Background. β-thalassemia is caused by mutations in the β-globin locus resulting in loss of, or reduced, Hemoglobin A (Adult hemoglobin, HbA, α2β2) production. Hydroxyurea treatment increases fetal γ-globin (Fetal Hemoglobin, HbF, α2γ2) expression in postnatal life substituting for the missing adult β-globin, and is therefore an appealing therapeutic approach. Patients treated with Hydroxyurea fall into three categories: (i) Responders, who increase hemoglobin to therapeutic levels (ii) Moderate-responders, who increase hemoglobin levels but still need transfusions at longer intervals, and (iii) Non-responders, who do not reach adequate hemoglobin levels and remain transfusion-dependent. The mechanisms underlying these differential responses remain largely unclear. Design and Methods. We generated RNA expression profiles from erythroblast progenitors of 8 responder and 8 non-responder β-thalassemia patients. Results. These profiles revealed that Hydroxyurea treatment induced differential expression of many genes in cells from non-responders while it had little impact on cells from responders. Part of the gene program upregulated by Hydroxyurea in non-responders was already highly expressed in responders before Hydroxyurea treatment. Baseline HbF expression was low in non-responders, and Hydroxyurea treatment induced significant cell death. Conclusions. We conclude that cells from responders have adapted well to constitutive stress conditions and display a propensity to proceed to the erythroid differentiation program.