Somatic mutations of the MEN1 tumor suppressor gene in sporadic gastrinomas and insulinomas.

Abstract

Gastrinomas and insulinomas are frequent in multiple endocrine neoplasia type 1 (MEN1). The MEN1 tumor suppressor gene was recently identified. To elucidate the etiological role of the MEN1 gene in sporadic enteropancreatic endocrine tumorigenesis, we analyzed tumors (28 gastrinomas and 12 insulinomas) from 40 patients for MEN1 gene mutations and allelic deletions. One copy of the MEN1 gene was found to be deleted in 25 of 27 (93%) sporadic gastrinomas and in 6 of 12 (50%) sporadic insulinomas. MEN1 gene mutations were identified in 9 of 27 (33%) sporadic gastrinomas and 2 of 12 (17%) insulinomas and were not seen in corresponding germ-line DNA sequence. A specific MEN1 mutation was detected in one gastrinoma and in the corresponding germ-line DNA of a patient who had no family history of MEN1. Somatic MEN1 gene mutations and deletions play a critical role in the tumorigenesis of sporadic gastrinomas and may also contribute to the development of a subgroup of insulinomas.

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1997;57:4682-4686. Published online November 1, 1997.Cancer Res


Zhengping Zhuang, Alexander O. Vortmeyer, Svetlana Pack, et al.
Sporadic Gastrinomas and Insulinomas
Tumor Suppressor Gene in
MEN1
Somatic Mutations of the



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(CANCERRESEARCH57. 4682-4686. November 1, 19971
Advances in Brief
Somatic Mutations of the MEN1 Tumor Suppressor Gene in Sporadic Gastrinomas
and Insulinomas
Zhengping
Won-Sang Park, Sunita K. Agarwal, Larisa V. Debelenko,MaryBeth Kester, Siradanahalli C. Guru,
PachiappanManickam, Shodimu-EmmanuelOlufemi, Fang Yu, Christina Heppner, Judy S. Crabtree,
Monica C. Skarulis,David J. Venzon, MichaelR. Emmert-Buck,
FrancisS. Collins,A. Lee Burns,StephenJ. Marx,
Zhuang, Alexander0. Vortmeyer,Svetlana Pack, Steve Huang, Thu A. Pham, Chaoyu Wang,
Allen M. Spiegel, Settara
T. Jensen,Lance
C. Chandrasekharappa,
IrmaA. Lubensky'Robert A Liotta,and
Laboratory
National
Research Institute (S. C. G., P. M., S-E. 0., J. S. C., S. C. C., F. S. C.); Digestive Diseases (F. Y., R. T. J.] and Diabetes Branches (M. C. S.], National institute of Diabetes and
Digestive and Kidney Diseases; and Biostatistics and Data Management Section, National Cancer Institute (D. J. V.1. NiH, Bethesda, Maryland 20892
of Pathology.
Institute of Diabetes
National Cancer Institute (1 1,
and Digestive
A. 0. V., S. P.. S. H., T. A. P., C. W, W-S. P., L V. D., M. R. E-B., L A. L, I. A. LI:
and Kidney Diseases(S. K. A., M. K., C. H., A. M. S., A. L B., S. J. Mi;
Metabolic Diseases
Human Genome
Branch,
Laboratory of Gene Transfer, National
Abstract
Gastrinomas and insulinomasare frequent in multipleendocrineneo
plasia type 1 (MEN1). The MEN1 tumor suppressor gene was recently
identified. To elucidate the etiological role of the MEN1 gene in sporadic
enteropancreaticendocrine tumorigenesis, we analyzed tumors (28 gas
trinomas and 12 insulinomas)from 40 patIents
andallelicdeletions.OnecopyoftheMENIgenewasfoundtobedeleted
in 25 of 27 (93%) sporadic gastrinomas and in 6 of 12 (50%) sporadic
insulinomas.MENJgene mutationswere identifiedin 9 of 27 (33%)
sporadicgastrinomasand 2 of 12 (17%) insulinomas
corresponding germ-line DNA sequence. A specific MENJ mutation was
detectedin one gastrinomaand in the corresponding
patientwho had no family historyof MEN!.
tions and deletions play a critical role in the tumorigenesis of sporadic
gastrinomas and may also contribute to the development of a subgroup of
insulinomas.
for MEN1 gene mutations
and were not seen in
germ-line
MENJ
DNA of a
gene muteSomatic
Introduction
Gastrinomas and insulinomas occur sporadically (1) and as a part of
an autosomaldominantsyndrome,
follow a benign clinical course and are cured by resection
tumor. In contrast, gastrinomashave high malignant potential, with
regional lymph node or liver metastases developing in up to 90% of
the cases, and are curable in the long term in only 30% of the cases.
The role of tumor suppressor genes/oncogenes
endocrine tumorigenesisremains uncertain (3, 4). The Knudson tumor
suppressor gene hypothesis states that hereditary cancers develop due
to the inheritance of a mutated tumor suppressor gene and that a
somatic mutational event involving the wild-type allele of the gene
leads to a neoplasm (5). The model predicts that sporadic tumors of
the same cell type would arise after somatic inactivation
copies of the gene responsible for the familial tumor (5). The MENJ
gene was linked to chromosome11q13 in 1988 (6). Previous studies
demonstratedLOH with polymorphic
gastrinomasandinsulinomas,suggesting
play a role in sporadic enteropancreatic endocrine tumorigenesis
(3, 7).
MEN12(2). Insulinomas usually
of the
in enteropancreatic
of both
markers on llql3
that
in sporadic
genethe MENJmay
Received 7/31/97; accepted 9/22/97.
The costs of publication of this article were defrayed in part by the payment of page
charges. This article must therefore be hereby marked advertisement
18 U.S.C. Section 1734 solely to indicate this fact.
in accordance with
I To whomrequestsfor reprints shouldbe addressed,at Laboratoryof Pathology,
National Cancer Institute, NIH, Building
MD20892.Phone:(301)496-0549; Fax:(301)402-0043.
2Theabbreviations usedare:MENI, multipleendocrineneoplasia type 1;LOH, loss
of heterozygosity;ZES, Zollinger-Ellison syndrome; P'FH, parathyroid hormone; SSCP,
single-strand conformation polymorphism; FISH, fluorescence in situ hybridization.
10, Room 2N212, 10 Center Drive, Bethesda,
The MENJ tumor suppressor gene was recently identified (8), and
germ-line mutations in the gene were detected in 47 of 50 kindreds
with familial MEN1 (9). To evaluate whether sporadic gastrinomas
and insulinomas are associatedwith functional
MENJ gene, the gene was analyzed for deletions and mutations in 40
sporadic enteropancreatictumors.
inactivation of the
Patients and Methods
Subjects. Tumors from 40 patients who underwentexploratory laparotomy
for pancreatic or duodenal endocrine tumors at the NIH were included in the
study. All patients gave informedconsent
NIDDKInstitutionalReview Board.Clinical andfamily history werereviewed
in each case, and all 40 patients were thought to have a sporadic neoplasm at
the time of surgery. The diagnosis of insulinoma required a fasting blood sugar
of less than 45 mg/dl, hypoglycemia symptoms,
plasma insulin level of >5 microunits/mI(10). The diagnosis of ZES (gastri
noma) was established as described previously (11) and required meeting at
least two of the following criteria: an elevated fasting serum gastrin level; a
basal acid output of more than 15 mEq/h if the patient had not had previous
gastric surgery or of more than 5 mEq/h if the patient had had previous gastric
surgery; and an increasein the serum gastrin level of 200 pg/mI after i.v.
secretin, an increase in serum gastrin level of 395 pg/mI after an iv. calcium
infusion, or a positive histologicaldiagnosis
A diagnosis of sporadic insulinoma was made by an absence of a family
history of endocrinopathies associated with MEN1 (renal stones, hyperpara
thyroidism,pituitary disease,or pancreatic
serum calciumand albuminmeasurements.
prolactin and gastrin measurements and family screening were done in some
cases. A diagnosis of sporadic gastrinoma required a negative family history
(as described above)and normal values
calcium, albumin, plasma PTH level (mid-PTH portion), plasma PTH (intact
PTH), serum prolactin, urinary cortisol, and normal computed tomography
magnetic resonance imaging of the sella turcica (12).
Tumors. Twenty-eightfrozengastrinomas (4 pancreatic, 8duodenal, and 1
common bile duct) and metastases (14 lymph node and 1liver) and 12benign
pancreatic insulinomas were obtained from the file of the Laboratory of
Pathology, National Cancer Institute, NIH (Bethesda, MD). All tumors were
frozen in liquid nitrogen at the time of surgery and stored at —70°C.
normal DNA was obtained from frozen normal exocrine pancreas, duodenum,
lymph node, or blood of each patient. Histological evaluation of tumors
revealed characteristic neuroendocrine features and positive staining for chro
mogranin A (Boehringer Mannheim, Indianapolis, IN) and/or synaptophysin
(Zymed,SanDiego,CA)byimmunohistochemistry. Theclinicaldiagnosesof
gastrmnoma and insulinoma were confirmed by positive immunostain for gas
tin (DAKO Corp., Carpenteria,CA) and insulin (BioGenex, San Ramon, CA),
respectively.
Tumor and normal cells were selected from 5-sm-thick H&E-stained his
tological slides, microdissected under direct light microscope visualization as
in a protocol approvedby the
and a simultaneouselevated
of gastrinoma.
endocrine
If results were unclear,
tumors)and normal
serum
for total serum calcium, ionized
or
Paired
4682
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Table 1 PCR primer sets usedfor MENI gene mutationanalysisExonSize
Annealing
temperaturePrimer sequence(bp)(‘C)2k'
2W'
2C―
3
4
5
6
7
8
9
bA―
lOB―
lOC―
a Seve5'-ttgccttgcaggccgccgcc-3'
ml primer sets were used to amplify exons 2 and 10due to their large size.
201 60
5'-tggtagggatgacgcggttg-
5'-ggcttcgtggagcattttct-3'
5,-ctcgagga tagagggacagg- 3'
5'-ttcaccgcccagatccgagg--3'
5'-taagattcccacctactggg-3'
5'-attacctcccccttccacag-3'
5,-c tggggggagggaacaa
5'-cataatgatctcatcccccc-3'
5'-attggctcagccctcacctg-3'
5'-gttccgtggctcataactct-3'
5'- tggccacttccctctactga-
5'-ggcagcctgaattatgatcc-3'
5'-ttctgcaccctccttagatg-3'
5'-ggactccttgggatcttcctgtg-3'
5'-atcctcactcctggatgacagtg-3'
5'-cagagaccccactgctctca-3'
5' -ggctggagctccagcctttc-
5'-ctgctaaggggtgagtaagagac-3'
5'-gtc tgacaagcccgtggctgctg-
5'-tcaccttgctctccccactg-3'
5‘-ccaggcccttgtccagtgct-3'
5'-ccaagaagccagcactggac-3'
5'-cactctggaaagtgagcact-
5'-ctgaaggtggcagcacggct-3'
5'-gtagtcactaggggtggaca-
3'
202 60
29758
24955
tac -3'
171
98
58
58
3'
142 58
18360
181 65
3'
225 60
3'
189 60
183 62
3'
219 60
3'
MEN!ALTERATIONSIN SPORADIC GASTRINOMAAND INSULINOMA
described
study, dissections were not performed to obtain pure populations of tumor cells
but to enrich tumor samples sufficiently to perform mutational analysis by
SSCP. Procured cells were resuspended in 30 jxlsolution containing Tris-HCI
(pH 8.0), 0.1MEDTA (pH 8.0), 1%Tween 20, and 0.1 mg/ml proteinase K
and incubated overnight at 37°C.Following thermal inactivation of proteinase
K (95°C for 5 mm), 1—1.5 @.d of the DNA extract were used for PCR
amplification.
MENJ Allelic Deletion Analysis. FISHwasperformedusingcosmidclone
clOBll(size, 40 kb) containing the MENJ gene as a probe (15). Touch
preparationsof frozentumor specimenswere made on glass slidesand used for
FISH analysis as described previously
scored usinga Zeiss Axiophotepifluorescence
images were captured on a Photometrics charge-coupled device camera (Pho
tometrics, Ltd., Tucson, AZ) using IP Lab image software (Signal Analytics
Corporation, Vienna, VA). At least 100interphases with strong hybridization
signals were scored for each tumor. The presence
signal at a frequency of more than 30% was interpreted as an allelic deletion
(loss of one copy of the MENJ gene). Seventy % of the cases with allelic
deletion detectedby FISH showed,upon touch preparation,
MEN1 signal at a frequency of more than 70%, and 30% of the cases showed
cells with one MEN1 signal at a frequency of 35—70%. The percentage of
tumor and somatic cells in each frozen tissue imprint correlated with percent
age of tumor and somatic cells in the H&E-stainedfrozen tissue sections from
which the touch preparations had been prepared.
(exocrine and endocrine) tissue controls showed cells with one MENI signal
at a frequency of 3%.
In cases with no detectable MEN] allelic deletion by FISH, DNA from
precisely microdissected tumor and normal samples (13) was screened for
LOHusing threepolymorphicmarkerson llql3 as describedpreviously(3,
7). The markersincludedintragenicmarker
5'-flanking region, and the flanking markers PYGMand D11S449 (18, 19).
MENJ Mutation Analysis by SSCP and Sequencing. PCR-SSCPanaly
sit was performed using 13 primer sets (Table I) with intronic sequences
designed to amplify exons 2—10 from tumor and normal DNA. After detection
of a mutant allele, DNA was extracted from the excised aberrant band of the
SSCP gel by overnight incubationin 100 @xl of distilled water at room
temperature and subsequentlyreamplified by PCR (14, 20).The PCR products
were directly sequenced (Cyclin Sequencing kit, Perkin-Elmer), and normal
and tumor DNA sequences were compared. Tumor and peripheral blood DNA
from familial MENI patients with known germ-line mutations (8) in each exon
were run in parallel with each SSCP assay gel as positive controls.
previously (13), and used for PCR-SSCPanalysis(14). For this
(16, 17). Hybridization
microscope,
signals were
andtwo-color
of cells with one MENI
cells with one
Normal frozen pancreatic
D11S4946,located in the MEN]
Results
Gastrinomas
age
patients (7 males and 5 females; mean age — 47 years, range = I7—81
years)were examinedfor allelic deletions
gene.
One copy of the MEN] gene was found to be deleted in 25 of 27
(93%) sporadic gastrinomas and in 6 of 12 (50%) sporadic insulino
mas as determined by FISH using a probe containing the MEN! gene
(Fig. 1). MEN] gene somatic mutations were identified in 33% (9 of
27) of sporadic gastrinomas and 17% (2 of I2) of sporadic insulino
mas analyzed(Table 2 and Fig. 2). All but two of the tumors
mutations also showed MEN] gene deletion by FISH, indicating that
both copies of the gene were inactivated (Table 2). The absence of
allelic deletion in tumors G-1236 and In- 1257 was confirmed by LOH
analysisusingan intragenicmicrosatellite
MEN! 5' region and two flanking
D!!S449 (data not shown). No mutation in the MEN!
detectedby SSCP analysis in any exons from 17 of 27 sporadic
gastrinomas and 5 of 12 insulinomas
Observed MEN!gene mutationswere limited
not detected in germ-line DNA in any case, except for one patient who
showed an exon 8 mutation in both tumor (0-1229)
DNA (Table 2). This patient had no family history of MEN I. Preop
eratively, this patient'sMEN1 status was unclear because his serum
calcium, prolactin, and (mid-PTH portion) PTH levels were normal;
however, he had a slightly elevated intact PTH level and a history of
neck surgery for an unknown reason 15 years prior to admission.
Mutations observed among enteropancreatic
the study included four deletions, two insertions, and five missense
mutations (Table 2 and Fig. 2). No tumor was found to have more than
one MEN] gene mutation. Mutations were identified in exons 2, 3, 4,
8, and 10 (Table 2). Five of nine (56%) somatic mutations in sporadic
gastrmnomas occurred in exon 2. Two insulinomas showed mutations,
one in exon 2 and one in exon 10. Deletions or insertions resulting in
a frameshift of the protein coding sequence accounted for 55% (6 of
11) of the mutations. These mutations
resulting in protein truncation with predicted loss of function. The five
missensemutations resultedin amino
not polymorphismsbecause matched normal DNA from the same
individuals showed only wild-type MEN] sequence (Fig. 2C), and the
missensemutationswere not seen in an analysis
chromosomes(8). Characterization
nm will be necessaryto define the significance
missense mutations. MEN! mutations and deletions were detected in
both localized (duodenal and pancreatic) gastrinomas
(lymph node) gastrmnomas examined (Table 2).
We identified three benign polymorphisms
D418D (GAC/GAT), and A541T (GCA/ACA)
respectively, in threesporadic insulinoma
phisms wereidentical to thethree
observed previously (8, 9).
from 28 patients (15 males and I3 females;
50 years, range = I5—7 1 years) and insulinomas
mean
from 12
and mutationsof the MEN]
with
marker(Dl !S4946)
markers,
at the
and llql3PYGM
gene was
that showed allelic deletion.
to tumorDNA and were
and germ-line
endocrinetumors in
produce altered transcripts,
acid substitutions.These were
of 142 normal
of the functional domains of me
of the observed
and metastatic
R17 1Q (CGG/CAG),
in exons 3, 9 and 10,
cases. These
common polymorphisms
polymor
most
Discussion
The role of the MEN! gene in sporadic enteropancreatic
tumorigenesis was suggested
7). Our previous work using 10 polymorphic
area of the putative MEN! gene on chromosome
that 44% of sporadic gastrinomas
flanking markers used were PYGM (centromeric)
PPPJCA (telomeric), each located more than 100 kb from the MEN!
gene. Here, we tested for allelic deletions at the precise location of the
endocrine
on I 1q13 (3,previouslyby LOH studies
markers flanking the
I 1q13 demonstrated
showed LOH (3, 7). The closest
and DI !S449 and
4683
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Table 2MEN! gene mutations and allelic deletions in gastrinomasinsulinomasTumorAllelicExonTumorand
code°locationMutationbdeletionc2G-344Duodenum358del4―+2G-58Lymph
nodee358de121+20-187Duodenum186F
TTC)+2G-
I351Pancreas483delAT+2G-l236PancreasW126G
GGG)-2In-I
147Pancreas545insT+3G-40Lymph
TGT)+4G-520Lymph
nodee875insA+8
(ATC —‘
(TGG —@
nodeeF159C(lTr—@
8G-149l G-1229Lymph
+10In-l257PancreasA535V
GTF)—10G-555Lymph
nodee
Lymph nodee+
(OCT —‘
(TCA—‘
nodeeS543L
Tl'A)+
In, insulinoma.
abbreviations
MEN! ALTERATIONSIN SPORADIC GASTRINOMAAND INSULINOMA
A
B
#@,@
,@‘
S
C
0
Fig.I.Representative resultsofFISHinlymphoblast celllineandsporadic enteropancreatic endocrine tumors.Greensignal,a-satellitecentromeric marker;redsignal,chromosome
I1q13probe containing the MEN! gene (cosmid cl0Bl 1).A, the MEN! gene localized to 11q13on metaphasechromosome preparation from a normal lymphoblastcell line by FISH.
B—D, interphase touch preparations of sporadic enteropancreatic endocrine tumors. B, sporadic insulinoma (In-756) showing no deletion of the MEN! gene by FISH. Two red signals,
two alleles of the MEN! gene. C, allelic deletion of the MEN! gene detected in sporadic insulinoma cells (In-l 147:one red signal) as compared to a normal somatic cell with two
red signals (left). D. allelic deletion of the MEN! in sporadic gastrinoma cells (G-l544; one red signal).
gene. The significant
indicates that a substantial subset of gastrinomas may contain limited
interstitialdeletions of a 300 kb or less in size that would not be
evident at flanking markers PYGM, D]!S449,
ingly, the LOH profile observed in gastrinomas differs from that seen
in MEN1-associatedparathyroid
large deletions of several megabases or more (13). The significance,
if any, of the different deletion profiles between the two tumor types
is currently not clear.
The high frequency of MEN] mutations (33%) and allelic deletions
(93%) in sporadic gastrinomasindicates that, analogous to MEN1-
associated tumors, MEN! gene alterations are critical events in spo
radic gastrinoma tumorigenesis. Mutational inactivation of one MEN!
allele coupled with deletion of the second allele strongly implicates
the MEN! gene in the pathogenesis of these tumors, as defined in the
two-hit recessive (loss-of-function)
were detected in localized duodenal and pancreatic gastrinomas
increasein deletionrate, 93% versus 44%,
and PPPJCA.Interest
tumors,which exclusivelyshow
mechanism (5). MEN] mutations
and
a G,gastrinoma;
b Mutation follow standard nomenclature(27).
C Presence(+) or absence(—) ofMEN! allelicdeletion asdetected byFISH.
d Deleted nucleotidesare C'FGT.
C Metastasis.
1Deleted nucleotides are TGTCTA TCATCGCCGC CCTCTATGC.
S Deleted nucleotides areGCCAATG.
4684
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C
G
C
T
A
C
I
A
I
Ca
Ta -@
--@...-.@
@
C
\@
Tumor
GATC
Normal
GATC
Tumor
GATC
Normal
GATC/@
__ ,-. C
@
.@C
.—@-.C
A
C
C
I
C
C
C
@
C.
MEW! ALTERATIONS IN SPORADIC GASTRINOMAAND INSULINOMA
ABC
TN TNTN
E
F
C
A
C
C
C
C
I
ICA
C
C
I
C
C
C
C
D
A
C'@\ GATC
T ‘@
Tumor
TI ‘—#.-.@,
aT
V
G
C
C
Fig. 2. Detection of somatic mutations in the MEN! gene in sporadic enteropancreatic
(A), G-344 (B), and G-555 (C) are shown. Arrows, mutant alleles. The mutations were found in the tumor (Lanes 7) but not in the corresponding normal (Lanes N) tissue. SSCP gels
(A and C) show the presence of both normal and mutated allele in the tumor (Lanes 7) samples due to normal tissue contamination. D—F, corresponding DNA sequence of aberrant
band from SSCP gel.Arrowheads, changes in tumor DNA sequence as comparedto normal tissue DNA.D, mutation545insT in insulinoma In-1147.E, mutation 358del4 ingastrinoma
G-344. F, missense mutationTCA—+TI'A (S543L) in gastrinoma 0-555. In this instance. clear excision of the abnormal band was not possible, so the sequence reveals both the mutant
and wild-type allele.
endocrine tumors. A—C, SSCP changes for three MEN! gene mutations in tumors In- I 147
metastatic (lymph node) gastrmnomas examined (Table 2), suggesting
that they occur at an early stage of sporadic gastrinoma tumorigenesis.
In the presentstudy, we observedan apparentclusteringof muta
tions in exon 2 of the MEN] gene in sporadic gastrinomas. Exon 2 is
the first coding exon in the MEN! gene, and four of the five exon 2
mutations observed in the gastrinomas
in a protein that, even if biologically
wild-type amino acid sequence. Thus, it is possible that tumors which
share essentially complete knockout of menin may also share pheno
typic characteristics. Clearly, the number of cases examined is small
and needs to be expanded before firm genotypic/phenotypic
sions can be drawn.
Here, a MEN] gene mutation was found in exon 8 in germ-line
DNA in a patient who had no family history of MEN1. The patient
had ZES with normal serum calcium and prolactin levels, conflicting
levels of plasma PTH values, and an unclear past history of neck
surgery. Interestingly,the E363del mutation in this patient was iden
tical to a germ-line mutation previously described in two unrelated
MEN1 families (8, 9). Patients with MEN1 can initially present with
enteropancreatic endocrine tumors as the sole manifestation
disease (12, 21) and can, therefore, be difficult to distinguish from
patients with sporadic tumors without MENI. In one study, one-third
of the 28 patients with ZES with MEN! initially presented with ZES
and only developed hyperparathyroidism
later (12). Because 20—25%of patients with ZES have MEN! and
because their clinical management
patients with the sporadic form of the disease, the ability to screen
patients with ZES for MEN] germ-line mutations will be a valuable
tool for evaluation of ZES patients in the future.
Somatic MEN] mutations were detected in 2 of 12 (17%) sporadic
insulinomas.One of the two insulinomas
second copy of the MEN] gene. Five insulinoma cases had a loss of
one copy of the MEN! gene without a detectable MEN] gene mutation
produce frameshifts resulting
stable, contains little of the
conclu
of their
or pituitary disease years
differs significantly from that of
also showed a loss of the
in the other copy. In 50% (6 of 12) of insulinomas,
mutation nor deletion could be detected. Thus, the MEN! gene ap
pearsto be alteredin a subsetof sporadicinsulinomas.Other,as yet
unidentified tumor suppressor genes/oncogenes
pathogenesis of the majority of insulinomas
here for insulinomas are similar to recent data obtained in a group of
sporadic parathyroid tumors analyzed for MEN! gene mutations (23).
MEN] mutations were identified in 7 of 33 (21%) sporadic parathy
roid tumors, and LOH on l!ql3
parathyroid tumor samples.
No mutation in the MEN! gene was detected by SSCP analysis in
any exons from 17 of 27 sporadic gastrinomas and 5 of 12 insulino
mas that showed allelic deletion. This work may, in fact, underesti
mate the prevalence of MEN] somatic mutations in sporadic entero
pancreatic endocrine tumors, as the sensitivity rate of SSCP analysis
for detection of single base substitutions
Moreover, some of these tumors may contain a mutation in regions of
the MEN! gene that were not screened, such as the promoter. In the
absence of a mutation or deletion, an alternative mechanism, such as
hypermethylationof a CpG island as described in other tumor sup
pressor genes (25, 26), may inhibit transcription of the second copy of
the MEN! gene. Lastly, another as yet unidentified tumor suppressor
gene that is importantfor pathogenesis
endocrine tumors may exist near the MEN! locus. Further studies are
necessary to investigate such alternative mechanisms in enteropancre
atic endocrine tumorigenesis.
In summary, a high frequency
deletions was observed in localized and metastatic sporadic gastrino
mas. The findingthat the MEN!
etiological role in sporadic gastrinoma should lead to a better under
standing of the molecular pathogenesis
should aid in methods of diagnosis
gastrinoma. The study suggeststhat MEN!
4685
neither MEN!
may contribute to the
(22).The data reported
was detectedin 13 of 33 (39%)
is estimatedat 80% (24).
of some enteropancreatic
of MEN!mutations and allelic
tumor suppressorgene has an
of this malignant tumor and
and treatmentof patients with
mutations and allelic
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