The Functional Effect of Repeated Cryopreservation on Transduced CD34 Cells from Patients with Thalassemia.

Stable gene marking and effective engraftment of gene-modified CD34 hematopoietic stem cells is a prerequisite for gene therapy success but may be challenged by the inevitable cryopreservation of the final product prior to extensive quality assurance testing. We investigated the β-globin gene transfer potency in fresh and cryopreserved CD34 cells from mobilized patients with β-thalassemia, as well as the qualitative impact of repeated freeze/thaw cycles on the functionality of cultured and unmanipulated CD34 cells in terms of engrafting capacity in a xenotransplantation model, under partial myeloablation. Cells transduced fresh or after one freeze-thaw cycle yielded similar clonogenic and gene transfer frequencies.

Poor stem cell harvest may not always be related to poor mobilization: lessons gained from a mobilization study in patients with β-thalassemia major.

Background: Hematopoietic stem cell mobilization and leukapheresis in adult patients with β-thalassemia have recently been optimized in the context of clinical trials for obtaining hematopoietic stem cells for thalassemia gene therapy. In some patients, however, the yield of cluster of differentiation 34-positive (CD34+) cells was poor despite successful mobilization, and a modification of apheresis settings was mandatory for harvest rescue.

Study Design And Methods: Data were analyzed from 20 adult patients with β-thalassemia who were enrolled in a clinical trial of optimizing mobilization strategies for stem cell gene therapy.

Plerixafor+G-CSF-mobilized CD34+ cells represent an optimal graft source for thalassemia gene therapy.

Globin gene therapy requires abundant numbers of highly engraftable, autologous hematopoietic stem cells expressing curative levels of β-globin on differentiation. In this study, CD34+ cells from 31 thalassemic patients mobilized with hydroxyurea+granulocyte colony-stimulating factor (G-CSF), G-CSF, Plerixafor, or Plerixafor+G-CSF were transduced with the TNS9.3.

Hematopoietic stem cell mobilization for gene therapy: superior mobilization by the combination of granulocyte-colony stimulating factor plus plerixafor in patients with β-thalassemia major.

Successful stem cell gene therapy requires high numbers of genetically engineered hematopoietic stem cells collected using optimal mobilization strategies. Here we focus on stem cell mobilization strategies for thalassemia and present the results of a plerixafor-based mobilization trial with emphasis on the remobilization with granulocyte-colony stimulating factor (G-CSF)+plerixafor in those patients who had previously failed mobilization. Plerixafor rapidly mobilized CD34(+) cells without inducing hyperleukocytosis; however, 35% of patients failed to reach the target cell dose of ≥6×10(6) CD34(+) cells/kg.

Hematopoietic stem cell mobilization for gene therapy of adult patients with severe β-thalassemia: results of clinical trials using G-CSF or plerixafor in splenectomized and nonsplenectomized subjects.

The safety and efficacy of hematopoietic stem cell (HSC) mobilization was investigated in adult splenectomized (SPL) and non-SPL patients with thalassemia major, in two clinical trials, using different mobilization modes: granulocyte-colony-stimulating factor (G-CSF)-alone, G-CSF following pretreatment with hydroxyurea (HU), plerixafor-alone. G-CSF-mobilization was both safe and effective in non-SPL patients. However, in SPL patients the procedure resulted in excessive response to G-CSF, expressed as early hyperleukocytosis necessitating significant dose reduction, and suboptimal CD34(+) cells yields.