p38 (IC50 = 50 nM); p38β2 (IC50 = 500 nM)

SB203580 has an IC50 of 3-5 μm and blocks the proliferation of primary human T cells, murine CT6 T cells, or BAF F7 B cells when IL-2 is present. Although the concentration needed is a little bit higher and the IC50 is above 10 μM, SB203580 also inhibits IL-2-induced p70S6 kinase activation. With an IC50 in the 3-10 μM range, SB203580 also inhibits the activity of PDK1 in a dose-dependent manner. MAPKAPK2 stimulation by p38-MAPK is inhibited by SB203580 with an IC50 of roughly 0.07 μM, whereas total SAPK/JNK activity is inhibited with an IC50 of 3–10 μM. Higher concentrations of SB203580 cause the ERK pathway to be activated, which then improves the transcriptional activity of NF-κB. Human hepatocellular carcinoma (HCC) cells are induced to undergo autophagy by SB203580.

SB203580 protects the pig myocardium in an in vivo model from ischemic damage. In the MRL/lpr mouse model of systemic lupus erythematosus (SLE), SB203580 is effective in preventing and treating the illness.

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Proteinuria is prevented in SB203580 treated MRL/lpr mice. [4]
ALT and AST are not influenced by SB203580 in MRL/lpr mice. [4]
BUN but not Cr is decreased in SB203580 treated MRL/lpr mice. Renal but not splenic weight is reduced in SB203580 treated MRL/lpr mice. [4]
Renal pathologic changes are attenuated in SB203580 treated MRL/lpr mice. [4]
Hepatic pathologic changes are relieved in SB203580 treated MRL/lpr mice. [4]
Splenic pathologic changes are relieved in SB203580 treated MRL/lpr mice. [4]
Glomerular IgG, IgM, IgA and C3 depositions are reduced in SB203580 treated MRL/lpr mice. [4]

4 μg of sheep anti-PKBα is immobilized on 25 μL of protein G-Sepharose overnight (or 1.5 hours) and washed in Buffer A (50 mm Tris, pH 7.5, 1 mm EDTA, 1 mm EGTA, 0.5 mm Na3VO4, 0.1% β-mercaptoethanol, 1% Triton X-100, 50 mm sodium fluoride, 5 mm sodium pyrophosphate, 0.1 mm phenylmethylsulfonyl fluoride, 1 μg/mL aprotinin, pepstatin, leupeptin, and 1 μm microcystin). The immobilized anti-PKB is then incubated with 0.5 ml of lysate (from 5 × 106 cells) for 1.5 hours and washed three times in 0.5 mL of Buffer A supplemented with 0.5 m NaCl, two times in 0.5 mL of Buffer B (50 mm Tris-HCl, pH 7.5, 0.03% (w/v) Brij-35, 0.1 mm EGTA, and 0.1% β-mercaptoethanol), and twice with 100 μl of assay dilution buffer; 5× assay dilution buffer is 100 mm MOPS, pH 7.2, 125 mm β-glycerophosphate, 25 mm EGTA, 5 mm sodium orthovanadate, 5 mm DTT. To the PKB enzyme immune complex is added 10 μL of assay dilution buffer, 40 μm protein kinase A inhibitor peptide, 100 μm PKB-specific substrate peptide, and 10 μCi of [γ-32P]ATP, all made up in assay dilution buffer. The reaction is incubated for 20 minutes at room temperature with shaking, then samples are pulse spun, and 40 μL of reaction volume are removed into another tube to which is added 20 μL of 40% trichloroacetic acid to stop the reaction. This is mixed and incubated for 5 minutes at room temperature, and 40 μL is transferred onto P81 phosphocellulose paper and allowed to bind for 30 seconds. The P81 piece is washed three times in 0.75% phosphoric acid then in acetone at room temperature. γ-32P incorporation is then measured by scintillation counting.

In the absence of growth factors, antibiotics, or β-mercaptoethanol supplements, CT6 cells and BA/F3 F7 cells are rested by washing three times in RPMI and culturing overnight in RPMI with 5% fetal calf serum. Preincubation with SB203580 or a vehicle control, as indicated in the figure legends, is performed on 2-5 106 rested CT6 cells in 2 mL of RPMI, 5% fetal calf serum. Following a 5-minute incubation period at 37 °C with 20 ng/ml recombinant human IL-2 stimulation, cells are pelleted in a minifuge for 30 seconds, the medium is aspirated, and the pellet is lysed in the proper buffer. BA/F3 cells are maintained in glutamine-containing RPMI that is additionally supplemented with 5% fetal calf serum and 0.2 μg/mL G418 and stably express deletion mutants of the IL-2 β receptor chain. The cells are then thoroughly washed, allowed to rest for the night, and then washed once more before being activated with IL-2. Such cell preparations contain >90% T cells. The incorporation of [3H]thymidine is measured in cellular proliferation assays.

Six-week-old female atymic Nu/Nu mice CAL27 p38WT and p38TM tumors[1]
5 mg/kg/day
Intra peritoneal injected daily for 16 consecutive days

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Systemic lupus erythematosus (SLE) are established in female MRL/lpr mice and female C57BL/6 mice
0.1 M/day
Orally administered

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Female MRL/lpr mice were randomized into two groups (n = 10 per group) and were fed control diet (named as group 2 in the following) or diet with SB203580 (named as group 3 in the following) starting at the age of 14 weeks and continuing for up to 22 weeks. Adezmapimod (SB203580) was dissolved in drinking water (250 μmol/L), was orally administered (0.4 ml/day). Ten C57BL/6 female mice were used as negative controls (named as group 1 in the following). Two mice in MRL/lpr group 2 were dead at 16 weeks and 18 weeks of age respectively. Two mice in MRL/lpr group 3 were dead at 19 weeks of age. Significant increase of urine protein (300–2000 mg/dl) was found in each mouse before death, indicating a probable renal failure be the cause of death. Ultimately, 10 mice in group 1, 8 mice in group 2 and group 3 were included in statistical analysis.[4]

1. Solubility and Formulation
o Solubility: SB203580 is highly soluble in DMSO (43 mg/mL or 113.92 mM), but insoluble in water and ethanol.
o Formulation: Usually available as a powder or DMSO solution for research use.
2. Absorption and Bioavailability
o Oral Administration: In animal studies, SB203580, administered orally (e.g., dissolved in drinking water at a concentration of 250 μM), showed efficacy in disease models.
o Intraperitoneal Injection (IP): Systemic activity was observed in mice at a dose of 5 mg/kg/day.
3. Metabolism and Half-Life
o Metabolic Stability: Direct data on metabolic pathways are not available, but storage conditions (-20°C, protected from light) suggest it is prone to degradation.
o In vivo efficacy: In mice and rats, at doses of 15–60 mg/kg, it effectively inhibits inflammatory cytokines, with an in vivo IC50 of 15–25 mg/kg.
4. Distribution and protein binding
o Cell permeability: SB203580 can permeate cell membranes and inhibit intracellular p38 MAPK (intracellular IC50 = 600 nM).
o Tissue effects: It can alleviate inflammation in collagen-induced arthritis and endotoxin shock models, indicating its broad tissue distribution.
5. Excretion and clearance
o Specific data on the excretion route are not available, but its effects in animal models suggest moderate clearance (e.g., daily dosing is required to maintain activity).

1. Acute Toxicity and Safety Hazards
• Oral Toxicity: Category 4 (Hazard Statement H302) – Harmful if swallowed.
• Ocular Toxicity: Causes severe eye damage (Category 1, H318); direct contact requires immediate rinsing and medical attention.
• Handling Precautions: Due to the risk of inhalation or skin contact, protective equipment (gloves, goggles, mask) is required.
________________________________________
2. In Vivo Toxicity Studies
Ocular Exposure (Conjunctival Injection)
• Study: Subconjunctival injection of 50 μM SB203580 in rats did not show significant corneal toxicity (e.g., intact epithelium, normal stromal arrangement), but caused transient conjunctival anemia, which subsided within 24 hours.
• Conclusion: Low short-term ocular toxicity, but local irritation may occur.
Systemic Administration
• Asthma Model: In asthmatic rats exposed to smoke, SB203580 reduced airway inflammation and improved lung function, with no adverse reactions reported at the tested dose.
• Pancreatitis Model: In a rat model of severe acute pancreatitis, inhibition of TNF-α and apoptosis of pancreatic acinar cells was high, suggesting therapeutic potential, and no significant toxicity was observed.
________________________________________
3. Biochemical and Cytotoxic Effects
• Off-target Effects: At high concentrations (>10 μM), SB203580 may nonspecifically inhibit kinases such as PKB or PDK1 and anomalously activate the ERK/NF-κB pathway.
• Cell Culture: Cytotoxicity was observed in certain cell lines (e.g., hepatocytes) when concentrations exceeded its p38 MAPK IC50 (0.3–0.5 μM).
________________________________________
4. Environmental and Disposal Risks
• Storage: Stable at -20°C, but degrades under high temperature, humidity, or light.
• Disposal: Must be incinerated or treated as hazardous waste to avoid environmental pollution.

[1]. Specificity and mechanism of action of some commonly used protein kinase inhibitors. Biochem J. 2000 Oct 1;351(Pt 1):95-105.

[2]. The pyridinyl imidazole inhibitor SB203580 blocks phosphoinositide-dependent protein kinase activity, protein kinase B phosphorylation, and retinoblastoma hyperphosphorylation in interleukin-2-stimulated T cells independently of p38 mitogen-activated protein kinase. J Biol Chem. 2000 Mar 10;275(10):7395-402.

[3]. A role for p38 MAPK in head and neck cancer cell growth and tumor-induced angiogenesis and lymphangiogenesis. Mol Oncol. 2014 Feb;8(1):105-18.

[4]. The selective p38 mitogen-activated protein kinase inhibitor, SB203580, improves renal disease in MRL/lpr mouse model of systemic lupus. Int Immunopharmacol. 2011 Sep;11(9):1319-26.

Pyridinyl imidazole inhibitors, particularly SB203580, have been widely used to elucidate the role of p38 mitogen-activated protein kinase (MAP kinase, p38/HOG/SAPKII) in various biological systems. Our study and studies by other teams have shown that SB203580 possesses antiproliferative activity against cytokine-activated lymphocytes. However, we recently reported that the antiproliferative effect of SB203580 is independent of p38 MAP kinase activity. This study demonstrates that SB203580 can inhibit the phosphorylation of retinoblastoma protein in interleukin-2 stimulated T cells, a key cell cycle event. Further investigation of the phosphatidylinositol 3-kinase/protein kinase B (PKB) (Akt/Rac) kinase pathway, a proximal regulator of this event, shows that SB203580 blocks PKB phosphorylation and activation by inhibiting the PKB kinase—phosphatidylinositol-dependent protein kinase 1. The concentration of SB203580 required to block PKB phosphorylation (IC50 of 3-5 μM) was only about 10 times higher than the concentration required to inhibit p38 MAP kinase (IC50 of 0.3-0.5 μM). These data reveal a novel activity of the drug and suggest that extra caution should be exercised when interpreting data for SB203580 concentrations above 1-2 μM. [2] We have recently gained significant insights into the mutational profile of head and neck squamous cell carcinoma (HNSCC). However, the nature of the dysregulated signaling pathway network leading to the progression of head and neck squamous cell carcinoma (HNSCC) remains unclear. This study focuses on the roles of the mitogen-activated protein kinase (MAPK) family, extracellular signal-regulated kinase (ERK), c-Jun terminal kinase (JNK), and p38 MAPK in HNSCC. Immunohistochemical analysis of a large number of human HNSCC tissues revealed that the phosphorylated forms of ERK1/2 and JNK were elevated in less than 33% and 16% of cases, respectively. However, it is noteworthy that high levels of active phosphorylated p38 were observed in most (79%) of the hundreds of tissues analyzed. We explored the biological function of p38 in HNSCC cell lines using three independent approaches: treatment with the specific p38 inhibitor SB203580; a reverse inhibition strategy involving the co-expression of an inhibitor-insensitive p38α mutant with SB203580; and short hairpin RNA (shRNA) targeting p38α. We found that specific blocking of the p38 signaling pathway significantly inhibited the proliferation of HNSCC cells in vitro and in vivo. Indeed, we observed that inhibition of p38 in HNSCC cancer cells reduced tumor growth in a tumor xenograft model and significantly reduced intratumoral angiogenesis and lymphangiogenesis. We concluded that p38α acts as a positive regulator of HNSCC in the tumor microenvironment, controlling cancer cell growth as well as tumor-induced angiogenesis and lymphangiogenesis. [3]

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Systemic lupus erythematosus (SLE) is an autoimmune disease with excessive inflammatory responses in multiple organs. Aberrant activation of p38 MAPK is thought to be associated with inflammation in systemic lupus erythematosus (SLE), leading to progressive tissue and organ damage and ultimately developing into lupus nephritis and autoimmune hepatitis. To determine the efficacy of p38 MAPK inhibitors in an SLE mouse model, we orally administered the p38 MAPK-specific inhibitor SB203580 to 14- to 22-week-old MRL/lpr mice. Subsequently, we evaluated renal and hepatic function, as well as pathological changes in vital organs including the kidneys, liver, and spleen, in the MRL/lpr mice. The results showed that SB203580 reduced proteinuria and serum blood urea nitrogen (BUN) levels, improved renal function, alleviated renal pathological changes, and reduced the deposition of immunoglobulins (Ig) and C(3) in the kidneys. Furthermore, administration of SB203580 inhibited hepatocellular necrosis and the recruitment and proliferation of leukocytes in the liver and spleen. Therefore, activation of p38 MAPK may be partly responsible for the exacerbation of autoimmune kidney, liver, and spleen damage, while its inhibitors may reduce the autoimmune attack on these vital organs and improve kidney function. Our study shows that selective blocking of p38 MAPK can effectively prevent and treat the disease in this SLE model. [4]

C₂₁H₁₇CLFN₃OS

413.90

413.076

C, 60.94; H, 4.14; Cl, 8.56; F, 4.59; N, 10.15; O, 3.87; S, 7.75

869185-85-3

Adezmapimod;152121-47-6

16760644

Light yellow to yellow solid powder

6.349

2

5

4

28

500

0

Cl.O=S(C)C1C=CC(C2NC(C3C=CC(F)=CC=3)=C(C3C=CN=CC=3)N=2)=CC=1

WOSGGXINSLMASH-UHFFFAOYSA-N

InChI=1S/C21H16FN3OS.ClH/c1-27(26)18-8-4-16(5-9-18)21-24-19(14-2-6-17(22)7-3-14)20(25-21)15-10-12-23-13-11-15;/h2-13H,1H3,(H,24,25);1H

4-[4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-1H-imidazol-5-yl]pyridine;hydrochloride

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: Please store this product in a sealed and protected environment, avoid exposure to moisture.

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

Solubility in Formulation 1: ≥ 2.5 mg/mL (6.04 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 (6.04 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: 4% DMSO+30% PEG 300+5% Tween 80+ddH2O: 5mg/mL

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