Murine transgenic cells lacking DNA topoisomerase IIbeta are resistant to acridines and mitoxantrone: analysis of cytotoxicity and cleavable complex formation.
Murine transgenic cell lines lacking DNA topoisomerase II (topo II)beta have been used to assess the importance of topo IIbeta as a drug target. Western blot analysis confirmed that the topo IIbeta -/- cell lines did not contain topo IIbeta protein. In addition, both the topo IIbeta +/+ and topo IIbeta -/- cell lines contained similar levels of topo IIalpha protein. The trapped in agarose DNA immunostaining assay (TARDIS) was used to detect topo IIalpha and beta cleavable complexes in topo IIbeta -/- and topo IIbeta +/+ cells. These results show that both topo IIalpha and beta are in vivo targets for etoposide, mitoxantrone, and amsacrine (mAMSA) in topo IIbeta +/+ cells. As expected, only the alpha-isoform was targeted in topo IIbeta -/- cells. Clonogenic assays comparing the survival of topo IIbeta -/- and topo IIbeta +/+ cells were carried out to establish whether the absence of topo IIbeta caused drug resistance. Increased survival of topo IIbeta -/- cells compared with topo IIbeta +/+ cells was observed after treatment with amsacrine (mAMSA), methyl N-(4'-[9-acridinylamino]-2-methoxyphenyl) carbamate hydrochloride (AMCA), methyl N-(4'-[9-acridinylamino]-2-methoxyphenyl)carbamate hydrochloride (mAMCA), mitoxantrone, and etoposide. These studies showed that topo IIbeta -/- cells were significantly more resistant to mAMSA, AMCA, mAMCA, and mitoxantrone, than topo IIbeta +/+ cells, indicating that topo IIbeta is an important target for the cytotoxic effects of these compounds.
Errington
F
F
School of Biochemistry and Genetics, The Medical School, The Cookson Building, University of Newcastle upon Tyne, United Kingdom.
Willmore
E
E
Tilby
M J
MJ
Li
L
L
Li
G
G
Li
W
W
Baguley
B C
BC
Austin
C A
CA
D000166:Acridines / Q000494:pharmacology*; D000677:Amsacrine / Q000494:pharmacology; D000818:Animals; D000951:Antigens, Neoplasm; D000970:Antineoplastic Agents / Q000494:pharmacology; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D004250:DNA Topoisomerases, Type II / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins; D005047:Etoposide / Q000494:pharmacology; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008822:Mice, Transgenic; D008942:Mitoxantrone / Q000494:pharmacology*; D059005:Topoisomerase II Inhibitors
D000166:Acridines; D000951:Antigens, Neoplasm; D000970:Antineoplastic Agents; D004268:DNA-Binding Proteins; D007527:Isoenzymes; D059005:Topoisomerase II Inhibitors; D000677:Amsacrine; D005047:Etoposide; D008942:Mitoxantrone; D004250:DNA Topoisomerases, Type II