Journal: Cancers

Article Title: The m 7 G Reader NCBP2 Promotes Pancreatic Cancer Progression by Upregulating MAPK/ERK Signaling

doi: 10.3390/cancers15225454

Figure Lengend Snippet: NCBP2 is significantly expressed in PDAC patients and correlated with poor prognosis. ( A ) Analysis of NCBP2 transcript levels in PDAC and normal tissues based on the RNA-seq data from GEPIA 2.0 cohort. ( B ) Analysis of NCBP2 transcript levels in PDAC and normal tissues based on DNA microarray data from GEO cohorts (GSE15471, GSE28735, GSE16515). ( C , D ) Overall survival and disease-free survival curves for PDAC patients with high or low NCBP2 expression in GEPIA 2.0 cohort. ( E , F ) The RNA and protein expression level of NCBP2 in HPDE and five PDAC cell lines. ( G ) The expression level of NCBP2 in PDAC and para-carcinoma tissues from tissue microarray, * p < 0.05 and *** p < 0.001.

Article Snippet: We obtained human embryonic kidney epithelial cell line 293T (HEK293T) and PDAC cell lines (Panc 05.04, PANC-1, AsPC-1, BxPC-3, MIA PaCa-2) from the American Type Culture Collection (ATCC, Manassas, VA, USA).

Techniques: RNA Sequencing, Microarray, Expressing

Journal: Cancers

Article Title: The m 7 G Reader NCBP2 Promotes Pancreatic Cancer Progression by Upregulating MAPK/ERK Signaling

doi: 10.3390/cancers15225454

Figure Lengend Snippet: NCBP2 promotes PDAC cell growth in vitro. ( A – C ). The cell counting, 3D sphere, and colony formation assay results in control and NCBP2-knockdown Panc 05.04 and PANC-1 cells. ( D – F ). The cell counting, 3D sphere and colony formation assay results in control and NCBP2-overexpression AsPC-1 and BxPC-3 cells, * p < 0.05, ** p < 0.01, and *** p < 0.001.

Article Snippet: We obtained human embryonic kidney epithelial cell line 293T (HEK293T) and PDAC cell lines (Panc 05.04, PANC-1, AsPC-1, BxPC-3, MIA PaCa-2) from the American Type Culture Collection (ATCC, Manassas, VA, USA).

Techniques: In Vitro, Cell Counting, Colony Assay, Control, Knockdown, Over Expression

Journal: Cancers

Article Title: The m 7 G Reader NCBP2 Promotes Pancreatic Cancer Progression by Upregulating MAPK/ERK Signaling

doi: 10.3390/cancers15225454

Figure Lengend Snippet: NCBP2 promotes PDAC cell growth in-vivo. ( A , B ) Tumor growth curves of xenograft models established from control or stable NCBP2-knockdown Panc 05.04 cells. ( C ) Assessment of tumor weight from control and NCBP2-knockdown groups. ( D , E ) The expression level of Ki-67 in tumor tissues from control and NCBP2-knockdown groups. ( F ) Body weight of nude mice in control and NCBP2-knockdown group, * p < 0.05, and ** p < 0.01.

Article Snippet: We obtained human embryonic kidney epithelial cell line 293T (HEK293T) and PDAC cell lines (Panc 05.04, PANC-1, AsPC-1, BxPC-3, MIA PaCa-2) from the American Type Culture Collection (ATCC, Manassas, VA, USA).

Techniques: In Vivo, Control, Knockdown, Expressing

Journal: Cancers

Article Title: The m 7 G Reader NCBP2 Promotes Pancreatic Cancer Progression by Upregulating MAPK/ERK Signaling

doi: 10.3390/cancers15225454

Figure Lengend Snippet: NCBP2 upregulates c-JUN to activate MEK/ERK signaling in a m 7 G-dependent manner. ( A ) KEGG analysis of RNA-seq data in PDAC patients with high or low NCBP2 expression from TCGA cohort. ( B ) The GSEA enrichment plot of “MAPK signaling pathway” in PDAC patients with high or low NCBP2 expression from TCGA cohort. ( C ) Immunoblotting for protein levels of total JNK/phosphorylated JNK (Thr183/Tyr185), total p38/phosphorylated p38 (Thr180/Tyr182), and total ERK/phosphorylated ERK (Thr202/Tyr204) in control and NCBP2-knockdown PDAC cells. Tubulin was used as the internal control. ( D ) Immunoblotting for protein levels of total/phosphorylated MEK and total/phosphorylated ERK (Thr202/Tyr204) in control and c-JUN-knockdown Panc 05.04 and PANC-1 cells. Tubulin was used as the internal control. ( E ) The mRNA expression levels of NCBP2 and c-JUN in control and NCBP2-knockdown Panc 05.04 and PANC-1 cells. ( F ) Immunoblotting for protein levels of NCBP2 and c-JUN in control and NCBP2-knockdown Panc 05.04 and PANC-1 cells. Tubulin was used as the internal control. ( G ) Polysome profiling results of the control and NCBP2-knockdown Panc 05.04 and PANC-1 cells. ( H ) Gene-specific m 7 G qPCR results for the m 7 G methylation levels of c-JUN in PANC 05.04 and PANC-1 cells, *** p < 0.001.

Article Snippet: We obtained human embryonic kidney epithelial cell line 293T (HEK293T) and PDAC cell lines (Panc 05.04, PANC-1, AsPC-1, BxPC-3, MIA PaCa-2) from the American Type Culture Collection (ATCC, Manassas, VA, USA).

Techniques: RNA Sequencing, Expressing, Western Blot, Control, Knockdown, Methylation

Journal: Journal of Translational Medicine

Article Title: The long non-coding RNA HOTTIP promotes progression and gemcitabine resistance by regulating HOXA13 in pancreatic cancer

doi: 10.1186/s12967-015-0442-z

Figure Lengend Snippet: HOTTIP levels are up-regulated in PDAC tissues and cell lines. (A) Total RNA from eight cases of pancreatic ductal adenocarcinoma (PDAC) and four cases of chronic pancreatitis were used for microarray analysis. Long noncoding RNAs (lncRNAs) upregulated >10-fold in PDAC tissues (n = 8) compared with chronic pancreatitis tissues (n = 4) are shown. (B) HOTTIP expression levels in 90 paired PDAC tissues and corresponding adjacent non-neoplastic tissues was examined via quantitative reverse transcription-polymerase chain reaction (qRT-PCR). β-actin was used as internal control. Relative gene expression was determined using the comparative delta-delta CT method, and data are presented as △△Ct. (C) HOTTIP expression was evaluated in five pancreatic cancer cell lines compared with immortalized human ductal epithelial cells by qRT-PCR. HOTTIP mRNA levels were normalized to GAPDH . Data represent the mean ± s.d. from three independent experiments. **p < 0.01, Student’s t-test.

Article Snippet: The human pancreatic cancer cell lines, PANC-1, MIA PaCa-2, Capan-2, SW1990, and BxPC-3, were purchased from the American Type Culture Collection and grown in complete growth medium as recommended by the manufacturer, supplemented with 10% FBS and 1% penicillin/streptomycin.

Techniques: Microarray, Expressing, Reverse Transcription, Polymerase Chain Reaction, Quantitative RT-PCR, Control, Gene Expression

Journal: Journal of Translational Medicine

Article Title: The long non-coding RNA HOTTIP promotes progression and gemcitabine resistance by regulating HOXA13 in pancreatic cancer

doi: 10.1186/s12967-015-0442-z

Figure Lengend Snippet: HOTTIP knockdown enhances the chemosensitivity of human pancreatic cancer cells to gemcitabine in vitro and in vivo. (A) SW1990 cells were infected with control shRNA or an shRNA against HOTTIP as indicated, and then treated with gemcitabine (0.1, 1, or 10 μM) for 72 h. Cell survival rates were then measured using CCK-8 assays. The values are presented as the means ± s.d. **P < 0.01. (B) The effect of HOTTIP suppression on the IC50 of SW1990 cells was calculated. (C) SW1990 cells were infected with control shRNA or shRNA against HOTTIP as indicated and then treated with gemcitabine (10 μM) or PBS (100 μL) for 14 days. Colony formation on plastic was then assessed. Representative plates are shown. The number of colonies on each plate was calculated using Image J. The first group was set as the control group, and served as the baseline for colony number normalization. (D) SW1990 cells were transduced with control shRNA or shRNA against HOTTIP as indicated. Cells (3 × 10 5 ) were then subcutaneously injected into mice, and gemcitabine or PBS were injected intraperitoneally once every 3 days for five cycles. Representative images of tumor-bearing mice are shown. (E) Images of tumors from all mice in each group. (F) Tumor volumes were measured on the indicated days. (G) Tumor weights were determined. In vitro data are represented as the mean ± s.d. from three independent experiments. **p < 0.01, Student’s t-test.

Article Snippet: The human pancreatic cancer cell lines, PANC-1, MIA PaCa-2, Capan-2, SW1990, and BxPC-3, were purchased from the American Type Culture Collection and grown in complete growth medium as recommended by the manufacturer, supplemented with 10% FBS and 1% penicillin/streptomycin.

Techniques: Knockdown, In Vitro, In Vivo, Infection, Control, shRNA, CCK-8 Assay, Transduction, Injection

Journal: PLoS ONE

Article Title: The WSB1 Gene Is Involved in Pancreatic Cancer Progression

doi: 10.1371/journal.pone.0002475

Figure Lengend Snippet: Microarray signal intensities recorded for WSB1 (arbitrary units)

Article Snippet: The human pancreatic cancer-derived Mia-PaCa2, BxPC3, Panc-1, Capan-1 and Capan-2, and the Phoenix Amphotropic viral packaging cell lines were cultivated as recommended by American Type Culture Collection.

Techniques: Microarray

Journal: PLoS ONE

Article Title: The WSB1 Gene Is Involved in Pancreatic Cancer Progression

doi: 10.1371/journal.pone.0002475

Figure Lengend Snippet: Expression of WSB1 isoforms 1+2 and 3 mRNAs was assessed by qRT-PCR in pancreatic cancer cell lines and in mice xenografted tumors (A), in human pancreatic cancer samples (B), and in Mia-PaCa2 cells after treatment with stress-inducing agents (C). Values are expressed as the mean±S.D. of combined results from two independent experiments performed in triplicate. (*, p<0.05).

Article Snippet: The human pancreatic cancer-derived Mia-PaCa2, BxPC3, Panc-1, Capan-1 and Capan-2, and the Phoenix Amphotropic viral packaging cell lines were cultivated as recommended by American Type Culture Collection.

Techniques: Expressing, Quantitative RT-PCR

Journal: PLoS ONE

Article Title: The WSB1 Gene Is Involved in Pancreatic Cancer Progression

doi: 10.1371/journal.pone.0002475

Figure Lengend Snippet: A. Mia-PaCa2 cells were transfected with a siRNA directed against WSB1 mRNA and expression of WSB1 isoforms 1+2+3, 1+2 and 3 mRNAs was analyzed by qRT-PCR. B. Mia-PaCa2 cells were transfected with the WSB1 siRNA and growth was analyzed by direct cell counting (B), MTT analysis (C) or by BrdU incorporation (D) as described in Material and Methods section. E. Mia-PaCa2 cells were transfected with the WSB1 siRNA and caspase 3 activity was measured in untreated and after treatment with staurosporine or doxorubicin. Values are expressed as the mean±S.D. of combined results from two independent experiments performed in triplicate. (*, p<0.05).

Article Snippet: The human pancreatic cancer-derived Mia-PaCa2, BxPC3, Panc-1, Capan-1 and Capan-2, and the Phoenix Amphotropic viral packaging cell lines were cultivated as recommended by American Type Culture Collection.

Techniques: Transfection, Expressing, Quantitative RT-PCR, Cell Counting, BrdU Incorporation Assay, Activity Assay

Journal: PLoS ONE

Article Title: The WSB1 Gene Is Involved in Pancreatic Cancer Progression

doi: 10.1371/journal.pone.0002475

Figure Lengend Snippet: A. Mia-PaCa2 cells were transduced with retrovirus expressing WSB1 isoforms 1, 2 or 3 as EGFP fusion protein or EGFP alone as a control and selected with puromycin. Cell growth was measured by direct counting after plating 10 5 cells in 48-well dishes every day during 4 days. B. After synchronization with 24 h serum-starvation, cells were incubated in serum containing medium for 24 h, stained with propidium iodide and cell cycle was analysed with a flow cytometer. C. S/G1 ratio. Values are expressed as the mean±S.D. of combined results from two independent experiments performed in triplicate (A) or trice in triplicate (B and C) (*, p<0.05).

Article Snippet: The human pancreatic cancer-derived Mia-PaCa2, BxPC3, Panc-1, Capan-1 and Capan-2, and the Phoenix Amphotropic viral packaging cell lines were cultivated as recommended by American Type Culture Collection.

Techniques: Transduction, Expressing, Incubation, Staining, Flow Cytometry

Journal: PLoS ONE

Article Title: The WSB1 Gene Is Involved in Pancreatic Cancer Progression

doi: 10.1371/journal.pone.0002475

Figure Lengend Snippet: Mia-PaCa2 cells were transduced with retrovirus expressing WSB1 isoforms 1, 2 or 3 as EGFP fusion protein or EGFP as control and selected with puromycin. Cells were treated with gemcitabine (100 and 150 µM) for 48 hours or with doxorubicin (150 and 350 µM) for 6 hours. Cell viability was measured by MTT. Values are expressed as the mean±S.D. of combined results from two independent experiments performed in triplicate. (*, p<0.05).

Article Snippet: The human pancreatic cancer-derived Mia-PaCa2, BxPC3, Panc-1, Capan-1 and Capan-2, and the Phoenix Amphotropic viral packaging cell lines were cultivated as recommended by American Type Culture Collection.

Techniques: Transduction, Expressing

Journal: Cell reports

Article Title: MIRO2 promotes cancer invasion and metastasis via MYO9B suppression of RhoA activity

doi: 10.1016/j.celrep.2024.115120

Figure Lengend Snippet:

Article Snippet: Human: PANC-1 , ATCC , Cat#CRL-1469; RRID:CVCL_0480.

Techniques: Control, Plasmid Preparation, Virus, Luciferase, Microarray, Recombinant, Viability Assay, CyQUANT Assay, Proliferation Assay, Activation Assay, Binding Assay, Software

Journal: Journal for Immunotherapy of Cancer

Article Title: Oncolytic peptide LTX-315 induces anti-pancreatic cancer immunity by targeting the ATP11B-PD-L1 axis

doi: 10.1136/jitc-2021-004129

Figure Lengend Snippet: ATP11B interacts with PD-L1 in a CMTM6-dependent manner. (A–B) ATP11B interaction with PD-L1 in vivo. Cell lysates from KPC and SW1990 cells were separately subjected to immunoprecipitation and western blotting using the indicated antibodies. (C) Lack of binding of ATP11B to PD-L1 in vitro. FLAG-tagged ATP11B and MYC-tagged PD-L1 purified from 293 T cell lysates and subjected to immunoprecipitation after co-incubation. (D) Positive correlation of ATP11B with CMTM6. Correlation of ATP11B and CMTM6 analyzed according to the pancreatic data sets of The Cancer Genome Atlas. (E–G) Representative images (E–F) and statistical results (G) of CMTM6 and ATP11B immunohistochemical staining in pancreatic cancer tissue microarray. (H) Western blotting of ATP11B, CMTM6, and PD-L1 expression in paired clinical tissue samples. (I–M) Interaction of ATP11B with PD-L1 in pancreatic cancer in a CMTM6-dependent manner. (I−J) cell lysates from KPC and SW1990 cells were separately subjected to immunoprecipitation and western blotting using the indicated antibodies. (K) Western blotting of ATP11B expression in pancreatic cancer cell lines after CMTM6 knockdown. (L–M) Cell lysates from CMTM6-KD KPC (L) and CMTM6-KD SW1990 (M) separately subjected to immunoprecipitation and western blotting using the indicated antibodies. CMTM6, CKLF-like MARVEL transmembrane domain containing 6; PD-L1 programmed cell death ligand 1.

Article Snippet: KPC, BXPC-3, and SW1990 cells were stably transfected or co-transfected with human ATP11B Double Nickase Plasmid (sc-411692-NIC, Santa Cruz), and ATP11B Double Nickase Plasmid (sc-429296-NIC, Santa Cruz) and KPC and SW1990 were stably transfected with CMTM6 Double Nickase Plasmid (sc-412381-NIC, Santa Cruz) and CMTM6 Double Nickase Plasmid (sc-426447-NIC, Santa Cruz) according to the manufacturer’s instructions.

Techniques: In Vivo, Immunoprecipitation, Western Blot, Binding Assay, In Vitro, Purification, Incubation, Immunohistochemical staining, Staining, Microarray, Expressing

Journal: Journal for Immunotherapy of Cancer

Article Title: Oncolytic peptide LTX-315 induces anti-pancreatic cancer immunity by targeting the ATP11B-PD-L1 axis

doi: 10.1136/jitc-2021-004129

Figure Lengend Snippet: ATP11B prevents lysosomal degradation of PD-L1 through the interaction with CMTM6. (A–B) regulation of PD-L1 expression by ATP11B via lysosome-mediated degradation. (A) PD-L1 expression was analyzed by western blotting in WT and ATP11B KO KPC treated with or without the proteasome inhibitor MG132. (B) PD-L1 expression was analyzed by western blotting in WT and ATP11B KPC KO treated with or without the lysosome inhibitor aloxistatin and pepstatin A. (C–F) regulation of CTMT6 expression by ATP11B in pancreatic cancer. CMTM6 expression was analyzed by western blotting in pancreatic cancer cell lines overexpressing ATP11B (C–D) and ATP11B KD/KO pancreatic cell lines (E–F). (G–K) Rescues of the decrease of PD-L1 by CMTM6 overexpression caused by ATP11B kD. Western blotting (G–I) and flow cytometry (J–K) analysis of PD-L1 expression in WT/ATP11B kD pancreatic cancer cell lines with or without CMTM6 overexpression. CMTM6, CKLF-like MARVEL transmembrane domain containing 6; KD, knockdown; KO, knockout; PD-L1, programmed cell death ligand 1; WT, wild type.

Article Snippet: KPC, BXPC-3, and SW1990 cells were stably transfected or co-transfected with human ATP11B Double Nickase Plasmid (sc-411692-NIC, Santa Cruz), and ATP11B Double Nickase Plasmid (sc-429296-NIC, Santa Cruz) and KPC and SW1990 were stably transfected with CMTM6 Double Nickase Plasmid (sc-412381-NIC, Santa Cruz) and CMTM6 Double Nickase Plasmid (sc-426447-NIC, Santa Cruz) according to the manufacturer’s instructions.

Techniques: Expressing, Western Blot, Over Expression, Flow Cytometry, Knock-Out

Journal: Frontiers in Pharmacology

Article Title: A senescence-associated signature refines the classification of different modification patterns and characterization of tumor immune microenvironment infiltration in triple-negative breast cancer

doi: 10.3389/fphar.2023.1191910

Figure Lengend Snippet: The hub gene FAM3b and its prognostic value in TNBC. (A) Number of trees showing the importance proportion of SASP regulator genes. (B) FAM3B expression in paired tumor and normal tissues in BRCA from the TCGA database. (C) FAM3B expression according to the molecular subtypes of breast cancer. (D) UMAP plot of intratumoral immune cells and FAM3B showing the correlation between infiltration of different immune cells and FAM3B expression in the Alex, EMTAB8107, GSE150660 and GSE161529 datasets. (E,F) The impact of FAM3B on OS and DFS in breast cancer tissue microarray using Kaplan-Meier analysis.

Article Snippet: IHC staining of FAM3B protein expression in the tissue microarray was performed by incubation with rabbit polyclonal antibodies against human FAM3B antibody (27131-1-AP, Proteintech, 1:200) overnight, followed by incubation with goat monoclonal antibody against rabbit antibody (111-035-003, JACKSON, 1:1,000) for 1 h at room temperature.

Techniques: Expressing, Microarray