Abstract
ICRF-187 is a bisdioxopiperazine anticancer drug that inhibits the catalytic activity of DNA topoisomerase (topo) II without stabilizing DNA-topoII cleavable complexes. To better understand the mechanisms of action of and resistance to topoII catalytic inhibitors, human leukemic CEM cells were selected for resistance to ICRF-187. The clones CEM/ICRF-8 and CEM/ICRF-18 are approximately 40- and 69-fold resistant to ICRF-187, and 12- and 67-fold cross-resistant to ICRF-193, respectively, but are sensitive to other topoII catalytic inhibitors (merbarone and aclarubicin), as well as collaterally sensitive to the DNA-topoII complex-stabilizing drug etoposide (VP-16). Both the number of VP-16- induced DNA-topoII complexes formed and the amount of in vitro topoII catalytic activity are enhanced in the drug-resistant cells. The ICRF-187-resistant clones contain ∼5-fold increase in topoIIα protein levels and ∼2.2-fold increase in topoIIα mRNA levels. Furthermore, CEM/ICRF-8 expresses ∼3.5-fold increase in topoIIα promoter activity, suggesting that up-regulation of topoIIα in this clone occurs at the transcriptional level. Treatment of the drug-resistant or -sensitive cells with equitoxic doses of merbarone or teniposide results in a G2/M arrest. In marked contrast, when treated with equitoxic ICRF-187 doses, the drug-resistant clones exhibit either a transient arrest or completely lack the G2/M checkpoint compared with the drug-sensitive cells. This aberrant cell cycle profile is associated with a 48-h delay in drug-induced apoptotic cell death, as revealed by fluorescent-end labeling of DNA and poly (ADP-ribose) polymerase cleavage. In summary, resistance to ICRF-187 in CEM cells is associated with increased levels of catalytically active topoIIα and altered G2/M checkpoint and apoptotic responses.
Footnotes
-
Send reprint requests to: William T. Beck, Ph.D., Department of Molecular Genetics (M/C 669), University of Illinois at Chicago College of Medicine, 900 S. Ashland Ave., Chicago, IL 60607-7173. E-mail: wtbeck{at}uic.edu.
-
This work was supported in part by research Grants CA40570 and CA30103 from the National Cancer Institute (to W.T.B.), a research fellowship from the American Medical Association and Research Foundation (to S.E.M.), Cancer Center of the University of Illinois at Chicago, and research Grant CA31566 from the American Lebanese Syrian Associated Charities (ALSAC; to S.C.R.). S.E.M. was the (1998) Florence A. Carter Fellow of the American Medical Association.
- Abbreviations:
- topo
- topoisomerase
- VM-26
- teniposide
- VP-16
- etoposide
- MTT
- 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium
- FBS
- fetal bovine serum
- PARP
- poly (ADP-ribose) polymerase
- PCR
- polymerase chain reaction
- RT
- reverse transcriptase
- BrdU
- 5-bromo-2-deoxyuridine
- FITC
- fluorescein isothiocyanate
- CPT
- camptothecin
- Received August 11, 1999.
- Accepted October 20, 1999.
- The American Society for Pharmacology and Experimental Therapeutics
MolPharm articles become freely available 12 months after publication, and remain freely available for 5 years.Non-open access articles that fall outside this five year window are available only to institutional subscribers and current ASPET members, or through the article purchase feature at the bottom of the page.
|
