GP7 induces internucleosomal DNA fragmentation independent of caspase activation and DNA fragmentation factor in NB4 cells

Abstract

DNA fragmentation into internucleosomal fragments is the best recognized biochemical event of apoptosis. Two major caspase pathways have been identified in the signal transduction leading to DNA fragmentation: the receptor pathway and the mitochondrial pathway. DNA fragmentation factor (DFF) has been identified as a major apoptotic endonuclease in the internucleosomal DNA fragmentation process. However, the potential roles of caspases and DFF in internucleosomal DNA fragmentation induced by specific stimuli still need to be investigated since caspaseindependent pathways and nuclease(s) other than DFF also play important roles during this process. In the present study, we investigated the activity of GP7 (4-[4"-(2",2",6",6"-tetramethyl-l"-piperidinyloxy) amino]-4'-demethyl epipodophyllotoxin), a new spin-labeled derivative of podophyllotoxin semi-synthesized by our university, to induce apoptosis of the human leukemia cell line NB4. GP7 induced the release of cytochrome-c from mitochondria, activations of caspase-3,-8, and-9, cleavage of DFF45/inhibitor of caspase-activated DNase, activation of DFF40/caspase-activated DNase, and apoptotic DNA fragmentation in NB4 cells. The broadspectrum caspase inhibitor zVAD-fmk abrogated GP7induced caspase-3,-8, and-9 activations but could not inhibit GP7-induced apoptotic DNA fragmentation in NB4 cells. Our findings suggest that GP7-induced apoptotic DNA fragmentation in NB4 cells is independent of caspase activation and DFF, although they are closely involved in this process.

Contemporary Challenges in the Design of Topoisomerase II Inhibitors for Cancer Chemotherapy

“…Indeed, it is known that selenite induces topoisomerase II–DNA complexes in cells, with a potential link to the induction of apoptosis Nitroxide-free radical to generate spin-labeled probes. − Different series of spin-labeled etoposides have been described, including compounds with immunosuppressive properties and, recently, molecules such as the antioxidant cytotoxic derivative 15 (Figure ) and GP-7 characterized as a proapoptotic agent capable of down-regulating PgP Aroylthiourea derivatives, able to interfere with both tyrosine kinases and topoisomerases.…”

“…Nitroxide-free radical to generate spin-labeled probes. − Different series of spin-labeled etoposides have been described, including compounds with immunosuppressive properties and, recently, molecules such as the antioxidant cytotoxic derivative 15 (Figure ) and GP-7 characterized as a proapoptotic agent capable of down-regulating PgP…”

Recent Progress on C‐4‐Modified Podophyllotoxin Analogs as Potent Antitumor Agents

“…Qi et al. reported that GP‐7, a new spin‐labeled derivative of PPT, activated the caspase signaling pathway by releasing cytochrome c . In tumor necrosis factor α (TNF‐α) induced human aortic smooth muscle cells, deoxy‐PPT strongly inhibited matrix metalloproteinases (MMP‐9) expression and migration, as well as mRNA transcription of MMP‐9 gene expression and phosphorylation of ERK 1, ERK 2, p38, and JNK, as reported by Suh et al .…”

Fucoidan from Lessonia trabeculata Induces Apoptosis through Caspase Dependent and Caspase-Independent Activation in 4T1 Breast Adenocarcinoma In Vitro

“…The 4T1 cells (2 × 10 5 cells/well) treated with the IC 50 as described in (item 3.5) were washed and resuspended in lysis buffer (2 mM ethylenediaminetetraacetic acid, 10 mM Tris-HCl, 10 mM potassium chloride, 10 mM magnesium chloride) according to [ 41 , 42 , 43 , 44 ] with slight modifications. The pellet was resuspended in lysis buffer, 10% SDS, RNase (10 mg/mL), and Proteinase K (20 mg/mL) and incubated at 55 °C and 37 °C for 10 min.…”