1. Whitcomb 2000
Pancreatic Diseases:
Insights from Hereditary
Pancreatitis
Written byWritten by
David C Whitcomb MD PhDDavid C Whitcomb MD PhD
Department of Medicine, UniversityDepartment of Medicine, University ofof PittsburghPittsburgh
Educational Use ONLY
2. Whitcomb 2000
Hereditary Pancreatitis
Hereditary pancreatitisHereditary pancreatitis (HP) is an unusual form of(HP) is an unusual form of
acute and chronic pancreatitis that runs in families.acute and chronic pancreatitis that runs in families.
The risk of pancreatic cancer is >50 times normal.The risk of pancreatic cancer is >50 times normal.
Although HP is only responsible for 2-3% of allAlthough HP is only responsible for 2-3% of all
cases of chronic pancreatitis, study of this diseasecases of chronic pancreatitis, study of this disease
has revolutionized our understanding of pancreatichas revolutionized our understanding of pancreatic
diseasesdiseases
PancreasPancreas
3. Whitcomb 2000
Discovery of the Pancreatitis Gene
ccaccaccagtcaggcac
actctaccaccATGAA
TCCACTCCTGAT
CCTTACCTTTGT
GG/ACAGCTGC
TCgtgagtatcatgccct
gcctcaggccccaaccac
ccccccgttcctggccgaDNA
1. Family 2. Genetic Mapping 3. Mutation 4. Mechanism
Chromosome 7
Hereditary
Pancreatitis
gene
Functional
significance
determined
Mutation in the
trypsinogen
DNA sequenceRecruitment
Trypsinogen
7. Whitcomb 2000
HP is caused by “Super-Trypsin”
Whitcomb et al,Whitcomb et al, Nature GeneticsNature Genetics 19961996
Trypsin
Active trypsin in the pancreas
will cause the pancreas to digest
itself.
Normally, the pancreas is
protected because active trypsin
will destroy itself by cutting at
R117. This will split the trypsin
and inactivate it.
In HP, R117 is mutated to H117.
This creates a “super-trypsin”
that cannot be inactivated and
leads to acute pancreatitis.
8. Whitcomb 2000
Conclusions
Discovery of theDiscovery of the hereditary pancreatitishereditary pancreatitis
gene is an example of how genetic researchgene is an example of how genetic research
can revolutionize our understanding of acan revolutionize our understanding of a
disease process.disease process.
9. Whitcomb 2000
Pancreatitis
PancreasPancreas -- an organ that makesan organ that makes bicarbonatebicarbonate to nutralize gastric acid,to nutralize gastric acid,
enzymesenzymes to digest the contents of a meal andto digest the contents of a meal and insulininsulin to signal the body toto signal the body to
store ingested nutrients.store ingested nutrients.
Acute PancreatitisAcute Pancreatitis - An acute, potentially life-threatening condition- An acute, potentially life-threatening condition
presenting with severe abdominal pain in which the pancreas appears topresenting with severe abdominal pain in which the pancreas appears to
digest itself. It is usually caused by gallstones, alcohol or is idiopathic.digest itself. It is usually caused by gallstones, alcohol or is idiopathic.
Chronic PancreatitisChronic Pancreatitis - an- an irreversible scarring of the pancreas withirreversible scarring of the pancreas with
permanent loss of pancreatic function that typically causes unrelentingpermanent loss of pancreatic function that typically causes unrelenting
abdominal pain.abdominal pain.
Hereditary PancreatitisHereditary Pancreatitis - a unusual form of acute and chronic pancreatitis- a unusual form of acute and chronic pancreatitis
that runs in families. The risk of pancreatic cancer is >50 times normal.that runs in families. The risk of pancreatic cancer is >50 times normal.
PancreasPancreas
12. Whitcomb 2000
Implications of the HP Discovery
Acute pancreatitisAcute pancreatitis starts with trypsin activation within thestarts with trypsin activation within the
pancreatic acinar cells -pancreatic acinar cells - not from digestive enzymes leakingnot from digestive enzymes leaking
from injured pancreatic ducts!from injured pancreatic ducts!
Chronic pancreatitisChronic pancreatitis develops from multiple attacks ofdevelops from multiple attacks of
acute pancreatitis -acute pancreatitis - not from duct obstruction by calcium.not from duct obstruction by calcium.
Pancreatic CancerPancreatic Cancer - occurs from chronic inflammation of- occurs from chronic inflammation of
the pancreas -the pancreas - not from an oncogenenot from an oncogene
TherapyTherapy should be directed at preventing excessive,should be directed at preventing excessive,
premature trypsin activation.premature trypsin activation.
13. Whitcomb 2000
** WARNING **
Educational Use Only
No portion of this presentation is
to be used without the express
written consent of Dr Whitcomb
Editor's Notes
Title slide. The talk is based on the following work : Whitcomb, DC, RA Preston, CE. Aston, MJ Sossenheimer, PS Barua, Y Zhang, A Wong-Chong, GJ White, PG. Wood, L Gates, C Ulrich, SP. Martin, JC Post and GD. Ehrlich, A Gene for Hereditary Pancreatitis Maps to an 19 cM Locus on Chromosome 7q35. Gastroenterology 110:1975-1980, 1996 Whitcomb, DC, MC Gorry, RA Preston, W Furey, MJ Sossenheimer, CD Ulrich, SP Martin, LK Gates, ST Amann, PP Toskes, R Liddle, K McGrath, G. Uomo, JC Post and GD Ehrlich. Hereditary pancreatitis is caused by a mutation in the cationic trypsinogen gene. Nature Genetics, 13(6): 141-145, 1996 Lowenfels, AB, P. Maisonneuve, EP DiMagno, Y Elitsur, LK Gates, J Perrault, DC Whitcomb. Hereditary pancreatitis and the risk of pancreatic cancer. J.Nat. Cancer Inst. 89 (6):442-446, 1997 Sosssenheimer, MJ, C Aston, GD Ehrlich, L Gates, S Martin, C Ulrich, R Preston, DC Whitcomb. Clinical characteristics of hereditary pancreatitis in a large family based on high-risk haplotype. Am. J. Gastro. 92(7):1113-1116, 1997 Gorry, MC, D Gabbaizedeh, W Furey, LK Gates Jr, RA Preston, CE Aston, Y Zhang, C Ulrich, GD Ehrlich, and DC Whitcomb, Mutations in the cationic trypsinogen gene are associated with recurrent acute and chronic pancreatitis. Gastroenterology. 113:1063-1068, 1997.
The major steps to accomplish a genetic linkage study are listed (This is also an outline of the presentation). First, a family must be identified with a well characterized disease. Second, a careful survey of the 23 human chromosomes IN EACH FAMILY MEMBER is made to identify the one segment of DNA that is ALWAYS inheredted with the disease. This identifies a single region of a chromosome that contains the affective gene. Third, the genes in the segment of DNA identified in step 2 are sequenced in order to identify the disease-causing mutation. Fourth, once the mutation is identified, then the genetic code is solved to reveal the normal protein and abnormal protein caused by the mutation. Scientist and physicians can then determine how the mutation caused the inherited disease.
This slide represents a partial pedigree of the S-family used to map the hereditary pancreatitis gene. This portion covers seven generations and individuals were chosen to maximize the mapping process. Those family members who had characteristic pancreatitis are shown in the symbols filled in white. Clinically unaffected individuals who carried the hereditary pancreatitis disease gene are shown in light blue. For the entire family the 80% of the individuals with the gene had clinical symptoms as reported by others. The entire family tree had over 500 individuals
To find the chromosome region containing the affective gene, microsatellite markers from a number of chromosomes were sequentially tested for linkage with the disease gene. This led to the identification of the hereditary pancreatitis locus on chromosome 7. This figure illustrates the LOD score plot of microsatellite markers along chromosome 7. This is a statistic plot that proved that this region of DNA was inherited through the family tree with the hereditary pancreatitis trait. The hereditary pancreatitis gene was on the same chromosome as the cystic fibrosis gene, and at the exact location of the T-cell receptor beta chain gene.
The T-cell beta chain gene was sequenced by a group at the University of Washington and was over 685,000 base pairs in length. Within this gene were 8 trypsin-like genes. One of the trypsinogen genes had the hereditary pancreatitis mutation in it. (red letter is an “A” in the mutated gene).
The trypsin molecule is illustrated in yellow and blue and is shaped like a pac-man with the jaws opening to the left. If a protein chain with an arginine or lysine inters the jaw it is rapidly cut into two peices. Tryspinogen is made by the pancreas in an inactive form. It is one of the most important enzymes because one it is activated to trypsin in the intestine, it is responsible for activating nearly all of the other digestive enzymes to their active form, in the intestine . If trypsinogen becomes active in the pancreas it could activate all of the other digestive enzymes and cause the pancreas to digest itself. Forturnately, the pancreas also makes a trypsin inhibitor (PSTI - in red) that plugs the active site and prevents trypsin from activating other trypainogen molecules and all the other enzymes. However, the amount of PSTI is limited, so another protective mechanism is necessary. If too much trypsin is active, trypsin begins attacking itself at arginine #117 (R117). Cutting the connecting chain splits the trypsinogen molecule in half, permanently inactivating it, and saving the pancreas from auto-digestion. Solving the genetic code for hereditary pancreatits proved that the mutation was at R117 changing it to Histidine 117.
The pancreas has 3 main functions. I makes: Bicarbonate to nutralize gastric acid as the meal enters the small intestine, Digestive enzymes - enough to turn an intire meal to liquid so that it can be absobed, and Insulin. Acute pancreatitis can be defined clinically as an acute condition typically presenting with abdominal pain and is usually associated with elevated pancreatic enzymes in blood or urine, owing to inflammatory disease of the pancreas . Chronic pancreatitis can be defined as a continuing inflammatory disease of the pancreas characterized by irreversible morphologic change and typically causing pain and/or permanent impairment of function. Clinically, about 80% present with recurrent or persistent abdominal pain. In more advanced stages of chronic pancreatits patients develop maldigestion or loss of endocrine function leading to diabetes mellitus. Morphologically, chronic pancreatitis has areas of fibrosis with loss of parenchyma in focal, segmental or diffuse patterns and is usually associated with ductal changes. Hereditary pancreatitis is an unusual form of pancreatitis that runs in families. Affected family members have BOTH acute pancreatitis and chronic pancreatitis that is indistinguishable from other forms of pancreatitis. These family members also have a 50-fold increase risk of pancreatic cancer.
The trypsin molecule is illustrated in yellow and blue and is shaped like a pac-man with the jaws opening to the left. If a protein chain with an arginine or lysine inters the jaw it is rapidly cut into two peices. Tryspinogen is made by the pancreas in an inactive form. It is one of the most important enzymes because one it is activated to trypsin in the intestine, it is responsible for activating nearly all of the other digestive enzymes to their active form, in the intestine . If trypsinogen becomes active in the pancreas it could activate all of the other digestive enzymes and cause the pancreas to digest itself. Forturnately, the pancreas also makes a trypsin inhibitor (PSTI - in red) that plugs the active site and prevents trypsin from activating other trypainogen molecules and all the other enzymes. However, the amount of PSTI is limited, so another protective mechanism is necessary. If too much trypsin is active, trypsin begins attacking itself at arginine #117 (R117). Cutting the connecting chain splits the trypsinogen molecule in half, permanently inactivating it, and saving the pancreas from auto-digestion. Solving the genetic code for hereditary pancreatits proved that the mutation was at R117 changing it to Histidine 117.
Protective mechanism against premature trypsin activation. Since active trypsin has the potential of driving the pancreatic digestive enzyme activation cascade, a number of mechanism are employed to protect the pancreas and body from autodigestion. (1) The first defense is the synthesis of all digestive enzymes as inactive proenzymes, except for amylase and lipase. (2) Since trypsin activation is calcium sensitive, the intracellular calcium levels are kept low. (3) To prevent activation of the zymogens by lysosomal enzymes, the zymogens are separated in a different intracellular compartment. The diagram shows the rough endoplasmic reticulum (RER) where proenzymes and other acinar cell proteins are synthesized, followed by transport to the golgi apparatus (GA) for sorting and packaging in condensing vacuoles (CV) which condense to form the zymogen granules (G). (4) If trace levels of trypsin activation occur in the zymogen granules, then trypsin is inhibited by pancreatic secretory trypsin inhibitor (PSTI) which is a protein synthesized along with trypsinogen and can inactivate up to 20% of potential trypsin. (5) If excessive levels of trypsin activation occur in the zymogen granules there is a mechanism for trypsin to autodigests itself. (6) The normal activation of trypsin is removed from the pancreas to the intestine where enterokinase is located. (7) The serum proteins alpha-1-antitrypsin and beta-2-microglobulin inactivate any trypsin that escapes the pancreatic acinar cells and limits damage from zymogens.
