Isobutyramide activates transcription of human fetal gamma- and murine embryonic epsilon(y)-globin genes.

OBJECTIVE

To examine the effect of isobutyramide synthesized in our laboratory on human and murine globin gene expression and to test cell toxicity of the drug.

METHODS

MEL cells were transfected with the recombinant construct muLCRAgammapsibetadeltabeta and the stable transformants were cultured in the medium with different concentrations of isobutyramide. The experimental mice and rabbit were injected with different doses of isobutyramide. The globin mRNAs were analyzed by RNase protection assay. The hematological toxicity and electrolyte toxicity of the drug were tested.

RESULTS

An inducible and dose-dependent expression of the human gamma-, beta- and mouse alpha-globin gene was observed in the transfected MEL cells. The induction of the human gamma-globin gene is significant stronger than that of the beta-globin gene. With 2.5 approximately 5 mmol/L isobutyramide, the induction of the human gamma-globin gene is even more effective than that of mouse alpha-globin gene. After a 15-day injection under the doses of 500 approximately 900 mg x kg(-1) x d(-1), the level of the mouse embryonic epsilon(y)-globin mRNA could be significantly induced up to 3 approximately 4 fold of that of uninjected controls. The changes of hemoglobin(Hb), RBC, hematocrit(HCT), WBC, derived from mice injected with different doses of isobutyramide at the interval of 24 hours for 2 approximately 4 weeks, were generally within the normal range. In rabbits injected with isobutyramide in the same regiment for 2 weeks, the concentration of blood K+, Na+, Cl- and CO2 were all within normal range and serum ionic osmotic pressure remained stable as well.

CONCLUSION

Our results suggested that isobutyramide is a weak inducer of cell differentiation, but it can selectively activate transcription of human gamma-globin gene at a certain degree, and it can act on early stages of erythroid progenitor differentiation in adult mice and activate transcription of embryonic epsilon(y)-globin gene and have no hematological toxicity. Our results have further proved the potential value of isobutyramide in treatment of beta-thalassemia and sickle cell disease.