Arsenic trioxide induces differentiation and apoptosis of malignant cells in vitro and in vivo, but the mechanisms by which such effects occur have not been elucidated. In the present study we provide evidence that arsenic trioxide induces activation of the small G-protein Rac1 and the ␣ and ␤ isoforms of the p38 mitogenactivated protein (MAP) kinase in several leukemia cell lines. Such activation of Rac1 and p38-isoforms results in downstream engagement of the MAP kinase-activated protein kinase-2 and is enhanced by pre-treatment of cells with ascorbic acid. Interestingly, pharmacological inhibition of p38 potentiates arsenic-dependent apoptosis and suppression of growth of leukemia cell lines, suggesting that this signaling cascade negatively regulates induction of antileukemic responses by arsenic trioxide. Consistent with this, overexpression of a dominant-negative p38 mutant (p38␤AGF) enhances the antiproliferative effects of arsenic trioxide on target cells. To further define the relevance of activation of the Rac1/p38 MAP kinase pathway in the induction of arsenic-dependent antileukemic effects, studies were performed using bone marrows from patients with chronic myelogenous leukemia. Arsenic trioxide suppressed the growth of leukemic myeloid (CFU-GM) progenitors from such patients, whereas concomitant pharmacological inhibition of the p38 pathway enhanced its growth-suppressive effects. Altogether, these data provide evidence for a novel function of the p38 MAP kinase pathway, acting as a negative regulator of arsenic trioxide-induced apoptosis and inhibition of malignant cell growth.

Myelosuppressive cytokines, in particular IFN-γ and TNF-α, play an important role in the pathogenesis of idiopathic aplastic anemia in humans. It is unknown whether these negative regulators of hemopoiesis suppress stem cells by activating a common signaling cascade or via distinct nonoverlapping pathways. In this study, we provide evidence that a common element in signaling for IFN-γ and TNF-α in human hemopoietic progenitors is the p38/MapKapK-2 signaling cascade. Our studies indicate that pharmacological inhibition of p38 reverses the suppressive effects of IFN-γ and TNF-α on normal human bone marrow-derived erythroid and myeloid progenitors. Most importantly, inhibition of p38 strongly enhances hemopoietic progenitor colony formation from aplastic anemia bone marrows in vitro. Thus, p38 appears to play a critical role in the pathogenesis of aplastic anemia, suggesting that selective pharmacological inhibitors of this kinase may prove useful in the treatment of aplastic anemia and other cytokine-mediated bone marrow failure syndromes.

The myelodysplastic syndromes (MDSs) are collections of heterogeneous hematologic diseases characterized by refractory cytopenias as a result of ineffective hematopoiesis. Development of effective treatments has been impeded by limited insights into any unifying pathogenic pathways. We provide evidence that the p38 MAP kinase is constitutively activated or phosphorylated in MDS bone marrows. Such activation is uniformly IntroductionThe myelodysplastic syndromes (MDSs) comprise a spectrum of stem-cell malignancies characterized by cytologic dysplasia and ineffective hematopoiesis. [1][2][3] Although approximately one third of patients may experience progression to acute leukemia, refractory cytopenias are the principal cause of morbidity and mortality. MDS can be divided into low-and high-risk subtypes using the International Prognostic Scoring System (IPSS), based on features such as the number of hematopoietic deficits, the percentage of marrow blasts, and cytogenetic pattern. 4 Approximately two thirds of patients present with lower-risk disease (Low and Int-1 IPSS scores) characterized by increased rates of apoptosis in the progenitor and differentiated cell compartments in the marrow. [5][6][7][8] High intramedullary apoptosis leads to ineffective hematopoiesis and peripheral cytopenias. Higher grade or more advanced disease categories (Int-2 and High IPSS scores) are associated with a significant risk of leukemia transformation with a corresponding lower apoptotic index and higher percentage of marrow blasts.Cytokines play important roles in the regulation of normal hematopoiesis, and a balance between the actions of hematopoietic growth factors and myelosuppressive factors is required for optimal production of different hematopoietic-cell lineages. Excess production of inhibitory cytokines contributes in part to ineffective hematopoiesis in MDS. Tumor necrosis factor-␣ (TNF␣) has been implicated in the increased stem-cell apoptosis seen in MDS, 9,10 and high expression of TNF receptors and TNF mRNA have been reported in MDS bone marrows. [11][12][13][14] Transforming growth factor-␤ (TGF␤), interleukin-6 (IL-6), vascular endothelial growth factor (VEGF), and interferon (IFN-␥ and -␣) are also myelosuppressive, and these proinflammatory cytokines have been found to be elevated in serum of patients with MDS in various studies and are hypothesized to play a role in suppressing hematopoiesis in this disease. 9,11,[15][16][17] Because multiple cytokines are involved in promoting abnormal hematopoietic development in MDS, targeting one single cytokine may not yield appreciable clinical benefit. In fact, anti-TNF therapeutic strategies (monoclonal antibodies and TNFR blockers) have only shown minimal efficacy. [18][19][20][21] Thus, it is imperative to identify common targetable pathways that regulate many different cytokines. Our previous studies have shown that myelosuppressive cytokines such as interferons (IFN-␣, -␤, and -␥), TGF␤, and TNF␣ can all activate the p38 mitogen activated protein kinase (MAPK) in ...