WJH-C19 (compound 10e12) (25 hours) exhibits potent concentration-dependent protective effects against necrosis and apoptosis in various cell lines, with an EC50 value of 0.28 nM in L929 cells, 4.0 nM in HT22 cells, 1.6 nM in HT29 cells, and 2.0 nM in U937 cells; in addition, this compound reduces necrosis and apoptosis in L929 cells in a dose-dependent manner [1]. WJH-C19 can effectively inhibit the kinase activity of RIPK1, with an IC50 value of 5.7 nM [1]. WJH-C19 (0.5-50 nM) concentration-dependently inhibited TNFα/z-VAD-FMK (G) (TZ)-induced phosphorylation of RIPK1, RIPK3 and MLKL in L929 cells and mTNFα/Smac mimic/z-VAD-FMK (TSZ)-induced phosphorylation in HT29 cells, thereby blocking the formation of necrosomes and downstream necroptosis signaling pathways [1].

WJH-C19 (1.25-2.5 mg/kg; orally; once daily; for 7 days) significantly improved dextran sulfate sodium (DSS)-induced colitis in female C57BL/6 mice, resulting in a 47.5% recovery of body weight, a 42.9% reduction in disease activity index (DAI), and significant inhibition of the RIPK1/RIPK3/MLKL signaling pathway [1]. WJH-C19 (10-20 mg/kg; orally; once daily; for 14 days) significantly improved Freund's complete adjuvant (CFA)-induced rheumatoid arthritis in female C57BL/6 mice, reducing paw swelling, joint inflammation, and bone destruction, and inhibiting the RIPK1/RIPK3/MLKL signaling pathway [1].

Animal/Disease Models:C57BL/6 (female, 10-12 weeks old, approximately 28 g, DSS-induced colitis) [1]
Doses: 2.5 mg/kg; 1.25 mg/kg
Route of Administration: Oral; once daily; 7 days
Experimental Results: The 2.5 mg/kg dose group recovered 47.5% of body weight, reduced the disease activity index (DAI) by 42.9%, and recovered 60.5% of colon length. The 1.25 mg/kg dose group recovered 33.2% of body weight, reduced DAI by 32.1%, and recovered 35.1% of colon length. Both doses reduced the levels of pro-inflammatory cytokines (IL-1β, IL-6, TNFα) in colon tissue and increased the level of the anti-inflammatory cytokine IL-10. Both doses inhibited the phosphorylation of RIPK1, RIPK3, and MLKL in colon proteins, with the high dose group showing the strongest inhibitory effect. Both doses reduced mucosal damage, maintained glandular structural integrity, and restored the expression of tight junction protein-1 (ZO-1) and closure protein, with the high-dose group showing near-normal morphology.

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Animal/Disease Models:C57BL/6 (female, 8 weeks old, CFA-induced rheumatoid arthritis) [1]
Doses: 20 mg/kg; 10 mg/kg
Route of Administration: Oral; daily; 14 days
Experimental Results: Both doses reduced paw swelling percentage, ankle temperature and arthritis index score in a dose-dependent manner, with efficacy comparable to or better than celecoxib. Reduced osteophyte formation and reversal of bone-destructive changes, including improvements in bone mineral density (BMD), cortical bone density, trabecular number (Tb.N), connectivity density (Conn.D), bone volume (BV), cortical volume (CV), trabecular spacing (Tb.Sp), porosity (Po.N), cortical area (Ct.Ar), and cortical thickness (Ct.Th), were observed in both dose groups, with the most significant improvement observed in the high-dose group. Inflammatory cell infiltration, synovial hyperplasia, and cartilage erosion were reduced in both dose groups, with the histological score in the high-dose group approaching normal levels. The M1/M2 macrophage ratio in the spleen decreased in a dose-dependent manner in both dose groups, shifting towards the anti-inflammatory M2 phenotype. The expression of phosphorylated RIPK1, RIPK3, and MLKL in the ankle joint was inhibited in both dose groups, with the highest inhibitory effect observed in the high-dose group.

[1]. From Hit to Lead: Discovery of Novel Selective RIPK1 Inhibitor with Pyridoimidazole Scaffold for the Treatment of Autoimmune Diseases through Phenotypic Screening and Structural Optimization. J Med Chem. 2026;69(3):3252-3288.

C27H22CLN5O2S

516.01

Solid

O=C(C1=CN=C2C=CC(C3=CC=C4N=C(NC(C5CC5)=O)SC4=C3)=CN21)N[C@H](C6=CC=C(Cl)C=C6)C

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 : ~50 mg/mL (~96.90 mM; with sonication)

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.

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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).

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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)

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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).

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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

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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.

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