Download Development of a New Method to Prioritise Gene Analysis in

Development of a New Method
to Prioritise Gene Analysis in
Familial Hypertrophic
Cardiomyopathy
Jayne Duncan
West of Scotland Regional Genetics
Service, Glasgow
Familial Hypertrophic
Cardiomyopathy (FHC)
• Autosomal dominant disorder showing
variable penetrance and age of onset.
• Affects approximately 1/500 adults and is
the most common cause of sudden death
in young healthy individuals.
• So far mutations in over 20 genes have
been associated with FHC
Primary Clinical Features of FHC
• Left ventricular hypertrophy, “a thickening of the tissue
due to increased size of the constituent cells”.
• Myocyte/myofibrillar disarray caused by the abnormal
shapes, intracellular connections and arrangement of the
hypertrophic myocytes and fibrosis.
1 http://www.maxshouse.com
2 Arad et al 2002 Hum Mol Genet. 11. (20) 2499-2506
Genotype Phenotype Correlation
The Heterogeneous Nature of
FHC
• HCM is caused by dominant mutations in the sarcomeric
genes.
• de novo mutations occur rarely and account for
approximately 10% of cases.
• Mutations in the sarcomeric genes account for ~55% of
cases of HCM.
• Syndromes such as the Glycogen storage disorders and
Friedreich ataxia can mimic HCM.
Glasgow Linkage Exclusion
Analysis Method (GLEAM)
• Glasgow Linkage Exclusion Analysis Method
(GLEAM)
• Novel method to prioritise gene analysis in
heterogeneous disorders
• A gene is excluded from analysis when affected
relatives are oppositely homozygous for SNPs in
and around the gene of interest
GLEAM
I:1
I:2
II:1
II:2
II:3
BB
AB
AA
III:1
BB
III:2
AB
II:4
AB
III:3
III:4
AB
AA
• A and B represent alleles at a susceptibility locus for a
dominantly inherited disorder affecting individuals II:2,
II:3, III:1 and III4.
• Since III:1 has no allele in common with II:3 or III:4 it
effectively rules out this locus as being responsible for
the disease in this family.
Genes analysed in the FHC Project
Gene Name
Chromosome
No Exons
No SNPs
TTN
2
363
212
MYH7
14
38
76
MYH6
14
37
77
MYBPC3
11
35
147
RAF1
3
16
163
PRKAG2
7
16
146
TPM1
15
16
132
TNNT2
1
15
116
MYLK2
20
12
94
TNNI3
19
8
92
MYL3
3
7
89
MYL2
12
7
94
CAV3
3
2
98
SNP Analysis Platform
Sentrix Array Matrix
• 96 fibre optic bundles on each plate
• Each fibre contains a bead that corresponds to
each SNP
• Image taken from www.Illumina.com
Results- Raw Data
Raw data for one patient sample
Results- Raw Data
AA
AB
BB
Clustered patient SNP data for a single SNP locus
Results- Genotype Comparisons
Results
Relationship
Number of pairs
Average number
of genes excluded
Sibs
Aunt/Niece
Nephew
49
22
3
5
First Cousins
First Cousins once
removed
10
5
7
8
Second Cousins
Grandparent/
Grandchild
9
1
7
2
Results
Gene
Number of times
gene excluded
Percentage
number of times
gene excluded
TTN
34/96
35%
MYH6 and MYH7
33/96
34%
MYBPC3
28/96
29%
RAF1
23/96
24%
PRKAG2
47/96
49%
TPM1
29/96
30%
TNNT2
39/96
41%
MYLK2
40/96
42%
TNNI3
34/96
35%
MYL3
33/96
34%
MYL2
26/96
27%
CAV3
40/96
42%
Interesting Case
I:1
II:1
III:1
IV:1
IV:2
H15.1 H15.4
I:2
II:2
II:3
III:2
III:3
III:4
IV:3
IV:4
IV:5
H15.7
TNNI3/
TNNI3/ MYBPC3
MYBPC3 MYBPC3
H15.14
TNNI3
H15.15
TNNI3
II:4
III:5
III:6
III:7
IV:6
H15.16
TNNI3
H15.12
TNNI3
Interesting Case
• Familial mutation in TNNI3 was not excluded in all
affected family members.
• Comparisons between H15.1, H15.4 and H15.7 did not
exclude MYBPC3.
• MYBPC3 was excluded when H15.1, H15.4 and H15.7
were compared against other family members who did
not have this mutation.
• Testing for the TNNI3 mutation in H15.7 would have
been negative and suggested a second mutation
prompting further analysis.
Conclusions
• For all the pedigrees with one known mutation, this gene
was not excluded in any of the analyses performed.
• More genes tend to be excluded when more distantly
related individuals such as first cousins or aunt/niece,
nephew pairs are considered, rather than more closely
related sibs
• GLEAM can be used to determine the order in which
genes are sequenced in heterogeneous disorders
Acknowledgements
• Scottish Health Innovations Ltd
• Dr Wai Lee & Dr Stewart Lang, British Heart
Foundation, Glasgow Cardiovascular Research
Centre, University of Glasgow.
 Dr Petros Syrris, Department of Medicine,
University College London