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Genotype-phenotype correlation
In PKU
Szeged, December 12, 2011.
Introduction
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HPA – BH4-responsive – classic
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Severity depends partially on the mutation of PAH enzyme
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atypical (BH4-dependent)
Genotype correlates with the metabolic phenotype in most of the cases –
homozygous, severe null mutations
… or „mild” mutations
Most of the patients are compound heterozygous
(or – in Hungary - R408W homozygous)
There are significant genotype-phenotype inconsistencies between identical
PAH mutation bearing patients
So the PKU/HPA phenotype manifestation is more complex than it could be
predictable based on the Mendelian inheritance of the mutated alleles
PAH enzyme
N-terminal regulatory domain: AA 1–142
Catalytic domain: AA 143–410
C-terminal domain: AA 411–452
→ tetramerization
PAH gene mutations
Missense 60,5%
Deletion 13,5%
Splice
11%
Silent
5,7%
Nonsense
5,0%
Insertion
3,2%
Unknown
0,7%
Silent/splice
0,5%
0
50
www.pahdb.mcgill.ca
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12q22-q24.1, 13 exon
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564 mutation
100
150
200
250
300
350
400
PAH gene mutations
Metabolic phenotype
BH4-responsiveness
Mutation type
mild
Mostly responsive (80%)
mild/mild
moderate
May be responsive, depending of the
„mild” mutation type
mild/null
severe (classic)
Non-responsive
null/null
BH4-responsive PAH mutations
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~70 known mutation
Mostly point-mutation (never
splice, insertion or deletion)
Mostly in the catalytic
domain, but not in the
cofactor-binding site
BH4-responsive PAH
mutations
mutáció
R408W
A403V
allél
mutáció
45 null
5 may be responsive
allél
E390G
1
responive
T380M
1
responive
Y414C
1
may be responsive
R252W
1
null
IVS12+1G>A
5 splice, null
R261Q
5 may be responsive
IVS4+47C>T
1
L48S
4 responsive
P225T
1
IVS10-11G>A
3 splice, null
IVS10+94G>A
1
silent
IVS7+1G>A
1
splice
I174V
1
V177L
1
IVS3-22C>T
3 silent
R158Q
3
IVS7-4delT
3 unknown, splice?
IVS4-5C>G
1
c.15delCT
2
S350Y
1
P281L
2 null
Q172X
1
null
IVS11+1G>A
2 splice, null
G148D
1
unknown
M276T
1
I95F
1
R243X
1
null
V245A
1
HPA
F302V
1
unknown
c.39delC
2
IVS4+5G>T
2 splice
splice
Our cases: null mutations
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Since 2006 we could not have the patients' DNA sequenced
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52 patients' DNA sequence and mutations are known
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Classic PKU: null mutations
Allél 1
Allél 2
esetszám mutáció típus
R408W
R408W
14
missense
IVS12+1G>
A
IVS11+1G>A
2
splice
R408W
IVS10-11G>A
1
missense/splice
R408W
IVS12+1G>A
1
missense/splice
Non-null mutations
Trefz et al. J Inherit Metab Dis (2009) 32:22–26
Mutation
BH4-response
(allele %)
In vitro residual
enzyme activity (%)
L48S
83
39
Regulatory domain
A403V
100
32
Catalytic domain
E390G
100
72
Catalytic domain, enzyme surface
Y414C
93
36
Tetramerization
R261Q
86
38
Active site secondary structure,
tetramerization
R158Q
56
10
Catalytic domain
Same genotype – different BH4-responsiveness
Y414C (mild)/R408W (null) → moderate PKU?
Y414C
BH4-resp: 93%
Residual enzyme
activity: 36%
BH4 loading with 20 mg/kg bw in 3 patients, all genotype Y414C/R408W.
Lindner M et al, Mol Genet Metab 2001; 73:104-106.
HPA cases
Allele 1
Allele 2
Phe level (mmol/l)
BH4 loading test?
T380M
wt
130
not performed
E390G
A403V
180
not performed
no diet
S350Y
A403V
240
not performed
on diet
Interesting cases
KA
Allele 1 Allele 2 Phe level
Remark
R408W
on diet
R261Q
600-1200
Matalon 2005
BH4-responsive
Desviat 2004
non-responzive
Erlandsen 2004
partially responsive
DG
R252W
?
300-1200
Tesztelt BH4-reszponzív esetek
Allél 1
Allél 2
Phe
szint
Megjegyzés
SZB & SZF
IVS7-4delT
A403V
360
BH4-reszponzív
NÁ
R408W
A403V
240
BH4-reszponzív
UIA
V245A
R158Q
240
BH4-reszponzív
Genotype-phenotype in PKU
Pathogenic mutation
phenotype 1
phenotype 2
phenotype 3
Genetically determined factors:
other intragenic differences, polymorphisms
modifying genes: aminoacid transport, competition,
metabolism, excretion
Non-genetically determined factors
Conclusion
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Confirming the genotype may help in the judgement and
predicition of the severity of the disease
There are significant genotype-phenotype inconsistencies
between similar PAH mutation bearing patients: mainly the
regulatory domain mutations lead to unpredictable phenotype, in
the cases of compound heterozygosy
BH4-loading test result may vary in a patient - intracellular
environment, metabolic status influence the mutated enzyme
activity
The standard(ized) Phe+BH4 loading test is the gold standard for
planning the treatment!