Recent developments in diagnostics and treatment of neonatal cholestasis

Abstract

Neonatal cholestasis is characterized by conjugated hyperbilirubinemia in the newborn and young infant and is a sign common to over 100 hepatobiliary and/or metabolic disorders. A timely evaluation for its etiology is critical in order to quickly identify treatable causes such as biliary atresia, many of which benefit from early therapy. An expanding group of molecularly defined disorders involving bile formation, canalicular transporters, tight junction proteins and inborn errors of metabolism are being continuously discovered because of advances in genetic testing and bioinformatics. The advent of next generation sequencing has transformed our ability to test for multiple genes and whole exome or whole genome sequencing within days to weeks, enabling rapid and affordable molecular diagnosis for disorders that cannot be directly diagnosed from standard blood tests or liver biopsy. Thus, our diagnostic algorithms for neonatal cholestasis are undergoing transformation, moving genetic sequencing to earlier in the evaluation pathway once biliary atresia, "red flag" disorders and treatable disorders are excluded. Current therapies focus on promoting bile flow, reducing pruritus, ensuring optimal nutrition, and monitoring for complications, without addressing the underlying cause of cholestasis in most instances. Our improved understanding of bile formation and the enterohepatic circulation of bile acids has led to emerging therapies for cholestasis which require appropriate pediatric clinical trials. Despite these advances, the cause and optimal therapy for biliary atresia remain elusive. The goals of this review are to outline the etiologies, diagnostic pathways and current and emerging management strategies for neonatal cholestasis.

Keywords: Alagille syndrome; Biliary atresia; Genomics; Neonatal cholestasis; Progressive familial intrahepatic cholestasis.

Figure 1.

Molecular regulation of the enterohepatic circulation of bile acids.In hepatocytes, bile acids are taken up by NTCP, OSTα/β or OATP1B1–1B3 on the basolateral membrane or are synthesized from cholesterol by CYP7A1. The ATP-dependent BSEP transports bile acids into bile across the canalicular membrane. FXR, a nuclear hormone receptor, is a master switch that upregulates mRNA expression of BSEP and downregulates NTCP and CYP7A1 (through SHP). MRP2 (conjugated bilirubin transporter), MDR3 (phospholipid transporter) and FIC1 (phosphotidylserine flippase) participate in bile formation across the canalicular membrane. MRP3 and MRP4 transport bile acids from the hepatocyte to the sinusoids. Tight junction proteins (TJP2 and CLDN1) maintain the boundary between canalicular bile and the hepatocyte, preventing the toxicitiy of secreted bile acids. In ileal enterocytes, ASBT promotes absorption of luminal bile acids across the enterocyte brush border which are transported by OSTα/β or MRP3 into portal blood to return to the liver where they interact with FXR to suppress further bile acid synthesis. Within the enterocyte, absorbed bile acids may also activate FXR which upregulates synthesis and secretion of FGF19 into the portal circulation. When it reaches the liver FGF19 binds to the FGFR4/ βKL receptor on the hepatocyte basolateral membrane and triggers suppression of CYP7A1. In cholangiocytes, ASBT in the apical membrane may transport bile acids from bile back into the cell and subsequently into the portal circulation by OSTα/β or MRP3 in the basolateral membrane. CTFR actively secretes chloride into bile. Perturbations of bile formation can result from genetic variants in many of these proteins (FIC1, BSEP, MDR3, MRP2, CFTR, TJP2, CLDN1) and from adaptive changes in transporter expression and function in response to inflammatory, obstructive, or drug-induced insults. . BA = bile acid, PC = phosphatidylcholine, Bili = conjugated bilirubin, Cl = chloride, PS= phosphatidylserine.

Figure 2.

Current paradigm for evaluation of neonatal cholestasis.In the current sequential evaluation of neonatal cholestasis, alpha-1 antitrypsin deficiency (A1AT), biliary atresia, and treatable causes of cholestasis (such as choledochal cyst or urinary tract infection) are excluded in a timely fashion, which may require liver biopsy and intraoperative cholangiogram to determine if biliary atresia is present. If a “red flag” points to a specific diagnosis (Table 2), evaluation for that disease should proceed promptly.If no diagnosis if found, the next tier of testing may include multiple blood and urine tests to evaluate for infectious, genetic and metabolic causes. If no diagnosis is confirmed, targeted gene panels or specific gene testing is performed and whole exome sequencing (WES) is reserved for those cases without a diagnosis after this exhaustive evaluation. R/O= rule out, HPE= hepatoportoenterostomy, UTI= urinary tract infection, PFIC= progressive familial intrahepatic cholestasis, TGP= targeted gene panel, WES= whole exome sequencing.

Figure 3.

Future evaluation of neonatal cholestasis. In the near future, it is anticipated that following exclusion of biliary atresia and A1AT deficiency, investigation of “red flags” pointing to specific etiologies, and testing for treatable conditions, rapid turn-around genotyping via whole exome sequencing (WES) or whole genome sequencing (WGS) will be utilized to evaluate for common and rare genetic causes. R/O= rule out, HPE= hepatoportoenterostomy, UTI= urinary tract infection, PFIC= progressive familial intrahepatic cholestasis, TGP= targeted gene panel, WES= whole exome sequencing, WGS= whole genome sequencing.

Figure 4.

Holistic Management of Neonatal Cholestasis. Different stages of neonatal cholestasis are defined by the rate of progression from early (neonatal) to chronic (> 6 months of cholestasis) to end-stage liver disease. The latter is characterized by progressive portal fibrosis and synthetic liver failure or complications of portal hypertension. Diagnostic testing may be continued as new etiologies are discovered over time. Disease-specific therapy (if available) should be instituted. Medical therapies can be utilized to improve or treat pruritus, cholangitis and portal hypertension. In the future, new choleretic, anti-fibrotic, anti-inflammatory and bile acid-modifying agents might become available. Medium-chain triglyceride containing infant formula and fat-soluble vitamin supplementation are essential for most infants who remain cholestatic. Monitoring and therapy are initiated for complications of portal hypertension; screening for hepatocellular carcinoma is initiated as indicated. Immunization schedules are accelerated if liver transplantation is planned. Developmental services and family support are provided as needed. MCT= medium chain triglycerides, NG = nasogastric, PN = parenteral nutrition, HCC = hepatocellular carcinoma, HPE = hepatoportoenterostomy, NTBC = 2-(2-nitro-4-trifluoromethylbenzoyl)-1,3-cyclohexanedione

Figure 5.

Emerging therapeutic targets for the treatment of cholestasis. Inhibitors of NTCP may reduce the bile acid uptake by hepatocytes and subsequent toxicity. FXR agonists may upregulate hepatocyte efflux of bile constituents through BSEP, MRP2, MRP3, MRP4, and OSTα/β. UCDA, molecular chaperones such as 4-phenylbutyrate or other agents may reduce the toxic bile acid burden of the hepatocyte and improve bile flow and fat absorption. PPARα agonists may induce canalicular MDR3 expression and phospholipid secretion protecting cholangiocytes against bile acid toxicity. Alternatively, another strategy that would reduce hepatocyte bile acid levels is the inhibition of bile acid synthesis through suppressing CYP7A1 by FGF19 agonists, FXR agonists, or short interfering RNAs. In the ileal enterocyte, inhibiting ASBT will increase fecal excretion of bile acids, lower the bile acid pool size, change bile acid composition and alter enterocyte FXR signaling. In the cholangiocyte, nor-UCDA may protect the cholangiocyte from bile acid-induced injury by altering bile pH. 4-PB = 4-phenylbutyrate, BA = bile acid, PC = phosphatidylcholine, Bili = conjugated bilirubin, Cl = chloride, PS= phosphotidylserine, UDCA = ursodeoxycholic acid.