1. Copyright © 2006 American Medical Association. All rights reserved.
From: Small Molecule Inhibition of HDM2 Leads to p53-Mediated Cell Death in Retinoblastoma Cells
Arch Ophthalmol. 2006;124(9): doi: /archopht
Figure Legend:
Activation of p53 cascade in WERI-RB-1 and Y79 cells. A, Confocal indirect immunofluorescence microscopy showing minimal p53 in WERI-RB-1 cells (left) and Y79 cells (right). There is increased nuclear and cytoplasmic p53 (green) after 8 hours of exposure to 5 μM of Nutlin 3A in both cell lines, but no discernable response to Nutlin 3B. B, Western blot analysis of Y79 cells after 8 hours of exposure to both Nutlin 3A and 3B at 0 (dimethyl sulfoxide vehicle), 1, 5, and 10 μM. After Nutlin 3A treatment, p53 levels rise in association with increases in both HDM2 and p21. There are minimal effects after Nutlin 3B treatment.
Date of download: 10/23/2017
Copyright © 2006 American Medical Association. All rights reserved.

2. Copyright © 2006 American Medical Association. All rights reserved.
From: Small Molecule Inhibition of HDM2 Leads to p53-Mediated Cell Death in Retinoblastoma Cells
Arch Ophthalmol. 2006;124(9): doi: /archopht
Figure Legend:
Nutlin 3A causes cell death in a dose-dependent manner. Y79 and WERI-RB-1 cells were incubated with 0 (dimethyl sulfoxide vehicle), 1, 5, or 10 μM of Nutlin 3A. Viable cells were identified by trypan blue exclusion, counted over 5 days, and expressed graphically as cells per high-power field (HPF). Experiments were done in duplicate and Y error bars correspond to standard deviations from the mean. In both experiments, Nutlin 3A at 10 μM results in near complete cell death.
Date of download: 10/23/2017
Copyright © 2006 American Medical Association. All rights reserved.

3. Copyright © 2006 American Medical Association. All rights reserved.
From: Small Molecule Inhibition of HDM2 Leads to p53-Mediated Cell Death in Retinoblastoma Cells
Arch Ophthalmol. 2006;124(9): doi: /archopht
Figure Legend:
Apoptosis in Nutlin 3A–treated WERI-RB-1 cells. A, WERI-RB-1 cells treated with dimethyl sulfoxide (DMSO) for 36 hours demonstrate little Annexin V and propidium iodide (PI) staining. B, WERI-RB-1 cells treated with 10 μM of Nutlin 3A for 36 hours demonstrate a shift to the lower-right quadrant, indicating Annexin V but not PI positivity, which is consistent with early apoptotic activity.
Date of download: 10/23/2017
Copyright © 2006 American Medical Association. All rights reserved.

4. Copyright © 2006 American Medical Association. All rights reserved.
From: Small Molecule Inhibition of HDM2 Leads to p53-Mediated Cell Death in Retinoblastoma Cells
Arch Ophthalmol. 2006;124(9): doi: /archopht
Figure Legend:
p53-deficient cell line demonstrates resistance to Nutlin 3A. MDA-MB-435, Y79, and WERI-RB-1 cells were incubated with 0 (dimethyl sulfoxide [DMSO] vehicle), 1, or 10 μM of Nutlin 3A, and viability was assessed on day 5 by the trypan blue exclusion assay. Viability was expressed as a percentage of viable cells as compared with the DMSO control. MDA-MB-435 cells demonstrated resistance to Nutlin 3A–induced cell death. This experiment was done as a single experiment; therefore, there are no error bars.
Date of download: 10/23/2017
Copyright © 2006 American Medical Association. All rights reserved.

5. Copyright © 2006 American Medical Association. All rights reserved.
From: Small Molecule Inhibition of HDM2 Leads to p53-Mediated Cell Death in Retinoblastoma Cells
Arch Ophthalmol. 2006;124(9): doi: /archopht
Figure Legend:
Y79 with short interfering RNA (siRNA) to p53 is resistant to Nutlin 3A. A, Western blots demonstrating decreased p53 expression in siRNA infected cells vs vector control and Y79 cells. B, Y79 cells with vector alone were plated and incubated with 0 (dimethyl sulfoxide vehicle), 1, 5, or 10 μM of Nutlin 3A for 5 days. Viable cells were counted using the trypan blue exclusion assay. C, Y79 cells with siRNA to p53 were incubated with Nutlin 3A as described and found to have an attenuated response to the small molecule. HPF indicates high-power field.
Date of download: 10/23/2017
Copyright © 2006 American Medical Association. All rights reserved.

6. Copyright © 2006 American Medical Association. All rights reserved.
From: Small Molecule Inhibition of HDM2 Leads to p53-Mediated Cell Death in Retinoblastoma Cells
Arch Ophthalmol. 2006;124(9): doi: /archopht
Figure Legend:
Absence of phosphorylation at serine-15 p53. Western blot analysis on protein lysate from WERI-RB-1 and Y79 cells treated with dimethyl sulfoxide (DMSO), 5 μM of Nutlin 3A, or 1 μM of doxorubicin (Dox) for a period of 24 hours. Nutlin 3A showed no evidence of inducing phosphorylation of Serine-15 p53 in contrast to doxorubicin, implying that p53 activation by Nutlin 3A does not involve DNA damage.
Date of download: 10/23/2017
Copyright © 2006 American Medical Association. All rights reserved.