Results: 5

1.
Fig. 1.

Fig. 1. From: Phosphoaspirin (MDC-43), a novel benzyl ester of aspirin, inhibits the growth of human cancer cell lines more potently than aspirin: a redox-dependent effect.

Chemical structures of aspirin, m-phosphoaspirin (MDC-63) and p-phosphoaspirin (MDC-43). An aromatic linker molecule binds diethylphosphate to the carboxyl group of conventional aspirin. The position of diethylphosphate on the benzene ring of the linker moiety defines the two isomers.

Wenping Zhao, et al. Carcinogenesis. 2009 March;30(3):512-519.
2.
Fig. 3.

Fig. 3. From: Phosphoaspirin (MDC-43), a novel benzyl ester of aspirin, inhibits the growth of human cancer cell lines more potently than aspirin: a redox-dependent effect.

The effect of P-ASA on ROS levels in SW480 colon cancer cells. (A) SW480 cells were preloaded with a molecular probe for ROS as indicated and treated with P-ASA (MDC-43) for 1 h. DCFDA is a general ROS probe; DHE detects superoxide anion in cells; MitoSOX Red detects specifically mitochondrial superoxide anion and DAF-FM detects NO. (B) Superoxide anion levels in mitochondria detected by MitoSOX Red were decreased following pretreatment with NAC. Values are the mean ± SEM of four independent experiments; *P < 0.05. (C) Cells were stained with MitoSOX Red and MitoTracker Green, a stain specific for mitochondria. The overlay images (lower row) establish the mitochondrial origin of the increased superoxide anion levels in response to P-ASA.

Wenping Zhao, et al. Carcinogenesis. 2009 March;30(3):512-519.
3.
Fig. 2.

Fig. 2. From: Phosphoaspirin (MDC-43), a novel benzyl ester of aspirin, inhibits the growth of human cancer cell lines more potently than aspirin: a redox-dependent effect.

The cell kinetic effect of P-ASA on SW480 colon cancer cells. SW480 cells were grown overnight and treated with P-ASA (MDC-43) as shown. (A) Cell proliferation assay based on bromodeoxyuridine (BrdU) incorporation into DNA during the S-phase of the cell cycle. The percentage of bromodeoxyuridine positive cells is shown in the right upper corner of each panel. (B) Cell cycle analysis by PI staining for DNA content of cells treated with and without P-ASA. Results, quantified in (C), demonstrate the induction of a G2/M to G0/G1 block by P-ASA. (D) Flow cytometric analysis of cells stained with PI and Annexin V (A). A(−)/PI(−) cells are viable cells; A(+)/PI(−) are early apoptotic; A(+)/PI(+) are late apoptotic and A(−)/PI(+) are necrotic. The numbers inside each panel represent the percentage of cells in each category. NAC 20 mM was used to pretreat the cells for 4 h. (E) The effect of pretreatment with NAC on cell viability in response to P-ASA was determined by trypan blue staining and cell counting. Figures are representative of two experiments, whose results were within 10%.

Wenping Zhao, et al. Carcinogenesis. 2009 March;30(3):512-519.
4.
Fig. 5.

Fig. 5. From: Phosphoaspirin (MDC-43), a novel benzyl ester of aspirin, inhibits the growth of human cancer cell lines more potently than aspirin: a redox-dependent effect.

Cell signaling effects of P-ASA in SW480 colon cancer cells. (A and B) SW480 cells were grown overnight and treated with 25 μM P-ASA (MDC-43) for up to 24 h, and the proteins shown were detected by immunoblot in whole-cell extracts. Phosphorylated (p-) and total p38, JNK, ERK and AKT were assayed. Loading control in A–D: β-actin. (C and D) Pretreatment with NAC 20 mM for 4 h reduces the activation of MAPKs and the induction of COX-2 in response to P-ASA treatment. (E) NF-κB-DNA binding was detected by an enzyme-linked immunosorbent assay method in SW480 cells treated with several concentrations of P-ASA for 4 h. Values are the mean ± SEM of three independent experiments; *P < 0.01 compared with control. (F) Electrophoretic mobility shift assay for NF-κB in SW480 cells treated with 20 μM P-ASA for 4 h. NAC 20 mM was used to pretreat the cells for 4 h. To determine the specificity of the NF-κB-DNA complex, the control nuclear fraction was incubated before the binding assay with 100-fold molar excess of unlabeled oligonucleotide containing the sequence for NF-κB (lane labeled +NF-κB) or an unrelated transcription factor (lane labeled +SP-1).

Wenping Zhao, et al. Carcinogenesis. 2009 March;30(3):512-519.
5.
Fig. 4.

Fig. 4. From: Phosphoaspirin (MDC-43), a novel benzyl ester of aspirin, inhibits the growth of human cancer cell lines more potently than aspirin: a redox-dependent effect.

P-ASA decreases thiol levels and induces intrinsic apoptosis in SW480 colon cancer cells. (A) SW480 cells were grown overnight and treated with various concentrations of P-ASA (MDC-43) for 4 h. GSH levels, determined as in Materials and Methods, were decreased in a concentration-dependent manner. Values are the mean ± SEM of three independent experiments. *P < 0.05 compared with control. (B) Overnight pretreatment with 20 mM NAC restores GSH levels in P-ASA-treated cells. BSO, an inhibitor of GSH synthase, was used as a control for GSH depletion. (C) SW480 cells were treated with or without BSO for 24 h, followed by treatment with P-ASA for 18 h. Data (mean ± SEM of three experiments) are expressed as percent of control. (D) Inmunoblots for procaspase 8, procaspase 9 and caspase 9 in SW480 cells treated for 18 h with 1×, 1.5× or 2× IC50 P-ASA. Loading control: β-actin. (E) SW480 cells treated with P-ASA for 18 h following pretreatment with 20 mM NAC or vehicle for 4 h. Procaspase 9, caspase 9 and β-actin were detected by immunoblot. (F) SW480 cells were treated with P-ASA 1.5× IC50 for 3 h and their mitochondria membrane potential was determined by flow cytometry as described in Materials and Methods. Upper panel: fluorescence histograms of control SW480 cells and cells treated with P-ASA; the latter show a shift to the right indicating increased green fluorescence and thus collapsed mitochondrial membrane potential [the corresponding geometric means are as follows: control = 152 ± 17, P-ASA = 258 ± 27 (mean ± SEM)]. Lower panel: flow cytometry of SW480 cells stained as in Materials and Methods for mitochondrial membrane potential. Abscissa, FL1 (green fluorescence); ordinate, FL2 (red fluorescence). The shift toward green fluorescence indicates collapsed mitochondrial membrane potential.

Wenping Zhao, et al. Carcinogenesis. 2009 March;30(3):512-519.

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