congenital central hypoventilation syndrome ( cchs ) is a life - threatening disorder characterized by an idiopathic failure of the automatic control of breathing , particularly during sleep . in cchs , patients breathe with insufficient depth due to impaired sensitivity to hypercapnea and hypoxia .
hirschsprung 's disease ( hd ) , also known as congenital aganglionic megacolon , is characterized by the absence of ganglion cells in any part of the bowel wall .
the absence of ganglion cells in hirschsprung 's disease inhibits normal peristalsis , resulting in a functional intestinal obstruction .
the combination of cchs and hd was reported first by haddad in 1978 and it has been extremely rare with approximately 60 cases reported in the worldwide literature ( 1 , 2 ) .
previous studies described that de novo mutation of the phox2b gene was involved in cchs pathogenesis ( 3 , 4 ) .
many patients with cchs are heterozygous for a polyalanine expansion mutation in the second polyalanine repeat residue of the phox2b gene , with one allele having 20 repeats and the affected allele having 25 - 33 repeats of the polyalanine sequence ( 3 , 4 ) .
some authors reported that a subset of patients had other mutations in the phox2b gene ( 5 , 6 ) . in korean population ,
only one case has been reported , however , genetic analysis has not been performed ( 7 ) . here
, we report a haddad syndrome with the phox2b gene mutation for the first time in a korean .
this 41-week - gestation male was born to a 30-yr - old mother with good health by normal spontaneous vaginal delivery in november 2009 .
there was neither family history of congenital anomalies nor maternal exposure to drugs during pregnancy .
the baby had apgar scores of 7 and 5 at 1 and 5 min , respectively .
the baby was immediately transferred to our hospital for further evaluation and management after stabilization .
for the next few days , we tried extubation several times , but failed to succeed because the baby suffered from recurrent apnea and hypercarbia leading to the clinical suspicious of cchs . on day 8 of life ,
a cranial ultrasound study was performed and it did not show evidence of hemorrhage or mass lesions . during the second day of life , it was noted that the patient had distended abdomen and was unable to pass meconium without rectal stimulation .
a gastrografin contrast enema study showed multiple meconium plugs in the colon suggesting meconium plug syndrome ( fig .
meconium was passed and abdominal distension was relieved after contrast study . on day 6 of life , he showed sustained abdominal distension with poor bowel movement .
a barium contrast enema study was needed to rule out other causes of lower gastrointestinal obstruction .
he was noted to have long segment hd with a transition zone at mid descending colon .
seromuscular biopsies of the bowel taken 3 cm proximal and distal to the transition zone revealed the presence of ganglion cells in the proximal biopsy specimen only .
the cytogenetic study was performed clinically in the infant and his parents and their karyotypes were normal .
genomic dna of the patient and his parents was isolated from peripheral blood using the wizard genomic dna purification kit according to the manufacturer 's instructions ( promega , madison , wi , usa ) . amplification of the coding sequence of the phox2b gene by polymerase chain reaction ( pcr ) was carried out as previously described with minor modifications ( 4 ) .
polymerase chain reaction - single strand conformational polymorphism ( pcr - sscp ) was performed as previously described ( 8) .
pcr products in the patient , his parents , and normal controls were electrophoresed through 8 and 10% polyacryamide gels ( fig .
a mutant band was found , however , his parents had no mutant band . to determine the sequences of the dna sample showing the mutant band , direct sequencing was performed by macrogen inc , in korea
however , the quality of the result was not clear . to get apparent result of sequencing ,
paisley , united kingdom ) and 10 single colonies were collected for extracting plasmid dna .
direct sequencing revealed expanded alleles containing polyalanine 26 repeats in exon 3 of the phox2b gene , further supporting the diagnosis cchs ( fig .
importantly , we were able to prove a phox2b gene mutation containing polyalanine 26 repeats in this patient .
hirschsprung 's disease often produces intestinal obstruction in the neonate , characterized by abdominal distention , bilious vomiting , and failure to pass meconium within the first 48 hr .
cchs is characterized by relatively normal ventilation when the infant is awake and hypoventilation , even apnea , during sleep .
haddad syndrome is interpreted not so much as a combination of two diseases but rather as a complicated form of cchs . when hd is associated with cchs , it is different from classical hd , characterized by extensive intestinal aganglionosis ( 9 ) .
the incidence of cchs is extremely low in hd cases , however , the incidence of hd in cchs cases varies from 16% to 50% according to previous studies ( 3 , 5 , 6 , 9 ) .
additional symptoms are ophthalmic abnormalities , esophageal dysmotility , sensorineural hearing loss , neural crest tumors ( ganglioneuromas , neuroblastomas ) , and dysmorphic facial features ( 3 , 5 , 9 ) .
however , additional symptoms were not found in our case . according to previous report in korean
, a haddad syndrome patient also had no additional symptoms except congenital ptosis ( 7 ) .
( 3 ) , and its rate was over 90% among cchs patients ( 4 , 6 ) .
polyalanine expansions are the most common mutations in patients with cchs , though rare mutations have been reported such as frameshift mutation , nonsense mutation or missense mutation ( 3 - 5 ) . in our case , his parents showed no phox2b gene mutation , suggesting de novo mutation of the phox2b gene in haddad syndrome .
interestingly , genotype - phenotype correlation has been described in cchs patients with the phox2b gene mutation ( 4 ) .
increasing polyalanine repeat mutation size is associated with a more severe clinical phenotype ( 4 ) .
( 10 ) described that haddad syndrome ( cchs + hd type ) showed usually expansions of + 6 and + 7 alanines , and frameshift or missense phox2b mutations may predispose to neuroblastoma .
clinical phenotype of our patient was severe , presenting long segment hd and no capacity to maintain sufficient oxygenation and ventilation for at least 30 - 60 min of sleep after removing mechanical ventilation support ( 4 ) . however , our case with expansion of + 6 alanines had no additional symptoms like neural crest tumors . though the symptoms of cchs and hd were severe , no additional symptom in the patient with this mutation type was in agreement with the hypothesis by trochet et al .
this result supports that the assay for phox2b polyalanine repeat mutation represents a highly sensitive and specific technique for confirming the diagnosis of cchs and estimating the prognosis . in summary
, this report presents a rare case of haddad syndrome with de novo mutation of the phox2b gene in korean for the first time . considering the difficulty of diagnosis of this syndrome because of overlapped of symptoms and signs in an infant
 congenital central hypoventilation syndrome with hirschsprung 's disease , also known as haddad syndrome , is an extremely rare disorder with variable symptoms . 
 recent studies described that congenital central hypoventilation syndrome had deep relation to the mutation of the phox2b gene in its diagnosis and phenotype . 
 we report a newborn male infant with clinical manifestations of recurrent hypoventilation with hypercapnea and bowel obstruction . 
 these clinical manifestations were compatible with congenital central hypoventilation syndrome and hirschsprung 's disease , and polyalanine 26 repeats in the phox2b gene supported the diagnosis of congenital central hypoventilation . 
 we described a first case of haddad syndrome in korean and its clinical and genetic characteristics were discussed . 
INTRODUCTION
CASE DESCRIPTION
DISCUSSION