Ex vivo expansion of human mobilized peripheral blood stem cells using epigenetic modifiers

Background Epigenetic modifications likely control the fate of hematopoietic stem cells (HSCs). The chromatin‐modifying agents (CMAs), 5‐aza‐2′‐deoxycytidine (5azaD) and trichostatin A (TSA), have previously been shown to expand HSCs from cord blood and marrow. Here we assessed whether CMA can also expand HSCs present in growth factor–mobilized human peripheral blood (MPB). Study Design and Methods 5azaD and TSA were sequentially added to CD34+ MPB cells in the presence of cytokines, and the cells were cultured for 9 days. Results After culture, a 3.6 ± 0.5‐fold expansion of CD34+CD90+ cells, a 10.1 ± 0.5‐fold expansion of primitive colony‐forming unit (CFU)‐mix, and a 2.2 ± 0.5‐fold expansion of long‐term cobblestone‐area–forming cells (CAFCs) was observed in 5azaD/TSA‐expanded cells. By contrast, cells cultured in cytokines without 5azaD/TSA displayed no expansion; rather, a reduction in CD34+CD90+ cells (0.7 ± 0.1‐fold) and CAFCs (0.3 ± 0.1‐fold) from their initial numbers was observed. Global hypomethylation corresponding with increased transcript levels of several genes implicated in HSC self‐renewal, including HOXB4, GATA2, and EZH2, was observed in 5azaD/TSA‐expanded MPB cells in contrast to controls. 5azaD/TSA‐expanded MPB cells retained in vivo hematopoietic engraftment capacity. Conclusion MPB CD34+ cells from donors can be expanded using 5azaD/TSA, and these expanded cells retain in vivo hematopoietic reconstitution capacity. This strategy may prove to be potentially useful to augment HSC numbers for patients who fail to mobilize.