Biliary Atresia - Advances in Etiology

and Early Diagnosis

Ronald J. Sokol, MD, FAASLD

Pediatric GI, Hepatology, Nutrition

Digestive Health Institute

Children’s Hospital Colorado

University of Colorado School of Medicine

Disclosure

• Consultant for Retrophin, Shire, Alexion and Albireo

• Grant support from NCATS and NIDDK of NIH, Alpha

One Foundation, Cystic Fibrosis Foundation

Objectives

• Biliary atresia – epidemiology and

endotypes

• Etiology & Pathogenesis

• Early Diagnosis and Screening

Biliary Atresia

• Most frequent and important liver disease causing obstructive

jaundice in 1st 3 mo.

• 1:8,000 (Asia) to 1:18,000 live births (Europe)

– U.S. ~ 1:13,000 rural = urban

– 300-400 new cases per year in U.S., 8-12 in Israel

• More common in girls, Asians, African-Americans, preterm

infants

• >40% of all liver transplants in children

• ~ 200 per year in U.S.

Illustration courtesy of

Robert E. Kramer, MD

BILIARY ATRESIA

Inflammatory

Fibrosing

Obstruction

Extrahepatic BD

First 3 months

Intrahepatic BDs

Phenotypes of Biliary Atresia

1. BASM

Polysplenia or Asplenia -

laterality defects

(situs, malrotation, midline

liver, interrupted IVC)

• Frequency: 8-12%, 3-4% in

Asia

• Association: IDM in UK

• Morphogenesis defect

2. BA with Multiple

Malformations

• At least one other major

congenital malformation,

not BASM

• Frequency: 10%

• Morphogenesis defect

Schwarz et al. Hepatology 2013

Phenotypes of Biliary Atresia

3. Isolated BA

• No other congenital anomalies

• Frequency: 75-80%

• BD formed but then was damaged by insult pre- or

postnatal

• ? Viral, toxin, immune/autoimmune, vascular etiology

or genetic

• Outcome better than BASM

Phenotypes of Biliary Atresia

4. Cystic BA

• Choledochal cyst (usually

hilar) + BA

• Frequency: 5-10%

• Prenatal identification

• Outcome: ? Improved

• CDC may lead to delayed

IOC and HPE

5. CMV-associated BA

CMV + IgM Ab

• Frequency: ~10%

• At presentation: older, higher

bilirubin, AST and APRI and

↑inflammation and fibrosis

on liver bx

• Poorer outcome (normal bili

post HPE in 15% vs. 52%)

J Pediatr Surg 50 (2015) 1739–1745

CMV-associated BA

• CMV not detected in

liver

• Brindley et al. did

show specific T-cell

reactivity to CMV in

50% BA patients

Hepatology 2012;55:1130–8

• ? Any role for

ganciclovir treatment

Candidate Etiologies of Biliary Atresia

• Defective morphogenesis (embryonic)

• Laterality sequence genes, others

• PKD1L1 Gene Variants in BASM

• Infectious

• Immune or autoimmune

• Toxin - ? Biliatresone-like substance

• Vascular

New Paradigm for BA Pathogenesis

Bezerra et al., NIH Workshop 2017, Hepatology 2018

Jaundice

Acholic Stools

New Paradigm for BA Pathogenesis

Bezerra et al., NIH Workshop 2017, Hepatology 2018

New Paradigm for BA Pathogenesis

Bezerra et al., NIH Workshop 2017, Hepatology 2018

New Paradigm for BA Pathogenesis

Bezerra et al., NIH Workshop 2017, Hepatology 2018

Genes: PKD1L1 and Biliary Atresia

• WES on 58 BASM trios (pt, MOC, FOC) –

from NIDDK-supported ChiLDReN

• 2,017 genes - ciliary structure or function

and putative BA susceptibility loci

• 5 subjects – rare, potentially deleterious homozygous, compound

heterozygous, or heterozygous missense variants in PKD1L1

• PKD1L1 interacts with PKD2 - involved in left-right embryonic

patterning, is present and functions in cholangiocyte cilium

• PKD1L1 – candidate gene causing BASM

Berauer J-P et al. Hepatology 2019

PKD1L1 IHC Expression in Bile Ducts

A. BA with

PKD1L1

variants D. ALGS

BD Paucity

C. Control

Livers

E-G. Normal

Infant Liver

H and E,

CK7,

PKD1L1

PKD1L1 IHC Expression in Bile Ducts

A. BA with

PKD1L1

variants D. ALGS

BD Paucity

C. Control

Livers

E-G. Normal

Infant Liver

H and E,

CK7,

PKD1L1

PKD1L1 IHC Expression in Bile Ducts

A. BA with

PKD1L1

variants D. ALGS

BD Paucity

C. Control

Livers

E-G. Normal

Infant Liver

H and E,

CK7,

PKD1L1

Infectious Etiology of Biliary Atresia

• Main candidate Viruses

– Rotavirus

– Reovirus

– Cytomegalovirus (CMV)

• Mouse models – perinatal infection causes bile

duct injury and obstruction

• Human data – association of evidence of

increased infection rates with BA vs. controls

Courtesy of Cara Mack

Asai, A. et al. Nat. Rev. Gastroenterol. Hepatol. 12, 342–352 (2015)

Viral-Induced Pathogenesis of Biliary Atresia

Innate and Adaptive

Immunity Pathways

Toxin: Biliatresone – Toxic Cholangitis & BA

• Outbreaks of BA in lambs during droughts in Australia

• R. Wells team harvested plants in New South Wales during

2007-2008 drought

• Dysphania – sequential fractionation of the plants, guided by

a zebrafish bioassay

• Isolated a previously unknown toxin that causes selective

extrahepatic biliary tree atresia in zebrafish

Lorent K et al., Science Transl Med 2015;7:286ra67

Lorent K et al., Science Transl Med 2015;7:286ra67

Zebrafish Assay

Inflammatory vs. Fibrotic Endotypes

• Gene expression profiling of liver done on BA infants

• Identified two transcriptional profiles (endotypes)

correlating with liver biopsy findings: inflammatory vs.

fibrotic

• Different outcomes (SNL) for the two endotypes

• Could this guide treatment after Kasai HPE?

• For example, are there subtypes that might respond to anti-

inflammatory treatments (precision Rx)?

Moyer K, et al. Genome Medicine 2010; 2: 33

Transcriptional Endotypes of BA

Evaluation of Neonatal Cholestasis

Nat Rev

GI Hep

2019

Feldman

Sokol

Diagnostic Tests for Biliary Atresia

• Liver Blood tests

• Exclude other diseases

• Ultrasonography

• Scintigraphy

• Duodenal bile collection

• Liver biopsy

• Cholangiography

intraoperative, ERCP, transhepatic

cholecystography, MRCP

Diagnostic Tests for Biliary Atresia

• Liver Blood tests

• Exclude other diseases

• Ultrasonography

• Scintigraphy

• Duodenal bile collection

• Liver biopsy

• Cholangiography

intraoperative, ERCP, transhepatic

cholecystography, MRCP

A1AT

ALGS

CF

RRV

Mouse

ModelLiver - BA

patients

Liver Expression of MMP7 in BA

J Surg Res. 2010 April ; 159(2):

611–617

Modern Pathology (2005) 18,

941–950

Serum MMP7 Elevated in BA

AUC ROC = 0.96

J Pediatr 2019; 208: 30-37

MMP7 Clinical Test Available

Treatment & Outcomes of Biliary

Atresia

• Progressive liver injury with cirrhosis in 95% of adult biliary

atresia patients with native liver

Lykavieris et al., Hepatology, Vol 41, 2005

• Kasai portoenterostomy: removal of occluded

extrahepatic bile duct and Roux-en-Y jejunum

to porta hepatis

• Succesful bile drainage in 30-70% cases

• 70-80% of children with biliary atresia will

require liver transplant in childhood

< 31 days : 99 patients

31 to 60 days : 435 patients

61 to 90 days: 361 patients

> 90 days : 132 patients

1027 patients 1986-2009

P<0.0001

Age (years)

Age at Kasai operation:

0

,2

,4

,6

,8

1

0 5 10 15 20 25

J Hepatol 2013; 58(6):1209-17

Survival with Native Liver & Age at Kasai

Operation - France

Goal for Timing of BA Diagnosis

• Goal:

– Establish diagnosis of BA before 30-45 days of life

– Portoenterostomy prior to 30-45 days of life

• Education efforts over past 30 years have failed to

change average age at Dx

– 65-70 days in ChiLDReN research network (15 sites)

Wang KS, et al. Newborn Screening for Biliary Atresia. Pediatrics 2015:136:e1663

No Change in Average Age at Kasai France 743 BA patients – 1986-2002

Serdinet et al, Pediatrics 2009;123:1280

We need to move the needle!!!!

HOW?

Stool Color Card

Neonatal Conjugated Bilirubin Level

Acholic Stools

Age at HPE (Kasai)

74% have Kasai surgery before 60 days of age

Better Survival with Native Liver

Less need for Liver transplantation

Would Stool Color Card Screening Work

in Israel?

• BA is 3x more prevalent in Taiwan – knowledge greater

• A 1-month WCC visit (for the HBV vaccine) is required in

Taiwan, not in U.S.

• What would be adherence?

• Would it be cost-effective? YES

• Needs to be piloted

• PoopMD App may help standardize “acholic” Mogul DB, PLoS One.

2015 Jul 29;10(7):e0132270

How about using Serum

Conjugated Bilirubin as a

Newborn Screen?

Newborn Conjugated Bilirubin

as Screen for BA

• Texas Children’s Hospital 2007-2010

• Serum bili (total & direct/conjug) measured on every newborn in some

hospitals in Houston, TX

• 73 BA pts. & 75 normal newborn controls (300 values)

• Look back at serum bilirubin levels and % conjugated or direct

in 1st 96 hours

• 34 of 73 BA patients had bilirubin levels available (27 direct and

7 conjugated)

Harpavat S et al. Pediatrics 2011; e1428

Total and Direct Bilirubin Levels in 1st 48

hours of Life

J Pediatr 2011

Direct Bilirubin 1st 96 hours in BA

J Pediatr 2011

20%

BA Patients

DB/TB Ratio at 24-48 hrs.

J Pediatr 2011

20%

79% BA

< 20%

BA Patients

DB/TB Ratio at 24-48 hrs.

J Pediatr 2011

20%

> 99% Controls

< 10% DB/TB

10%

BA Patients

Questions Raised

• Why were so many BA infants with elevated Direct Bilirubin

as newborn not f/u’d?

– perhaps the DB:TB felt to be normal

• Are we using the wrong cholestasis cut-off for DB/TB ratio in

first 4 days of life?

– Should it be Direct > 10% of Total Bilirubin (not 20%)

• Should all infants get direct as well as total bilirubin?

– “Newborn” screening for cholestasis

?

Take Home Points

• Etiology of Biliary Atresia still not clear, however

genomics technology will lead to discovery of new

genetic causes for some cases

• Etiology of bulk of BA cases (isolated) still not clear

• Diagnosis needs to be made before 30-45 days for best

outcomes from Kasai HPE

• Screening will be only effective way to earlier diagnosis

Thank You