Journal: Chemmedchem

Article Title: Borcalein: a Carborane‐Based Analogue of Baicalein with 12‐Lipoxygenase‐Independent Toxicity

doi: 10.1002/cmdc.202100588

Figure Lengend Snippet: Selectivity index values of borcalein and baicalein. The selectivity index was calculated as IC 50 MRC‐5/IC 50 tested compound.

Article Snippet: Cultivation of the cells : Human melanoma A375, mouse melanoma B16 and B16F10, human colon cancer HCT116 and SW480, mouse colon cancer CT26CL25 and non‐malignant MRC‐5 cell lines were purchased from American Type Culture Collection (ATCC, Rockville, MD, USA).

Techniques:

Journal: Chemmedchem

Article Title: Borcalein: a Carborane‐Based Analogue of Baicalein with 12‐Lipoxygenase‐Independent Toxicity

doi: 10.1002/cmdc.202100588

Figure Lengend Snippet: Mode of cytotoxicity of borcalein in comparison to baicalein. CT26CL25 cells were treated with IC 50 doses of borcalein and baicalein for 48 h and then subjected to: ( A ) CFSE assay, ( B ) annexin/propidium iodide, ( C ) apostat, and ( D ) acridine orange staining. Cells were analysed by flow cytometry, one representative experiment out of three is shown.

Article Snippet: Cultivation of the cells : Human melanoma A375, mouse melanoma B16 and B16F10, human colon cancer HCT116 and SW480, mouse colon cancer CT26CL25 and non‐malignant MRC‐5 cell lines were purchased from American Type Culture Collection (ATCC, Rockville, MD, USA).

Techniques: Comparison, CFSE Assay, Staining, Flow Cytometry

Journal: Frontiers in Pharmacology

Article Title: Natural Polysaccharides and Their Derivates: A Promising Natural Adjuvant for Tumor Immunotherapy

doi: 10.3389/fphar.2021.621813

Figure Lengend Snippet: The immunoregulatory activity of natural polysaccharides and their derivates on immune cells.

Article Snippet: , C57BL/6 (6 weeks old), BALB/c, OT-I and OT-II TCR transgenic mice and C57BL/6-Ly5.1 (CD45.1) congenic mice; TLR2, TLR4 and SR-A-KO mice; the murine melanoma cell line B16F10 (ATCC, CRL-6475) expressing OVA (B16-OVA) and murine carcinoma cell line CT26 (ATCC, CRL-2639) , Increase levels of co-stimulatory molecule expression and pro-inflammatory cytokine production in spleen DCs dependent on TLR4; enhance ovalbumin (OVA) antigen (Ag)-specific immune activation in tumor-bearing mice , .

Techniques: Activity Assay, Functional Assay, Expressing, Activation Assay, Membrane, Gene Expression, In Vivo, Protein-Protein interactions, Concentration Assay, Transgenic Assay, In Vitro, Phospho-proteomics, Isolation, Cell Culture, Modification, Control, Algae

Journal: Cancer immunology research

Article Title: Human GUCY2C-targeted chimeric antigen receptor (CAR)-expressing T cells eliminate colorectal cancer metastases

doi: 10.1158/2326-6066.CIR-16-0362

Figure Lengend Snippet: (A) Recombinant 5F9 antibody was assessed by ELISA for specific binding to hGUCY2CECD or BSA (negative control) plated at 1 μg/mL. Two-way ANOVA; ****p<0.0001. (B) Flow cytometry analysis was performed on parental CT26 mouse colorectal cancer cells or CT26 cells engineered to express hGUCY2C (CT26.hGUCY2C) and stained with 5F9 antibody. (C) Schematic of the third-generation murine CAR construct containing murine sequences of the BiP signal sequence, 5F9 scFv, CD8α hinge region, the transmembrane and intracellular domain of CD28, the intracellular domain of 4-1BB (CD137), and the intracellular domain of CD3ζ (5F9.m28BBz). The CAR construct was inserted into the MSCV retroviral plasmid pMIG upstream of an IRES-GFP marker. (D) Murine CD8+ T cells transduced with a retrovirus containing a control (1D3.m28BBz) CAR or CAR derived from the 5F9 antibody (5F9.m28BBz) were labeled with purified 6xHis-hGUCY2CECD (10 μg/mL), detected with anti-5xHis-Alexa Fluor 647 conjugate. Flow plots were gated on live CD8+ cells. (E) 6xHis-hGUCY2CECD binding curves for 5F9-derived or control (1D3) CARs, gated on live CD8+GFP+ cells (Supplementary Fig. S5). Combined from 3 independent experiments.

Article Snippet: Cell lines and reagents CT26 and β-galactosidase–expressing CT26.CL25 mouse colorectal cancer cell lines and the human colorectal cancer cell lines T84 and SW480 were obtained from ATCC and large stocks of low-passage cells were cryopreserved.

Techniques: Recombinant, Enzyme-linked Immunosorbent Assay, Binding Assay, Negative Control, Flow Cytometry, Staining, Construct, Sequencing, Retroviral, Plasmid Preparation, Marker, Transduction, Control, Derivative Assay, Labeling, Purification

Journal: Cancer immunology research

Article Title: Human GUCY2C-targeted chimeric antigen receptor (CAR)-expressing T cells eliminate colorectal cancer metastases

doi: 10.1158/2326-6066.CIR-16-0362

Figure Lengend Snippet: (A–E) Murine CD8+ T cells were left non-transduced (None) or transduced with control 1D3.m28BBz or 5F9.m28BBz CAR constructs as indicated. (A) Gating strategy for all analyses in B–D. (B) Representative CAR-T cell phenotyping plot based on CD45RA and CD62L. Two-way ANOVA; NS: not significant; Bars: mean ± SD from 2–3 independent experiments; Tn/scm: naïve or T memory stem cells; Tcm: central memory T cells; Tem: effector memory T cells; Temra: effector memory T cells expressing CD45RA. (C–D) 106 CAR-T cells were stimulated for 6 hours with plate-coated antigen (BSA or hGUCY2C) or PMA and ionomycin (PMA/IONO). T-cell activation markers (CD25, CD69, or CD44) and intracellular cytokine production (IFNγ, TNFα, IL2, and MIP1α) were then quantified by flow cytometry. Graphs indicate the mean ± SD (C) activation marker upregulation (MFI) and (D) polyfunctional cytokine production (% of CAR+ cells) from 3 independent experiments. (E) Parental CT26 or CT26.hGUCY2C mouse colorectal cancer cells in an E-Plate were treated with CAR-T cells (5:1 E:T ratio), media, or 10% Triton-X 100 (Triton), and the relative electrical impedance was quantified every 15 minutes for 10 hours to quantify cancer cell death (normalized to time=0). Percent specific lysis values were calculated using impedance values following the addition of media and Triton for normalization (0% and 100% specific lysis, respectively). Two-way ANOVA, B–E; *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001.

Article Snippet: Cell lines and reagents CT26 and β-galactosidase–expressing CT26.CL25 mouse colorectal cancer cell lines and the human colorectal cancer cell lines T84 and SW480 were obtained from ATCC and large stocks of low-passage cells were cryopreserved.

Techniques: Transduction, Control, Construct, Expressing, Activation Assay, Flow Cytometry, Marker, Lysis

Journal: Cancer immunology research

Article Title: Human GUCY2C-targeted chimeric antigen receptor (CAR)-expressing T cells eliminate colorectal cancer metastases

doi: 10.1158/2326-6066.CIR-16-0362

Figure Lengend Snippet: (A–E) BALB/c mice were injected with 5x105 CT26.hGUCY2C cells via the tail vein to establish lung metastases. Control (4D5.m28BBz) or 5F9.m28BBz CAR constructs were transduced into murine CD8+ T cells. (A) Mice were treated 3 days later with 5 Gy total body irradiation (TBI) followed by 106–107 5F9.m28BBz (N=7–8/group) or 107 control (N=6) CAR-T cells. (B) Mice were treated on day 3 (D3) or day 7 (D7) with 5 Gy TBI followed by 107 control (N=10/group) or 5F9.m28BBz (N=9–10/group) CAR-T cells. (C) Mice were treated on day 7 with 5 Gy TBI followed by 107 control (N=10) or 5F9.m28BBz (N=12) CAR-T cells on day 7 and day 14. (D) Mice treated on day 7 with 5 Gy TBI and PBS or 107 control or 5F9.m28BBz CAR-T cells were sacrificed on day 18, lungs stained with India ink, and tumors/lung enumerated. One-way ANOVA; *p<0.05. (E) Surviving mice from B and C treated with 5F9.m28BBz CAR-T cells or naïve mice were challenged with 5x105 CT26 (N=4–7/group) or CT26.hGUCY2C (N=7/group) cells (re-challenge occurred 16–40 weeks after initial challenge). Log-rank Mantel-Cox test, A–C and E; **p<0.01, ***p<0.001, ****p<0.0001. Up arrows indicate CAR-T cell treatment days. Each panel indicates an independent experiment.

Article Snippet: Cell lines and reagents CT26 and β-galactosidase–expressing CT26.CL25 mouse colorectal cancer cell lines and the human colorectal cancer cell lines T84 and SW480 were obtained from ATCC and large stocks of low-passage cells were cryopreserved.

Techniques: Injection, Control, Construct, Irradiation, Staining