TY  - JOUR
AU  - Vučićević, Ljubica
AU  - Misirkić Marjanović, Maja
AU  - Ćirić, Darko
AU  - Martinović, Tamara
AU  - Jovanović, Maja
AU  - Isaković, Aleksandra
AU  - Marković, Ivanka
AU  - Šaponjić, Jasna
AU  - Foretz, Marc
AU  - Rabanal-Ruiz, Yoana
AU  - Korolchuk, Viktor I.
AU  - Trajković, Vladimir
PY  - 2020
UR  - http://link.springer.com/10.1007/s00018-019-03356-2
UR  - https://radar.ibiss.bg.ac.rs/handle/123456789/3528
AB  - We investigated the role of autophagy, a controlled lysosomal degradation of cellular macromolecules and organelles, in glutamate excitotoxicity during nutrient deprivation in vitro. The incubation in low-glucose serum/amino acid-free cell culture medium synergized with glutamate in increasing AMP/ATP ratio and causing excitotoxic necrosis in SH-SY5Y human neuroblastoma cells. Glutamate suppressed starvation-triggered autophagy, as confirmed by diminished intracellular acidification, lower LC3 punctuation and LC3-I conversion to autophagosome-associated LC3-II, reduced expression of proautophagic beclin-1 and ATG5, increase of the selective autophagic target NBR1, and decreased number of autophagic vesicles. Similar results were observed in PC12 rat pheochromocytoma cells. Both glutamate-mediated excitotoxicity and autophagy inhibition in starved SH-SY5Y cells were reverted by NMDA antagonist memantine and mimicked by NMDA agonists D-aspartate and ibotenate. Glutamate reduced starvation-triggered phosphorylation of the energy sensor AMP-activated protein kinase (AMPK) without affecting the activity of mammalian target of rapamycin complex 1, a major negative regulator of autophagy. This was associated with reduced mRNA levels of autophagy transcriptional activators (FOXO3, ATF4) and molecules involved in autophagy initiation (ULK1, ATG13, FIP200), autophagosome nucleation/elongation (ATG14, beclin-1, ATG5), and autophagic cargo delivery to autophagosomes (SQSTM1). Glutamate-mediated transcriptional repression of autophagy was alleviated by overexpression of constitutively active AMPK. Genetic or pharmacological AMPK activation by AMPK overexpression or metformin, as well as genetic or pharmacological autophagy induction by TFEB overexpression or lithium chloride, reduced the sensitivity of nutrient-deprived SH-SY5Y cells to glutamate excitotoxicity. These data indicate that transcriptional inhibition of AMPK-dependent cytoprotective autophagy is involved in glutamate-mediated excitotoxicity during nutrient deprivation in vitro.
T2  - Cellular and Molecular Life Sciences
T1  - Transcriptional block of AMPK-induced autophagy promotes glutamate excitotoxicity in nutrient-deprived SH-SY5Y neuroblastoma cells.
VL  - 77
DO  - 10.1007/s00018-019-03356-2
SP  - 3383
EP  - 3399
ER  - 

@article{
author = "Vučićević, Ljubica and Misirkić Marjanović, Maja and Ćirić, Darko and Martinović, Tamara and Jovanović, Maja and Isaković, Aleksandra and Marković, Ivanka and Šaponjić, Jasna and Foretz, Marc and Rabanal-Ruiz, Yoana and Korolchuk, Viktor I. and Trajković, Vladimir",
year = "2020",
abstract = "We investigated the role of autophagy, a controlled lysosomal degradation of cellular macromolecules and organelles, in glutamate excitotoxicity during nutrient deprivation in vitro. The incubation in low-glucose serum/amino acid-free cell culture medium synergized with glutamate in increasing AMP/ATP ratio and causing excitotoxic necrosis in SH-SY5Y human neuroblastoma cells. Glutamate suppressed starvation-triggered autophagy, as confirmed by diminished intracellular acidification, lower LC3 punctuation and LC3-I conversion to autophagosome-associated LC3-II, reduced expression of proautophagic beclin-1 and ATG5, increase of the selective autophagic target NBR1, and decreased number of autophagic vesicles. Similar results were observed in PC12 rat pheochromocytoma cells. Both glutamate-mediated excitotoxicity and autophagy inhibition in starved SH-SY5Y cells were reverted by NMDA antagonist memantine and mimicked by NMDA agonists D-aspartate and ibotenate. Glutamate reduced starvation-triggered phosphorylation of the energy sensor AMP-activated protein kinase (AMPK) without affecting the activity of mammalian target of rapamycin complex 1, a major negative regulator of autophagy. This was associated with reduced mRNA levels of autophagy transcriptional activators (FOXO3, ATF4) and molecules involved in autophagy initiation (ULK1, ATG13, FIP200), autophagosome nucleation/elongation (ATG14, beclin-1, ATG5), and autophagic cargo delivery to autophagosomes (SQSTM1). Glutamate-mediated transcriptional repression of autophagy was alleviated by overexpression of constitutively active AMPK. Genetic or pharmacological AMPK activation by AMPK overexpression or metformin, as well as genetic or pharmacological autophagy induction by TFEB overexpression or lithium chloride, reduced the sensitivity of nutrient-deprived SH-SY5Y cells to glutamate excitotoxicity. These data indicate that transcriptional inhibition of AMPK-dependent cytoprotective autophagy is involved in glutamate-mediated excitotoxicity during nutrient deprivation in vitro.",
journal = "Cellular and Molecular Life Sciences",
title = "Transcriptional block of AMPK-induced autophagy promotes glutamate excitotoxicity in nutrient-deprived SH-SY5Y neuroblastoma cells.",
volume = "77",
doi = "10.1007/s00018-019-03356-2",
pages = "3383-3399"
}

Vučićević, L., Misirkić Marjanović, M., Ćirić, D., Martinović, T., Jovanović, M., Isaković, A., Marković, I., Šaponjić, J., Foretz, M., Rabanal-Ruiz, Y., Korolchuk, V. I.,& Trajković, V.. (2020). Transcriptional block of AMPK-induced autophagy promotes glutamate excitotoxicity in nutrient-deprived SH-SY5Y neuroblastoma cells.. in Cellular and Molecular Life Sciences, 77, 3383-3399.
https://doi.org/10.1007/s00018-019-03356-2

Vučićević L, Misirkić Marjanović M, Ćirić D, Martinović T, Jovanović M, Isaković A, Marković I, Šaponjić J, Foretz M, Rabanal-Ruiz Y, Korolchuk VI, Trajković V. Transcriptional block of AMPK-induced autophagy promotes glutamate excitotoxicity in nutrient-deprived SH-SY5Y neuroblastoma cells.. in Cellular and Molecular Life Sciences. 2020;77:3383-3399.
doi:10.1007/s00018-019-03356-2 .

Vučićević, Ljubica, Misirkić Marjanović, Maja, Ćirić, Darko, Martinović, Tamara, Jovanović, Maja, Isaković, Aleksandra, Marković, Ivanka, Šaponjić, Jasna, Foretz, Marc, Rabanal-Ruiz, Yoana, Korolchuk, Viktor I., Trajković, Vladimir, "Transcriptional block of AMPK-induced autophagy promotes glutamate excitotoxicity in nutrient-deprived SH-SY5Y neuroblastoma cells." in Cellular and Molecular Life Sciences, 77 (2020):3383-3399,
https://doi.org/10.1007/s00018-019-03356-2 . .

TY  - JOUR
AU  - Jovanović-Tucović, Maja
AU  - Harhaji-Trajković, Ljubica
AU  - Dulović, Marija
AU  - Tovilović-Kovačević, Gordana
AU  - Zogović, Nevena
AU  - Jeremić, Marija
AU  - Mandić, Miloš
AU  - Kostić, Vladimir
AU  - Trajković, Vladimir
AU  - Marković, Ivanka
PY  - 2019
UR  - https://www.sciencedirect.com/science/article/pii/S0014299919306296?via%3Dihub
UR  - https://radar.ibiss.bg.ac.rs/handle/123456789/3482
AB  - We investigated the interplay between the intracellular energy sensor AMP-activated protein kinase (AMPK), prosurvival kinase Akt, oxidative stress, and autophagy in the cytotoxicity of parkinsonian neurotoxin 1-methyl-4-phenyl piridinium (MPP+) towards SH-SY5Y human neuroblastoma cells. MPP+-mediated oxidative stress, mitochondrial depolarization, and apoptotic cell death were associated with rapid (within 2 h) activation of AMPK, its target Raptor, and prosurvival kinase Akt. Antioxidants N-acetylcysteine and butylated hydroxyanisole suppressed MPP+-induced cytotoxicity, AMPK, and Akt activation. A genetic or pharmacological inhibition of AMPK increased MPP+-triggered production of reactive oxygen species and cell death, and diminished Akt phosphorylation, while AMPK activation protected SH-SY5Y cells from MPP+. On the other hand, genetic or pharmacological inactivation of Akt stimulated MPP+-triggered oxidative stress and neurotoxicity, but did not affect AMPK activation. At later time-points (16-24 h), MPP+ inhibited the main autophagy repressor mammalian target of rapamycin, which coincided with the increase in the levels of autophagy marker microtubule-associated protein 1 light-chain 3B. MPP+ also increased the concentration of a selective autophagic target sequestosome-1/p62 and reduced the levels of lysosomal-associated membrane protein 1 and cytoplasmic acidification, suggesting that MPP+-induced autophagy was coupled with a decrease in autophagic flux. Nevertheless, further pharmacological inhibition of autophagy sensitized SH-SY5Y cells to MPP+-induced death. Antioxidants and AMPK knockdown reduced, whereas genetic inactivation of Akt potentiated neurotoxin-triggered autophagy. These results suggest that MPP+-induced oxidative stress stimulates AMPK, which protects SH-SY5Y cells through early activation of antioxidative Akt and late induction of cytoprotective autophagy.
T2  - European Journal of Pharmacology
T1  - AMP-activated protein kinase inhibits MPP+-induced oxidative stress and apoptotic death of SH-SY5Y cells through sequential stimulation of Akt and autophagy.
VL  - 863
DO  - 10.1016/j.ejphar.2019.172677
SP  - 172677
ER  - 

@article{
author = "Jovanović-Tucović, Maja and Harhaji-Trajković, Ljubica and Dulović, Marija and Tovilović-Kovačević, Gordana and Zogović, Nevena and Jeremić, Marija and Mandić, Miloš and Kostić, Vladimir and Trajković, Vladimir and Marković, Ivanka",
year = "2019",
abstract = "We investigated the interplay between the intracellular energy sensor AMP-activated protein kinase (AMPK), prosurvival kinase Akt, oxidative stress, and autophagy in the cytotoxicity of parkinsonian neurotoxin 1-methyl-4-phenyl piridinium (MPP+) towards SH-SY5Y human neuroblastoma cells. MPP+-mediated oxidative stress, mitochondrial depolarization, and apoptotic cell death were associated with rapid (within 2 h) activation of AMPK, its target Raptor, and prosurvival kinase Akt. Antioxidants N-acetylcysteine and butylated hydroxyanisole suppressed MPP+-induced cytotoxicity, AMPK, and Akt activation. A genetic or pharmacological inhibition of AMPK increased MPP+-triggered production of reactive oxygen species and cell death, and diminished Akt phosphorylation, while AMPK activation protected SH-SY5Y cells from MPP+. On the other hand, genetic or pharmacological inactivation of Akt stimulated MPP+-triggered oxidative stress and neurotoxicity, but did not affect AMPK activation. At later time-points (16-24 h), MPP+ inhibited the main autophagy repressor mammalian target of rapamycin, which coincided with the increase in the levels of autophagy marker microtubule-associated protein 1 light-chain 3B. MPP+ also increased the concentration of a selective autophagic target sequestosome-1/p62 and reduced the levels of lysosomal-associated membrane protein 1 and cytoplasmic acidification, suggesting that MPP+-induced autophagy was coupled with a decrease in autophagic flux. Nevertheless, further pharmacological inhibition of autophagy sensitized SH-SY5Y cells to MPP+-induced death. Antioxidants and AMPK knockdown reduced, whereas genetic inactivation of Akt potentiated neurotoxin-triggered autophagy. These results suggest that MPP+-induced oxidative stress stimulates AMPK, which protects SH-SY5Y cells through early activation of antioxidative Akt and late induction of cytoprotective autophagy.",
journal = "European Journal of Pharmacology",
title = "AMP-activated protein kinase inhibits MPP+-induced oxidative stress and apoptotic death of SH-SY5Y cells through sequential stimulation of Akt and autophagy.",
volume = "863",
doi = "10.1016/j.ejphar.2019.172677",
pages = "172677"
}

Jovanović-Tucović, M., Harhaji-Trajković, L., Dulović, M., Tovilović-Kovačević, G., Zogović, N., Jeremić, M., Mandić, M., Kostić, V., Trajković, V.,& Marković, I.. (2019). AMP-activated protein kinase inhibits MPP+-induced oxidative stress and apoptotic death of SH-SY5Y cells through sequential stimulation of Akt and autophagy.. in European Journal of Pharmacology, 863, 172677.
https://doi.org/10.1016/j.ejphar.2019.172677

Jovanović-Tucović M, Harhaji-Trajković L, Dulović M, Tovilović-Kovačević G, Zogović N, Jeremić M, Mandić M, Kostić V, Trajković V, Marković I. AMP-activated protein kinase inhibits MPP+-induced oxidative stress and apoptotic death of SH-SY5Y cells through sequential stimulation of Akt and autophagy.. in European Journal of Pharmacology. 2019;863:172677.
doi:10.1016/j.ejphar.2019.172677 .

Jovanović-Tucović, Maja, Harhaji-Trajković, Ljubica, Dulović, Marija, Tovilović-Kovačević, Gordana, Zogović, Nevena, Jeremić, Marija, Mandić, Miloš, Kostić, Vladimir, Trajković, Vladimir, Marković, Ivanka, "AMP-activated protein kinase inhibits MPP+-induced oxidative stress and apoptotic death of SH-SY5Y cells through sequential stimulation of Akt and autophagy." in European Journal of Pharmacology, 863 (2019):172677,
https://doi.org/10.1016/j.ejphar.2019.172677 . .

TY  - JOUR
AU  - Isaković, Anđelka M
AU  - Petričević, Sasa M.
AU  - Ristić, Slavica M.
AU  - Popadić, Dušan M.
AU  - Kravić-Stevović, Tamara K
AU  - Zogović, Nevena
AU  - Poljarević, Jelena M.
AU  - Živanović Radnić, Tatjana V
AU  - Sabo, Tibor J.
AU  - Isaković, Aleksandra J.
AU  - Marković, Ivanka D.
AU  - Trajković, Vladimir S.
AU  - Misirlić-Denčić, Sonja T
PY  - 2018
UR  - https://insights.ovid.com/crossref?an=00008390-201802000-00002
UR  - https://radar.ibiss.bg.ac.rs/handle/123456789/3004
AB  - Melanoma, an aggressive skin tumor with high metastatic potential, is associated with high mortality and increasing morbidity. Multiple available chemotherapeutic and immunotherapeutic modalities failed to improve survival in advanced disease, and the search for new agents is ongoing. The aim of this study was to investigate antimelanoma effects of O,O-diethyl-(S,S)-ethylenediamine-N,N'di-2-(3-cyclohexyl) propanoate dihydrochloride (EE), a previously synthesized and characterized organic compound. Mouse melanoma B16 cell viability was assessed using acid phosphatase, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, sulforhodamine B, and lactate dehydrogenase assays. Apoptosis and autophagy were investigated using flow cytometry, fluorescence and electron microscopy, and western blotting. In vivo antitumor potential was assessed in subcutaneous mouse melanoma model after 14 days of treatment with EE. Tumor mass and volume were measured, and RT-PCR was used for investigating the expression of autophagy-related, proapoptotic, and antiapoptotic molecules in tumor tissue. Investigated organic compound exerts significant cytotoxic effect against B16 cells. EE induced apoptosis, as confirmed by phosphatidyl serine externalisation, caspase activation, and ultrastructural features typical for apoptosis seen on fluorescence and electron microscopes. The apoptotic mechanism included prompt disruption of mitochondrial membrane potential and oxidative stress. No autophagy was observed. Antimelanoma action and apoptosis induction were confirmed in vivo, as EE decreased mass and volume of tumors, and increased expression of several proapoptotic genes. EE possesses significant antimelanoma action and causes caspase-dependent apoptosis mediated by mitochondrial damage and reactive oxygen species production. Decrease in tumor growth and increase in expression of proapoptotic genes in tumor tissue suggest that EE warrants further investigation as a candidate agent in treating melanoma.
PB  - Melanoma Research
T2  - Melanoma Research
T2  - Melanoma Research
T1  - In vitro and in vivo antimelanoma effect of ethyl ester cyclohexyl analog of ethylenediamine dipropanoic acid.
IS  - 1
VL  - 28
DO  - 10.1097/CMR.0000000000000409
SP  - 8
EP  - 20
ER  - 

@article{
author = "Isaković, Anđelka M and Petričević, Sasa M. and Ristić, Slavica M. and Popadić, Dušan M. and Kravić-Stevović, Tamara K and Zogović, Nevena and Poljarević, Jelena M. and Živanović Radnić, Tatjana V and Sabo, Tibor J. and Isaković, Aleksandra J. and Marković, Ivanka D. and Trajković, Vladimir S. and Misirlić-Denčić, Sonja T",
year = "2018",
abstract = "Melanoma, an aggressive skin tumor with high metastatic potential, is associated with high mortality and increasing morbidity. Multiple available chemotherapeutic and immunotherapeutic modalities failed to improve survival in advanced disease, and the search for new agents is ongoing. The aim of this study was to investigate antimelanoma effects of O,O-diethyl-(S,S)-ethylenediamine-N,N'di-2-(3-cyclohexyl) propanoate dihydrochloride (EE), a previously synthesized and characterized organic compound. Mouse melanoma B16 cell viability was assessed using acid phosphatase, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, sulforhodamine B, and lactate dehydrogenase assays. Apoptosis and autophagy were investigated using flow cytometry, fluorescence and electron microscopy, and western blotting. In vivo antitumor potential was assessed in subcutaneous mouse melanoma model after 14 days of treatment with EE. Tumor mass and volume were measured, and RT-PCR was used for investigating the expression of autophagy-related, proapoptotic, and antiapoptotic molecules in tumor tissue. Investigated organic compound exerts significant cytotoxic effect against B16 cells. EE induced apoptosis, as confirmed by phosphatidyl serine externalisation, caspase activation, and ultrastructural features typical for apoptosis seen on fluorescence and electron microscopes. The apoptotic mechanism included prompt disruption of mitochondrial membrane potential and oxidative stress. No autophagy was observed. Antimelanoma action and apoptosis induction were confirmed in vivo, as EE decreased mass and volume of tumors, and increased expression of several proapoptotic genes. EE possesses significant antimelanoma action and causes caspase-dependent apoptosis mediated by mitochondrial damage and reactive oxygen species production. Decrease in tumor growth and increase in expression of proapoptotic genes in tumor tissue suggest that EE warrants further investigation as a candidate agent in treating melanoma.",
publisher = "Melanoma Research",
journal = "Melanoma Research, Melanoma Research",
title = "In vitro and in vivo antimelanoma effect of ethyl ester cyclohexyl analog of ethylenediamine dipropanoic acid.",
number = "1",
volume = "28",
doi = "10.1097/CMR.0000000000000409",
pages = "8-20"
}

Isaković, A. M., Petričević, S. M., Ristić, S. M., Popadić, D. M., Kravić-Stevović, T. K., Zogović, N., Poljarević, J. M., Živanović Radnić, T. V., Sabo, T. J., Isaković, A. J., Marković, I. D., Trajković, V. S.,& Misirlić-Denčić, S. T.. (2018). In vitro and in vivo antimelanoma effect of ethyl ester cyclohexyl analog of ethylenediamine dipropanoic acid.. in Melanoma Research
Melanoma Research., 28(1), 8-20.
https://doi.org/10.1097/CMR.0000000000000409

Isaković AM, Petričević SM, Ristić SM, Popadić DM, Kravić-Stevović TK, Zogović N, Poljarević JM, Živanović Radnić TV, Sabo TJ, Isaković AJ, Marković ID, Trajković VS, Misirlić-Denčić ST. In vitro and in vivo antimelanoma effect of ethyl ester cyclohexyl analog of ethylenediamine dipropanoic acid.. in Melanoma Research. 2018;28(1):8-20.
doi:10.1097/CMR.0000000000000409 .

Isaković, Anđelka M, Petričević, Sasa M., Ristić, Slavica M., Popadić, Dušan M., Kravić-Stevović, Tamara K, Zogović, Nevena, Poljarević, Jelena M., Živanović Radnić, Tatjana V, Sabo, Tibor J., Isaković, Aleksandra J., Marković, Ivanka D., Trajković, Vladimir S., Misirlić-Denčić, Sonja T, "In vitro and in vivo antimelanoma effect of ethyl ester cyclohexyl analog of ethylenediamine dipropanoic acid." in Melanoma Research, 28, no. 1 (2018):8-20,
https://doi.org/10.1097/CMR.0000000000000409 . .

TY  - JOUR
AU  - Misirlić-Dencić, Sonja T
AU  - Poljarević, Jelena M
AU  - Vilimanović, Uros
AU  - Bogdanović, Andrija D
AU  - Isaković, Aleksandra J
AU  - Kravić-Stevović, Tamara K
AU  - Dulović, Marija
AU  - Zogović, Nevena
AU  - Isaković, Anđelka M
AU  - Grgurić-Sipka, Sanja R
AU  - Bumbaširević, Vladimir Z
AU  - Sabo, Tibor J
AU  - Trajković, Vladimir S
AU  - Marković, Ivanka D
PY  - 2012
UR  - https://radar.ibiss.bg.ac.rs/handle/123456789/1196
AB  - We investigated the cytotoxicity of recently synthesized (S,S)-ethyleridiamine-N,N'-di-2-(3-cyclohexyl)propanoic acid esters toward human leukemic cell lines and healthy blood mononuclear cells. Cell viability was assessed by acid phosphatase assay, apoptosis, and differentiation were analyzed by flow cytometry and electron microscopy, while intracellular localization of apoptosis-inducing factor (AIF) was determined by immunoblotting. It was demonstrated that methyl, ethyl, and n-propyl esters were toxic to HL-60, REH, MOLT-4, KG-1, JVM-2, and K-562 leukemic cell lines, while the nonesterified parental compound and n-butyl ester were devoid of cytotoxic action. The ethyl ester exhibited the highest cytotoxic activity (IC50 10.7 mu M-45.4 mu M), which was comparable to that of the prototypical anticancer drug cisplatin. The observed cytotoxic effect in HL-60 cells was associated with an increase in superoxide production and mitochondrial membrane depolarization, leading to apoptotic cell death characterized by phosphatidylserine externalization and DNA fragmentation in the absence of autophagic response. DNA fragmentation preceded caspase activation and followed AIF translocation from mitochondria to nucleus, which was indicative of caspase-independent apoptotic cell death. HL-60 cells treated with subtoxic concentration of the compound displayed morphological signs of granulocytic differentiation (nuclear indentations and presence of cytoplasmic primary granules), as well as an increased expression of differentiation markers CD11b and CD15. The cyclohexyl analogues of ethylenediamine dipropanoic acid were also toxic to peripheral blood mononuclear cells of both healthy controls and leukemic patients, the latter being more sensitive. Our data demonstrate that the toxicity of the investigated cyclohexyl compounds against leukemic cell lines is mediated by caspase-independent apoptosis associated with oxidative stress, mitochondrial dysfunction, and AIF translocation.
T2  - Chemical Research in Toxicology
T1  - Cyclohexyl Analogues of Ethylenediamine Dipropanoic Acid Induce Caspase-Independent Mitochondrial Apoptosis in Human Leukemic Cells
IS  - 4
VL  - 25
EP  - 939
UR  - https://hdl.handle.net/21.15107/rcub_ibiss_1196
ER  - 

@article{
author = "Misirlić-Dencić, Sonja T and Poljarević, Jelena M and Vilimanović, Uros and Bogdanović, Andrija D and Isaković, Aleksandra J and Kravić-Stevović, Tamara K and Dulović, Marija and Zogović, Nevena and Isaković, Anđelka M and Grgurić-Sipka, Sanja R and Bumbaširević, Vladimir Z and Sabo, Tibor J and Trajković, Vladimir S and Marković, Ivanka D",
year = "2012",
abstract = "We investigated the cytotoxicity of recently synthesized (S,S)-ethyleridiamine-N,N'-di-2-(3-cyclohexyl)propanoic acid esters toward human leukemic cell lines and healthy blood mononuclear cells. Cell viability was assessed by acid phosphatase assay, apoptosis, and differentiation were analyzed by flow cytometry and electron microscopy, while intracellular localization of apoptosis-inducing factor (AIF) was determined by immunoblotting. It was demonstrated that methyl, ethyl, and n-propyl esters were toxic to HL-60, REH, MOLT-4, KG-1, JVM-2, and K-562 leukemic cell lines, while the nonesterified parental compound and n-butyl ester were devoid of cytotoxic action. The ethyl ester exhibited the highest cytotoxic activity (IC50 10.7 mu M-45.4 mu M), which was comparable to that of the prototypical anticancer drug cisplatin. The observed cytotoxic effect in HL-60 cells was associated with an increase in superoxide production and mitochondrial membrane depolarization, leading to apoptotic cell death characterized by phosphatidylserine externalization and DNA fragmentation in the absence of autophagic response. DNA fragmentation preceded caspase activation and followed AIF translocation from mitochondria to nucleus, which was indicative of caspase-independent apoptotic cell death. HL-60 cells treated with subtoxic concentration of the compound displayed morphological signs of granulocytic differentiation (nuclear indentations and presence of cytoplasmic primary granules), as well as an increased expression of differentiation markers CD11b and CD15. The cyclohexyl analogues of ethylenediamine dipropanoic acid were also toxic to peripheral blood mononuclear cells of both healthy controls and leukemic patients, the latter being more sensitive. Our data demonstrate that the toxicity of the investigated cyclohexyl compounds against leukemic cell lines is mediated by caspase-independent apoptosis associated with oxidative stress, mitochondrial dysfunction, and AIF translocation.",
journal = "Chemical Research in Toxicology",
title = "Cyclohexyl Analogues of Ethylenediamine Dipropanoic Acid Induce Caspase-Independent Mitochondrial Apoptosis in Human Leukemic Cells",
number = "4",
volume = "25",
pages = "939",
url = "https://hdl.handle.net/21.15107/rcub_ibiss_1196"
}

Misirlić-Dencić, S. T., Poljarević, J. M., Vilimanović, U., Bogdanović, A. D., Isaković, A. J., Kravić-Stevović, T. K., Dulović, M., Zogović, N., Isaković, A. M., Grgurić-Sipka, S. R., Bumbaširević, V. Z., Sabo, T. J., Trajković, V. S.,& Marković, I. D.. (2012). Cyclohexyl Analogues of Ethylenediamine Dipropanoic Acid Induce Caspase-Independent Mitochondrial Apoptosis in Human Leukemic Cells. in Chemical Research in Toxicology, 25(4).
https://hdl.handle.net/21.15107/rcub_ibiss_1196

Misirlić-Dencić ST, Poljarević JM, Vilimanović U, Bogdanović AD, Isaković AJ, Kravić-Stevović TK, Dulović M, Zogović N, Isaković AM, Grgurić-Sipka SR, Bumbaširević VZ, Sabo TJ, Trajković VS, Marković ID. Cyclohexyl Analogues of Ethylenediamine Dipropanoic Acid Induce Caspase-Independent Mitochondrial Apoptosis in Human Leukemic Cells. in Chemical Research in Toxicology. 2012;25(4):null-939.
https://hdl.handle.net/21.15107/rcub_ibiss_1196 .

Misirlić-Dencić, Sonja T, Poljarević, Jelena M, Vilimanović, Uros, Bogdanović, Andrija D, Isaković, Aleksandra J, Kravić-Stevović, Tamara K, Dulović, Marija, Zogović, Nevena, Isaković, Anđelka M, Grgurić-Sipka, Sanja R, Bumbaširević, Vladimir Z, Sabo, Tibor J, Trajković, Vladimir S, Marković, Ivanka D, "Cyclohexyl Analogues of Ethylenediamine Dipropanoic Acid Induce Caspase-Independent Mitochondrial Apoptosis in Human Leukemic Cells" in Chemical Research in Toxicology, 25, no. 4 (2012),
https://hdl.handle.net/21.15107/rcub_ibiss_1196 .