diagnostic testing for familial pancreatitis emma mccarthy clinical scientist merseyside &...
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Diagnostic Testing for Familial Pancreatitis
Emma McCarthyClinical Scientist
Merseyside & Cheshire Regional Molecular Genetics Laboratory
Aims
• Overview of pancreatitis both hereditary and idiopathic
• Genes involved & diagnostic service
• PRSS1 dosage analysis
Pancreatitis
• Inflammatory disease – Severe abdominal pain– Nausea– Vomiting fever– Acute or chronic
• Premature activation digestive enzymes
• Common triggers– Excessive alcohol consumption– Smoking– Gallstones
TRYPSINOGEN
TRYPSIN
AUTODIGESTION
CHRONIC PANCREATITIS
PERMANENT LOSS OF FUNCTION
DIABETES / PANCREATIC CANCER
multiple attacks
premature activation enzymes
activation
irreversible scarring
advanced stages
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
Hereditary Pancreatitis (HP)
• Autosomal dominant
• Reduced penetrance
• Childhood:– recurrent episodes acute pancreatitis
• Adulthood:– chronic pancreatitis
• Mutations of cationic trypsinogen gene (PRSS1)
Cationic Trypsinogen
• PRSS1 gene (protease serine 1)
• Trypsin– Activates digestive enzymes– Self regulating
• Autocatalytic activation• Feedback inhibition - cleaves at Arg122
• R122H mutation - “super trypsin”
N29I L41L C48C
p.N29I c.86A>T
p.C48C c.144T>C
p.L41L c.121C>T
p.N54S c.161A>G
p.V59V c.177A>G
p.G62G c.186C>T
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%)
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
SPINK1
• Inhibits 20% pancreatic trypsin
• p.N34S mutation– 22% ICP patients– 2.5% general population– ?disease modifier
SPINK1 Analysis
• Pyrosequencing assay– c.101A>G; p.N34S
• 9/60 mutation carriers– 0/8 CFTR mutations
• no CF testing on 1 positive case
Normal
2 copies N34S
1 copy N34S
PRSS1 Dosage Analysis
• Triplication of trypsinogen locus reported– Le Maréchal et al., Nature Genetics 2006; 38:1372-4
• Fluorescent dosage analysis PRSS1 exon 1
• 40 families (EUROPAC)
• Duplication in mother & daughter