Deregulated sequential motion of centromeres induced by antitumor agents may lead to genome instability in human peripheral blood lymphocytes.
OBJECTIVE
Segregation of chromosomes in anaphase is preceded by a sequential order of centromere separation. Alteration of the sequence of centromere separation or premature centromere division (PCD) has been found to be significantly higher in populations exposed to various xenobiotics. The purpose of this study was to investigate if PCD induced by various cytostatics can alter the stability of chromosomes and lead to aneuploidy.


METHODS
Peripheral blood lymphocytes of 10 healthy, non smoking subjects were exposed to 8-Cl-cAMP at a dose of 1, 5 and 15 microM, paclitaxel at a dose of 0.01, 0.05 and 0.2 microM, and cycloheximide (CX) at a dose of 5, 10 and 25 microg/ml. By using the cytohalasin B (CB)-micronucleus (MN) test in vitro, in combination with fluorescent in situ hybridization (FISH), the presence of MN was analyzed in 1000 binuclear cells for each experimental and negative control group. For analysis of MN content we used the alpha-centromeric probe for chromosome 18.


RESULTS
8-Cl-cAMP and paclitaxel induced an increase in the frequency of MN in peripheral blood lymphocytes. 8-Cl-cAMP and paclitaxel proved clastogenic, i.e. they increased the frequency of MN and induced PCD in all respective doses. CX proved not clastogenic in the respected doses when using the CB-MN test in vitro, although CX is a specific PCD inducer. No correlation of PCD and aneuploidy of chromosome 18 was found in cells exposed to 8-Cl-cAMP and paclitaxel by using FISH. In cells exposed to CX we found PCD of chromosome 18 in binuclear cells and single signals in scarce MN. These findings were not statistically significant compared to the negative control group.


CONCLUSIONS
Our results show that the properties of the investigated antitumor agents to induce PCD in peripheral blood lymphocytes and, therefore, aneuploidy and genome instability, is highly based on the nature of the alteration of centromere function, i.e. the temporal order of centromere kinetics are more regulated through the sequence of centromere separation than by the segregation processes. We suggest that PCD induced by novel antitumor agents could be included in preclinical and clinical genetic risk assessment analysis.
Bajic
V
V
Institute for Biomedical Research, Galenika a.d., Belgrade, Serbia. vladanpfd@eunet.yu
Spremo-Potparevic
B
B
Milicevic
Z
Z
Zivkovic
L
L
D015124:8-Bromo Cyclic Adenosine Monophosphate / Q000031:analogs & derivatives / Q000633:toxicity; D000328:Adult; D000782:Aneuploidy; D000970:Antineoplastic Agents / Q000633:toxicity*; D002455:Cell Division / Q000187:drug effects / Q000235:genetics; D002478:Cells, Cultured; D002503:Centromere / Q000187:drug effects* / Q000378:metabolism; D043171:Chromosomal Instability / Q000187:drug effects*; D020090:Chromosome Segregation / Q000187:drug effects*; D002887:Chromosomes, Human, Pair 18 / Q000187:drug effects*; D003513:Cycloheximide / Q000633:toxicity; D004305:Dose-Response Relationship, Drug; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D008214:Lymphocytes / Q000187:drug effects* / Q000378:metabolism; D008297:Male; D015162:Micronucleus Tests; D009153:Mutagens / Q000633:toxicity*; D017239:Paclitaxel / Q000633:toxicity
D000970:Antineoplastic Agents; D009153:Mutagens; D015124:8-Bromo Cyclic Adenosine Monophosphate; D003513:Cycloheximide; C044258:8-chloro-cyclic adenosine monophosphate; D017239:Paclitaxel