anti lig3  (R&D Systems)

Journal: bioRxiv

Article Title: A Novel miR-4745 -KLC2 Axis Regulates Cancer Stem Cell Traits in Colorectal Cancer

doi: 10.64898/2026.01.07.697660

Figure Lengend Snippet: (a) Quantification of the CD44v6-APC-positive population in CPP1 (n=7) and CTC44 (n=5) cells transfected with miR-4745-3p ( miR4745 ) or miRVana microRNA Mimic Negative Control #1 (miCTRL) mimics (right panel). Percentage of CD44v6-APC positive CPP1 cells is indicated in inset boxes for a representative experiment (left panel). (b) Representative images of tumorspheres (upper panel) and the percentage (lower panel) of sphere-forming cells in patient-derived CPP1 and circulating tumor (CTC44) colon cancer cells transfected with miR-4745-3p ( miR4745 ) or miCTRL mimics. Data from individual replicates (n=15 per experiment) are presented, along with the mean ± SEM from six independent experiments. (c) Percentage of sphere-forming cells in patient-derived CPP1 colon cancer cells transfected with miR4745 or negative control (miCTRL) mimics and, maintained as first-generation colonospheres (S1) or after several passages (S2, S3). Data from individual replicates (n=12 per experiment) are presented, along with the mean ± SEM from three independent experiments. (d) Percentage of surviving cancer stem cells (Aldefluor-positive) measured 48 hours following treatment with specified concentrations of FIRI (1X = 50 µM 5-FU + 500 nM SN38) in vitro . Sorted CPP1 Aldefluor-positive cells were first transfected with 50 nM miR-4745-3p ( miR-4745 ) or miRVana microRNA Mimic Negative Control #1 (miCTRL) mimics, 24 hours prior to FIRI treatment. Data are expressed as mean ± SEM (n = 3). (e) Percentage of Aldefluor-positive cells (% ALDH + cells) in CPP1 cells transfected with miR-4745 or miCTRL mimics and then exposed for 72 hours to chemotherapy (Firi=5µM 5-FU + 50nM SN38). Data are expressed as mean ± SEM (n=6). *, p<0.05; **, p<0.005, ***, p < 0.001, Mann Whitney test.

Article Snippet: CD44v6-APC (clone REA706, 130-111-238; Macs Miltenyi) antibody was incubated with 100 000 cells in PBS containing 5% FBS for 20 min at 4°C.

Techniques: Transfection, Negative Control, Derivative Assay, In Vitro, MANN-WHITNEY

Journal: bioRxiv

Article Title: Suppression of Glucosylceramide Synthase Reverses Drug Resistance in Cancer Cells Harbor Homozygous p53 Mutants

doi: 10.1101/2025.11.02.686136

Figure Lengend Snippet: Cells were planted in 5% FBS EMEM medium containing 6 μM IRI alone or combined 4 μM Genz-161 for 6 days. A, Imaging flow cytometry analysis for colon CSCs (CD44v6 + /CD133 + ). BF, bright field. B, GCS inhibition decreased CSC population. *, p <0.001 compared to WiDr cells treated with vehicle; **, p <0.001 compared to WiDr cells treated with IRI alone. C, Representative CSC plots of cancer cells with various treatments.

Article Snippet: After treatments, suspended cells (10 6 cells/ml) were incubated with human CD44v6 Alexa Fluor ® 488-conjugated antibody (2F10; mouse IgG1; from R&D Systems, Minneapolis, MN, USA) and human CD133 APC-conjugated antibody (170411; mouse IgG2b; from R&D Systems) in 1% BSA-containing PBS at 4°C for 45 min. After washing, cells were resuspended in 1% BSA PBS (5 x 10 5 cells/150 μL) and analysed using an Amnis Imagestream Mark II Imagestream software, and the data were further analysed using the IDEAS v6.2 program.

Techniques: Imaging, Flow Cytometry, Inhibition

Journal: bioRxiv

Article Title: Suppression of Glucosylceramide Synthase Reverses Drug Resistance in Cancer Cells Harbor Homozygous p53 Mutants

doi: 10.1101/2025.11.02.686136

Figure Lengend Snippet: A, Tumor growth of mice treated with oxaliplatin. Mice-bearing tumors generated from WiDr or WiDr/UGCG - cells were treated with vehicle, oxaliplatin (Oxa 2 mg/kg, i.p , once every 6 days) and combination (Oxa 2 mg/kg, i.p, once every 6 days and Genz-161 4 mg/kg, i.p, once every 3 days) for 37 days. *, p <0.01 compared with WiDr tumors treated with vehicle or Oxa. B, Tumor growth of mice treated with irenotecan. Mice-bearing tumors generated from WiDr cells were treated with vehicle, irenotecan (IRI 6 mg/kg, i.p , once every 6 days) and combination (IRI 6 mg/kg i.p , once every 6 days and Genz-161 4 mg/kg, i.p, once every 3 days) for 30 days. *, p <0.01 compared with tumors treated with vehicle; **, p<0.01 compared with tumors treated with IRI. C, H&E and immunofluorescence staining of tumors. Tumor sections were stained with H&E or fluorescent antibodies for CSC markers (CD44v6/CD133). Green, Alexa Fluor 448−CD44v6; red, APC-CD133; blue, DAPI nuclear counterstain. Images were magnified 200x, scale bar represents to 50 μm. D, GCS mRNA levels of tumors. *, p <0.01 compared with WiDr-tumors treated with vehicle or oxaliplatin. E, Tumor stem cell clusters in snRNA. Tumor-bearing mice were treated with Oxa (2 mg/kg, i.p, once every 6 days) for 37 days. *, p <0.001 compared with WiDr tumors treated with Oxa.

Article Snippet: After treatments, suspended cells (10 6 cells/ml) were incubated with human CD44v6 Alexa Fluor ® 488-conjugated antibody (2F10; mouse IgG1; from R&D Systems, Minneapolis, MN, USA) and human CD133 APC-conjugated antibody (170411; mouse IgG2b; from R&D Systems) in 1% BSA-containing PBS at 4°C for 45 min. After washing, cells were resuspended in 1% BSA PBS (5 x 10 5 cells/150 μL) and analysed using an Amnis Imagestream Mark II Imagestream software, and the data were further analysed using the IDEAS v6.2 program.

Techniques: Generated, Immunofluorescence, Staining

Journal: bioRxiv

Article Title: Suppression of Glucosylceramide Synthase Reverses Drug Resistance in Cancer Cells Harbor Homozygous p53 Mutants

doi: 10.1101/2025.11.02.686136

Figure Lengend Snippet: Cells were planted in 5% FBS EMEM medium containing 6 μM IRI alone or combined 4 μM Genz-161 for 6 days. A, Imaging flow cytometry analysis for colon CSCs (CD44v6 + /CD133 + ). BF, bright field. B, GCS inhibition decreased CSC population. *, p <0.001 compared to WiDr cells treated with vehicle; **, p <0.001 compared to WiDr cells treated with IRI alone. C, Representative CSC plots of cancer cells with various treatments.

Article Snippet: For detection of colon CSCs, the blocked slides were incubated with CD44v6 Alexa-Fluor 488-conjugated antibody (2F10; mouse IgG1; purchased from R&D Systems, Minneapolis, MN) and CD133/2 APC-conjugated antibody (293C3, mouse IgG2b; purchased from Miltenyi Biotec, San Diego, CA) (1:100).

Techniques: Imaging, Flow Cytometry, Inhibition

Journal: bioRxiv

Article Title: Suppression of Glucosylceramide Synthase Reverses Drug Resistance in Cancer Cells Harbor Homozygous p53 Mutants

doi: 10.1101/2025.11.02.686136

Figure Lengend Snippet: A, Tumor growth of mice treated with oxaliplatin. Mice-bearing tumors generated from WiDr or WiDr/UGCG - cells were treated with vehicle, oxaliplatin (Oxa 2 mg/kg, i.p , once every 6 days) and combination (Oxa 2 mg/kg, i.p, once every 6 days and Genz-161 4 mg/kg, i.p, once every 3 days) for 37 days. *, p <0.01 compared with WiDr tumors treated with vehicle or Oxa. B, Tumor growth of mice treated with irenotecan. Mice-bearing tumors generated from WiDr cells were treated with vehicle, irenotecan (IRI 6 mg/kg, i.p , once every 6 days) and combination (IRI 6 mg/kg i.p , once every 6 days and Genz-161 4 mg/kg, i.p, once every 3 days) for 30 days. *, p <0.01 compared with tumors treated with vehicle; **, p<0.01 compared with tumors treated with IRI. C, H&E and immunofluorescence staining of tumors. Tumor sections were stained with H&E or fluorescent antibodies for CSC markers (CD44v6/CD133). Green, Alexa Fluor 448−CD44v6; red, APC-CD133; blue, DAPI nuclear counterstain. Images were magnified 200x, scale bar represents to 50 μm. D, GCS mRNA levels of tumors. *, p <0.01 compared with WiDr-tumors treated with vehicle or oxaliplatin. E, Tumor stem cell clusters in snRNA. Tumor-bearing mice were treated with Oxa (2 mg/kg, i.p, once every 6 days) for 37 days. *, p <0.001 compared with WiDr tumors treated with Oxa.

Article Snippet: For detection of colon CSCs, the blocked slides were incubated with CD44v6 Alexa-Fluor 488-conjugated antibody (2F10; mouse IgG1; purchased from R&D Systems, Minneapolis, MN) and CD133/2 APC-conjugated antibody (293C3, mouse IgG2b; purchased from Miltenyi Biotec, San Diego, CA) (1:100).

Techniques: Generated, Immunofluorescence, Staining