Flurbiprofen interacts with aldehyde dehydrogenase, specifically mitochondrial aldehyde dehydrogenase (ALDH2) and aldehyde dehydrogenase family 1 member B1 (ALDH1B1). The interaction was identified via affinity purification and confirmed by Western blotting. [3]

In a concentration- and time-dependent manner, flurbiprofen (2–20 nM; 12-48 hours) strongly inhibits the growth of SW620 cells [1]. For 24 hours, flurbiprofen (10 nM) decreases COX-2 expression [1]. By blocking COX-2, flurbiprofen (10 nM; 24 hours) reduces the expression of inflammatory factors [1]. By blocking COX-2, flurbiprofen (10 nM; 24 hours) encourages apoptosis in colorectal cancer cells [1]. An experiment on cell proliferation [1]

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Flurbiprofen (100 µM) exhibited strong chaperone activity, significantly attenuating the aggregation of reduced-lactalbumin and heat-aggregated lysozyme in vitro. This activity was stronger than the positive control 4-phenylbutyrate (4-PBA). [3]
Other NSAIDs (aspirin, ibuprofen, meloxicam at 100 µM) did not exhibit significant chaperone activity in the same assay. [3]
In SH-SY5Y neuroblastoma cells, flurbiprofen (100 µM) significantly inhibited ER stress-induced cell death caused by tunicamycin (1 µg/ml) or brefeldin A, as measured by LDH leakage and crystal violet assays. [3]
Flurbiprofen (100 µM) inhibited the ER stress-induced unfolded protein response (UPR) in SH-SY5Y cells, as evidenced by reduced induction of CHOP mRNA, XBP1 splicing, and HERP mRNA. [3]
In SH-SY5Y-Ob-Rb cells, ER stress impaired leptin-induced STAT3 phosphorylation. Co-treatment with flurbiprofen (100 µM) reversed this impairment and restored nuclear phospho-STAT3 staining. [3]
In HEK293-Ob-Rb cells, flurbiprofen (100 µM) increased aldehyde dehydrogenase enzyme activity. [3]
Knockdown of ALDH2/ALDH1B1 in HEK293-Ob-Rb cells attenuated the ability of flurbiprofen to inhibit ER stress-induced IRE1 phosphorylation and to restore leptin-induced STAT3 phosphorylation under ER stress conditions. [3]

In rats with adrenalectomy, flurbiprofen (0.3–4.8 mg/kg; oral; 4-5 doses) exhibits immediate anti-inflammatory effects [2]. In mice with high-fat diets, flurbiprofen (10 mg/kg; intraperitoneal injection; daily; for 6 days) reduces obesity [3].

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In C57BL/6 mice fed a high-fat diet (HFD) for 8 weeks, oral administration of flurbiprofen (10 mg/kg/day, mixed in drinking water) significantly attenuated HFD-induced body weight gain compared to the HFD control group. This effect was not observed in mice on a normal chow diet. [3]
Flurbiprofen treatment (10 mg/kg/day for 8 weeks) in HFD-fed mice significantly reduced the total weight of visceral fat and inhibited the accumulation of both visceral and subcutaneous adipose tissue, as assessed by computed tomography (CT) scans, without affecting muscle volume. [3]
Flurbiprofen treatment (10 mg/kg/day for 8 weeks) normalized the HFD-induced elevation of circulating leptin levels in mice. [3]
In female ob/ob mice (leptin-deficient), intraperitoneal co-administration of flurbiprofen (10 mg/kg/day) with recombinant leptin (1 mg/kg/day) for 6 days significantly enhanced leptin-induced body weight reduction compared to leptin treatment alone. Flurbiprofen alone had only a slight effect on body weight. [3]
Other NSAIDs (aspirin 10 mg/kg/day, ibuprofen 60 mg/kg/day, meloxicam 1 mg/kg/day) administered orally with HFD did not show a significant anti-obesity effect on body weight gain. [3]

For measurement of aldehyde dehydrogenase activity, HEK293-Ob-Rb cells were harvested and incubated with or without flurbiprofen (100 µM) for 60 minutes. Enzyme activity was then measured using a commercial aldehyde dehydrogenase activity colorimetric assay kit according to the manufacturer's directions. [3]

Cell proliferation experiment [1]
Cell Types: SW620 cell
Tested Concentrations: 2 nM, 4 nM, 10 nM, 20 nM
Incubation Duration: 12 hrs (hours), 24 hrs (hours), 48 hrs (hours)
Experimental Results: Inhibition of colorectal cancer cell proliferation.

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Western Blot Analysis[1]
Cell Types: SW620 Cell
Tested Concentrations: 10 nM
Incubation Duration: 24 hrs (hours)
Experimental Results: The protein and mRNA levels of COX-2 were Dramatically diminished.

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RT-PCR[1]
Cell Types: SW620 Cell
Tested Concentrations: 10 nM
Incubation Duration: 24 hrs (hours)
Experimental Results: diminished COX-2 mRNA expression levels Cell apoptosis analysis[1]
Cell Types: SW620 Cell
Tested Concentrations: 10 nM
Incubation Duration: 24 hrs (hours)
Experimental Results: The expression of Bcl2 was Dramatically diminished, and the expression of Bax and cleaved-caspase3 was Dramatically increased, but there was no effect on total caspase-3.

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For chaperone activity using α-lactalbumin aggregates, the rate of aggregation of reduced-lactalbumin was induced by BSA aggregates and monitored by measuring turbidity at 488 nm in the presence or absence of reagents (e.g., flurbiprofen, 4-PBA, other NSAIDs) using a microplate reader. [3]
For chaperone activity based on heat-induced aggregation of lysozyme, lysozyme was dissolved in phosphate buffer and mixed with flurbiprofen (final concentration 30 mM) or vehicle. Samples were heated at 98°C for 10 minutes. After cooling, aggregated protein was separated by centrifugation, and the concentration of soluble protein in the supernatant was measured using the BCA method. [3]
For chaperone activity based on heat-induced aggregation of ALDH2, recombinant ALDH2 was mixed with flurbiprofen (final concentration 30 mM) or vehicle, heated at 70°C for 10 minutes, cooled, and centrifuged. The soluble protein concentration in the supernatant was measured. [3]
For dynamic light scattering (DLS) analysis, lysozyme with or without flurbiprofen (50 mM) was dissolved, filtered, and injected into an optical cell. DLS measurements were performed before and after heating the sample at 42°C for 5 minutes to analyze changes in the hydrodynamic radius (Rh) of protein aggregates. [3]
For the lactate dehydrogenase (LDH) leakage assay, SH-SY5Y cells were treated with ER stress inducers (tunicamycin or brefeldin A) in the presence or absence of flurbiprofen for 48 hours. Cell culture supernatant was collected, and LDH activity was measured using a cytotoxicity detection kit according to the manufacturer's directions. [3]
For the crystal violet assay, SH-SY5Y cells treated with ER stress inducers and flurbiprofen were washed, fixed, and stained with 0.1% crystal violet. After washing and drying, the stained dye was solubilized with SDS, and absorbance was measured at 590 nm. [3]
For gene expression analysis of UPR markers (CHOP, XBP1 splicing, HERP), SH-SY5Y cells were treated with tunicamycin and flurbiprofen. Total RNA was isolated, reverse transcribed into cDNA, and specific transcripts were amplified by PCR. PCR products were resolved by polyacrylamide gel electrophoresis, stained, and visualized. [3]
For Western blot analysis of STAT3 phosphorylation, SH-SY5Y-Ob-Rb cells were exposed to ER stress inducers and flurbiprofen for 4 hours, then stimulated with leptin. Cell lysates were prepared, proteins separated by SDS-PAGE, transferred to membranes, and probed with anti-phospho-STAT3 and anti-STAT3 antibodies, followed by detection using an enhanced chemiluminescence system. [3]
For immunohistochemistry of nuclear phospho-STAT3, SH-SY5Y-Ob-Rb cells were treated, fixed, permeabilized, blocked, and incubated with an anti-phospho-STAT3 primary antibody overnight at 4°C. After washing, cells were incubated with a fluorophore-conjugated secondary antibody, counterstained with propidium iodide, and visualized by confocal microscopy. [3]
For RNAi experiments, SH-SY5Y, HEK293-Ob-Rb, or HEK293T cells were transfected with siRNAs targeting ALDH2 and ALDH1B1 or a control siRNA using a lipid-based transfection reagent according to the manufacturer's directions. Cells were harvested 72-96 hours post-transfection for subsequent experiments. [3]

Animal/Disease Models: Rat[2]
Doses: 0.3 mg/kg, 0.6 mg/kg, 1.2 mg/kg, 2.4 mg/kg, 4.8 mg/kg
Route of Administration: po (po (oral gavage)) 4-5 times
Experimental Results: Inhibition of acute inflammation.

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Adult male C57BL/6 mice were fed either a normal chow diet (NCD) or a high-fat diet (HFD) for 8 weeks. Flurbiprofen was dissolved in sterilized water containing NaOH and mixed into the drinking water. The concentration was adjusted based on average water intake to achieve a daily dose of 10 mg/kg. Body weight was measured once a week. [3]
For the ob/ob mouse experiment, nine-week-old female ob/ob mice were housed individually. After acclimation, mice received intraperitoneal injections once daily: saline for 3 days, followed by recombinant mouse leptin (1 mg/kg) and/or flurbiprofen (10 mg/kg) for 6 days. Body weight was monitored. [3]
For fat content analysis, mice from the HFD study were euthanized after 7-8 weeks of treatment. Visceral fat was dissected and weighed. Alternatively, visceral and subcutaneous fat volumes were measured in live anesthetized mice using computed tomography (CT) scanning. [3]
For plasma leptin measurement, blood samples were collected from mice by decapitation at the end of the experiments. Plasma was separated by centrifugation, and leptin levels were quantified using a commercial ELISA kit according to the manufacturer's guidelines. [3]

Absorption, Distribution and Excretion
Flurbiprofen is rapidly and almost completely absorbed after oral administration. Peak plasma concentrations are reached 0.5–4 hours after oral administration. Very little flurbiprofen is excreted into human breast milk. After administration, less than 3% of flurbiprofen is excreted unchanged in the urine, and approximately 70% of the dose is excreted unchanged as well as its metabolites. Renal excretion is the primary route of excretion for flurbiprofen metabolites. 14 L [Normal healthy adults] 12 L [Elderly patients with arthritis] 10 L [Patients with end-stage renal disease] 14 L [Patients with alcoholic cirrhosis] 0.12 L/kg

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Metabolism/Metabolites Liver. Cytochrome P450 2C9 plays a crucial role in the metabolism of flurbiprofen to its major metabolite, 4-hydroxyflurbiprofen. In animal models of inflammation, the 4-hydroxyflurbiprofen metabolite exhibits almost no anti-inflammatory activity. Liver. Cytochrome P450 2C9 plays a crucial role in the metabolism of flurbiprofen to its major metabolite, 4-hydroxyflurbiprofen. In animal inflammation models, the 4-hydroxyflurbiprofen metabolite exhibits almost no anti-inflammatory activity. Elimination pathway: Flurbiprofen is rarely secreted into human milk. After administration, less than 3% of flurbiprofen is excreted unchanged in the urine, and approximately 70% of the dose is excreted unchanged as well as its metabolites. Renal excretion is a major pathway for the clearance of flurbiprofen metabolites. Half-life: R-flurbiprofen, 4.7 hours; S-flurbiprofen, 5.7 hours.

Toxicity Summary
Similar to other nonsteroidal anti-inflammatory drugs (NSAIDs), flurbiprofen's anti-inflammatory effect is achieved through reversible inhibition of cyclooxygenase (COX). COX is an enzyme responsible for converting arachidonic acid to prostaglandin G2 (PGG2) and then to prostaglandin H2 (PGH2) in the prostaglandin synthesis pathway. This effectively reduces the concentration of prostaglandins involved in inflammation, pain, swelling, and fever. Flurbiprofen is a non-selective COX inhibitor that inhibits the activity of both COX-1 and COX-2. It is also one of the most potent NSAIDs in terms of prostaglandin inhibitory activity. Hepatotoxicity
Prospective studies have shown that up to 15% of patients taking flurbiprofen may experience a slight increase in serum transaminase levels, but these are usually transient, mild, and asymptomatic, and often subside with continued use. Significant elevation of aminotransferases (more than 3-fold increase) occurred at a probability score of C (likely a rare cause of clinically significant liver damage). Pregnancy and Lactation Effects ◉ Overview of Use During Lactation Due to the low concentration and short half-life of flurbiprofen in breast milk, adverse effects on breastfed infants are unlikely, especially if the infant is older than 2 months. No adverse effects are expected on breastfed infants from maternal use of flurbiprofen eye drops. To significantly reduce the amount of medication entering breast milk after using eye drops, press the tear duct at the corner of the eye for 1 minute or longer, then wipe away excess medication with absorbent tissue. ◉ Effects on Breastfed Infants A retrospective medical record study in Taiwan compared the effects of acetaminophen (n = 348) and flurbiprofen (n = 132) on postpartum analgesia after vaginal delivery in full-term breastfed infants. There was no statistically significant difference in the incidence of hyperbilirubinemia between infants whose mothers took flurbiprofen (0.76%) and infants whose mothers took acetaminophen (2.01%). A study of full-term vaginally delivered, breastfed newborns compared newborns whose mothers took acetaminophen (n = 348) and those whose mothers took flurbiprofen (n = 132) for postpartum analgesia. Seven newborns (2%) who took acetaminophen had hyperbilirubinemia, compared to one newborn (0.76%) who took flurbiprofen. The difference was not statistically significant. Effects on lactation and breast milk: As of the revision date, no relevant published information was found. Protein binding: 99% binding, primarily to albumin. Unlike anticoagulants, sulfonamides, and phenytoin sodium, this product has a different major binding site to albumin.

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Toxicity Data>
LD50: 10 mg/kg (oral, dog) (A308)

[1]. Flurbiprofen suppresses the inflammation, proliferation, invasion and migration of colorectal cancer cells via COX2. Oncol Lett. 2020 Nov; 20(5): 132.

[2]. The pharmacology of 2-(2-fluoro-4-biphenylyl)propionic acid (flurbiprofen). A potent non-steroidal anti-inflammatory drug. Agents Actions. 1973 Nov;3(4):210-6.

[3]. Flurbiprofen ameliorated obesity by attenuating leptin resistance induced by endoplasmic reticulum stress. EMBO Mol Med, 2014.

Pharmacodynamics

Flurbiprofen is a propionic acid nonsteroidal anti-inflammatory drug (NSAID) with a structure and pharmacological action similar to fenprofen, ibuprofen, and ketoprofen, and has similar pharmacological action to other typical NSAIDs. Flurbiprofen has anti-inflammatory, analgesic, and antipyretic effects. Commercially available flurbiprofen is a racemic mixture of (+)S- and (-)R-enantiomers. The S-enantiomer appears to have most of the anti-inflammatory activity, while both enantiomers may have analgesic activity.

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Flurbiprofen is a 2-arylpropionic acid nonsteroidal anti-inflammatory drug (NSAID) that has traditionally been used to treat pain, inflammation, and fever by inhibiting cyclooxygenase (COX). [3]
This study reveals a novel, COX-independent pharmacological property of flurbiprofen: it acts as a chemical chaperone, inhibiting protein aggregation and thus alleviating endoplasmic reticulum (ER) stress. [3]
By reducing endoplasmic reticulum stress, flurbiprofen can attenuate leptin resistance (characterized by impaired STAT3 signaling pathway upon leptin stimulation), thereby counteracting high-fat diet-induced obesity in mice. [3]
The identified target aldehyde dehydrogenase (ALDH2/ALDH1B1) specifically interacts with flurbiprofen, but not with other tested nonsteroidal anti-inflammatory drugs (aspirin, ibuprofen, meloxicam). Studies have shown that flurbiprofen can inhibit heat-induced ALDH2 aggregation and increase its enzyme activity. [3]
The observed anti-obesity effect is thought to be mediated by this unique molecular chaperone activity and its interaction with aldehyde dehydrogenase, representing a potential new mechanism for treating obesity and related metabolic disorders. [3]

C15H13FO2

244.2609

244.089

5104-49-4

Tarenflurbil;51543-40-9;Flurbiprofen-d3;1185133-81-6;Flurbiprofen-d5;215175-76-1;Flurbiprofen-13C,d3;2747917-55-9

3394

White to light yellow solid powder

1.2±0.1 g/cm3

376.2±30.0 °C at 760 mmHg

110-112 °C(lit.)

181.3±24.6 °C

0.0±0.9 mmHg at 25°C

1.568

4.11

1

3

3

18

286

0

SYTBZMRGLBWNTM-UHFFFAOYSA-N

InChI=1S/C15H13FO2/c1-10(15(17)18)12-7-8-13(14(16)9-12)11-5-3-2-4-6-11/h2-10H,1H3,(H,17,18)

2-(3-fluoro-4-phenylphenyl)propanoic acid

Cebutid, dl-Flurbiprofen; Ansaid, Froben, Flurbiprofen, Antadys

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

DMSO : ≥ 100 mg/mL (~409.40 mM)

Solubility in Formulation 1: ≥ 2.5 mg/mL (10.23 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 2: ≥ 2.5 mg/mL (10.23 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

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Solubility in Formulation 3: ≥ 2.5 mg/mL (10.23 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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 (Please use freshly prepared in vivo formulations for optimal results.)