Download (Final) Report of GCC (UAEU/SQU) Research Grant Project Title

(Final) Report of GCC (UAEU/SQU) Research Grant
Project Title: Molecular Epidemiology and Diagnosis Study of Common Inborn Errors of
Metabolism Disorders in Oman and UAE
Research Affairs Reference: SQU-UAEU 1248-08-10
Principal Investigators:
Dr. Bassam R. Ali, Department of Pathology, Faculty of Medicine and Health Sciences, UAE
University, UAE
Dr. Said S. Al-Yahyaee, Department of Genetics, Sultan Qaboos University, Sultanate of
Oman
Background: Inherited metabolic disorders or inborn errors of metabolism are a group of
inherited, mostly recessive, disorders that encompass several branches of intermediates
biochemistry and metabolic pathways in the body associated with amino acid metabolism, fatty
acid oxidation, pyruvate and carbohydrate metabolism. Many of the metabolic disorders or
inborn errors of metabolism carry serious clinical consequences to the affected neonates or
young infants which include mild to severe irreversible mental retardation, physical handicaps
or even fatality. This of course results in significant effects on the quality of life and the
socioeconomic consequences to affected individuals, their families and the healthcare systems.
Due to their mode of inheritance, inborn errors of metabolism disorders are more common in
highly consanguineous populations such as UAE and Oman. The molecular causes of
metabolic disorders in both populations were largely unknown. The complications associated
with most inborn errors of metabolism disorders are preventable when detected early and
therefore early diagnosis is of paramount importance. In addition, the occurrence of new cases
of most inborn errors of metabolism disorders is potentially preventable by providing families
at risk with (1) accurate diagnosis of carriers, (2) effective genetic counseling and (3)
providing pre-marital testing, pre-implantation and prenatal diagnosis services.
Aim: Establish the molecular causes of inborn errors of metabolism disorders in UAE and
Sultanate of Oman and develop molecular diagnostic tests for their detection.
Objectives:
1.
Create a database of inborn errors of metabolism cases in Oman and UAE
2. Recruit and characterize families and individuals with common inborn errors of
metabolism
3. Perform mutational analysis by direct DNA sequencing of defective genes in affected
individuals
4.
Correlate mutation pattern with severity of the disorder
5.
Develop economical molecular diagnostic tests for those disorders
6.
Initiate mutation detection analysis to establish the carrier status of at risk individuals
1
Summary of Materials and Methods:
Patients. Individuals and families affected by metabolic disorders have been identified and
recruited for this study from Tawam Hospital (Al-Ain, UAE) and hospitals in the Sultanate of
Oman. The patients have been diagnosed by specialists in metabolic or clinical genetics.
Blood sample collection and DNA isolation. Peripheral blood samples have been collected
from subjects in EDTA tubes and the genomic DNA has been isolated following standard
procedures using FlexiGene kit.
Molecular Biology and Bioinformatics Techniques. Oligonucleotide primers for all the exons
and exon/intron junctions of the genes implicated in the identified metabolic disorders have
been designed using Primer3 (http://frodo.wi.mit.edu). The primers covered the coding regions
and the splice sites of the suspected genes. The designed primers have been custom-made
using commercial suppliers and were used to amplify those regions using the polymerase chain
reaction (PCR) technique. The amplified DNA fragments (amplicons) have been sequenced
using ABI 3130xl Gene Analayzer or equivalent instrument. Sequences obtained from each
subject have been compared using CLUSTALW2 to the reference sequence of the gene
deposited at the human genome browser and discrepancies in the sequences were carefully
analyzed using available softwares (e.g. Polyphen-2 and SIFT) to establish whether they were
pathogenic or non-pathogenic polymorphic variants.
Results and publications:
More than 70 mutations have been identified in patients with various IEM disorders from both
Oman and UAE (see tables 1-10 in appendix A for details). Parts of the generated data have
been published in three articles in international refereed biomedical journals as shown bellow:
1. Ali BR, Hertecant JL, Al-Jasmi F, Hamdan MA, Khuri S, Akawi NA & Al-Gazali L
(2011) New and known mutations associated with Inborn Errors of Metabolism in a
heterogeneous Middle Eastern population. Saudi Medical Journal, 32: 353-359.
2. Ben-Rebeh I, Hertecant, J, Al-Gazali L, Al-Jasmi F, Aburawi HE, Al-Yahyaee SA and
Ali BR (2012) Identification of Mutations Underlying Twenty Inborn Errors of
Metabolism in UAE population. Genetic Testing and Molecular Biomarkers, In Press.
3. Hertecant JL, Ben-Rebeh I, Marah MA, Abbas T, Ayadi L, BenSalem S, Al-Jasmi FA, AlGazali L, Al-Yahyaee SA and Ali BR (2012) Clinical and molecular analysis of Isovaleric
Acidaemia patients in the United Arab Emirates reveals remarkable phenotypes and four
novel mutations in the IVD gene. European Journal of Medical Genetics. Under review.
Copies of those publications are attached and more publications are expected. In addition, the
unpublished data are presented in tables 1 and 2 whereas the published data are presented ijn
tables 3-10..
Conclusions and limitations of the study: We largely achieved the first 4 objectives.
However, we have dealt with many more samples and genetic conditions than initially
anticipated. This is largely due to the larger number of patients affected by Inborn Errors of
Metabolism disorders than expected and the significant diversity nature of those conditions in
the UAE and Omani populations. Objectives 5 and 6 were not fully achieved because we gave
higher priority to identifying the mutations in patients before we start screening relative to
establish their carrier status (objective 6). For significant correlation studies between genotypes
2
and phenotypes, we need more samples to be analyzed. Developing economical diagnostic
tests should be straight forward by either using targeted DNA sequencing of the regions
harboring the mutations or by using restriction enzymes.
Budgets and Expenditure
UAEU Group
Salaries
Material and Equipment
Travel
Total
Budgeted (AED)
120,000
75,000
5,000
200,000
Actual (AED)
74,792.82
120,319*
4650
199,761.82
*This number is approximate as some of the invoices have not been fully settled yet. Since the
volume of samples and patients was above the anticipated number, we used more resources to
consumables.
SQU Group
Budgeted
UAE Dirhams
Actual (1st Year+2nd
Year)
Omani Rials
Salaries
120,000
5500+7700=13200
Material (consumables)
75,000
4300+2200=6500
Equipment
000.0+ 50
Travel
5,000
200+50=250
Total
200,000
20,000
3
Appendix A. Tables of results
Table 1. Mutations identified in metabolic disorders in Omani population
Disorder
(OMIM)
Propionic Acedimia
MIM 232000
Gene access
number
PCCA
NM_000282.3
MIM 232050
PCCB
NM_000532.4
Maple Syrup Urine
Disease
MIM 608348
BCKDHA
NM_000709.3
Glutaric Aciduria II
MIM 231675
ETFDH
NM_004453.2
Mucopolysaccharidosis
IIIB
MIM 252920
NAGLU
NM_000263.3
Mucopolysaccharidosis
IIID
MIM 252940
GNS
NM_252940
Homocystinuria
MIM 613381
Classical Galactosimia
MIM 606999
CBS
NM_001178008.1
GALT
NM_000155.2
Glycogen Storage
Disease I
MIM 613742
G6PC
NM_000151.2
SLC37A4
NM_001164277.1
MIM 602671
Methylmalonic
Acedimia
MIM 251000
Isovaleric Acedimia
MIM 60736
3-Hydroxy-3Methylglutaryl-CoA
Lyase Deficiency
MIM 613898
Tyrosinemia I
MIM 276700
Nucleotide
change
c.11901194del
AAGT
c.1195C>T
c.985-990ins
T
Protein
change
p.E397Vfs
p.R399W
p.E331X
p.A434T
c.1300G>A
c.332T>C
c.554del T
c.787-789del
TTC
p.L111P
p.L185Wfs
p.F263del
c.1414G>A
p.G472R
c.617del A
c.1597C>T
p.N206Tfs
p.R533X
c.19451947del TGG
p.W649del
c.2C>T
p.R2W
c.19del C
Novelty
%
Completed
%
Remain
Availability of
restriction site*
Novel
AgsI (2 sites)
MspJI (20 sites)
28.5%
Reported
Perez et
al.,(2003)
Novel
Novel
Novel
Novel
Reported
Olsen et
al.,(2003)
Novel
Reported
Mangas et
al.,(2008)
Novel
14.3%
MspJI (69sites)
57.1%
StuI (1 site)
61.1%
38.8%
TspRI (1sites)
BmrI (2 sites)
MboII (2 sites)
66.6%
33.3%
BtsIMutI (3 sites)
BsrI (3sites)
BanI (1 site)
---
80%
20%
StyI (1site)
Novel
100%
---
TauI (3 sites)
p.Q7Rfs
Novel
50%
50%
BsaI (3 sites)
c.404C>G
p.S135L
Reported
Fridovich et
al.,(1995)
100%
---
Hpy188I (1 site)
c.326G>C
p.C109S
Novel
12.5%
Hpy188I (1 site)
50%
c.1510T>C
p.L251P
Novel
37.5%
p.Y146Xfs
p.G427S
Novel
Novel
100%
---
TsoI (2 sites)
MnII (2 sites)
IVD
NM_002225.3
c.436-437del
TA
c.1279G>A
c.1175G>A
c.1184G>A
p.R392H
p.R395Q
Novel
Novel
88.8%
11.11%
AciI (3 sites)
TaqI (1 site)
HMGCL
NM_000191.2
c.914-915del
TT
p.F305Yfs
Reported
Mitchel et
al.,(1998)
100%
---
----
FAH
NM_000137.2
c.1142G>A
p.R381K
Novel
20%
80%
BssKI ( 4 sites)
MUT
NM_000255.3
Unpublished data
4
EcoNI (1 site)
Table 2. Mutations recently identified in inborn errors of metabolism disorders among Emirati
nationals.
Disorder
Gene
Nucleotide
change
Protein
change
National
Origin
Novelty
Microvillus inclusion
disease
MYO5B
c.1966C>T
p.R656C
UAE
Reported
Citrullinemia type 1
ASS1
c.535T>C
p.W179R
UAE
Reported
Medium chain acyl-CoA
dehydrogenase deficiency
ACADM
c.985A>G
p.K329E
UAE
Reported
Pyridoxine-dependent
seizures
ALDH7A1
Deletion of
exon 7
-
UAE
Novel
Mitochondrial DNA
Depletion
PLOG
c.3286C>T
p.R1096C
UAE
Reported
Isovaleric Acidemia
IVD
c.1175G>A
p.R392H
UAE
Novel
PKU
PAH
c.168+5G>C
Splicing
UAE
Reported
Unpublished data
5
Table 3. Mutations identified in amino acid and electrolyte metabolic disorders in the
United Arab Emirates population.
Disorder
(OMIM)
Gene access
number
Nucleotide
change
Protein change
National
Origin
Novelty
c.499+1G>C
Splice
UAE
Novel
c.1034C>G
p.P345R
Jordan
Novel
Lysinuric protein
intolerance
MIM 222700
SLC7A7
NM_003982
Citrullinemia
MIM 215700
ASS
NM_000050.4
c.349G>A
p.G117S
Pakistan
Reported
Berning et
al.,2008
Maple Syrup urine
disease
MIM 248600
BCKDHA
NM_000709.3
Deletion of exons
2-4
-
Egypt
Reported
Quental et
al.,2008
Carbamoyl phosphate
synthase I deficiency
MIM 238970
CPS1
NM_001875.2
c.1590dupT
p.Val531CysfsX9
Oman
Novel
Biotinidase
Deficiency
MIM 266150
BTD
NM_000060
c.1330G>C/
p.D444H/
UAE
c.557G>A
p.C186Y
Reported
Pomponio et
al., 2000
c.505+1G>A
Splice
Palestine
Reported
Christensen et
al.,2004
c.1169G>C
p.G390A
Iran
Reported
Mushimoto et
al.,2011
GCDH
NM_013976.2
Glutaric Acidemia I
MIM 231670
Published data
Ben-Rebeh I, Hertecant, J, Al-Gazali L, Al-Jasmi F, Aburawi HE, Al-Yahyaee SA and Ali BR (2012)
Identification of Mutations Underlying Twenty Inborn Errors of Metabolism in UAE population.
Genetic Testing and Molecular Biomarkers, In Press
6
Table 4. Mutations identified in Phenylketonuria (PKU, MIM 261600) disorder in the
United Arab Emirates population.
Nucleotide change
Protein change
National Origin
Novelty
c.970-2A>G
Splice
Palestine
Novel
c.842+1G>A
(IVS7+1G>A)
Splice
UAE
Reported
Dianzani et al.,1991
c.592-613del22/
C.581T>G
p.Y198Sfs136/
p.L194R
Egypt
Reported
Aurora et al.,2009
c.168+5G>C
(IVS2+5G>C)
Splice
UAE
Reported
Santos et al.,2008
c.592-613del22/ c.721C>T
p.Y198Sfs136/
p.R241C
Egypt
Reported
Aurora et al.,2009
c.143T>C/
c.842C>T
p.L48S/
p.P281L
Egypt
Reported
Karacic et al.,2009
c.143T>C/
c.727C>T
p.L48S/
p.R243X
Palestine
Reported
Karacic et al.,2009
c.169-171delGAG/
c.1066-11G>A
p.E57del/
Splice
Palestine
Reported
Dobrowolski et al.,2010
c.691 >C
p.S231P
Yemen
Reported
Diazani et al.,1992
c.441+5G>T
(IVS4+5G>T)
Splice
Iran
Reported
Zekanowski et al., 1996
c.472C>T
p.R231W
India
Reported
Takarada et al.,1993
c.1055del G/c.1066 11G>A
(IVS10-11G>A)
p.G352Vfs/
Splice
Morocco
Reported
Benit et al.,1994
Published data:
Ben-Rebeh I, Hertecant, J, Al-Gazali L, Al-Jasmi F, Aburawi HE, Al-Yahyaee SA and Ali BR (2012)
Identification of Mutations Underlying Twenty Inborn Errors of Metabolism in UAE population.
Genetic Testing and Molecular Biomarkers, In Press
7
Table 5. Mutations identified in Lipid, Steroid, Metal and mitochondrial energy
metabolic disorders in the United Arab Emirates population.
Disorder
(OMIM)
Gene access
number
Nucleotide
change
Protein change
National
Origin
Novelty
Oman
Reported
Franssen et al.,2010
Iraq
Reported
Aviram et al.,2000
Lipid disorder
Chylomicronemia
familial
MIM 118830
GPIHBP1
NM_178172.3
c.149G>A
p.G56Y
Steroid disorder
X-linked Ichthyosis
STS
MIM 308100
NM_000351.3
Deletion of the
entire gene
-
Mitochondrial energy disorders
Medium Chain Acyl
CoA Dehydrogenase
Deficiency(MCADD)
MIM 201450
ACADM
NM_000016.4
c.985A>G
p.K985E
Palestine
Reported
Matsubara et
al.,2003
Mitochondrial DNA
depletion syndrome
MIM 256810
MPV17
NM_002437.4
c.278A>C
p.G93P
Syria
Novel
Palestine
Reported
Deguti et al., 2004
Metal disorder
Wilson Disease
MIM 277900
ATP7B
NM 000053.2
c.122A>G
p.N41S
Published data:
Ben-Rebeh I, Hertecant, J, Al-Gazali L, Al-Jasmi F, Aburawi HE, Al-Yahyaee SA and Ali BR (2012)
Identification of Mutations Underlying Twenty Inborn Errors of Metabolism in UAE population.
Genetic Testing and Molecular Biomarkers, In Press
8
Table 6. Mutations identified in lysosomal storage disorder and miscellaneous metabolic
disorder in the UAE population.
Disorder
(OMIM)
Gene access
number
Nucleotide
Change
Protein
Change
National
Origin
Novelty
Lysosomal Storage disorder
Multiple Sulfatase
deficiency
MIM 272200
SUMF1
NM_001164674.1
c.603-2A>G
Splice
Iran
Novel
Cystinosis
MIM 219800
CTNS
NM_001031681.2
c.681+1G>A
(IVS9+1G>A)
Splice
UAE
Novel
Niemann-Pick type B
MIM 257220
SMPD1
NM_000543.4
c.1244C>T
p.A415V
UAE
Reported
Lan et al.,2009
c.2368C>T
p.Q790X
UAE
Novel
Alpha –Mannosidosis
MIM 248500
MAN2B1
NM_000528
c.2119C>T
p.Q707X
UAE
Novel
Sandhoff
GM2-Gangliosidosis
MIM 268800
HEXB
NM_000521.3
16 kb deletion
Unknown
Reported
GM1-Gangliosidosis
MIM 230500
GLB1
NM_000404.2
c.914+4A>G
Zhang et al.,1994
Splice
Palestine
Reported
Georgiou et al.,
2004
Miscellaneous disorder
Congenital Chloride
Diarrhea1 (CLD1)
MIM 214700
SLC26A3
NM_000111.2
c.559G>T
p.G187X
UAE
Reported
Hoglund et al.,
1998
Melanocortin-4 Receptor
Deficiency
MIM 601665
MC4R
NM_005912.2
c.485C>T
p.T187I
UAE
Reported
Tan et al.,2009
Published data:
Ben-Rebeh I, Hertecant, J, Al-Gazali L, Al-Jasmi F, Aburawi HE, Al-Yahyaee SA and Ali BR (2012)
Identification of Mutations Underlying Twenty Inborn Errors of Metabolism in UAE population.
Genetic Testing and Molecular Biomarkers, In Press
9
Table 7.Mutation in the IVD Gene causing Isovaleric Acidaemia (IVA) in the United Arab
Emirates.
Exon
DNA change
Protein change
Patients
Gender
Family #
Origin
Exon 11
c.11361138+4delTTGGTGA
p.F382fs
IV-3
F
F1
UAE
Exon 12
c.1184G>A
p.R395Q
IV-1
M
F2
UAE
IV-3
F
F4
UAE
Exon 12
c.1175G>A
p.R392H
IV-5
M
F5
UAE
Exon 12
c.1222G>A
p.E408K
IV-3
M
F3
Egypt
Data submitted for publication
Hertecant JL, Ben-Rebeh I, Marah MA, Abbas T, Ayadi L, BenSalem S, Al-Jasmi FA, AlGazali L, Al-Yahyaee SA and Ali BR (2012) Clinical and molecular analysis of Isovaleric
Acidaemia patients in the United Arab Emirates reveals remarkable phenotypes and four novel
mutations in the IVD gene. European Journal of Medical Genetics. Under Review.
10
Table 8. Mutations identified in amino acid metabolic disorders in the UAE population.
Disorder
Gene Details
Nucleotide
Change
Protein
Change
National
Origin
c.165delT
F55Lfs
Syrian
c.168+5G>C
(IVS2+5G>C)
Splice
UAE
R243X
Pakistan
c.728G>A
R243Q
Pakistani
c.755G>A
R252Q
UAE
R261Q
UAE
c.727C>T
Pheylketonuria
(PKU)
MIM#261600
PAH
NM_000277.1
MIM *612349
c.782 G>A
Canavan Disease
MIM #271900
Alkaptonuria
MIM #203500
ASPA
NM_000049.2
MIM *608034
HGD
NM_000187.2
MIM* 607474
Reference
Reports in other
populations
This Study
British;Tyfield et al.
1997
This Study
German; Zygulska et
al. 1993
This Study
Hungarian; Wang et
al. 1990
This Study
Chinese; Wang et al.
1991
This Study
French; Benit et al.
1994
This Study
Mediterranean;
Abadie et al. 1989
Lithuania;
Kasnauskiene et al
2003;
Poland; Dobrowolski
et al. 2009
Non-Jewish of
European descent;
Kaul et al. 1994
Spanish; BeltranValero de Bernabe et
al. 1999
c.1066-11G>A
Splice
UAE
This Study
c.914C>A
A305E
NA
This Study
c.174delA
(c.342delA)
R58 fs
UAE
Abdulrazzaq
et al. 2009
del exons 9 & 10
-
Pakistan
This Study
Novel
European and North
American; Perez et al.
2003
Maple Syrup
Urine Disease
(MSUD) MIM
#248600
BCKDHB
NM_183050.1
MIM *284611
Propionic
Acidemia
MIM# 606054
PCCB
NM_000532.3
MIM *232050
c.991dupT
E331X
Oman
This Study
Citrullinaemia,
classic
MIM#215700
Argininemia
(Arginase
Deficiency)
MIM #207800
ASS1
NM_000050.4
MIM*603470
c.349G>A
G117S
Pakistan
This Study
ARG1
NM_000045.2
MIM* 608313
c.93delG
L31 fs
Pakistan
Hertecant et
al. 2009
Japanese and
American; Gao et al.
2003
Published data:
Ali BR, Hertecant JL, Al-Jasmi F, Hamdan MA, Khuri S, Akawi NA & Al-Gazali L (2011) New
and known mutations associated with Inborn Errors of Metabolism in a heterogeneous Middle
Eastern population. Saudi Medical Journal, 32: 353-359.
11
Table 9. Mutations identified in carbohydrate and mitochondrial energy metabolic
disorders in the UAE population.
Disorder
Gene Details
Nucleotide
Change
Protein
Change
Disorders of carbohydrate metabolism
Glycogen storage
G6PC2
Disease Type
NM_000151.2 c.59A>G
1A
MIM*608058
MIM +232200
Hereditary
Fructose
Intolerance
MIM #229600
ALDOB
NM_000035.3
MIM*612724
Q20R
c.524C>A
Fructose-1,6FBP1
bisphosphate
c.616_619delAA
NM_000507.2
deficiency
AG
MIM*611570
MIM #229700
Disorders of mitochondrial energy metabolism
c.460G>A/c.578
β-Ketothiolase
ACAT1
T>G
Deficiency
NM_000019.3
(compound
MIM#203750
MIM*607809
heterozygousity)
Medium Chain
Acyl-CoA
ACADM
Dehydrogenase
c.431_434delAG
NM_000016.4
Deficiency
TA
MIM*607008
(MCADD)
MIM #201450
National
Origin
UAE
Reference
Reports in other
populations
This Study
German; Seydewitz &
Matern 2000
A175D
Jordan
This Study
German,
Mediterranean , Swiss,
Italian and Yugoslav;
Santer et al. 2005;
Cross et al. 1990
K206fs
UAE
This Study
German; Herzog et al.
2001
E154K
M193R
India
This Study
Novel
K144fs
Palestine
This Study
German; Ensenauer et
al. 2005
Published data:
Ali BR, Hertecant JL, Al-Jasmi F, Hamdan MA, Khuri S, Akawi NA & Al-Gazali L (2011) New
and known mutations associated with Inborn Errors of Metabolism in a heterogeneous Middle
Eastern population. Saudi Medical Journal, 32: 353-359.
12
Table 10. Mutations identified in lysosomal storage disorders and other IEM conditions in
the UAE population.
Disorder
Glycogen storage
Disease Type II
(GSDII, Pompe
Disease)
MIM#232300
Gene Details
GAA
NM_000152.3
MIM*606800
I-Cell Disease
(Mucolipidosis
type II) MIM
#252500
GNPTAB
NM_024312.3
MIM* 607840
Sandhoff Disease
MIM#268800
HEXB
NM_000521.3
MIM*606873
GM1Gangliosidosis,
Type I
MIM#230500
GLB1
NM_000404.2
MIM *611458
Peroxisomal disorders
Zellweger
PEX6
Syndrome
NM_000287.3
MIM#214100
MIM*601498
Nucleotide
Change
Protein
Change
National
Origin
Reference
Reports in other
populations
Dutch , British,
Australian, Greek,
Swiss and Israeli;
Hermans et al. 2004
Dutch and American;
Kroos et al. 2008
c.340_341insT
K114fs
Palestine
c.1327-2A>G
Splice
UAE
Hamdan et
al. 2008
c.3503_3504del
TC
L1168fs
UAE
This Study
Caucasian Arab;
Kudo et al. 2006
c.850C>T
R284X
Pakistan
This Study
Italian; Zampieri et al.
2009
c.1465_1466del
AT
I489fs
India
This Study
Novel
c.451G>T
D151Y
UAE
Georgiou et
al. 2004
c.914+4A>G
Splice
UAE
Georgiou et
al. 2004
c.611C>G
S204X
UAE
This Study
Novel
c.993A>G
(1073A>G)
Q331R
UAE
This Study
Irish; Moghrabi et al.
1993
This Study
Disorders of bilirubin metabolism
Crigler-Najjar II
MIM# 606785
UGT1A1
NM_000463.2
MIM*191740
Published data:
Ali BR, Hertecant JL, Al-Jasmi F, Hamdan MA, Khuri S, Akawi NA & Al-Gazali L (2011) New
and known mutations associated with Inborn Errors of Metabolism in a heterogeneous Middle
Eastern population. Saudi Medical Journal, 32: 353-359.
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