Evaluation of Potential role for Valproic Acid in IL-3-Mediated Megakaryopoiesis and Erythropoiesis in Serum-Free and Serum-Containing Cultures.

Abstract 2746

Poster Board II-722

Although attention has been paid to the antitumor activity of histone deacetylase (HDAC) inhibitors in hematologic disease, a role for HDAC in normal hematopoiesis has not been clearly defined. Previous study have shown that the potent HDAC inhibitor FK228 (romidepsin, depsipeptide) stimulates IL-3-mediated erythropoiesis in serum-free cultures. Here, we precisely examined the effects of well-tolerated antiepileptic drug valproic acid (VPA) on IL-3-mediated megakaryopoiesis and erythropoiesis in serum-free and serum-containing cultures. After incubation of CD34⁺ cells in serum-free medium containing IL-3+SCF, CD61⁺GPA⁻ megakaryocytic, a CD61⁺GPA⁺ mixture of megakaryocytic and erythroid, and CD61⁻GPA⁺ erythroid precursors were generated. Pharmacological concentration of VPA (100 μg/ml) and FK228 (0.45 ng/ml) remarkably increased the number of CD61⁺GPA⁺ cells by 7-fold and 9-fold, respectively. Other populations were also increased by FK228 and VPA. VPA and FK228 enhanced the generation of megakaryocytic and erythroid precursors from more mature CD34⁺ cell-derived CD36⁺ erythroid and megakaryocytic precursors. Thus, VPA was found to be as potent as FK228 in stimulating the IL-3-mediated megakaryopoiesis and erythropoiesis. In the presence of GM-CSF+SCF, CD61⁻GPA⁺ erythroid precursors were mostly developed. Addition of VPA to the cultures mainly promoted the generation of CD61⁻GPA⁺ erythroid precursors, while the generation of megakaryocytic precursors was also moderately enhanced. In serum-containing cultures, there were only low numbers of CD61⁺ or GPA⁺ cells detected even in the presence of IL-3+SCF. Nevertheless, addition of VPA to the cultures containing IL-3+SCF increased the numbers of CD61⁺GPA⁻, CD61⁺GPA⁺, and CD61⁻GPA⁺ cells by 3-fold, 18-fold, and 4-fold, respectively from three independent experiments. In megakaryopoiesis and erythropoiesis, GATA-1 and GATA-2 are shown to play important roles in the proliferation and differentiation process, respectively. By quantitative RT-PCR analysis, after incubation of CD34⁺ cells with IL-3+SCF, GATA-2 m-RNA expression was increased at day1 but returned to initial levels at day3. VPA increased the expression level of GATA-2 gene by 1.6-fold and 1.7-fold at day1 and day3, respectively. While GATA-1 gene expression was remarkably elevated at day1 and day3, its expression was not affected by VPA. Our data reveal a potential for VPA to promote the ability of IL-3 to stimulate megakaryopoiesis and erythropoiesis in serum-free and serum-containing cultures and thus implicate a novel therapeutic approach of epigenetic therapy for treatment of hematologic disease such as myelodysplastic syndrome.