NR1H4

Detailed Information

The NR1H4 gene, also known as Farnesoid X Receptor(FXR), is located on human chromosome 12q23.1 and comprises 11 exons. It encodes a nuclear receptor transcription factor composed of 486 amino acids. The protein contains a conserved DNA-binding domain (DBD)(amino acids 66–140) and a ligand-binding domain (LBD) (amino acids 249–472). The DBD includes two zinc finger motifs that recognize the specific DNA sequence known as the IR-1 response element (5'-AGGTCA N TGACCT-3'), while the LBD forms a hydrophobic pocket that undergoes conformational change upon binding ligands such as bile acids, leading to corepressor release and coactivator recruitment. There are four major splice variants of NR1H4: FXRα1 and FXRα2 contain the complete AF-2 activation domain, whereas FXRα3 and FXRα4 lack portions of the LBD and function as dominant-negative regulators.

NR1H4 exhibits ligand-specific activation. Among primary bile acids, chenodeoxycholic acid (CDCA) is the most potent natural agonist (EC50 ≈ 10 μM), followed by deoxycholic acid (DCA) and cholic acid (CA). In contrast, lithocholic acid (LCA), a secondary bile acid, acts as an antagonist. The synthetic ligand obeticholic acid (OCA)is 100 times more potent than CDCA. Upon ligand binding, NR1H4 primarily forms heterodimers with retinoid X receptor alpha (RXRα)to bind IR-1 elements in target gene promoters. Additionally, monomeric NR1H4 can competitively bind DR1 elements to suppress genes such as APOA1.

Regulation of Bile Acid Homeostasis

Canonical Negative Feedback Pathway

NR1H4 is a master regulator of bile acid homeostasis. In hepatocytes, elevated bile acid concentrations activate NR1H4, which induces the expression of small heterodimer partner (SHP, encoded by NR0B2). SHP subsequently inhibits two key transcription factors—liver receptor homolog-1 (LRH-1) and liver X receptor alpha (LXRα)—thus repressing the transcription of the rate-limiting enzymes CYP7A1 and CYP8B1 involved in bile acid synthesis. Concurrently, NR1H4 directly upregulates bile acid transporters ABCB11 (BSEP) and ABCC2 (MRP2), promoting bile acid excretion into bile canaliculi. This dual negative feedback regulation helps maintain a stable bile acid pool.

Figure 1. The importance of FXR in the enterohepatic circulation of bile acids. (Jiang L, et al., 2021)

Species-Specific Differences

Functional differences in NR1H4 between humans and mice stem from variations in bile acid composition. Murine liver expresses Cyp2a12 and Cyp2c70, which convert hydrophobic bile acids into hydrophilic muricholic acids (MCAs), a detoxifying mechanism absent in humans. Researchers at Tokai University developed a Cyp2a12/Cyp2c70 double-knockout (CYPDKO) mouse model that mimics the human bile acid profile. Hepatocyte-specific Nr1h4 knockout using AAV8-SaCas9 in this model led to severe liver injury, with a 30% increase in liver-to-body weight ratio, significantly elevated serum ALT/AST levels, disrupted hepatic cord structure, hepatocyte swelling, and upregulation of inflammatory cytokines such as TNF-α and TGF-β. Mechanistic studies showed that Nr1h4 deficiency resulted in rebound expression of Cyp7a1/Cyp8b1, and reduced expression of transporters Abcb11 and Ntcp by 50%, causing intrahepatic accumulation of hydrophobic bile acids.

Disease Associations and Molecular Mechanisms

Cholestatic Diseases

Loss-of-function mutations in NR1H4 lead to progressive familial intrahepatic cholestasis type 5 (PFIC5), characterized by early-onset intractable pruritus, elevated serum bile acid levels, and rapidly progressive liver fibrosis. Mutations such as R149Q and M1T impair DNA binding or nuclear localization, downregulate SHP expression, and lead to uncontrolled CYP7A1 overexpression. Interestingly, traditional Nr1h4 knockout mice do not replicate this phenotype, but hepatocyte-specific knockout on a CYPDKO background accurately models PFIC5, highlighting the importance of humanized bile acid profiles in disease modeling.

Tumorigenesis and Cancer Progression

NR1H4 plays a dual role in gastrointestinal cancers. In chronic atrophic gastritis (CAG), NR1H4 was identified as a key differentially expressed gene, positively correlating with neutrophil infiltration (r = 0.68), possibly via regulation of the IL-8/CXCL1 chemokine pathway, contributing to inflammation-driven carcinogenesis. Bioinformatic screening identified natural compounds such as columbianadin and stigmasterol with high binding affinity to NR1H4 (docking scores < -8.5 kcal/mol), suggesting potential therapeutic relevance in CAG.

In pancreatic ductal adenocarcinoma (PDAC), NR1H4 exhibits a pro-tumorigenic role. Among 176 tissue samples, NR1H4 mRNA and protein positivity rates were 68.75% and 77.27%, respectively. Expression correlated significantly with clinical stage (P = 0.020) and tumor differentiation (P = 0.010), with higher expression in advanced (stage III/IV) and well-differentiated tumors. Survival analysis showed that high NR1H4 expression shortened median overall survival by 6.3 months (15.2 vs. 21.5 months, P < 0.05). Mechanistically, NR1H4 promotes SHP expression, suppresses p53transcriptional activity, and upregulates multidrug resistance protein 1 (MDR1), contributing to chemotherapy resistance.

FXR Agonists

Obeticholic acid (OCA) was the first NR1H4 agonist approved by the FDA (2016) for primary biliary cholangitis. It activates the NR1H4-SHP pathway to suppress bile acid synthesis and improve liver function. In the phase III POISE trial, OCA achieved an alkaline phosphatase (ALP) response (<1.67×ULN) in 47% of patients versus 10% in the placebo group (P < 0.001). However, 28% experienced dose-dependent pruritus and 10% had elevated LDL cholesterol. Non-bile acid FXR agonists such as Cilofexor aim to reduce these side effects.

In non-alcoholic steatohepatitis (NASH), combination therapy with Efruxifermin (an FGF21 analog) and NR1H4 agonists showed synergistic effects. NR1H4 activation upregulated hepatic β-klotho expression, enhancing FGF21 sensitivity. In a phase IIb trial (NCT04929483), the combination reduced liver fat by 52.3% (MRI-PDFF) and improved fibrosis in 41.7% of patients (NASH CRN score).

SHP-Targeted Therapy

In the CYPDKO/Nr1h4-KO mouse model, overexpression of SHP via rAAV2 significantly alleviated cholestatic liver injury: serum ALT decreased by 40% (P < 0.05), and intrahepatic bile acid accumulation was reduced. Mechanistically, SHP selectively suppressed Cyp8b1 without affecting Cyp7a1, reducing cholic acid synthesis and preserving bile acid hydrophilicity. The SHP agonist SHP626 (Volixibat) has entered phase II trials for PFIC.

FXR Antagonists

NR1H4 antagonists have shown promise in cancer treatment. The small molecule ZLY28 competitively binds the LBD (Kd = 0.38 μM), blocking NR1H4 transcriptional activity. In PDAC xenograft models, ZLY28 combined with gemcitabine reduced tumor volume by 62.7%, and significantly enhanced apoptosis by releasing SHP-mediated p53 suppression (3.1-fold increase in TUNEL⁺ cells).

Challenges and Future Directions

One major challenge in targeting NR1H4 is tissue-specific effects. Intestinal NR1H4 activation induces FGF19, which suppresses hepatic bile acid synthesis, whereas hepatic NR1H4 directly regulates transporter expression. This dichotomy complicates systemic therapy. Potential solutions include gut-restricted FXR agonists and liver-targeted delivery systems.

Another hurdle is nuclear receptor compensation. Chronic NR1H4 inhibition may upregulate NR1H2 (FXRβ) and PPARα, diminishing therapeutic efficacy. This has prompted the development of dual-target modulators—for example, INT-767, which activates both NR1H4 and TGR5 (GPBAR1), demonstrating anti-inflammatory synergy in primary sclerosing cholangitis models.

Future research will delve into NR1H4's role in the gut-liver axis. Gut microbiota-derived secondary bile acids, such as DCA, are partial NR1H4 agonists, and antibiotic-induced microbiome changes can modulate FXR signaling. Moreover, circadian rhythm influences NR1H4 activity; the BMAL1/CLOCK complex directly binds the NR1H4 promoter, underlying the diurnal pattern of bile acid synthesis.