Tumour‐cell apoptosis after cisplat in treatment is not telomere dependent

Abstract

Cisplatin is a major chemotherapeutic agent, especially for the treatment of neuroblastoma. Telomeres with their sequence (TTAGGG) n are probable targets for cisplatin intrastrand crosslinking, but the role of telomeres in mediating cisplatin cytotoxicity is not clear. After exposure to cisplatin as single dose or continuous treatment, we found no loss of telomeres in either SHSY5Y neuroblastoma cells (telomere length, 4 kbp), HeLa 229 cells (telomere length, 20 kbp) or in the acute lymphoblastic T cell line 1301 (telomere length, 80 kbp). There was no induction of telomeric single strand breaks, telomeric overhangs were not degraded and telomerase activity was down-regulated only after massive onset of apoptosis. In contrast, cisplatin induced a delayed formation of DNA strand breaks and induced DNA damage foci containing c-H2A.X at nontelomeric sites. Interstitial DNA damage appears to be more important than telomere loss or telomeric damage as inducer of the signal pathway towards apoptosis and/or growth arrest in cisplatin-treated tumour cells. ' 2005 Wiley-Liss, Inc.Key words: telomeres; telomerase; cisplatin; neuroblastoma; apoptosis; DNA damage Cisplatin is a chemotherapeutic agent for the treatment of neuroblastoma, a common solid extracranial malignancy of childhood and infancy. Cisplatin binds to DNA to cause a biological effect by forming cisplatin-DNA adducts and is a potent inducer of apoptosis.1,2 The major DNA adduct formed is the bifunctional intrastrand cross-link between adjacent guanines. It is unclear how the cisplatin-DNA adducts induce cytotoxicty, though this is widely attributed to the formation of various types of cross-links, especially intrastrand cross-links, 3 which are by far the most frequent lesions, 4 although interstrand cross-links might be more cytotoxic. 5Telomeres, the DNA-protein structures that form the ends of chromosomes, play an important role for the stability of the genome. Their shortening with each round of DNA replication is caused by different mechanisms, one of these being their sensitivity to DNA damage. 6 Cisplatin has been shown to bind preferentially to runs of 2 or more consecutive guanines.7 Human telomeric DNA contains triple runs of guanines at high density, therefore cisplatin may preferentially target telomeric DNA. Telomeres end in single stranded overhangs of the G-rich strand, which appear to be essential for telomeric higher order structure 8 and for the generation of DNA damage signals from telomeres.9-11 Telomerase, a reverse transcriptase that uses an RNA template to add telomeric repeats onto the ends of chromosomes, 12,13 is expressed in 94% of neuroblastomas 14 and has been shown to be a powerful independent prognostic factor.15 Thus a specific role of the telomere/telomerase complex in mediating cisplatin-induced neuroblastoma cell death may be expected and has been more or less directly indicated by some data, [16][17][18][19][20][21][22] but refuted by others. 23-25Given these contradictory results, we decided to examine whether there is a...

Modification of Cyclic NGR Tumor Neovasculature-Homing Motif Sequence to Human Plasminogen Kringle 5 Improves Inhibition of Tumor Growth

“…Other findings have also suggested that vascular targeting could increase vascular permeability, alter tumor barriers and increase the penetration of chemotherapeutic drugs [46], [47]. Cisplatin is widely used in the treatment of human tumors [48]. However, the potential for tumor control with cisplatin chemotherapy must always be carefully balanced with the risk for normal tissue damage [49], [50].…”

IP3 receptor antagonist, 2‐APB, attenuates cisplatin induced Ca2+‐influx in HeLa‐S3 cells and prevents activation of calpain and induction of apoptosis

“…Several authors have considered that DNA damage is the main cause of the anti‐proliferative effect of cisplatin ( Mandic et al ., 2003 ; Jeyapalan et al ., 2005 ). These authors also discussed different apoptotic, cisplatin‐triggered, pathways in inner ear hair cells: (1) reactive oxygen species and free radicals; (2) activation of caspases; and (3) activation of calpain.…”

Improved procedure for the measurement of telomere length in whole cells by PNA probe and flow cytometry

“…Figure 3b,c show, respectively, EB (total DNA) and Cy5 (telomeric DNA) fluorescences at nuclear level in the same cell. In 1301 cells, that had telomeres ranging from 25 to 80 kb (4,14), S phase events calculated from the telomeric DNA histogram are little more than half the events calculated from the total DNA histogram. 3d.…”

“…4, the more sam-ples had long telomeres the more distributions of total and telomeric DNA that appeared to be dissimilar. In 1301 cells, that had telomeres ranging from 25 to 80 kb (4,14), S phase events calculated from the telomeric DNA histogram are little more than half the events calculated from the total DNA histogram. Most S cells were converted into G 2 + M cells when the Cy5 histogram was deconvoluted, increasing G 2 + M phase from 6% to 19%.…”