mouse colonic epithelial cell line ct26  (ATCC)

Journal: Journal for immunotherapy of cancer

Article Title: Intraperitoneal administration of a modified vaccinia virus Ankara confers single-chain interleukin-12 expression to the omentum and achieves immune-mediated efficacy against peritoneal carcinomatosis.

doi: 10.1136/jitc-2023-006702

Figure Lengend Snippet: Figure 3 Combination with anti-PD-L1 and dose-dependence of the antitumor efficacy of MVA.scIL-12 against aggressive PCa mouse models. (A) BALB/c mice were challenged i.p. with 2×105 CT26 colon cancer cells. Seven days later, mice were treated i.p. with 5×107 TCID50 of MVA.empty or MVA.scIL-12. C57BL/6 mice were challenged i.p. with 1×106 ID8.Vegf/GFP ovarian cancer cells. Five days later, mice were treated i.p. with 5×107 TCID50 of MVA.empty or MVA.scIL-12. Mice were treated in combination with 200 µg of the antibody anti-PD-L1 by i.p. administrations on days +1, +3, and +6 post-MVA treatment. InVivoMAb polyclonal rat IgG (Isotype) was used as a control. The Kaplan-Meier survival curves are shown. (B) As in A, vectors were administered on days 7, 10, and 13 (CT26) or on days 5, 8, and 11 (ID8.Vegf/GFP). Data are represented as mean±SEM. Log-rank test. *p<0.05; **p<0.01; ****p<0.0001. i.p., intraperitoneal; MVA, modified vaccinia virus Ankara; MVA.scIL-12, MVA encoding scIL-12; PCa, peritoneal carcinomatosis; PD-L1, programmed death-ligand 1; scIL-12, single-chain interleukin 12; TCID50, 50% tissue culture infectious dose.

Article Snippet: The CT26 colon cancer cell line was purchased from American Type Culture Collection (CRL- 2638 cell lines).

Techniques: Control, Modification, Virus

Journal: eLife

Article Title: A novel immunopeptidomic-based pipeline for the generation of personalized oncolytic cancer vaccines

doi: 10.7554/elife.71156

Figure Lengend Snippet: Figure 2. Properties of the peptides eluted from the CT26 tumor model. (A) Unique peptides, 7–13 specimen, and their respective source proteins are reported as finite number and depicted as bar plots. (B) Overall peptides’ amino acid length distribution is shown as function of number (left y-axis) and percentage of occurrence (right y-axis). (C) The eluted 9mers were analyzed in regard to their binding affinity to H2Kd and H2Dd. Binders and not binders were defined in NetMHCpan 4.0 Server (applied rank 2%). (D) Major histocompatibility complex (MHC) I consensus binding motifs. The consensus binding motifs among the eluted 9mers peptides were deconvoluted through Gibbs clustering analysis. The reference motif (according to NetMHCpan motif viewer) is depicted in the upper square. The clusters with the optimal fitness (higher KLD values, orange star) are shown, and the sequence logo is represented.

Article Snippet: Murine colon carcinoma CT26 cell line was purchased from ATCC (ATTC CRL- 2639) and cultured in RPMI- 1640 supplemented with 1% GlutaMAX (Gibco, Invitrogen, Carlsbad, CA), 10% heat- inactivated fetal bovine serum (Gibco), and 1% penicillin- streptomycin (10,000 U/mL) (Gibco).

Techniques: Binding Assay, Immunopeptidomics, Sequencing

Journal: eLife

Article Title: A novel immunopeptidomic-based pipeline for the generation of personalized oncolytic cancer vaccines

doi: 10.7554/elife.71156

Figure Lengend Snippet: Figure 4. Differential expression and HEX analysis for the major histocompatibility complex (MHC) I ligand candidates. (A, B) Differential gene expression profile (DESeq) in CT26 versus medullary thymic epithelial cell (mTEC) (A) and CT26 versus healthy Balb/c colon (B) is depicted as volcano plot of -log10 of p-adj-values versus log2 ratio (fold change). The source proteins of MHC-I ligands from our dataset are marked in red, and the differential expression is considered significant for a fold change of 1.5 and a padj-value of 0.05 (red square). (C) Scatter plot comparing the fold change of the source proteins found statistically overexpressed in both DESeq analysis and the average binding affinity score for both H2Kd and H2Dd allotypes. The values were considered significant for >-log10 0.5 H_average ranks and the third quartile of average fold change (red marked). (D, E) The peptides were stratified based on their binding affinity expressed as -log10 and on the weighted score to prioritize similarity between more central amino acids in the peptide (D) or on the percentage of similarity to viral peptides (E). Binding affinity <50 nM and weighted score and similarity >0.8 were considered as the threshold to select tumor peptides similar to viral epitopes.

Article Snippet: Murine colon carcinoma CT26 cell line was purchased from ATCC (ATTC CRL- 2639) and cultured in RPMI- 1640 supplemented with 1% GlutaMAX (Gibco, Invitrogen, Carlsbad, CA), 10% heat- inactivated fetal bovine serum (Gibco), and 1% penicillin- streptomycin (10,000 U/mL) (Gibco).

Techniques: Quantitative Proteomics, Immunopeptidomics, Gene Expression, Binding Assay

Journal: eLife

Article Title: A novel immunopeptidomic-based pipeline for the generation of personalized oncolytic cancer vaccines

doi: 10.7554/elife.71156

Figure Lengend Snippet: Figure 7. PeptiCRAd improved the tumor growth control in both injected and not injected lesions. (A) A schematic representation of the animal experiment setting is depicted. Immunocompetent Balb/c mice were subcutaneously injected with the syngeneic tumor model CT26 in the left (0.6 × 106 cells) and right flanks (1 × 106). PeptiCRAd was intratumorally administrated four times, 2 days apart. (B) The CT26 tumor growth was followed until the end of the experiment, and the tumor size is presented as the mean ± SEM. Statistically significant difference was assessed with two-way ANOVA (*p<0.05; ***p<0.001; ****p<0.0001; ns, nonsignificant).

Article Snippet: Murine colon carcinoma CT26 cell line was purchased from ATCC (ATTC CRL- 2639) and cultured in RPMI- 1640 supplemented with 1% GlutaMAX (Gibco, Invitrogen, Carlsbad, CA), 10% heat- inactivated fetal bovine serum (Gibco), and 1% penicillin- streptomycin (10,000 U/mL) (Gibco).

Techniques: Control, Injection

Journal: Molecular Therapy Oncolytics

Article Title: A Retroviral Replicating Vector Encoding Cytosine Deaminase and 5-FC Induces Immune Memory in Metastatic Colorectal Cancer Models

doi: 10.1016/j.omto.2017.12.001

Figure Lengend Snippet: Toca 511 in Combination with 5-FC Was Efficacious in a Multifocal Liver Metastasis Model of mCRC (A) Representative radiance intensities (photon/s/mouse) from the PBS and 5-FC treatment group at various time points throughout the study. After liver metastases were established with Toca 511 pre-transduced CT26-Luc cells delivered intrasplenically, each mouse was imaged by IVIS at indicated time points. 5-FC treatment (500 mg/kg, i.p., BID) was initiated on day 13 for 5 days on and 2 days off for a total of six cycles. The control group received PBS (see also Figure S1 ). (B) CT26-Luc bioluminescence average signal intensity at indicated time points (n = 9/group). (C) Kaplan-Meier survival analysis. After liver metastases were established with Toca 511 pre-transduced CT26 cells delivered intrasplenically, PBS or 5-FC treatment was started at day 13 post cell inoculation (n = 9/group). A total of six cycles of PBS or 5-FC treatment were administered and survival was examined out to 90 days. The table is the summary of the survival analysis out to 90 days post tumor cell implantations. (D) Cured mice from survival studies (Toca 511/CT26+5-FC; n = 5) were subcutaneously challenged with wild-type CT26 cells on day 0 (day 90 post initial tumor inoculation) at a cell dose of 5 × 10 5 . As a control, CT26 cells were implanted into naive BALB/cJ mice (n = 9) (*p = 0.028; naive versus Toca 511/CT26+5-FC). *Statistical significance was defined as p < 0.05. Error bars represent SEM.

Article Snippet: The mouse colon carcinoma cell line CT26 (CRL 2638) was purchased from American Type Culture Collection (ATCC) (Manassas, VA).

Techniques: Control

Journal: Molecular Therapy Oncolytics

Article Title: A Retroviral Replicating Vector Encoding Cytosine Deaminase and 5-FC Induces Immune Memory in Metastatic Colorectal Cancer Models

doi: 10.1016/j.omto.2017.12.001

Figure Lengend Snippet: Upon Intrasplenic, Intravenous, or Intraportal Delivery, GFP-Expressing Vector Concentrates in Metastatic Foci in the Liver and Not in Normal Liver Tissue (A) Representative images showing luciferase (CT26-Luc cells) and GFP (Toca GFP vector) signal in livers from mice receiving intrasplenic, intravenous, or intraportal delivery of Toca GFP vector. (B) Flow cytometric analysis of excised metastatic lesions from livers collected 18 days post vector delivery. (C) CT26-Luc tumor-bearing animals received 1 day or 3 days of intravenous delivery of Toca GFP vector. Flow cytometric analysis of excised metastatic lesions from livers collected 14 or 22 days post vector delivery (n = 9/group). Numbers above the columns indicate the average GFP + cells per group. Error bars represent SEM.

Article Snippet: The mouse colon carcinoma cell line CT26 (CRL 2638) was purchased from American Type Culture Collection (ATCC) (Manassas, VA).

Techniques: Expressing, Plasmid Preparation, Luciferase

Journal: Molecular Therapy Oncolytics

Article Title: A Retroviral Replicating Vector Encoding Cytosine Deaminase and 5-FC Induces Immune Memory in Metastatic Colorectal Cancer Models

doi: 10.1016/j.omto.2017.12.001

Figure Lengend Snippet: Intravenous Delivery of Toca 511, Followed by Treatment with 5-FC, Was Efficacious against Colorectal Liver Metastasis (A) Kaplan-Meier survival analysis. Mice inoculated with CT26-Luc cells intrasplenically were then administered Toca 511 i.v. for 5 consecutive days starting on day 3 post cell inoculation. PBS or 5-FC treatment started at day 12 post cell inoculation. Six cycles of PBS or 5-FC treatment were administered, and survival was examined out to 90 days. The table is the summary of the survival analysis out to 90 days post tumor cell implantations. Toca 511 and 5-FC treated animals lived significantly longer compared to Toca 511 and PBS control animals (p = 0.037). (B) Radiance intensities (photon/s/mouse) from representative animals at indicated time points. 5-FC treatment (500 mg/kg, i.p., BID) started on day 13 for 5 days on and 2 days off for 6 cycles. Control group received PBS (see also Figure S2 ). (C) Cured mice from survival studies (Toca 511+5-FC; n = 5) were subcutaneously challenged with wild-type CT26 cells on day 0 (day 90 post initial tumor inoculation) at a cell dose of 5 × 10 5 . As a control, CT26 cells were implanted into naive BALB/cJ mice (n = 9) (*p = 0.04; naive versus Toca 511+5-FC). *Statistical significance was defined as p < 0.05. (D) WBCs, LYMs, NEUs, and PLTs were determined in the PBS and 5-FC treatment groups. Dotted line represents LLN for age-matched mice. Age-matched mouse LLN = WBC, 6 × 10 9 cells/L; LYM, 3.4 × 10 9 cells/L; PLT, 200 × 10 9 cells/L; and NEU, 3.4 × 10 9 cells/L. Error bars represent SEM.

Article Snippet: The mouse colon carcinoma cell line CT26 (CRL 2638) was purchased from American Type Culture Collection (ATCC) (Manassas, VA).

Techniques: Control

Journal: Molecular Therapy Oncolytics

Article Title: A Retroviral Replicating Vector Encoding Cytosine Deaminase and 5-FC Induces Immune Memory in Metastatic Colorectal Cancer Models

doi: 10.1016/j.omto.2017.12.001

Figure Lengend Snippet: Toca 511 and 5-FC Resulted in Concentrated 5-FU within Liver Metastases in a Murine Model of mCRC LC/MS measurement of 5-FC and 5-FU in plasma and tumor from mice with Toca 511 pre-transduced CT26-Luc liver metastases treated for 3.5 days with 5-FC (500 mg/kg, i.p., BID). Numbers above columns indicate average values for each group. Error bars represent SEM.

Article Snippet: The mouse colon carcinoma cell line CT26 (CRL 2638) was purchased from American Type Culture Collection (ATCC) (Manassas, VA).

Techniques: Liquid Chromatography with Mass Spectroscopy, Clinical Proteomics

Journal: Molecular Therapy Oncolytics

Article Title: A Retroviral Replicating Vector Encoding Cytosine Deaminase and 5-FC Induces Immune Memory in Metastatic Colorectal Cancer Models

doi: 10.1016/j.omto.2017.12.001

Figure Lengend Snippet: Systemic 5-FU Treatment Was Not Efficacious in a Mouse Model of Colorectal Liver Metastasis (A) Kaplan-Meier analysis showing the survival of animals with liver metastasis treated with 5-FC (500 mg/kg, i.p., BID) or 5-FU (20 mg/kg, i.p., SID) for 5 consecutive days, followed by 2 days without drug for a total of 4 cycles. Animals received no vector in this experiment. 5-FC, without the presence of vector, has no effect on survival. (B) Hematologic analysis, including values for WBCs, LYMs, and NEUs from animals treated with 5-FC or 5-FU. Animals had established CT26 liver metastases but were not given Toca 511 vector. Error bars represent SEM.

Article Snippet: The mouse colon carcinoma cell line CT26 (CRL 2638) was purchased from American Type Culture Collection (ATCC) (Manassas, VA).

Techniques: Plasmid Preparation

Journal: Molecular Therapy Oncolytics

Article Title: A Retroviral Replicating Vector Encoding Cytosine Deaminase and 5-FC Induces Immune Memory in Metastatic Colorectal Cancer Models

doi: 10.1016/j.omto.2017.12.001

Figure Lengend Snippet: Toca 511 and 5-FC Prolonged Survival and Promoted Long-Term Anti-tumor Immunity in a Brain Metastasis Model of mCRC (A) Kaplan-Meier analysis. BALB/cJ mice received Toca 511 pre-transduced CT26 cells intracranially and were subsequently treated with cycles of 5-FC (500 mg/kg, i.p., BID) (n = 12) or PBS (n = 5). Treatments started at day 10 post cell inoculation and were for 7 days on and 7 days off for four cycles. Survival was examined out to 150 days. (B) Long-term survivors (n = 5) were rechallenged subcutaneously at day 65 post intracranial tumor implant with parental CT26 cells, and tumor growth was monitored over time. As a control, CT26 cells were implanted into naive, age-matched, BALB/cJ mice (n = 5) (*p = 0.0001; naive versus Toca 511+5-FC). (C) Recipient athymic animals received splenocytes from either cured or naive animals and were inoculated subcutaneously with parental CT26 cells and tumor growth was monitored over time (n = 6/group) (*p = 0.04). *Statistical significance was defined as p < 0.05. Error bars represent SEM.

Article Snippet: The mouse colon carcinoma cell line CT26 (CRL 2638) was purchased from American Type Culture Collection (ATCC) (Manassas, VA).

Techniques: Control

Journal: Molecular Therapy Oncolytics

Article Title: A Retroviral Replicating Vector Encoding Cytosine Deaminase and 5-FC Induces Immune Memory in Metastatic Colorectal Cancer Models

doi: 10.1016/j.omto.2017.12.001

Figure Lengend Snippet: Myeloid-Derived Suppressor Cells Decreased with Toca 511 and 5-FC Treatment in a Brain Metastasis Model of mCRC BALB/c mice received Toca 511 pre-transduced CT26 cells intracranially and were treated with 5-FC (500 mg/kg, i.p., BID) or PBS. Treatments started at day 10 post cell inoculation and were 7 days on and 7 days off for one cycle. (A and B) The tumor (A) (*p < 0.0001) and spleen (B) (*p = 0.03) were collected for flow cytometric analysis 14 days post cell implantation. Representative flow analysis from PBS and 5-FC treatment groups separating the populations of Cd11b + Gr-1 + Ly6C + cells in the tumor (A) and spleen (B). Error bars represent SEM.

Article Snippet: The mouse colon carcinoma cell line CT26 (CRL 2638) was purchased from American Type Culture Collection (ATCC) (Manassas, VA).

Techniques: Derivative Assay

Journal: Oncotarget

Article Title: Elevated O-GlcNAcylation promotes colonic inflammation and tumorigenesis by modulating NF-κB signaling

doi:

Figure Lengend Snippet: Wild-type mice were administered 2% DSS in drinking water for the indicated days. (A) Western blots of lysates from wild-type mice treated with DSS during a 9-day time course. Lysates for each time point were isolated from three separate mice. Two representative samples were loaded in each lane. (B) Densitometry was performed on immunoblots. The ratio of O-GlcNAc to β-actin was determined. Error bars represent the S.D. ( n = 3) and (**) P < 0.005, (*) P < 0.05 (Student's t -test). (C) Immunofluorescence staining of the colon with O-GlcNAc mAb (RL2) in wild-type mice not treated or treated with DSS for 7 days (green, O-GlcNAc modified proteins; blue, nuclei). Bars =50 μm. (D) Measurement of Uridine diphosphate-N-acetylglucosamine (UDP-GlcNAc; B) levels in control (n=3) and DSS-treated (n=6) colon tissues by HPLC analysis (left). Matched O-GlcNAcylation levels (Immunoblots with RL2 antibody) (right). Error bars represent ± SEM. (*) P < 0.05 (Student's t -test). (E) Caco2 and CT26 cells were treated with 3% DSS for up to 48 hours. O-GlcNAcylation levels were analyzed by Western blot analysis.

Article Snippet: The human (Caco-2) and mouse (CT26) colonic epithelial cell lines, and RAW264.7 cell lines were purchased from the American Type Culture Collection (ATCC); (Caco2, 2010), (CT26, 2010), and Cell lines were cultured in the recommended ATCC media.

Techniques: Western Blot, Isolation, Immunofluorescence, Staining, Modification, Control

Journal: Oncotarget

Article Title: Elevated O-GlcNAcylation promotes colonic inflammation and tumorigenesis by modulating NF-κB signaling

doi:

Figure Lengend Snippet: (A) CT26 cells were transfected with an NF-κB promoter–reporter construct before LPS (100 ng/mL) and/or Thiamet G (1μM) treatment; luciferase activity was measured at 6 h. (B and C) SYBR Green-based RT-PCR analysis was carried out after treatment with LPS (100 ng/mL) and/or Thiamet G (1 μM) and expression levels of IL-1β and IL-6 were determined by comparison with GAPDH. (D and E) Control and OGA knockdown Caco2 cell lines were stimulated with LPS and the mRNA levels of IL-1β and IL-6 were measured at 6h. (F-H) OGA +/+ and OGA −/− MEFs were stimulated by LPS (100 ng/ml) and the mRNA levels of TNF-α, IL-1β and IL-6 were measured at the indicated times post-stimulation. Error bars represent the S.D. ( n = 3). (**) P < 0.005, (*) P < 0.05 (Student's t -test.

Article Snippet: The human (Caco-2) and mouse (CT26) colonic epithelial cell lines, and RAW264.7 cell lines were purchased from the American Type Culture Collection (ATCC); (Caco2, 2010), (CT26, 2010), and Cell lines were cultured in the recommended ATCC media.

Techniques: Transfection, Construct, Luciferase, Activity Assay, SYBR Green Assay, Reverse Transcription Polymerase Chain Reaction, Expressing, Comparison, Control, Knockdown