Abstract 3234

Poster Board III-171

Gene therapy has been recently postulated as a realistic therapeutic potential for thalassemia and the mobilized autologous hematopoietic stem cells (HSCs) may represent the preferable source of stem cells for genetic modification due to the higher yield of HSCs compared to conventional bone marrow (bm) harvest. We have previously shown (manuscript under revision) that G-CSF mobilization in the HBBth-3 thalassemic mouse model is less efficient compared to normal C57Bl6 strain, mainly due to increased trapping of hematopoietic stem (Lin-sca-1+ckit+–LSK) and progenitor cells (CFU-GM) in the enlarged thalassemic spleen. The novel mobilizer, AMD3100 (plerixafor, mozobil), has been shown to reversibly bind to CXCR4 and inhibit the interaction between SDF-1 and CXCR4 within the bm microenvironment, resulting in the egress of CD34+ cells into the circulation of healthy donors and cancer patients. The addition of AMD to G-CSF results in even greater increases in circulating CD34+cells. We explored in the current study whether AMD alone or in combination with G-CSF improves the mobilization efficiency of thalassemic mice. C57 and HBBth-3 mice received G-CSF-alone at 250microgr/kgX7 days, AMD-alone at 5mg/kgX3 days or the combination of two with AMD administered in the evening of days 5-7 of G-CSF administration. Hematopoietic tissues (blood, bm, spleen) were collected and the absolute LSK and CFU-GM numbers were calculated based on their frequency within tissues (by FCM and clonogenic assays) in relation to the individual cell count per tissue. AMD-alone didn't significantly affect the HSC yield as compared to G-CSF mobilization in thal mice (LSK/μl blood: 103±85 vs 69±26 p=ns), although it significantly increased the circulating Colony Forming Cells (CFU-GM/ml blood: 1205±533 vs 330±87, p=0,05). In contrast, the AMD+G-CSF combination significantly improved the mobilization efficiency of HBBth-3 mice over the G-CSF-treated group (LSK cells/μl blood: 224±104 vs 69±26 p=0,04, CFU-GM/ml blood: 1671±984 vs 330±87 p=0,05, respectively) at levels comparable to normal mice treated with G-CSF (LSK cells/ μl blood: 241±167, CFU-GM/ml blood: 1235±1140, respectively). AMD induced a “detachment” of stem cells from the bm because reduced numbers of bm LSK cells were counted in the AMD-alone group as compared to the untreated group (LSK/2 femurs×103: 692±429 vs 1687±1016, respectively, p=0,05). This was in contrast to the marrow hyperplasia caused by G-CSF over the steady-state condition (LSK/2 femurs×103: 2684±1743 vs 1687±1016 p=0,02 / CFU-GM/2femurs:111.841±15.391 vs 76.774±31.728 p=0,01). Consequently, the combination of AMD+G-CSF resulted in increased numbers of circulating stem and progenitor cells without inducing marrow hyperplasia as compared to steady-state condition (LSK/2femurs×103: 1681±862 vs 1686±1017, p=ns / CFU-GM/2femurs: 76.774±31.728 vs 82.905±26.277, p=ns). AMD, also in contrast to G-CSF, did not cause increased trapping of stem and progenitor cells in the spleen compared to the untreated condition (LSK cells/spleen×103: 4738±2970 vs 8303±4166 p=ns / CFU-GM/spleen:146.269±93.174 vs 98.518±25.549, p=ns). However, the combination of AMD+G-CSF still resulted in splenic sequestration of progenitor cells (CFU-GM/spleen: 412.176±157.417 vs 98.518±25.549, p=0,0003) but not of LSK cells (LSK cells/spleen×103: 10.200±7.260 vs 8.300±4.166 p=ns). Overall, the combination of AMD3100+G-CSF seems to restore the less efficient mobilization in a thalassemic mouse model. This combination may prove beneficial in a GT setting for obtaining the high numbers of HSCs needed for genetic correction. In addition, the combination of AMD3100+G-CSF, by avoiding the marrow hyperplasia induced by G-CSF alone, indicates a better safety profile because it will not further burden the hyperplastic –due to the increased erythroid demand and the intramarrow destruction of erythroblasts-thalassemic bone marrow.


No relevant conflicts of interest to declare.

Author notes


Asterisk with author names denotes non-ASH members.