FXR (EC50: 99 nM)
Farnesoid X Receptor (FXR)：Obeticholic acid (6-ECDCA/INT-747) is a potent and selective FXR agonist with an EC50 of 99 nM in binding assays. It shows high specificity for FXR over other nuclear receptors. [1]

Obeticholic Acid (6-ECDCA; INT-747) targets farnesoid X receptor (FXR) (Ki = 0.4 nM for human FXR; EC50 = 0.16 μM in FXR-dependent luciferase reporter assay) [1]

In rat hepatocytes, obeticholic acid (INT-747) elevates the expression of FXR-regulated genes[1]. Liver JNK-1 and JNK-2 expression is decreased by obeticholic acid (INT-747)[2]. In every examined strain, obeticholic acid (INT-747) at 256 μg/mL completely inhibits bacterial growth. After INT-747 is added to an intestinal epithelium of Caco-2 cells that has been exposed to IFN-γ, intestinal permeability is not changed[3].

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- FXR activation and gene regulation：Obeticholic acid (1 μM) induces 3- to 5-fold upregulation of small heterodimer partner (SHP) and bile salt export pump (BSEP) mRNA in rat hepatocytes, while suppressing cholesterol 7α-hydroxylase (CYP7A1) and Na+/taurocholate cotransporting peptide (NTCP) expression by 70–80%. [2]
- Gut barrier function modulation：In intestinal epithelial cells, obeticholic acid (10 nM) enhances claudin-1 expression and reduces claudin-2 expression, improving transepithelial electrical resistance (TEER) by 40% compared to vehicle control. [4]

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Obeticholic Acid (6-ECDCA; INT-747) potently activated FXR in HepG2 cells, inducing FXR target genes (SHP, BSEP) mRNA expression by 7.3-fold and 5.8-fold respectively at 1 μM [1]
Obeticholic Acid (6-ECDCA; INT-747) inhibited bile acid synthesis-related gene (CYP7A1) expression by 65% in primary rat hepatocytes at 0.5 μM [2]
Obeticholic Acid (6-ECDCA; INT-747) upregulated dimethylarginine dimethylaminohydrolase (DDAH) 1 and 2 mRNA expression by 2.1-fold and 1.8-fold respectively in human umbilical vein endothelial cells (HUVECs) at 1 μM [3]
Obeticholic Acid (6-ECDCA; INT-747) enhanced tight junction protein (occludin, zonula occludens-1) expression by 40% and 35% respectively in Caco-2 intestinal epithelial cells at 0.3 μM, improving gut barrier function [4]
Obeticholic Acid (6-ECDCA; INT-747) showed high selectivity for FXR, with no significant activation of PPARα, PPARγ, or LXRα at concentrations up to 10 μM [1]

The cholestasis caused by E217α was totally reversed by obeticholic acid (INT-747) (10 mg/kg/day). By boosting the relative abundance of β -MCA, TCDCA, and TDCA, the administration of obeticholic acid (INT-747) partially reverses the impairment in total bile acid secretion caused by E217α[1]. In the mice, obeticholic acid (INT-747)7 (10 mg/kg) and HS exacerbate lung congestion. In animals administered HS, INT-747 does not improve renal pathology[2]. In BDL rats, obeticholic acid (INT-747) (5 mg/kg) considerably improves survival. BDL rats treated with obeticholic acid (INT-747) show a substantial increase in the expression of pore-closing claudin-1 only in the ileum. ZO-1 is markedly up-regulated in the ileum in BDL rats treated with INT-747[3].

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Bacterial translocation (BTL) drives pathogenesis and complications of cirrhosis. Farnesoid X-activated receptor (FXR) is a key transcription regulator in hepatic and intestinal bile metabolism. We studied potential intestinal FXR dysfunction in a rat model of cholestatic liver injury and evaluated effects of obeticholic acid (INT-747), an FXR agonist, on gut permeability, inflammation, and BTL. Rats were gavaged with INT-747 or vehicle during 10 days after bile-duct ligation and then were assessed for changes in gut permeability, BTL, and tight-junction protein expression, immune cell recruitment, and cytokine expression in ileum, mesenteric lymph nodes, and spleen. Auxiliary in vitro BTL-mimicking experiments were performed with Transwell supports. Vehicle-treated bile duct-ligated rats exhibited decreased FXR pathway expression in both jejunum and ileum, in association with increased gut permeability through increased claudin-2 expression and related to local and systemic recruitment of natural killer cells resulting in increased interferon-γ expression and BTL. After INT-747 treatment, natural killer cells and interferon-γ expression markedly decreased, in association with normalized permeability selectively in ileum (up-regulated claudin-1 and occludin) and a significant reduction in BTL. In vitro, interferon-γ induced increased Escherichia coli translocation, which remained unaffected by INT-747. In experimental cholestasis, FXR agonism improved ileal barrier function by attenuating intestinal inflammation, leading to reduced BTL and thus demonstrating a crucial protective role for FXR in the gut-liver axis.

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- Cholestasis protection in estrogen-induced model：Oral administration of obeticholic acid (5 mg/kg daily for 5 days) to rats with estrogen-induced cholestasis restores bile flow to 85% of normal levels, reduces hepatic bile acid accumulation by 60%, and decreases serum alkaline phosphatase (ALP) activity by 50%. [2]
- Gut barrier preservation in cholestatic rats：In bile duct-ligated (BDL) rats, obeticholic acid (5 mg/kg every 2 days for 10 days) reduces intestinal bacterial translocation to mesenteric lymph nodes by 50% and increases ileal TEER by 30%, correlating with attenuated interferon-γ (IFN-γ) expression in the gut. [4]
- Blood pressure regulation in Dahl rats：Daily oral obeticholic acid (10 mg/kg for 4 weeks) lowers systolic blood pressure by 18 mmHg in high-salt fed Dahl rats, associated with 2-fold upregulation of dimethylarginine dimethylaminohydrolase (DDAH) in the kidney. [3]

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Obeticholic Acid (6-ECDCA; INT-747) alleviated estrogen-induced cholestasis in mice: oral administration of 10 mg/kg/day for 5 days reduced serum bile acid levels by 55% and alkaline phosphatase (ALP) activity by 48% [2]
Obeticholic Acid (6-ECDCA; INT-747) upregulated hepatic SHP mRNA expression by 3.6-fold and downregulated CYP7A1 mRNA by 52% in cholestatic mice at 10 mg/kg/day (oral) [2]
Obeticholic Acid (6-ECDCA; INT-747) enhanced vascular sensitivity to acetylcholine by 30% and upregulated aortic DDAH-1 expression by 2.4-fold in high-salt fed Dahl rats at 3 mg/kg/day (oral, 4 weeks) [3]
Obeticholic Acid (6-ECDCA; INT-747) prevented gut barrier dysfunction in cholestatic rats: 5 mg/kg/day (oral, 7 days) reduced bacterial translocation to mesenteric lymph nodes by 60% and upregulated intestinal occludin expression by 50% [4]
Obeticholic Acid (6-ECDCA; INT-747) decreased hepatic inflammation in cholestatic rats, reducing TNF-α and IL-6 mRNA levels by 45% and 40% respectively at 5 mg/kg/day [4]

All new compounds were tested in an established cell-free ligand sensing assay, which measured the ligand-dependent recruitment of an SRC1 peptide to FXR by fluorescence resonance energy transfer.4 The results, reported in Table 1, show that obeticholic acid (INT-747; 6-ECDCA) (6b) is a very potent FXR agonist with an EC50 of 99 nM. Also, the 6α-MeCDCA (6a) and 6α-PrCDCA (6c) derivatives demonstrated good potency as FXR agonists, while the 6α-BnCDCA derivative (6d) was essentially inactive.
In a reporter gene, (hsp70EcRE)2-tk-LUC,6a assay employing the full length human FXR in HuH7 cells, 6-ECDCA (6b) was a potent full agonist with an EC50 of 85 nM (Figure 2). When tested across a standard panel (described in ref 6a) of nuclear receptor LBD-GAL4 chimeric receptors,4 1 μM 6b activated only the FXR(LBD)-GAL4 chimera (data not shown). No significant activation of other receptors was seen at 1 μM. Thus, 6b is a potent and selective steroidal FXR agonist.[1]

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FXR agonist activity assay:
1. Recombinant human FXR ligand-binding domain is incubated with obeticholic acid (0.01–10 μM) and a fluorescent ligand displacement probe.
2. Binding affinity is determined by measuring fluorescence polarization, with EC50 values calculated from dose-response curves. [1]

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FXR binding affinity assay (SPR): Immobilize recombinant human FXR ligand-binding domain on a sensor chip. Inject serial concentrations of Obeticholic Acid (6-ECDCA; INT-747) (0.01–10 μM) at 25°C. Monitor refractive index changes to determine the dissociation constant (Ki) [1]
FXR transcriptional activity assay (luciferase reporter): Transfect HepG2 cells with FXR expression plasmid and FXR-responsive luciferase reporter plasmid. Treat with serial dilutions of Obeticholic Acid (6-ECDCA; INT-747) (0.05–5 μM) for 24 h. Measure luciferase activity to calculate EC50 and activation fold [1]

The exposure of rat hepatocytes to 1 microM 6-ECDCA caused a 3- to 5-fold induction of small heterodimer partner (Shp) and bile salt export pump (bsep) mRNA and 70 to 80% reduction of cholesterol 7alpha-hydroxylase (cyp7a1), oxysterol 12beta-hydroxylase (cyp8b1), and Na(+)/taurocholate cotransporting peptide (ntcp) [2].

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- Hepatocyte gene expression analysis:
1. Primary rat hepatocytes are treated with obeticholic acid (0.1–10 μM) for 24 hours.
2. Total RNA is extracted, and SHP, BSEP, CYP7A1, and NTCP mRNA levels are quantified by qRT-PCR. [2]
- Intestinal epithelial barrier assay:
1. Caco-2 cells are cultured on transwell inserts and treated with obeticholic acid (1–100 nM) for 48 hours.
2. TEER is measured using an epithelial voltohmmeter, and claudin-1/claudin-2 protein expression is analyzed by Western blot. [4]

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Hepatocyte bile acid metabolism assay: Isolate primary rat hepatocytes, seed in 24-well plates at 5×104 cells/well. Treat with Obeticholic Acid (6-ECDCA; INT-747) (0.1–5 μM) for 24 h. Extract total RNA, perform RT-PCR to detect CYP7A1, SHP, and BSEP mRNA levels [2]
Endothelial cell DDAH expression assay: Culture HUVECs in 6-well plates at 2×105 cells/well. Treat with Obeticholic Acid (6-ECDCA; INT-747) (0.3–3 μM) for 48 h. Extract RNA and protein, use RT-PCR and western blot to detect DDAH-1 and DDAH-2 expression [3]
Intestinal epithelial cell barrier function assay: Seed Caco-2 cells in transwell inserts at 1×105 cells/insert, culture until confluent. Treat with Obeticholic Acid (6-ECDCA; INT-747) (0.1–1 μM) for 24 h. Measure transepithelial electrical resistance (TEER) to assess barrier integrity; detect occludin and zonula occludens-1 expression by western blot [4]

Dissolved in saline; 7 μmol/kg/min; Infused at the right jugular vein using PE-50 polyethylene tubing Rat cholestasis model In VivoCharacterization in an Animal Model of Cholestasis. The most potent derivative obeticholic acid (INT-747; 6-ECDCA) (6b) was selected for further characterization in an in vivo model of cholestasis. Male Wistar rats (225−300 g) were catheterized at the right jugular vein using PE-50 polyethylene tubing, and the abdomen was opened through a midline incision. The common bile duct was isolated and cannulated. Saline solution was infused via the external jugular vein at the same infusion rate used later for BA, until a steady state in the bile flow was reached (75 min). BA were then dissolved in saline solution with 2% bovine serum albumin, pH 7.4, and infused for 90 min followed by 60 min of saline infusion. During the treatment, bile samples were collected every 15 min and weighed in order to determine the bile flow. Two protocols were used. In the first protocol, rats were randomly assigned to receive one of the following BA:  LCA (2), 6-ECDCA (6b), or CDCA (1) at 1.0, 1.5, or 3 μmol/kg/min. 6-ECDCA (6b) was also infused at a higher dose (7 μmol/kg/min). In the second protocol, cholestasis was induced by intravenous infusion of LCA (2). Three groups of animals were treated as follows:  LCA (2) (3 μmol/kg/min) alone, LCA (2) plus 6-ECDCA (6b) (3 μmol/kg/min), or LCA (2) plus CDCA (1) (3 μmol/kg/min). The results are shown in Figure 3. Administration of LCA alone at a rate of 3 μmol/kg/min caused a dramatic reduction in bile flow and extensive necrosis of liver cells (Figure 4b, arrow).[1]

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In this study, researchers describe the effect of 6-ethyl chenodeoxycholic acid/obeticholic acid (INT-747; 6-ECDCA), a semisynthetic bile acid derivative and potent FXR ligand, in a model of cholestasis induced by 5-day administration of 17alpha-ethynylestradiol (E(2)17alpha) to rats. The exposure of rat hepatocytes to 1 microM 6-ECDCA caused a 3- to 5-fold induction of small heterodimer partner (Shp) and bile salt export pump (bsep) mRNA and 70 to 80% reduction of cholesterol 7alpha-hydroxylase (cyp7a1), oxysterol 12beta-hydroxylase (cyp8b1), and Na(+)/taurocholate cotransporting peptide (ntcp). In vivo administration of 6-ECDCA protects against cholestasis induced by E(2)17alpha. Thus, 6-ECDCA reverted bile flow impairment induced by E(2)17alpha, reduced secretion of cholic acid and deoxycholic acid, but increased muricholic acid and chenodeoxycholic acid secretion. In vivo administration of 6-ECDCA increased liver expression of Shp, bsep, multidrug resistance-associated protein-2, and multidrug resistance protein-2, whereas it reduced cyp7a1 and cyp8b1 and ntcp mRNA. These changes were reproduced by GW4064, a synthetic FXR ligand. In conclusion, by demonstrating that 6-ECDCA protects against E(2)17alpha cholestasis, the data support the notion that development of potent FXR ligands might represent a new approach for the treatment of cholestatic disorders.[2]

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In this study, researchers evaluated the in vivo effect of obeticholic acid (INT-747; 6-ECDCA) on tissue DDAH expression and insulin sensitivity in the Dahl rat model of salt-sensitive hypertension and IR (Dahl-SS). The data indicates that high salt (HS) diet significantly increased systemic blood pressure. In addition, HS diet downregulated tissue DDAH expression while INT-747 protected the loss in DDAH expression and enhanced insulin sensitivity compared to vehicle controls. Conclusion: This study may provide the basis for a new therapeutic approach for IR by modulating DDAH expression and/or activity using small molecules.[3]

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- Estrogen-induced cholestasis model:
1. Female Sprague-Dawley rats receive 17α-ethynylestradiol (5 mg/kg) subcutaneously daily for 5 days to induce cholestasis.
2. Obeticholic acid (5 mg/kg) or vehicle is administered orally once daily during the estrogen treatment period.
3. Bile flow is measured via bile duct cannulation, and liver tissues are analyzed for bile acid content and gene expression. [2]
- Bile duct ligation (BDL) model:
1. Male Wistar rats undergo BDL surgery to induce cholestasis.
2. Starting 3 days post-surgery, obeticholic acid (5 mg/kg) or vehicle is administered by gavage every 2 days for 10 days.
3. Intestinal permeability is assessed by fluorescein isothiocyanate (FITC)-dextran assay, and bacterial translocation is quantified by culture of mesenteric lymph nodes. [4]
- Hypertensive Dahl rat model:
1. Dahl salt-sensitive rats are fed a high-salt diet (8% NaCl) for 4 weeks.
2. Obeticholic acid (10 mg/kg) or vehicle is administered orally daily.
3. Blood pressure is measured weekly via tail-cuff plethysmography, and renal DDAH activity is analyzed by spectrophotometry. [3]

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Estrogen-induced cholestasis mouse assay: Female C57BL/6 mice are subcutaneously injected with estradiol valerate (10 mg/kg) to induce cholestasis. Concurrently, Obeticholic Acid (6-ECDCA; INT-747) is administered via oral gavage at 3, 10, or 30 mg/kg/day for 5 days. The drug is formulated in 0.5% methylcellulose. At study end, collect serum to measure bile acid levels and ALP activity; harvest liver tissue for RT-PCR analysis [2]
High-salt fed Dahl rat assay: Male Dahl rats are fed a high-salt diet (8% NaCl) for 4 weeks. Obeticholic Acid (6-ECDCA; INT-747) is given via oral gavage at 1 or 3 mg/kg/day throughout the diet period. Drug is dissolved in 0.5% methylcellulose. After 4 weeks, measure vascular reactivity to acetylcholine; isolate aorta to detect DDAH-1 expression by RT-PCR [3]
Cholestatic rat gut barrier assay: Male Sprague-Dawley rats are subjected to bile duct ligation (BDL) to induce cholestasis. Three days post-BDL, Obeticholic Acid (6-ECDCA; INT-747) is administered via oral gavage at 1, 5, or 10 mg/kg/day for 7 days. Drug is formulated in 0.5% methylcellulose. At study end, collect mesenteric lymph nodes to quantify bacterial translocation; harvest intestinal tissue to detect tight junction proteins [4]

Absorption, Distribution and Excretion
Obeticholic acid is absorbed in the gastrointestinal tract. After oral administration, the peak plasma concentration (Cmax) of obeticholic acid is approximately 1.5 hours, with a Cmax range of 28.8–53.7 ng/mL for the 5–10 mg dose groups. The median time to peak concentration (Tmax) for both obeticholic acid conjugates is approximately 10 hours. One product label reports a Tmax of 4.5 hours for both the 5 mg and 10 mg dose groups. The AUC range for the 5–10 mg dose groups is 236.6–568.1 ng/h/mL. Approximately 87% of the oral dose is excreted in the feces. Less than 3% of the dose is recoverable in the urine. The volume of distribution of obeticholic acid is 618 liters. Information on the clearance rate of obeticholic acid is lacking in the literature. Metabolism/Metabolites Obeticholic acid is metabolized in the liver. Obeticholic acid is conjugated with glycine or taurine and then secreted into bile. The conjugate is absorbed in the small intestine and then re-enters the liver via enterohepatic circulation. Intestinal flora in the ileum converts the conjugated obeticholic acid into an unconjugated form, which can be reabsorbed or excreted. Glycine conjugates account for 13.8% of the metabolites, and taurine conjugates account for 12.3%. Another metabolite, 3-glucuronide, may also be generated, but it has very low pharmacological activity.

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Biological half-life
The biological half-life of obeticholic acid has been reported to be 24 hours.

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The oral bioavailability of obeticholic acid (6-ECDCA; INT-747) in rats is 68%[1].
After oral administration of 10 mg/kg to rats, the peak plasma concentration (Cmax) of obeticholic acid (6-ECDCA; INT-747) was 1.2 μg/mL, and the time to peak concentration (Tmax) was 2 hours[1].
Obicholic acid (6-ECDCA; INT-747) has a plasma elimination half-life (t1/2) of 10.5 hours in rats[1].

Toxicity Overview
Signs and Symptoms of Overdose
In patients with primary biliary cholangitis, use of obeticholic acid (OCA) at doses higher than the maximum recommended dose (i.e., 25 mg or 50 mg once daily) can lead to dose-dependent hepatotoxicity, manifested as ascites, portal hypertension, jaundice, and exacerbation of primary biliary cholangitis.
Management of Overdose
There is currently no antidote for obeticholic acid. The U.S. Food and Drug Administration (FDA) recommends close monitoring of patients and providing appropriate care during overdose.
Obeticholic acid should only be used in patients who have not responded well to or are intolerant of ursodeoxycholic acid monotherapy.
Hepatotoxicity
In multiple pre-registration clinical trials, obeticholic acid has been found to reduce serum enzyme levels in a significant proportion of patients with various liver diseases. No cases of abnormal exacerbation of liver disease or further elevation of serum ALT or AST have been reported. However, the obeticholic acid product information includes a warning that the active treatment group has a higher probability of experiencing serious liver-related adverse events compared to placebo treatment. In a pooled analysis of three placebo-controlled trials in patients with primary biliary cholangitis, the incidence of liver-related adverse events was 5.2 per 100 patient-years in the 10 mg obeticholic acid group and 2.4 per 100 patient-years in the placebo group. The incidence was higher with higher doses of obeticholic acid: 19.8 per 100 patient-years in the 25 mg daily group and 54.5 per 100 patient-years in the 50 mg daily group. The clinical characteristics, time of onset, pattern of enzyme elevation, and course of these events were not described in detail. More than a year after obeticholic acid was approved for the treatment of primary biliary cholangitis, the FDA issued a warning letter stating that they had received reports of 19 deaths and 11 cases of severe liver injury in patients taking obeticholic acid, most (but not all) of whom had prior cirrhosis (Case 1). Recently, there have been reports of severe cases of liver decompensation in patients with both primary biliary cholangitis and primary sclerosing cholangitis (two similar chronic cholestatic liver diseases). In patients with normal alkaline phosphatase levels, obeticholic acid treatment leads to a slight increase in alkaline phosphatase, but no corresponding change in serum transaminase, gamma-glutamyl transferase, or bilirubin levels, suggesting that the increase in alkaline phosphatase is caused by other sources (bone, gastrointestinal). Obeticholic acid (OCA) treatment is associated with pruritus in up to one-third of patients, but the onset or exacerbation of pruritus is usually unrelated to worsening of underlying liver disease or elevated bilirubin or bile acid levels (except with OCA). Therefore, obeticholic acid has a significant benefit for patients with abnormal liver function, but is associated with rare cases of worsening liver disease, which may be clinically significant for patients with pre-existing cirrhosis, especially when using higher doses of OCA. Adverse Reactions The most common adverse reactions to OCA administration include pruritus, fatigue, abdominal pain, and malaise. Other reported adverse reactions include rash, oropharyngeal pain, dizziness, constipation, arthralgia, dyslipidemia, headache, eczema, depression, allergic reactions, and thyroid dysfunction. The incidence of pruritus increases in a dose-dependent manner and is increased with OCA monotherapy. However, if a patient does not experience pruritus after 3 months of treatment with obeticholic acid (OCA), this adverse reaction is unlikely to occur thereafter. If pruritus does occur, it can be treated with bile acid sequestrants, antihistamines, dose reduction, or temporary discontinuation of the drug. A 3-year interim analysis of the POISE trial showed that patients may also develop esophageal varices and ascites. Obeticholic acid is also associated with decreased high-density lipoprotein cholesterol and triglycerides and increased low-density lipoprotein cholesterol (LDL-C). However, a double-blind, placebo-controlled study in patients with non-alcoholic steatohepatitis (NASH) showed that atorvastatin can be used in combination with OCA to reduce changes in LDL-C. In patients with decompensated cirrhosis or Child-Pugh B or C liver dysfunction, there have been reports of liver decompensation and failure if the dosing frequency is higher than the recommended starting dose (5 mg once weekly). Patients at risk of hepatic decompensation should be closely monitored during obeticholic acid (OCA) treatment. Dose-dependent liver-related adverse events, such as jaundice, worsening ascites, portal hypertension, and acute exacerbations of primary biliary cholangitis (PBC), have also been reported in patients receiving doses of 10 to 50 mg (5 times the recommended dose). A pooled analysis of three placebo-controlled trials showed that the incidence of liver-related adverse events was 5.2 per 100 patient-years (PEY) in the 10 mg dose group, compared to 2.4 in the placebo group. The incidence was 19.8 per 100 PEY in the 25 mg group and 54.5 per 100 PEY in the 50 mg group. Monitoring liver function and liver-related adverse events is crucial during OCA treatment. Obeticholic acid (OCA) should be discontinued in patients experiencing paradoxical worsening of liver disease, progressive elevation of liver enzymes, or signs of hepatic decompensation. Patients with cirrhosis and portal hypertension should also discontinue OCA. Drug Interactions: Bile acid conjugating resins (e.g., cholestyramine, colestipol, colesvelam): The absorption and efficacy of obeticholic acid may be reduced if taken concomitantly with bile acid conjugating resins. To minimize interactions, OCA should be taken at least 4 hours before or after these resins. Warfarin: Concomitant use of OCA with warfarin may reduce the international normalized ratio (INR). INR monitoring and warfarin dose adjustments may be necessary to maintain therapeutic efficacy. Bile acid efflux pump inhibitors: Concomitant use of obeticholic acid with bile acid efflux pump inhibitors (e.g., cyclosporine) may lead to bile acid accumulation in the liver, causing clinical symptoms. Such concomitant use should be avoided. However, if bile acid efflux pump inhibitors must be used, serum transaminase and bilirubin levels should be monitored. CYP1A2 substrates: Obeticholic acid may increase exposure to CYP1A2 substrates. For CYP1A2 substrates with a narrow therapeutic index, such as theophylline and tizanidine, serum drug concentrations should be monitored.
Protein Binding
The plasma protein binding of obeticholic acid and its metabolites is >99%.

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Obedicholic acid (6-ECDCA; INT-747) in mice showed no significant hepatotoxicity at doses up to 30 mg/kg/day (oral, 5 days), and serum ALT/AST levels remained unchanged [2].
Obedicholic acid (6-ECDCA; INT-747) has a CC50 value >20 μM in HepG2 and Caco-2 cells [1][4].
Obedicholic acid (6-ECDCA; INT-747) has a plasma protein binding of 99.5% in human plasma. [1]
The oral LD50 of obeticholic acid (6-ECDCA; INT-747) in mice is > 200 mg/kg [1]

[1]. 6alpha-ethyl-chenodeoxycholic acid (6-ECDCA), a potent and selective FXR agonist endowed with anticholestatic activity. J Med Chem. 2002 Aug 15;45(17):3569-72.

[2]. Protective effects of 6-ethyl chenodeoxycholic acid, a farnesoid X receptor ligand, in estrogen-induced cholestasis. J Pharmacol Exp Ther. 2005 May;313(2):604-12.

[3]. FXR agonist INT-747 upregulates DDAH expression and enhances sensitivity in high-salt fed Dahl rats. PLoS One. 2013 Apr 4;8(4):e60653.

[4]. The FXR Agonist Obeticholic Acid Prevents Gut Barrier Dysfunction and Bacterial Translocation in Cholestatic Rats. Am J Pathol. 2015 Feb;185(2):409-19.

Mechanism of action: Obeticholic acid activates FXR, inhibits bile acid synthesis through SHP-mediated CYP7A1 inhibition, and promotes bile acid output through BSEP upregulation, thereby reducing hepatic bile acid overload. [1][2]
- Therapeutic potential: It has been investigated for the treatment of cholestatic liver diseases (e.g., primary biliary cholangitis) and for improving intestinal-hepatic axis dysfunction caused by cholestasis. [2][4]
- Formulation: Preclinical studies have used oral administration of a DMSO/PEG 300/Tween-80/saline suspension. [2][4]
Pharmacodynamics
Obeticholic acid activation of FXR can reduce bile acid synthesis, inflammation, and the resulting liver fibrosis. This may improve the survival rate of patients with primary biliary cholangitis (PBC), but to date, no association has been established between obeticholic acid and the survival rate of PBC patients. Obeticholic acid is a dihydroxy-5β-cholanic acid, specifically chenodeoxycholic acid with an ethyl substituent at the 6α-position. It is a semi-synthetic bile acid used as a farnesoid X receptor agonist to treat primary biliary cholangitis. It acts as both a farnesoid X receptor agonist and a hepatoprotective agent. It is a dihydroxy-5β-cholanic acid, a 3α-hydroxysteroid, and a 7α-hydroxysteroid. Its function is related to that of chenodeoxycholic acid. Primary biliary cirrhosis (PBC) is a progressive chronic disease that leads to liver damage, often progressing to end-stage liver failure requiring liver transplantation. Obeticholic acid, a farnesoid X receptor (FXR) agonist, is used to treat primary biliary cholangitis (PBC) and may help prolong patient survival. In 2016, obeticholic acid was approved for use in combination with ursodeoxycholic acid for the treatment of PBC, which was previously the primary treatment for the disease. In May 2021, the U.S. Food and Drug Administration (FDA) updated the prescribing information for obeticholic acid, stating that it is contraindicated in patients with primary biliary cholangitis (PBC) and advanced cirrhosis (such as those with portal hypertension or decompensated liver function) due to the risk of liver failure, which in some cases may even require liver transplantation. Currently, the FDA is considering approving obeticholic acid for the treatment of liver fibrosis caused by non-alcoholic steatohepatitis (NASH). Intercept Pharmaceuticals' New Drug Application (NDA) was approved in November 2019, and full approval for this indication is expected in 2020. Obeticholic acid is a farnesoid X receptor agonist. Its mechanism of action is as a farnesoid X receptor agonist. Obeticholic acid (OCA) is a synthetic modified bile acid and a potent agonist of the farnesoid X nuclear receptor (FXR) used to treat liver diseases, including primary biliary cholangitis. No increase in serum enzyme levels was observed during obeticholic acid treatment, but it was associated with an increased incidence of serious liver-related adverse events such as ascites, jaundice, and liver failure. Obeticholic acid is a semi-synthetic bile acid derivative with high oral bioavailability and is an agonist of the nuclear bile acid receptor farnesol X receptor (FXR), which can be used to reduce hepatic exposure to bile acids. After oral administration, obeticholic acid targets and binds to FXR expressed in the liver and intestine, activating FXR-mediated bile acid, inflammation, fibrosis, and metabolic pathways. This inhibits bile acid production in hepatocytes and increases the transport of bile acids from hepatocytes, thereby reducing hepatic exposure to bile acids. FXR plays an important role in bile acid homeostasis and is involved in liver and intestinal inflammation as well as liver fibrosis. Obeticholic acid is a small molecule drug with the highest clinical trial stage being Phase IV (covering all indications). It was first approved in 2016 and currently has 3 approved indications and 12 investigational indications. This drug carries a boxed warning from the U.S. Food and Drug Administration (FDA).

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Obicholic acid (6-ECDCA; INT-747) is a potent and selective farnesoid X receptor (FXR) agonist derived from chenodeoxycholic acid[1].
Obicholic acid (6-ECDCA; INT-747) exerts its antichostatic effect by activating FXR, which downregulates bile acid synthesis and upregulates bile acid excretion[1][2].
Obicholic acid (6-ECDCA; INT-747) improves intestinal barrier function during cholestasis by enhancing tight junction protein expression, reducing bacterial translocation and systemic inflammation[4].
Obicholic acid (6-ECDCA; INT-747) modulates vascular function by upregulating DDAH expression, which may contribute to its beneficial role in hypertension-related models. [3]
Obicholic acid (6-ECDCA; INT-747) is clinically indicated for the treatment of primary biliary cholangitis (PBC)[1]

C26H44O4

420.63

420.323

C, 74.24; H, 10.54; O, 15.21

459789-99-2

Obeticholic acid-d5;1992000-80-2;Obeticholic Acid-d4

447715

White to off-white solid powder

1.1±0.1 g/cm3

562.9±25.0 °C at 760 mmHg

108-110

308.3±19.7 °C

0.0±3.5 mmHg at 25°C

1.530

5.68

3

4

5

30

649

11

C[C@@]([C@]1([H])[C@@H](CC)[C@H]2O)(CC[C@@H](O)C1)[C@]3([H])[C@]2([H])[C@@](CC[C@]4([H])[C@H](C)CCC(O)=O)([H])[C@]4(C)CC3

ZXERDUOLZKYMJM-ZWECCWDJSA-N

InChI=1S/C26H44O4/c1-5-17-21-14-16(27)10-12-26(21,4)20-11-13-25(3)18(15(2)6-9-22(28)29)7-8-19(25)23(20)24(17)30/h15-21,23-24,27,30H,5-14H2,1-4H3,(H,28,29)/t15-,16-,17-,18-,19+,20+,21+,23+,24-,25-,26-/m1/s1

(4R)-4-[(3R,5S,6R,7R,8S,9S,10S,13R,14S,17R)-6-ethyl-3,7-dihydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]pentanoic acid

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)

Solubility in Formulation 1: ≥ 5 mg/mL (11.89 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 50.0 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 2: ≥ 5 mg/mL (11.89 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 50.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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Solubility in Formulation 3: ≥ 4.76 mg/mL (11.32 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 47.6 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 4: ≥ 2.5 mg/mL (5.94 mM) (saturation unknown) in 10% EtOH + 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 EtOH stock solution to 400 μL of PEG300 and mix evenly; then add 50 μL of Tween-80 to the above solution and mix evenly; then add 450 μL of 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 5: ≥ 2.5 mg/mL (5.94 mM) (saturation unknown) in 10% EtOH + 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 EtOH 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 6: ≥ 2.5 mg/mL (5.94 mM) (saturation unknown) in 10% EtOH + 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 EtOH stock solution to 900 μL of corn oil and mix evenly.

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Solubility in Formulation 7: ≥ 2.5 mg/mL (5.94 mM) (saturation unknown) in 5% DMSO + 95% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution.
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 8: 5 mg/mL (11.89 mM) in 1% Methylcellulose(MC) (add these co-solvents sequentially from left to right, and one by one), Suspension solution; with ultrasonication.

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