In Vitro Chemotactic Index and Response to CXCR4 Antagonist AMD3100 Correlate with CD34⁺ Cell Yield from Leukapheresis Products: Implications for Poor Mobilizers.

While autologous hematopoietic stem/progenitor cell (HSPC) transplantation has been used successfully for many years, new approaches are being explored to boost the number of HSPC (CD34⁺ cells) collected by leukapheresis to permit more rapid hematopoietic recovery. Still, a considerable number of patients may be excluded from this procedure because a sufficient quantity of HSPC cannot be obtained by standard mobilization agents. Among factors regulating mobilization is stromal cell-derived factor (SDF)-1 constitutively produced by bone marrow (BM) stromal cells and which is known to retain HSPC in the BM. Responsiveness to an SDF-1 gradient can be measured in vitro using the chemotactic index, defined as the ratio of the number of cells that migrate towards SDF-1 and the number of cells that migrate towards media alone. We hypothesized that (i) a high chemotactic index would indicate a greater ability of the HSPC to be retained in their BM niches and therefore more difficulty in coaxing them out into the circulation, as could be the case in poor mobilizers; and that (ii) AMD3100, an antagonist of the SDF-1 receptor CXCR4, will have a greater disruptive effect on SDF-1-dependent signalling in poor mobilizers. Leukapheresis products were obtained from patients diagnosed with Hodgkin’s or non-Hodgkin’s lymphoma and CD34⁺ cells isolated by positive selection were loaded onto Boyden chambers and allowed to migrate across bare filters towards a gradient of SDF-1 (200 ng/mL). In some experiments cells were incubated with AMD3100 (10 mg/mL) for the duration of the assay. We observed broad inter-patient differences in in vitro migratory ability of CD34⁺ cells, ranging from 3.1 ± 0.6 to 21.7 ± 2.1 % for spontaneous or passive percentage migration; and from 11.1 ± 0.7 to 58.6 ± 10.7 % for SDF-1-directed chemotaxis, neither of which bore any correlation with the number of CD34⁺ cells/kg obtained from the leukapheresis product. However, a negative correlation (r=−0.7) was found between the chemotactic index and the percentage of CD34⁺ cells obtained from the leukapheresis product, i.e., the good mobilizers responded poorly to an SDF-1 gradient whereas the poor mobilizers responded better. Also a higher expression of CXCR4 was observed (by flow cytometry) in samples from poor mobilizers. Chemotaxis towards SDF-1 was reduced by 10 to 80% by AMD3100 and there was a positive correlation (r=0.6) between chemotactic index and percentage inhibition, i.e., chemotaxis of poor mobilizers was significantly more inhibited by this CXCR4 antagonist. Thus our results suggest that

• the chemotactic index could be employed as a predictor of good vs. poor mobilization;

• optimal mobilization especially in poor mobilizers may be achieved by a protocol with both G-CSF and AMD3100;

• for good mobilizers, including AMD3100 for mobilization could lead to a reduced requirement for volume of leukapheresis product and number of collections; and finally

• in an allogeneic transplant setting the chemotactic index could be used to predict whether a normal donor of HSPC will be a good or poor mobilizer.