STAT1: a modulator of chemotherapy-induced apoptosis.
The anthracyclines, such as doxorubicin, are widely used in the treatment of breast cancer. Previously, we showed that these drugs could activate the transcription factor, nuclear factor kappaB, in a DNA damage-dependent manner. We now show that these drugs can potentiate the activation of signal transducer and activator of transcription 1 (STAT1) in MDA-MB 435 breast cancer cells treated with IFN-gamma. We observed that key markers of STAT1 activation, including tyrosine 701 and serine 727 phosphorylation, were enhanced in the presence of doxorubicin. This potentiation resulted in enhanced nuclear localization of activated STAT1 and led to an increase in the nuclear binding of activated STAT complexes. The observed potentiation was specific for STAT1 and IFN-gamma, as no effects were observed with either STAT3 or STAT5. Furthermore, the type I IFNs (alpha and beta) had little or no effect. The observed effects on STAT1 phosphorylation have previously been linked with maximal transcriptional activation and apoptosis. Cell viability was assessed by crystal violet staining followed by analysis with CalcuSyn to determine combination index values, a measure of synergy. We confirmed that significant synergy existed between IFN-gamma and doxorubicin (combination index = 0.34) at doses lower than IC(50) values for this drug (0.67 micromol/L). In support of this, we observed that apoptotic cell death was also enhanced by measuring poly(ADP-ribose) polymerase and caspase-3 cleavage. Finally, suppression of STAT1 expression by small-interfering RNA resulted in a loss of synergistic apoptotic cell death compared with cells, where no suppression of STAT1 expression was attained with scrambled small-interfering RNA control. We conclude that doxorubicin potentiates STAT1 activation in response to IFN-gamma, and that this combination results in enhanced apoptosis in breast cancer cells.
Thomas
Michelle
M
Centre for Cancer Research, Queens University Belfast, Belfast City Hospital, Belfast, Northern Ireland.
Finnegan
Clodagh E
CE
Rogers
Katherine M-A
KM
Purcell
James W
JW
Trimble
Anne
A
Johnston
Patrick G
PG
Boland
Marion P
MP
D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D001483:Base Sequence; D001943:Breast Neoplasms / Q000473:pathology; D045744:Cell Line, Tumor; D017931:DNA Primers; D004268:DNA-Binding Proteins / Q000378:metabolism / Q000502:physiology*; D004317:Doxorubicin / Q000494:pharmacology; D024202:Electrophoretic Mobility Shift Assay; D006801:Humans; D008856:Microscopy, Fluorescence; D010766:Phosphorylation; D050794:STAT1 Transcription Factor; D015534:Trans-Activators / Q000378:metabolism / Q000502:physiology*
D000970:Antineoplastic Agents; D017931:DNA Primers; D004268:DNA-Binding Proteins; D050794:STAT1 Transcription Factor; C494079:STAT1 protein, human; D015534:Trans-Activators; D004317:Doxorubicin