Induction of differentiation and growth arrest associated with nascent (nonoligosomal) DNA fragmentation and reduced c-myc expression in MCF-7 human breast tumor cells after continuous exposure to a sublethal concentration of doxorubicin.
The effects on DNA integrity of continuous (72-h) exposure of human MCF-7 breast adenocarcinoma cells to 50 nM doxorubicin (a concentration which can be maintained in the plasma by continuous infusion) were characterized by bisbenzimide spectrofluorophotometry, cell flow cytometry, agarose gel electrophoresis, and neutral elution. Spectrofluorophotometry and cell flow cytometry indicated the presence of DNA fragmentation, which was maximal at 24 h. Resolution of these fragments on agarose gels failed to demonstrate "laddered" oligosomal profiles. Neutral elution analysis at 24 h indicated that doxorubicin induced fragmentation of nascent, but not mature, double-stranded DNA. Drug-treated cells exhibited endoreduplication and significant shifts in cell cycle distribution, (i.e., increased G0/G1 and G2/M fractions and a markedly reduced S-phase fraction). These alterations occurred without inhibiting the incorporation of [3H]dThd into cellular DNA; in fact, both the rate and magnitude of [3H]dThd incorporation increased progressively. Doxorubicin also produced a sustained decline in c-myc mRNA levels that paralleled both growth arrest and induction of DNA fragmentation. Ultrastructural examination revealed morphological alterations consistent with the induction of differentiation (e.g., increased lipid content and mitochondrial density, appearance of tight junctions, and secretory ducts) and further suggested the possibility of autocatalysis (e.g., lipofuschin-containing vacuoles). A gradual decline in cell number was observed, with loss of approximately 35% of the cell population after 72 h.(ABSTRACT TRUNCATED AT 250 WORDS)
Fornari
F A
FA
Department of Medicine, Medical College of Virginia, Richmond 23298.
Jarvis
W D
WD
Grant
S
S
Orr
M S
MS
Randolph
J K
JK
White
F K
FK
Mumaw
V R
VR
Lovings
E T
ET
Freeman
R H
RH
Gewirtz
D A
DA
D000230:Adenocarcinoma / Q000473:pathology*; D017209:Apoptosis; D001943:Breast Neoplasms / Q000473:pathology*; D002453:Cell Cycle / Q000187:drug effects; D002454:Cell Differentiation / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D004249:DNA Damage; D004273:DNA, Neoplasm / Q000187:drug effects*; D004317:Doxorubicin / Q000494:pharmacology*; D005260:Female; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects*; D016259:Genes, myc / Q000187:drug effects; D006801:Humans; D009363:Neoplasm Proteins / Q000096:biosynthesis* / Q000235:genetics; D016271:Proto-Oncogene Proteins c-myc / Q000096:biosynthesis* / Q000235:genetics; D014407:Tumor Cells, Cultured
D004273:DNA, Neoplasm; D009363:Neoplasm Proteins; D016271:Proto-Oncogene Proteins c-myc; D004317:Doxorubicin