Ex Vivo Expansion of Mobilized Peripheral Blood Myeloid Progenitor Cells: Pretreated Patients vs Normal Donors.

Background: Additional infusion of ex vivo differentiated myeloid progenitor may reduce or abrogate severe neutropenia following high-dose chemotherapy and peripheral blood (PB) transplantation. Normal HPC as well as HPC from patients in clinical remission can be used as source of cells. The aims of our study were to compare the ex vivo expansion of myeloid progenitor cells starting from pretreated patients (PP) and normal donors (ND) and to evaluate if the cells collected from poor mobilizers -for which the ex vivo expansion is particularly interesting- present similar ex vivo expansion capacities than cells from good mobilizers.

Methods: In this study, 11 normal donors (ND) and 17 patients (PP) were evaluated (5 NHL, 2 HK, 3 MM, 3 lymphoma, 1AML, 1 CLL and 2 solid tumor). The CD34⁺ cells were plated for 7 days in serum-free medium with SCF, IL-3, G-CSF and FLT3- ligand. The expression of CD34, CD13, CD15 and CD16 Ags and the presence of primary (myeloperoxydase, MPO) and secondary granules (lactoferrin, LF) were used to evaluate maturation into myeloid lineage. Functional capacities of the ex vivo generated cells were also analysed using oxydative burst activity and clonogenic capacities (CFU-GM).

Results: (1) the proliferation capacities of total leucocytes were higher in ND than in PP (28.8±5.6 vs 17.2±4.0 fold, p<0.04) with a comparable CD34⁺ cell expansion (8.7±3.0 vs 7.8±2.7 fold, NS); (2) in patients, 65.3±5.8% of the expanded cells were CD13⁺ and 23.2±4.1% remained CD34⁺ with 7.8±2.5 % of immature CD34⁺CD13⁻ cells; these results were similar to that observed in ND (72.7±7.5% CD13⁺, 23.0±5.3% CD34⁺, 9.0±3.2% CD34⁺CD13⁻); (3) the number of myeloid progenitors produced per seeded CD34⁺ cell consisted of 19.0±3.7 CD13⁺, 11.8±3.7 CD15⁺, 10.2±3.2 CD16⁺ and 15.4±4.4 MPO⁺/LF⁺cells in PP and 25.3±5.4 CD13⁺, 12.5±3.8 CD15⁺, 7.6±2.1 CD16⁺ and 21.7±7.2 MPO⁺/LF⁺ cells in ND (NS); (4) these cells, essentially promyelocytes and metamyelocytes, were able to proliferate in clonogenic assay and presented oxydative burst activity; (5) we failed to observe a correlation between the cell expansions (total leucocytes, CD34⁺ cells and myeloid progenitor cells) and the diagnosis; (6) no correlation was observed between the mobilization of CD34⁺ cells into PB and the capacities of cells to be expanded in vitro.

Conclusions: in our culture system, the generation of cells committed to the myeloid lineage from CD34⁺ cells is feasible from ND as well as from PP, whatever is the diagnosis. The ex vivo expanded cell population consisted of cells at all stages of differentiation including immature hematopoietic progenitors. The mobilisation rate doesn’t correlate with the proliferative capacities of cells, suggesting that ex vivo expansion can be performed even in subjects with poor mobilization.