TRβ-1 (EC50 = 0.16 μM); TRα-1 (EC50 = 0.58 μM)
Thyroid hormone receptor beta (TRβ) - selective agonist [1]
- Thyroid hormone receptor alpha (TRα) - weak agonist [1]

In a bacterial biosensor assay using engineered E. coli strains expressing human TRα-1 or TRβ-1 ligand-binding domains (LBDs), GC-1 demonstrated agonistic activity. It induced a dose-dependent growth response in both TRα-1 and TRβ-1 biosensor strains when cultured in thymine-deficient (-Thy) medium, indicating activation of the thymidylate synthase reporter enzyme via ligand binding to the TR LBDs. [1]
- The half-maximal effective concentration (EC50) of GC-1 for the TRα-1 biosensor was determined to be 0.58 μM. For the TRβ-1 biosensor, the EC50 was 0.16 μM. This corresponds to a selectivity ratio (TRα/TRβ) of approximately 3.63, confirming its preferential activity for TRβ. [1]
- Compared to the natural ligand T3 (EC50 = 0.52 μM for TRα-1; 0.58 μM for TRβ-1), GC-1 was approximately 3-fold more potent at TRβ-1 and showed similar potency at TRα-1. [1]
- The agonistic activity and TRβ selectivity of GC-1 observed in this bacterial biosensor system qualitatively match those reported in the literature from mammalian cell reporter gene assays and in vitro binding studies. [1]

Sobetirome (GC-1) is a selective agonist for the thyroid hormone receptor β (TRβ) and liver uptake. In euthyroid mice, sobetirome (48 nmol/kg) lowers levels of very low-density lipoprotein (VLDL) triglycerides and high-density lipoprotein (HDL) cholesterol. In hypercholesterolemic mice, sobetirome lowers triglyceride and HDL cholesterol levels. In hypercholesterolemic mice, sobetirome stimulates the synthesis of bile acid and increases the number of HDL receptors in the liver[2]. Treatment with 10× Sobetirome (GC-1) causes only a 21% (1.7 g) increase in fat mass, while treatment with 20× Sobetirome (GC-1) causes a 20% (1.3 g) decrease in fat mass[3].

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In euthyroid male C57BL/6 mice, daily intraperitoneal (i.p.) administration of GC-1 for 8 days resulted in dose-dependent reductions in serum cholesterol (up to 25% at the highest dose of 97 nmol/kg/day) and serum triglycerides (up to 75% at the highest dose). The cholesterol-lowering effect was stronger for GC-1 than for equimolar doses of T3. [2]
- Lipoprotein profile analysis showed that GC-1 primarily reduced HDL cholesterol levels and reduced VLDL and LDL triglycerides, with no effect on VLDL cholesterol. [2]
- In mice fed a high-cholesterol diet or a diet containing cholesterol and cholic acid, treatment with GC-1 (97 nmol/kg/day for 8 days) markedly attenuated diet-induced hypercholesterolemia, reducing serum cholesterol levels to those seen in chow-fed controls. [2]
- GC-1 treatment significantly increased the protein expression of the hepatic HDL receptor SR-BI in liver membranes, as detected by immunoblotting. This effect was stronger than that observed with T3. SR-BI mRNA levels were either unaffected or reduced, indicating post-transcriptional regulation. [2]
- GC-1 stimulated reverse cholesterol transport (RCT) by increasing bile acid synthesis. It increased serum levels of 7α-hydroxy-4-cholesten-3-one (C4), a marker of CYP7A1 activity, and increased fecal excretion of bile acids. Hepatic CYP7A1 mRNA levels were generally increased. [2]
- GC-1 treatment did not significantly affect the expression or activity of the LDL receptor, nor did it alter hepatic cholesterol levels or the expression of SREBP-2 and its target genes (e.g., HMG-CoA reductase). [2]
- Hepatic triglyceride content was reduced by GC-1 treatment in mice fed a high-cholesterol diet. [2]
- GC-1 reduced hepatic mRNA levels of SHP (small heterodimer partner), which may contribute to the upregulation of CYP7A1 by relieving feedback inhibition. [2]

CYP7A1 Activity (Surrogate Marker - C4): The activity of cholesterol 7α-hydroxylase (CYP7A1), the rate-limiting enzyme in bile acid synthesis, was assessed indirectly by measuring serum levels of 7α-hydroxy-4-cholesten-3-one (C4). Serum samples were pooled for each group. C4 was analyzed by HPLC after solid-phase extraction, using 7β-hydroxy-4-cholesten-3-one as an internal standard. [2]
- Fecal Bile Acid and Neutral Sterol Analysis: Feces were collected from mice for 24 hours before and during treatment. Pooled dried feces (1 gram) from each group were analyzed for neutral sterols (coprostanol, coprostanone, and cholesterol) and bile acids by quantitative gas-liquid chromatography. [2]

Bacterial Biosensor Assay for TR Agonism: The assay utilizes E. coli strain D1210ΔthyA, which is deficient in thymidylate synthase (TS) and requires thymine for growth. This strain was transformed with plasmids encoding chimeric biosensor proteins. For TRα-1, the plasmid pMIT::TRα-1 expressed a fusion protein consisting of maltose-binding protein (MBP), a mini-intein splicing domain with the human TRα-1 LBD (amino acids E149 to D407) inserted, and the T4 TS reporter enzyme. For TRβ-1, the plasmid pMIT::TRβ-1 expressed an analogous fusion protein containing the human TRβ-1 LBD (amino acids E203 to D461). [1]
- Fresh transformant colonies were used to inoculate LB medium with ampicillin and thymine. Cultures were grown to an OD600 of 1.3-1.7, then diluted 1:200 into -Thy medium (a defined minimal medium lacking thymine). The diluted cells were aliquoted into 96-well plates (198 μL/well). Test compounds, including GC-1, were dissolved in DMSO and added to wells (2 μL/well) to achieve desired final concentrations, with the DMSO concentration kept constant at 1% (v/v). Plates were incubated at 34°C with agitation (150 rpm) and 80% humidity. Cell growth was monitored over time by measuring OD600 using a spectrophotometer. Increased growth in -Thy medium indicates ligand binding to the TR LBD, which allosterically activates the TS reporter enzyme. [1]
- To confirm the specificity of the growth phenotype, cells were also grown in TTM medium (-Thy medium supplemented with thymine and trimethoprim). In this medium, ligand binding leads to decreased growth, providing a mirror-image confirmation of TS-specific effects. [1]
- For EC50 determination, dose-response curves were generated by testing GC-1 at various concentrations via serial dilution. Growth rates were normalized and fitted to a standard sigmoidal dose-response equation using nonlinear regression. The selectivity for TRβ over TRα was calculated as the ratio of EC50(TRα) to EC50(TRβ). Each dose-response series was performed in triplicate on each plate, and results from three separate plates were used to calculate final EC50 values. [1]

Animals:** 3-month-old male C57BL/6 mice were used. They were kept in standardized conditions with free access to water and normal chow. [2]
- **Drug Preparation and Administration:** GC-1 was dissolved in 20% DMSO or propylene glycol and administered via daily intraperitoneal (i.p.) injection at 9:00 a.m. for 8 days. Control mice received vehicle only. [2]
- **Dose-Response Study:** Nine groups of six mice were treated with vehicle, or 5.4, 24, 48, and 97 nmol/kg/day of either T3 or GC-1. [2]
- **Diet-Induced Hypercholesterolemia Study:** One group of seven mice was given no additional supplementation. Three groups of seven mice received a diet containing 10% corn oil and 2% cholesterol (cholesterol diet). Three groups of seven mice received a diet containing 10% corn oil, 2% cholesterol, and 0.5% cholic acid (cholic acid diet). For each diet, one group was treated with vehicle, one with GC-1 (97 nmol/kg/day), and one with T3 (97 nmol/kg/day). [2]
- **Bile Acid Excretion Study:** Three groups of five chow-fed mice were treated with vehicle, 48 nmol/kg/day of GC-1, or T3 for 5 days. Feces were collected group-wise 24 hours before treatment and during the last 24 hours of the experiment. [2]
- **Tissue Collection:** Food was withdrawn 5 hours before sacrifice. Blood was drawn by cardiac puncture under light isoflurane anesthesia. Animals were killed by cervical dislocation. Livers were immediately frozen in liquid nitrogen. [2]

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Animals: 3-month-old male C57BL/6 mice were used. They were kept in standardized conditions with free access to water and normal chow. [2]
- Drug Preparation and Administration: GC-1 was dissolved in 20% DMSO or propylene glycol and administered via daily intraperitoneal (i.p.) injection at 9:00 a.m. for 8 days. Control mice received vehicle only. [2]
- Dose-Response Study: Nine groups of six mice were treated with vehicle, or 5.4, 24, 48, and 97 nmol/kg/day of either T3 or GC-1. [2]
- Diet-Induced Hypercholesterolemia Study: One group of seven mice was given no additional supplementation. Three groups of seven mice received a diet containing 10% corn oil and 2% cholesterol (cholesterol diet). Three groups of seven mice received a diet containing 10% corn oil, 2% cholesterol, and 0.5% cholic acid (cholic acid diet). For each diet, one group was treated with vehicle, one with GC-1 (97 nmol/kg/day), and one with T3 (97 nmol/kg/day). [2]
- Bile Acid Excretion Study: Three groups of five chow-fed mice were treated with vehicle, 48 nmol/kg/day of GC-1, or T3 for 5 days. Feces were collected group-wise 24 hours before treatment and during the last 24 hours of the experiment. [2]
- Tissue Collection: Food was withdrawn 5 hours before sacrifice. Blood was drawn by cardiac puncture under light isoflurane anesthesia. Animals were killed by cervical dislocation. Livers were immediately frozen in liquid nitrogen. [2]

The study notes that GC-1 was designed to avoid the cardiac side effects (e.g., tachycardia) associated with non-selective TR activation, and previous work has shown it can lower plasma lipids without obvious adverse effects on the heart, bone, or muscle. [2]

[1]. Bacterial biosensors for screening isoform-selective ligands for human thyroid receptors α-1 and β-1. FEBS Open Bio. 2012 Aug 15;2:247-53.

[2]. Selective thyroid receptor modulation by GC-1 reduces serum lipids and stimulates steps of reverse cholesterol transport in euthyroid mice. Proc Natl Acad Sci U S A. 2005 Jul 19;102(29):10297-302.

[3]. Thyroid hormone receptor beta-specific agonist GC-1 increases energy expenditure and prevents fat-mass accumulation in rats. J Endocrinol. 2007 Apr;193(1):21-9.

Sobetirome is a diarylmethane compound. It is a liver-selective analog of the β1 subtype of thyroid hormone receptor, possessing lipid-lowering and anti-atherosclerotic activities. In animal models, Sobetirome lowers blood lipids and cholesterol levels, and stimulates reverse cholesterol transport, promoting the reverse transport of cholesterol from atherosclerotic macrophages back to the liver for excretion. In human trials, Sobetirome lowers plasma LDL cholesterol and plasma triglyceride levels; its liver-selective action helps avoid many of the side effects common to other thyroid hormone analogs.

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GC-1 (3,5-dimethyl-4-(4'-hydroxy-3'-isopropylbenzyl)-phenoxy acetic acid) is a synthetic thyromimetic compound developed as a potentially therapeutic isoform-selective TRβ agonist. [1]
- It is designed to lower LDL cholesterol levels without the cardiovascular side effects (e.g., tachycardia) associated with non-selective thyroid hormone receptor activation, due to its selectivity for the TRβ isoform, which is dominant in the liver, over the TRα isoform, which is dominant in the heart. [1]
- The study validates the use of a simple, economical bacterial biosensor system to detect the agonistic activity and subtype selectivity of GC-1. The results obtained (EC50 = 0.58 μM for TRα-1; 0.16 μM for TRβ-1; selectivity ~3.6-fold for TRβ) are qualitatively consistent with more complex and expensive mammalian cell-based assays, demonstrating the utility of this method for primary screening of potential TR subtype-selective therapeutics. [1]

C20H24O4

328.40216

328.167

C, 73.15; H, 7.37; O, 19.49

211110-63-3

211110-63-3

9862248

White to yellow solid powder

1.152g/cm3

510.156ºC at 760 mmHg

178.945ºC

0mmHg at 25°C

1.575

4.186

2

4

6

24

396

0

O=C(O)COC1=CC(C)=C(CC2=CC=C(O)C(C(C)C)=C2)C(C)=C1

QNAZTOHXCZPOSA-UHFFFAOYSA-N

InChI=1S/C20H24O4/c1-12(2)17-9-15(5-6-19(17)21)10-18-13(3)7-16(8-14(18)4)24-11-20(22)23/h5-9,12,21H,10-11H2,1-4H3,(H,22,23)

2-[4-[(4-hydroxy-3-propan-2-ylphenyl)methyl]-3,5-dimethylphenoxy]acetic acid

Sobetirome; GC-1; GC 1; GC1; QRX-431; QRX431; QRX 431

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: ~100 mg/mL (~304.5 mM)

Solubility in Formulation 1: ≥ 2.5 mg/mL (7.61 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.

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Solubility in Formulation 2: ≥ 2.5 mg/mL (7.61 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.

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Solubility in Formulation 3: ≥ 2.5 mg/mL (7.61 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 of corn oil and mix evenly.

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