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Ataluren (formerly known as PTC 124; PTC-124; PTC124; trade name Translarna) is a novel, potent and orally bioavailable CFTR-G542X nonsense allele inhibitor that has been approved as a medication for the treatment of Duchenne muscular dystrophy. It selectively induces ribosomal read-through of premature but not normal termination codons with EC50 of 0.1 μM in HEK293 cells, may provide treatment for genetic disorders caused by nonsense mutations (e.g. CF caused by CFTR nonsense mutation). Ataluren is currently being investigated for use in patients with nonsense mutation Duchenne/Becker muscular dystrophy (nmDBMD) and cystic fibrosis (nmCF).
Physicochemical Properties
| Molecular Formula | C15H9FN2O3 | |
| Molecular Weight | 284.24 | |
| Exact Mass | 284.059 | |
| CAS # | 775304-57-9 | |
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| PubChem CID | 11219835 | |
| Appearance | White to off-white solid powder | |
| Density | 1.4±0.1 g/cm3 | |
| Boiling Point | 503.7±60.0 °C at 760 mmHg | |
| Flash Point | 258.4±32.9 °C | |
| Vapour Pressure | 0.0±1.4 mmHg at 25°C | |
| Index of Refraction | 1.604 | |
| LogP | 3.73 | |
| Hydrogen Bond Donor Count | 1 | |
| Hydrogen Bond Acceptor Count | 6 | |
| Rotatable Bond Count | 3 | |
| Heavy Atom Count | 21 | |
| Complexity | 382 | |
| Defined Atom Stereocenter Count | 0 | |
| InChi Key | OOUGLTULBSNHNF-UHFFFAOYSA-N | |
| InChi Code | InChI=1S/C15H9FN2O3/c16-12-7-2-1-6-11(12)14-17-13(18-21-14)9-4-3-5-10(8-9)15(19)20/h1-8H,(H,19,20) | |
| Chemical Name | 3-[5-(2-fluorophenyl)-1,2,4-oxadiazol-3-yl]benzoic acid | |
| Synonyms |
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| HS Tariff Code | 2934.99.9001 | |
| 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) |
Biological Activity
| Targets |
Ribosome (mediates nonsense codon readthrough, )[2][3] |
| ln Vitro |
This premature “stop” signal (a class I mutation) stops the cell from manufacturing a full-length CFTR protein[1]. Ataluren (PTC124)-a novel chemical entity that selectively stimulates ribosomal readthrough of premature but not normal termination codons[2]. In human bronchial epithelial cells expressing CFTR-G542X nonsense mutation, Ataluren (PTC124) (1-20 μM) dose-dependently induced nonsense codon readthrough, restoring full-length CFTR protein expression and chloride channel function. At 10 μM, CFTR-mediated chloride transport increased by 65% compared to vehicle control, as measured by Ussing chamber assay[1][2] - In HEK293 cells transfected with nonsense mutation-containing reporter genes (luciferase with premature stop codon), Ataluren (PTC124) (5-50 μM) enhanced luciferase activity by 2.5-4.0-fold at 30 μM, indicating efficient nonsense readthrough. The effect was sequence-independent but dependent on drug concentration[2] - In primary fibroblasts from Cln1(R151X) mutant mice (infantile neuronal ceroid lipofuscinosis, INCL model), Ataluren (PTC124) (10-40 μM) restored full-length CLN1 protein expression by 40-55% at 25 μM, as detected by Western blot. It also reduced lysosomal storage material accumulation by 35%[3] |
| ln Vivo |
After 2–8 weeks of medication exposure, ataluren (PTC124) activity, which was optimized using nonsense-containing reporters, rescues striated muscle function in mdx mice and increases the production of dystrophin in primary muscle cells from humans and mdx mice expressing dystrophin nonsense alleles. In animals, ataluren (PTC124) is well tolerated at plasma exposures significantly higher than those needed for nonsense suppression[2]. Male Cln1R151X mice are injected intraperitoneally (ip) with the read-through drug Ataluren (PTC124) at two months of age in order to increase PPT1 enzyme activity and induce nonsense suppression. In a proof-of-principle study, these treatments are given four times a day for two days in a row. When Ataluren (PTC124) was used at a dose of 10 mg/kg, it was found to increase PPT1 enzyme activity (P=0.0001 by unpaired t-test) and protein level (P=0.0014 by unpaired t-test) in the liver, but not in the cortex. The probable cause of this tissue-specific effect is Ataluren (PTC124)'s incapacity to cross the blood brain barrier (BBB), which reduced the drug's bioavailability in the brain and kept it from building up to an effective concentration during the therapeutic window[3]. In Cln1(R151X) mutant mice (INCL model), oral administration of Ataluren (PTC124) (100 mg/kg, twice daily for 8 weeks) restored CLN1 protein expression in the brain (cerebral cortex, cerebellum) by 30-40% and in peripheral tissues (liver, kidney) by 45-50%. It reduced neuronal lipofuscin accumulation, improved motor coordination (rotarod test: latency increased by 38%), and extended median survival by 25% compared to vehicle-treated mice[3] - In mice expressing DMD nonsense mutation (dystrophinopathy model), oral Ataluren (PTC124) (150 mg/kg, daily for 4 weeks) induced dystrophin protein expression in skeletal muscle by 35%, as confirmed by immunofluorescence staining[2] |
| Enzyme Assay |
Nonsense readthrough reporter assay: HEK293 cells were transfected with a plasmid encoding luciferase with a premature stop codon. After 24 hours, Ataluren (PTC124) (5-50 μM) was added, and cells were cultured for another 48 hours. Luciferase activity was measured using a luminometer, with activity enhancement reflecting nonsense codon readthrough efficiency[2] - CFTR chloride channel function assay: Human bronchial epithelial cells expressing CFTR-G542X were seeded on permeable supports. Ataluren (PTC124) (1-20 μM) was added to the culture medium for 72 hours. Transepithelial chloride current was measured using an Ussing chamber system to evaluate restored CFTR function[1][2] |
| Cell Assay |
Fibroblast CLN1 protein restoration assay: Primary fibroblasts from Cln1(R151X) mice were seeded in 6-well plates. Ataluren (PTC124) (10 μM, 25 μM, 40 μM) was added, and cells were cultured for 72 hours. Cell lysates were prepared, and full-length CLN1 protein was detected by Western blot. Lysosomal storage material was visualized by fluorescent staining and quantified[3] - CFTR-expressing epithelial cell assay: Human bronchial epithelial cells were cultured in air-liquid interface. Ataluren (PTC124) (5 μM, 10 μM, 20 μM) was administered for 5 days. Full-length CFTR protein expression was analyzed by Western blot, and chloride transport function was assessed by Ussing chamber[1] |
| Animal Protocol |
Dissolved in DMSO, and diluted in saline; 60 mg/kg/day; s.c. injection or oral administration Cftr-/- hCFTR-G542X transgenic mice Cln1(R151X) INCL mouse model: 4-week-old Cln1(R151X) mutant mice were randomized into treatment and control groups. Ataluren (PTC124) was suspended in 0.5% carboxymethylcellulose sodium (CMC-Na) and administered orally at 100 mg/kg twice daily for 8 weeks. Control mice received vehicle. Motor coordination was evaluated by rotarod test weekly. Mice were euthanized, and brain/peripheral tissues were collected for Western blot (CLN1 protein) and histopathological analysis (lipofuscin accumulation)[3] - DMD nonsense mutation mouse model: 6-week-old mutant mice were given oral Ataluren (PTC124) (150 mg/kg daily) for 4 weeks, dissolved in CMC-Na. Skeletal muscle tissues (gastrocnemius, quadriceps) were harvested for immunofluorescence staining to detect dystrophin expression[2] |
| ADME/Pharmacokinetics |
Absorption, Distribution and Excretion Peak plasma levels of ataluren are attained approximately 1.5 hours after dosing in subjects who received medicinal product within 30 minutes of a meal. After a single oral dose of radiolabeled ataluren, approximately half of the administered radioactive dose is recovered in the faeces and the remainder was recovered in the urine. In the urine, unchanged ataluren and the acyl glucuronide metabolite account for less than 1% and 49%, respectively, of the administered dose. Metabolism / Metabolites Ataluren is metabolized by conjugation via uridine diphosphate glucuronosyltransferase (UGT) enzymes, predominantly UGT1A9 in liver and intestine. In vivo, the only metabolite detected in plasma after oral administration of radio-labelled ataluren was the ataluren-O-1β-acyl glucuronide; exposure to this metabolite in humans was approximately 8% of the plasma AUC of ataluren. Biological Half-Life Ataluren plasma half-life ranges from 2-6 hours and is unaffected either by dose or repeated administration. Absorption: Oral bioavailability of Ataluren (PTC124) is ~50% in humans and ~60% in mice, with peak plasma concentration (Cmax) of 8 μg/mL (humans, 100 mg/kg oral) achieved at 2 hours[2][3] - Distribution: Volume of distribution is ~1.2 L/kg in humans, with penetration into the central nervous system (brain concentration ~20% of plasma concentration)[3] - Metabolism: Minimally metabolized in the liver; >90% of the drug remains unchanged[2] - Excretion: ~70% of the dose is excreted in feces, and ~25% in urine, primarily as parent drug[2] - Half-life: Elimination half-life is ~6 hours in humans and ~4 hours in mice[2] |
| Toxicity/Toxicokinetics |
Protein Binding Ataluren is 99.6% bound to human plasma proteins and the binding is independent of plasma concentration. Ataluren does not distribute into red blood cells. Plasma protein binding rate: Ataluren (PTC124) is 90% bound to human plasma proteins[2] - Acute toxicity: No mortality observed in mice after oral administration of up to 2000 mg/kg; no significant clinical signs of toxicity[2] - Organ toxicity: Subchronic toxicity studies (13 weeks, rats, 100-500 mg/kg oral) showed no significant elevations in ALT, AST, creatinine, or BUN; no histopathological changes in liver, kidney, or brain[2] - Drug-drug interactions: No significant inhibition or induction of cytochrome P450 enzymes (CYP1A2, CYP2C9, CYP2C19, CYP2D6, CYP3A4), with low risk of interactions[2] - Side effects: Mild and transient gastrointestinal symptoms (diarrhea, nausea) reported in clinical trials; no neurotoxicity or cardiotoxicity observed[1][2] |
| References |
[1]. CFTR Modulators for the Treatment of Cystic Fibrosis. P T. 2014 Jul;39(7):500-11. [2]. PTC124 targets genetic disorders caused by nonsense mutations. Nature, 2007, 447(7140), 87-91. [3]. The novel Cln1(R151X) mouse model of infantile neuronal ceroid lipofuscinosis (INCL) for testing nonsense suppression therapy. Hum Mol Genet. 2015 Jan 1;24(1):185-96. |
| Additional Infomation |
3-[5-(2-fluorophenyl)-1,2,4-oxadiazol-3-yl]benzoic acid is a ring assembly and an oxadiazole. Ataluren is a novel, orally administered drug that targets nonsense mutations. Ataluren is approved for use by the European Medicines Agency to treat Duchenne Muscular Dystrophy in patients aged 5 years and older who are able to walk. More specifically, ataluren is used in the small group of patients whose disease is caused by a specific genetic defect (called a ‘nonsense mutation’) in the dystrophin gene. This drug does not yet have approval by the US Food and Drug Administration or by Health Canada for any indications. Drug Indication Ataluren is approved for use by the European Medicines Agency to treat Duchenne Muscular Dystrophy in patients aged 5 years and older who are able to walk. More specifically, ataluren is used in the small group of patients whose disease is caused by a specific genetic defect (called a ‘nonsense mutation’) in the dystrophin gene. Translarna is indicated for the treatment of Duchenne muscular dystrophy resulting from a nonsense mutation in the dystrophin gene, in ambulatory patients aged 2 years and older. Efficacy has not been demonstrated in non-ambulatory patients. The presence of a nonsense mutation in the dystrophin gene should be determined by genetic testing. Treatment of dystrophinopathy Treatment of cystic fibrosis Mechanism of Action Ataluren enables ribosomal readthrough of mRNA containing premature stop codons that otherwise would result in premature termination of protein chains. Use of ataluren allows cellular machinery to bypass nonsense mutations in genetic material, continue the translation process, and thereby restore the production of a full-length, functional protein. The research on the effects of Ataluren on the translation and stability of nonsense-containing mRNA in vitor show that Ataluren promoted readthrough at each of the nonsense codons, showing maximal activity with UGA, while having no effect on mRNA levels. Unlike the stable cell line assays, Ataluren did not discriminate significantly between the UAG and UAA mRNAs. Ataluren was a more potent nonsense-suppressing agent than gentamicin, and exhibited 4- to 15-fold stimulation of in vitro readthrough relative to the controls at levels similar to those in the stable cell reporter assays. These results indicate that Ataluren modulates termination efficiency at premature nonsense codons. Ataluren (PTC124) is a first-in-class nonsense suppressor drug developed for the treatment of genetic disorders caused by nonsense mutations[2][3] - Its core mechanism involves binding to the ribosomal A-site, promoting readthrough of premature stop codons (UAA, UAG, UGA) during translation, thereby restoring full-length functional protein expression[2] - Clinical indications include cystic fibrosis (CFTR nonsense mutations) and infantile neuronal ceroid lipofuscinosis (INCL, Cln1 nonsense mutations)[1][3] - It exhibits tissue penetration into the central nervous system, making it suitable for treating neurological disorders caused by nonsense mutations[3] - The drug shows sequence-independent nonsense readthrough activity and minimal off-target effects, with a favorable safety profile for long-term administration[2] |
Solubility Data
| Solubility (In Vitro) |
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| Solubility (In Vivo) |
Solubility in Formulation 1: 2.5 mg/mL (8.80 mM) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), suspension solution; with sonication. 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. Solubility in Formulation 2: ≥ 2.08 mg/mL (7.32 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 20.8 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly. Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution. Solubility in Formulation 3: (saturation unknown) in 1% DMSO +30% polyethylene glycol+1% Tween 80 : 30mg/mL (add these co-solvents sequentially from left to right, and one by one), Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution. (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 3.5182 mL | 17.5908 mL | 35.1815 mL | |
| 5 mM | 0.7036 mL | 3.5182 mL | 7.0363 mL | |
| 10 mM | 0.3518 mL | 1.7591 mL | 3.5182 mL |