had high viability (>95%) and MSC identity with positive expression (>95%) of CD73, CD90 and CD105 and negative expression (<5%) of CD14, CD19, CD34, CD45 and HLA-DR. UC-MSCs also demonstrated high proliferative capacity (EdU+ >45%;DBT = 22h) and enhanced IDO expression (ΔΔCq?>18) when treated with IFN-γ. For the final product, UC-MSCs were thawed, plated and cultured for 24 to 120 h before harvesting to produce a batch of the final drug product formulated as 2.5×106 fresh UC-MSCs/mL suspended in PlasmaLyte A containing 5% Human Albumin, to be infused within 48h. Batches were tested for viability, endotoxin level, ACE-2 expression, tissue factor activity, sterility and mycoplasma. Sixteen batches of UC-MSCs were produced for a total of 41 cell doses (16 doses of 25M;13 doses of 50M;12 doses of 90M cells each). Twenty-seven of the 41 doses (9 doses of 25M, 50M and 90M cells each) were used to treat trial participants. The remaining doses were used for stability studies. All drug products had high viability (> 95%), endotoxin levels of <0.2 EU/mL and tested negative for mycoplasma and bacterial contaminants. All UC-MSC batches were negative for ACE-2 expression (Cq?>35;GAPHD Cq: 15±2;no detectable levels by western blotting) and had tissue factor activity levels between 250-310pM. UC-MSC drug product was stable for up to 96h (>80% viability) and had?>90% viability up to 48h in all 3 dose panels. This study demonstrates the feasibility of manufacture and delivery of a multi-dose fresh cell product in an emergent ICU setting.
Manufacturing freshly cultured umbilical cord-derivedmesenchymal stromal cells (UC-MSCS) for a phase 1, multiple-dose clinical trial for COVID-19-induced acute respiratory distress syndrome (ARDS)