Comparative studies between the effects of mitozolomide and two novel tetrazepinones PYRCL and QUINCL on NIH:OVCAR-3 cells.
Cytotoxicity, reduction of macromolecule synthesis and cell cycle perturbations by two novel 3-(2-chloroethyl)-tetrazepinones, PYRCL and QUINCL were compared with those produced by the structurally related 3-(2-chloroethyl)-tetrazinone, mitozolomide, in the OVCAR-3 cell line.


METHODS
Macromolecule synthesis was determined by incorporation of 3H-thymidine, 3H-uridine and 3H-leucine into acid-precipitable fractions of OVCAR-3 cell extracts. Maxam-Gilbert sequencing was used to compare the DNA alkylating sites induced by the tetrazepinones, with those created by mitozolomide. Alkaline sucrose-density sedimentation was employed to detect genomic DNA damage. Also, the effects of the tetrazepinones on the cell cycle were determined by univariate flow cytometry.


RESULTS
At 3 h post-treatment, mitozolomide appeared as a selective inhibitor of DNA synthesis, while both tetrazepinones inhibited the synthesis of all three macromolecules. At 24 h post-treatment, the inhibition of DNA synthesis was observed to increase in cells treated with mitozolomide, while it decreased in those previously exposed to the tetrazepinones. Also at 24 h post-treatment, mitozolomide induced accumulation of cells in S(late)/G2M at low concentrations and in S-middle at high concentrations. In contrast, at the same recovery time, cells treated with the tetrazepinones accumulated specifically in G2M, the strength of the block being dose-dependent. At an equimolar concentration, the tetrazepinones induced weaker guanine N-7 alkylation than mitozolomide. By 24 h after treatment, cells exposed to the tetrazepinones showed significantly greater DNA fragmentation than those previously treated with mitozolomide.


CONCLUSIONS
In summary, based on (a) their effects on DNA, RNA, protein synthesis and on the cell cycle, (b) their alkylating power and (c) their interactions with DNA, the 3-(2-chloroethyl)tetrazepinones appeared to kill tumor cells by a novel mechanism which may significantly differ from that of their 3-(2-chloroethyl)-tetrazinone counterpart, mitozolomide.
Jean-Claude
B J
BJ
Department of Oncology, McGill University, Montreal, Quebec, Canada.
Mustafa
A
A
Damian
Z
Z
De Marte
J
J
Vasilescu
D E
DE
Yen
R
R
Chan
T H
TH
Leyland-Jones
B
B
D000478:Alkylation; D000970:Antineoplastic Agents / Q000494:pharmacology*; D001381:Azepines / Q000494:pharmacology*; D002453:Cell Cycle; D004247:DNA / Q000187:drug effects; D004249:DNA Damage; D004354:Drug Screening Assays, Antitumor; D005260:Female; D005434:Flow Cytometry; D006147:Guanine; D006801:Humans; D009588:Nitrogen Mustard Compounds / Q000494:pharmacology*; D010051:Ovarian Neoplasms / Q000188:drug therapy* / Q000473:pathology; D011725:Pyridines / Q000494:pharmacology*; D011804:Quinolines / Q000494:pharmacology*; D014407:Tumor Cells, Cultured
D000970:Antineoplastic Agents; D001381:Azepines; D009588:Nitrogen Mustard Compounds; C112464:PYRCL tetrazepinone; D011725:Pyridines; C112466:QUINCL tetrazepinone; D011804:Quinolines; D006147:Guanine; D004247:DNA; C040369:mitozolomide