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Orismilast (LEO32731) is a novel, oral and potent PDE4 (phosphodiesterase-4) inhibitor with a broad spectrum of anti-inflammatory activity. It is being investigated in clinical trials for treating atopic dermatitis (AD), psoriasis, hidradenitis suppurativa (HS). It acts early in the inflammation cascade to induce a broad range of anti-inflammatory effects. Orismilast has been selected as lead molecule based on its attractive therapeutic window (i.e., the combination of improved efficacy and tolerability) and because of its potential to inhibit many inflammatory pathways involved in immunologic diseases due to the broad range of anti-inflammatory effects. Over the course of Phase 1 and Phase 2 trials, orismilast has been demonstrated to be safe in both oral and topical dosage forms and the PDE4 mode-of-action has been verified as an effective, well-tolerated approach to treatment of patients across multiple diseases, with no need for monitoring.
| Targets |
Orismilast is a selective inhibitor of phosphodiesterase 4 (PDE4). It demonstrates potent inhibition of PDE4B and PDE4D subtype splice variants, with IC₅₀ values for most isoforms below 10 nmol/L. The compound shows reduced potency for the PDE4A10 subtype (IC₅₀ = 52 nmol/L) and PDE4C2 subtype (IC₅₀ = 104 nmol/L) [1].
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| ln Vitro |
Orismilast selectively inhibits PDE4, showing near full inhibition of the catalytic domain of PDE4 at a single concentration of 308 nmol/L, with significantly less inhibition of other PDE families (PDE1-3, PDE5-11) [1].
In radiometric PDE4 profiling assays, orismilast potently inhibited multiple splice variants: PDE4B1 (IC₅₀ = 1 nmol/L), PDE4B2 (IC₅₀ = 6 nmol/L), PDE4B3 (IC₅₀ = 9 nmol/L), PDE4D1 (IC₅₀ = 9 nmol/L), PDE4D2 (IC₅₀ = 5 nmol/L), PDE4D3 (IC₅₀ = 8 nmol/L), PDE4D4 (IC₅₀ = 5 nmol/L), PDE4D5 (IC₅₀ = 8 nmol/L), PDE4D7 (IC₅₀ = 3 nmol/L), while being less potent against PDE4A10 (IC₅₀ = 52 nmol/L) and PDE4C2 (IC₅₀ = 104 nmol/L) [1]. In human whole blood assays, orismilast inhibited aCD3/aCD28-induced TNFα release with an IC₅₀ of 30 nmol/L [1]. In human peripheral blood mononuclear cell (PBMC) assays, orismilast inhibited LPS-induced TNFα release with an IC₅₀ of 10 nmol/L [1]. In cytokine profiling assays using human PBMC, orismilast demonstrated broad anti-inflammatory effects by inhibiting secretion of Th1 cytokines (IFNγ relative IC₅₀ = 4 nmol/L, TNFα relative IC₅₀ = 2 nmol/L), Th2 cytokines (IL-13 relative IC₅₀ = 47 nmol/L, IL-4 relative IC₅₀ = 124 nmol/L, IL-5 relative IC₅₀ = 16 nmol/L), Th17 cytokines (IL-17A relative IC₅₀ = 71 nmol/L, IL-22 relative IC₅₀ = 57 nmol/L, IL-23 relative IC₅₀ = 19 nmol/L), and innate cytokines (IL-1α relative IC₅₀ = 10 nmol/L, IL-1β IC₅₀ = 114 nmol/L). IL-8 secretion was not inhibited at concentrations up to 10 μmol/L. Cell viability was not impaired at the highest tested concentration (10 μmol/L) [1]. |
| ln Vivo |
In a murine chronic oxazolone-induced ear inflammation model, oral administration of orismilast at doses of 10 mg/kg and 30 mg/kg significantly reduced ear thickness over time (p < 0.0001 for both doses compared to vehicle). The mean reduction in ear thickness with 30 mg/kg oral orismilast was comparable to that observed with dexamethasone at 2 mg/kg (AUC = 3.4 day×mm vs. 2.7 day×mm) [1].
In the same model, orismilast treatment at both 10 mg/kg and 30 mg/kg significantly reduced concentrations of inflammatory cytokines in ear tissue, including IL-1β, IL-4, IL-5, TNFα, mKC (murine homologue of IL-8), and IFNγ, showing on-par or greater reduction compared to dexamethasone at 2 mg/kg. Treatment with orismilast had no impact on body weight during the treatment period [1]. |
| Enzyme Assay |
PDE1-11 Enzymatic Activity Assay: The inhibitory activity of orismilast against PDE1 through PDE11 was measured using immobilized metal affinity for phospho-chemicals (IMAP) technology, which is based on the high-affinity binding of phosphate by immobilized metal coordination complexes on nanoparticles. The assay was conducted using a single concentration of 308 nmol/L orismilast to determine inhibitor potency, expressed as percent inhibition. For PDE6-AB, a radiometric assay was used instead of the IMAP technology [1].
PDE4 Subtype Profiling Radiometric Assay: A radiometric assay based on the two-step method of Thompson and Appleman, adapted to a 96-well plate format, was performed to determine the PDE4 profile of orismilast. Partially purified PDEs were obtained by clonal cDNA expression in S. frugiperda insect cells using a baculovirus expression system. Multiple splice variants of the four PDE4 subtypes were investigated. A seven-point, half-log dilution series of orismilast was used to determine the IC₅₀ value for each splice variant. IC₅₀ values were calculated in GraphPad Prism using a four-parameter logistic equation. Standard inhibitors served as experimental quality controls [1]. |
| Cell Assay |
Human Whole Blood TNFα Secretion Assay: Freshly drawn heparin-stabilized human peripheral blood was diluted with RPMI medium and supplemented with anti-CD3, anti-CD28, and IL-2. Orismilast diluted in DMSO was mixed with the blood to a final DMSO concentration of 0.1% and incubated at 37°C with 5% CO₂ for 24 hours. TNFα levels in the culture supernatant were quantified by AlphaLISA. A plasma aliquot was incubated with acceptor beads for 2 hours at room temperature, followed by addition of donor beads and incubation for 3 hours. TNFα levels were measured on an EnVision detection system [1].
Human PBMC TNFα Secretion Assay: PBMC were isolated from buffy coat blood using density gradient centrifugation, washed, frozen, and later thawed and suspended in serum-free assay medium with 0.1% BSA. Orismilast was diluted in DMSO and further diluted in assay medium before being transferred to 384-well tissue culture plates. TNFα levels were measured in the culture supernatant from LPS-treated PBMC (10 ng/mL final concentration) after 18 hours of incubation at 37°C. TNFα levels were quantified by AlphaLISA [1]. Cytokine Inhibition Profiling in Human PBMC: PBMC were isolated from buffy coat blood and resuspended in RPMI-10 medium. Cells were seeded in white 96-well plates and stimulated with soluble anti-CD3 (for IFNγ, IL-13, IL-17A, IL-22, IL-4, and IL-5) or LPS (for IL-1α, IL-1β, IL-23, TNFα, and IL-8) at a final concentration of 1 μg/mL. The cells were incubated for 24 hours (or 72 hours for IL-1β) with an eight-point dilution series of orismilast or 0.1% DMSO at 37°C with 5% CO₂. Supernatants were analyzed using a 12-plex Luminex assay. Cell viability was assessed using a luminescent cell viability assay according to supplier instructions, with measurements performed on a plate reader. Relative and absolute IC₅₀ values were calculated using an asymmetric 5PL curve in GraphPad Prism [1]. |
| Animal Protocol |
Murine Chronic Oxazolone Model: Female BALB/cABomTac mice were sensitized on the right ear with a single application of 10 μL oxazolone (0.8% in acetone) on Day 7. Repeated challenges with 10 μL oxazolone (0.4% in acetone) on the right ear were initiated 7 days later, on days 0, 3, 5, 7, 10, 12, 14, 16, 18, and 20. On day 10, 44 mice were randomized based on ear thickness measured on Day 10 to receive vehicle, control, or oral orismilast treatment. From Day 10-20, mice were orally administered vehicle (methylcellulose) (n=10), orismilast at 10 mg/kg and 30 mg/kg (n=10 per group), or dexamethasone at 2 mg/kg (n=10). Four mice were kept as naive, non-sensitized and non-treated animals. Ear thickness was measured on days 10, 12, 14, 17, 19, and 21 using a micrometer. One pharmacokinetic blood sample was taken 2 hours after the last dose by cardiac puncture under isoflurane anesthesia to determine serum concentration of orismilast. Serum was separated by centrifugation, and concentrations were analyzed using liquid chromatography-mass spectrometry. Tissue from the right ear was sampled to determine cytokine concentrations. After weighing, tissue was transferred to cryotubes with ice-cold sterile 0.9% NaCl containing protease inhibitor, homogenized, and centrifuged. Cytokine analysis was carried out using a Mouse Th1/Th2 Tissue kit [1].
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| ADME/Pharmacokinetics |
In the murine chronic oxazolone model, serum concentration of orismilast followed a dose-response relationship with an average concentration of 423 ng/mL at the 10 mg/kg dose and 1009 ng/mL at the 30 mg/kg dose, measured 2 hours after the last oral dose [1].
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| Toxicity/Toxicokinetics |
In human PBMC cytokine inhibition assays, cell viability was not impaired at the highest concentration of orismilast tested (10 μmol/L) [1].
In the murine chronic oxazolone model, oral administration of orismilast at doses of 10 mg/kg and 30 mg/kg had no impact on body weight during the treatment period, indicating no significant adverse effects on general health [1]. The manuscript notes that in a phase 1 trial, the development of an orismilast modified-release (MR) formulation attenuated gastrointestinal (GI)-related adverse events, which are common side effects of PDE4 inhibitors, in healthy volunteers [1]. |
| References |
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| Additional Infomation |
Orismilast is a potent and selective next-generation phosphodiesterase 4 (PDE4) inhibitor. By acting high in the inflammatory cascade, PDE4 inhibition has the potential to modulate multiple autoimmune pathways implicated in dermatologic diseases. The safety and tolerability profile of PDE4 inhibitors is well established, supported by two approved oral therapies and one topical formulation currently on the market.
Despite their established use, currently marketed oral PDE4 inhibitors face two key limitations: modest overall efficacy within their approved indications and dose-dependent gastrointestinal side effects that constrain dose escalation, thereby limiting therapeutic potential. Orismilast was designed to enhance anti-inflammatory potency while minimizing gastrointestinal tolerability issues. Proof-of-concept clinical efficacy has been demonstrated in both psoriasis and atopic dermatitis (AD) patients.
In human PBMC cytokine inhibition assays, cell viability was not impaired at the highest concentration of orismilast tested (10 μmol/L) [1]. In the murine chronic oxazolone model, oral administration of orismilast at doses of 10 mg/kg and 30 mg/kg had no impact on body weight during the treatment period, indicating no significant adverse effects on general health [1]. The manuscript notes that in a phase 1 trial, the development of an orismilast modified-release (MR) formulation attenuated gastrointestinal (GI)-related adverse events, which are common side effects of PDE4 inhibitors, in healthy volunteers [1]. |
| Molecular Formula |
C19H15CL2F2NO7S
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| Molecular Weight |
510.28
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| Exact Mass |
508.991
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| Elemental Analysis |
C, 44.72; H, 2.96; Cl, 13.89; F, 7.45; N, 2.74; O, 21.95; S, 6.28
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| CAS # |
1353546-86-7
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| PubChem CID |
54765967
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| Appearance |
White to off-white solid powder
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| Density |
1.7±0.1 g/cm3
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| Boiling Point |
788.8±60.0 °C at 760 mmHg
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| Flash Point |
430.9±32.9 °C
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| Vapour Pressure |
0.0±2.8 mmHg at 25°C
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| Index of Refraction |
1.649
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| LogP |
0.82
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| Hydrogen Bond Donor Count |
0
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| Hydrogen Bond Acceptor Count |
9
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| Rotatable Bond Count |
5
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| Heavy Atom Count |
32
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| Complexity |
788
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| Defined Atom Stereocenter Count |
0
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| SMILES |
C1CS(=O)(=O)CCC12OC3=C(C=CC(=C3O2)OC(F)F)C(=O)CC4=C(C=[N+](C=C4Cl)[O-])Cl
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| InChi Key |
ZININGNRPUGNSL-UHFFFAOYSA-N
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| InChi Code |
InChI=1S/C19H15Cl2F2NO7S/c20-12-8-24(26)9-13(21)11(12)7-14(25)10-1-2-15(29-18(22)23)17-16(10)30-19(31-17)3-5-32(27,28)6-4-19/h1-2,8-9,18H,3-7H2
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| Chemical Name |
2-(3,5-dichloro-1-oxidopyridin-1-ium-4-yl)-1-[7-(difluoromethoxy)-1',1'-dioxospiro[1,3-benzodioxole-2,4'-thiane]-4-yl]ethanone
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| Synonyms |
LEO32731; Orismilast; UNII-JH1CX8SG5V; LEO-32,731; JH1CX8SG5V; 1353546-86-7; Orismilast; LEO 32731; LEO-32731
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| HS Tariff Code |
2934.99.9001
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| Storage |
Powder -20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month |
| Shipping Condition |
Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)
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| Solubility (In Vitro) |
DMSO : ~100 mg/mL (~195.97 mM)
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| Solubility (In Vivo) |
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 Formulations
Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)(e.g. IP/IV/IM/SC) *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)] Oral Formulations
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 (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 1.9597 mL | 9.7985 mL | 19.5971 mL | |
| 5 mM | 0.3919 mL | 1.9597 mL | 3.9194 mL | |
| 10 mM | 0.1960 mL | 0.9799 mL | 1.9597 mL |
*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.
Calculation results
Working concentration： mg/mL；
Method for preparing DMSO stock solution： mg drug pre-dissolved in μL DMSO (stock solution concentration mg/mL). Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug.
Method for preparing in vivo formulation:：Take μL DMSO stock solution, next add μL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH2O，mix and clarify.
(1) Please be sure that the solution is clear before the addition of next solvent. Dissolution methods like vortex, ultrasound or warming and heat may be used to aid dissolving.
(2) Be sure to add the solvent(s) in order.
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