Development of a surrogate angiogenic potency assay for clinical-grade stem cell production

Background aims. Clinical results from acute myocardial infarction (AMI) patients treated with MultiStem ® , a large-scale expanded adherent multipotent progenitor cell population (MAPC), have demonstrated a strong safety and benefi t profi le for these cells. The mechanism of benefi t with MAPC treatment is a result, in part, of its ability to induce neovascularization through trophic support. Production of clinical-grade stem cell products requires the development of lot-release criteria based on potency assays that directly refl ect the fundamental mechanistic pathway underlying the therapeutic response to verify manufacturing process consistency and product potency. Methods and Results . Using an in vitro endothelial tube formation assay, a potency assay has been developed that refl ects MAPC pro-angiogenic activity. Serum-free conditioned media collected from MAPC culture induced endothelial tube formation. A proteomic survey of angiogenic factors produced by the cells in vitro revealed candidate factors linked to angiogenic potency. Three cytokines, chemokine (C-X-C motif) ligand 5 (CXCL5), interleukin 8 (IL-8) and vascular endothelial growth factor (VEGF), were required for this angiogenic activity. Depletion of any of these factors from the media prevented tube formation, while adding back increasing amounts of these cytokines into the depleted serum-free conditioned media established the lower limits of each of the cytokines required to induce angiogenesis. Conclusions. A necessary threshold of angiogenic factor expression was established using an in vitro angiogenesis assay. By correlating the levels of the cytokines required to induce tube formation in vitro with levels of the factors found in the spent media from manufacturing production runs, detection of these factors was identifi ed as a surrogate potency assay with defi ned pass/fail criteria.