Defining direct targets of transcription factors and regulatory pathways is key to understanding their roles in physiology and disease. We combined SLAM-seq thiol(SH)-linked alkylation for the ...metabolic sequencing of RNA, a method for direct quantification of newly synthesized messenger RNAs (mRNAs), with pharmacological and chemical-genetic perturbation in order to define regulatory functions of two transcriptional hubs in cancer, BRD4 and MYC, and to interrogate direct responses to BET bromodomain inhibitors (BETis). We found that BRD4 acts as general coactivator of RNA polymerase II-dependent transcription, which is broadly repressed upon high-dose BETi treatment. At doses triggering selective effects in leukemia, BETis deregulate a small set of hypersensitive targets including MYC. In contrast to BRD4, MYC primarily acts as a selective transcriptional activator controlling metabolic processes such as ribosome biogenesis and de novo purine synthesis. Our study establishes a simple and scalable strategy to identify direct transcriptional targets of any gene or pathway.
Objective.Utilizing ovarian cancer cell lines, we examined the effect of IFN-γ on each type of TNF receptor. Additionally, we sought to determine the effect of receptor modulation on TNF-α-mediated ...cytolysis.
Methods.Ovarian cancer cell lines Caov-3, A2780, and SK-OV-3 were employed. The number of TNF receptors was determined by a TNF-α binding assay utilizing125I-labeled TNF-α. Monoclonal antibodies specific for the 55- to 60-kDa (TR60) and the 75- to 80-kDa (TR80) TNF receptors were used to determine the relative density of each receptor type. Northern blot analyses were performed employing cDNA probes for the TR60 and TR80 mRNAs. To elucidate which receptor(s) was responsible for mediating the signal for cytolysis, 24-h MTT cytolytic assays were performed in the presence of receptor-specific monoclonal antibodies.
Results.IFN-γ treatment resulted in an increase in TNF receptors in the cell lines A2780 and Caov-3 (P< 0.001), but not SK-OV-3. Northern blot analyses suggested distinct regulatory mechanisms for the two receptors. In Caov-3 and SK-OV-3 cells a synergistic increase in TNF-α-mediated cytolysis was seen when cells were pretreated with IFN-γ. In both cell lines, pretreatment with IFN-γ markedly enhanced the ability of the TR60 receptor to mediate cell lysis. Conversely, under similar treatment conditions, the TR80 receptor did not appear capable of generating a cytolytic signal.
Conclusions.TNF receptor modulation by IFN-γ appears to be unique to individual cell lines. The TR60 TNF receptor plays a central role in the synergistic cytolytic effects of IFN-γ and TNF-α. Sequential therapy with IFN-γ and TNF-α and specific TNF receptor activation may provide novel translational strategies for the use of cytokines in the treatment of ovarian cancer.
OBJECTIVE:
To evaluate utility and cost-effectiveness of preoperative autologous blood donation in gynecologic and gynecologic oncology patients.
METHODS:
Pheresis unit records were retrospectively ...reviewed to identify all women who performed autologous blood donation. Clinical charts were abstracted. Use rate (number of units used/number of units donated) and quality-adjusted life years were calculated. Statistical analysis consisted of χ
2, Student
t, and Fisher exact tests.
RESULTS:
A total of 106 women with benign (
n = 63) and malignant disease (
n = 43) donated 143 units (1.4 units per patient) of which 126 (88%) were discarded. Fifteen patients (14%) were transfused a total of 24 units, 17 autologous (71%) and seven allogeneic (29%). Those transfused had a significantly higher estimated blood loss (700 mL versus 275 mL,
P < .001), lower nadir hemoglobin (7.9 versus 9.6,
P < .001), and longer hospital stay (4.9 days versus 4.0 days,
P = .05). Despite similar estimated blood loss (370 mL versus 310 mL), the use rate for malignant versus benign disease was significantly greater (0.31 versus 0.07,
P = .005). Radical versus nonradical surgery had a significantly higher estimated blood loss (620 mL versus 250 mL,
P = .001) and use rate (0.26 versus 0.11,
P = .001) as well. Estimated cost per quality-adjusted life years for autologous blood donation for each category exceeded $1,000,000.
CONCLUSION:
Autologous blood donation is an expensive medical practice and does not guarantee that exposure to allogeneic blood will not occur. If pursued, it should be directed towards those who have a known malignancy or those for whom radical surgery is anticipated. Other methods of blood conservation may be safer and more cost-effective.
PDF
