Diagnostic Testing for Familial Pancreatitis

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Diagnostic Testing for Familial Pancreatitis

• Emma McCarthy
• Clinical Scientist
• Merseyside Cheshire
• Regional Molecular Genetics Laboratory

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Aims

• Overview of pancreatitis both hereditary and
  idiopathic
• Genes involved diagnostic service
• PRSS1 dosage analysis

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Pancreatitis

• Inflammatory disease
• Severe abdominal pain
• Nausea
• Vomiting fever
• Acute or chronic
• Premature activation digestive enzymes
• Common triggers
• Excessive alcohol consumption
• Smoking
• Gallstones

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• TRYPSINOGEN
• TRYPSIN
• AUTODIGESTION
• CHRONIC PANCREATITIS
• PERMANENT LOSS OF FUNCTION
• DIABETES / PANCREATIC CANCER

activation
premature activation enzymes
multiple attacks
irreversible scarring
advanced stages
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Reproduced from Rosendahl et al. Orphanet
Journal of Rare Diseases 2007 21
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Diagnostic service

• PRSS1
• Fluorescent sequencing exons 2 3
• SPINK1
• p.N34S mutation (pyrosequencing)
• CFTR
• 28 mutations (Elucigene CFHT)
• EUROPAC www.liv.ac.uk/surgery/europac.html

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Hereditary Pancreatitis (HP)

• Autosomal dominant
• Reduced penetrance
• Childhood
• recurrent episodes acute pancreatitis
• Adulthood
• chronic pancreatitis
• Mutations of cationic trypsinogen gene (PRSS1)

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Cationic Trypsinogen

• PRSS1 gene (protease serine 1)
• Trypsin
• Activates digestive enzymes
• Self regulating
• Autocatalytic activation
• Feedback inhibition - cleaves at Arg122
• R122H mutation - super trypsin

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N29I
L41L
C48C
p.N29I c.86AgtT
p.L41L c.121CgtT
p.C48C c.144TgtC
p.N54S c.161AgtG
p.V59V c.177AgtG
p.G62G c.186CgtT
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Laboratory Data

• Prior 2007
• PCR / restriction digest - p.A16V, p.N29I,
  p.R122H
• Feb 2007
• fluorescent sequencing exons 2 3
• Pathogenic mutations detected - 7/77 (9.1)
• 2/77 unclassified variants
• Total sequence changes identified - 9/77 (11.7)

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Idiopathic Chronic Pancreatitis (ICP)

• 30 pancreatitis cases - unknown cause
• Susceptibility genes
• SPINK1 - Serine Protease Inhibitor Kazal type 1
• CFTR mild mutations ICP specific mutations?
• ?polygenic model

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SPINK1

• Inhibits 20 pancreatic trypsin
• p.N34S mutation
• 22 ICP patients
• 2.5 general population
• ?disease modifier

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SPINK1 Analysis

• Pyrosequencing assay
• c.101AgtG p.N34S
• 9/60 mutation carriers
• 0/8 CFTR mutations
• no CF testing on 1 positive case

Normal
2 copies N34S
1 copy N34S
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PRSS1 Dosage Analysis

• Triplication of trypsinogen locus reported
• Le Maréchal et al., Nature Genetics 2006
  381372-4
• Fluorescent dosage analysis PRSS1 exon 1
• 40 families (EUROPAC)
• Duplication in mother daughter

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Normal Sample
Proband
PRSS2 exon 4
Normal Sample
Proband
PRSS1
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Pedigree
70
78
84
DUP
Awaiting sample
nmd
nmd
72
44
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DUP
DUP
21
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nmd
DUP
DUP
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Acknowledgements

• Roger Mountford
• Victoria Blake
• DNA Laboratory Staff

• Chris Grocock
• Bill Greenhalf
• John P. Neoptolemos