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Indian Pediatr 2017;54: 467-471 |
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Clinical and Molecular
Characterization of Children with Neonatal Diabetes Mellitus at
a Tertiary Care Center in Northern India
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Vandana Jain, Amit Satapathy, Jaivinder Yadav, Rajni
Sharma, *Venkatesan Radha, *Viswanathan Mohan,
#Elisa De Franco and
#Sian Ellard
From Division of Pediatric Endocrinology, Department
of Pediatrics, AIIMS, New Delhi, and *Madras Diabetes Research
Foundation, Chennai, India; and #Institute of Biomedical and Clinical
Science, University of Exeter Medical School, Exeter, UK.
Correspondence to: Prof. Vandana Jain, Division of
Pediatric Endocrinology, Department of Pediatrics,
AIIMS, Delhi-110 029, India.
Email: [email protected]
Received: May 19, 2016;
Initial review: August 31, 2016;
Accepted: March 28, 2017.
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Objective: To study the genetic mutations and
clinical profile in children with neonatal diabetes mellitus Methods:
Genetic evaluation, clinical management and follow-up of infants
with neonatal diabetes Results: Eleven infants were studied of
which eight had permanent neonatal diabetes. Median age at presentation
was 8 weeks and mean (SD) birth weight was 2.4 (0.5) kg. Pathogenic
genetic mutations were identified in 7 (63.6%) children; 3 infants with
mutations in KCNJ11 gene and 1 in ABCC8 were switched to
oral sulfonylureas; 2 infants had mutations in INS and 1 in
ZFP57. Conclusion: Neonatal diabetes mellitus is a
heterogeneous disorder. Identification of genetic cause guides
clinical management.
Keywords: Diabetic ketoacidosis, Genetic analysis, HbA1c,
Sulfonylureas.
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Neonatal diabetes mellitus (NDM) is a
constellation of rare monogenic disorders, with an estimated prevalence
of 1 in 89,000[1]. The condition usually manifests within the first 6
months of life, and occasionally in second half of the first year with
polyuria, dehydration and diabetic ketoacidosis [1]. In the past three
decades, monogenic mutations in more than 20 genes have been identified
that lead to impaired development, reduced function or progressive
destruction of the pancreatic beta cells [2,3].
There are two distinct subtypes of NDM: (i)
permanent; and (ii) transient form characterized by resolution of
diabetes within weeks to months [4]. Activating mutations in the genes
ABCC8 and KCNJ11, which encode for the Kir6.2 and SUR1
subunits, respectively, of the ATP-sensitive potassium (KATP) channel of
the pancreatic beta cells are the commonest cause of permanent NDM [2].
Of the transient NDM cases, nearly 70% of are caused by loss of
methylation at the differentially imprinted locus on chromosome 6q24
[4], while KCNJ11 and ABCC8 mutations account for majority
of the remaining cases [5]. The present study was conducted with the
objective of describing the genetic mutation profile and the clinical
presentation, management and outcome of children with neonatal diabetes
mellitus at our institute over the last seven years.
Methods
We reviewed the case records of all infants with NDM
presenting to All India Institute of Medical Sciences (AIIMS), New Delhi
from January 2009 to December 2015. NDM was diagnosed when the child
presented within the first 6 months of life with symptoms of polyuria
and dehydration with random blood sugar more than 200 mg/dL, or when the
presentation was later in the first year but proven to be of monogenic
etiology [1,2].
All infants were hospitalized at presentation.
Diabetic ketoacidosis (DKA) was managed as per protocol [6]. Subsequent
management was with 0.3 to 0.7 U/kg/day of subcutaneous insulin in
multiple divided doses. Majority of the infants needed 1-2 doses of
Neutral Protamine Hagedorn (NPH) insulin and 3-4 doses of Regular
insulin. Parents were instructed to dilute the insulin with normal
saline and administer it using insulin syringe. They were given simple
written instructions to monitor blood glucose at home, manage hypo- and
hyper-glycemia, and check urinary ketones when baby was sick [7]; close
follow-up was maintained.
Genetic mutation analysis was done after obtaining
informed consent from the parents. Genetic testing for six infants
(cases 1, 6, 7, 8, 9 and 11 in Table I) was carried out at
Department of Molecular Genetics, Royal Devon and Exeter NHS Foundation
Trust, Exeter, UK. It consisted of rapid Sanger sequencing of the
KCNJ11, ABCC8 and INS genes with additional EIF2AK3
analysis in infants born to related parents. In patient 8, a custom
designed targeted next generation sequencing of all known NDM genes [3]
was also done; and methylation analysis for chromosome 6q24 was
conducted at Wessex Regional Genetics Laboratory, Salisbury [8,9].
Mutation testing for cases 2, 3, 4 and 5 was done at Department of
Molecular Genetics, Madras Diabetes Research foundation, Chennai.
Sequencing was carried out on ABI 3500 Genetic Analyzer (Applied
Biosystems, Foster City, CA) using the Big Dye terminator V3.1, and the
sequences were compared with the database. Published primers were used
to amplify the DNA for KCNJ11, ABCC8 and INS [10,11].
TABLE I Summary of the Clinical Profile and Mutations in the Patients with Neonatal Diabetes Mellitus
Case No. |
City, State |
Gender, Birth weight, Age at presentation |
Family history
|
Mutation
|
Management
|
Current age and HbA1c |
Index case |
Parents |
1. |
Jalandhar, Punjab |
M, 2.90 Kg,4 weeks |
Paternal grandfather and great grandmother had type 2 DM |
Compoundheterozygous missense mutationsin
ABCC8(p.Arg168Cys andp.Gly1256Ser) |
Heterozygous missense mutation in ABCC8 in both parents
(p.Arg168Cysin mother and p.Gly1256Ser in father) |
Glibenclamide @0.4 mg/kg/day |
7 y; 6.8% |
2 |
Delhi |
F,1.50 Kg,16 weeks |
Adopted child |
No mutation in KCNJ11,ABCC8 or INS |
Not tested |
Insulin @0.6 u/kg/day |
6 y;7.6% |
3 |
Delhi |
F,3.10 Kg44 weeks |
Nil |
Heterozygous missense mutation in INS (p. Arg89Cys) |
No mutation |
[email protected] u/kg/day |
4 y; 7.1% |
4 |
Indore, Madhya Pradesh |
M,2.50 Kg3 weeks |
Paternal grand mother had type 2 DM |
Heterozygous mutation in KCNJ11 (p. Arg50Gln) |
Heterozygous mutation in KCNJ11 (p.Arg50Gln)in mother, No
mutation in father |
Glibenclamide @2 mg/kg/day |
3 y;6.2% |
5 |
Manipur |
M, 2.20 Kg,10 weeks |
Nil |
No mutation in ABCC8, KCNJ11 or INS |
Not done |
[email protected] u/kg/day |
2.5 y; 8.9% |
6 |
Jaipur Rajasthan |
M,3.20 Kg,12 weeks |
Nil |
Heterozygous missensemutation in KCNJ11 (p. Phe333Leu) |
No mutation |
Glibenclamide @1 mg/kg/day |
2 y;7.4% |
7 |
Ranchi,
Chattishgarh |
F,2.05 Kg,4 weeks |
Nil |
Heterozygous missensemutation in KCNJ11 (p.Arg201Cys) |
No mutation |
Glibenclamide @0.9 mg/kg/day |
2 y;6.2% |
8 |
Delhi |
M, 2.30 Kg,1 week |
Sibling died on day21 due to delayed diagnosis of neonatal
diabetes |
Homozygous frameshiftmutation in ZFP57(p.Gln469fs) |
Not done |
Off insulin after4 months of age |
1.5 y; 6.0% |
9 |
Jhansi
Uttar Pradesh |
F,1.85 Kg8 weeks |
Nil |
No mutation in KCNJ11, ABCC8 and INS |
No mutation |
Off insulin after
4 months of age |
1 y;8.1% |
10 |
MeerutUttar Pradesh |
M, 2.70 Kg10 weeks |
Nil |
Not done |
Not done |
[email protected] u/kg/day |
Died at 14 weeks |
11 |
Delhi |
M,1.75 Kg,4 weeks |
Nil |
Heterozygous missense mutationin INS (p.Ser101Cys) |
No mutation |
[email protected]/kg/day |
6 mo;7.1% |
After receiving the genetic report, four infants with
mutations in KCNJ11 or ABCC8 were switched to oral
sulfonylureas as per the protocol [12]. All children were followed-up
for growth, development and glycemic control.
Results
Eleven infants (8 boys) were diagnosed with NDM. The
clinical, demographic and genetic mutation profile of the cases is
summarized in Table I. The median (range) age at initial
presentation was 8 (1- 44) weeks. Mean (SD) birth weight was 2.4 (0.5)
kg, with 7 (63.6%) infants being low birth weight. Six (54.5%) infants
had presented with DKA; 4 (36.4%) with polyuria, irritability and
increased demand for feeds, and one was diagnosed by surveillance of
blood glucose in view of previous sibling’s death with NDM.
Eight (72.7%) infants in our cohort had permanent
NDM, 2 had transient NDM and the type was uncertain in one child who
died at 3 months. Pathogenic genetic mutations were identified in seven
(63.6%) infants (Table
I). The most commonly affected gene was
KCNJ11 in three cases, followed by INS gene in two cases. One
patient each had mutation in ABCC8 and ZFP57 genes. None
of the common mutations were identified in three infants. In one
patient, initial sample was insufficient and the baby died before repeat
sample could be obtained.
We were able to successfully transfer all four
children with mutations in either KCNJ11 or ABCC8 genes
from insulin to oral glibenclamide. The age at transfer ranged from 2 to
9 months. The initial dose of glibenclamide on which glycemic control
was achieved ranged from 0.5 to 2 mg/kg/day, and was reduced on
follow-up. Glycemic control was excellent in these children with no
episode of DKA or symptomatic hypoglycemia. At last follow-up, the mean
(SD) HbA1c level in these 4 children was 6.6 (0.5)%. Two infants having
mutations in the INS gene were continued on subcutaneous regular
and NPH insulin, and are under regular follow-up.
Case 8 was diagnosed within the 1 st
week of life. He was born out of consanguineous marriage, with the
conspicuous history of death of a sibling with hyperglycemia at 3 weeks
of age. A homozygous frameshift mutation in the ZFP57 gene
(c.1405del, p.Gln469fs) was identified. Complete loss of maternal
methylation at the transient neonatal diabetes differentially methylated
region on 6q24 was confirmed. Insulin could be stopped at 4 months of
age. In the two other children in whom a mutation in the more common NDM
genes was not identified (cases 2 and 5), insulin was continued, and
glycemic control was satisfactory. We lost one baby in our series at 3
months of age to probable sepsis and DKA. The current median (range) age
of our cohort is 27 (6-84) months, and the mean (SD) HbA1c is 7.1
(0.9)%. Growth and development of all the children is within normal
limits.
Discussion
We have presented the clinical and molecular
characteristics for our series of 11 infants with NDM; pathogenic
mutations were identified in seven (63.6%). Four cases of permanent NDM
were caused by mutations in the K ATP
channel (KCNJ11 and ABCC8) genes, and could be
successfully switched over to oral glibenclamide. INS gene
mutations were present in 2 children and transient NDM related to loss
of 6q24 methylation due to a ZFP57 homozygous mutation was
identified in one child. Seven out of the 11 cases in our cohort had low
birth weight. This is a relatively consistent feature of NDM, and
reflects on the role of insulin as an important growth factor in fetal
life [2].
Limitations of this study are: small sample size and
incomplete genetic evaluation for some of the patients. Of the four
cases without confirmed genetic mutation, two had undergone evaluation
for only the three most common genes (KCNJ11, ABCC8 and
INS), one had transient NDM but could not be tested for loss of
methylation and one baby died before genetic testing.
In a recent large study, it was reported that in
offspring of non-consanguineous parents, mutations in KCNJ11,
ABCC8, INS and 6q24 methylation abnormalities accounted for
approximately 28%, 18%, 10% and 10%, respectively of the NDM cases;
while among children of consanguineous parents, EIF2AK3 mutations
were the commonest [2]. In an earlier Indian series, Jahnavi, et al.
[14] had reported a total of 12 mutations in 33 children with
infancy-onset diabetes; ABCC8 in 7, KCNJ11 in 3 and INS
in 2. In another recent series from Chennai [15], mutation profile of 10
children with NDM (six of whom were born to consanguineous parents) was
presented. Nine of these had permanent NDM, attributable to mutations in
ABCC8 and INS in 2 each, and KCNJ11, EIF2AK3, SLC19A2,
NEUROD1 and PDX1 in one each; while one had transient NDM due
to loss of methylation in chromosome 6q24.
The identification of mutations in KCNJ11 or
ABCC8 genes highlights the importance of establishing early
genetic diagnosis, as these patients can be switched over to oral
sulfonylureas, that are not only more convenient, but also associated
with reduction in episodes of hypo- and hyper-glycemia [12,13]. Insulin
gene (INS) mutations are associated with synthesis of misfolded
preproinsulin or proinsulin that is retained in endoplasmic reticulum
and results in apoptosis of the beta cells, leading to permanent
insulin-dependence. The age at onset is variable with many patients
presenting beyond 6 months or even beyond infancy. INS mutations
are typically sporadic and heterozygous [10], as in both our cases.
Abnormalities in 6q24 are typically associated with moderate
intrauterine growth retardation and diabetes within first week of life.
Hyperglycemia remits by few weeks to months, but may recur during
illness [4], and nearly half relapse during adolescence or early
adulthood [16].
To conclude, genetic testing is essential in children
with neonatal diabetes mellitus. It not only confirms the diagnosis, but
also defines the prognosis of the affected child, and predicts the risk
in subsequent pregnancies. Since mutations in KCNJ11 and ABCC8
have major therapeutic implications, these genes should be screened
first. Testing of 6q24 must be considered in infants with NDM who had
intrauterine growth retardation, and go into spontaneous remission. The
role of adequate counseling and education of the parents in appropriate
management of the young infants with NDM cannot be over-emphasized.
Early diagnosis, identification of the genetic defect and appropriate
clinical management can be very rewarding.
Acknowledgements: Dr Deborah Mackay and Prof
Karen I Temple for performing the methylation specific analysis, and to
Mr Anil Kumar for extracting DNA and ensuring safe transport of the
clinical samples from AIIMS to the laboratories in Chennai or Exeter.
Contributors: VJ: conceptualized the study
design, critically revised the paper and will act as guarantor; VJ, AS:
drafted the paper, VJ, AS, JY, RS: acquired the data; VR, VM, EDF, SE:
performed and interpreted the mutation analysis. All authors contributed
to manuscript writing and approved the final draft.
Competing interests: None stated.
Funding: SE is the recipient of a Wellcome Trust
Senior Investigator award (grant number: WT098395/Z/12/Z). The genetic
analysis carried out at Institute of Biomedical and Clinical Science,
Exeter has been supported by this grant.
What This Study Adds?
• The commonest cause of permanent neonatal
diabetes mellitus in our series of north Indian infants was
mutations in the KATP channel (KCNJ11 and ABCC8)
genes, and these children could be switched over to oral
sulfonylureas.
• Mutations in INS gene were the next
common and affected children require life-long insulin therapy.
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