Rotor syndrome (RS, OMIM*237450) is a rare and benign genetic disease characterized by low-grade, chronic or fluctuating, predominantly conjugated hyperbilirubinemia. It has no other features of hepatobiliary disorder (1, 2). The prevalence of RS is unknown but is very low (<1:1,000,000) (3). First described by Rotor and Florentin (4) in 1948, it is inherited in an autosomal recessive digenic manner. Biallelic pathogenic mutations in solute carrier organic anion transporter family member 1B1 (SLCO1B1) and SLCO1B3 genes cause RS. Organic anion transporting polypeptide 1B1 (OATP1B1) and OATP1B3 are encoded by SLCO1B1 and SLCO1B3 genes, respectively. They serve as transporters for hepatic uptake of conjugated bilirubin. Inactivation of both proteins together leads to RS, which does not affect life expectancy and usually requires no treatment (1). To date, 51 mutations in SLCO1B1 and 30 variants in SLCO1B3 have been described in the Human Gene Mutation Database (HGMD; http://www.hgmd.cf.ac.uk/ac/index.php). In the present study, we reported three patients with RS and tested two disease-causing mutations: SLCO1B1 (NM_006446.5): c.1738C>T (p.R580*) and SLCO1B3 (NM_019844.4): c.481+22insLINE.

This study included two male and one female patient, aged from 14 to 20 years. They were born to nonconsanguineous parents (Figures 1A–C) and presented with mild intermittent jaundice. The liver function test only showed predominantly conjugated hyperbilirubinemia. No abnormalities were observed in viral serologies (HBV and HCV), hemolysis test, coagulation function, autoimmune liver disease-associated antibodies, immunoglobulin G, serum ceruloplasmin testing, or abdominal ultrasound examination. The major clinical manifestations of the three patients with Rotor syndrome are summarized in Table 1. The WES indicated that all patients harbored the same homozygous c.1738C>T mutation in SLCO1B1 (Figure 1D) and c.481+22insLINE variant in SLCO1B3.

OATP1B1/3 are expressed in the hepatocyte basolateral membrane, which are also called SLCO1B1/3. They uptake endogenous substances, such as conjugated bilirubin, bile acids (BAs), eicosanoids, prostaglandins, and hormones. Unconjugated bilirubin (UCB) enters hepatocytes through passive diffusion and/or transporters, which may include OATP1B1/3. Uridine-diphospho glucuronosyl transferase 1A1 (UGT1A1) catalyzes the conversion of UCB to bilirubin glucuronides (BG) in the endoplasmic reticulum. BG is secreted into bile by ABCC2 and ABCG2. A substantial fraction of BG is rerouted by ABCC3 to the blood. It can be taken up by downstream hepatocytes via OATP1B1/3 transporters (2). In RS, the absence or dysfunction of the OATP1B1/3 may disrupt the uptake of BG (Figure 2), which causes predominantly conjugated hyperbilirubinemia.

The accurate diagnosis of RS is of paramount clinical importance, as it directly influences pharmaceutical safety by preventing unwarranted exposure to drugs that rely on functional OATP1B1/3 transporters for hepatic uptake and systemic clearance. The dysfunction of OATP1B1/3 transporters can profoundly impact numerous drug metabolisms, particularly statins, ezetimibe, methotrexate, irinotecan, cabazitaxel, sunitinib, and sartans (3, 6–8). They could exhibit significantly increased systemic exposure in RS patients. After a confirmed diagnosis of RS, clinicians can proactively select alternative drugs with minimal OATP dependency and implement therapeutic drug monitoring (TDM) for high-risk agents.

The c.1738C>T is a nonsense variant in SLCO1B1, which was very strong evidence of pathogenicity (PVS1). The mutation is located in a mutational hotspot (PM1). It has been reported in multiple clinical cases of RS (6, 9–12). Prediction software, specifically MutationTaster (13), predicted a deleterious effect of the variant (PP3). Therefore, according to ACMG guidelines, the variant is classified as a pathogenic mutation.

The c.481+22insLINE mutation in SLCO1B3 is a long-interspersed element (LINE) insertion variant about 6.1 kb in size. It could affect normal editing of mRNA and cause abnormal skipping of exons (PS3) (10). It is a common mutation of RS in Asian populations (PM1) (10). The variant is not found in either the 1000G or EXAC databases (PM2). The patients carried both the homozygous c.1738C>T variant in SLCO1B1 and the c.481+22insLINE mutation in SLCO1B3. The findings were consistent with the digenic recessive pattern of RS (PM3). Therefore, the variants were also classified as a pathogenic mutation.

This study provided evidence that the detected genetic mutations in SLCO1B1 and SLCO1B3 are common in Rotor syndrome, which is consistent with previous research (6, 9–12, 14). However, precise information on the frequencies of c.1738C>T (SLCO1B1) and c.481+22insLINE (SLCO1B3) mutations is limited. There is insufficient data to assess its distribution in RS and the general population. In resource-limited settings across East Asia, targeted PCR assays for the two mutations might be considered as a screening method for RS; however, validation in larger sample sizes is required.

In conclusion, RS is a rare inherited disorder that causes predominantly conjugated hyperbilirubinemia. Genetic testing is a useful tool for efficient diagnosis. This study supported that c.1738C>T in SLCO1B1 and c.481+22insLINE in SLCO1B3 are common pathogenic mutations in the East Asian RS patients.