Inhibition of anchorage-independent growth of pancreatic cancer cells by YAP1 siRNA oligonucleotides.

<p>A) Percentage of colonies relative to the Cells Only control on soft agar after 21 days of growth from initial seeding. p-values were calculated by comparing the YAP1 siRNA (YAP1_1 and YAP1_2) to the Neg siRNA (independent two tailed <i>t</i> test, two-tailed). B) qPCR analysis of YAP1 mRNA expression after 48 hours of siRNA transfection. C) Western blotting analysis of YAP1 expression after siRNA-treatment for 72 hrs. The samples for each cell line were separated and analyzed on the same blot.* denotes p<0.05, and ** denotes p<0.01.</p

Fig. S4 from Nuclear PTEN Localization Contributes to DNA Damage Response in Endometrial Adenocarcinoma and Could Have a Diagnostic Benefit for Therapeutic Management of the Disease

Supplementary Figure 4. Effects of Zeocin treatment on levels of �H2AX and accumulation of cells in G2/M phase in PTEN-null Ishikawa cells after the overexpression of four different PTEN plasmids. Cells were transfected with plasmids for 24 h, treated with Zeocin for 16 h, then assayed. A-P Levles of �H2AX as determined by co-immunocytostaining with anti-PTEN antibody followed by Alexa Fluor 488 conjugated secondary antibody (green) and anti-phospho H2AX (Ser 139) antibody followed by Alexa Fluor 546 conjugated secondary antibody (red). Nuclei were stained with DAPI (blue). A-D, pEGFP-EV. E-H, PTEN-WT. I-L, PTEN-NLS. M-P, PTEN-NES. Scale bar; 200 μm. Q, Comparison of % �H2AX+ cells as determined by flow cytometry analysis in PTEN transfected cells with or without Zeocin treatment. R, Comparison of the percentage of cells in G2/M as determined by flow cytometry analysis in PTEN transfected cells with or without Zeocin treatment. Statistical analysis performed was paired T-test for each plasmid P<0.05</p

Summary of YAP1 immunostaining in pancreatic tumors and adjacent normal samples.

<p>Summary of YAP1 immunostaining in pancreatic tumors and adjacent normal samples.</p

Brain (1.5T MRI appearance of multiple brain metastasis over time.

<p>Top row contains axial images from the T1-FLAIR (Flip Angle Inversion Recovery) sequences. Bottom row displays corresponding axial contrast-enhanced (T1-GAD) images of selected metastasis that were the target of rapid autopsy analysis. Notice that the right precentral gyrus brain metastasis evolved into a more solid appearing lesion over time (panels A, C and E bottom row, red arrow) and incited significant white matter edema on the final scan (panel E, top row, red arrow). The left inferior parietal lesion (panels B, C and E panels-white arrows) evolved to become more necrotic over time with mild incitement of white matter edema on T1-FLAIR on the final image (panel E, top row, white arrow) and an enhancing rind of tumor surround the necrotic portion of the lesion (panel E, bottom row-white arrow). Such behavior on MRI might be anticipated given the distinct clonal nature of each of the tumor metastasis.</p

Clinicopathological and molecular details for the samples from the MSC patient.

<p>Clinicopathological and molecular details for the samples from the MSC patient.</p

Fig. S1 from Nuclear PTEN Localization Contributes to DNA Damage Response in Endometrial Adenocarcinoma and Could Have a Diagnostic Benefit for Therapeutic Management of the Disease

Supplementary Figure 1. A, Oncoprints of PTEN and TP53 in four molecular subtypes of EndoCA based on TCGA. B, Mutational landscape of PTEN in EndoCA. DSPc; dual specificity phosphatase domain, C2; cell membrane binding domain. Results were generated using C-BioPortal website algorithms (48, 49).</p

Fig. S3 from Nuclear PTEN Localization Contributes to DNA Damage Response in Endometrial Adenocarcinoma and Could Have a Diagnostic Benefit for Therapeutic Management of the Disease

Supplementary Figure 3. PTEN localization in cytoplasm and nuclei of epithelial cells in ANT and EndoCA by grade. PTEN immunostaining was performed on adjacent normal tissues (ANT) and matched primary endometrial tumors (EndoCA). Quantification of staining intensities was assessed by histo-scoring (H-score) in cytoplasm and nuclei of epithelial cells. Average intensities are expressed as median with 95% CI, and statistical significance are determined by Wilcoxon matched-pairs signed rank test. A, n=10 matched ANT and Grade1 tumors. B, n=9 matched ANT and Grade 2 tumors. C, n=10 matched ANT and Grade 3 tumors.</p

YAP1 protein expression and localization in pancreatic cancer cell lines.

<p>As cell density increases, the pancreatic cancer cell lines, BxPC-3 and PANC-1, response to the deactivation of YAP1 as a transcription cofactor by cytosolic sequestration is less substantial than HPDE6. Cells were extracted at increasing cell densities after 48 hours of initial seeding. PARP serves as the loading control for the nuclear fraction. α-tubulin serves as the loading control for the whole cell lysates and cytosolic fraction.</p

Effect of YAP1 targeted siRNAs on the proliferation and apoptosis of pancreatic cancer cell lines.

<p>The cell growth curves of A) BxPC-3 and B) PANC-1 transfected with YAP1 siRNA oligonucleotides. The independent two-sample <i>t</i> test (two-tailed) was utilized to calculate the statistical significance at the 96-hour time point compared to Neg siRNA. * denotes p<0.05, and ** denotes p<0.01. C) The Caspase-Glo 3/7 Assay (Promega) was used to determine the level of apoptosis in pancreatic cancer cells transfected with YAP1 siRNA oligonucleotides after 72 hours. ** denotes an independent two-sample <i>t</i> test (two-tailed) p-value of <0.01 when compared to the negative siRNA control (Neg siRNA).</p

Clonal analyses of the autopsy samples from the maxillary sinus cancer patient.

<p>A) DAPI-based DNA content analysis detected a 3.8N clonal population in brain right frontal pole brain left cerebellar and lung left lower lobe samples, while a 2.4N clonal population was seen in the jejunum sample. The diploid and aneuploid populations were sorted for subsequent aCGH studies. <b>B</b>) Zoomed in chromosome view showing the <i>PKP4</i> gene locus in the above four samples. <b>C</b>) Zoomed in chromosome view showing the <i>pre-miR-651 (labeled miR-651)</i> gene locus status in the above four samples.</p