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Leflunomide

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Leflunomide is a laboratory reagent used for research purposes. It is a chemical compound that can be used in various experimental setups. The core function of Leflunomide is to serve as a research tool, but its specific applications and intended uses should not be extrapolated.

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Lab products found in correlation

Tetrahydrouridine thu, by Cayman Chemical (2 mentions) Thapsigargin, by Cell Signaling Technology (2 mentions) Dulbecco modified eagle medium (dmem), by Merck Group (2 mentions) Il 13, by Thermo Fisher Scientific (1 mentions) Dimethysulfoxide dmso, by Merck Group (1 mentions) Purecol bovine collagen solution, by Advanced BioMatrix (1 mentions) Williams e medium, by Merck Group (1 mentions) Antibiotic antimycotic, by Thermo Fisher Scientific (1 mentions) Pd184352, by LC Laboratories (1 mentions) Sp600125, by LC Laboratories (1 mentions) Fetal bovine serum (fbs), by Atlanta Biologicals (1 mentions) Uridine, by Merck Group (1 mentions) Ki67 antibody ab1667, by Abcam (1 mentions) Mouse monoclonal anti β actin antibody a1978, by Merck Group (1 mentions) Aicar, by LGC (1 mentions) Dab substrate kit, by Vector Laboratories (1 mentions) Matrigel, by Corning (1 mentions) Bongkrekic acid, by Merck Group (1 mentions) Cell culture media and supplements, by Thermo Fisher Scientific (1 mentions) A77 1726, by Enzo Life Sciences (1 mentions)

7 protocols using leflunomide

1

Ginsenoside Rb1 Regulates Macrophage Activation

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Peritoneal macrophages were pre‐treated with indicated concentrations of Ginsenoside Rb1 (Fleton Natural Products, Chengdu, Sichuan, China) for 1 h prior to the incubation of lipopolysaccharide (LPS; Sigma‐Aldrich®, Merck, Shanghai, China, 1 μg/ml) for indicated time period. To examine the effects of IL‐4 (3 μg/ml; eBioscience®, ThermoFisher Scientific, Shanghai, China) and IL‐13 (3 μg/ml; eBioscience®, ThermoFisher Scientific, Shanghai, China) neutralizing antibodies, the two antibodies were added in medium at the same time with Rb1 (20 μM) for indicated period of time. To address the role of signal factor STAT6, cells were pre‐treated with 50 μM leflunomide (Enzo Life Sciences, Farmingdale, NY, USA), a STAT6 inhibitor, for 12 hrs before exposure to Rb1.
Zhang X., Liu M., Qiao L., Zhang X., Liu X., Dong M., Dai H., Ni M., Luan X., Guan J, & Lu H. (2017). Ginsenoside Rb1 enhances atherosclerotic plaque stability by skewing macrophages to the M2 phenotype. Journal of Cellular and Molecular Medicine, 22(1), 409-416.
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2

Cellular Signaling Pathway Modulation

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Williams’ Medium E, 4-phenylbutyrate acid (4-PBA), and dimethy-sulfoxide (DMSO) were obtained from Sigma-Aldrich (St. Louis, MO). Fetal bovine serum (FBS) was purchased from Atlanta Biologicals (Lawrenceville, GA). Antibiotic-antimycotic was obtained from Life Technologies (Grand Island, NY). PureCol Bovine Collagen Solution was from Advanced BioMatrix (San Diego, CA). Leflunomide was purchased from Enzo Life Sciences (Farmingdale, NY). SP600125 (JNK inhibitor) and PD184352 (ERK1/2 inhibitor) were from LC laboratories (Woburn, MA).
Ren Z., Chen S., Qing T., Xuan J., Couch L., Yu D., Ning B., Shi L, & Guo L. (2017). Endoplasmic reticulum stress and MAPK signaling pathway activation underlie leflunomide-induced toxicity in HepG2 Cells. Toxicology, 392, 11-21.
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3

Investigating Leflunomide and AICAR Effects

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Leflunomide was purchased from Enzo life sciences (ALX-4300–095-G001). AICAR was purchased from Toronto Research Chemicals Inc (North York, Canada). Uridine was purchased from Sigma (St. Louis, MO). Antibodies against LKB1 (3047s), Caspase-3 (9962s), Cleaved caspase-3 (9664s), PARP (9542s), Kras-G12D (14429), p53 (2524s) were purchased from Cell Signaling Technology (Beverly, MA). Ki67-antibody (ab1667) and TTF-1 antibody (ab76013) were purchased from Abcam (Cambridge, UK). DAB substrate kit was purchased from Vector Laboratories (SK-4100, Burlingame, CA). Mouse monoclonal anti-β-actin antibody (a1978) was purchased from Sigma (St. Louis, MO). Matrigel was purchased from Corning (Cat#356255, Corning, NY).
Jin R., Liu B., Liu X., Fan Y., Peng W., Huang C., Marcus A.I., Sica G.L., Gilbert-Ross M., Liu Y, & Zhou W. (2020). Leflunomide Suppresses the Growth of LKB1-inactivated Tumors in the Immune-Competent Host and Attenuates Distant Cancer Metastasis. Molecular cancer therapeutics, 20(2), 274-283.
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4

Validation of Pancreatic Cancer Cell Lines

Cited 1 time
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Pancreatic cancer cell lines Capan-2, T3M4, PATU8902 and CFPAC have been already described (36 (link)). Cell lines were validated by STR profiling by the Genetics Core at the University of Arizona. All cell lines were cultured in Dulbecco’s modified Eagle medium (DMEM) (Sigma-Aldrich, St Louis, MO USA) supplemented with 10% fetal bovine serum (FBS), 100 I.U/ml penicillin, 100 μg/ml streptomycin, and incubated at 37°C in a humidified incubator with 5% CO2. Thapsigargin was purchased from Cell Signaling Technology (Danvers, MA USA) and stocks were diluted in DMSO. Tetrahydrouridine (THU) was purchased from Cayman Chemical (Ann Arbor, MI USA), Leflunomide was from Enzo Life Sciences (Farmingdale, NY USA). Deoxycytidine and deoxyuridine were obtained from MP Biomedical (Irvine, CA USA) and Sigma-Aldrich, respectively.
Olou A.A., King R.J., Yu F, & Singh P.K. (2020). MUC1 Oncoprotein Mitigates ER Stress via CDA-mediated Reprogramming of Pyrimidine Metabolism. Oncogene, 39(16), 3381-3395.
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5

Validation of Pancreatic Cancer Cell Lines

Cited 1 time
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Pancreatic cancer cell lines Capan-2, T3M4, PATU8902 and CFPAC have been already described (36 (link)). Cell lines were validated by STR profiling by the Genetics Core at the University of Arizona. All cell lines were cultured in Dulbecco’s modified Eagle medium (DMEM) (Sigma-Aldrich, St Louis, MO USA) supplemented with 10% fetal bovine serum (FBS), 100 I.U/ml penicillin, 100 μg/ml streptomycin, and incubated at 37°C in a humidified incubator with 5% CO2. Thapsigargin was purchased from Cell Signaling Technology (Danvers, MA USA) and stocks were diluted in DMSO. Tetrahydrouridine (THU) was purchased from Cayman Chemical (Ann Arbor, MI USA), Leflunomide was from Enzo Life Sciences (Farmingdale, NY USA). Deoxycytidine and deoxyuridine were obtained from MP Biomedical (Irvine, CA USA) and Sigma-Aldrich, respectively.
Olou A.A., King R.J., Yu F, & Singh P.K. (2020). MUC1 Oncoprotein Mitigates ER Stress via CDA-mediated Reprogramming of Pyrimidine Metabolism. Oncogene, 39(16), 3381-3395.
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6

Procurement of Small Molecule Compounds

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Leflunomide (≥98% purity) and A77 1726 (≥98% purity) were purchased from Enzo Life Sciences (Farmingdale, NY). Bongkrekic acid, cyclosporine A, and dimethyl sulfoxide (DMSO) were obtained from Sigma-Aldrich (St. Louis, MO). Cell culture media and supplements were purchased from Life Technology (Grand Island, NY) and Atlanta Biologicals (Lawrenceville, GA).
Xuan J., Ren Z., Qing T., Couch L., Shi L., Tolleson W.H, & Guo L. (2018). Mitochondrial dysfunction induced by leflunomide and its active metabolite. Toxicology, 396-397, 33-45.
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7

THP-1 Macrophage Differentiation and Apoptosis

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THP-1 cells were grown to confluence and differentiated into macrophage-like cells using 150 nM of 1α,25-Dihydroxy-Vitamin D3 for 24 hours. Once the cells were semi-adherent, 160 nM of phorbol 12-myristate 13-acatate (PMA) in sterile DMSO was applied to the cells for 30 minutes to initiate maturation from monocyte to macrophage. Following maturation, THP-1 cells were treated for one hour with 50 μM Leflunomide in DMSO, a selective STAT6 inhibitor (Enzo Life Sciences, Farmingdale, NY) [35 (link)] or 4 μM InSolution JAK Inhibitor I in DMSO, a broad JAK/STAT inhibitor (EMD Millipore) [36 (link)]. THP-1 cells were then primed with LPS and exposed to MWCNTs and/or the combination of recombinant IL-4 and IL-13 for 24 hours. Cell supernatants were removed and cells washed with sterile DPBS. After washing, THP-1 cells were exposed to 50mJ UVB to induce apoptosis. Following UVB treatment, fresh media was added and cells were allowed to incubate at 37°C for 16 hours. Apoptotic cells were then collected and lysed, and analysis of caspase-1 activity was performed using the Caspase-1 Colorimetric Assay Kit from R&D Systems (#BF14100) and following the manufacturer’s protocol.
Shipkowski K.A., Taylor A.J., Thompson E.A., Glista-Baker E.E., Sayers B.C., Messenger Z.J., Bauer R.N., Jaspers I, & Bonner J.C. (2015). An Allergic Lung Microenvironment Suppresses Carbon Nanotube-Induced Inflammasome Activation via STAT6-Dependent Inhibition of Caspase-1. PLoS ONE, 10(6), e0128888.
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