Pediatr Blood Cancer 2010;54:627–628

BRIEF REPORTPersistent Jaundice in an Infant With Homozygous Beta Thalassemia

Due to Co-Inherited Crigler–Najjar Syndrome

Varun Aggarwal, MD,1 Anju Seth, MD,1* Sunita Sharma, MD,2 Satinder Aneja, MD,1

Pietro Sammarco, MD,3 and Carmelo Fabiano, PhD3

INTRODUCTION

Homozygotes for b-thalassemia show a marked variability in

serum unconjugated bilirubin levels. This variability may be related

either to the transfused red blood cell destruction rate, ineffective

erythropoeisis, hemolysis, or to bilirubin elimination capacity

which depends partially on the bilirubin glucuronidation activity.

Crigler-Najjar syndrome is a rare disorder of bilirubin metabolism

with two distinct forms: type 1 and type 2. This syndrome is caused

by the mutation in the gene by a defect in UDP glucuronosyl

transferase 1A1 (UGT1A1). Glucuronidation is essential for biliary

elimination of bilirubin, and UGT1A1 is the only physiologically

relevant isoform in bilirubin glucuronidation. We report a case of

b-thalassemia major coexisting with Crigler-Najjar syndrome

type 2.

CASE REPORT

A patient, 6-month-old male was the product of a full-term home

delivery to a 19-year-old primigravida. There was no history of

consanguinity in parents. He was noticed to have jaundice

on second day of life which had persisted till the day of presentation

to our Center. He had never received any treatment in the form of

phototherapy, exchange transfusion or medication. At presentation

the child weighed 6.2 kg and measured 65.5 cm in length. He was

pale and had jaundice. His development was normal for age.

The liver was enlarged 3 cm (span 11 cm) and spleen 4 cm below

the costal margin. There were no features like edema, ascites, or

systemic bleeds suggestive of chronic liver disease of the systemic

examination was normal.

At presentation, hemoglobin of the patient was 5.0 g/dl with a

corrected reticulocyte index of 0.5%. MCV, MCH, and MCHC were

78.9 fl, 23.9 pg, and 30.5 g/dl, respectively. The peripheral smear

showed predominantly microcytic hypochromic cells with moder-

ate aniso-poikilocytosis, few tear drop cells, elliptocytes, and

occasional polychromatophils. Few red cells showed Howell Jolly

bodies, Cabot rings, and basophilic stippling. Twenty-three

nucleated red cells were seen per hundred leucocytes. Hemoglobin

electrophoresis revealed HbF 94.7%, HbA2, 2.0%, and HbA only

0.2% confirming presence of b-thalassemia major in the patient.

Hemoglobin electrophoresis of parents revealed b-thalassemia

minor in both.

Total serum bilirubin in the child at presentation was 23.6 mg/dl;

conjugated fraction being 1.5 mg/dl. The serum alkaline phospha-

tase, alanine and aspartate transaminase levels were normal. Direct

and indirect Coomb’s tests were negative. Serum G6PD activity was

12.8 U/gm Hb, within the normal range of 4.6–13.50 U/gm Hb. The

thyroid function tests were normal. Serological tests for toxoplasma,

rubella, cytomegalovirus virus, herpes virus, syphilis, hepatitis A, B,

C, and E were non-reactive. The urine tested negative for reducing

substances. DNA sequencing analysis of the patient for UGT1A-1

gene mutations showed the presence of a homozygous base

substitution at position 362 (GGT>AGT) in exon 3. This mutation

resulted in replacement of glycine amino acid (at position 362 in

exon 3) by the amino acid serine. The diagnosis of Crigler–Najjar

syndrome was thus established. The promoter region of the gene

carried the normal (TA)6/(TA)6 configuration, thus, ruling out

Gilbert syndrome.

The patient was transfused with packed red cells and started on a

therapeutic trial of phenobarbitone. His total serum bilirubin

declined from 23.6 to 5.4 mg/dl over a period of 20 days. After

discontinuing phenobarbitone, the serum bilirubin increased to

11.2 mg/dl over the next week. The child was discharged after the

parents were counseled regarding need for regular transfusions and

follow-up.

It would have been of interest to observe the influence of regular

transfusion on reducing ineffective erythropoiesis, hemolysis and

therefore on serum bilirubin levels. Unfortunately, the child never

reported for follow-up. On contacting the parents it was found that

they never got the child transfused after the initial transfusions

carried out in the hospital. The child expired at home 3 months after

the diagnosis was made.

Clinically apparent jaundice is unusual in patients with b-thalassemia major. Co-inheritance of Gilbert syndrome has beenreported to cause hyperbilirubinemia in these subjects. Crigler–Najjar syndrome is another rare disorder of bilirubin metabolismcaused by mutation in the gene coding the enzyme UGT1A1. We

report a patient of b-thalassemia major who presented with persistentjaundice due to co-inherited Crigler–Najjar syndrome type 2 secon-dary to a novel mutation in UGT1A1 gene [homozygous basesubstitution at position 362 (GGT>AGT) in exon 3]. Pediatr BloodCancer 2010;54:627–628. � 2009 Wiley-Liss, Inc.

Key words: Crigler–Najjar; hemoglobin disorders; jaundice; thalassemia

� 2009 Wiley-Liss, Inc.DOI 10.1002/pbc.22313Published online 1 December 2009 in Wiley InterScience(www.interscience.wiley.com)

——————1Department of Pediatrics, Lady Hardinge Medical College, New

Delhi, India; 2Department of Pathology, Lady Hardinge Medical

College, New Delhi, India; 3Genetic Laboratory, ‘‘V Cervello’’

Hospital, Palermo, Italy

*Correspondence to: Anju Seth, Department of Pediatrics, Lady

Hardinge Medical College, New Delhi 110001, India.

E-mail: anjuseth.peds@gmail.com

Received 24 May 2009; Accepted 2 September 2009

DISCUSSION

Mild elevation in serum bilirubin levels may be observed in

patients with thalassemia due to ineffective erythropoiesis and

destruction of transfused red cells. Clinical jaundice is unusual

and serum bilirubin levels rarely exceed 3–4 mg/dl. Higher levels

usually indicate the presence of co-existing hepato-biliary dys-

function.

Galanello et al. [1] have shown that, in patients with transfusion

dependent thalassemia major with normal liver functions, the serum

indirect bilirubin levels are related to the configuration of the

polymorphic A(TA)nTAA motif of the UGT1A1 promoter. Those

patients with the (TA)7/(TA)7 configuration (the sequence associ-

ated with Gilbert syndrome) have levels higher serum bilirubin

(3.82� 1.40 mg/dl) than those with (TA)6/(TA)6 or (TA)6/(TA)7

genotype (serum bilirubin 0.80� 0.26 mg/dl). They concluded that

the variability in the serum bilirubin of patients with homozygous b-

thalassemia can be partly explained by Gilbert syndrome. Similar

observation has been reported by Tzetis et al. [2]. Coexistence of

Gilbert syndrome has also been reported in patients with sickle cell

disease [5], heterozygous b-thalassemia [3–5], and G6PD defi-

ciency [3] leading to variable serum bilirubin levels.

Our patient had persistent jaundice since early neonatal period.

We found only two other reported cases of b-thalassemia major

developing neonatal jaundice [6,7]. The etiology of jaundice in

these patients was not clearly defined. Our patient hadb-thalassemia

with associated Crigler–Najjar syndrome, a rare disorder of

bilirubin metabolism with two distinct forms. The more severe

Crigler–Najjar type I is characterized by severe chronic non-

hemolytic unconjugated hyperbilirubinemia with high levels of

serum bilirubin owing to the absence of bilirubin UGT activity. In

the milder Crigler–Najjar type II, bilirubin UGT activity is only

decreased and a significant reduction in serum bilirubin levels is

obtained after treatment with phenobarbitone, a phenomenon that

does not occur in Crigler–Najjar type I. Crigler–Najjar syndrome is

caused by the mutation in the UDP glucuronosyltransferase 1A1

(UGT1A1) [8] gene. To date nearly 50 mis-sense or non-sense

mutations in the UGT1A1 have been reported to cause Crigler–

Najjar syndrome [9]. A review of previous reports indicates that

mutation at codon 362 in exon 3 (causing base substitution

GGT>AGT), as found in our patient is a novel mutation. To

establish if the change was a polymorphism we studied as control 50

healthy individuals (100 chromosomes), who had no history of

jaundice, to study the population frequency. Of all the samples, only

five were heterozygous for other mutations already described, while

all others were negative for mutations in all five exons and in their

flanking splice junctions.

Since the serum bilirubin level in the patient reduced

promptly and remarkably on administration of phenobarbitone,

the diagnosis of Crigler–Najjar-2 as opposed to Crigler–Najjar-1 is

suggested.

The clinical findings of this case indicate that co-inheritance

of Crigler–Najjar syndrome is capable of modifying some

aspects of the clinical phenotype of homozygous b-thalassemia

and represents an example of the role of co-inherited modifying

gene(s) in the determination of clinical heterogeneity of a mono-

genic disorder.

REFERENCES

1. Galanello R, Cipollina MD, Dessi C, et al. Co-inherited Gilbert’s

syndrome: A factor determining hyperbilirubinemia in homozygous

b thalassemia. Hematologica 1999;84:103–105.

2. Tzetis M, Kanavakis E, Tsezou A, et al. Gilbert syndrome associated

with beta thalassemia. Pediatr Hematol Oncol 2001;18:477–484.

3. Maurizio S, Loredana L, Luca P, et al. The expression of Uridine

diphosphate glucuronosyltransferase gene is a major determinant of

bilirubin levels in heterozygous beta thalassemia and in glucose-6-

phosphate dehydrogenase deficiency. Br J Hematol 1997;99:437–

439.

4. Galanello R, Perseu L, Melis MA, et al. Hyperbilirubinaemia in

heterozygous beta-thalassaemia is related to co-inherited Gilbert’s

syndrome. Br J Hematol 1997;99:433–437.

5. Kalotychou V, Antonatou K, Tzanetea R, et al. Analysis of the

A(TA)(n)TAA configuration in the promoter region of the UGT1A1

gene in Greek patients with thalassemia intermedia and sickle cell

disease. Blood Cells Mol Dis 2003;31:38–42.

6. Furbetta M, Cossu P, Angius A, et al. Early onset of homozygous

beta-thalassaemia associated with neonatal jaundice. Arch Dis Child

1978;53:250–252.

7. Hazir T, Qazi SA, Abbas KA. Homozygous beta thalassemia

presenting as neonatal jaundice. J Pak Med Assoc 1995;45:306–

307.

8. Kakadol A, Ghosh SS, Sappal BS, et al. Genetic lesions of bilirubin

uridine-diphosphoglucuronate glucuronosyltransferase (UGT1A1)

causing Crigler-Najjar and Gilbert’s syndromes: Correlation of

genotype to phenotype. Hum Mutat 2000;16:297–306.

9. The Human Gene Mutation Database at the Institute of medical

genetics in Cardiff: Gene symbol UGT1A1, downloaded on 22/03/

2009.

Pediatr Blood Cancer DOI 10.1002/pbc

628 Aggarwal et al.