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Purity: ≥98%
PT2385 (PT-2385) is a nove, potent, selective and orally bioactive inhibitor of hypoxia inducible factor (HIF-2α) with anticancer and anti-hepatic steatosis activity. It inhibits HIF-2α with luciferase EC50 of 27 nM and no significant off-target activity. PT2385 can allosterically binds to HIF-2α, thereby preventing HIF-2α heterodimerization and its subsequent binding to DNA. Human-intestine biopsies from individuals with or without obesity revealed that intestinal HIF-2α signaling was positively correlated with body-mass index and hepatic toxicity. The causality of this correlation was verified in mice with an intestine-specific disruption of Hif2a, in which high-fat-diet-induced hepatic steatosis and obesity were substantially lower as compared to control mice. PT2385 had preventive and therapeutic effects on metabolic disorders that were dependent on intestine HIF-2α. Intestine HIF-2α inhibition markedly reduced intestine and serum ceramide levels. Mechanistically, intestine HIF-2α regulates ceramide metabolism mainly from the salvage pathway, by positively regulating the expression of Neu3, the gene encoding neuraminidase 3. These results suggest that intestinal HIF-2α could be a viable target for hepatic steatosis therapy.
| Targets |
HIF-2α (Kd<50 nM)[1]
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| ln Vitro |
HIF-2 is selectively antagonistic to PT-2385 (PT2385), but not HIF-1. HIF-1α is not inhibited by PT-2385[1].
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| ln Vivo |
Tumor regression is accelerated and dose-dependent when PT-2385 (30 or 100 mg/kg; oral gavage; twice daily) is administered[3]. In vivo, PT-2385 (PT2385) exhibits dose-dependent inhibition of HIF-2α-regulated gene expression. In 786-O xenografts, tumor regression was achieved with PT-2385 (3 and 10 mg/kg, oral, twice daily dosages). Moreover, PT-2385 (1, 3 and 10 mg/kg) reduced the amounts of tumor-derived VEGFA protein. Treatment with 10 mg/kg of PT-2385 decreases angiogenesis (CD-31) and proliferation (Ki67) [1].
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| Enzyme Assay |
The potent, selective, and orally active small-molecule inhibitor PT2385 as a specific antagonist of HIF2α that allosterically blocks its dimerization with the HIF1α/2α transcriptional dimerization partner ARNT/HIF1β. PT2385 inhibited the expression of HIF2α-dependent genes, including VEGF-A, PAI-1, and cyclin D1 in ccRCC cell lines[3].
Isothermal titration calorimetry: Human HIF2α-B was expressed and purified as described earlier. Rat HIF2α differs from human HIF2α-B by only three amino acids. So, the expression vector for human HIF2α-B was mutated at these residues (T262L, I 265V, and I326V) to make rat HIF2α-B. Pas-B domain of HIF1α was unstable, and the complex of HIF1α-B*:ARNT-B* was used for isothermal titration calorimetry (ITC). The complex was coexpressed in Escherichia coli harboring pET28-HIF1α-B* and pGB1-ARNT-B* and copurified in a single step of Ni-affinity chromatography. The binding affinity between PT2385 and PAS-B domains was determined using ITC on an iTC200 system. PAS-B at 0.4 mmol/L was titrated into 40 μmol/L of PT2385 in the cell in buffer consisting of 20 mmol/L Tris-HCl, pH 8.0, 150 mmol/L KCl, and 1% DMSO[3]. |
| Cell Assay |
786-O, A498, Hep3B, and Caki-1 cell lines were purchased from ATCC. All were acquired in 2012 and were maintained in culture for no more than 30 continuous passages. The cells were cultured in DMEM supplemented with 10% FBS, 100 units penicillin, and 100 μg/mL streptomycin.[3]
For compound treatment, 5 × 105 cells were plated into 6-well cell culture plates in 2 mL of medium. Compound dissolved in DMSO was added as the cultures reached confluence with the final concentration of DMSO at 0.1%. For hypoxia-treated cells, cell cultures were placed in the chamber supplied with 1% oxygen and 5% CO2 for more then 4 hours before compound addition and maintained under the hypoxic condition for the duration of compound treatment.[3] All relevant human cell lines used in experiments were obtained from ATCC. ATCC authenticated cell lines with short tandem repeat profiling.[3] |
| Animal Protocol |
Animal/Disease Models: SCID/beige mice with 786-O and A498 RCC cell lines [3]
Doses: 30 or 100 mg/kg Route of Administration: po (oral gavage); twice (two times) daily Experimental Results: Caused rapid, dose-dependent tumor regression . |
| References |
[1]. Eli Wallace, Ph.D. PT2385: HIF-2α Antagonist for the Treatment of VHL Mutant ccRCC. 12th International VHL Medical Symposium April 8, 2016.
[2]. Xie C, et al. Activation of intestinal hypoxia-inducible factor 2α during obesity contributes to hepatic steatosis. Nat Med. 2017 Nov;23(11):1298-1308. [3]. Wallace EM, et al. A Small-Molecule Antagonist of HIF2α Is Efficacious in Preclinical Models of Renal Cell Carcinoma. Cancer Res. 2016 Sep 15;76(18):5491-500. |
| Additional Infomation |
PT-2385 is under investigation in clinical trial NCT03108066 (PT2385 for the Treatment of Von Hippel-lindau Disease-associated Clear Cell Renal Cell Carcinoma).
Nonalcoholic fatty liver disease is becoming the most common chronic liver disease in Western countries, and limited therapeutic options are available. Here we uncovered a role for intestinal hypoxia-inducible factor (HIF) in hepatic steatosis. Human-intestine biopsies from individuals with or without obesity revealed that intestinal HIF-2α signaling was positively correlated with body-mass index and hepatic toxicity. The causality of this correlation was verified in mice with an intestine-specific disruption of Hif2a, in which high-fat-diet-induced hepatic steatosis and obesity were substantially lower as compared to control mice. PT2385, a HIF-2α-specific inhibitor, had preventive and therapeutic effects on metabolic disorders that were dependent on intestine HIF-2α. Intestine HIF-2α inhibition markedly reduced intestine and serum ceramide levels. Mechanistically, intestine HIF-2α regulates ceramide metabolism mainly from the salvage pathway, by positively regulating the expression of Neu3, the gene encoding neuraminidase 3. These results suggest that intestinal HIF-2α could be a viable target for hepatic steatosis therapy. |
| Molecular Formula |
C17H12F3NO4S
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| Molecular Weight |
383.34
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| Exact Mass |
383.04
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| Elemental Analysis |
C, 53.27; H, 3.16; F, 14.87; N, 3.65; O, 16.69; S, 8.36
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| CAS # |
1672665-49-4
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| Related CAS # |
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| PubChem CID |
91754484
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| Appearance |
White to off-white solid powder
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| Density |
1.6±0.1 g/cm3
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| Boiling Point |
524.7±50.0 °C at 760 mmHg
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| Flash Point |
271.1±30.1 °C
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| Vapour Pressure |
0.0±1.4 mmHg at 25°C
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| Index of Refraction |
1.613
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| LogP |
1.8
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| SMILES |
CS(C(C=C1)=C2C(CC(F)(F)[ C@H]2O)=C1OC3=CC(C#N)=CC(F)=C3)(=O)=O
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| InChi Key |
ONBSHRSJOPSEGS-INIZCTEOSA-N
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| InChi Code |
InChI=1S/C17H12F3NO4S/c1-26(23,24)14-3-2-13(12-7-17(19,20)16(22)15(12)14)25-11-5-9(8-21)4-10(18)6-11/h2-6,16,22H,7H2,1H3/t16-/m0/s1
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| Chemical Name |
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| Synonyms |
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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 |
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| 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) |
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| Solubility (In Vivo) |
Solubility in Formulation 1: 2.87 mg/mL (7.49 mM) in 5% DMSO + 40% PEG300 + 5% Tween80 + 50% Saline (add these co-solvents sequentially from left to right, and one by one), suspension solution; with sonication.
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 (6.52 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. View More
Solubility in Formulation 3: ≥ 2.5 mg/mL (6.52 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. Solubility in Formulation 4: ≥ 2.5 mg/mL (6.52 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (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 corn oil and mix evenly. |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 2.6087 mL | 13.0433 mL | 26.0865 mL | |
| 5 mM | 0.5217 mL | 2.6087 mL | 5.2173 mL | |
| 10 mM | 0.2609 mL | 1.3043 mL | 2.6087 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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The ceramide-synthesis-related geneNeu3is a novel HIF-2α target gene in the small intestine.Nat Med.2017 Nov;23(11):1298-1308. |
Intestinal HIF-2α deficiency reduces ceramide synthesis in the small intestine. (a) Score scatter plot of a PCA model of the intestinal metabolites betweenHif2αfl/fl(circle) andHif2αΔIE(square) mice.Nat Med.2017 Nov;23(11):1298-1308. |
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