SLCO1B1 c.388A > G variant incidence and the severity of hyperbilirubinemia in Indonesian neonates

Neonatal hyperbilirubinemia (jaundice) occurs in about 60% of all term infants [1]. In general, most of the increase in bilirubin levels is physiological and usually will improve by the end of the first week of life. An increase in total serum bilirubin level above 5 mg/dL can cause jaundice that is visible to observers [2]. This condition can be severe if the infant was born prematurely, where the incidence increases to 80% [2]. In these infants, hyperbilirubinemia can develop into severe neonatal jaundice (SNNJ) that can cause bilirubin-induced neurological damage (BIND) and ultimately lead to irreversible neurodevelopmental impairment or even death [3].

Aside from prematurity, genetic polymorphisms have been reported as an important factor in the development of SNNJ [4‐6]. The most commonly-studied gene is the uridine diphosphate glucuronosyltransferase 1A1 (UGT1A1) that is closely associated with Crigler-Najjar’s syndrome type I and II, and Gilberts syndrome. These conditions are frequently found in Caucasian and African populations [7, 8]. Another gene that has been gaining attention, one that is mostly found in Asian populations, is OATP2/SLCO1B1 [9]. SLCO1B1 (solute carrier organic anion transporter family member 1b1) encodes the solute carrier organic anion transporter family member 1B1 and is responsible for the absorption of unconjugated and conjugated bilirubin in the liver. This gene, located in chromosome 12p12, consists of 15 exons (1 coding exon and 1 non-coding exon) and 14 introns, as well as containing 2073 nucleotides in the gene-coding area [9]. Neonates carrying this gene polymorphism are associated with reduced bilirubin clearance which induces hyperbilirubinemia [10]. Rotor Syndrome, an autosomal recessive disease associated with elevated conjugated bilirubin, is an example of a disease frequently associated with mutations in SLCO1B1 and SLCO1B3. The most frequent polymorphisms found in this gene are SLCO1B1*1B (388A > G), SLCO1B1*4 (463C > A), and SLCO1B1*5 (521 T > C), and the most commonly associated with hyperbilirubinemia is the SLCO1B1*1B (388A > G) [9, 11‐13].

Based on previous studies, there is an association between SLCO1B1 with racial variety in regard to the severity of unconjugated neonatal hyperbilirubinemia. In Indian, Japanese, Taiwanese, and Chinese neonates, the incidence of polymorphism is higher in infants with higher bilirubin levels compared to those with lower bilirubin levels [9, 11, 12, 14]. However, in studies of Malaysian and Thai hyperbilirubinemic neonates, this SNP was not frequently found [15, 16]. In Indonesia, polymorphism studies on neonatal hyperbilirubinemia has yet to be conducted extensively. Therefore, this study provides valuable insight into the association of genetic factors especially SLCO1B1 with the severity of neonatal hyperbilirubinemia in Indonesia.

Physiological conditions such as the short neonatal red blood cell’s lifespan, absorption of bilirubin by the liver and a non-optimal conjugation process of bilirubin can cause hyperbilirubinemia [1, 2]. Premature birth carries a higher risk of severe hyperbilirubinemia because of the immaturity of the red blood cells and an impaired hepatic cell function responsible for neonatal the bilirubin metabolism in neonates [2, 20]. The cause for this condition also varies between populations due to suspected contributions of genetic factors towards the pathogenesis of hyperbilirubinemia.

SLCO1B1 is a sinusoidal membrane protein expressed at the basolateral membrane of hepatocytes and is putatively involved in facilitating the hepatic uptake of unconjugated bilirubin [9, 13]. The exon 5 mutation of 388G > A is a nonsynonymous SNP with a missense mutation that causes a single amino acid substitution (asparagine to aspartic acid) encoded by codons 130 of SLCO1B1. This change disrupts the function of the SLCO1B1 transporter in transporting bilirubin from blood to the liver, leading to bilirubin concentration remaining high in the blood, which, in turn, causes hyperbilirubinemia [12].

Aside from the well-established risk factors of low gestational age and low birth weight, we found a significant association between the severity of hyperbilirubinemia and difference of populations across Jakarta, Bengkulu, and Papua, as well as with delivery location.

SLCO1B1*1B in both the heterozygous and homozygous variants was predominantly found (94.32%) in the neonates that we studied. Compared with the study of UGT1A1*60 and UGT1A1*6 from our previous study, SCLO1B1*1B was similar to *60 which showed a high incidence albeit with no significant association with the higher bilirubin level (severity) in neonatal hyperbilirubinemia [19]. Other studies on SCLO1B1 in different countries also showed similar findings to our study concerning the incidence of polymorphisms. In a study in India, there were 77.7% hyperbilirubinemic neonates having the variant *1B in hyperbilirubinemic neonates, and 87.7% in Malaysia. All these studies demonstrate that more than half of the studied population has the *1B variant [14, 16, 21]. Both Asian countries share a lot in common in terms of genetic makeup with Indonesia.

Regarding the association between SLCO1B1*1B and higher bilirubin level in infants, there are still inconclusive evidence from different studies of different populations. In a study by Huang et al. in 2004, they reported the *1B SNP to be more frequent among hyperbilirubinemic infants with an OR of 2.01. In the study by Liu et al., the Chinese variant of *1B was also found to be associated with hyperbilirubinemia in the Guangdong population, but not in the Yunnan population [9]. Whereas other studies in Fujian and Taiwan also failed to prove the association of *1B with neonatal hyperbilirubinemia, one study in Taiwan showed that Taiwanese neonates with the minor allele of *1B were at high risk to develop severe hyperbilirubinemia [22‐24].

Some ethnicities are shown to be more at risk for severe hyperbilirubinemia than others such as African-American and Asian [25]. This is most likely caused by the different prevalence of genetic polymorphisms responsible for metabolism or transport pathways [26]. Variations of specific genes can also differ between the individuals in a smaller population group. An example of this is the intra-ethnic differences of UGT1A1 polymorphism in the Chinese populations of Han, She, and Dong origin [27]. Global analysis of SLCO1B1 also suggests that SLCO1B1 diversity is greater within populations than between populations [28]. Conversely, we did not find any intra-ethnic differences in our three populations for SLCO1B1 polymorphism.

From the comparison of genotypic distribution and clinical factor among the three different centers comprising population of different sub-ethnic origins, we observed a significant association with the severity of hyperbilirubinemia independent from the history of exclusive breastfeeding and other established clinical risk factors. This suggests the presence of a potentially undiscovered important risk factor for the development severe hyperbilirubinemia in the Indonesian population. Further research effort in Indonesia is warranted to investigate other potential genetic factors in different pathways leading to hyperbilirubinemia, including BLVRA and HMOX1 [29‐32].

SLCO1B1*1B was found to be a common occurrence in Indonesian hyperbilirubinemic neonates. There was no statistically significant differences between occurrence of the SLCO1B1*1B variant and severity of neonatal hyperbilirubinemia. There was, however, a significant difference in the severity depending on the center where the neonate was enrolled. Future studies are required that includes a more diverse population to represent the entirety of the ethnically-diverse Indonesian population and exploring additional genetic risk factors is highly recommended.