| Size | Price | Stock | Qty |
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| 1mg |
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| 5mg |
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| 10mg |
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| Other Sizes |
| Targets |
Smoothened (SMO) (EC₅₀ = 3 nM)
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| ln Vitro |
- In SHH-Light2 cells, SAG (0.1 nM - 100 μM; 30 h) induced firefly luciferase expression with an EC₅₀ of 3 nM, but inhibited expression at higher concentrations. Additionally, SAG (1 - 1000 nM; 1 h) competed for the binding of BODIPY-cyclopamine to SMO-expressing COS-1 cells, yielding an apparent dissociation constant (Kd) of 59 nM for the SAG/SMO complex [2].
- SAG (250 nM; 48 h) significantly increased SMO mRNA and protein expression in MDA-MB-231 cells. It also increased CA XII mRNA expression in MDA-MB-231 cells at 24 h under normoxic and hypoxic conditions, and enhanced cell migration at 24 h [3]. - SAG (10 μM) treatment of primary mouse astrocytes for 24 hours increased Gli1 mRNA levels by 2.3-fold and PTCH1 mRNA levels by 2.5-fold. It also reduced GLT-1 protein levels by 50% and GFAP protein levels by 40% [2]. With an EC50 of 3 nM, SAG (0.1 nM-100 μM; 30 h) stimulates firefly luciferase expression in Shh-LIGHT2 cells, which is subsequently inhibited at higher concentrations[1]. When SAG (1-1000 nM; 1 h) competes with BODIPY-cyclopamine to bind to Smo-expressing Cos-1 cells, the SAG/Smo complex has an apparent dissociation constant (Kd) of 59 nM[1]. Robotnikinin-induced pathway activation by ShhN is inhibited by SAG (100 nM)[2]. In MDAMB231 cells, SAG (250 nM; 48 h) dramatically raises SMO mRNA and protein expression[3]. In MDAMB231 cells, SAG (250 nM; 24 and 48 hours) boosts the expression of CAXII mRNA at 24 hours under both normoxic and hypoxic conditions[3]. MDAMB231 cell migration is increased by SAG (250 nM; 24 h)[3]. |
| ln Vivo |
- Systemic administration of SAG (15-20 mg/kg; i.p.) induced pre-axial polydactyly in a dose-dependent manner in C57BL/6J mice [5].
- In a mouse model of glucocorticoid-induced neonatal cerebellar injury, SAG (1.0 mM) prevented neurotoxic effects by activating the SHH-SMO pathway. It increased 11β-HSD2 expression and promoted cerebellar granule neuron precursor survival and proliferation. Treatment with SAG did not interfere with glucocorticoid-induced lung maturation and did not promote tumor formation after 1-week treatment [2]. - SAG (1.0 mM) induced more osteogenesis at the defect borders and increased bone volume/tissue volume (BV/TV) at the eight-week time point in CD-1 mice [2]. - Combining SAG (1.0 mM) with NEL-like protein-1 (NELL-1) in a collagen scaffold enhanced bone healing in critical-sized calvarial defects in mice, with significantly higher bone volume and mineral density compared to single-agent treatment [4]. In CD-1 mice, SAG (1.0 mM) causes a notable rise in BV/TV at the eight-week timepoint and increased osteogenesis, primarily at the defect borders[4]. In mice, SAG (15–20 mg/kg; ip) dose-dependently causes pre-axial polydactyly to occur[5]. |
| Enzyme Assay |
To determine the binding affinity of SAG to SMO, a competition binding assay was performed. COS-1 cells expressing SMO were incubated with BODIPY-cyclopamine (10 nM) and increasing concentrations of SAG (1 - 1000 nM) for 1 hour at room temperature. Fluorescence polarization was measured to determine the displacement of BODIPY-cyclopamine by SAG, yielding a Kd of 59 nM [2].
- An enzyme reporter assay was conducted using SHH-Light2 cells, which stably express a Gli-responsive luciferase reporter. Cells were treated with SAG (0.1 nM - 100 μM) for 30 hours, and luciferase activity was measured. The EC₅₀ for SAG-induced luciferase expression was 3 nM [2]. |
| Cell Assay |
- Primary mouse astrocytes were treated with SAG (10 μM) for 24 hours. Total RNA was extracted, and qPCR was performed to measure Gli1, PTCH1, GLT-1, and GFAP mRNA levels. SAG increased Gli1 and PTCH1 mRNA levels and decreased GLT-1 and GFAP mRNA levels. Western blot analysis confirmed the reduction in GLT-1 and GFAP protein levels [2].
- MDA-MB-231 cells were treated with SAG (250 nM) for 24 or 48 hours. Total RNA was extracted, and qPCR was performed to measure SMO and CA XII mRNA levels. SAG increased both SMO and CA XII mRNA levels. Cell migration was assessed using a wound healing assay, where SAG-treated cells showed increased migration compared to controls [3]. |
| Animal Protocol |
For the glucocorticoid-induced cerebellar injury model, neonatal mice (postnatal day 0) received daily intraperitoneal injections of SAG (15 mg/kg) or vehicle for 7 days. Glucocorticoids (dexamethasone, 0.5 mg/kg) were administered subcutaneously daily for 7 days starting on postnatal day 3. Mice were sacrificed on postnatal day 10, and cerebella were analyzed for histological changes, cell proliferation (Ki-67 staining), and apoptosis (TUNEL assay) [2].
- For the polydactyly induction study, pregnant C57BL/6J mice (gestational day 10.5) received a single intraperitoneal injection of SAG (20 mg/kg). Offspring were evaluated for limb malformations at birth [5]. - For the bone regeneration study, CD-1 mice with critical-sized calvarial defects were treated with SAG (1.0 mM) in a collagen scaffold implanted into the defect site. Mice were sacrificed at 8 weeks, and micro-CT analysis was performed to assess bone volume and mineral density [2]. - In the combined SAG and NELL-1 study, C57BL/6J mice with calvarial defects received a collagen scaffold containing SAG (1.0 mM) and NELL-1 (10 μg/mL). Mice were sacrificed at 12 weeks, and bone healing was evaluated by micro-CT and histological analysis [4]. Animal/Disease Models: Pregnant C57BL/6J mice[5] Doses: 15, 17, 20 mg/kg Route of Administration: A single ip Experimental Results: Effective in ca. 80% of the embryos and increased Gli1 and Gli2 mRNA expression in the limb bud, with Gli1 mRNA being the most upregulated at the dose of 20 mg/kg. |
| ADME/Pharmacokinetics |
After intraperitoneal injection, the half-life of SAG in mouse plasma is about 2 hours. It can be rapidly distributed to various tissues, with the highest concentrations in the liver, kidneys and brain. SAG is mainly metabolized by the cytochrome P450 system, with less than 10% excreted unchanged in the urine [2].
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| Toxicity/Toxicokinetics |
- In acute toxicity studies, intraperitoneal injection of SAG at doses up to 500 mg/kg did not cause death or significant adverse reactions in mice. Subchronic toxicity studies (14 days) showed no hepatotoxicity or nephrotoxicity (normal liver and kidney function indicators) or hematological abnormalities [2]. - SAG has low plasma protein binding in mouse plasma (less than 20%) [2].
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| References |
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| Additional Infomation |
SAG is a small molecule agonist of the smoothed receptor that activates the Hedgehog (Hh) signaling pathway. It has been investigated for use in regenerative medicine, particularly in cartilage and bone repair and neuroprotection against glucocorticoid-induced damage [2]. SAG activation of the Hh signaling pathway promotes the proliferation and survival of various cell types, including chondrocytes, neuronal progenitors, and astrocytes. However, chronic activation of the Hh signaling pathway is associated with tumorigenesis, but short-term treatment with SAG did not promote tumor formation in animal models [2]. Studies have shown that SAG can enhance osteogenic activity in vivo, making it a potential drug for treating bone defects. Its ability to activate the Hh signaling pathway in stem cells/progenitor cells contributes to its regenerative effects [2,4]. Mice exposed to SAG during pregnancy developed preaxial polydactyly, highlighting the teratogenic potential of Hh pathway activation during embryonic development [5].
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| Molecular Formula |
C28H30CL3N3OS
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|---|---|
| Molecular Weight |
562.98
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| CAS # |
2702366-44-5
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| Related CAS # |
SAG;912545-86-9;SAG hydrochloride;2095432-58-7;(Rac)-SAG;364590-63-6
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| PubChem CID |
154884292
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| Appearance |
Light yellow to light brown solid powder
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| Hydrogen Bond Donor Count |
3
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| Hydrogen Bond Acceptor Count |
4
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| Rotatable Bond Count |
6
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| Heavy Atom Count |
36
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| Complexity |
666
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| Defined Atom Stereocenter Count |
0
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| SMILES |
O=C(C1=C(Cl)C2=CC=CC=C2S1)N([C@H]3CC[C@H](NC)CC3)CC4=CC=CC(C5=CC=NC=C5)=C4.[H]Cl
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| Synonyms |
SAG dihydrochloride; SAG 2HCl
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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 Note: Please store this product in a sealed and protected environment, avoid exposure to moisture. |
| 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) |
H2O: 100 mg/mL (177.63 mM)
DMSO: 33.33 mg/mL (59.20 mM) |
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| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 2.5 mg/mL (4.44 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. Solubility in Formulation 2: ≥ 2.5 mg/mL (4.44 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. View More
Solubility in Formulation 3: ≥ 2.5 mg/mL (4.44 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution. Solubility in Formulation 4: 50 mg/mL (88.81 mM) in PBS (add these co-solvents sequentially from left to right, and one by one), clear solution; with ultrasonication. |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 1.7763 mL | 8.8813 mL | 17.7626 mL | |
| 5 mM | 0.3553 mL | 1.7763 mL | 3.5525 mL | |
| 10 mM | 0.1776 mL | 0.8881 mL | 1.7763 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.
| NCT Number | Recruitment | interventions | Conditions | Sponsor/Collaborators | Start Date | Phases |
| NCT02051413 | COMPLETED | Drug: Venlafaxine extended release | Major Depressive Disorder Major Depressive Episode |
Institut National de la Santé Et de la Recherche Médicale, France | 2014-02-18 | Phase 4 |
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