Abstract:
In this study, we attempt to target the mitogen-activated protein kinase (MAPK) pathway in acute myeloid leukemia (AML) cells using a recombinant anthrax lethal toxin (LeTx). Around 15,000 new case of Acute Myeloid Leukemia are diagnosed each year with a fatality rate of 65%. The poor prognosis rate is due to the high proliferative and quick progressive characteristics of AML. However, most AML patients eventually relapse due to persistence of chemotherapy-resistant blasts in the bone marrow hence the need for alternative approaches employing novel, more selective mechanisms for targeting AML blasts. One such approach consists of targeting the mitogen-activated protein (MAP) kinase (MAPK) pathway in AML cells. LeTx consists of protective antigen (PrAg) and lethal factor (LF). PrAg binds cells, is cleaved by furin, oligomerizes, binds three to four molecules of LF, and undergoes endocytosis, releasing LF into the cytosol. LF cleaves MAPK kinases, inhibiting the MAPK pathway. The MAKP pathway is a conserved pathway between eukaryotes. Through a wide range of extracellular signals, the MAPK pathway regulates growth, proliferation, differentiation and death. Its constitutive activation, particularly the Ras/Raf/MEK1/2/ERK1/2 cascade promotes proliferation and survival of most human cancer cells. We tested potency of LeTx on a panel of 11 human AML cell lines. Seven cell lines showed cytotoxic responses to LeTx. Cytotoxicity of LeTx was mimicked by the specific mitogen-activated protein/extracellular signal–regulated kinase kinase 1/2 (MEK1/2) inhibitor U0126, indicating that LeTx-induced cell death is mediated through the MEK1/2–extracellular signal–regulated kinase (ERK1/2) branch of the MAPK pathway. The four LeTx-resistant cell lines were sensitive to the phosphatidylinositol 3-kinase inhibitor LY294002. Flow cytometry analysis of MAPK pathway activation revealed presence of phospho-ERK1/2 only in LeTx-sensitive cells. In this study, we have shown that a majority of AML cell lines are sensitive to the LF-mediated inhibition of the MAPK pathway. Furthermore, we have demonstrated that LeTx-induced cytotoxicity in AML cells is non-apoptotic and dependent on phospho-ERK1/2 levels.
