Combination therapy of erythropoietin, hydroxyurea, and clotrimazole in a β thalassemic mouse: A model for human therapy

Abstract

β thalassemia (β thal) in DBA/2J mice is a consequence of the spontaneous and complete deletion of the β major globin gene. Homozygous β thal mice have clinical and biological features similar to those observed in human β thal intermedia. Erythrocytes in human p thai are characterized by a relative cell dehydration and reduced K+ content. The role of this erythrocyte dehydration in the reduced erythrocyte survival, which typifies the disease, has not previously been evaluated. We examined for 1 month the effects on the anemia and the erythrocyte characteristics of β thal mice of daily treatment with either clotrimazole (CLT), an inhibitor of red blood cell (RBC) dehydration via the Gardos channel, or human recombinant erythropoietin (r-HuEPO), or hydroxyurea (HU). The use of either r-HuEPO or HU induced a significant increase in hemoglobin (Hb), hematocrit (Hct), erythrocyte K+ and a decrease in percent reticulocytes, suggesting improved erythrocyte survival. CLT alone decreased only mean corpuscular hemoglobin concentration (MCHC) and cell density and increased cell K+. Thus, though the Gardos channel plays a major role in cell dehydration of murine β thal erythrocytes, its activity does not contribute to reduced erythrocyte survival. Combination therapy with r-HuEPO plus HU produced no incremental benefit beyond those of single drug therapy. However, addition of CLT to r- HuEPO, to HU, or to combined r-HuEPO plus HU led to statistically significant increases in Hb, Hct, and erythrocyte K+ compared with any of the regimens without CLT. These results suggest that CLT not only inhibits erythrocyte dehydration, but also potentiates the erythropoietic and cellular survival responses to r-HuEPO and HU.