Extracellular ATP recently emerged as constituent of tumor microenvironment modulating tumor cell growth and progression (Adinolfi E. et al., 2015) Immunogenic cell death (ICD) is a form of cancer cell death caused by chemotherapeutic agents such as anthracyclines that has been associated to extracellular ATP (eATP) release (Martins I. et al., 2014). ATP release from solid cancer dying cells induces an antitumor immune response (Michaud M. et al., 2011). Although anthracyclines such as daunorubicine are widely used as chemotherapy in leukemia, nothing is known about ICD in these diseases. Here we analyzed the effect of two different drugs i.e. daunorubicine and ara-C, which are known respectively to induce or not ICD on eATP release in a model of leukemia. eATP release was determined thanks to the use of plasma membrane luciferase (PmeLUC), a luciferase-derived probe that was engineered to be expressed on the outer facet of the plasma membrane and emits photons in an ATP dependent-manner (Pellegatti P. et al., 2005, 2008). WEHI-3B mouse leukemia cells were transfected with PmeLUC and stable clones were obtained (WEHI-3B PmeLUC). In a first set of in vitro experiments WEHI-3B PmeLUC were treated with either vehicle, 100 ng/ml daunorubicine and 15 μg/ml ara-C for 4, 24 and 48 hours and luminescence emission was monitored by luminometer. After 48 hours incubation the levels of eATP were higher in the daunorubicine group than in the vehicle and ara-C treated cells. Nevertheless cell proliferation was reduced by both chemotherapeutics. In a second set of in vivo experiments WEHI-3B PmeLUC were injected in syngeneic BALBCJ mice and luminescence emission was in vivo monitored thanks to an IVIS lumina total body luminometer. After a first tumor mass detection placebo, daunorubicine and ara-C (600 μg/ml and 10 mg/ml respectively) were administered to mice every other day for twelve days. Systemic treatment with both drugs caused a significant reduction of tumor growth. However eATP levels were more than doubled in the daunorubicine treated group compared to placebo or ara-C treated mice. These data suggest that eATP levels can be specifically modulated by anthracyclines in leukemia such as in solid tumors, giving a first indication that ICD could be occurring also in these lympho-proliferative disorders.
Extracellular ATP modulation by daunorubicin in a murine leukemia model
DE MARCHI, Elena;ORIOLI, Elisa;DI VIRGILIO, Francesco;ADINOLFI, Elena
2015
Abstract
Extracellular ATP recently emerged as constituent of tumor microenvironment modulating tumor cell growth and progression (Adinolfi E. et al., 2015) Immunogenic cell death (ICD) is a form of cancer cell death caused by chemotherapeutic agents such as anthracyclines that has been associated to extracellular ATP (eATP) release (Martins I. et al., 2014). ATP release from solid cancer dying cells induces an antitumor immune response (Michaud M. et al., 2011). Although anthracyclines such as daunorubicine are widely used as chemotherapy in leukemia, nothing is known about ICD in these diseases. Here we analyzed the effect of two different drugs i.e. daunorubicine and ara-C, which are known respectively to induce or not ICD on eATP release in a model of leukemia. eATP release was determined thanks to the use of plasma membrane luciferase (PmeLUC), a luciferase-derived probe that was engineered to be expressed on the outer facet of the plasma membrane and emits photons in an ATP dependent-manner (Pellegatti P. et al., 2005, 2008). WEHI-3B mouse leukemia cells were transfected with PmeLUC and stable clones were obtained (WEHI-3B PmeLUC). In a first set of in vitro experiments WEHI-3B PmeLUC were treated with either vehicle, 100 ng/ml daunorubicine and 15 μg/ml ara-C for 4, 24 and 48 hours and luminescence emission was monitored by luminometer. After 48 hours incubation the levels of eATP were higher in the daunorubicine group than in the vehicle and ara-C treated cells. Nevertheless cell proliferation was reduced by both chemotherapeutics. In a second set of in vivo experiments WEHI-3B PmeLUC were injected in syngeneic BALBCJ mice and luminescence emission was in vivo monitored thanks to an IVIS lumina total body luminometer. After a first tumor mass detection placebo, daunorubicine and ara-C (600 μg/ml and 10 mg/ml respectively) were administered to mice every other day for twelve days. Systemic treatment with both drugs caused a significant reduction of tumor growth. However eATP levels were more than doubled in the daunorubicine treated group compared to placebo or ara-C treated mice. These data suggest that eATP levels can be specifically modulated by anthracyclines in leukemia such as in solid tumors, giving a first indication that ICD could be occurring also in these lympho-proliferative disorders.I documenti in SFERA sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
