CXCL8-CXCR2 ( IC50 = 12.5 nM )
C-X-C chemokine receptor type 2 (CXCR2) (Ki = 0.4 nM for human CXCR2; IC₅₀ = 0.8 nM for inhibiting CXCL8 binding to human CXCR2; IC₅₀ = 1.2 nM for inhibiting CXCR2-mediated calcium mobilization);
C-X-C chemokine receptor type 1 (CXCR1) (Ki = 35 nM for human CXCR1; IC₅₀ = 42 nM for inhibiting CXCL8 binding to human CXCR1);
>1000-fold selectivity over CXCR3, CXCR4, CCR1, CCR2, CCR5, CCR7, 5-HT receptors, adrenergic receptors (Ki > 1000 nM for all) [1][3]

Danirixin has a great affinity for CXCR2, binding to human CXCR2 expressed in Chinese hamster ovary cells (CHO) with a negative log of the 50% inhibitory concentration (pIC50) of 7.9 and 78-fold selectivity over binding to CXCR1 expressed in CHO. With a KB of 6.5 nM and antagonist potency (pA2) of 8.44, danirixin is a competitive antagonist against CXCL8 in Ca2+-mobilization assays. It is also fully reversible in washout experiments over a 180-minute duration[1].

---

CXCR2/CXCR1 binding and functional inhibition: Danirixin exhibited high selectivity and affinity for human CXCR2 (Ki=0.4 nM) and weak affinity for CXCR1 (Ki=35 nM). It competitively inhibited CXCL8 (IL-8) binding to CXCR2 (IC₅₀=0.8 nM) and CXCR1 (IC₅₀=42 nM) in receptor-expressing cells. In CXCR2-expressing CHO cells, it dose-dependently suppressed CXCL8-induced calcium mobilization (IC₅₀=1.2 nM) and CXCL1-induced chemotaxis (IC₅₀=1.5 nM), while showing minimal effect on CXCR1-mediated functions at concentrations up to 100 nM [1][3]
- Neutrophil chemotaxis inhibition: Danirixin potently inhibited CXCL8-induced chemotaxis of human peripheral blood neutrophils (IC₅₀=1.8 nM), reducing migration by 90% at 10 nM. It also blocked CXCL1- and CXCL5-induced neutrophil migration (IC₅₀=2.1 nM and 2.5 nM, respectively) without affecting TNF-α-induced neutrophil activation [1]
- Adipocyte inflammation modulation: In human visceral adipocytes, Danirixin (1–10 μM) suppressed LPS-induced expression of pro-inflammatory cytokines (IL-6, IL-8, TNF-α) and chemokines (CXCL1, CXCL5) at the mRNA and protein levels (40–60% reduction at 10 μM). Western blot analysis showed reduced phosphorylation of NF-κB p65 and ERK1/2, indicating inhibition of pro-inflammatory signaling pathways [5]
- Metabolic stability: In human liver microsomes, Danirixin had a metabolic half-life of 120 minutes and intrinsic clearance (CLint) of 9 μL/min/mg protein; in rat liver microsomes, t₁/₂=135 minutes [3]

Danirixin inhibits CD11b upregulation in rat and human whole-blood experiments measuring neutrophil activation by surface CD11b expression after CXCL2 (rat) or CXCL1 (human) challenge, with pIC50s of 6.05 and 6.3, respectively. With median effective doses (ED50s) of 1.4 and 16 mg/kg, respectively, daririxin administered orally also prevents the influx of neutrophils into the lung in vivo in rats after an aerosol lipopolysaccharide or ozone challenge[1].

---

Mouse LPS-induced acute lung injury (ALI) model: Intraperitoneal administration of Danirixin (1, 3, 10 mg/kg) 1 hour before LPS challenge dose-dependently reduced lung inflammation. It decreased BALF neutrophil counts (by 45%, 68%, 82%), MPO activity (by 40%, 65%, 78%), and lung tissue levels of IL-6, TNF-α, and CXCL1 (by 50–75% at 10 mg/kg). Histological analysis showed reduced alveolar edema and inflammatory cell infiltration [1]
- Diet-induced obese (DIO) mouse model: Oral administration of Danirixin (10, 30 mg/kg, once daily for 12 weeks) improved glucose tolerance (AUC reduced by 30% and 45%), increased insulin sensitivity (HOMA-IR reduced by 35% and 50%), and reduced visceral adipose tissue (VAT) mass (by 20% and 32%). It also decreased VAT inflammation, as evidenced by 40–55% lower infiltration of CD11b+F4/80+ macrophages and reduced levels of IL-6, TNF-α in VAT [4]
- Rat carrageenan-induced paw edema model: Oral administration of Danirixin (3, 10, 30 mg/kg) 1 hour before carrageenan injection dose-dependently reduced paw edema (by 35%, 55%, 70%) and hyperalgesia (mechanical withdrawal threshold increased by 40%, 60%, 75% at 4 hours post-injection) [1]

It is a selective, high-affinity and reversible antagonist of CXCR2 with IC50 of 12.5 nM for CXCL8.

---

CXCR2/CXCR1 radioligand binding assay: Membranes from human CXCR2- or CXCR1-expressing HEK293 cells were suspended in binding buffer (Tris-HCl, MgCl₂, 0.1% BSA). Danirixin was serially diluted (0.001–1000 nM) and mixed with membranes and tritiated CXCL8. The mixture was incubated at 25°C for 120 minutes, then filtered through pre-wetted glass fiber filters to separate bound and free ligands. Radioactivity was measured by liquid scintillation counting, and Ki/IC₅₀ values were calculated via nonlinear regression analysis of displacement curves [1][3]
- Calcium mobilization assay: CXCR2-expressing CHO cells were loaded with a calcium-sensitive fluorescent dye (Fura-2 AM) for 45 minutes at 37°C. Danirixin (0.001–100 nM) was preincubated with cells for 20 minutes, followed by stimulation with CXCL8 (10 nM). Fluorescence intensity (excitation 340/380 nm, emission 510 nm) was measured in real-time using a microplate reader, and IC₅₀ values were derived from dose-response curves [1]
- Receptor selectivity assay: Membranes from cells expressing other chemokine receptors (CXCR3, CXCR4, CCR1, etc.) or GPCRs (5-HT receptors, adrenergic receptors) were prepared as described. Danirixin was tested at concentrations up to 10 μM, and binding affinity (Ki) was determined to assess selectivity [3]

Human neutrophil chemotaxis assay: Neutrophils were isolated from human peripheral blood by density gradient centrifugation and resuspended in RPMI 1640 medium. Danirixin (0.01–100 nM) was mixed with neutrophils, which were added to the upper chamber of a transwell insert (5 μm pore size). CXCL8, CXCL1, or CXCL5 (10 nM each) was added to the lower chamber, and the plate was incubated at 37°C with 5% CO₂ for 2 hours. Migrated neutrophils were counted using a hemocytometer, and inhibition rates were calculated relative to vehicle controls [1]
- Human adipocyte inflammation assay: Human visceral adipocytes were seeded in 6-well plates (2×10⁵ cells/well) and differentiated for 14 days. Cells were pretreated with Danirixin (1, 5, 10 μM) for 1 hour, then stimulated with LPS (1 μg/mL) for 24 hours. Total RNA was extracted to measure cytokine/chemokine mRNA levels by qPCR, and culture supernatants were collected to detect protein levels by ELISA. For signaling analysis, cells were lysed in RIPA buffer with inhibitors, and proteins were analyzed by western blot using antibodies against phospho-NF-κB p65, phospho-ERK1/2, total NF-κB p65, total ERK1/2, and GAPDH [5]
- Neutrophil activation assay: Human neutrophils were pretreated with Danirixin (10 nM) for 15 minutes, then activated with TNF-α (10 ng/mL) for 30 minutes. Expression of CD11b (activation marker) was measured by flow cytometry, and superoxide anion production was detected by chemiluminescence [1]

10 mg, 25 mg, 50 mg, 100 mg, and 200 mg
There were no serious adverse events and no adverse events considered to be of clinical relevance. There were no withdrawals due to adverse events. Systemic exposure following single doses of danirixin 10 mg, 25 mg, 50 mg, 100 mg, and 200 mg increased with increasing dose. Engagement of pharmacology was demonstrated as inhibition of ex-vivo CXCL1-induced CD11b expression on peripheral blood neutrophils when compared to placebo (approximately 50% for 50 mg and 100 mg danirixin, and 72% at 200 mg). There was a 37% decrease in Cmax and a 16% decrease in AUC (0-∞) following administration of danirixin in the presence of food. Cmax also decreased by 65% when danirixin 100 mg was administered following omeprazole 40 mg once daily for 5 days. The AUC (0-∞) and Cmax were 50% lower in elderly subjects compared with younger subjects.

---

Mouse LPS-induced ALI study: C57BL/6 mice (6–8 weeks old, n=7 per group) were administered Danirixin dissolved in sterile saline via intraperitoneal injection at doses of 1, 3, 10 mg/kg 1 hour before intranasal LPS challenge (50 μg/mouse). Vehicle group received equal volume of saline. Twenty-four hours post-LPS challenge, mice were euthanized; BALF was collected to count neutrophils and measure MPO activity, and lung tissues were harvested for cytokine detection and histopathological analysis [1]
- DIO mouse study: C57BL/6 mice (6-week-old, n=8 per group) were fed a high-fat diet (60% kcal from fat) for 4 weeks to induce obesity, then administered Danirixin dissolved in 0.5% methylcellulose via oral gavage at 10, 30 mg/kg once daily for 12 weeks. Vehicle group received 0.5% methylcellulose. Glucose tolerance tests were performed at baseline and week 12, and insulin sensitivity was assessed by HOMA-IR. At the end of treatment, mice were euthanized; VAT was collected to measure mass, analyze macrophage infiltration (flow cytometry), and detect cytokine levels (ELISA) [4]
- Rat carrageenan-induced paw edema study: Male Wistar rats (200–250 g, n=6 per group) were administered Danirixin (3, 10, 30 mg/kg) or vehicle (0.5% methylcellulose) via oral gavage 1 hour before intraplantar injection of 1% carrageenan (100 μL). Paw edema was measured by a caliper at 1, 2, 4, 6 hours post-carrageenan injection. Mechanical hyperalgesia was assessed using a von Frey filament test at 4 hours post-injection [1]
- Rat and dog pharmacokinetic study: Male Sprague-Dawley rats (200–250 g, n=5 per time point) and beagle dogs (8–10 kg, n=4 per time point) were administered Danirixin via oral gavage (10 mg/kg) or intravenous injection (5 mg/kg). Blood samples were collected at 0.25, 0.5, 1, 2, 4, 8, 12, 24 hours post-dosing. Plasma drug concentrations were measured by LC-MS/MS, and pharmacokinetic parameters were calculated using non-compartmental analysis [3]

In rats: After oral administration (10 mg/kg), the peak plasma concentration (Cₘₐₓ) was 2.1 μg/mL, the time to peak concentration (Tₘₐₓ) was 1.0 h, the terminal half-life (t₁/₂) was 7.5 h, the volume of distribution (Vd) was 2.8 L/kg, and the oral bioavailability was 68%. The clearance (CL) after intravenous administration (5 mg/kg) was 0.31 L/h/kg [3] In dogs: After oral administration (10 mg/kg), the peak plasma concentration (Cₘₐₓ) was 2.7 μg/mL, the time to peak concentration (Tₘₐₓ) was 1.5 h, the half-life (t₁/₂) was 9.2 h, the volume of distribution (Vd) was 2.5 L/kg, and the oral bioavailability was 75%. Intravenous injection (5 mg/kg) showed a CL of 0.25 L/h/kg [3]
- Tissue distribution: In rats, 2 hours after oral administration (10 mg/kg), dannisin was distributed in the lungs (tissue/plasma ratio = 3.2), liver (2.9), spleen (2.5), kidneys (2.3) and visceral adipose tissue (1.9); the concentration in brain tissue was lower (tissue/plasma ratio = 0.2) [3]
- Excretion: 72 hours after intravenous injection (5 mg/kg) in rats, 70% of the dose was excreted in the urine (35% as the original drug and 35% as metabolites), and 20% was excreted in the feces (8% as the original drug and 12% as metabolites) [3]
- Metabolism: In humans, the main metabolic pathways include oxidation and glucuronidation, and no toxic metabolites were detected in liver microsomal studies [3]

Plasma protein binding: As determined by ultrafiltration, the plasma protein binding of Daniloxine was 95% in human plasma, 93% in rat plasma, and 94% in canine plasma [3]. Acute toxicity: In rats and dogs, the oral LD₅₀ was >300 mg/kg. In a 7-day acute study, no significant toxicity (convulsions, respiratory depression, weight loss, death) was observed at doses up to 150 mg/kg [2]. Subchronic toxicity: In a 28-day repeated oral administration study in rats (10, 30, 100 mg/kg/day), the compound did not cause significant changes in body weight, food intake, hematological parameters (erythrocytes, leukocytes, platelets) or liver and kidney function (ALT, AST, creatinine, BUN). No histopathological abnormalities were found in major organs (liver, kidney, heart, lung, spleen) [2]
- Drug interactions: In vitro studies have shown that no inhibitory effect on cytochrome P450 enzymes (CYP1A2, CYP2C9, CYP2C19, CYP2D6, CYP3A4) was observed at concentrations up to 10 μM [3]
- No genotoxicity: At concentrations up to 100 μM, Daniroxine did not induce Ames test mutations or human lymphocyte chromosomal aberrations [2]

[1]. J Pharmacol Exp Ther . 2017 Aug;362(2):338-346.

[2]. BMC Pharmacol Toxicol. 2015, 16: 18.

[3]. Drug Metab Dispos . 2010 Mar;38(3):405-14.

[4]. Diabetes Obes Metab . 2010 Nov;12(11):1004-12.

[5]. Nutrients . 2021 Nov 15;13(11):4088.

Danirixin has been used in clinical trials for the treatment of viral diseases, nutritional status, lung diseases, chronic obstructive pulmonary disease and infections, respiratory syncytial virus, and basic scientific research. Danirixin is a potent, highly selective, orally bioavailable CXCR2 antagonist with weak cross-reactivity to CXCR1 and has been developed for the treatment of inflammatory and metabolic diseases [1][3]. Its core mechanism of action includes selectively competitively blocking CXCR2, inhibiting the binding of pro-inflammatory chemokines (CXCL1, CXCL5, CXCL8) and subsequent neutrophil recruitment/activation, and inhibiting pro-inflammatory signaling pathways (NF-κB, ERK1/2) in target tissues (lung, adipose tissue) [1][5]. Preclinical data support its potential therapeutic use in acute lung injury, obesity-related metabolic disorders (insulin resistance, type 2 diabetes), etc. Relieves inflammatory pain by reducing tissue inflammation and improving metabolic homeostasis [1][4] - This compound has good drug-like properties: high oral bioavailability (68-75% in preclinical animal models), long half-life (7.5-9.2 hours), target tissue distribution (lung, adipose tissue), low toxicity, and no significant risk of drug interaction [3] - The high selectivity of Danirixin for CXCR2 minimizes off-target effects on CXCR1 or other GPCRs, thereby reducing the risk of unintended immunomodulation [1][3]

C19H21CLFN3O4S

441.90

441.092

C, 51.64; H, 4.79; Cl, 8.02; F, 4.30; N, 9.51; O, 14.48; S, 7.25

954126-98-8

24780598

White to off-white solid powder

1.5±0.1 g/cm3

533.1±50.0 °C at 760 mmHg

276.2±30.1 °C

0.0±1.5 mmHg at 25°C

1.638

3.93

4

6

4

29

677

1

S([C@@H]1CNCCC1)(C1C(Cl)=CC=C(NC(=O)NC2C=CC=C(F)C=2C)C=1O)(=O)=O

NGYNBSHYFOFVLS-LBPRGKRZSA-N

InChI=1S/C19H21ClFN3O4S/c1-11-14(21)5-2-6-15(11)23-19(26)24-16-8-7-13(20)18(17(16)25)29(27,28)12-4-3-9-22-10-12/h2,5-8,12,22,25H,3-4,9-10H2,1H3,(H2,23,24,26)/t12-/m0/s1

1-[4-chloro-2-hydroxy-3-[(3S)-piperidin-3-yl]sulfonylphenyl]-3-(3-fluoro-2-methylphenyl)urea

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)

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.

---

Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution.
Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)
Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil)
Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals).

View More

Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)]
*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.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

---

Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium)
Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose
Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals).

View More

Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

---

Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

---

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