5 fluorouracil

Manufactured by Selleck Chemicals

Sourced in United States, China

5-fluorouracil is a chemical compound that is commonly used in laboratory settings. It is a fluorinated pyrimidine analog that inhibits the synthesis of DNA and RNA, thereby disrupting cellular processes. This product is primarily used for research purposes in various fields, such as biochemistry, molecular biology, and pharmacology.

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Lab products found in correlation

31 protocols using 5 fluorouracil

Cell Viability Assay with 5-FU and Cisplatin

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MTT assays were performed to examine the viability of cells. Briefly, cells were diluted and seeded at 1×10⁴ cells/100 µl in 96-well plates for 12 h for attachment, and then incubated with 5-fluorouracil (5-FU; Selleck Chemicals) or cisplatin (Selleck Chemicals) at various concentrations for 48 h at 37°C with 5% CO₂. Absorbance measurements following DMSO resolution were performed at a wavelength of 490 nm (Bio-Rad Laboratories, Inc.).

Guo B., Wang L., Qin X., Shen Y, & Ma C. (2019). Jumonji domain-containing protein 6 functions as a marker of head and neck squamous cell carcinoma at advanced stage with no effect on prognosis. Oncology Letters, 18(6), 5843-5852.

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Culturing HUVECs, HIECs, and HCT116 Cells

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Human umbilical vein endothelial cells (HUVECs) were obtained from the Institute of Biological Sciences, Chinese Academy of Sciences (Shanghai). HUVECs were kept in RPMI-1640 medium from Invitrogen in Gaithersburg, Maryland, which also contained 10% fetal bovine serum, 2 mM l-glutamine, 100 U/mL penicillin, and 100 mg/mL streptomycin (complete medium). Human intestinal epithelial cells (HIECs) were supplied by Guangzhou Jennio Biotech Co., Ltd. Both HIEC and HCT116 (ATCC #CCL-247) cell lines were cultured in DMEM (Dulbecco’s modified Eagle’s medium, Thermo Fisher Scientific) containing 10% FBS at 37 °C in a humidified environment consisting of 95% air and 5% CO₂. All cell lines were passaged for fewer than 6 months after being revived. Artesunate (S2265), 5-fluorouracil (S1209), and insulin (S6955) were supplied by Selleck. H₂O₂ (10011208) was purchased from Wokai Biotechnology (Shanghai, China). They were reconstituted in dimethyl sulfoxide (DMSO) and kept at 20 °C for each experiment. The ultimate 5-FU concentration employed was 10 mmol/L in all experiments.

Xia J., Dai Q.L., He S., Jia H.J., Liu X.G., Hua H., Zhou M, & Wang X. (2023). Artesunate alleviates 5-fluorouracil-induced intestinal damage by suppressing cellular senescence and enhances its antitumor activity. Discover. Oncology, 14, 139.

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Chemical Compounds Characterization and Procurement

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Gemcitabine (purity > 98%), cycloheximide (purity > 93%) and MTT (purity > 98%) were purchased from Sigma-Aldrich (St. Louis, USA). LY294002 (purity > 98%) was purchased from Beyotime Institute of Biotechnology (Shanghai, China). Digoxin (purity > 97%), etoposide (purity > 99%), paclitaxel (purity > 99%), cisplatin (purity > 99%), 5-Fluorouracil (5-FU, purity > 99%), cytarabine (ara-C, purity > 99%), doxorubicin (purity > 99%) and MG132 (purity > 97%) were purchased from Selleck Chemicals (Houston, USA). Actinomycin D (purity > 95%) was purchased from KeyGen (Nanjing, China). Anti-NQO1, anti-HO-1 and anti-GCLC antibodies were obtained from Santa Cruz Biotechnology (Texas, USA). Anti-Keap1, anti-Nrf2, anti-p-Akt, anti-Akt, anti-p-P38 and anti-P38, anti-p-ERK1/2, anti-ERK1/2, anti-p-JNK and anti-JNK antibodies were purchased from Cell Signaling Technology (Danvers, USA). Anti-ABCC1 and anti-ABCC5 antibodies were purchased from ABclonal (Wuhan, China). Anti-β-actin and anti-ubiquitin antibodies were obtained from Bioworld (Minnesota, USA). Anti-lamin A antibody was obtained from Sigma-Aldrich (St. Louis, USA).

Zhou Y., Zhou Y., Yang M., Wang K., Liu Y., Zhang M., Yang Y., Jin C., Wang R, & Hu R. (2019). Digoxin sensitizes gemcitabine-resistant pancreatic cancer cells to gemcitabine via inhibiting Nrf2 signaling pathway. Redox Biology, 22, 101131.

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Multi-drug Tumor Slice Culture Assay

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Stock solutions of irinotecan (Selleck Chemicals), oxaliplatin (Selleck Chemicals) and 5-fluorouracil (Selleck Chemicals) were prepared in DMSO (Sigma-Aldrich) and stored in aliquots. Prior to drug treatment, working solutions were prepared fresh by diluting the stock solutions with medium to a final concentration of 1 μg/ml 5-fluorouracil, 1 μg/ml oxaliplatin and 2 μg/ml irinotecan. Tumor slices were treated with 1 μg/ml 5-fluorouracil in combination with 1 μg/ml oxaliplatin in the FX group or 1 μg/ml 5-fluorouracil in combination with 2 μg/ml irinotecan in the FI group. Control group consisted of slices treated with 0.2% DMSO in medium. Multi-well plates containing slices were set on the PS-3D fixed tilt 3D platform rotator (Grant Instruments) for a smooth motion and incubated at 37°C for 72 hours. Every treatment group or control consisted of at least three slices treated similarly in different wells. Consecutive slices were shown to be ‘identical’ biologic replicates,¹⁷ (link) and they were evenly distributed across treatment groups to maintain equal tumor representation for each group to account for intra-tumor heterogeneity. Media were replaced every 24 hours with freshly prepared treatment or control media in relevant groups.

Jabbari N., Kenerson H.L., Lausted C., Yan X., Meng C., Sullivan K.M., Baloni P., Bergey D., Pillarisetty V.G., Hood L.E., Yeung R.S, & Tian Q. (2020). Modulation of Immune Checkpoints by Chemotherapy in Human Colorectal Liver Metastases. Cell Reports Medicine, 1(9), 100160.

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Anti-LGR5 Antibody-Drug Conjugate Generation

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The anti-LGR5 ADC (anti–LGR5-mc-vc-PAB-MMAE) with drug-antibody ratio of 4 was generated as described (3 (link)). Irinotecan and 5-fluorouracil were purchased from Selleck and Acros Organics, respectively. Stattic was purchased from Tocris, Cryptotanshinone and Gefitinib from Selleck, Crizotinib from Cell Signaling, and XAV939 from Cayman Chemical. The plasmid encoding myc-LGR5 was generated previously (9 (link)). Constitutively active mouse STAT3 (STAT3-CA) plasmid Stat3-C Flag pRc/CMV was a gift from Jim Darnell (Addgene, 8722).

Posey T.A., Jacob J., Parkhurst A., Subramanian S., Francisco L.E., Liang Z, & Carmon K.S. (2023). Loss of LGR5 through therapy-induced downregulation or gene ablation is associated with resistance and enhanced MET-STAT3 signaling in colorectal cancer cells. Molecular cancer therapeutics, 22(5), 667-678.

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Evaluation of Combination Therapies for CR-CSphCs

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CR-CSphCs were treated with 5-fluorouracil 10 μM (Selleckchem) and oxaliplatin 10 μM (Sigma-Aldrich), NORA234 (0,3 μM), and LY2603618 0,1μM (rabusertib, Selleckchem) alone or in combination. All the compounds were replenished in culture media every 48 h. To determine the IC50 or drug-combination efficiency of LY2603618 and NORA234, 6×10³ CR-CSphCs were seeded in 96-well plates and, after 24 h, treated with different concentrations (vehicle, 0.01 μM, 0.1 μM, 1 μM, 10 μM) up to 96 h. For drug screening, CR-CSphCs were treated with vehicle or different neo-synthetic alkaloid compounds at the indicated concentrations (Vehicle, 0.03 μM, 0.3 μM, 3 μM).

Di Franco S., Parrino B., Gaggianesi M., Pantina V.D., Bianca P., Nicotra A., Mangiapane L.R., Lo Iacono M., Ganduscio G., Veschi V., Brancato O.R., Glaviano A., Turdo A., Pillitteri I., Colarossi L., Cascioferro S., Carbone D., Pecoraro C., Fiori M.E., De Maria R., Todaro M., Screpanti I., Cirrincione G., Diana P, & Stassi G. (2021). CHK1 inhibitor sensitizes resistant colorectal cancer stem cells to nortopsentin. iScience, 24(6), 102664.

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Cell Cycle Analysis of 5-FU and Radiation

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Cells were cultured in 6-well plates in the presence or absence of 5-fluorouracil (Selleck, Shanghai, China) and irradiation (2 Gy) treatment (CT/RT). The 5-fluorouracil was dissolved in dimethyl sulfoxide (DMSO) (Millipore Sigma). After a 48 h treatment, cells were collected and resuspended, then stained with propidium iodide (PI) (Dojindo) for 15 min, followed by flow cytometric analysis.

Chen H., Yin L., Yang J., Ren N., Chen J., Lu Q., Huang Y., Feng Y., Wang W., Wang S., Liu Y., Song Y., Li Y., Jin J., Tan W, & Lin D. (2023). Genetic polymorphisms in genes regulating cell death and prognosis of patients with rectal cancer receiving postoperative chemoradiotherapy. Cancer Biology & Medicine, 20(4), 297-316.

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Organoid-Based Cytotoxicity Assay

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Organoids were dissociated as described in the previous section. Ten microliters
of dissociated cells/BME (1000 cells/µL) was seeded in each well of a 96-well
plate (Corning Costar, Oneonta, NY; 3904). After 4 days of culturing, organoids
were treated with different doses of staurosporine (ST; Sigma-Aldrich; 569396),
irinotecan (IR; Sigma-Aldrich; I1406), 5-fluorouracil (5-FU; Selleck Chemicals,
Houston, TX; S1209), or SN-38 (Sigma; 7-ethyl-10-hydroxycamtothecin, H0165).
DRAQ7 (Abcam, Cambridge, MA; ab109202) was added to each well at 5 µM final
concentration 1 h prior to imaging. Drugs and dye were refreshed at 3 days after
treatments.

Kim S., Choung S., Sun R.X., Ung N., Hashemi N., Fong E.J., Lau R., Spiller E., Gasho J., Foo J, & Mumenthaler S.M. (2020). Comparison of Cell and Organoid-Level Analysis of Patient-Derived 3D Organoids to Evaluate Tumor Cell Growth Dynamics and Drug Response. Slas Discovery, 25(7), 744-754.

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MTT Assay for Tumor Cell Growth Inhibition

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The growth inhibitory effect of different compounds on tumor cells was examined by MTT assay [41 (link)]. Cells were seeded on a 96-well plate at a final density of 1 × 10⁵ cells/well and cultured. Subsequently, the supernatant was discarded, and cells were incubated in dulbecco’s modified eagle medium (DMEM) medium containing different concentrations of compounds (10, 100, 1000, 5000 and 10,000 nM) for 72 h. 20 μL of MTT (5 mg/mL) was added to each well. The supernatant of each pore was discarded. Adding 150 μL DMSO to each pore, the supernatant was shaken for 10 min. The OD₅₇₀ value was determined by the enzyme-labeled instrument after crystallization was completely dissolved. The IC₅₀ values were calculated by GraphPad Prism 7.0. 5-Fluorouracil (Selleck Chemicals, Houston, TX, USA) was used as a positive control.

Wu D.L., Liao Z.D., Chen F.F., Zhang W., Ren Y.S., Wang C.C., Chen X.X., Peng D.Y, & Kong L.Y. (2019). Benzophenones from Anemarrhena asphodeloides Bge. Exhibit Anticancer Activity in HepG2 Cells via the NF-κB Signaling Pathway. Molecules, 24(12), 2246.

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Inhibition of SIRT1 by EX-527 in 5-FU Treatment

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5-Fluorouracil (2,4-dihydroxy-5-fluoropyrimidine, 5-FU) and the SIRT1 inhibitor EX-527 were purchased from Selleck Chemicals (Houston, TX, USA). EX-527 inhibited SIRT1 in a concentration-dependent manner with an IC50 of 38 nM, but it has much lower potency against SIRT2 (IC50, 19.6 μM) or SIRT3 (IC50, 48.7 μM) [16 (link)], and therefore, only SIRT1 expression was detected in this study. The following antibodies were used in immunohistochemical staining: USP22, SIRT1, MRP3 and Cyclin B1; all the antibodies were from Abcam (Cambridge, MA, USA).

Wen X., Ling S., Wu W., Shan Q., Liu P., Wang C., Wei X., Ding W., Teng X, & Xu X. (2019). Ubiquitin-Specific Protease 22/Silent Information Regulator 1 Axis Plays a Pivotal Role in the Prognosis and 5-Fluorouracil Resistance in Hepatocellular Carcinoma. Digestive Diseases and Sciences, 65(4), 1064-1073.

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