Journal: Science Advances

Article Title: Interferon response and epigenetic modulation by SMARCA4 mutations drive ovarian tumor immunogenicity

doi: 10.1126/sciadv.adk4851

Figure Lengend Snippet: ( A ) Up-regulated Hallmark and Gene Ontology (GO) pathways in ID8 cell single-guide SMARCA4 (sg SMARCA4 ) compared to those in sgNTC. Ribodeplete RNA sequencing was performed. Statistical analysis was based on hypergeometric test and performed using ClusterProfiler. IL-6, interleukin-6; JAK, Janus kinase; STAT3, signal transducer and activator of transcription 3; TNFA, tumor necrosis factor–α; FDR, false discovery rate. ( B ) Gene expression heatmap of type I IFN pathway–related genes in ID8 cells. Reads per kilobase of transcript per million mapped reads values were scaled to z -score for visualization. Gene expression fold change of sg SMARCA4 versus sgNTC cells is color coded according to the legend. ( C ) Quantitative reverse transcription polymerase chain reaction (qRT-PCR) validation results for IFN genes in ID8 cells (sgNTC and four sg SMARCA4 clones). Expression levels were normalized to β-actin expression, and comparisons of mRNA expression levels were performed relative to control (sgNTC). n = 3 independent experiments. ( D ) MHC1 expression in ID8 cells with or without IFN-ɣ by flow cytometry. ( E ) PD-L1 expression in ID8 cells with or without IFN-ɣ by flow cytometry. MFI, median fluorescence intensity. Statistical analysis was performed using two-tailored unpaired t test [(C) to (E)]. * P < 0.05, ** P < 0.01, *** P < 0.001, and **** P < 0.0001; n.s., not significant. Error bars represent ± SEM. Samples in duplicates [(A) and (B)] and triplicates [(D) and (E)]. KO, knockout; KO_1, target exon 14 clone 1; KO_2, target exon 14 clone 2; KO_3, target exon 23 clone 3; KO_4, target exon 23 clone 4; NTC, non-target control; NTC_1, NTC clone 1.

Article Snippet: Knockouts of STING were generated using gene-specific CRISPR-Cas9–knockout plasmids for STING (no. SC428364, Santa Cruz Biotechnology) and for NTC (SC-421688, Santa Cruz Biotechnology).

Techniques: RNA Sequencing, Gene Expression, Reverse Transcription, Polymerase Chain Reaction, Quantitative RT-PCR, Biomarker Discovery, Clone Assay, Expressing, Control, Flow Cytometry, Fluorescence, Knock-Out

Journal: Science Advances

Article Title: Interferon response and epigenetic modulation by SMARCA4 mutations drive ovarian tumor immunogenicity

doi: 10.1126/sciadv.adk4851

Figure Lengend Snippet: ( A ) Changes in genomic site accessibility in sg SMARCA4 (KO1) versus sgNTC cells. Log 2 fold change and FDR-adjusted P value (Wald test P values from DESeq2 with Benjamini-Hochberg correction). ( B ) Gene set enrichment analysis (GSEA) of immune pathways with increased accessibility in sg SMARCA4 versus sgNTC cells. Kolmogorov-Smirnov statistic with Benjamini-Hochberg correction. Exact q values indicated in each panel. ( C ) Changes in chromatin accessibility at transcription start sites (TSSs) of ISGs (CXCL10 and CCL2) in sg SMARCA4 versus sgNTC cells. ( D ) Motifs enriched in open chromosomal regions affected by SMARCA4 deficiency. P value and binomial test were performed using Homer2. Experiments performed in duplicates. GO, Gene Ontology; KO, knockout; KO_1, target exon 14 clone 1; NTC, non-target control; TF, transcription factor.

Article Snippet: Knockouts of STING were generated using gene-specific CRISPR-Cas9–knockout plasmids for STING (no. SC428364, Santa Cruz Biotechnology) and for NTC (SC-421688, Santa Cruz Biotechnology).

Techniques: Knock-Out, Control

Journal: Science Advances

Article Title: Interferon response and epigenetic modulation by SMARCA4 mutations drive ovarian tumor immunogenicity

doi: 10.1126/sciadv.adk4851

Figure Lengend Snippet: ( A ) IFNAR-1 neutralizing assay in ID8 single-guide NTC (sgNTC) compared to that in sg SMARCA4 cells (four sg SMARCA4 clones). Cells were treated with Mock or IFNAR-1 antibody for 48 hours at 10 μg/ml. ( B ) IRF3 expression levels in sg SMARCA4 versus sgNTC cells by qRT-PCR. ( C ) IRF3 expression in doxycycline-inducible short hairpin NTC (shNTC) and shIRF3-transfected sg SMARCA4 cells by qRT-PCR. ( D ) qRT-PCR quantification of ISGs in ID8 sgNTC and sg SMARCA4 cells. The latter were transfected with shIRF3 or shNTC (with and without POLYI:C). ( E ) Enriched IRF motif at TSSs of ISG locus from ATAC-seq analysis of ID8 sgNTC versus sg SMARCA4 cells. Experiments performed in duplicates. ( F ) Analysis of publicly available ChIP-Atlas data of IRF3 DNA binding sites on ISGs in murine immune cell lines. Statistical analysis was performed using two-tailored unpaired t test [(A) to (D)]. * P < 0.05, ** P < 0.01, *** P < 0.001, and **** P < 0.0001. Error bars represent ± SEM. n = 3 independent experiments in (A) to (D). Expression levels were normalized to β-actin expression, and comparisons of mRNA expression levels were performed relative to control [sgNTC_1 in (A), (B), and (D) and sg SMARCA4 transfected with shNTC in (C), as indicated]. KO, knockout; KO_1, target exon 14 clone 1; KO_2, target exon 14 clone 2; KO_3, target exon 23 clone 3; KO_4, target exon 23 clone 4; LPS, lipopolysaccharide stimulated, NTC, non-target control; NTC_1 = NTC clone 1; POLYI:C, polyinosinic-polycytidylic acid stimulated.

Article Snippet: Knockouts of STING were generated using gene-specific CRISPR-Cas9–knockout plasmids for STING (no. SC428364, Santa Cruz Biotechnology) and for NTC (SC-421688, Santa Cruz Biotechnology).

Techniques: Neutralizing Assay, Clone Assay, Expressing, Quantitative RT-PCR, Transfection, Binding Assay, Control, Knock-Out

Journal: Science Advances

Article Title: Interferon response and epigenetic modulation by SMARCA4 mutations drive ovarian tumor immunogenicity

doi: 10.1126/sciadv.adk4851

Figure Lengend Snippet: ( A ) Volcano plots for differential expression of TEs in ID8 cells. ( B ) Heatmap of long terminal repeats (LTR) with adjusted P < 0.05. ( C ) Representative pictures (top) of double-stranded RNA (dsRNA) identification by immunofluorescence (IF) in sg SMARCA4 and sgNTC ID8 cells with quantification (bottom). RNAse, ribonuclease. ( D ) qRT-PCR quantification of ISGs in ID8 sgNTC cells and sg SMARCA4 cells transfected with shMAVS or shNTC. ( E ) MAVS expression in doxycycline-inducible shNTC and shMAVS-transfected sg SMARCA4 cells by qRT-PCR. Expression levels were normalized to β-actin expression, and comparisons of mRNA expression levels were performed relative to control [sg SMARCA4 transfected with shNTC in (C) and sgNTC_1 in (D), as indicated]. Statistical analysis was performed using two-tailored unpaired t test [(C) to (E)]. * P < 0.05, ** P < 0.01, *** P < 0.001, and **** P < 0.0001. Error bars represent ± SEM. Samples in duplicates [for (A) and (B)]. n = 3 independent experiments [in (C) and (D)]. Red line in (A) represents a cutoff of an adjusted P value of <0.05. KO, knockout; KO_1, target exon 14 clone 1; KO_2, target exon 14 clone 2; KO_4, target exon 23 clone 4; NTC, non-target control; NTC_1, NTC clone 1.

Article Snippet: Knockouts of STING were generated using gene-specific CRISPR-Cas9–knockout plasmids for STING (no. SC428364, Santa Cruz Biotechnology) and for NTC (SC-421688, Santa Cruz Biotechnology).

Techniques: Quantitative Proteomics, Immunofluorescence, Quantitative RT-PCR, Transfection, Expressing, Control, Knock-Out

Journal: Science Advances

Article Title: Interferon response and epigenetic modulation by SMARCA4 mutations drive ovarian tumor immunogenicity

doi: 10.1126/sciadv.adk4851

Figure Lengend Snippet: ( A ) Workflow for the OC tumor model. C57BL/6 (Cg)–Tyr c-2J /J mice were inoculated intraperitoneally with 10 million ID8 sgNTC or sg SMARCA4 ( KO_4 ) tumor cells, and spectral flow cytometry was performed 21 days later on harvested ascites samples. ip, intraperitoneal. ( B ) Frequency of tumor PD1 + CD4 + T cells and PD1 + CD8 + T cells. ( C ) Frequency of NK1.1 + cells expressing Granzyme B + . ( D ) Frequency of tumor dendritic cells (of CD45 + cells) and MHCII-expressing dendritic cells. ( E ) Frequency of tumor macrophages (of CD45 + cells) and PD-L1–expressing macrophages. ( F ) In vivo bioluminescence imaging of tumor burden in sg SMARCA4 versus sgNTC tumors (unpaired t test of area under the curve). Statistical analysis was performed using two-tailored unpaired t test [(B) to (F)]. * P < 0.05, ** P < 0.01 and **** P < 0.0001. Error bars represent ± SEM. n = 10 mice per group. KO, knockout; KO_4, target exon 23 clone 4; NTC, non-target control; NTC_1, NTC clone 1.

Article Snippet: Knockouts of STING were generated using gene-specific CRISPR-Cas9–knockout plasmids for STING (no. SC428364, Santa Cruz Biotechnology) and for NTC (SC-421688, Santa Cruz Biotechnology).

Techniques: Flow Cytometry, Expressing, In Vivo, Imaging, Knock-Out, Control

Journal: Science Advances

Article Title: Interferon response and epigenetic modulation by SMARCA4 mutations drive ovarian tumor immunogenicity

doi: 10.1126/sciadv.adk4851

Figure Lengend Snippet: ( A ) Workflow for the B16-F10 tumor model. C57BL/6 (Cg)–Tyr c-2J /J mice were inoculated subcutaneously with 150,000 shNTC, sgEV, sh SMARCA4_1 , sh SMARCA4_2 , or sg SMARCA4 tumor cells, and spectral flow cytometry was performed 21 days later on harvested tumor samples. ( B ) Frequency of tumor CD8 + T cells and MFI of ICOS + -expressing CD8 + T cells in B16-F10 model. ( C ) Frequency of NK1.1 + cells in B16-F10 knockout and knockdown models (of CD45 + cells). ( D ) MFI of MHCII in macrophages in B16-F10 SMARCA4 knockout and knockdown models. ( E ) Tumor volume of sh SMARCA4 versus shNTC B16 tumors (unpaired t test of final time points). n = 5 mice per group, three biological replicates. ( F ) Frequency of NK1.1 + cells in sh SMARCA4 versus shNTC tumors (of live cells) in RAG2 −/− mice. ( G ) MFI of MHCII-expressing macrophages in B16-F10 sh SMARCA4 versus shNTC tumors in RAG2 −/− mice. ( H ) Tumor volumes of sh SMARCA4 versus shNTC B16 tumors in RAG2 −/− mice (unpaired t test of final time points). n = 3 to 5 mice per group, three biological replicates. Statistical analysis was performed using two-tailored unpaired t test. * P < 0.05, ** P < 0.01, *** P < 0.001. Error bars represent ± SEM. Dox, doxycycline; EV, empty vector; NTC, non-target control; SC, subcutaneous.

Article Snippet: Knockouts of STING were generated using gene-specific CRISPR-Cas9–knockout plasmids for STING (no. SC428364, Santa Cruz Biotechnology) and for NTC (SC-421688, Santa Cruz Biotechnology).

Techniques: Flow Cytometry, Expressing, Knock-Out, Knockdown, Plasmid Preparation, Control

Journal: bioRxiv

Article Title: Increasing Cas9-mediated homology-directed repair efficiency through covalent tethering of DNA repair template

doi: 10.1101/231035

Figure Lengend Snippet: ( a ) Schematic of Cas9 fused to the HUH endonuclease PCV with a covalently attached ssODN. ( b ) SDS-PAGE of Cas9 variants reacted with an Alexa 488 fluorescently labelled ssDNA containing the PCV recognition sequence. The top panel is the coomassie stained gel, and the bottom panel is the identical fluorescently imaged gel. PCV is fused to either the carboxyl (Cas9-PCV) or amino (PCV-Cas9) terminus of Cas9. Cas9-PCV(Y96F) represents catalytically inactive PCV(Y96F) fused to Cas9. (C) SDS-PAGE gel shift assay of Cas9 reacting with two ssDNA templates containing the PCV recognition sequence of differing lengths in a 1:1 ssDNA:Cas9 molar ratio.

Article Snippet: Streptococcus pyogenes Cas9 was amplified out of the plasmid pET15_SP-Cas9 (a gift from Niels Geijsen, Addgene plasmid #62731) and inserted in pTD68_SUMO-PCV2 at the BamHI site using Infusion cloning (Clontech) to create C-terminally fused Cas9-PCV.

Techniques: SDS Page, Sequencing, Staining, Electrophoretic Mobility Shift Assay

Journal: bioRxiv

Article Title: Increasing Cas9-mediated homology-directed repair efficiency through covalent tethering of DNA repair template

doi: 10.1101/231035

Figure Lengend Snippet: ( a ) Schematic of split luciferase insertion. The C-terminus of NanoLuc nanoluciferase (HiBiT) is encoded on the 200bp ssODN along with the 5’ PCV recognition sequence and targeted to the 3’ end of GAPDH . ( b ) Assaying luminescence using different Cas9 variants when inserting HiBiT into GAPDH in HEK-293T cells. PCV is fused to either the amino (PCV-Cas9) or carboxyl (Cas9-PCV) terminus of Cas9. Transfections were performed with ssODN lacking the PCV recognition sequence (PCV- ssODN) or ssODN containing the PCV recognition sequence (PCV+ ssODN). Units are displayed in relative light units (RLU) normalized to Cas9. ( c ) The calculated fold change from (b) between the PCV-ssODN and PCV+ ssODN is shown for each variant. ( d ) Targeting the GAPDH locus in U2-OS cells. ( e ) Targeting a locus in vinculin in HEK-293T cells using an ssODN containing the PCV recognition sequence. ( f ) Fold change in RLU compared to Cas9 when varying the amount of RNP (equimolar ssODN). All graphs represent data from one of multiple independent experiments exhibiting similar results. Data are shown as mean +/− SD (n=3). Significance calculated using 2-tailed Student’s t-test: ** P < 0.01, *** P < 0.001, ns = no significance (P >0.05).

Article Snippet: Streptococcus pyogenes Cas9 was amplified out of the plasmid pET15_SP-Cas9 (a gift from Niels Geijsen, Addgene plasmid #62731) and inserted in pTD68_SUMO-PCV2 at the BamHI site using Infusion cloning (Clontech) to create C-terminally fused Cas9-PCV.

Techniques: Luciferase, Sequencing, Transfection, Variant Assay

Journal: bioRxiv

Article Title: Increasing Cas9-mediated homology-directed repair efficiency through covalent tethering of DNA repair template

doi: 10.1101/231035

Figure Lengend Snippet: ( a ) HEK-293T cells stably expressing a mutant mCherry-GFP reporter are edited by HDR through a frameshift correction, restoring mCherry activity. ( b ) Representative microscopy images of fluorescent reporter editing. ( c ) The percent of mCherry positive cells determined by flow cytometry at two different RNP concentrations using an ssODN containing the PCV recognition sequence ( d ) RNP transfections at 3 pmol in the presence or absence of ssODN. Data are shown as mean +/− SD (n=3). For (c) and (d), the statistical significance of %mCherry positive cells between PCV-fusions of Cas9 and Cas9 alone was <0.001, calculated using 2-tailed Student’s t-test.

Article Snippet: Streptococcus pyogenes Cas9 was amplified out of the plasmid pET15_SP-Cas9 (a gift from Niels Geijsen, Addgene plasmid #62731) and inserted in pTD68_SUMO-PCV2 at the BamHI site using Infusion cloning (Clontech) to create C-terminally fused Cas9-PCV.

Techniques: Stable Transfection, Expressing, Mutagenesis, Activity Assay, Microscopy, Flow Cytometry, Sequencing, Transfection

Journal: Stem Cells and Development

Article Title: A Novel Human Pluripotent Stem Cell-Derived Neural Crest Model of Treacher Collins Syndrome Shows Defects in Cell Death and Migration

doi: 10.1089/scd.2017.0234

Figure Lengend Snippet: NC cells transfected with TCOF1 siRNA impair regular migration of NC and MSC. (A) H9s-derived NC cells were transiently transfected with a siRNA to TCOF1. Left panel : Flow cytometry was performed for CD44 and P75 following 5 days of differentiation from NC to MSC, demonstrating that TCOF1 KD does not impair MSC differentiation. Right panel : Flow cytometric analysis of CD44 and P75 in NC transiently transfected with a siRNA to TCOF1. Red contour plots represent CD44+P75 double stained populations, and blue contour plots represent isotype control staining. (B) MTT cell proliferation assay was performed using TCOF1 KD NC cells (NC siRNA TCOF1) and siRNA scramble NC cells (NC siRNA SCRAMBLE) during 4 days. Results are presented as mean ± SD of three independent experiments. * P < 0.05; ** P < 0.01. Two-sided Student's t -test. (C) Representative images of scratch wound assays of HESC-derived NC cells transiently transfected with Scramble siRNA or TCOF1 siRNA. Images were collected 4 days following transfection, at the indicated time points. Scale bar: 100 μm. (D) Box plot depicting the quantification of chemotaxis potential and migration of NC cells transfected with TCOF1 siRNA or Scramble siRNA during a 6 h CytoSelect Cell Migration Assay. FGF8B was used as a NC chemoattractant. DMEM/F12 + 10% fetal bovine serum (SERUM) was used as a positive control for cell migration. Data are expressed relative to the NC cells transfected with TCOF1 siRNA, maintained in FSB medium. * P < 0.05; ** P < 0.01. Two-sided Student's t -test. (E) Representative images of scratch wound assays of NC-derived MSC transiently transfected with Scramble siRNA or TCOF1 siRNA. Images were collected after 5 days of differentiation, at the indicated time points. Scale bar: 100 μm. KD, knockdown; MTT, 3-(4, 5-dimethylthiazolyl-2)-2, 5-diphenyltetrazolium bromide; siRNA, small interfering RNA. Color images available online at www.liebertpub.com/scd

Article Snippet: For gene targeting, 2.5 × 10 6 HIPSC were electroporated with 1 μg of generated TCOF1 targeting Cas9 plasmid in 100 μL of nucleofection mix from the P3 Primary Cell 4D-Nucleofector X Kit (Lonza) using a 4D-nucleofector system (Lonza).

Techniques: Transfection, Migration, Derivative Assay, Flow Cytometry, Staining, MTT Cell Proliferation, Chemotaxis Assay, Cell Migration Assay, Positive Control, Small Interfering RNA

Journal: Stem Cells and Development

Article Title: A Novel Human Pluripotent Stem Cell-Derived Neural Crest Model of Treacher Collins Syndrome Shows Defects in Cell Death and Migration

doi: 10.1089/scd.2017.0234

Figure Lengend Snippet: Generation of TCOF1 heterozygous knockout HIPSC using CRISPR/Cas9. (A) Schematic of CRISPR/Cas9-mediated nonhomologous end joining strategy to generate INDELs leading to HIPSC TCOF1 knockout clones. (B) Representative genomic sequencing of HIPSC transfected with specific CRISPR TCOF1 gRNA. Heterozygous knockout clones showed the same deletion leading to a change in codon reading frame and the introduction of a premature STOP codon in TCOF1 Exon 1. (C) Immunoblot for Treacle protein demonstrating that NC derived from HIPSC TCOF1 +/− clones (C12 and C24) showed a reduction in Treacle compared with NC derived from HIPSC TCOF1 +/+ (C8) and NC derived from H9s (WT). (D) Time course MTT proliferation assay of TCOF1 +/+ NC derived from H9s (WT), HIPSC (C8), and NC derived from TCOF1 +/− HIPSC (C12 and C24), over a 4-day period. Proliferation rate decreased significantly in mutated cells compared with WT. Results are presented as mean ± SD of three independent experiments. ** P < 0.01, *** P < 0.001, two-sided Student's t -test. (E) Flow cytometric analysis of Annexin V staining depicting the apoptotic rate in TCOF1 +/+ NE-derived cells from H9s (WT), HIPSC (C8), and NE derived from TCOF1 +/− HIPSC (C12 and C24). Red histograms represent Annexin V staining, and blue histograms represent the unstained population. (F) Single cell analysis of cell migration in NC derived from TCOF1 +/+ HIPSC (C8) and TCOF1 +/− HIPSC (C24). TCOF1 +/− NC demonstrates impaired migration and reduced directionality of movement. Each dot and tail represent a single cell analyzed, with 30 cells analyzed from both conditions, over a 12-h period. gRNA, guide RNA; HIPSC, human induced pluripotent stem cell; INDELs, insertions or deletions; WT, wild type. Color images available online at www.liebertpub.com/scd

Article Snippet: For gene targeting, 2.5 × 10 6 HIPSC were electroporated with 1 μg of generated TCOF1 targeting Cas9 plasmid in 100 μL of nucleofection mix from the P3 Primary Cell 4D-Nucleofector X Kit (Lonza) using a 4D-nucleofector system (Lonza).

Techniques: Knock-Out, CRISPR, Clone Assay, Genomic Sequencing, Transfection, Western Blot, Derivative Assay, Proliferation Assay, Staining, Single-cell Analysis, Migration

Journal: Cancers

Article Title: Differential Dependency of Human Pancreatic Cancer Cells on Targeting PTEN via PLK 1 Expression

doi: 10.3390/cancers12020277

Figure Lengend Snippet: Expression of polo-like kinase 1 (PLK1) as a companion biomarker with phosphatase and tensin homolog (PTEN) expression in pancreatic cancer. ( a ) Overall survival was analyzed using Kaplan-Meier curves depending on the differential expression of PTEN in PTEN-adverse data-sets, PAAD-US-TCGA, GSE78229, and GSE62452 ( p value was calculated using Log-rank (Mantel-Cox) Test). ( b ) Overall survival was analyzed using Kaplan-Meier curves depending on the differential expression of PTEN in PTEN favorable data-sets, PACA-AU and GSE21501 ( p value was calculated using Log-rank (Mantel-Cox) Test). ( c ) PLK1 and PTEN proteins in AsPC-1, Capan-2, Miapaca-2, SNU-213, CFPAC-1, Panc-1, Miapaca-2, and H6c7 cells were detected using the Western blot. GAPDH was used as a control. Relative pixel intensities of PLK1 were measured using ImageJ analysis software (PLK1/GAPDH). Data represent three individual experiments. ( d ) AsPC-1 and SNU-213 cells were transfected with a vector control or the PLK1 overexpression construct. After 48 h of transfection, AsPC-1 and SNU-213 cells were incubated with different doses of SF1670 for an additional 72 h. Viability was measured by the WST-1 assay ( n = 3, Tukey’s post hoc test has been used to determine the significant group effects in ANOVA, p < 0.0001, asterisks indicate a significant difference compared with 0% inhibition, * p < 0.05, ** p < 0.01). ( e ) Miapaca-2 and Panc-1 cells were transfected with scrambled or PLK1-specific siRNA. At 48 h post-transfection, cells were incubated with different doses of SF1670 for an additional 72 h. Viability was measured by the WST-1 assay ( n = 3, Tukey’s post hoc test has been used to determine the significant group effects in ANOVA, p < 0.0001. Asterisks indicate a significant difference compared with 0% inhibition, * p < 0.05, ** p < 0.01). ( f ) After 48 h of transfection with the vector control or PLK1 overexpression construct, AsPC-1 and SNU-213 cell lysates were subjected to Western blot analysis using antibodies specific for PLK1, PTEN, p-AKT, AKT, and Survivin. GAPDH was used as a control. ( g ) Miapaca-2 and Panc-1 cells were transfected with scrambled or PLK1-specific siRNA. After 48 h of transfection, Miapaca-2 and Panc-1 cell lysates were subjected to immunoblot analysis using antibodies specific for PLK1, PTEN, p-AKT, AKT, and Survivin. GAPDH was used as a control.

Article Snippet: Scrambled control (sc-37007) and PLK1 overexpression construct (sc-400411) have been purchased from Santa Cruz Biotechnology.

Techniques: Expressing, Biomarker Discovery, Quantitative Proteomics, Western Blot, Control, Software, Transfection, Plasmid Preparation, Over Expression, Construct, Incubation, WST-1 Assay, Inhibition

Journal: Cancers

Article Title: Differential Dependency of Human Pancreatic Cancer Cells on Targeting PTEN via PLK 1 Expression

doi: 10.3390/cancers12020277

Figure Lengend Snippet: IPPScores compared with PLK1 expression levels in human pancreatic cancer.

Article Snippet: Scrambled control (sc-37007) and PLK1 overexpression construct (sc-400411) have been purchased from Santa Cruz Biotechnology.

Techniques: Expressing

Journal: Cancers

Article Title: Differential Dependency of Human Pancreatic Cancer Cells on Targeting PTEN via PLK 1 Expression

doi: 10.3390/cancers12020277

Figure Lengend Snippet: Prognosis associated with phosphatase and tensin homolog (PTEN) and polo-like kinase 1 (PLK1) expressions in human pancreatic cancer. ( a ) Overall survival of pancreatic cancer patients was analyzed using Kaplan-Meier curves depending on the PLK1 and PTEN expressions in GSE17891 data-set ( p value was calculated using the Log-rank (Mantel-Cox) Test). ( b ) Prognostic relevance was analyzed using the log-rank test and its non-parametric version of curves on the differential expression of PLK1 and PTEN in the GSE17891 data-set. ( c ) Transcriptional levels of PLK1 in low PTEN and high PTEN groups were analyzed using GSE78229 and GSE62452 data sets. (a.u. indicates arbitrary unit using the UPCs method. The p -value was evaluated with a Student’s t test). ( d ) Transcriptional levels of PLK1 in low PTEN and high PTEN groups were analyzed using PACA-AU and GSE21501 data sets. (a.u. indicates an arbitrary unit using the UPCs method. The p -value was evaluated with a Student’s t test).

Article Snippet: Scrambled control (sc-37007) and PLK1 overexpression construct (sc-400411) have been purchased from Santa Cruz Biotechnology.

Techniques: Quantitative Proteomics

Journal: Cancers

Article Title: Differential Dependency of Human Pancreatic Cancer Cells on Targeting PTEN via PLK 1 Expression

doi: 10.3390/cancers12020277

Figure Lengend Snippet: Selection of Polo-like kinase1 (PLK1) using easily applicable techniques and whole blood samples of pancreatic cancer patients. ( a ) soluble PLK1s (sPLK1) were selected by a sandwich ELISA analysis using the same volumes of whole bloods samples from 16 healthy donors (N1–N16) and 12 pancreatic cancer patients (C1–C12). ( b ) Soluble PLK1s (sPLK1) were selected by Western blot analysis using the same volumes of whole blood samples from 10 healthy donors (N1–N10) and 12 pancreatic cancer patients (C1–C12).

Article Snippet: Scrambled control (sc-37007) and PLK1 overexpression construct (sc-400411) have been purchased from Santa Cruz Biotechnology.

Techniques: Selection, Sandwich ELISA, Western Blot