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Aggressive systemic mastocytosis (ASM) and mast cell leukemia (MCL) are advanced hematopoietic neoplasms with poor prognosis. In these patients, neoplastic mast cells (MCs) are resistant against various drugs. We examined the effects of 2 demethylating agents, 5-azacytidine and decitabine on growth and survival of neoplastic MCs and the MC line HMC-1. Two HMC-1 subclones were used, HMC-1.1 lacking KIT D816V and HMC-1.2 exhibiting KIT D816V. Both agents induced apoptosis in HMC-1.1 and HMC-1.2 cells. Decitabine, but not 5-azacytidine, also produced a G(2)/M cell-cycle arrest in HMC-1 cells. Drug-induced apoptosis was accompanied by cleavage of caspase-8 and caspase-3 as well as FAS-demethylation and FAS-re-expression in neoplastic MCs. Furthermore, both demethylating agents were found to synergize with the FAS-ligand in inducing apoptosis in neoplastic MCs. Correspondingly, siRNA against FAS was found to block drug-induced expression of FAS and drug-induced apoptosis in HMC-1 cells. Neither 5-azacytidine nor decitabine induced substantial apoptosis or growth arrest in normal MCs or normal bone marrow cells. Together, 5-azacytidine and decitabine exert growth-inhibitory and proapoptotic effects in neoplastic MCs. These effects are mediated through "FAS-re-expression" and are augmented by the FAS-ligand. Whether epigenetic drugs produce antineoplastic effects in vivo in patients with ASM and MCL remains to be determined.

Background: In advanced systemic mastocytosis the response of neoplastic mast cells to conventional drugs is poor and the prognosis is bad. Current research is, therefore, attempting to identify novel drug targets in neoplastic mast cells. Polo-like kinase-1 is a serine/threonine kinase that plays an essential role in mitosis and has recently been introduced as a new target in myeloid leukemias and solid tumors., Design and Methods: In the present study, we analyzed the expression and function of Polo-like kinase-1 in neoplastic mast cells in systemic mastocytosis., Results: As determined by immunostaining, primary neoplastic mast cells as well as the human mast cell leukemia cell line HMC-1 displayed phosphorylated Polo-like kinase-1. In addition, neoplastic mast cells expressed Polo-like kinase-1 mRNA. Polo-like kinase-1-specific small interfering RNA induced apoptosis in neoplastic mast cells, whereas no effect was seen with a control small interfering RNA. BI 2536, a drug targeting Polo-like kinase-1, was found to inhibit proliferation in HMC-1 cells in a dose-dependent manner. BI 2536 also inhibited the growth of primary neoplastic mast cells and cells of the canine mastocytoma cell line C2. The growth-inhibitory effects of BI 2536 on neoplastic mast cells were found to be associated with mitotic arrest and subsequent apoptosis. Finally, BI 2536 was found to synergize with the KIT-targeting kinase inhibitor midostaurin (PKC412) in inhibiting the growth of neoplastic mast cells. In control experiments, BI 2536 did not induce apoptosis in normal cultured mast cells., Conclusions: Collectively, our data show that Polo-like kinase-1 is a potential therapeutic target in neoplastic mast cells. Targeting Polo-like kinase-1 may be an attractive pharmacological concept in the management of advanced systemic mastocytosis.

Objective: In most patients with systemic mastocytosis (SM), including aggressive SM (ASM) and mast cell (MC) leukemia (MCL), neoplastic cells express the oncogenic KIT mutation D816V, which confers resistance to imatinib. Cladribine (2CdA) is a nucleoside analog that has been introduced as a promising agent for treatment of advanced SM., Materials and Methods: We examined the in vitro effects of 2CdA on growth of neoplastic MC, and the in vivo effects of 2CdA (0.13 mg/kg/day intravenously, days 1-5; three to eight cycles) in seven patients with advanced SM., Results: Cladribine was found to inhibit growth of primary MC and the MC line HMC-1 in a dose-dependent manner, with lower IC(50) values recorded in HMC-1.2 cells harboring KIT D816V (IC(50): 10 ng/mL) compared to HMC-1.1 cells lacking KIT D816V (IC(50): 300 ng/mL). In two patients with progressive smoldering SM, 2CdA produced a long-lasting response with a sustained decrease in serum tryptase levels, whereas in patients with progressive ASM or MCL, 2CdA showed little if any effects. The drug was well-tolerated in most cases. However, one patient developed a massive generalized purulent long-lasting skin rash. The antiproliferative effects of 2CdA on MC were found to be associated with morphologic signs of apoptosis and caspase cleavage. Cladribine did not counteract the kinase activity of KIT D816V or KIT-downstream signaling molecules., Conclusions: Cladribine may be a promising agent for treatment of progressive smoldering KIT D816V(+) SM. In rapidly progressing ASM or MCL, additional or alternative drugs are required to induce long-lasting antineoplastic effects.